Abstracts Book - The 32nd World Congress of Biomedical

2 016
IFBLS
Kobe Japan
I F B L S 2 016
Abstracts Book
http://www.ifbls2016.org/
The 32nd World Congress of
Biomedical Laboratory Science
International Innovation of Laboratory Medicine
−Basic and Advanced−
Date :
August
31 (Wed.)-September 4 (Sun.), 2016
Venue : Kobe International Conference Center, JAPAN
Congress Organizer
International Federation of Biomedical Laboratory Science
Japanese Association of Medical Technologists
CONTENTS
Keynote Speech ……………………………………………………………… 5
Invited Lecture … …………………………………………………………… 6
Special Lecture… …………………………………………………………… 8
Educational Lecture… ……………………………………………………… 9
Symposium …………………………………………………………………… 14
Case Conference … ………………………………………………………… 30
Oral Presentation … ………………………………………………………… 33
Poster Presentation … ……………………………………………………… 43
National Action Plan on Antimicrobial Resistance in Japan … …… 189
Keynote Speech
Invited Lecture
Special Lecture
Educational Lecture
Symposium
Case Conference
Applications of Mass Spectrometry in Laboratory Medicine
Koichi Tanaka
Shimadzu Corporation, Japan
Invited Lecture
Special Lecture
Educational Lecture
Laboratory medicine commonly requires analytical instruments that can quickly, easily, and inexpensively identify compounds and
their forms associated with diseases with the highest sensitivity, quantitative performance, and specificity using minimally invasive
techniques. Advancements in technical innovation for mass spectrometers (MS) have now led to techniques that meet such requirements.Besides identifying known substances, other purposes and advantages of MS that are not well known to the public include using MS as a tool to discover unknown phenomena and compounds. An example is clarifying the mechanisms of human diseases. The
human body has approximately 100 thousand types of proteins, and there may be more than 10 million types of post-translationally
modified proteins and their metabolites. Most of them have yet to be discovered and their discovery may give birth to new academic
fields and lead to a better understanding of diseases, development of new drugs, and other advancements.For example, using the MS
system developed under “Contribution to drug discovery and diagnosis by next generation of advanced mass spectrometry system,”
one of the 30 projects funded by the “Funding Program for World-Leading Innovative R&D on Science and Technology” (FIRST program), and using other individual elemental and basic technologies, we succeeded in discovering new disease biomarker candidates,
such as for Alzheimer’s disease and cancers.Further contributions by MS to laboratory medicine can be expected through the development and improvement of new techniques, efforts to verify discoveries, and through multidisciplinary communication, especially
with researchers and engineers directly involved in using instruments for medical applications.
Keynote Speech
KS
WHO-Mini Lecture
Symposium
Case Conference
Oral Presentation
Poster Presentation
5
Jennifer Young
Invited Lecture
As the HPV vaccination rises in popularity and new advances in molecular diagnostics make primary HPV testing a reality; the cytopathology community is faced with a responsibility to understand the implications of primary HPV screening and associated clinical
trials. The Cytopathology Education and Technology Consortium (CETC) issued a formal statement to the U.S. Food and Drug Administration (FDA) expressing safety and efficacy concerns around HPV as a primary screening tool – underlining the key considerations
when contemplating using HPV testing alone. Concerns focused on quality control, HPV negative cervical cancer, HPV testing methodologies, and appropriate triage of high risk HPV patients. This lecture will provide a review of the HPV virus, associated progression, and vaccine; current recommended screening guidelines; and the Athena trial and ancillary studies on HPV as a primary screening tool.
Special Lecture
HPV DNA Testing in Primary Cervical Cytology Screening: The
Controversies in its Clinical Applications
Keynote Speech
IL01
Objectives:
1. Describe the pros and cons of primary HPV testing versus co-testing
2. Understand the natural history of Cervical Cancer/HPV progression
3. Describe HPV vaccine options
4. Describe outcomes and limitations of the Athena trial and other related studies on co-testing
5. Understand CETC Safety and Efficacy Concerns with HPV Primary Screening
Director, International Strategy and Operations, American Society for Clinical Pathology, United States
Educational Lecture
WHO-Mini Lecture
IL02
Circulating cell-free DNA as a diagnostic tool in transplantation and
cancer
Michael Oellerich
Symposium
University Medicine Göttingen (UMG), Institute for Clinical Pharmacology, Germany
Case Conference
Oral Presentation
High-qualitiy genomic analysis is critical for a personalized medicine approach to cancer patient management. Tumor-specific
genomic alterations identified in cell-free DNA (cfDNA) from patient blood samples seem to be helpful to monitor relapse or response
to treatments. Noninvasive analysis of acquired resistance to cancer therapy is possible by detection of somatic mutations in plasma
cfDNA and allows for the identification of specific mutations selected by treatment such as EGFR T790M , in patients with NSCLC
treated with gefitinib. Gains and losses of chromosomal regions have been detected in plasma tumor-specific cfDNA as copy number
aberrations and can be used to compute a genomic copy number instability index (CNI) of cfDNA. The CNI obtained by massive parallel sequencing discriminated prostate cancer from both controls, and benign prostatic disease. CNI change may also serve as a potentially early predictor of response to standard chemotherapy for various other cancer types (e.g. NSCLC, colorectal cancer, pancreatic
ductal adenocarcinomas). Transplantation biomarkers have especially attracted attention, because there are still unresolved problems (e.g. irreversible chronic allograft dysfunction, side effects of standard immunosuppression) that limit long-term outcome. There
are limitations to how immunosuppressive drugs are currently monitored. Therapeutic drug monitoring is more useful to prevent
toxicity than to predict efficacy. Biomarkers are needed that can be used to facilitate personalized immunosuppression. A particularly
promising new approach for early detection of graft rejection is based on the determination of graft-derived circulating cfDNA, using
droplet digital PCR. This assay has the advantage that it directly interrogates the health of the donor organ (“liquid biopsy”). It may
also be useful to guide changes in immunosuppression and to monitor immunosuppression minimization. In the future, such personalized medicine approaches will shift emphasis from reaction to prevention, provide actionable health care information and could
improve outcomes at lower healthcare costs.
Poster Presentation
6
Mobile and digital health to support Universal Health Care and the
Sustainable Development Goals
Per Erlend Hasvold
World Health Organization, Switzerland
Invited Lecture
Special Lecture
Non-communicable diseases (NCDs) are causing 36 million deaths annually. Many of these are premature deaths, costing the societies enormous amounts in terms of lost productivity, in addition to the obvious tragedy and pain to the families and dependents. Also,
we need to consider the loss of quality of life and the burden of disease from people living with complicated and chronic conditions.
NCDs are not a typical rich-mans problem. In fact, a larger percentage of premature deaths occur in low and lower-middle income
countries.
There are currently more mobile phone subscriptions than there are people on this planet. Mobile phones provide a means to reach
further and to more people than most other channels of communication and information.
The United Nations have defined 17 Sustainable Development Goals (SDGs). These emphasize the importance that health has on all
parts of society. SDG 3 is: Ensure healthy lives and promote well-being for all at all ages. SDG target 3.8 aims at achieving Universal
Health Care (UHC) for all. Health is woven into almost all the other SDGs as well.
The SDGs are also about partnerships, and sees partnerships as a key element of achieving the goals.
Another key element of the SDGs is the emphasis on building good systems for health and other sectors of society.
The Be Healthy Be Mobile joint WHO and ITU initiative on mHealth for NCDs is now supporting national scale, sustainable mHealth
programs in nine countries. These national programs demonstrate how mHealth can be used to reach patients, the general public,
and healthcare providers.
Keynote Speech
IL03
Educational Lecture
WHO-Mini Lecture
Symposium
Case Conference
Oral Presentation
Poster Presentation
7
Keynote Speech
SL01
Challenges and Prospects for International Standardization of
Molecular-genetic Testing in the Era of Genomic Medicine
Hayato Miyachi
Tokai University School of Medicine, Department of Laboratory Medicine, Japan
Invited Lecture
Special Lecture
Educational Lecture
WHO-Mini Lecture
Elucidation of the molecular pathogenesis of diseases and application of emerging technologies for testing and therapy have resulted in
a series of paradigm shifts in patient care, from conventional to personalized medicine, towards genomic medicine. This has been promoted by companion diagnostics and molecular targeted therapy, tailoring treatment to individual characteristics of each patient. Precision medicine has been accelerated by integrating the enhanced resolution of molecular analysis, mechanism clarity, and therapeutic relevance through genomic knowledge.As clinical applications of molecular-diagnostic testing is expanding and disseminating worldwide,
its international standardization becomes increasingly important. Now a number of standards are proposed and developed for moleculargenetic testing as a result of regional and international efforts. The 2012 version of ISO 15189 (ISO 15189: 2012) (Medical laboratories
- requirements for quality and competence) expanded its scope, covering genetic testing. ISO standards under development include that
for microbial pathogens, preanalytic process of various types of samples such as blood, frozen tissues and FFPE, and multiplex molecular
analysis.In Japan, medical laboratory accreditation program has been implemented on the basis of ISO 15189, under the cooperation of
the Japanese Committee for Clinical Laboratory Standards (JCCLS) and the Japan Accreditation Body (JAB). It is carried out in accordance with the specific guidance document Guide RM300: 2014. There are major issues in ISO 15189 medical laboratory accreditation in
Japan. As for the requirement in medical policy, conventionally the accreditation is not mandatory. Regarding the subject in the accreditation program, it is currently limited to common laboratory tests, which are covered by health insurance, with regulatory approved,
and not applied to the molecular-genetic tests based on emerging technology. In 2015, clinical research core hospital was established
to serve as the bases for conducting international-level clinical studies and playing a central role in implementing doctor initiated clinical trials, in order to create innovative drugs and medical devices. It is required to get accredited under ISO 15189. In order to achieve
the purpose of a clinical research core hospital, there is a need for development of an accreditation program to cover a lack of insurance
coverage, such as molecular-genetic testing. To this end, the development of a guidance document intended for the molecular-genetic
testing by which medical laboratories can implement a quality system to meet ISO 15189: 2012 is to be considered. In its clinical implementation, medical laboratories are faced with challenges for a new and pivotal role concerning accurate measurement using stored
samples, bioinformatics availability, practical decision-making algorithms, and ethical issues regarding incidental findings. In addition, it
is necessary to secure human resources who are familiar with implementation, operation and management of molecular-genetic testing.
This presentation will focus on provision of information and thoughts on current efforts and expectations of medical laboratory to solve
these quality issues on molecular-genetic testing in the era of genomic medicine with regards to the international standardization.
SL02
Bridging the gap between Clinical Medicines and Laboratory Medicine
on the Urine Sediment Microscopy
Yu Chu Su1,2
Symposium
1 Department of Laboratory Medicine, National Taiwan University Hospital, Taiwan
2 Institute of Biomedical Engineering, National Taiwan University
Case Conference
Urine sediment microscopy is important for evaluating patients with possible urinary tract or kidney diseases. However, there is no
consensus standard for urine sediment microscopy yet. Nephrologists and medical laboratory technologists apply distinct protocols
for the urine sediment microscopy and there are major discrepancies in the differential counts of the formed elements, which could
affect the clinical diagnoses. To resolve these issues and improve the agreement between physicians and medical laboratory scientists, we reviewed the most commonly followed guidelines and identified the sources of variability within those protocols. Next, we
proposed a new assessment method for glomerular bleeding based on a dysmorphic red blood cell score by bright-field microscopy.
Finally, we adopted the Sternheimer stain, which is recommended by European Confederation of Laboratory Medicine, Japanese
Association of Medical Technologists, and Taiwan Society of Laboratory Medicine, for the urine sediment microscopy because the
enhanced contrast between the formed elements in urine sediment and thus can provide better report quality of urine sediment microscopy for diagnosing, screening, and monitoring a wide spectrum of kidney and urinary tract diseases.
Oral Presentation
Poster Presentation
8
Laboratory Diagnosis of Dengue Fever Diseases: Current and Future
Perspectives
Chuan-Liang Kao
Department of Clinical Laboratory Sciences and Medical Biotechnology, College of Medicine, National Taiwan University, Taiwan,
ROC, Taiwan
Educational Lecture
WHO-Mini Lecture
Strengthening Laboratory Capacity in Africa.
Special Lecture
EL01-2
Invited Lecture
Dengue virus (DENV) belongs to the family Flaviviridae and consisted of four serotypes (DENV1-4). DENV infections in humans cause
a spectrum of illness ranging from in-apparent or mild febrile illness to severe and fatal hemorrhagic disease (Dengue hemorrhagic
fever/Dengue shock syndrome). An estimated 50 million people in the world widely experience dengue each year and approximately
500,000 of them are associated with severe, life-threatening disease. Since no specific treatments are available for dengue virus infections, accurate diagnosis is critical for the early initiation of specific preventive health measures to curtail epidemic spread and
reduce economic losses. Commonly used diagnosis methods for confirming dengue infection involve virus isolation or RNA detection
in plasma /serum/tissues, and the presence of dengue virus specific antibodies in serum and other body fluids. However, all of these
methods have disadvantages such as time-consuming and tedious work, non-specific outcome or high-cost instrument for amplification. Recently, several techniques have been developed for improving the turn-around time and simply the performance of laboratory diagnosis of dengue virus, including the flow cytometry method for early detection of cultured virus, real-time PCR to detect
DENV RNA and early detecting free viral non-structure antigens in blood samples. Newly established methods must be standardized
to maintain high quality laboratory performance. Laboratory diagnostics must be tailored to a specific laboratory environment, the
objectives of clinical needs and the availability of clinical specimens. Speed and accuracy of diagnosis must be balanced against test
cost and availability.
Recently, the emerging and spreading of Zika virus were recorded in Africa, the Americas, Asia and the Pacific. It caused a social
panic world widely. The clinical manifestation of Zika virus infection is similar to the dengue fever infection. Future challenge is to
set up laboratory methods for the differentiation diagnosis of dengue and Zika virus infections.
Keynote Speech
EL01-1
Patrick Joseph Chattad
This lecture presents an overview of medical laboratory practise in Africa and the concerted efforts of WHO and its partners towards
strengthening laboratory systems in recent years.
Symposium
FIMLS, Cameroon
Case Conference
Oral Presentation
Poster Presentation
9
Keynote Speech
EL01-3
New Professional Certification Program for Laboratory Biosafety
Professionals
Maureen Ellis
International Federation of Biosafety Associations, Canada
Invited Lecture
Special Lecture
Educational Lecture
The International Federation of Biosafety Associations is a worldwide non-governmental organization with 37 Member Biosafety
Associations in all regions of the world. The IFBA’s mission is safe, secure and responsible work with biological materials and the organization conducts its work in partnership with its Members, international agencies (e.g. World Health Organization) and like-minded professional associations (e.g. International Federation of Biomedical Laboratory Science). The IFBA is currently implementing a
Memorandum of Understanding with the IFBLS and its network of local Members to strengthen biosafety and biorisk management in
laboratories around the world. Most recently, the IFBA has launched a unique international certification program for laboratory professionals and scientists in the area of biosafety and biorisk management. This exciting new program creates a certification scheme
that establishes competencies in a variety of technical disciplines (e.g. biosafety, risk assessment, waste management) allowing a professional to advance his/her expertise and qualifications throughout their laboratory career. IFBA’s certificants bring increased value
to their employers by demonstrating competence to carry out their responsibilities and by achieving high standards of excellence,
professionalism, and continuous learning. By earning certifications from the IFBA, individuals reap the benefits of safer workplaces,
career advancement, and international recognition among colleagues. The IFBA’s Professional Certification in Biorisk Management
and Professional Certification in Biological Waste Management are both currently being offered and certifications in Biocontainment
Facilities, Biosafety Cabinets and Biosecurity will be released shortly. Exams are offered both online using a remote proctoring system and on paper in conjunction with Member conferences and events. IFBA’s Members are providing pre-exam training for IFBLS
Members. To date, 150 professionals have participated in the program from 25 countries around the world. Future expansion of the
program include translation into key languages including French, Spanish and Russian.
WHO-Mini Lecture
EL02-1
The approach for the safety of serological screening tests
Chitose Inoue, Yoshiharu Suzuki
Symposium
Japanese Red Cross Kanto-Koshinetsu Block Blood Center, Japan
Case Conference
Oral Presentation
Japanese Red Cross Society (JRCS) has performed several safety measures, i.e., donor identification, donor interviews, laboratory
screening for TTI, and inventory hold, to supply safety blood products for patients. Among them, laboratory screening has played
one of the most important roles to ensure the safety blood products. At JRCS blood centers, we have conducted 6 serological screening tests, i.e., HBV, HCV, HIV, Syphilis, HTLV-1, and B19, and NAT (Nuclei-acid testing) for HBV, HCV, and HIV. Herein, I would like to
share the efficacy of the laboratory screening and the challenge for the prevention of TTI in Japan.JRCS has recently developed noteworthy approaches for further safety measures such as the revision of the criteria for HBcAb in August 2012, and introduction of
individual NAT (ID-NAT) in August 2014. With respect to the criteria for HBV screening, we have used the combination of HBsAb and
HBcAb as well as HBsAg and NAT screening. Before the revision of the criteria, the blood samples with less than 12.0 C.O.I of HBcAb
had been viewed as low virus titer. Then, they had been accepted as a safety product even if their HBsAb level is less than 200mlIU/
ml. However, in order to eliminate any risks of TTI, the C.O.I. level of HBcAb was lowered to 1.0 C.O.I from 12 C.O.I. Since the revision
of HBcAb criteria, HBV past-infection has not been reported so far. Moreover, the employment of ID-NAT has been highly effective to
shorten the window periods of virus, resulting in no report for the cases of TTI from August in 2014 to December in 2015.Theoretically, as compared to 20-pool NAT, which was employed from 2004 to 2014, the window periods with ID-NAT were shortened to 34
days from 44 days, 23 days from 24.5 days, and 11 days from 13.5 days for HBV, HCV, and HIV, respectively. In fact, even highly
low-concentrated of HBV-DNA, which could not be detected with 20-pool NAT, have been detected in 13 blood samples by making
use of ID-NAT from August in 2014 to March in 2016. Therefore, ID-NAT has been valued as the most advanced technology to directly detect even low levels of virus in blood samples.Employing this high-technology has significantly contributed to the safety products; however, we have been still straggling with the risks of TTI due to window periods. To challenge it, it will be quite important to
promote the blood donation with responsibility.
Poster Presentation
10
Safety Measures to Blood for Transfusion implemented in the Blood
Preparation Process: Leukocyte Reduction, Irradiation and Visual Inspection
Hideto Ogawa
Japanese Red Cross Kinki Block Blood Center, Japan
Invited Lecture
Special Lecture
Safety of blood and blood components for transfusion should be secured as much as possible by implementing safety measures both
in blood program and clinical transfusion practice from blood collection to transfusion to the recipient. Actually, blood center has
achieved great improvement in safety of blood products by implementing many safety measures in the different processes, including
blood collection, laboratory testing, preparation and distribution.
The main safety measures in the preparation process are pre-storage leukocyte reduction, irradiation and visual inspection of blood
products under the preparation process. The pre-storage leukocyte reduction by use of leukocyte reduction filters or on the basis of
apheresis device significantly reduces the residual leukocytes. The leukocyte – reduced blood products contain only less than 1 ×
106 residual white blood cells per bag in Japan. It has been reported that pre – storage leukocyte reduction is helpful in reducing adverse effects, such as febrile reactions. Irradiation to blood products by Gamma or X – ray irradiation prevents transfusion – associated graft versus host disease, which is a rare but fatal complication of transfusion. Red blood cell and platelet components (but not
fresh frozen plasma,) are irradiated at the dose of 15Gy – 50Gy in Japan. Visual inspection of blood and blood products are mainly
for discoloration, bacteria contamination, hemolysis, contamination of foreign substances, and lipemia. However, lipemia is acceptable for transfusion. Visual inspection needs to be performed not only in preparation process, but also before entering preparation
processes and before shipping the blood products.
I will give a full detail of pre-storage leukocyte reduction, irradiation and visual inspection of producing blood products in the seminar.
Keynote Speech
EL02-2
Educational Lecture
Experiences regarding usage of apheresis in a small blood bank – How to get
the best utilization of blood donors using apheresis –
WHO-Mini Lecture
EL02-3
Henriette L Michelsen
Oral Presentation
Poster Presentation
11
Case Conference
The vast majority of Norway’s blood component preparation are decentralized, and there are many factors to take into account when
deciding on which kind of production line is most suitable. In Norway there are no national guidelines to what kind of production
line the hospitals are required to follow.
In the blood bank in Kristiansand we have based our production line on a combination between whole blood collecting and apheresis. We do not prepare platelet concentrates from buffy coat, because we use blood bags with in line filter. All our platelet concentrates are collected from apheresis.In addition to platelets, we also collect red blood cells, plasma and multi component. Some of the
reasons why we collect red blood cells and plasma, is to maintain our expertise on the use of the apheresis machine and venipuncture technique.
National wise Kristiansand is a small blood bank with approx. 3200 red blood cell units collected each year. Approx. 30 % of that
is collected by apheresis which makes Kristiansand blood bank dominating in the usage of apheresis in Norway. Over the years our
focus on apheresis have built up a high level of expertise and giving us increased knowledge in selecting donors for apheresis, so we
can optimize the utilization of our donors.
Collecting blood by apheresis gives us more flexibility and the preparation of components does not demand the same amount of resources. Most importantly, we utilize each donor so we get the most out of each collection.
Symposium
Sørlandet sykehus Kristiansand, Norway
Keynote Speech
EL03-1
Standardization of Cytomorphologic Judgement and Training in Clinical
Laboratory
Leila Lany Florento1,2
1 United Laboratories, Inc, Philippines
2 The Philippine Association of Medical Technologists, Inc. (PAMET)
Invited Lecture
Special Lecture
Educational Lecture
Cytomorphology is the study of structure of cells. Morphological cell analysis is a key issue for abnormality identification and classification, early cancer detection, and dynamic changes analysis under specific environmental stress. The quantitative results and
primary, objective, and reliable, which is beneficial to pathologists in making the final diagnosis and providing fast observation and
automated analysis systems.The Medical Technologists are well-trained in the preparation of smears and staining techniques, reading and evaluation of blood cell morphology as part of Hematology subject and preparation of slides and staining techniques in Histopathology subject to complete the Bachelor of Science in Medical Technology. For cytological studies, skills in special techniques
are learned during actual work. The pathologists read the slides and make evaluation. In other countries, there is Cytotechnology
program to train the Med Techs in reading and evaluation of the slides.The laboratory interpretation of blood film morphology
is frequently a rapid, accurate, and cost effective final-stage of blood count analysis. However, the interpretation of findings often
rests with a single individual, and errors can carry significant impact. Cell identification and classification skills are well supported
by existing resources. A peripheral blood film evaluation should be part of complete blood counts. Proper sample collection, slide
preparation, and staining are essential to accurately evaluate a blood film, as is the correct use of a high-quality microscope.Medical
technologists perform morphologic assessment of blood cells using microscope to identify abnormalities in all cell lines. This part of
the hemogram is the most subjective and difficult to quantitate. If abnormalities are found, these are semi-quantified as mild, moderate and marked or few, moderate and many, depending on the abnormality. Proficiency in recognizing and interpreting morphologic abnormalities comes only with practice and familiarity with normal features of each species. An important part of blood smear
examination is to separate artifact from pathologic changes. This can be challenging and also takes experience and proficiency. All
expertise trainings are obtained during actual practice under the supervision of laboratory supervisors.Cytology is a useful clinical
tool for investigation of disease processes, and the techniques and their interpretation have developed into an entire discipline. Full
cytologic interpretation requires a good quality sample. The technique appears to be simple, but consistently obtaining good-quality
samples requires practice. If samples are sent to a laboratory for interpretation, sending more than one is recommended. This requires a good staining technique and a good quality microscope with a range of objectives, including oil immersion.This field is being
shaped predominantly by new techniques for imaging and for acquiring and processing samples, advances in molecular diagnosis
and therapeutics.
WHO-Mini Lecture
EL03-2
Microscopy quality control and an associated education system at our
hospital blood laboratory
Takako Tamura
Symposium
Department of Blood Tests, Clinical Laboratory, Tokyo Women’s Medical University Hospital, Japan
Case Conference
Oral Presentation
Poster Presentation
Quality control is very important in laboratory examinations. It is relatively easy to check the numerical data output by automatic
analyzers. However, quality control in the morphological field is left to the judgment of medical technologists, and the management
of their abilities is very difficult. Abnormal cells are frequently seen in the peripheral blood and bone marrow of patients with blood
disorders, and blood test technologists are likely to be the first to discover them; thus, the abilities of technologists that perform
microscopic examinations can influence diagnostic outcomes. Therefore, we developed a microscopy quality control system and a
related educational program at our blood laboratory. In this system, we evaluate the abilities of technologists who routinely perform
microscopic examinations and provide them with appropriate instructions. An outline of the contents of this presentation is shown
below:
1, Introduction to Tokyo Women’s Medical University Hospital laboratory
2, The education provided to new staff at the blood laboratory
3, The ongoing training provided to technologists who routinely conduct microscopic examinations
4, Instructions regarding microscopy given to night duty staff
5, Holding of meetings between affiliated hospitals to present educational research papers
Above all, the evaluations of the technologists’ abilities and the education provided to them are carried out in a positive manner at
our blood laboratory. It is very important for blood test technologists to read and understand patients’ overall data, and to be aware
of disease characteristics as well as being able to distinguish between different cell types.Therefore, we created an original test. During the test, microscopic examinations of blood samples are performed to evaluate diseases, but the technologist must also consider
other test results, such as biochemical data or the findings produced by an automated hematological analyzer. As a result, the ability
of the technologist, and the topics on which they require further instruction become clear. The accumulation of such learning leads
to the elucidation of findings that “a medical technologist should not overlook”, and we consider that it will improve the performance
of the blood laboratory, which is relied on by medical attendants. The improvements in the abilities of technologists brought about
by our system encourage the technologists to try to get certified qualifications, and 70% of the technologists in our laboratory acquire certified qualifications in some kind of hematology. Also, we hold meetings with affiliated hospitals to present research papers
once a year. In particular, we utilize these meetings to provide new technologists with an opportunity to gain experience of making
presentations.
12
Performance Evaluation of Liquid Chromatography-Tandem Mass
Spectrometry as Confirmatory Tests for Drugs of Abuse from Urine Samples
Minji Yoon, M.T.1, Sun-Hee Jun, M.T.1, Kyunghoon Lee, M.D.1,2, Sang Hoon Song, M.D.2, and Junghan Song, M.D.1,2
1 Department of Laboratory Medicine, Seoul National University Bundang Hospital, Seongnam, Korea
2 Department of Laboratory Medicine, Seoul National University College of Medicine, Seoul, Korea
WHO-Mini Lecture
The role of mass spectrometry in clinical laboratory: focusing on vitamin
D testing
Educational Lecture
EL04-2
Special Lecture
Key words: drugs of abuse, multiplex assay, tandem mass spectrometry
Invited Lecture
Background: Recently, liquid chromatography-tandem mass spectrometry (LC-MS/MS) has become a more efficient tool for quantitative and confirmatory tests of drugs of abuse (DOA), as it does not require any additional derivatization reaction. In this study, we
evaluated the analytical performance of the multiplexing LC-MS/MS method for 8 major metabolites: amphetamine, methamphetamine, codeine, 6-acetylmorphine, morphine, phencyclidine (PCP), benzoylecgonine, and tetrahydrocannabinol carboxylic acid (THCCOOH).
Methods: Random urine samples were processed for analysis by solid phase extraction. And then all prepared samples were analyzed
on Waters UPLC system with ACQUITY UPLC HSS T3 column (2.1*100mm, 1.8 µm; Waters, Watford, UK). All drug concentrations
were determined by Quattro premier mass spectrometer (Waters). The assay performance was evaluated including imprecision, limit
of quantification (LOQ), and linearity. Stored external quality control materials were measured for comparison to the peer group
mean values.
Results: The coefficients of variation of inter-assay and intra-assay were 3.3–5.6% and 3.7–17.2%, respectively. The LOQ of amphetamine, methamphetamine, codeine, 6-acetylmorphine, morphine, PCP, benzoylecgonine, and THC-COOH were 50, 50, 25, 5, 25, 10,
12.5, 5 ng/mL, respectively. Linearity was acceptable at 5 levels of each analyte (R 2 = 0.978–0.996). Correlation with the external
quality control materials were acceptable (r = 0.91 – 0.99).
Conclusions: The performance of our multiplex DOA test was clinically acceptable. This method is simple, efficient and accurate
enough to be used as confirmatory tests for DOA. According to Korean laws, we have performed our method as confirmatory test
when the drug screening test is positive.
Keynote Speech
EL04-1
Mamoru Satoh, Fumio Nomura
Oral Presentation
Poster Presentation
13
Case Conference
Mass spectrometry (MS) is a powerful analytical tool used in an increasing number of clinical laboratories around the world. One
of the most familiar applications to clinical laboratory would be the use of matrix-assisted laser desorption ionization–time of flight
mass spectrometry (MALDI-TOF MS) based microorganisms identification. The methodology based on the difference in protein
expression profiles of microorganisms is simple and quick. Rapid and accurate identification of microorganisms, a prerequisite for
appropriate patient care and infection control, is a critical function of clinical microbiology laboratory. Chiba University hospital
has actually set up a MALDI-TOF MS system for use in routine testing. One of the most effective events, the time required to identify
microorganisms from a blood culture bottle has been shortened to half. Indeed, there has been a revolutionary shift in clinical diagnostic microbiology. Moreover, Liquid chromatography (LC) coupled with tandem mass spectrometry (MS/MS) has been used for
newborn screening, toxicology, therapeutic drug monitoring, endocrinology, and more recently for measurement of targeted proteins
and peptides. Measurement by LC-MS/MS using the isotope-dilution LC-MS/MS (ID LC-MS/MS) is a method of internal standardization to spike the stable isotope labeling of the analyte to be measured in all samples. Sensitivity, stability at low concentration range,
and linearity are excellent as compared to those of immunoassays because the analyte can be measured directly by MS without
the antibody. As described above, distance between clinical testing and mass spectrometry is now very close. In this presentation, I
would like to discuss our experiences of the clinical application of LC-MS/MS for vitamin D measurements.
Symposium
Division of Clinical Mass Spectrometry, Chiba University Hospital, Japan
Keynote Speech
SY01-1
Morphology of 9p21 Homozygous Deletion-Positive Pleural Mesothelioma Cells
Analyzed Using FISH and Virtual Microscope System in Effusion Cytology
Shinji Matsumoto, Katsumi Kobata, Makoto Hamasaki, Kazuki Nabeshima
Department of Pathology, Fukuoka University Hospital, Japan
Invited Lecture
Special Lecture
BACKGROUND: In malignant pleural mesothelioma (MPM), most patients first present with pleural effusion; thus, cytologic analysis is the primary diagnostic approach. However, the cytologic distinction between MPM and reactive mesothelial cells (RMCs) in effusions can be extremely difficult. Homozygous deletion of the 9p21 locus, the site of the cyclin dependent kinase inhibitor 2A/p16
(CDKN2A/p16) gene, frequently occurs in MPM but has never been reported in RMCs. The aim of this study was to define the cytomorphological characteristics of MPM cells, identified by the presence of 9p21 homozygous deletion by fluorescence in situ hybridization (FISH).METHODS: For this purpose, cells on smear preparations were recorded using a virtual microscope system and were
subjected to FISH analysis. Thereafter, 9p21 homozygous deletion-positive cells were identified in the recorded virtual slides, followed by analysis of their morphological characteristics. RESULTS: Mesothelioma cells positive for the 9p21 homozygous deletion
exhibited significantly more frequent cell-in-cell engulfment(with or without hump-like appearance), multi-nucleation (more than 2
nuclei), and larger multicellular clusters composed of more than 10 cells than did 9p21 deletion-negative RMCs. Possible cutoff values are also proposed for these morphological markers to differentiate MPM cells from RMCs. CONCLUSIONS: These morphological
differences and cutoff values are useful for cytological differentiation of mesothelioma cells from RMCs. We also introduced a novel
methodology to analyze tumor morphology using a combination of the virtual microscope system and FISH.
Educational Lecture
Symposium
SY01-2
Characterization of cellular and molecular radiation effects in cancer
Experimental study in lymphoma and lung cancer
Fernando Mendes1,2,3,4, Ana Margarida Abrantes1,3, Cátia Domingues3,5, Paulo Rodrigues-Santos3,6,7,8, Ana Cristina Gonçalves3,5, Tiago Sales1, Rita Silva1, Jéssica Estrela2,
João Encarnação1, Ana Salome Pires1,3,4, Mafalda Laranjo1,3,4, Vera Alves6, Ricardo Teixo1, Ana Bela Sarmento3,5,7, Maria Filomena Botelho1,3, Manuel Santos Rosa6
Case Conference
1 Biophysics and Biomathematics Institute, IBILI-Faculty of Medicine, University of Coimbra, Portugal
2 Polytechnic Institute of Coimbra, ESTESC-Coimbra Health School, Department Biomedical Laboratory Sciences, Coimbra, Portugal
3 CIMAGO, FMUC-Faculty of Medicine, University of Coimbra, Portugal
4 CNC.IBILI, Universidade de Coimbra, Portugal
5 Applied Molecular Biology and Clinical University of Hematology, Faculty of Medicine of University of Coimbra, Coimbra, Portugal
6 Immunology Institute, Faculty of Medicine, University of Coimbra, Coimbra, Portugal
7 Hematology Clinic Department Centro Hospitalar Universitário de Coimbra, Coimbra, Portugal
8 Immunology and Oncology Laboratory, Center for Neurosciences and Cell Biology, University of Coimbra, Coimbra Portugal
Oral Presentation
Poster Presentation
Radiotherapy (RT) is a common modality for cancer treatment. Lung cancer (LC) is a solid tumour while diffuse large B-cell lymphoma (DLBCL) is a hematopoietic tumour both with higher incidence and mortality rates.
We aim to determine and characterize RT effects in cell lines of small cell LC (H69, P53Mut), non-small cell LC (A549, P53Wild, and H1299, P53Null) and DLBCL
(Farage, P53Wild). We assessed viability (trypan blue assay), proliferation (Alamar blue assay), survival (clonogenic assay), cell death mechanisms (Annexin
V and propidium iodide double staining, BAX/BCL-2 ratio and mitochondrial membrane potential (MMP)), and cell morphology alterations (May GrünwaldGiemsa staining). We evaluated RT effects in oxidative stress (OS) and anti-oxidant defences. DNA damage was determined by comet assay, and expression of
phosphorylated and total P53 protein was assessed by Western blot.
We also aimed to evaluate RT effects on immune system (IS) of LC and DLBCL patients. We studied 8 LC and 9 DLBCL patients. After collecting peripheral
blood (PB), leukocyte, lymphocyte and regulatory T cells (Tregs) counts were determined by Lymphogram® and immunophenotyping, respectively. We evaluated 34 cytokines and chemokines relevant to IS response through ProcartaPlex™ Immunoassay Magnetic Beads kit.
RT decreased proliferation, viability and cell survival. Survival was adjusted to different cellular injury models (linear quadratic model for H1299 and Farage, and linear model for H69 and A549 cells). RT induced cell death was dose and P53 expression dependent. Farage and A549 cells presented cell death by
apoptosis. A549 and Farage cells had augmented P53 and phosphorylated P53 levels. Disruption of MMP and increased BAX/BCL-2 ratio indicates intrinsic
apoptotic cell death. H1299 and H69 cells showed necrotic cell death at higher doses. We observed arrested cell cycle in G0/G1 and S phases in A549 and Farage, and G2/M arrest in H69 and H1299 cells. OS and DNA damage stood out as very important processes involved in RT effects.
We observed that IS response to RT was cancer type dependent. Assessment of leucocyte count of LC patients showed changes in leukocytes, lymphocyte and
monocytes during treatment. No significant differences in leucocytes were found in DLBCL patients. LC patients presented changes in B cells, Natural Killer (NK)
and cytotoxic NK cells. In DLBCL patients there were changes in induced Tregs cells. Regarding Th1 cytokines and chemokines, we observed an increase in
INF- γ concentration in DLBCL patients. LC patients showed a stronger Th1 profile than DLBCL patients, which is confirmed by higher levels of IL-2, INF- γ
and IL-1 β . Th2 profile was characterized by highest concentration of IL-5 in DLBCL patients. There was an increase in IL-27 and IL-7 in LC patients.
We can have concluded that response to IR is dependent on cellular and molecular characteristics of tumour cells. Better knowledge and understanding of molecular characteristics of tumour and mechanisms of response to treatment is an asset concerning therapeutic decision and prognosis evaluation. In addition,
tumour microenvironment, as well as activity of IS on tumour cells in RT treatments, may contribute to better therapeutic outcome and long term survival.
Keywords: Radiotherapy; Ionizing radiation, LC; Large diffuse B cell Lymphoma, Oxidative stress; Cell cycle; Cell death; P53, Immune system
14
THE ROLE OF LABORATORY MEDICINE IN DIAGNOSIS, TREATMENT AND
MONITORING OF BREAST CANCER
Mirjana Stupnisek1,2, Ivan Milas1,3
1 Faculty of Medicine, J.J. Strossmayer University of Osijek, Osijek, Croatia
2 University Hospital for Infectious Diseases „Dr. Fran Mihaljevic“, Zagreb, Croatia
3 University Hospital for Tumors, Sestre milosrdnice University Hospital Center, Zagreb, Croatia
Educational Lecture
Symposium
Quality assurance of staining in the histology laboratory
Special Lecture
SY01-4
Invited Lecture
Breast cancer is the most common type of cancer among women worldwide (in both developed and developing countries), with nearly 1.7 million new cases diagnosed in 2012,
and is the leading cause of cancer death in women (0.5 million). In 2012, the estimated annual incidence of breast cancer in European countries was 92.8/100 000 and the mortality 23.1/100 000. In Croatia, incidence was 83.0/100 000 and the mortality 24.5/100 000 (EUCAN).
Early diagnosis and more effective treatment of invasive breast cancer resulted in significant mortality reduction, improvement of survival and the quality of life of the patients.
This paper discusses the role of laboratory medicine in the diagnosis and management of breast cancer in women, with emphasis on testing and biologic characteristics of the
tumour.
The diagnosis of breast cancer is based on clinical examination in combination with imaging, and confirmed by pathological assessment. Apart from imaging, pre-treatment disease evaluation includes pathological examination of the primary tumour and cytology/histology of the axillary nodes, if involvement is suspected. Other assessments include:
complete personal medical history, family history relating to breast/ovarian and other cancers, physical examination, a full blood count, liver and renal function tests, alkaline
phosphatase and calcium levels.
Pathological diagnosis should be based on a core needle biopsy, obtained preferably by ultrasound or stereotactic guidance. A core needle biopsy (if this is not possible, at least a
fine needle aspiration indicating carcinoma) must be obtained before any type of treatment is initiated. If preoperative systemic therapy is planned, a core needle biopsy is mandatory to ensure a diagnosis of invasive disease and assess biomarkers.
Final pathological diagnosis should be made according to the World Health Organization (WHO) classification and the tumour–node–metastases (TNM) staging system. The
pathological report should include the histological type, grade, immunohistochemical (IHC) evaluation of oestrogen receptor (ER) status and, for invasive cancer, IHC evaluation
of progesterone receptor (PgR) and human epidermal growth factor 2 receptor (HER2) gene expression. HER2 gene amplification status may be determined directly from all
invasive tumours using in situ hybridisation, replacing IHC or only for tumours with an ambiguous (2+) IHC score. Proliferation markers such as the Ki67 labelling index may
supply additional useful information, particularly if the assay can be standardised. ER/PgR and HER2 are the validated predictive factors, allowing the selection of patients for
endocrine therapies and anti-HER2 treatments.
Current progress in laboratory medicine, allows different tests that can be used for diagnosis, determining prognosis, selecting and monitoring treatment, and predicting and
detecting recurrence of breast cancer. Successful treatment is most likely to be achieved through cooperation of multidisciplinary team of experts (e.g. pathologist, oncologist,
BLSs). The clinical guidelines are used in order to standardize the procedures and criteria for diagnosis, management, treatment and monitoring of patients with breast cancer in
the Republic of Croatia.
The role of laboratory medicine is extremely important because, without it, it is impossible to make an exact diagnosis of breast cancer and to monitor disease course and outcomes of applied treatment.
Keynote Speech
SY01-3
Berit W. Revaa
Oral Presentation
Sustaining good quality requires both internal and external quality assurance measures.
Internal quality assurance includes knowledge about the methods, validation/verification, use of controls and microscopy. The selection of staining-methods reflects the material analyzed. The biomedical laboratory scientist has the knowledge about the methods
and is responsible for the staining results. The result depends on every step in the histotechnical process from fixation to staining.
External quality assurance could be arranged by independent companies or laboratories sending slides to each other for comparison.
Laboratories need external quality assurance to be accredited by ISO 15189.
Case Conference
Department of Pathology, Vestfold Hospital Trust, Norway
Poster Presentation
15
Keynote Speech
SY02-1
Explore the Possibility of POCT
Wen-Shyang Hsieh
President, Taiwan Society of Laboratory Medicine, Taiwan
Invited Lecture
Special Lecture
In the traditional process of clinical examination, the patient’s body fluid, excreta and tissue are managed for testing at the central
controlled and regulated laboratory (Central Lab). Meanwhile, the related laboratory quality system certification is also developed
and accredited in the same patterns.
However, with the development of science and technology and demand for clinical services, in vitro diagnostic instruments (IVD)
performed located near the patient are used increasingly. The simple and convenient features undoubtedly have an impact against
the existing health care environment, namely the Point of Care Test (POCT), or Near-patient testing.
In the past, perhaps limited by the effectiveness of the examination and clinical needs, POCT mostly is just a referential simple test
on health care. However, because these test results have become possible to change the decisions in the patient’s care, in particular
the treatment decisions. And should become part of the official clinical laboratory reports, its proper quality management system is
necessary to prepare, execute, and ensure efficiency and correction that in order to reduce risk and improve safety.
Simultaneously, clinical laboratory provide appropriate equipment, processes, in order to reach a proper quality management and
service needs for long-term care is an immediate and important issue now. So where for about POCT status of development and application is bound to become medical laboratory scientist duty that how to explore the possibility of POCT, and provide professional
services to long-term care team, through professional thinking and discussion.
Educational Lecture
Symposium
SY02-2
Usefulness of POCT at the time of the disaster
Mitsuaki Nagasawa
Case Conference
Department of Medical Technology and Sciences, School of Health Sciences at Narita, International University of Health and Welfare,
Japan
Oral Presentation
The clinical inspection support at the time of the disaster varies regarding the support contents and the support system at the time a
disaster occurs.
In Japan, there are many natural disasters such as earthquakes. After the Great East Japan Earthquake that occurred in 2011, and
the earthquake in Kumamoto this April, clinical laboratories were severely damaged.
In order to provide clinical inspection support in the stricken areas, the Japanese Association of Medical Technologists (JAMT) and
the Japanese Society of Laboratory Medicine (JSLM) offered to inspect clinical equipment and reagents (point-of-care testing (POCT),
in particular), to support the health care of refugees.
I report on the usefulness of the POCT support in the stricken areas.
A summary of strong earthquakes in Japan and clinical inspection support
Type of the earthquake
Poster Presentation
January 17, 1995
Name
The Great HanshinAwaji Earthquake
Characteristic of the
earthquake
Crushed to death
under buildings
Dead person / Missing
person
The cause of death
Medical correspondence
Earthquake with
tsunami type
Near-field urban earthquake
Date of onset
April 16, 2016
The Great East Japan
Earthquake
Tsunami, Nuclear
power generation
disaster of the wide
area
Mudslides
6,437 / 3
59 / 3
Crushed and burned
to death
15,894 / 2,561
Drowning, Frozen to
death
Crushed to death
Super urgent measures
－
－
Subacute chronic measures
Needs of laboratory test
Blood gas analysis,
Blood type, CK,
Creatinine, EKG
none
Hospital building damage
300 hospitals
234 hospitals
Operation method of the
inspection equipment
March 11, 2011
Kumamoto
Earthquake
Secondary inspection
380 hospitals
It is supported by the
import of the simple
apparatus and POCT
reagent
Cope with inspection equipment for emergency
(Dry chemistry)
16
POCT as a global health
Keynote Speech
SY02-3
Kaoru Terashima
Fujifilm Corporation, Japan
Educational Lecture
Symposium
How to improve local citizen-centered healthcare services using new
diagnostic and laboratory solutions
Special Lecture
SY02-4
Invited Lecture
We have developed a system that specializes in POCT fields such as the biochemistry and various immunoassay since 1984.POCT
is an important entrance for early diagnosis, treatment, and recovery by testing on the spot to care the patient. If a highly accurate
testing will be made in early stage, it will be expected to treat sooner, to prevent severity of the patient’s own, and to halt minimized
the impact of infection to the surroundings. To produce such a higher effect, it is necessary to increase its sensitivity and specificity
in order to perform a more reliable testing. However usually, there is a trade-off relationship between a simple and quick testing and
a higher accuracy testing. It is a common that the higher accuracy testing is needed more precise equipment and well-equipped facilities. In order to do this, it takes much time for testing, and we will lose the therapeutic opportunity if the case was a severe disease.
By using our own photographic developing technology we has succeed to develop higher sensitive immunoassay system using lateral
flow technology which is most popular, simple and rapid diagnostic reagent. Thus, especially in infectious diseases areas, it has become possible to contribute significantly to the definite diagnosis at an earlier stage.In Japan, launched the influenza diagnostic reagent using this technology from four years ago, it has contributed greatly to the positive rate improvement in the onset of an early
stage.On the other hand, from the point of view of global health, it is becoming increasingly serious infections problem of developing
countries that a lot of people are suffering from infectious diseases as HIV, malaria,and tuberculosis. We have started the application
of the Ebola hemorrhagic fever and tuberculosis using this technology.As a result, a testing that required the well-equipped location of equipment such as genetic testing, microscopic examination and culture test, will move to a testing more close to the patient
which can realize the simple fast and high sensitivity and high specificity, and we can expect to identify the presence of infection at
an early stage.In addition, for the BSL4 specimen such as Ebola hemorrhagic fever, we also developed the blood sampling method for
medical safety for the entire related peoples including MSF. At the same time, we will introduce a workflow leading to a safe diagnosis and treatment.
Martina Jurs
Poster Presentation
17
Oral Presentation
A new approach for local citizen-centered diagnostic partners using POCT, by The Danish Association of BLS’es.
A new ambitious Danish project makes it possible to improve both health- and life quality, create a better patient flow and reduce
pressure on the hospitals. By the use of new POCT technology and mobile laboratories it has proven possible to change the existing approach and paradigm. Now patients do not necessarily need to come to the hospital; the hospital can come to them!A hospital
reform in Denmark will soon result in fewer hospitals, thus increasing the need to improve local health care services. At the same
time Denmark face a growing number of elderly citizens as well as patients with chronical conditions consequently increasing the
pressure on the hospitals. The rapid improvement in POCT technology makes it, however, possible to perform diagnostic analysis at
a local level: fast, safe and accurate (if used by skilled professionals). This is suggested, by an independent study, facilitated through
mobile sampling, mobile laboratories and pre-diagnostic teams, resulting in a more effective use of existing resources and improved
health care services altogether.From this outset The Danish Association of Biomedical Laboratory Scientists works on a new strategy
to offer local citizens diagnostic analysis of the highest quality, outside hospitals. This is made possible through biomedical laboratory scientists in new functions.At the moment a new project involving a mobile laboratory runs as a two-year project: Two biomedical laboratory scientists work together with a nurse in a special-designed unit, which can be ordered to a patient’s home by the local
doctor or hospital. Almost fifty para clinical studies can be performed using POCT and other equipment placed inside the bus, this
with a response time of less than thirty minutes.This project is still work in progress. However, the current results point to the possibility of improving:
-The quality of diagnostic analysis
-The welfare of the patients
-The health budget.
Technology and the use of biomedical laboratory scientists at a local level makes it possible. Next step is to change the mindset of
decision makers in the heath sector.
Case Conference
dbio - Danish Association of Biomedical Laboratory Scientists, Denmark
Keynote Speech
SY03-1
Immunological and Histopathological Analysis of Graves' Disease
Patients with or without IgG4 Positive Cell Rich Infiltration
Keiko Inomata
Yamashita Thyroid & Parathyroid Clinic, Japan
Invited Lecture
Special Lecture
Educational Lecture
In order to clarify the etiology of a disease, we need to obtain a variety of information about the target disease. In this study, we analyzed the pathology
of IgG4 – related disease (IgG4 – RD) in the thyroid glands of patients with Graves’ disease, using immunological and histopathological techniques.
IgG4 – RD is newly recognized diseases, which are characterized by elevated serum IgG4 levels and the prominent infiltration of IgG4 – positive plasma
cells in the involved organs. They are in the group of diseases that exhibit similar histopathological features across multiple organs, such as pancreas,
liver, gallbladder, lacrimal gland, and salivary gland. In the thyroid, the cases that meet the diagnostic criteria for IgG4 – RD has been reported in Riedel’
s thyroiditis and Hashimoto’s thyroiditis, and they have been termed IgG4 – thyroiditis. Riedel’s thyroiditis is extremely rare, therefore, few cases of IgG4
– RD associated with Riedel’s thyroiditis have been reported. In contrast, because of the high prevalence of Hashimoto’s thyroiditis in Japan, cases of the
condition involving IgG4 – RD have been reported frequently. Features of IgG4 – thyroiditis in Hashimoto’s thyroiditis are significant fibrosis in the thyroid, rapid enlargement of the thyroid gland and pressure symptoms in the neck.
The diagnostic criteria for IgG4 – thyroiditis include an elevated serum IgG4 concentration (>135 mg/dL), an increased number of IgG4 – positive plasma cells (≥20/HPF), and an IgG4/IgG – positive cell ratio of >30% in thyroid tissue.
In Graves’ disease, some cases exhibit similar histopathological findings with Hashimoto’s thyroiditis, such as lymphocytic infiltration, stromal fibrosis,
follicle degeneration and eosinophilic changes in epithelial cells, namely Hashitoxicosis. We classified the cases of Hashitoxicosis into two groups depending on whether they met the diagnostic criteria for IgG4 – thyroiditis, and compared the thyroid function test results of the two groups, i.e., free
thyroxine, free triiodothyronine, thyroid stimulating hormone (TSH), TSH receptor antibody (TRAb), anti – thyroglobulin antibody (TgAb), anti – thyroid
peroxidase antibody (TPOAb), thyroid stimulating antibody (TSAb), thyroid stimulation blocking antibody and serum IgG4 levels. We also examined their
histopathological features using hematoxylin and eosin staining and immunohistochemical staining of IgG4. In addition, we examined the transition of
serological parameters, such as TgAb, TPOAb, TRAb and serum IgG4 concentrations serially before and after surgery in patients with Graves’ disease associated with IgG4 – thyroiditis.
It is not well known whether IgG4 is involved in the development and progress of IgG4 – RD or is merely a clinical marker for the present. In addition,
the mechanisms for the increases in the number of IgG4 – positive plasma cells and their subsequent tissue infiltration are not yet clarified. We investigated the recognition antigen of serum IgG4 in Graves’ disease patients with IgG4 – thyroiditis by measuring the reactivity of serum IgG4 with thyroid
antigens
I would like to report the results of the above research, and the role of IgG4 in IgG4 – thyroiditis is discussed by comparing the serological and histopathological features of the IgG4 – thyroiditis group and those of non IgG4 – thyroiditis group with Graves’ disease.
Symposium
SY03-2
Urinary exosomes: the future of biomarkers in renal disease
Aki Nakayama Howley
Case Conference
Bunkyo Gakuin University, Japan
Oral Presentation
Poster Presentation
Exosomes are vesicles (40–100 nm in diameter) that are released from most viable cells into various body fluids, including blood and
urine. Exosomes are involved in cell-to-cell communication in a variety of biological processes and can modify the cellular functions
of recipient cells. Exosomes are thought to represent a miniature version of the original cells, as they contain numerous cellular components such as proteins, lipids, and nucleic acids that are enclosed in lipid bilayer membranes. Pathophysiological alteration of cellular and extracellular components directly influences exosome contents. Therefore, they are considered a novel source of biomarkers for diseases. Conventional diagnostic tools for renal disease-related glomerular filtered proteins or creatinine lack specificity, and
invasive renal biopsy is the gold standard for diagnosis. Therefore, more specific and noninvasive renal tests are needed. Exosomes
can be isolated from biological fluids using ultracentrifugation, immune-affinity methods, or commercial available kits, and their
origin can be traced using surface markers, as the markers expressed on the exosomal surfaces are the same as those on the origin
cells. Exosomes in urine are derived from renal epithelial cells such as mesangial cells, the thick ascending limb of Henle’s loop,
and the collecting tubule. Therefore, urinary exosomes might represent a noninvasive replacement for renal biopsy. Furthermore,
exosome numbers and protein levels are constant in timed urine samples of healthy individuals. Here, we examined the urinary exosomes of healthy individuals by two-dimensional electrophoresis-based proteome analysis for renal markers. Approximately, 90
proteins were identified, including tropomyosin alpha-4 chain, which has been linked to osmotic stress in the thick ascending limb of
Henle’s loop. Development of immunoassays for these candidate exosomal protein markers will aid the design of a noninvasive and
specific evaluation test for renal dysfunction.
18
Newborn screening for primary immunodeficiencies – A quicker
diagnosis by screening test for a better state of health –
Claudia Finocchi, Elisa De Vitis
ANTeL-Assiatel/AITIC, Italy
Invited Lecture
Special Lecture
The immunodeficiencies (ID) are a set of diseases characterized by deficits of the defence immune system and by a high level of infections. The different types of immune disorders are rarely observed in clinical practice. Indeed, 5% of paediatric patients show suspect symptoms of an immunodeficiency that occur in 0,6% of all live births. Most of congenital immunodeficiencies can be identified
by researching new biological markers: T-cell excision circle (TRECs) and K-deleting Recombination Exision Circle (KRECs). TRECs
and KRECs are portions of DNA, which are removed during the recombination V(D)J in lymphocytes T and B. It has been shown that
their absence is capable of identifying children with immune deficiency directly on newborn spot. The method to show the presence
of TRECs and KRECs qualitatively and quantitatively is the Real-time PCR. The analytic method has been developed with 1200 samples from patients with no T and B cell deficiency (control group) and 20 samples from patients with immunodeficiency or deficiency
related to T-lymphocyte functionality. The analysis of sensitivity and specificity of this method has demonstrated over 1220 samples
a sensitivity of 100% and a specificity of at least 99,8%. This technique was then repeated on a screening study of people involving
the analysis of about 30 spots per day and amounting to more than 13000 spots until today.
In conclusion, the use of a newborn screening for immunodeficiencies can significantly improve the clinical course of the disease:
reaching a diagnosis as soon as possible allows actions that can be taken with a right therapeutic protocol aimed at improving the
patient’s state of health. In addition, the early diagnosis gives the opportunity to minimize lasting damage or mortality connected
with severe infections, occurring even in the first months of life. This allows a reduction of a premature morbidity and hospitalizations.
Keynote Speech
SY03-3
Educational Lecture
Seroprevalence and risk factors associated with Toxoplasmosis among HIV positive patients at TASO Entebbe, Uganda
Detection of Toxoplasma gondii using Rapid Testing Kits and ELISA Toxoplasma IgG and IgM Kits at TASO Entebbe, Uganda
Symposium
SY03-4
Joseph Ndarubweine, Polly Rwandekeye, Josephine Lunkuse
Poster Presentation
19
Oral Presentation
Introduction
Toxoplasmosis is an infection caused by a single celled parasite Toxoplasma gondii whose definitive host is the cat. It is estimated
that 10 to 30% of HIV 1 infected patients are seropositive for anti toxoplasma antibodies and 25 to 50% patients experience symptomatic infections in the absence of antimicrobial prophylaxis (Porter, 1992).
The AIDS Support Organization Entebbe in Uganda cares for 6,200 HIV positive clients with 1 to 2 patients diagnosed with toxoplasmosis per clinic irrespective being on AntiRetroviral Therapy and Cotrimoxazole prophylaxis.
Objective
The objective of this study was to determine the seroprevalence of toxoplasmosis among HIV positive patients and the associated
risk factors at TASO Entebbe.
Methodology
The study employed a cross sectional design and random sampling method was used.
Results
The seroprevalence of Toxoplasmosis was 23.5% using Toxoplasma rapid IgG and IgM test kits. Seroprevalence of Toxoplasmosis
increased with age ranging from 18.9% in the 18 to 25 years age group to 28.3% in greater or equal to 45 years age group. Toxoplasma seroprevalence was higher in male at 30.5% than in female at 20.2%. Toxoplasma seroprevalence was higher among the fishermen at 53.8% compared to other occupations.The toxoplasma seroprevalence was higher among the Muslims at 35.7% compared
to other religions at 15.0% to 23.8%. The toxoplasma seroprevalence among patients who have been on ART for 3 to 6 months was
higher at 41.7%. The seroprevalence for toxoplasmosis was high among clients who have ever been treated for toxoplasmosis at
40.0%.
The results of this study were similar to studies conducted in South Africa at 26%, (Sonneberg, Silber and Jentsch, 1998), and 24.6%,
Milongo et al, 2000.
It is recommended that all newly diagnosed HIV patients be tested for Toxoplasma gondii antibodies and confirmed with a molecular
diagnostic technique.
Case Conference
Programs, TASO Uganda, Uganda Laboratory Technology Association (UMLTA), Uganda
Keynote Speech
SY04-1
Precollection and collection
– Not only analytical but also preanalytical –
Kazumasa Isobe
Laboratory Medicine, Institute of Clinical Medicine, Tsukuba University, Japan
Invited Lecture
Special Lecture
Educational Lecture
Introduction
Hospital laboratories often use basic techniques of immunology and biochemistry to automatically analyze clinical samples from
patients. These laboratory tests have been developed and improved to perform rapid and sensitive assays. Concurrently, systems of
quality assurance have also been developed in the analytical phase. However, approximately 50 to 75% of the errors identified in the
average clinical laboratory’s total testing process occur in the preanalytical phase, outside the walls of clinical laboratory. Indeed, in
our laboratory (2009-2014), 59 to 71% of the quality control incidents identified occurred outside of the clinical laboratory.
Precollection phase
A variety of factors can influence laboratory determinations, such as sex, age, diurnal variation, exercise, fasting, diet, alcohol intake,
tobacco and drug use, posture and also genetic variations. As exempla of the influence of such factors, we will present data concerning laboratory tests concerning growth hormone and testosterone, which have sex difference and are age-dependent, and interindividual variability in carotenoid levels caused by genetic variations.
Collection phase
Issues that can occur in the collection phase include inappropriate test request, order entry errors, specimen misidentification, and
sample collection. Hemolysis is the most common issue to arise during sample collection for laboratory tests. In our hospital, hemolysis occurred in 8.6% of the total specimens. Of the 80 different laboratory tests we examined in this study, 11 tests showed increased
results and 7 showed decreased results due to apparent hemolysis.
Another issue is blood glucose levels, which decrease by 10 mg/dL over a 2-hour period despite the addition of an antiglycolytic
agent. This can be problematic when the specimen transportation time is prolonged, leading to errors in the laboratory test results.
Conclusion
A variety of factors can influence laboratory determinations. In order to address preanalytical problems, we need to investigate the
cause and effects of errors in laboratory testing, disseminate the information, and educate hospital staff involved with laboratory
testing.
Symposium
SY04-2
Applied ethics for Biomedical Laboratory Scientists – Use of ethical
reflection models as a systematic approach to ethical dilemmas –
Marie N Roald
Case Conference
NITO The Norwegian Institute of Biomedical Science, Oslo, Norway
Oral Presentation
Poster Presentation
Introduction
The international code of ethics for Biomedical Laboratory Scientists (BLS) was adopted in 1992, and revised in 2010 by the International Federation for Biomedical Laboratory Science (IFBLS). But a code of ethics on its own and knowledge of ethical theory does
not solve ethical problems: Constant work on values, ethical reflection and development of ethical awareness is required to maintain
an ethically sound profession.
Method
Ethical reflection models can be used as a systematic approach to ethical dilemmas. To help make ethical reflection accessible we
have developed a program for Biomedical Laboratory Scientists, where we combine theory and practical exercises in ethical reflection. We describe ethical dilemmas that BLS’ meets in their daily work situation, for use in group discussions, and use an adapted
ethical reflection model to discuss dilemmas.
The elements in the model include exploration of the situation and action options, clarifying who is involved in the dilemma, disclosure of values and principles involved, determining the consequences, and then selecting action. A reflection model is meant to
assist in the process of uncovering what a dilemma or problem consists of, evaluate alternatives and arrive at a possible solution.
Participants in our ethical reflection groups have reported that they found it useful and stimulating, as well as challenging, to discuss
relevant ethical dilemmas with a systematic approach.
Conclusion
Discussing ethical dilemmas using ethical reflection models gives Biomedical Laboratory Scientists the opportunity to discuss professional ethical dilemmas in a systematic, structured manner. The method can be used to discuss own and others’ dilemmas, reflect on
choices and values, and develop communication skills and perceptive abilities.
20
Specimen storage and transportation – Circumstance surrounding preanalytical quality assurance at a Japanese clinical testing company. –
Mie Nagatsuka
Satellite Laboratory Planning and Coordination Center, Clinical Laboratory Business Segment , LSI Medience corporation, Japan
Invited Lecture
Special Lecture
The pre-analytical quality assurance is very important for a series of processes from specimen sampling to result report of the clinical testing. We cannot avoid “transportation and storage” of the specimen between the processes. As the way to maintain the quality
of the specimen required for a test, I would like to introduce our process at LSI Medience which is based on ISO15189.
Our company has 67 branch offices in Japan, and transports the specimen to the central laboratory in Tokyo, where we do testing.
For transportation, we use three different purpose-built boxes of three different temperature zones, frozen, refrigerated, and room
temperature (or ambient). The specimens are stored following detailed instruction to protect them from vibration and shock during
transportation. We are currently developing a database that will accumulate temperature data of the specimen boxes during transportation as an approach to a new management system.
We use air or land transportation depending on the area. The specimens are brought in to the central laboratory from the whole
country before the next morning, and we release the test result report the next morning. Safe and superior transportation and highly
reliable logistic networks in Japan makes it possible to transport within the time limit while maintaining the storage quality.
The specimens brought in to our laboratory, are managed by an ID system. Specimen type, storage condition, and aliquot stage are
distinguished by the bar code. Storage conditions are maintained and the specimens are managed for a certain period so it will be
available for additional tests after testing.
In order to guarantee accuracy for pre-analytical process, the systemization of management will continue to evolve.
Keynote Speech
SY04-3
Educational Lecture
Real Time Reaction Trace system on JCA-BM Analyzers
Symposium
SY05-1
Nau Ishimine1, Kenji Kawasaki1, Yoko Usami1, Kazuhiro Nagata1, Mitsutoshi Sugano1, Takayuki Honda2
Poster Presentation
21
Oral Presentation
Objectives
In clinical chemistry testing, accuracy and precision of test results have been dramatically improved by technological innovation and automation of analyzers. Such analyzers can also process very high volume of samples, achieving
a much higher throughput. At the same time, abnormal reaction time-course may yield abnormal data, therefore, it is important to monitor reaction time-course to avoid false result reporting. However, it is not possible to monitor
every reaction time-course for all samples manually. To overcome this limitation, automated detection of abnormal reaction time-course has been created using the conventional functions of the analyzer.
In this session, I will present the three functions utilized in the “Active Trace System” on JCA-BM Analyzers presented by JEOL Ltd, and an example case of each function.
I. Sub-Analytical Conditions
When sub-analytical conditions are defined, the analyzer can simultaneously calculate data from one measurement with three different analytical conditions. While the main analytical condition monitors the main reaction of a test,
the second and the third analytical conditions can detect abnormal reactions, such as abnormal turbidity after mixing reagent and sample.
Case 1: A male patient in his fifties
The attending physician ordered immunoglobulin quantitation in addition to a standard chemistry testing. The results calculated by the analyzer were TP: 106 g/L, Alb: 30 g/L, IgG: 702 mg/dL, IgA: -76 mg/dL and IgM: >700 mg/dL.
Having detected abnormality in IgA and IgM reaction curves using the Active Trace System, it was confirmed that abnormal turbidity occurred after R1 was dispensed into the sample. The results of diluted samples were IgA: 176
mg/dL and IgM: 5,504 mg/dL. Monoclonal IgM was the cause of abnormal reaction in this case.
II. Variance and Limit Value Settings
The Active Trace System monitors the equilibrium state of end-point assays and reaction constancy of rate assays.
Case 2: Five-year-old female patient
The attending surgeon requested a standard chemistry testing before the surgery. TP showed an abnormally high value, 100 g/L and the Active Trace System detected an abnormal reaction time-course. The result of TP calculated
from a dilution sample was 72 g/L. Monoclonal immunoglobulins and RF were not observed. The cause of this abnormality is not completely determined, however, LpX is considered involved in the abnormal reaction in this case.
III. Variance Allowance in R1 and R2 Reaction Periods
The Active Trace System monitors the reaction constancy for R1 period and R2 period.
Case 3: Variance error occurred to one analyzer
Variance errors occurred only on Unit No.2 of JCA-BM6070 analyzer. Indicated by the Active Trace System, air bubbles or insufficient mixing was suspected to be the cause of the errors. After replacing the mixing rods for the reaction cuvettes, the error stopped occurring.
Conclusion
The demand for higher quantity and quality is rapidly growing in clinical chemistry testing. Having real-time processing and reaction time-course check systems adds great value to the laboratory systems in detection of abnormal
data and prevention of false result reporting.
Case Conference
1 Department of Laboratory Medicine, Shinshu University Hospital, Japan
2 Department of Laboratory Medicine, Shinshu University school of Medicine
Keynote Speech
SY05-2
Reliability assurance of individual measurements and detection of any
unusual reactions based on the Reaction Curve Fitting Method
Yoshikazu Yamamoto1, Shuji Matsuo1, Noriko Hatanaka1, Kumiko Kamihara2, Tomonori Mimura2, Akihiro Iguchi2
1 Tenri Health Care University, Japan
2 Hitachi High-Technologies Corporation
Invited Lecture
Special Lecture
Educational Lecture
Typical reaction checking performed using an automated biochemical analyzer can potentially encounter many samples for which the
failure to detect unusual reactions occurs due to the effects of sample turbidity, sample composition, and/or the presence of medications.
In order to eliminate such problems, a revolutionary approach known as the Reaction Curve Fitting Method has been developed. This
method fits specific reaction curves for individual samples to approximate numerical functions, to analyze whether the reaction processes in individual samples have been completed normally, thereby ensuring measurement reliability.The Reaction Curve Fitting Method is a
technique for approximating a reaction curve obtained from measurements by a model function derived from chemical kinetics, depending on reaction patterns characterized by each item, which thereby provides the numeric indicators.For the end-point assay and rate
assay, model functions A and B are applied, respectively:Model function A: ABS = A0 + A1(1-e-kt), andModel function B: ABS = pt + q + s/
(t + r).(ABS: absorbance, A0: initial absorbance, A1: change in absorbance, e: Napier’s constant, k: rate constant, t: time, p: slope, q: initial
absorbance, s, r: coefficients.)The numerically-expressed characteristics of the pattern of the reaction curve are called index factors, and
the terms A1, A0, k , and Err (the total sum of squares of the difference between the estimated approximate curve and the measured reaction curve) are defined from model function A, while terms p, q, D0 (value of the lag phase), Tl (time on lag phase), and Err are defined
from model function B.To verify the ability of this method, trials for the detection of any unusual reactions based on measured reaction
curves were conducted using the MiRuDa (developed by Hitachi High-Technologies Corporation), a software program to apply the Reaction Curve Fitting Method.In this symposium, the successful detection of unusual reactions caused by variable factors in the sample composition and analyzer, based on the Reaction Curve Fitting Method, are presented.Successful detection of unusual reactionsUnusual reaction caused by a reagent: In the T-CHO reagent, the fact that reduced concentrations in the reagent composition decrease reaction rates
was quantitatively detected based on the values of k (verification experiment).Unusual reaction caused by sample composition: In the
measurement of Fe, the nonspecific reaction of Fesin (Saccharated Ferric Oxide) remaining in the blood was detected based on the value
of k .Unusual reaction caused by the automatic analyzer: In the measurement of UN, reduced absorbance took place due to water inflow
into the reaction cell during the reaction, which resulted from trouble in the cell cleaning mechanism, and lead to the multiple detection
of deviation in the data for the q -value;DiscussionThis method features the ability to detect any unusual reaction caused by sample composition or otherwise derived from the reagents/analyzer, which has thus far eluded detection. It is evident that the capability to identify
specific reaction curves for individual samples will create a new type of quality assurance to ensure the reliability of measurements.
Symposium
SY05-3
Pitfalls in Utilization of HbA1c in SEA patients
Nilrat Wannasilp
Case Conference
Department of Clinical Pathology, Faculty of Medicine, Siriraj Hospital, Mahidol University, Thailand
Oral Presentation
Poster Presentation
Determination of glycohemoglobin (gHb) has been recommended by several clinical studies for diagnosis, and monitoring treatment
in patients with diabetes mellitus (DM). At present, there are several methods used worldwide that can be classified into two groups
based on differences in charge and structure. The first group includes High Performance Liquid Chromatography (HPLC), electrophoresis, isoelectric focusing, and IFCC reference methods, and the later are immunoassay, Boronate Affinity Chromatography (BAC), and
enzymatic assay.
More than 700 hemoglobin (Hb) variants have been reported and about half of these variants are clinically silent. This could be an
issue when using HbA1c to assess the glycemic control in diabetic patients with Hb variants, especially in South East Asia where incidence of Hb variants is very high such as HbE and HbCS. Although Α -thalassemia is present throughout Southeast Asia, its distribution is heterogeneous. The frequencies are higher in the northern than in the southern part of the region (30.6% in Chiengmai Province of Thailand and 0.5% in Indonesia, respectively), and in Laos than in Khon Kaen Province in Northeastern Thailand (42.8% and
5.5%, respectively). Β -Thalassemia in northern Thailand has an incidence of about 5-8%, in Laos of 3-9%, in Indonesia of 6-l0%, and
in Myanmar of 4.3%. The prevalence of HbE is about 50-60% at the junction with Thailand, Laos and Cambodia, and HbCS is about
1-8%.
In HPLC methods, HbE is co-elutes with HbA, whereas the HbE1c is resolved from HbA1c and this result produces either underestimation of HbA1c or no peak of HbA1c in some instruments. In some analyzers, HbE may elute as shoulder of HbA1c resulting in overestimation of HbA1c. In our pilot study, no peak of HbA1c was observed in the presence of homozygous HbE.
In other methods such as immunoassay, there is no indication of possible interferences unless result is out of clinical range. It is
therefore important for laboratories to be aware of Hb variants interferences on their method. By using HPLC, the availability of
chromatograms provides an added advantage to detect the presence of Hb variants, along with excellent precision.
22
Reflections on clinical education about pre-transfusion testing
Mitsuko Maruyama1, Koji Yamamoto2, Makoto Okuda3
1 Division of Blood Transfusion, Mie University Hospital, Japan
2 Saiseikai Matsusaka General Hospital
3 Toho University Omori Medical Center
Educational Lecture
Symposium
The Frequency of Unexpected Antibodies at the Seoul National University Hospital in Recent 3 Years
The Frequency and case of Unexpected Antibodies at the SNUH during recent 3 Years (March 2012-February 2015)
Special Lecture
SY06-2
Invited Lecture
It is important to prevent transfusion errors and transfusion reaction to ensure safety of blood transfusion, and pre-transfusion testing is essential. Medical Technologist has an important role in blood transfusion safety.
Request in clinical practice is depend on the situation such as the degree of urgency, human resources as adequate or number of
medical technologist, and environment as equipment or computer system. Thus, medical technologist an ability to adapt to specific
or case-by-case situation.Therefore, the clinical education about pre-transfusion testing should be set a goal to enhance the adaptability to various clinical situation as well as knowledge and technical capabilities. It is important to acquire medical knowledge other
than transfusion medicine and hands-on expertise like know-how to achieve a goal. Moreover, it is also important to remember the
necessity of building a trust between medical staffs.
Mie prefecture association of medical technologists organized, three practical traning seminars.
A)Delivery practical lecture: Specialist in Blood banking (a certified medical technologist for transfusion service) visited the laboratory of small and medium-scale hospital. They suggested the best way to acquire practical skills appropriate to the environment of
laboratory through practical training.
B)Practical training seminar using fictitious scenario: Participants were segmented into small groups and practiced based on fictitious scenario. Participants tried the communication skills, presentation skills and flexibility required in performing a procedure for
transfusion.
C)Practical training seminar aimed at fostering of trainer in pre-transfusion testing: This seminar was held at the initiative of Society
(Co-sponsored Japan association of transfusion and cell therapy and Japanese association of medical technologists).
In this symposium, I will address the details and verification based on questionnaire results of practical training seminars besides
future challenges in clinical education about pre-transfusion testing.
Keynote Speech
SY06-1
Ji-sang Kang1, Young-kuk Kwak1, Kyou-sup Han2
(Methods)
Unexpected antibodies tests performed in a Seoul National University Hospital during recent 3 Years (March 2012-February 2015)
were analysed : 207,247 tests for screening and 3,766 tests for identification. Antibodies were screened and identified parallely using the Autovue innova Autoanalyser Methods(Ortho-Clinical Diagastics, USA), LISS/Coombs Gel card methods (DiaMeD, Switzerland)
and Manual Tube Methods (Ortho-Clinical Diagastics. USA)
Oral Presentation
(Results)
A total of 207,247 (203,674 tests in SNUH and 3,533 tests referred from another Hospital) samples were screened for the presence
of unexpected antibodies. Antibody screening was positive in 3,766 (645 tests in SNUH 0.32% Positive and 3,121 tests referred from
another Hospital 88.3% Positive) cases. Among them, Unexpected antibodies were identified in 3,766 cases. The most frequently detected antibody was anti-E in 704 cases (18.7%), followed by anti-Lea 349 cases (9.2%), anti-E+c 265 cases (6.5%), anti-M 203cases
(5.4%), anti-P1 117 cases (3.1%) and warm auto Ab 63 cases (1.6%) etc. Other unusal Unexpected antibodies detected anti-Dia 14
cases (0.4%), anti-Xga 7 cases (0.2%), anti-Jra 26 cases (0.7%), anti-Lua 2 case(0.05%) and anti-HTLA 2 cases (0.05%) etc.
Poster Presentation
(Background) :
Unexpected antibodies test are very important pre-transfusion tests for preventing transfusion reactions. Patients with Unexpected
blood antibodies may be at increased risk for delayed transfusion reactions. For these reasons, it is essential to know the frequency
of unexpected antibodies among South Koreans for prompt and safe transfusion. This study aimed at frequency and case of Unexpected Antibodies, problem that should be solved for safe blood banks
Case Conference
1 Laboratory Medicine, Seoul National University Hospital, Korea
2 Laboratory Medicine, Seoul National University College
(Discussion)
A higher rate of alloimmunization was observed in females 2,917 cases(69.7%) than in males 1,267 cases(30.3%). This study demonstrated that the frequency of Unexpected antibodies was not different from that of references in South Korea (0.3%-1.73%). Unexpected antibodies were associated with multiple transfusion. Therefore, antibodies screening and identification test are crinical steps
in pre-transfusion tests
23
Keynote Speech
SY06-3
Isoagglutinin Titer for ABO-Incompatible Living Donor Liver
Transplantation
SUK WON SEO, HOI JOO YANG, SEOG WOON KWON
Asan Medical Center, Dept of Laboratory Medicine, Korea
Invited Lecture
Special Lecture
Educational Lecture
Background: ABO-incompatible living donor liver transplantation (ABO-i LDLT) has emerged to be an effective means to deal with the
shortage of organ donors. Reduction of pre- and post-operative isoagglutinin titer is important for preventing the antibody-mediated
humoral rejection, which could possibly lead to graft dysfunction. Effective administration of rituximab and plasmapheresis reduces
isoagglutinin titer and improves the clinical outcome of ABO-i LDLT. In this study, we analyzed the clinical significance of isoagglutinin titer .
Methods: A total of 69 patients who underwent ABO-i LDLT were analyzed. Preoperative IgM isoagglutinin titer was reduced to 1:8
or less by administration of anti-CD20 monoclonal antibody (rituximab) and performing plasmaphereis. For all patients, isoagglutinin
titers were measured at the time of admission, before and after each plasmapheresis, before transplantation, and at postoperative
follow-up days. Anti-A and anti-B IgM Isoagglutinin titers were determined using standardized tube test: incubating 0.1mL of type A
or B red blood cell suspensions in 0.1mL of saline containing two-fold serial dilution of patient serum at room temperature, followed
by centrifugation. The last dilution showing trace reactivity was defined as anti-A or anti-B isoagglutinin titer in each assay. For IgG
assays, anti-human globulin was added
Results: The graft survival was 97.1% for a median follow-up of 474 days (range, 301-701). Preoperatively, IgM isoagglutinin titer
was reduced to 1:8 or less Sixty eight patients (98.6%) are still alive without allograft dysfunction. One patient died after re-LT and
the other patient developed AMR. Patients’ blood groups had no significant effect in the clinical outcomes of ABO-i LDLT.
Conclusions: Rituximab and plasmapheresis reduced isoagglutinin titer and improved the clinical outcome of ABO-i LDLT. By reducing pre-LT IgM isoagglutinin titers to 1:8 or less, 1-year graft survival was excellent. Isoagglutinin titer is important for preventing
the antibody-mediated rejection in ABO-i LDLT.
Symposium
SY06-4
Blood safety and pretransfusion testing
Shyh-Chyi Lo
Case Conference
Department of Laboratory Medicine, National Taiwan University Hospital, Taipei, Taiwan
Oral Presentation
1) Employment of the column agglutination technology (CAT): CAT can improve pretransfusion testing safety and efficiencies. Its
value can be best demonstrated in cases of ABO discrepancy associated with mixed field reaction. One example is B3 blood type, the
most prevalent ABO subtype in Taiwan. Cells from patient of B3 blood type show a mixed-field reaction when testing against anti-B
sera. This could be easily misinterpreted by traditional tube method. Another example is from cases of ABO incompatible hematopoietic cell transplantation.
Pretransfusion testing, including ABO/Rh(D) typing, antibody screen, and cross-matching, must be performed prior to release of RBC
components. Pre-transfusion testing is an essential element of the entire transfusion process to enhance vein-to-vein safety. Continual improvement of safety of pretransfusion testing is the top priority of every transfusion service laboratory. During the last decade,
several improvements have been undertaken to maximize blood safety. We share some of our experiences:
Poster Presentation
2) Automation of the pre-transfusion testing: Human error associated with manual pretransfusion testing is a major cause of
transfusion related morbidity and mortality. Most of the human errors can be effectively reduced by automated systems. Automation
can reduce the number of process steps and cut back on the steps that require human intervention. Documentation processing, data
tracking, and security all can be improved.
3) Transfusion information technology (IT) systems: A validated IT system in hospital transfusion laboratories can help to improve blood safety. One key function is the verifying mechanism that doesn’t allow selection of ABO incompatible blood products.
However, the rule of selection for blood type compatibility should be modified for some clinical situations such as in patient undergoing ABO incompatible hematopoietic cell transplantation. We worked with IT system department to implement a special algorithm
to better manage the transfusion needs of patients undergoing ABO incompatible hematopoietic cell transplantation and recipients
of ABO incompatible kidney transplant.
24
¨What Is¨ and ¨How to Make¨Leader of Biomedical Laboratory Scientist?
Ryunosuke Ohkawa, Minoru Tozuka
Tokyo Medical and Dental University, Japan
Keynote Speech
SY07-1
Prologue
Japanese system in Medical Laboratory
What is ideal leader of BLS?
How to make each leader of BLS?
1. To acquire the ability to discover something and create new things, repeated practice of analyzing data and thinking logically is important. Regarding this, conducting research is the most appropriate
way to reach the goals. More students and BLS getting not only bachelor, but also master or doctoral degree are expected.
2. In Japan, even university, there is predominately passive education in class so that many students and BLS lack proactivity. To develop communication skills, project-based learning should be introduced. After graduation, senior BLS should encourage young BLS to attend congress and to practice giving their presentations and throwing many questions at such events.
Educational Lecture
1. Ameliorator and Pioneer
In routine laboratory test, there are many errors or problem buried in plenty of laboratory testing. Like an eagle catching fishes in a river, BLS is expected to find the error, analyze the data, determine
the cause and finally improve the testing or create the protocol to avoid making the error data.
2. ¨Clinical BLS¨
In Japan, formerly, BLS was not required to employ much communication skill. They were called, ¨powerful person in the background¨ meaning unsung hero. However, BLS having the skill more and
discussing about data with medical staffs as joining in medical team--infection control team, nutrition support team etc.--needs to be produced.
Special Lecture
In Japan, there is one type of job which is related to conducting laboratory tests professionally: Biomedical Laboratory Scientist (BLS). After students complete a three years program at technical school
or four years program at university, which includes clinical rotation within six months, students can take the paper exam of national certification of BLS. When they pass the exam, they can start to work
at a hospital and associated institutions. The work of BLS covers various fields of testing: clinical chemistry, hematology, immunology, microbiology, electro physiology, ultrasonography, transfusion
medicine and pathology.
Here, nowadays medical field is undergoing continuous changes, due to low birth rate, longevity, swelling medical expenses and so on. In the field of laboratory medicine, efficiency of testing without
any degradation of quality and staff downsizing would be required along with accelerating of automation. To correspond these changes, BLS must open the new door and educators including expert BLS
must make new generation of BLS who become leader heading other BLS.
Invited Lecture
Students sometime ask me, ¨The automation of clinical laboratory is accelerating in development. Will the job of medical technologists no longer be required in the future?¨ Actually, researchers from
the Oxford University reported 47 percent of total US employment including ¨Medical and Clinical Laboratory Technologists¨ will be at risk in regards to the probability of computerization in the
future. Is the automation beneficial for us? or is it a threat? Will Laboratory Technologist become a profession of the past? What action should we take towards preventing this?
I answer to students, ¨ Leaders of BLS including you would rather welcome the Automation and open new era.¨
Options for Delivering Biomedical Scientist Education and Training
Symposium
SY07-2
Alan Wainwright
Oral Presentation
Biomedical scientists are required to be autonomous, independent practitioners with responsibility to: manage their own workload;
plan routine scientific investigations; apply and maintain quality control and quality assurance processes; perform and validate a discrete range of manual, semi-automatic and automatic techniques and investigations; interpret results recognising the clinical significance of the outcome of discrete investigations and provide clinical, scientific and service advice. In addition they are expected to be
a flexible workforce capable of addressing future challenges in service delivery, and should aspire to give excellence in training that
directly links to improvements in patients´ outcomes.
This presentation will look at options for delivering fit for purpose education and training at all levels from support staff through
to consultant biomedical scientists. It will consider this in the context of the applied academic knowledge, vocational skills, and the
universal standards that are unique to our profession yet not always safeguarded by statutory regulation and career development.
Case Conference
Institute of Biomedical Science, United Kingdom
Poster Presentation
25
Marie Culliton1,2
Invited Lecture
Educating Biomedical Scientists for the Future.
Keynote Speech
SY07-3
Biomedical Scientists are a young and constantly adapting profession. Over the past 50 decades, or more, we have developed from
an assistant role to a profession in our own right. This has led to formal qualifications of Bachelors, Masters and Doctorate Degrees
and in some cases the attainment of professional qualifications previously reserved for Medical Doctors such as FRCPath (Fellow of
the Royal College of Pathologists).
The profession must continue to adapt to new and changing political, economic social and technological challenges if it is to survive
and take its rightful place as “Diagnostic Partners in Healthcare”.
This presentation will consider what education: undergraduate, postgraduate, formal, informal and continuous professional development is required to keep this profession dynamic and relevant in a rapidly changing environment. What knowledge, skills and competencies do we need to give our current scientists to equip them to evolve and reach their potential?
1 European Association for Professions in Biomedical Sciences, Brussels, Belgium
2 The Academy of Clinical Science and Laboratory Medicine, Dublin, Ireland
Special Lecture
Educational Lecture
Symposium
SY08-1
Improving Diagnoses--The Role of the Biomedical Laboratory Scientist
Catherine Otto
Case Conference
Shoreline Community College, United States
Oral Presentation
Poster Presentation
For the last sixteen years, healthcare delivery systems have focused upon improving their quality and patient safety as a result of
key reports published by the National Academies of Sciences, Engineering and Medicine beginning in 2000: To Err is Human, Crossing the Quality Chasm and Health Professions Education--A Bridge to Quality. From these reports we have adopted the definition of
quality as healthcare that is safe, effective, efficient, timely, patient-centered and equitable. These patient safety quality aims provide
the framework for evaluating and improving healthcare delivery processes. Future healthcare practitioners, including biomedical
laboratory scientists need specific competencies to improve healthcare delivery and patient safety. To deliver healthcare that meets
the quality aims, practitioners need to work in interprofessional teams, practice evidence-based medicine, deliver patient-centered
care, use information technology and focus upon quality improvement. Biomedical laboratory scientists have slowly adopted patient
safety quality aims and healthcare practitioners competencies into our curricula and practice, however as a result of the latest report, Improving Diagnosis in Health Care there is a greater urgency to become more involved in the diagnostic process. This latest
report from the National Academies identified eight recommendations for practitioners, healthcare delivery systems and regulators
to implement in order to decrease the harm that patients experience as a result of diagnostic errors. The key recommendation that
biomedical laboratory scientists need to focus upon is the first goal: “Facilitate more effective teamwork in the diagnostic process
among health care professionals, patients, and their families”. Working together with clinicians and other healthcare practitioners
will provide biomedical laboratory scientists the opportunity to develop effective laboratory test ordering and test interpretation
methodologies. Case studies will be used to demonstrate the application of the healthcare competencies to the practice of medical
laboratory science to reduce diagnostic errors.
26
Infection Control and Biomedical Laboratory Scientists in Japan
Shigeki Misawa
Department of Clinical Laboratory Medicine, Juntendo University Hospital, Japan
Educational Lecture
Symposium
Management for Pathology and Cytology Laboratory
―From the View point of Patient first―
Special Lecture
SY08-3
Invited Lecture
Infection control is one of the most important components of patient safety in hospitals. The clinical laboratory department plays an extremely important role in the prevention of hospital
infections. Biomedical laboratory scientists (BLS) in the microbiology laboratory can early detect pathogens and outbreaks of infectious diseases in the hospital. Therefore, microbiology laboratory must make efforts to provide information regarding the control of hospital infection.
In this symposium, I would like to introduce the activities of the microbiology laboratory aiming at hospital infection control in Japan.
Laboratory information for empiric therapy
The microbiology laboratory routinely provides laboratory information to help the initial treatment of diseases. Results of the smear examination of clinical specimens are rapidly reported,
particularly in cases of severe infections such as sepsis and bacterial meningitis. In cases where the blood culture is positive for microorganisms, smears containing the suspected microorganisms are immediately analyzed using the Gram stain morphology. In specimens with a turbid cerebrospinal fluid (CSF), Gram stain results are required to be ready within 1 h of specimen
reception. Specimens of positive blood culture or direct CSF smears are also cultured and tested for drug susceptibility, and an interim report is required to be ready within a day.
Definition and detection of alert organisms
Important microorganisms related to hospital infection control are defined as alert organisms. Alert organisms include the following: the Mycobacterium tuberculosis complex, Mycoplasma
pneumoniae , Shigella spp., Salmonella spp., Escherichia coli O157, respiratory and enteropathogenic viruses, and drug-resistant organisms such as MRSA, ESBL-producing organisms, carbapenem-resistant Enterobacteriaceae , and Clostridium difficile . If an alert organism is detected during routine microbiology, it is reported to the physicians and infection control division.
Analysis and provision of surveillance data
Surveillance data are regularly collected in the microbiology laboratory; these include blood culture test surveillance, target surveillance, and antimicrobial susceptibility surveillance. Blood
culture surveillance includes number of cultures per 1,000 patient-days, two culture set submission rate, positive rate, contamination rate, and isolates, which are monitored monthly. Target
surveillance monitors alert organisms mentioned above. In some hospitals, active surveillance culture for MRSA is performed on admission and is analyzed in relation to the consumption of
alcohol hand rubs. The screening test for ESBL-producing organisms in fecal specimens is also performed in a few hospitals. For antimicrobial susceptibility surveillance, cumulative antimicrobial susceptibility is accessed with an antibiogram of clinically important microorganisms and the trend in resistance at the local level is monitored. Data are analyzed in association with
antimicrobial usage in hospitals.
Participation in ICT activities
In Japan, ICT has been organized according to the following four occupations: doctors, nurses, BLS, and pharmacists. ICT is in charge of executing measures of infection control in hospitals.
ICT activities also involve wards and the outpatients department, investigation at the time of outbreak, and evaluation after the execution of each measure. The microbiology laboratory contributes to infection control by performing these actions.
I would like present and discuss the effects of these activities in the symposium.
Keynote Speech
SY08-2
Kyoko Komatsu1,2
Poster Presentation
27
Oral Presentation
In recent years, hospital laboratories are required to manage of patient’s safety, and quality management., and occupational health
management. The patient’s safety will be able to discuss as QC and RM and Infection control.
Quality control, or QC for short, is a process by which entities review the quality of all factors involved in production. ISO 9000
defines quality control as “A part of quality management focused on fulfilling quality requirements”. A peculiarity is the service for
people in the medical world is “the service is for life”. The tools for QC in the hospital are CPC (Clinico-Pathologic Conference), participating internal and external control, EBM(Evidence Based Medicine), and so on. Recently, Japanese government announced guideline of clinical trial. They suggested that the Lab data should be proved trustworthy for carrying out clinical trial. ISO 15189 or CAP
will accept as a good control management Laboratory. Now, JAMT created their own certification system for Lab data. It is good for
especially small hospitals. I will introduce the QC system in Japan.
Risk management (RM) : Increased requests from patients of medical accidents, hospital are reqired “Risk management (RM)”. RM
is the identification, assessment, and prioritization of risks followed by coordinated and economical application of resources to minimize, monitor, and control the probability and/or impact of unfortunate events or to maximize the realization of opportunities. Accident in pathology and cytology laboratory will give a severe damage to the patients. We analyze failures at any phase in design and
working system for avoiding human error.
Infection Control : Infection control addresses factors related to the spread of infections within the healthcare setting (whether
patient-to-patient, from patients to staff and from staff to patients, or among-staff), including prevention (via hand hygiene/hand
washing, cleaning/disinfection/sterilization, vaccination, surveillance), monitoring/investigation of demonstrated or suspected
spread of infection within a particular health-care setting (surveillance and outbreak investigation), and management (interruption
of outbreaks). It is on this basis that the common title being adopted within health care is “infection prevention and control.” Infection control teams (ICT) is the Group of specialists responsible, and Government support the system of ICT. I will introduce qualified
system as a specialist for infectious control staff which recognized by scientific organizations.
Occupational health management : Three occupational health management are occupational hygiene, health management for
employee, and work management. We use many kinds of toxic reagent such as formalin, xylene, methanol, and so on, in Pathology
Laboratory. Japanese government enacts the Industrial Safety and Health Law, and Measurement Law Working Environment Measurement Law for protecting labor’s health. I will introduce how our working place has developed from this point of view.
Case Conference
1 PAST President of IFBLS
2 Cancer Institute Hospital of JFCR, Japan
Keynote Speech
SY09-1
Detection of carbapenem-resistant Enterobacteriaceae and issues to
overcome
Sayoko Kawakami
Laboratory of Antimicrobial Agents and Resistance Department of Bacteriology 2, National institute of Infection Diseases, Japan
Invited Lecture
Special Lecture
Educational Lecture
1. CRE and CPE
Carbapenem-resistant Enterobacteriaceae (CRE) are increasing worldwide, so nosocomial infections and intractable infections have become a problem.
MIC breakpoints for CRE set by the European committee on Antimicrobial Susceptibility Testing (EUCAST) and the Clinical and Laboratory Standards Institute
(CLSI) are: imipenem (IPM) >8 micro g/mL, meropenem (MEPM) >8 micro g/mL, and doripenem >2 micro g/mL.
On the other hand, carbapenemase-producing Enterobacteriaceae (CPE) produce various carbapenemase, such as the IMP, NDM, VIM, KPC, and OXA type. Many
of the CPE is included in the CRE.There is also a strain with porin loss and AmpC-type beta-lactamase (AmpC) overproduction as CRE of non- CPE.
2. Classification of CRE
CRE can be classified by phenotype, MALDI-TOF MS, and genotype. Phenotyping can be done by the mercapto acetic acid disk method, Etest-EDTA method,
Carba NP test, Modified Hodge test, combination disk test (equivalent to EUCAST), and carbapenemase inactivation method(CIM). Genotyping is done by PCR/
real-time PCR or whole-genome sequencing.
3. Features and problems of these methods
The mercapto acetate disk method and Etest-EDTA method are simple tests for detection metallo-beta-lactamase (MBL) with high sensitivity and specificity,
and test results can be judged by the next day. The Carba NP test is simple and quick for detection of CPE, giving results in less than two hours, but sensitivity for the OXA-type is slightly lower. The modified Hodge test also detects CPE, but is cumbersome with low sensitivity for the NDM type and may show falsepositive results for the AmpC type. Results are available by the next day.The combination disk test assesses synergistic effects and can classify MBL, KPC, OXA,
and AmpC. It is simple and it shows high sensitivity and specificity for MBL and KPC. However, sensitivity for AmpC and specificity for OXA are slightly lower.
Results are available by the next day.
The CIM was introduced in 2015 for differentiation of CPE. An MEPM disk is immersed for 2 hours in a suspension of resistant bacteria and then placed on Mueller Hinton agar coated with E. coli ATCC25922. The diameter of the inhibitory zone is measured after 6 hours of incubation at 35 degrees. An inhibitory zone
is not formed in the presence of CPE. This method is simple and inexpensive, and the time required is about 8 hours.
In the MALTI-TOF MS CPE detection assay, to identify the peak corresponding to the IPM metabolites, after a 20 minutes incubation of the isolate with 0.5 mg/
mL IPM solution at 35 degrees.
PCR/real-time PCR for detection of CPE is a method based on multiplex PCR. It only requires about four hours, but is complicated and expensive, so use is limited to some laboratories. Whole-genome sequencing is another method which was recently developed. It provides an abundance of genetic information, such as
MLST analysis as well as data on drug resistance genes. In the future, it is expected to become more popular, but currently is only used in very few laboratories.
Symposium
SY09-2
Outer membrane protein C of Escherichia coli in carbapenem-resistance
and bacterial virulence
Jiunn-Jong Wu
Case Conference
Department of Biotechnology and Laboratory Science in Medicine Yang-Ming University, Taiwan
Oral Presentation
Carbapenems are often used to treat serious infections caused by multidrug-resistant strains of Enterobacteriaceae . The spread of
carbapenem resistance in Enterobacteriaceae may become a serious problem. In Escherichia coli , there are three major outer membrane proteins (OMPs), OmpA, OmpC, and OmpF, which function as passive diffusion channels for small molecules, like nutrients,
toxic salts, and antibiotics. The expression of OmpC and OmpF is regulated by osmolarity, and loss of OmpC or OmpF is related to
antibiotic resistance, particularly when combined with production of extended-spectrum β -lactamases and AmpC. The loss of OmpC
expression may be due to selection under antibiotic pressure. OmpA in E. coli is well known as a virulence determinant for increased
survival in macrophages, serum resistance, and invasion of brain microvascular endothelial cells. The deletion of ompC in E. coli isolated from Crohns disease was shown to decrease adherence and the ability to invade intestinal cells. OmpC is also characterized as a
lactoferrin binding protein. Recently, it has been shown that the loss of OmpK36 in Klebsiella pneumoniae leads to increased resistance to antibiotics and susceptibility to neutrophil phagocytosis. The aim of this talk will focus on characterize the role of OmpC in
carbapenem resistance and bacterial virulence in E. coli .
Poster Presentation
28
Trends of Carbapenem-Resistant Enterobacteriaceae and Its Detection
Techniques
Katsunori Yanagihara
Nagasaki University, Japan
Invited Lecture
Special Lecture
Educational Lecture
CRE, which stands for carbapenem-resistant Enterobacteriaceae, are a family of germs that are difficult to treat because they have
high levels of resistance to antibiotics. CRE have emerged as a global threat. Patients with CRE infections have adverse outcomes,
including high mortality. Although CRE are still relatively uncommon, the rate of carbapenem resistance among Enterobacteriaceae
is increasing. Surveillance data demonstrate a steady increase in the burden of disease from CRE, in particular carbapenem-resistant
Klebsiella pneumoniae, between 2000 and the present day. In 2013, the U.S. Centers for Disease Control and Prevention estimated
9,300 patients a year were infected with CRE, with an associated 610 deaths. Previous CRE outbreaks have occurred elsewhere in
many countries, including Japan.Despite the global emergence of CRE, no clear consensus has emerged in regard to the method of
detection. Because of early studies that showed that some CRE had carbapenem minimum inhibitory concentrations (MICs) in the
susceptible range, the Clinical and Laboratory Standards Institute (CLSI) recently lowered breakpoints for carbapenem antibiotics.
The CLSI requires that laboratories take additional steps to validate the lower breakpoints. However, many laboratories have not
adopted the new carbapenem breakpoints, potentially resulting in underestimation of CRE prevalence.Once carbapenem resistance
is identified through standard susceptibility testing, additional phenotypic tests can help to identify CRE. These include the modified
Hodge test (MHT), the Carba NP test and the carbapenem inactivation method (CIM). Another option is the use of matrix-assisted
laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) to detect carbapenemase activity. Like the MHT and
the Carba NP test, MALDI-TOF-based assays do not provide insight into which carbapenemase is present.Molecular assays for CRE
detection include PCR, LAMP (Loop-mediated isothermal amplification), microarrays, and whole-genome sequencing (WGS). These
methods have the benefit of determining the exact mechanism conferring carbapenem resistance, which can be especially helpful
during outbreak investigations. The primary limitation of molecular assays is that only known genes can be targeted; those encoding
novel carbapenemases will be missed with molecular approaches.
Keynote Speech
SY09-3
Symposium
Case Conference
Oral Presentation
Poster Presentation
29
Fumihiko Ookubo1, Yutaka Koga1,2, Yoshihiro Ohishi1,2, Yoshinao Oda1,2
Invited Lecture
Case presentation
Keynote Speech
CC01-1
Patients: Female in her sixties
Main complaint: Appetite loss
Clinical history
The patient complained of appetite loss. She was found to have a peri-ampullary submucosal tumor by gastrointestinal endoscopy at
an outside hospital. She was referred to our institute for further evaluation. Computed tomography (CT) showed an early enhancing
duodenal mass, measuring 17 × 22 mm. Endoscopic ultrasound (EUS) demonstrated a hypoechic mass mainly in the submucosal
layer, which was hypervascular with Doppler-mode. No pancreatic or biliary ductal dilatation was seen. EUS-guided fine needle aspiration was performed for definite diagnosis.
1 Division of Diagnostic Pathlogy, Kyushu University Hospital, Japan
2 Department of Anatomic Pathology, Pathological Sciences, Graduate School of Medical Sciences, Kyushu University, Japan
Special Lecture
Educational Lecture
Symposium
CC01-2
Cytology of uterine cervix
Junzo Fujiyama
Case Conference
Department of Cytology, Cancer Institute Hospital, Japan
Oral Presentation
Case : 70s year old women, para 1.
Clinical symptom was post-menopausal bleeding.
Physical examination revealed a cervical polypoid tumor.
Cervical cytologic and histologic specimen were taken from the cervical polypoid tumor and diagnosed as malignant tumor.
It was diagnosed as stage Ⅱ b of cervical cancer by MRI and pelvic examination. Then the operation was performed.
Poster Presentation
30
Fine needle aspiration cytology of lung mass
Hwa-jeong Ha, Jung-soon Kim, Seung-sook Lee, Jae-soo Koh
Department of Pathology, Korea Cancer Center Hospital, Korea
Invited Lecture
A 45-year old female patient complaining shoulder pain was transferred to our hospital due to abnormal chest CT findings. This
patient has a previous history of tuberculosis three years ago, which has been completely controlled. The chest CT showed airspace
consolidation in RUL with lymphadenopathy in right axillary,supraclavicular, internal mammary, and prevascular areas. Radiologic
differential diagnosis includes lymphoma, lung cancer, pneumonia, or Tb. FDG PET/CT scan revealed a large intense hypermetabolic
lesion on RUL and mild hypermetabolic lesions on RLL and LUL. There were multiple hypermetabolic lesions including thoracic and
intra-abdominal lymph nodes, abdominal wall and bone. The PET/CT scan study suggests lung cancer with multiple metastasis or
lung cancer with concurrent malignant lymphoma. The following slide was made from percutaneous fine needle aspiration procedure of the RUL pulmonary mass.
Keynote Speech
CC01-3
Special Lecture
Educational Lecture
Diagnostic pitfalls in breast fine-needle aspiration (FNA) cytology of
sclerosing adenosis
Department of Laboratory Medicine, National Taiwan University Hospital, Taiwan
Materials and Methods: From January 2006 to May 2016, we have identified 28 breast FNA cytology cases with histologic diagnosis of sclerosing adenosis at National Taiwan University Hospital. Among these cases, three were diagnosed as suspicious for carcinoma. The patient’s age were 40, 43 and 57. Lesion sizes were 0.8, 1.2 and 2.0 cm. The mammography findings were BI-RADS category
0, 4 and 4b.
Conclusions: Sclerosing adenosis may show clinical, radiologic and cytologic features that overlap with breast carcinoma. Although
FNA cytology has its diagnostic limitation, awareness of the cytologic features of sclerosing adenosis may help prevent overinterpretation. When encountered difficult cases, histologic confirmation are needed.
31
Poster Presentation
Results: After reviewing these three misdiagnosed cases, cytology smears showed crowded clusters of atypical cells with pleomorphic nuclei, which were ovoid, elongated or enlarged. Cell clusters were loosely arranged. Nuclear overlapping were frequently noted
within cell clusters. Few stromal fragments and naked nuclei in the background were also noted.
Oral Presentation
Introduction: Sclerosing adenosis is a benign and unusual breast lesion that can be clinically and mammographically confused with
breast carcinoma. Although core needle biopsy (CNB) has become the major diagnostic tool of breast tumors, FNA cytology remains
a cost-effective and relatively non-invasive procedure to evaluate indeterminate breast lesions. However, the cytologic smears of sclerosing adenosis may cause overinterpretation when discohesive clusters or individual cells accompanying with nuclear atypia are
present.
Case Conference
Chun-Ming Wang, Tsu-Yao Cheng, Ya-Ting Lee, Ping-Fung Chung, Min-Se Huang, Ming-Hsiang Weng,
I-Shiow Jan, Sow-Hsong Kuo
Symposium
CC01-4
Keynote Speech
CC02-1
Case 1
Aya Ogane
The University of Tokyo Hospital, Japan
Invited Lecture
Special Lecture
A 67·year·old woman suffering from primary sclerosing cholangitis (PSC) was referred to our ER due to a high fever of 38.5 °C. Two
weeks ago, she had taken percutaneous transhepatic biliary drainage (PTBD) and several prophylactic antibiotics had been given. At
the time of visit, her leukocyte count decreased to 1.7 × 109/L and neutrophil percentage was 0%. In addition, her C-reaction protein (CRP) increased to 15.83 mg/dL. Since these laboratory findings indicated she has had severe infection, she was admitted to our
hospital immediately.
Her past medical history included autoimmune hepatitis and hyperthyroidism at age 50, and steroid-induced diabetes at age 67. In
the previous admission for PTBD, she had itchy erythema over her entire body after the administration of a contrast agent and antibiotics, which might have been caused by an allergic reaction.
Hematological testing values on admission were as follows: white blood cell count, 1.7 × 109/L (differential: 0% neutrophils, 36.5%
eosinophils, 1.0% basophils, 14.0% monocytes, and 48.5% lymphocytes); red blood cell count, 3.44 × 1012/L; hemoglobin, 11.3 g/dL;
hematocrit, 33.7%; and platelet count, 283 × 109/L. Clinical chemistry findings showed lactate dehydrogenase of 218 U/L, AST(GOT)
77 U/L, ALT(GPT) 111 U/L, alkaline phosphatase 671 U/L, γ ·GT 1156 U/L, total bilirubin 331.7 μ mol/L(19.4 mg/dL), and CRP
158300 μ g/L(15.83 mg/dL). Three pathogens were detected in the bile culture: Pseudomonas aeruginosa (+), Stenotrophomonas
maltophilia (3+), and Enterococcus faecium (3+).
We immediately reported to the doctor that the patient had severe neutropenia.
Educational Lecture
Symposium
CC02-2
Case 2
Yuki Ohkawa
Case Conference
Okinawa Red Cross Hospital, Japan
Oral Presentation
History of Present Illness:A Japanese female in her 70s was referred to our hospital from a local clinic due to her anorexia and dull
headache lasting for a month. She was diagnosed with microcytic anemia with Hemoglobin (Hgb) 52 g/L (5.2g/dL) that required repeated blood transfusions. Iron deficiency was denied by further testing. Her endoscopic examinations of gastrointestinal tracts and
CT scan with contrast materials did not reveal any abnormal findings. Past Medical History: She has been a hepatitis B carrier. She
has had cervical spondylosis and osteoporosis. Past Surgical History: She had brain aneurysm that was treated with coil embolization. Family Medical History: None. Social Habits: She is a non-smoker and non-drinker. Laboratory Data: [Complete blood count
and Peripheral blood (PB) smear] RBC 3.02 × 10
12
Poster Presentation
/L(3.02 × 106/µL), Hgb 77g/L(7.7g/dL), Hct 0.24L/L(24.0%), MCV 78.1fL, MCH 25.5pg, MCHC 326g/L(32.6g/dL), Reticulocyte
0.5%, Plt 75 × 109/L(75 × 103/µL), WBC 2.6 × 109/L(2.6 × 103/µL), Neutrophil 65.5%, Lymphocyte 22.5%, Monocyte 10.5%,
Eosinophil 0.5%, Basophil 1.0%.PB smear showed anisocytosis, elliptocytes, acanthocytes, schizocytes, large platelets and platelets
with hypo granulation. [Coagulation]Prothrombin time (PT) 14.1s, PT-INR 1.20, activated partial thromboplastin time (APTT) 31.5s,
fibrinogen 2.44 g/L(244mg/dL), D-dimer 400 µg/L(0.4µg/mL), FDP 1500µg/L(1.5µg/mL) [Chemistry panels] Alb 46g/L(4.6g/dL), TBil 34.2µmol/L(2.0mg/dL), AST(GOT) 55U/L, ALT(GPT) 61U/L, LD 223U/L, ALP 228U/L, UA 244µmol/L(4.1mg/dL), UN 7.5mmol/
L(20.9mg/dL), CRE 71µmol/L(0.80mg/dL), Na 138 mmol/L, K 4.0 mmol/L, Ca 2.25 mmol/L(9.0mg/dL), CRP 500µg/L(0.05mg/dL),
Fe 38.9µmol/L(217µg/dL), TIBC 41.4µmol/L(231µg/dL), UIBC 2.5 µmol/L(14µg/dL), Ferritin 771pmol/L(343ng/mL) [Serology]IgG
14.80g/L(1480mg/dL), IgA 1.91g/L(191mg/dL), IgM 1.66g/L(166mg/dL), Hptoglobin 0.19g/L(19mg/dL)(type 1-1), Direct Antiglobulin test negative, HBsAg positive.
32
Oral Presentation
Association of Liver Enzymes and Bilirubin with Impaired
Blood Glucose
A Cross-sectional Survey of Patients in Mombasa Kenya
OL01-2
The prevalence of diabetes in the Rural Kenyan Community is not known.
To determine prevalence of the disease within the community living in
this region.
METHOD
A retrospective data analysis carried out to the patients who visited the
hospital and referred to the diabetic clinic for a period between July 2008
to December 2015.
RESULTS
Out of these tested 1559 (14%) their Random Blood sugar levels were
above 11.7mmols.
878 (7.8%) were newly diagnosed to be diabetic.
Of the total 6366(57%) were females and 4768 (43%) were males.
DISCUSSION / CONCLUSION
The study shows that there many people who are suffering from the disease and they are not aware. Woman are mostly affected by the disease.
This trend shows that there is need for more Public Health awareness. So
that people get checked regularly. The National prevalence of diabetes is
between 2.7% to 14%
World wide 285 million people are affected by diabetes.
Nearly 1.5 million Kenyans are living with the disease and more are not
aware.
Therefore immediate intervention must be put in place o reduce the burden.
Also reduce the cost of testing and treatment so that many more patient’s
can attend the hospital for monitoring and treatment.
GLYCATED HEAMOGLOBIN AND FBS RATIO IN TYPE II
DIABETES MELLITUS PATIENTS AT NNPC HOSPITAL,ABUJA.
Glycated heamoglobin, Fasting blood sugar.
OL01-4
Accuracy-based proficiency testing of serum creatinine in Taiwan
Accuracy assessment of serum creatinine with an isotope dilution
LC-tendam mass spectrometry method in Taiwan
Shu-Chu Shiesh1, Jau-Tsuen Kao2, Wen-Shyang Hsieh3,4, Sheng-Hsiang Yang4
1 Medical
Laboratory Science and Biotechnology, National Cheng Kung University, Taiwan
2 Clinical
Laboratory Sciences and Medical Biotechnology, National Taiwan Univeristy, Taipei, Taiwan
3 Department
of Laboratory Medicine, Taipei Medical University, Shuang Ho Hospital, New Taipei, Taiwan
4 Committee
of Proficiency Testing, Taiwan Society of Laboratory Medicine, Taipei, Taiwan
35
Poster Presentation
Estimated glomerular filtration rate (eGFR), derived from serum creatinine,
is used as a criterion for the diagnosis and staging of chronic kidney disease. Therefore, accurate and precise measurement of creatinine is critical
for proper clinical management. This study was aimed to establish a liquid
chromatography-isotope dilution tandem mass spectrometry (ID-MS/
MS) as the reference method for creatinine measurement, and assess the
performance of routine creatinine assays in Taiwan. The ID-MS/MS reference method was established using a separation on Xterra C18 column
with Agilent 1200 LC system and detection on API 5000 triple quadruple
mass spectrometer. The intra-run and inter-run imprecision (CV%) of the
established reference method was 0.4%-1.5% and 2.1%-3.8%, respectively.
The accuracy of the method was assessed by standard reference material
(SRM967) from NIST with the recovery of 98.5% and 98.6% at the level
of 0.86 and 3.93 mg/dL, respectively. Three fresh-pooled plasma samples
were sent to participating laboratories. The survey samples were run in
triplicate on 2 separate days by the established reference method; the
mean of all results was used as the assigned target value. A passing criterion of +/- 0.2 mg/dL or 9.8% of the target value was used. Acceptable
performance was deemed as all three survey samples within the acceptable limits. At the creatinine concentrations tested method-specific interlaboratory imprecision (CVs) were 1.6%-9.9%. Three methods had interlaboratory CVs less than one-third of the PT limit (3.3%) at creatinine
concentration of 2.49 mg/dL. Differences between target values and median values from the commonly used methods ranged from 0.006 mg/dL
to 0.146 mg/dL at creatinine concentration of 1.45 mg/dL; and 0.068 mg/
dL to 0.188 mg/dL at the concentration of 2.49 mg/dL. 84% of laboratories passed the survey. In conclusion, we developed a reference method for
serum creatinine and assessed the accuracy of routine assays in Taiwan.
Oral Presentation
Introduction: Diabetes mellitus has been one of the non communicable disease consuming developing nation with attendance high mortality rate in
Sub Sahara Africa. Many complications associated with the disease due to
poor health facilities and inadequate prognostic index to identify the likelihood of complication has been the bane of many patients .
Methodology: This study is carried out to identify the possibility of fasting
blood sugar(FBS) and glycated hemoglobin ratio (FBS/GLYHB)as prognostic index for predicting cardiovascular disease complication independent
of lipid profile .Reflotron plus analyzer and semi automated analyzer
were used for analysis of fasting blood sugar and lipid profile(enzymatic
method) while Clove A1C tm analyzer were used for the glycated heamoglobin. Two hundred and forty six diabetes patients and one hundred non
diabetes patients were involved in the study.
Result: The results shows statistical significant in major biochemical considered except HDL-C. There are significant negative correlation between
FBS/GLYHB and atherogenic index measured HDL-C percentage in total
cholesterol P ＜ 0.037;positive correlation with triglycerides P ＜ 0.004
and negative correlation with HDL-C though not statistically significant P
＞ 0.05.
Conclusion: Conclusively ,FBS/GLYHB ratio can safely be used as a prognostic index for determine the risk of cardiovascular disease in type ii
diabetes patients independent of lipid profile with the pattern above. Since
increase in ratio indicate safety to an extent.
Case Conference
ADEYEMI A ADESINA1, DAVID A BADEJO2, SAMUEL O OYEDEJI1
1 Department
of Chemical Pathology, Obafemi Awolowo University Teaching Hospital Complex, Nigeria
2 NNPC
Medical service Abuja,Nigeria
Symposium
OL01-3
A total 11,134 had their Random Blood Sugar level tested and those who
tested levels above 11.7 mmols were monitored.
Educational Lecture
Aspartate aminotransferase and alkaline phosphatase are associated with
increased risk of type 2 diabetes. In Kenya data on liver markers among
prediabetics is still limited, therefore this study examined the association
between aspartate aminotransferase (AST), alkaline phosphatase gammaglutamyltransferase (GGT) and total bilirubin with the risk for impaired
fasting blood glucose (IFG) among patients attending Macca Medical Clinic
in Mombasa, Kenya.
This was a cross sectional study, data collection was by review of laboratory records at Macca Medical Clinical, Mombasa between February 2013
and December 2015. Statistical analyses were done using Mann-Whitney
U and Spearmans to test for difference and correlation between continuous variables. The Fischer’s Exact test and logistic regression model were
used to compare categorical variables. A P-value ＜ 0.05 was considered
statistically significant.
79 participants were included, among them 22 (27.8%) were male and 57
(72.2%) female. 27 (34.2%) of the participants presented with IFG. Fasting blood glucose values were significantly higher among participants
aged ＞ 40 years (P = 0.025), however alkaline phosphatase values were
significantly lower among participants aged ＞ 40 years (P = 0.021). GGT
showed significant negative correlation with triglycerides (P= 0.033). We
observed no significant association between liver markers and IFG.
IFG was significantly higher among older participants and alkaline phosphatase also varied significantly by age group. GGT values correlated with
triglycerides. Therefore screening of liver markers among pre-diabetics is
prudent to prevent future diabetes related adverse outcomes.
Key word: Aspartate aminotransferase, alkaline phosphatase, gammaglutamyltransferase, Fasting blood glucose, Diabetes mellitus
Special Lecture
JOSEPH N MWANGI1, Ronard R Wigina2
1 Kenya
national blood transfusion services, ministry of health, Kenya
2 Mombasa
polytechnic University College
Invited Lecture
Nicholas Polle1, Felix Mwango2, Ali Mohamed2
1 Technical
University of Mombasa, Kenya
2 Macca
Medical Clinic
Keynote Speech
OL01-1
Keynote Speech
OL01-5
Effect of AK activity on the JSCC recommended CK activity
by double kinetic method
OL02-1
Radiation-induced angiosarcoma of the uterus: Report of a
secondary sarcoma case
Shoko Kitaaki, Kousuke Sakurai, Yuuki Watanabe, Itsuko Sato, Yuji Nakamachi, Nobuhide Hayashi,
Jun Saegusa
Shuhei Ishii1,2, Kyoko Komatsu1,2, Junzo Fujiyama2, Takahiko Itoh2, Rira Hoshi2, Marisa Yamada2,
Naoko Suzuki2, Noriyuki Furuta2, Yutaka Takazawa1, Yuichi Ishikawa1,2, Yuko Sugiyama2
Kobe University Hospital, Japan
1 Division
of Pathology, The Cancer Institute, Japan
2 Department
of Cytology, The Cancer Institute Hospital, Japanese Foundation for Cancer Research, Tokyo, Japan
Invited Lecture
Special Lecture
Educational Lecture
Background:Adenylate kinase (AK) is widely distributed among different organs, such as muscle, liver, and red blood cells, and has multiple
isozymes. Because AK give rises to positive error on the reported values
of creatine kinase (CK), Japan Society of Clinical Chemistry (JSCC) recommends minimizing its interference by adding AK inhibitors (AMP and
AP5A) in the assay. Further, CK activity is calculated by a so-called double
kinetic assay, in which the background activity in the absence of creatine
phosphate (first reaction, pH 6.6) is subtracted from the second reaction
in the presence of creatine phosphate (pH 6.8). Here we examined if AK
interfered with CK in those cases.Reagent and instrument:Cygnus Auto
CK, adherent to the JSCC recommended method (Shino-Test Corporation)
and an automated analyzer BM8040 (JEOL Ltd.) were used.Method and
results:Among 27,767 samples, only 7 samples showed relatively high AK
activity in the first reaction( Δ Abs/min ＞ 0.01). Negative errors of CK
assay of the 7 samples ranged from -27 to 0 U/L. We identified isozymes
of AK by calculating the inhibition rate of AK and by electrophoresis. The
inhibition rate of AK by AMP was dominant in 2 samples and they showed
AK2 activities. The inhibition rate of AK by AP5A was dominant in 5 samples. Among 5 samples, 4 samples showed AK3 activities and one showed
AK2 and AK3 activities. Discussion:Use of AK inhibitors and double kinetic
assay are good tools to minimize the error in CK assays, however, high AK
activities may cause negative errors in CK values. Since the occurrence of
such errors is rare, and the maximum negative error in CK in the present
study was -27 U/L, we concluded that the interference of AK in CK assay is
negligible in the clinical setting.
Symposium
OL02-2
[Introduction] Angiosarcoma is a rare soft tissue tumor. It may occur after
cancer therapies such as irradiation and chemotherapy and it is known
as a secondary sarcoma. Secondary angiosarcomas differ in many ways
including age, precipitating factors and presentation. Angiosarcoma per
se has a highly malignant behavior and a poor prognosis. Therefore,
although there are some related names such as hemangiosarcoma, hemangioendothelial sarcoma and malignant hemangioendothelioma, these
additional terms should be avoided to prevent confusion with epithelioid
hemangioendothelioma, which is a separate entity of low malignancy. [Patient] A 77-year-old Japanese woman who presented with abnormal genital bleeding. Her medical history included radiation therapy for squamous
cell carcinoma of the uterine cervix 13 years before. Magnetic resonance
imaging showed no tumor formation in the uterus and fluorodeoxyglucose
uptake was not observed by positron emission tomography. [Cytology]
Cytological examination of the uterine cervix showed malignant cells
composed of a small number of spindle cells. The differential diagnosis
included non-epithelial tumor, and the tumor was difficult to diagnose by
Pap stains. [Histology] The patient undertook open surgery with anterior
pelvic exenteration and construction with an urethrocutaneous fistula.
Histological diagnosis was angiosarcoma. [Discussion] This case is thought
to be a secondary angiosarcoma induced by irradiation because the tumor
was developed within the irradiation field and because there was a long
latent period after irradiation, 13 years. Results of immunohistochemistry
and comparisons with similar secondary tumors of other organs will be
presented.
Developing techniques for differentiating between melanoma
and spitz by using iCCD.
OL02-3
Developing techniques for differentiating between
adenocarcinoma and squamous cell carcinoma in lung.
Case Conference
Oral Presentation
Poster Presentation
Emmy Yanagita, Akikazu Endo, Hiroshi Yamada, Ryuko Tsukamoto, Tomoo Itoh
Emmy Yanagita, Akikazu Endo, Hiroshi Yamada, Ryuko Tsukamoto, Tomoo Itoh
Kobe University Hospital, Japan
Kobe University Hospital, Japan
[Background]Spitz and melanoma are characterized by similar clinical
presentations that are hard to differentiate. Because these two diseases require distinct regiments of therapy despite their similarity, differentiation
between them is crucial. To achieve this, we developed a multiplex immunohistochemical method that can simultaneously stain cells in the G1 and
S/G2/M phases and undergoing apoptosis using 3 markers cdt1, geminin,
and H2AX.[Method]The tissue speciments of spitz, melanoma and narmal
skin cells were fixed in 10% buffered neutral formalin and embedded in
paraffin. We used immunohistochemistry-based cell cycle detection (iCCD)
to compare the staining patterns of each specimen. (A Novel System to Visualize Cell Kinetics on Formalin-fixed Paraffin-embedded Tissue. Am J Surg
Pathol 2012;36:796-773)[Results]Specimens from normal skin and spitz
showed organized staining patterns of cells positive for these cell cycle
markers unlike those of melanoma. As compared to normal skin and spitz,
melanoma showed an increased proportion of geminin-positive cells and
decreased proportion of cdt1-positive cells. The result shows that iCCD is
superior to conventional single-color immunostaining, it allows examination for multi-cell populations at one time. In addition, unlike multicolor
immunofluorescence techniques, which requires the use of fluorescence
microscopes, iCCD only requires light microscopes. Western blotting quantitatively examines the expression of proteins but it cannot determine the
cellular origin.[Conclusion]In conclusion, the multicolor immunostaining
method such as iCCD enables the differentiation between two distinct pathologies with similar the differentiations, such as spitz and melanoma.
[background]The current Food and Drug Administration approved standard treatment for non-small cell carcinomas consists of carboplatin/taxol/
avastin. However, nowadays more specialized protocols, depending on tumor subtype, are being used for lung cancer patients. Therefore, accurate
differentiation between adenocarcinoma and squamous cell carcinoma
is essential for the selection of appropriate therapies. We designed a
rapid multiplex immunostaining method using a novel 4 antibody cocktail, YANA4. This antibody cocktail consists of the following monoclonal
antibodies:rabbit for thyroid transcription factor 1(TTF1), mouse for
napsin A, mouse for p63, and rabbit for CK14. All procedures can be completed within 2 hours. This method labels the nuclei of adenocarcinomas
as brown with TTF1, and cytoplasm as blue with napsin A. Squamous cell
carcinomas could be differentiated from adenocarcinomas with an inverse
staining pattern:blue nuclei with p63 and brown cytoplasm with CK14.
(Yanagita E, Itho T, et al.Rapid multiplex immunohistochemistry using the
4 antibody cocktail YANA4 in differentiating primary adenocarcinoma
from squamous cell carcinoma of the lung. Appl Immunohistochem Mol
Morphol. 2011 Dec:19(6):509-513.)We have recently developed a new
IHC method based on an alternating current electric field to facilitate
the antigen-antibody reaction (R-IHC). This technique was developed for
non catalytically mixing microdroplets. In this method, an alternatingcurrent (AC) electric field is used to promote the antigen antibody reaction within the microdroplet. According to this technique, it is possible to
immunostaining only 30min. [experimental]We have employed YANA4
and R-IHC to study differentiating between adenocarcinoma and SCC in
lung.1.Staining in 30min2.Comparing the staining pattern[results]The results showed the effectiveness of the proposed method.
36
A comparative evaluation of three phenotypic methods in the
detection of carbapenemase-producing gram-negative rods
OL02-5
Age-specific percentile-based reference curve of serum
procalcitonin concentrations in Japanese preterm infants
Yuki Ohno, Akihiro Nakamura, Abe Noriyuki, Eriko Hashimoto, Hiroko Matsutani, Saori Fukuda,
Hisashi Kohno, Fumihiko Nakamura
Noriko Fukuzumi1, Kayo Osawa2, Itsuko Sato1, Ruri Kouno1, Nobuhide Hayashi1, Ichiro Morioka3,
Jun Saegusa1
Div. of Clinical Bacteriology, Dept. of Laboratory Medicine, Tenri Hosp., Japan
1 Department
of Clinical Laboratory, Kobe University Hospital, Japan
2 Department
of Biophysics, Kobe University Graduate School of Health Sciences
3 Department
of Pediatrics, Kobe University Graduate School of Medicine
Examination of the maltose detection method as the examination of gastric mucosa disorder
Development of the enzymatic method and UHPLC method for maltose permeability test of gastric
mucosa using oral glucose tolerance samples
1 Division
of Medical Technology, Department of Health Sciences, Graduate school of Medicine, Kyushu University, Japan
2 Department
of Medical Technology and Sciences, School of Health and Sciences at Narita, International University of Health
and Welfare
3 Departmnt
of Medical Technology, Faculty of Health Sciences, Junshin Gakuen University
BACKGROUND
MALDI-TOF MS is increasingly used for routine bacterial and fungal identification in Japan, whereas application of LC-MS/MS in clinical chemistry
laboratories has remained very limited. Although immunoassays are often
used for measurement of serum estradiola (E2) when fast turnaround time is
required, more sensitive and specific measurements are needed for determination of menopausal status, estrogen deficiency and in the diagnosis of sex
hormone related disorders. The aim of this study is to develop and validate
liquid chromatography – tandem mass spectrometry (LC-MS/MS) method for
simultaneous measurement of E2, that compere tew immunoassay kits.
MATERIALS AND METHODS
Patient samples: fumale 60 samples
LC-MS/MS methods: For sample preparation, 100 μ L of calibration solution,
QC sample or patient serum were diluted, spiked by 20 μ L of internal standard (10ng/mL) and were loaded on the supported liquid extraction (SLE) plate
and were then extracted by using 1.8 mL of extraction solution. The extracted
samples were dried under nitrogen and were derivatized by dansyl chloride
acetone solution. An aliquot for 40µL was then subjected to LC-MS/MS.
The selected reaction monitoring (SRM) was performed with a Bruker EVOQ
Elite in electrospray ionization (ESI) and positive ion mode. The SRM transitions were 506 ＞ 171 for E2.
Immunoassay kits: CLIA Architect® Estradiol2, Abbott and CLIA ADVIA Centaur® E-Estradiol-6, Simens Healthcare were used.
RESULTS
Method comparison studies for All samples showed: [Architect] = 0.8597 [LCMS E2] +15.652 (n=60) , for samples with E2 ＜ 60pg/mL showed: [Architect
E2] = 0.7448[LC-MS E2] +15.295 (n= 20) and [Centaur] = 0.959[LC-MS E2]
+15.258 (n=60) , for samples with E2 ＜ 60pg/mL showed: [Centaur E2] =
0.7953 [LC-MS E2] +23.564 (n= 20) .
CONCLUSION
The immunoassay of overestemate is serious in the low concentration. We are
now planning to use this method for E1 and E2 measurement on a routine
basis in our university hospital.
Background: Early detection and treatment of gastric cancer are important. An easily and noninvasive screening test is required for detection in
the early stage of gastric cancer. Seimiya et al. reported the sucrose permeability test as a simple and noninvasive marker of gastric mucosal damage
in human subjects. Therefore, we aim to develop an enzymatic method for
a maltose permeability test of gastric mucosa using oral glucose tolerance
samples instead of sucrose. Moreover, we developed an Ultra High-Performance Liquid Chromatography (UHPLC) method for measuring maltose as
a reference method and estimated the present enzymatic method with it.
Method: ＜ Enzymatic method ＞ This method is comprised of two reagents: one is for removal of endogenous glucose by GOD and in the other
Α -Glucosidase acts for maltose and the formed glucose is detected. The
enzymatic assay is used with a Hitachi 7600 automated analyzer. Plasma
maltose was measured based on a 2-point end assay. ＜ UHPLC method ＞
The following conditions were used. Column; GL InertSustain C18 50 ×
2.0 mm l.D. Eluent; acetonitrile and 40 mmol/L acetic acid/sodium acetate
buffer (pH 4.2). Flow rate; 0.3 mL/min. Column temperature; 50 ℃ . UV
detect; at 271 nm. Injection volume; 5 µL.
Result: ＜ Enzymatic method ＞ Basic performance evaluations (withinrun CVs, recovery test, linearity and detection limits) were good. ＜ UHPLC
method ＞ Maltose was retained at 3.7 min. The present method was sufficient to detect maltose without interference from other components in
plasma. The correlation of the two methods is also good.
Conclusions: Our enzymatic assay was sensitive enough to monitor plasma
maltose concentrations during an oligosaccharide permeability test. We
consider Maltose has a potential that is available as the test of damaged
gastric mucosal instead of sucrose. We address the relationship between
the degree of gastric mucosa disorder and maltose concentration.
37
Poster Presentation
1 Department
of Molecular Diagnosis, Graduate School of Medicine, Chiba University, Japan
2 Division
of Clinical Mass Spectrometry, Chiba University Hospital, Japan
3 Departmen
of Reproductive Medicine, Graduate School of Medicine, Chiba University, Japan
4 Division
of Laboratory Medicine, Chiba University, Japan
Oral Presentation
Yui Nakano1, Eisaku Hokazono1, Eri Ohta1, Miki Kawano1,2, Takiko Tateishi1,3, Susumu Osawa2,
Yuzo Kayamori1
Case Conference
Yui Miyabayashi1, Mamoru Satoh2, Masaki Takiwaki2, Motoi Nishimura1,4, Kazuyuki Matsushita1,4,
Makio Shozu3, Fumio Nomura2
Symposium
OL03-2
Educational Lecture
Comparison of LC-MS/MS and immunoassays
due to cross-reactivity in estradiol
Special Lecture
OL03-1
Introduction: Procalcitonin (PCT) levels are elevated early after birth in
newborn infants; however, the physiological features and reference of
serum PCT concentrations have not been fully studied in preterm infants.
The aims of the current study were to determine the features and establish an age-specific percentile-based reference curve of serum PCT concentrations in preterm infants. Methods: Serum samples (n=1267) were
obtained from 283 Japanese newborn patients at birth to 136 days old.
The enrolled newborns were divided into three infant groups on the bases
of gestational age (GA) as follows: preterm (<34 weeks’ GA, n = 37), late
preterm (34 – 36 weeks’ GA, n = 61), and term infants ( > 37 weeks’ GA, n
= 185). Three late-onset bacterial infected preterm infants were enrolled
to analyse changes in serum PCT concentrations. The serum PCT concentration was measured by electrochemical luminescence immunoassay.Results: The PCT concentration peaked in infants at 1 day old and decreased
thereafter. At 1 day old, the serum PCT concentration in preterm infants
was higher compared with that in late preterm or term infants (50-percentile values were 11.1 ng/mL in preterm, 1.2 ng/mL in late preterm, and 2.2
ng/mL in term infants). Although the 50-percentile value in late preterm
and term infants reached the adult normal level (0.1 ng/mL) at 5 days old,
it did not in preterm infants (0.31 ng/mL). It took 9 weeks for preterm
infants to reach it. Serum PCT concentrations at onset in late-onset infected preterm infants were over the 95-percentile value. Conclusion: We
showed that the physiological feature in preterm infants was significantly
different from that in late preterm infants, even in those <37 weeks’ GA. To
detect late-onset bacterial infection and sepsis, an age-specific percentilebased reference curve may be useful in preterm infants.
Invited Lecture
Objective: Carbapenemase-producing gram-negative rods (CPGNRs) have
been spreading worldwide and has become a threat to healthcare systems.
The purpose of this study was to compare three phenotypic methods for
detecting CPGNRs: the Modified Hodge Test (MHT), the Carba NP Test,
and the Carbapenem Inactivation Method (CIM). Material and Methods:
A total of 49 carbapenem resistant GNRs from clinical specimens at Tenri
Hospital (Japan) were used. Twenty-eight CPGNR isolates (IMP-1 group,
n=27 [Klebsiella pneumoniae , n=10; Escherichia coli , n=7; Enterobacter
cloacae , n=3; Klebsiella oxytoca , n=1; Enterobacter aerogenes , n=1; Proteus
mirabilis , n=1; and Pseudomonas aeruginosa , n=4]) and VIM-2 group, n=1
[P. aeruginosa ]) and 21 non-CPGNR isolates (P. aeruginosa , n=20; and E.
aerogenes , n=1) were used. These isolates confirmed that it was not the
same clone in rep-PCR. We used the three phenotypic methods to test for
all of the above-mentioned strains, and evaluated the detection capability
of each of the methods. Results: The rates of agreement of the MHT, the
Carba NP Test and CIM were 96% (27/28), 93% (26/28), and 96% (27/28),
respectively, in the CPGNR group, and 95% (20/21), 100% (21/21), 100%
(21/21) in the non-CPGNR group. The three phenotypic methods provided
false-negatives for a P. aeruginosa isolate (VIM-2 group) while, the Carba
NP Test provided a false-negative for a P. mirabilis isolate. Conclusion: All
three phenotypic methods showed high discriminability. Furthermore the
Carba NP Test and CIM were comparable to MHT. The Carba NP Test was
the quickest of the three phenotypic methods. However, it should be used
with care because it may provide a false-negative for P. mirabilis .
Keynote Speech
OL02-4
Keynote Speech
OL03-3
Oxidation and glycation modulate HDL capacity to carry
Sphingosine 1-phosphate, an Anti-atherosclerotic Bioactive
Lipid.
OL03-4
Modification of apolipoprotein A-I in high-density lipoprotein
by myeloperoxidase and chymase.
Tamaki Kobayashi1, Makoto Kurano1, Takahiro Nojiri1, Ryunosuke Ohkawa2, Minoru Tozuka2,
Shigeo Okubo1, Yutaka Yatomi1
Maasa Morita1, Ryunosuke Ohkawa1, Megumi Satou1, Akira Yoshimoto1, Kouji Yano1, Takeshi Kasama2,
Minoru Tozuka1
1 The
University of Tokyo Hospital, Japan
2 Tokyo
Medical and Dental University
1 Analytical
Laboratory Chemistry, Graduate School of Health Care Sciences, Tokyo Medical and Dental University, Japan
2 Instrumental
Analysis Research Division, Research Center for Medical and Dental Sciences, Tokyo Medical and Dental
University
Invited Lecture
Special Lecture
Educational Lecture
Introduction: Sphingosine 1-phosphate (S1P) is a potent lysophospholipid
mediator, which is thought to possess anti-atherosclerotic properties. Since
two thirds of S1P are carried on high-density lipoprotein cholesterol (HDL)
in plasma, we investigated whether post-transcriptional modulation, such
as oxidation and glycation, would affect the S1P-binding capacity of HDL.
Methods: HDL fractions were prepared with the ultracentrifugation method from healthy human subjects. (1) We oxidized HDL with CuSO 4 (ox-HDL)
or glycated HDL with methylglyoxal (gly-HDL) for 24 hours and determined the S1P contents of HDL. (2) We also investigated the S1P-binding
capacity of each modulated HDL. We investigated how much C17S1P
transferred to the HDL by incubating C17 S1P bound to albumin with each
modulated HDL. (3) Apolipoprotein M (ApoM) is a carrier of S1P on HDL.
Therefore, we finally examined the modulation of ApoM by oxidation or
glycation with a western blot analysis. Results: (1) The S1P contents were
significantly decreased in ox-HDL and gly-HDL, compared with those of
na ïve HDL. (2) The S1P-binding capacity was significantly attenuated in
ox-HDL and gly-HDL; HDL/non-HDL ratio in Gly-HDL and Ox-HDL were
lower than that in na ï ve HDL. (3) The western blot analysis revealed that
ApoM in gly-HDL was detected at ＞ 250kDa, while that in ox-HDL and na
&iuml;ve HDL was located at around 25 kDa. When we glycated HDL with
more mild conditions, ApoM was detected at around 25 kDa, 50 kDa, and
100 kDa, suggesting that glycation might polymerize ApoM. Conclusion:
Post-transcriptional modulation such as oxidation and glycation affects
the S1P contents and S1P-binding capacity of HDL, which might at least
partly explain that anti-atherosclerotic properties of HDL were attenuated
by oxidation and glycation.
Symposium
OL03-5
Background: Apolipoprotein A-I (apoA-I), the primary component of highdensity lipoprotein (HDL), exerts antiatherogenic effects in atherosclerotic lesions. However, in the progression of atherosclerosis, apoA-I has
been exposed to modifications involving oxidation by myeloperoxidase
(MPO) mainly secreted from macrophages and truncation by chymase
released from activated mast cells. We previously showed some fragments of apoA-I with apparent molecular masses of around 10 kDa were
produced after the treatment of HDL with both MPO and chymase. The
aim of this study is to identify these fragments on the assumption that
they would be possible biomarkers for a progression of atherosclerosis.
Methods: HDL (1.063-1.210 g/mL) was isolated from healthy subjects
by ultracentrifugation. HDL was treated by sequential incubation with
MPO and chymase. Treated HDL was re-ultracentrifuged at the density of 1.063 g/mL. HDL sample was then separated by SDS-PAGE and
nondenaturing-PAGE, followed by Western blotting using anti-apoAI polyclonal antibody. To identify the cleavage sites, truncated apoAI bands separated by SDS-PAGE were subjected to in-gel digestion with
trypsin followed by MALDI-TOF-MS and MALDI-TOF-MS/MS analysis.
Results: Truncated apoA-I fragments with molecular masses of approximately 14.5 and 12 kDa were observed in MPO- and chymase-treated
HDL, however, those bands were not observed in only chymase-treated
HDL. After re-centrifugation of treated HDL samples, these fragments
remained binding to HDL without apparent change in the particle size.
Mass spectrometric analysis suggested that the 14.5 kDa fragment corresponded to C-terminal part of apoA-I and the 12 kDa fragment was the
mixture of N-terminal and C-terminal parts. Conclusion: MPO treatment
alters apoA-I in HDL particle susceptible to chymase digestion suggesting
that truncated apoA-I with molecular masses of approximately 14.5 and
12 kDa might be available as biomarkers reflecting the state of atherosclerotic lesions. Specification of the actual cleavage sites will be needed to
develop assay methods of these fragments.
Development of detection method for serum variant
transthyretins using MALDI-TOF mass spectrometry
OL03-6
Development of an assay for measuring diluted plasma sodium concentration in
fingertip blood samples collected
A novel home examination system based on trace blood sampling from the fingertip
Case Conference
Masayoshi Tasaki1, Mitsuharu Ueda2, Konen Obayshi1, Taro Yamashita3, Yukio Ando2
Susumu Osawa1, Sinya Sugimoto2
1 Department
of Morphological and Physiological Sciences, Graduate School of Health Science, Kumamoto University, Japan
2 Department
of Neurology, Graduate School of Medical Sciences, Kumamoto University
3 Diagnostic
Unit for Amyloidosis, Department of Neurology, Kumamoto University Hospital
1 Leisure,
Inc. International University of Health and Welfare, Japan
2 Leisure,
Inc.
Oral Presentation
BACKGROUND We newly produced a test kit for performing health
checkups based on 65- μ L fingertip blood samples for measuring biochemical parameters. The plasma sample dilution ratio was measured by
assessing the sodium level of each diluted sample. Development of the
dilution of plasma sodium was determined using an enzymatic rate assay.METHODS1. The DEMECAL kit The DEMECAL kit (Fujifilm, Japan) is
composed of a tube, dilution buffer solution, a lancet, a blood-aspiration
sponge, a cylinder with a blood cell separation filter, a cap, a swab, and a
sticking plaster. The dilution buffer solution is composed of HEPES buffer
and EDTA-2K.2. Instrument and measurement conditionsA JCA-BM6050
automated analyzer (JOEL Ltd., Japan) was used to obtain the biochemical
measurements with commercially available reagents. The sample volume
was greater than that of the original sample.RESULTS The sodium analysis
exhibited good linearity from 0-25 mmol/L. The proportionality of the
dilution rate of sodium was twenty fold admitted. The measurements were
performed 20 times, and the mean coefficient of variation at a dilution
rate of 9.8-fold was 2.2%. The correlations between the results obtained
with the DEMECAL kit and venous plasma analysis were as follows: AST:
0.990, ALT: 0.998, GGT: 0.998, total cholesterol: 0.973, HDL-cholesterol:
0.987, LDL-cholesterol: 0.990, triglycerides: 0.999, urea-nitrogen: 0.993,
creatinine: 0.966, uric acid: 0.994, and glucose: 0.994. The diluted plasma
remained stable in three different temperature conditions (4, 20, and 37
degrees Celsius).CONCLUSIONSThis measurement system can provide reliable clinical data about various molecules in diluted plasma derived from
65- μ L samples of fingertip blood collected at home. The DEMECAL kit
can be used to collect blood at distant locations, providing a suitable transport system is available. The DEMECAL kit can also be employed for other
biochemical and immunity tests.
Introduction: Familial amyloid polyneuropathy (FAP) is a hereditary amyloidosis caused by a variant transthyretin (TTR). To diagnose FAP, we have
performed the detection of serum variant TTR using mass spectrometry.
However, it takes 3 hours to obtain the results.In this study, we developed
a simple and rapid detection method for serum variant TTR in FAP. Methods: We used 121 serum samples obtained from FAP patients. To detect
serum variant TTRs, we used matrix assisted laser desorptionionization
time of flight mass spectrometry (MALDI TOF MS, Bruker). Procedures for
the assay, including matrices and serum concentrations, were optimized.
Results: TTR was efficiently detected from 100 fold diluted serum samples
by using MALDI TOF MS. Therefore, our data revealed that DHB was suitable for detection of serum TTR compared with others. This system allowed
analysis for serum TTR via a 30 minutes one step procedure. Several variant TTRs (Val30Met, Ser50Ile, Leu55Pro, Ile107Val and Tyr114Cys) were
detected by MS.Conclusion:Our novel method is very simple and rapid to
detect several serum variant TTRs.
Poster Presentation
38
Microbial diversity of Enterobacteriaceae in Mondego River
Microbial diversity in Mondego River
OL04-2
Bacterial Community ESBLs producers in water belonging to
rural and urban areas
ESBLs in urban areas
Introduction: Mondego is a Portuguese river where many socio-economical activities are based. Its route crosses some anthropogenic threats that
can negatively modify the microbiological water quality which may affect
public and environmental health. Enterobacteriaceae are used as contamination indicator to prevent the occurrence and spread of waterborne
diseases. This study intends to identify the microbial diversity and the antibiotic sensitivity profile of enterobacteria on Mondego River.
Material and methods: 233 isolates were collected from three different
places along the river. The genotyping of the isolates was based on BOX
PCR and the identification was achieved with RapIDTM ONE System. We
have also analyzed the antibiotic sensitivity profile of each isolate based
on Kirby-Bauer method.
Results: 65 of the 233 isolates were classified as enterobacteria, with special emphasis to Escherichia coli (16.9%), Hafnia alvei and Shigella Sonnei
(7.7%) and Salmonella choleraesuis (6.2%). The other enterobacteria species represent 13.3% and the remaining isolates (47.9%) were not identified. The results showed 46% and 26% of the isolates are ampicilin and
nalidixic acid resistant, respectively. Concerning the collection sites there
is an increase of antibiotic resistance in Local C and more bacterial diversity on Local B.
Discussion: A total of eleven species were isolated from 65 enterobacteria.
Therefore it was demonstrated the existence of opportunistic pathogens,
as E. coli, with capacity to originate severe infections mainly on susceptible individuals. It is also important to refer the resistance of this isolates
to some of the antibiotics commonly used in the clinical treatment to bacterial infections. This study indicates that anthropogenic threats are the
main factor for the decrease in water quality of the Mondego River and
also influence the antibiotic sensitivity profile of the bacteria present.
Keywords: Enterobacteriaceae, Public Health, Microbiological water quality
Introduction: The treatment of human drinking water are often ineffective
in removing some chemicals compounds, including antibiotics and their
metabolites. Thus, these aquatic environments become promising for the
development of resistant strains, as well as excellent places for the dissemination of resistance genes, between different bacterial species. The
spread has been high in genes encoding enzymes capable of cleaving betalactam antibiotics, in particular cephalosporins, known ESBLs.
Material and Methods: The collection of water samples was made in
fountains close to urban and rural areas belonging to the Viseu. The microorganisms were isolated from MacConkey medium supplemented with
Cefotaxime (2181.g/mL) and their identification was performed by Api
galleries. The antibiotic sensitivity tests were performed by the disk diffusion method.
Results: From a total of 29 microorganisms were able to identify approximately 15 different bacterial species. In urban areas of water quality
showed a greater bacterial diversity comparing to a rural areas, mostly belonging to the family not Enterobacteriaceae. These bacteria also showed a
resistance profile for some antibiotics tested.
Discussion/Conclusion: The water in urban areas have a higher bacterial
diversity related to EBSLs producing. We believe that the urban involvement promotes the increased of water contamination. Those environments
are most promising for maintenance and spread of resistant strains.
Key-words: Quality of water, Bacterial resistance and ESBLs
Volatile Metabolites as Early Diagnostics for Blood-Borne Pathogens?
The rapid evaluation of bacterial growth in blood cultures by SIFT-MS and
comparison with the BacT/ALERT automated blood culture system
OL04-4
Yatin J Mange1, Jenny M. Scotter1, Randall A. Allardyce2, Vaughan S. Langford1, Alex L. Hill1,
David R. Murdoch2,3
Immunologic property of specific chicken egg yolk antibody (IgY) against Methicillin Resistant Staphylococcus aureus
(MRSA)
Determination of the activity of egg-derived antibodies against Methicillin Resistant Staphylococcus aureus (MRSA)
Symposium
OL04-3
Educational Lecture
Biomedical Laboratory Sciences, Polytechnic Institute of Coimbra, ESTESC-Coimbra Health School, Department Biomedical
Laboratory Sciences, Coimbra, Portugal
Special Lecture
Fernando Mendes, Daniela Rouxinol, Ana Valado, Armando Caseiro, Antonio Gabriel, Nadia Osorio
Biomedical Laboratory Sciences, Polytechnic Institute of Coimbra, ESTESC-Coimbra Health School, Department Biomedical
Laboratory Sciences, Coimbra, Portugal
Invited Lecture
Fernando Mendes, Ana Lopes, Ana Valado, Armando Caseiro, Antonio Gabriel, Nadia Osorio
Keynote Speech
OL04-1
Oliver Shane R Dumaoal1,2, Ronalyn S Sanchez2
1 Syft
Technologies, New Zealand
2 Christchurch
School of Medicine and Health Sciences, University of Otago, New Zealand
3 Microbiology
Unit, Canterbury Health Laboratories, Christchurch, New Zealand
39
Poster Presentation
The timely laboratory diagnosis of bacteremia is central to effective management of bloodstream infections. Earlier detection and identification of
infective bacteria would allow earlier, more accurate antibiotic targeting,
leading to better patient outcomes and reduced cost per case.
Volatile metabolites produced by infecting organisms may offer a faster
path to diagnosis of infection and the causative organism if different organisms produce unique profiles of metabolites. To be effective, a rapid,
selective, and very sensitive analyzer is required. In this study, we have applied Selected Ion Flow Tube Mass Spectrometry (SIFT-MS), which fulfills
these requirements and is an industry-proven analytical technique that
instantly and directly analyzes air to part-per-trillion-by-volume (pptv) concentrations (Prince et al., 2010; Langford et al., 2014).
The aim of this study was to directly compare the time at which in vitro
growth of species commonly causing bacteremia was detected by SIFT-MS
with that of the BacT/ALERT system using identical culture conditions.
Results of this study indicate that the growth of less than 10 CFU of five
bacterial species can be demonstrated by SIFT-MS analysis of headspace
VOCs at 8 h using standard Biomerieux BacT/ALERT aerobic and anaerobic blood culture bottles, that measurement of VOCs may allow species differentiation, and that targeted selected ion mode analysis of 20 VOCs from
each bacterial culture headspace may be completed and recorded within
30 s without any prior sample preparation. Combining shorter incubation
times, rapid analysis, and the newly available autosampler-SIFT-MS systems, could yield a powerful early screening tool.
References:
Langford, V.S., Graves, I., & McEwan, M.J. (2014). Rapid Commun. Mass
Spectrom., 28, 10 – 18.
Prince, B.J., Milligan, D.B., & McEwan, M.J. (2010). Rapid Commun. Mass
Spectrom., 24, 1763-1769.
Oral Presentation
Methicillin Resistant Staphylococcus aureus (MRSA) is an increasingly predominant pathogen that causes severe infection and outbreaks. Its capacity to elaborate its virulence factor and develop its resistance to empirical
antibiotic treatment makes eradication difficult. One recourse is to use
monoclonal antibody but cross reaction phenomenon could lead to failed
treatment. This study explored the potential immunologic property of specific egg yolk antibodies known as immunoglobulin Y (IgY) in inhibiting
the growth of MRSA. Formalin-killed MRSA was used to immunize white
leghorn chickens (Gallus domesticus) and the IgY titer was determined by
micro-agglutination test (MAT). Growth inhibition assay was used to confirm anti-MRSA activity of the isolated IgY while its action on the MRSA
cellular structure was determined by transmission electron microscopy
(TEM). MAT revealed a titer of 1:2048 and the isolated IgY showed a
significant activity in the growth inhibition assay. The 30 microliters and
50 microliters concentration showed a significant difference among treatments at p value ＜ 0.001 while 40 microliters was significantly different
at p value ＜ 0.01. Further, the isolated IgY caused disruption of the cell
wall and eventual lysis of MRSA in TEM at a concentation of 50 microliters. Results of experiments suggest that the isolated IgY was effective in
inhibiting the growth of MRSA.
Case Conference
1 College
of Allied Medical Professions, Lyceum of the Philippines University - Batangas, Philippines
2 Graduate
School, Lyceum of the Philippines University - Batangas
Keynote Speech
OL04-5
Differentiation of toxigenic and non-toxigenicClostridium difficile by Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass
Spectrometry (MALDI-TOF)
Develop the utilization of mass spectrometry for toxigenicC. difficile identification and compare the performance of different testing modalities
OL04-6
Analysis the correlation between the turnaround time of mycobacterium in culture
and acid-fast stain results
The specimen source can affect the turnaround time of mycobacterium culture
Yi Chi Wu1, Yu Han Huang1, Ya Fen Chen1, Chia Lin Lee1, Yu Jiun Chan1,2
Ya-Yen Yu, Yi-Chun Hung, Fu-Ai Chang, Si-Yin Yu, Guo-Xing Hung
1 Division
of microbiology. Department of pathology and laboratory medicine, Taipei Veterans General Hospital, Taipei, Taiwan
2 School
of Medicine, National Yang-Ming University, Taipei, Taiwan
Department of Laboratory Medicine, Chang-Hua Hospital, Ministry of Health and Welfare , Taiwan
CDC in Taiwan recommends turnaround times (TAT) of mycobacterium
in culture within 21 days. The achievement rate in our laboratory was
about 70%. According to CLSI guidelines, it should be reviewed based on
the results of AFS. The aim of this study is to analysis the correlation between the TAT of culture positive and AFS results. All specimens, enrolled
between January 2014 to April 2015, were processed by NALC-NaOH, and
then concentrated smears were prepared and inoculated MGIT AFS and
LJ medium. Smears were screened by fluorescent stain, and positive result
were confirmed by Ziehl-Neelsen stain. In 34,013 specimens, AFS positive
rate and culture positive rate were 9.19% and 15.74%, respectively. MTBC
isolation rate was 5.70%, and non-MTBC was 10.04%. The mean TAT of
culture positives in MTBC groups with AFS result negative, scanty, 1+, 2+,
3+ and 4+ were 28.9, 25.3, 23.1, 20.6, 16.5 and 11.7 days. In non-MTBC
groups, they were 17.7, 14.7, 13.0, 10.3, 9.2, and 7.4 days. The regression coefficients were 0.974 and 0.969, respectively. Both MTBC and
non-MTBC TAT of culture and AFS titers showed significant negative correlation. The TAT required for MTBC culture and non-MTBC also showed
significant difference (p ＜ 0.05). The results indicate that a standardized
threshold (21 days) may not applicable to each laboratory due to variety
sources of the specimens. The amount of the group after treatment will
influence performances of each laboratory. Therefore, every laboratory
needs to calculate the TAT required for mycobacterium culture as a baseline data and as a self-monitoring goal. It is essential to consider external
factors such as whether there is too much pretreatment or poor quality of
culture reagents cause the prolonged of the TAT.
Clostridium difficile (C. difficile ) belongs to Gram positive anaerobic ba-
Invited Lecture
Special Lecture
Educational Lecture
cilli and its infection is highly associated with antibiotics overuse and will
cause diarrhea and colitis. Toxigenic C. difficile is one of the major pathogens for diarrhea among healthcare-associated infections and Clostridium
difficile infection(CDI) is an important indicator for antibiotic stewardship.
Rapid and accurate diagnosis of CDI cases is crucial for proper infection
control. However, traditional gold standard to identify toxigenic C. difficile
is time consuming. Therefore, to develop a convenient method for toxigenic C. difficile identification is crucial.
Current diagnostic methods forc include 1. Anaerobic culture for C. difficile , 2. Enzyme-linked immunoassays for toxins A and B, and 3. Molecular
methods for toxic genes. However, they all have limitations and standard
anaerobic culture methods failed to differentiate toxigenic from nontoxigenic strains. It has been a great success in identification of bacterial
isolates by mass spectrometry. We want to develop a novel technique for
toxigenic C. difficile identification by MALDI-TOF. A total of 100 stools
were included in the study and investigated in parallel by anaerobic culture, GDH - toxin A/B EIA testing of isolates and toxin B gene DNA amplification. Based on toxin B gene DNA amplification and anaerobic culture, a
total of 69 samples were classified as C. difficile positive. Only 39 strains
were identified as toxigenic by toxin B gene DNA amplification and GDHtoxin A/B EIA test. Then the 39 toxigenic strains and 30 non-toxigenic
strains were were further confirmed as C. difficile by MALDI-TOF RUO
mode. Furthermore, the spectrum comparison between the toxigenic and
non-toxigenic groups showed differently. There were 3 speaks present in
toxigenic group but not in non-toxigenic group, which were 3961 m/z,
7262 m/z and 7921 m/z. These results suggested that MALDI-TOF RUO
mode could differentiate C. difficile toxigenic strains from non-toxigenic
strains.
Symposium
OL05-1
Rapid meropenem susceptibility testing
Using MALDI Biotyper antibiotic susceptibility test rapid
assay (MBT-ASTRA).
OL05-2
Sussie Rasmussen1, Jens Joergen E Christensen1,2, Bent L Roeder1, Rimtas Dargis1, Mikkel B Nielsen1
Genetic mechanism underlying drug resistance inMycobacterium kyorinense andMycobacterium celatum
Investigations on the genes related to drug resistance inMycobacterium kyorinense andMycobacterium
celatum using whole genome sequencing
Hiroaki Ohnishi, Kouki Ohtsuka, Satsuki Matsushima, Shota Yonetani, Eriko Nozaki, Satoko Yamasaki,
Tomonori Kishino, Takashi Watanabe
Case Conference
1 Department
of Clinical Microbiology, Slagelse Hospital, Region Zealand, Denmark
2 Department
of Clinical Medicine, University of Copenhagen, Denmark.
Department of Laboratory Medicine, Kyorin University School of Medicine, Japan
Background
We applicated the MALDI Biotyper antibiotic susceptibility test rapid assay
(MBT-ASTRA) on 68 challenging Gram-negative rods for detecting susceptibility to meropenem.
Oral Presentation
Poster Presentation
＜ Introduction ＞ Mycobacterium kyorinense is a nonpigmented, slowly
growing mycobacterium that was initially isolated in 2007. Biochemical tests and genetic analyses showed that M. kyorinense is most closely
related to Mycobacterium celatum . Both strains exhibited relatively high
MICs for rifampin, ethambutol, and isoniazid, and relatively low MICs for
macrolides, aminoglycosides, and quinolones. To explore the genetic similarity of these 2 species and underlying genetic features related to drug
resistance, we performed whole genome sequencing of both species.
＜ Materials & Methods ＞ DNA of M. kyorinense type strain KUM060204T
and M. celatum type strain ATCC51131T were extracted and subjected to
whole genome sequencing using ION PGM system.
＜ Results & Discussion ＞ The assembled sequences of KUM060204T
comprised 453 contigs, with a combined length of 5,302,980 bp, with a
G+C ratio of 66.9%. The assembled sequences of ATCC51131T comprised
1,217 contigs, with a combined length of 4,662,059 bp and a G+C ratio of
66.9%. The percent identities of ahpC , inhA , embB , and rpsL between M.
celatum and M. kyorinense were 95.9%, 95.4%, 91.5%, and 97.9%, respectively, while those between M. celatum and Mycobacterium tuberculosis
were 83.7%, 85.5%, 81.7%, and 90.1%, respectively. These results suggested that M. celatum is closely related to, but distinct from, M. kyorinense .
Analysis of the rpoB gene revealed that of both strains harbor a Ser531Asp
amino acid substitution, the most frequent mutation in rifampin-resistant
M. tuberculosis . In contrast, we did not detect typical substitutions in inhA
and ahpC , which are commonly observed in isoniazid-resistant M. tuberculosis , in M. kyorinense and M. celatum . These results suggested that the
mechanisms underlying drug resistance in M. kyorinense and M. celatum
are similar to each other but partly different from those in M. tuberculosis .
Materials/methods
Antibiotic susceptibility testing: For each strain, three different 200 mcL
setups (0.5 McFarland bacterial suspensions) containing no antibiotics, 2
mcg and 16 mcg/mL meropenem, respectively, were incubated at 37C for
1-1.5 hour. Suspensions were washed with HPLC water and ethanol, pellets were air-dried, bacteria lysed with formic acid and acetonitrile containing an internal standard followed by mass spectrometry examination. Data
evaluation for presence of peaks indicating bacterial growth was done
manually and by ASTRA software (Bruker Daltonics).
Results.Of the 30 known carbapenemase producing strains MICs for meropenem for 21 strains were ＞ = 16 mcg/mL, MIC was ＞ = 2 and ＜ 16
mcg/mL for nine strains, and no strains had MICs below 2 mcg/mL. Of the
26 ESBL/AmpC positive strains, MIC for only one strain was ＞ = 16 mcg/
mL, MIC was ＞ = 2 and ＜ 16 mcg/mL for nine strains, and 16 strains had
MICs below 2 mcg/mL. Among the 12 strains without detected resistance
mechanisms no strains were with MICs ＞ = 16 mcg/mL, MIC was ＞ = 2
and ＜ 16 mcg/mL for six strains and six strains had MICs below 2 mcg/
mL. Of strains with Etest MICs ＜ 2 mcg/mL (n=42) 24 strains by MBTASTRA had MICs ＞ = 2 mcg/mL.
Conclusion. MBT-ASTRA results were obtained within 3-4 hours. The carbapenemase producing K. pneumoniae strains were strong producers with
13 of 14 strains having meropenem MICs ＞ = 16 mcg/mL by both methods, while the examined carbapenemase producing E. coli strains were
weak producers with meropenem MICs ＜ 16 mcg/mL, but by MBT-ASTRA
having meropenem MICs ＞ = 2 mcg/mL. Therefore, when suspected, the
MBT-ASTRA seems capable of detecting reduced sensitivity to meropenem
within hours.
40
Development of a soil DNA extraction method and detection
of Balamuthia mandrillaris genetic material
in soil from Aomori Prefecture, Japan
OL05-4
Toyoyasu Koriyama1,2, Munekazu Yamakuchi2, Kazunori Takenouchi2, Yoko Oyama2, Toshiaki Shimizu2,
Masakaze Matsushita1, Teruto Hashiguchi2
Hiroaki Arima, Kanako Yamanouchi, Koichi Ito, Kazuki Kanto, Yamato Sakamoto, Takashi Inaba
Division of Bioscience and Laboratory Medicine, Hirosaki University , Japan
1 Department
of Laboratory Medicine, Kagoshima University Hospital, Japan
2 Department
of Laboratory and Vascular medicine,Kagoshima University Graduate School of Medical and Dental Sciences
Rie Nagaoka1, Makoto Onodera2, Yumiko Koba2, Toshinori Hara2, Yumiko Joichi2, Maki Furushimo2,
Hiroki Ohge3, Michiya Yokozaki4
Comparison of Cefotaxime Resistant Enterobacteriaceae
Isolated from Domestic and Imported Broilers in Japan
Symposium
OL05-6
Educational Lecture
Serotypes, antimicrobial susceptibility, and molecular
epidemiology of invasive Streptococcus pneumoniae isolates
Special Lecture
Background:Legionella pneumophilia (L. pneumophilia ) is a Gram-negative bacterium that causes severe pneumonia. L. pneumophilia invades
and replicates within the alveolar macrophages, in which L. pneumophilia
blocks the normal process of phagocytosis. The mechanism by which L.
pneumophilia survives in macrophages and dysregulates the function is
still unknown. miRNA is a small non-coding RNA that regulates a series of
protein expressions post-transcriptionally. miRNAs are involved in many
pathological events, such as inflammation caused by bacterial infection.
Methods: L. pneumophilia (ATCC 33152 strain) were grown for 48h at 37
degrees in WYO-alpha agar. Human monocytic cells U937 derived macrophages were cultured in RPMI-1640 containing 10 % inactivated fetal
bovine serum (FBS) with phorbol 12-mirystate 13-acetate (PMA, 10ng/mL)
for 24h. The cells were infected by L. pneumophilia (MOI=0-100) at 37 degrees for 1-3 days. miRNAs expressions (miR-218, miR-155 and beta actin)
were measured by real time PCR. Protein levels (Rictor and CathepsinB)
were evaluated by Western blotting. Cell death was detected using by LDH
cytotoxicity detection kit (TaKaRa).
Results: miR-218 and miR-155 were upregulated by L. pneumophilia
infection in U937 derived macrophages. L. pneumophilia infected cells
at MOI ＞ 100 (24 h) induced cell death, whereas low MOI of L. pneumophilia infection did not affect cellular viability. Knockdown of endogenous
miR-218 decreased Rictor protein.
Conclusion: Rictor is one of target gene of miR-218 in macrophage. L.
pneumophilia increases miR-218 level, which suppresses Rictor expression. Since Rictor regulates cellular survival and proliferation, L. pneumophilia infection might change cellular growth and viability via miR-218
regulation.
Invited Lecture
Purpose:Various pathogenic microorganisms, such as bacteria, fungi, and
amoeba, are soil-dwelling. Among these pathogenic microorganisms, the
free-living amoeba Balamuthia mandrillaris, is known to cause balamuthia
amoebic encephalitis (BAE). There have been about 200 fatal cases of BAE
reported from autopsies worldwide to date, with a further 8 fatal cases in
Japan.Despite this, the habitat and distribution characteristics of B. mandrillaris remain unknown. In this study, we aimed to detect B. mandrillaris
gene in soil samples from Aomori, Japan. Although rare in the rest of the
world, the volcanic ash – derived Andosol soil type is common in Japan,
which made standard DNA extraction from the soil extremely difficult.
Therefore, we also developed a new DNA extraction method that enables
the removal of allophane, to which DNA is adsorbed, from Andosol.
Methods:Beads fracturing and thermal fracture were used for DNA extraction from the soil. To remove allophane, skim milk was added to soil
samples. Isopropyl alcohol and polyethylene glycol were used for DNA
precipitation. PCR sensitivity was verified by creating a spike sample with
Acantamoeba sp. 1 – 1000/g added to the soil. Detection of B. mandrillaris
was performed by PCR targeting the 16s rRNA region.
Results:High-purity purification of soil DNA using only the present kits has
been difficult. Our present study demonstrates the feasibility of using PCR
via the addition of a 3-step purification method. B. mandrillaris genetic
material was detected in 3 of 13 soil samples. Sequence analysis confirmed
98 – 99% homology to pathogenic B. mandrillaris. Future studies should
attempt to simplify this soil DNA extraction method and apply it to widearea epidemiological investigation.
OL05-5
Role of miR-218 in human monocytic cell derived macrophage.
Keynote Speech
OL05-3
Yuya Hasunuma, Nozomi Hiyoshi, Moe Yokemura, Yoshikazu Tokuoka
Faculty of Biomedical Engineering, Toin University of Yokohama, Japan
41
Poster Presentation
Purpose: Streptococcus pneumoniae induce invasive pneumococcal disease (IPD) that may cause serious sequelae or be fatal.We examined the
current status of IPD as well as capsular typing involved in the prediction
of vaccination efficacy, primarily based on the results of an epidemiological survey. Materials and methods: 17 strains were isolated from IPD patients since January 2010 to June 2016. We examined clinical background
of patients, antimicrobial susceptibility of strains, genotypes of penicillinresistant Streptococcus pneumoniae (PRSP) by PCR, capsular typing by the
Multiplex PCR, and Sequence Type (ST).Results: A total of 17 strains were
isolated from blood, and one case was isolated from both blood and cerebrospinal fluids. Patients with underlying disease were 82.4%. The median
age was 69 years old. The 30-days mortality rate was 52.9%.All strains
isolated from blood were susceptible to penicillin (PCG) in MIC results of
<= 2µg/ml. One strain was isolated from cerebrospinal fluids was PRSP in
MIC results of 0.5µg/ml. The prevalence rate of penicillin resistance genes
pbp1a +2x +2b , pbp1a +2x , pbp2x +2b and pbp2x was 41.2% (7 strains),
18% (3 strains), 12% (2 strains) and 2% (5 strains), respectively. The prevalence rate of macrolide resistance genes ermB , mefA and ermB +mefA was
71% (12 strains), 6% (1 strain) and 23% (4 strains), respectively. The most
isolated serotype was 6B with 17.6% (3 strains) followed by 15B, 19A, 23F
with 11.8% (each 2 strains). The pneumococcal polysaccharide vaccine
(PPV23) and the pneumococcal conjugate vaccine (PCV13) covered 70.7%
and 58.8%, respectively. As for the ST, 3,111,63,199,2,923 was 11.8%
each 2 case. Conclusion: Surveillance of pneumococcal infections including IPD is essential for monitoring the beneficial effects of pneumococcal
conjugate vaccine (PCV) implementation.We conclud that epidemiological
studies are needed in addition to vaccination, which is indispensable for
the control of IPD.
Oral Presentation
Many antimicrobial agents have been used for growth promotion of livestock throughout the world. Thereby, extended-spectrum ß-lactamases
(ESBL) producing Enterobacteriaceae is often isolated from chicken meat
in Japan. Moreover, the increase in ESBL producing Enterobacteriaceae
colonization in healthy adults is a significant concern. In this research, we
compared molecular epidemiological characterization of the third generation cepharosporins resistance Enterobacteriaceae between domestic and
imported broilers. Forty-two samples (28 domestic and 14 imported) were
purchased from retail stores in Tokyo and Kanagawa. The homogenized
sample was spread on MacConkey ager with 2 μ g/mL cefotaxime and incubated at 37°C for 18-24 hours. Isolates were identified with the Api20E
test system. The MIC was measured with an ager plate dilution method
involving ampicillin, cefazolin, cefmetazole, ceftazidime, cefotaxime, gentamycin, amikacin, and ciprofloxacin. ESBL and plasmid-mediated AmpC ßlactamase (pAmpC) genes were detected with PCR amplification of bla CTX-M,
bla TEM, and bla SHV genes and with six main groups of pAmpC-type genes, as
described previously. Cefotaxime resistant Enterobacteriaceae was isolated
from 68 % (19/28) of the domestic and from 64 % (9/14) of the imported
samples. Moreover, 24 and 17 strains were collected from the domestic
and the imported, respectively. The CAZ resistance rate of the strain in the
domestic broiler, 50.0 %, was higher than that in the imported, 33.3 %.
However, the CPFX and GM resistance rate of the strain in the domestic
broiler was lower than those in the imported: 20.8 % vs. 58.8 % for CPFX
and 8.3 % vs. 47.1 % for GM. The PCR amplification measurement indicates that the high CAZ resistance rate of the strain in the domestic broiler
is due to high frequency of its pAmpC-type gene. Consequently, we need
to investigate the dissemination of the plasmid coding broad-spectrum
cepharosporins resistance mechanisms with not only CTX-M type of ESBL
but also pAmpC genes.
Case Conference
1 Section
of Infection Diseases, Laboratory Division of Clinical Support, Hiroshima University Hospital, Japan
2 Sect.
of Infection Diseases, Laboratory Div. of Clinical Support, Hiroshima Univ. Hosp.
3 Dept.
of Infectious Diseases, Hiroshima Univ. Hosp.
4 Div. of Laboratory Medicine, Hiroshima Univ. Hosp.
Keynote Speech
OL06-1
Establishment of a hemolysis index cut-off value for determining blood
recollection
Deciding blood recollection by objective evaluation of the degree of hemolysis
OL06-2
Mitsuaki Nishioka1, Akiyo Ishiguro1, Toshihiko Kobayashi1, Aki Masakane1, Naoko Okayama1,
Hidekazu Mizuno1, Yutaka Suehiro2, Takahiro Yamasaki2
Chitoshi Sato1, Makiko Suzuki2, Akira Ishih3
1 Department
of Clinical Laboratory, Okazaki city Hospital, Japan
2 Department
of Medical Technology, Shizuoka College of Medical Care Science
3 Department
of Infectious Diseases, Hamamatsu University School of Medicine
1 Department
of Clinical Laboratory, Yamaguchi University Hospital, Japan
2 Dept.of
Oncology and Laboratory Medicine, Yamaguchi Univ. Graduate School of Medicine
Invited Lecture
Special Lecture
Recently, the rate of parasitic infections has dramatically decreased in
Japan, and there are few opportunities to perform parasitic examination in
most of medical laboratories as a result. Such a public health improvement
is good for us, which situation has, however, led to difficulty of succession
of the knowledge and the technique about the examination of parasites
to a new generation.Parasitic infections are spread especially around the
developing countries, and hence Japanese tourists have an opportunity of
parasite infection anytime in those countries as well as foreign tourists,
and an examination of parasites is needed in Japanese medical centers.
Intestinal parasites being common in the developing counties are usually
detected and identified by microscopic observation of stool samples. Microscopic observation to detect parasites is important method for medical
laboratories, but there are some problems including difficulty of preservation of positive samples.It is most important to examine intestinal parasite
eggs in fecal materials in the laboratory, and hence we made an attempt at
preparing permanent slides including parasite eggs which will be used in
education at hospitals and universities. We would like to report some results about morphological changes of the external and internal structures
of eggs in the process of fixation, dehydration and embedding.
Educational Lecture
Background Hemolysis is usually caused by inadequate blood collection
technique and affects various laboratory data. Although the hemolysis
index (HI) value is used to indicate the degree of hemolysis, an appropriate
HI cut-off value to determine blood recollection has not been established.
Therefore, we performed this study to establish a cut-off value of HI for determining blood recollection. Methods Routine venous blood samples were
collected into plastic evacuated serum separator test tubes. The serum
samples were analyzed for clinical chemistry analytes using a BM6070
automated analyzer (NIHON DENSHI KABUSHIKIKAISHA (JEOL Ltd.)),
which automatically measures HI using a spectrophotometric technique.
The amount of free hemoglobin (Hb) in these samples was measured using
a sulfhemoglobin assay kit (Wako Pure Chemical Industries, Ltd.). Results
(1) There was a linear relationship between the HI value and the amount
of free Hb (HI = 0.0677 x Hb (mg/dL) – 0.0277).(2) Hemolyzed blood
samples were prepared from 18 healthy volunteers. Simple linear regression analyses showed that 19 clinical chemistry analytes correlated with
Hb concentration with a correlation coefficient of > 0.8 (absolute value).
Of them, multiple linear regression analyses resulted in significant correlations of Hb concentration with LD, AST, K, Fe, and Na values. HI values
reflecting the upper or lower limit of the reference range of each clinical
chemistry analyte were 6.4 for LD, 19.5 for AST, 25.9 for K, 94.7 for Fe,
and 124.7 for Na. We determined 6.4 to be the minimum HI value for
deciding on blood recollection.(3) Analyses of blood samples from about
54,800 patients showed that an increase of 6.4 in the HI value resulted in
an increase of 122 IU/L for LD, which was consistent with the reference
range of LD (98 IU/L). Conclusion We recommend a HI value of 6.4 as a
cut-off value for determining blood recollection.
Symposium
OL06-3
Preparing permanent microscope slides for observation of
parasite eggs
Medical technology in disaster medicine
Consideration from a viewpoint of the experience of Japan
Medical Team for Disaster Relief activity
Chitoshi Sato1, Tomokazu Najima2, Hiroaki Yamazaki5, Rika Shimizu3, Shoji Uga4
Case Conference
1 Department
of Clinical Laboratory, Okazaki city Hospital, Japan
2 St.
Mary's Hosp.
3 Division
of Central Clinical Laboratory, Mie University Hospital
4 Faculty
of Nursing, Kobe Women's Univ.
Oral Presentation
The Japanese Government has established a Japan Disaster Relief Team
for an assistance of overseas disaster. This team consists of three subteams i.e. Rescue, Medical and Expert. Among them, we have registered to
the Medical Team (Japan Medical Team for Disaster Relief; abbreviate to
JMTDR). The JMTDR was first dispatched to Cambodia in 1979 and these
kind of volunteer activities are continued thereafter in foreign countries.
The JMTDR, however, is different from the Disaster Medical Assistance
Team (known as DMAT) which deals only with disasters that occurs in
Japan. Some of us have participated in JMTDR as a medical laboratory
scientist. Those were Pakistan in 2010 (water flood) and Nepal in 2015
(earthquake). The major difference of these activities was the equipment.
In the case of JMTDR activity in Nepal, we equipped both ward function
and operation equivalent which regulated by type 2 of the FMT standard,
WHO. From these experiences, we would like to talk about the role, problem and/or inspection items from the viewpoint of Medical Technology in
respective disasters.
Poster Presentation
42
Poster Presentation
Staphylococcus haemolyticus endocarditis: a case report
Staphylococcus haemolyticus endocarditis
PA-02
A rare case of Tsukamurella paurometabola infection in
immunocompromised patient
Case report
Yi-Chu Lo
KAI-HSIANG LIN1,2, JUI-HSIANG WANG1, JU-HUEI CHIEN1
Department of Cardiology,Tainan Municipal Hospital, Tainan, Taiwan
1 Taichung
Tzu Chi Hospital, Buddhist Tzu Chi Medical Foundation, Taiwan
2 Department
of Life Sciences, National Chung Hsing University, Taichung, Taiwan, ROC
Educational Lecture
Discussion
Once blood culture is positive with coagulase-negative staphylococci
(CoNS), especially more than two sets of blood culture positive with CoNS,
the possibility of bacterial endocarditis should be kept in mind. In this setting, heart echo, especially transesophageal heart echo, may be performed
to diagnose bacterial endocarditis.
Kluyvera ascorbata cholecystitis: a case report
Kluyvera ascorbata cholecystitis
PA-04
Nosocomial Enterobacter cloacae endocarditis: a case report
Enterobacter cloacae endocarditis
Yi-Chu Lo
Department of Cardiology,Tainan Municipal Hospital, Tainan, Taiwan
Introduction
Kluyvera ascorbata infections are rare in the literature. Herein, we reported a case of cholecystitis caused by K. ascorbata.
Introduction
The case numbers of Enterobacter cloacae endocarditis were rare in the
literature. Herein, we reported a case, the first case of E. cloacae and even
gram-negative bacilli endocarditis in this hospital.
Discussion
Kluyvera is known as its chromosome carrying CTX-M-type extendedspectrum beta-lactamase (ESBL) genes; hence, all Kluyvera should be
regarded as ESBL-producing organisms. In this case of cholecystitis, both
E. cloacae and C. freundii were susceptible to ceftriaxone, but K. ascorbata
was intermediate-resistant to ceftriaxone; hence, infection caused by K.
ascorbata might be the cause of ceftriaxone treatment failure. Herein,
we suggested that carbapenems may be given to used to treat infections
caused by Kluyvera.
45
Poster Presentation
Case report
A 47-year-old male patient was admitted under impression of diabetic
ketoacidosis on 21 November 2015. He had underdying diseases of liver
cirrhosis, hypertension, and diabetes. After admission, only conservative
therapy, including subcutaneous insulin injection, was given. On 27 November, nosocomial fever occurred, and then vancomycin and ceftazidime
were given as empiric antibiotics. Thereafter, fever had decreased gradually and subsided on two days later. On the same day, a transthoracic
heart echo was performed and revealed calcified aortic valve (AV) with
mild aortic stenosis (AS) and aortic regurgitation (AR) as well as highly
suspected vegetations on AV. On 28 November, a transesophageal heart
echo was performed and revealed multiple vegetations on AV with mild AS
and AR. On 30 November, all four blood cultures taken on 27 November,
the time of blood culture drawn was 13:40, 14:10, 17:20, and 17:59, respectively, were positive with E. cloacae . All four E. cloacae isolates had the
same antimicrobial susceptibility testing results, including resistance to
cefazolin, cefuroxime, and amoxicillin-clavulanate as well as susceptibility
to ceftriaxone, ceftazidime, piperacillin-tazobactam, gentamcin, amikacin,
levofloxacin, trimethoprin-sulmethoazole, and tigecycline. Consequently,
nosocomial E. cloacae endocarditis was diagnosed according to the modified Duke crteria, and then doripenem was given for definitive antibiotic
treatment. On 24 December, a 28-day treatment course was completed,
and the patient was discharged.
Discussion
According to the modified Duke criteria, persistently positive blood cultures with first and last blood culture taken at least one hour apart are
one major criteria to diagnose infectious endocarditis. Herein, we suggest
heart echo should be performed to diagnose infectious endocarditis once
encountering persistently positive blood cultures with gram-negative bacilli.
Oral Presentation
Case report
A 78-year-old male patient was admitted under a diagnosis of acute
cholecystitis complicated with septic shock and multiple organ failure on
16 March 2014. He had underlying diseases of coronary artery disease
and liver cirrhosis. After admission, ceftriaxone plus metronidazole was
given as empiric antibiotic. On 17 March, abdominal echo was done and
revealed gallbladder distension with sludge, and then percutaneous gallbladder drainage was performed. However, the patient's condition was still
gone downhill persistently. On 20 March, ceftriaxone plus metronidazole
was shifted to doirpenem. On 22 March, bile culture taken on 17 March
was positive with Kluyvera ascorbata, Enterobacter cloacae, and Citrobacter freundii. In vitro susceptibility testing results revealed K. ascorbata was
intermediate-resistant to ceftriaxone, but both E. cloacae and C. freundii
were susceptible to ceftriaxone. All three isolates were susceptible to ceftazidime, piperacillin-tazobactam, and carbapenems. After treatment with
doripenem, the patient's condition had improved gradually. However, total
bilirubin elevated persistently to 11.4 mg/dl on 23 March. On 24 March,
repeated bile culture did not reveal any organism. On 7 April, total bilirubin elevated to 32.7 mg/dl. On 8 April, doripenem was shifted to tigecycline. On 12 April, the patient expired due to multiple organ failure.
Case Conference
Yi-Ling Tai
Department of Internal Medicine, Tainan Municipal Hospital, Tainan, Taiwan
Symposium
PA-03
Special Lecture
Tsukamurella paurometabola is a weakly acid-fast, pleomorphic grampositive bacterium found in soil. To our knowledge, this extraordinary
case of bloodstream infection in immunocompromised patient is rarely
reported. It is diffcult to identified this Tsukamurella sp. by conventional
biochemical identification methods. The biochemical profile of this organism did not includ in VITEK 2 system and BD Crystal Rapid Gram-Positive
ID Kit. The proteins profile neither in VITEK MS and Bruker MALDI-TOF
detection systems. To correct identify this organism , the 16S ribosomal
DNA was amplied by PCR method,and also subjected to automated DNA
sequencing. The antibiotic susceptibility testing was performed by disk
diffusion method. Microbiological identification of Tsukamurella sp. is
challenging and laborious. Rare cases of this pathogens are isolated in
immunosuppressed patient, especially patients with central venous catheters or lung infection. Until now, there is no CLSI Guidline to treat this
pathogen infection.We treated this patient with empiric antibiotics , and
the patient was discharged after two months therapy. The potential role
of Tsukamurella paurometabola infection in bloodstream is still unknown.
Therefore, further study is necessary which will be benefit for physicians
to treat this infection.
Invited Lecture
Introduction
The case numbers of Staphylococcus haemolyticus endocarditis were rare
in the literature. Herein, we reported a case.
Case report
A 90-year-old male patient with uremia received hemodialysis treatment
was admitted due to fever on 20 April 2014. After admission, piperacillintazobactam was given for treatment. On 23 April, transthoracic heart
echo revealed a vegetation with a size of 2.5 cm × 1.83 cm located on the
mitral valve. On 24 April, the two sets of blood culture taken on 20 April
were positive with S. haemolyticus , identified by Phoenix automachine
(Becton Dickinson and Company, Sparks, MD). Antimicrobial susceptibility testing done by Phoenix automachine revealed the two isolates were
resistant to oxacillin with minimal inhibitory concentration (MIC) 0.5
mg/L, and susceptible to clindamycin with MIC<=0.5 mg/L , fusidic acid
with MIC<=0.5 mg/L, linezolid with MIC<=1 mg/L, and vancomycin with
MIC<=1 mg/L. Consequently, S. haemolyticus endocartitis was definitively
diagnosed according to the modified Duke criteria, and then vancomycin
was given for treatment. On the same day, another two sets of blood culture were taken again. Thereafter, fever still persisted despite treatment
with vancomycin. On 28 April, the two sets of blood culture taken on 24
April were also positive with S. haemolyticus , and the results of antimicrobial susceptibility testing were the same as previous isolates. Because the
patient's condition was gone downhill persistently, valve replacement was
suggested; however, the family refused this operation. Finally, the patient
expired on 5 May 2015.
Keynote Speech
PA-01
Keynote Speech
PA-05
Rhodotorula in blood culture: significance or non- significance
Rhodotorula mucilaginosa, contamination, case report
PA-06
Chung-Hsiang Hsu1, Yin-Tai Tsai3, Meng-Huei Liang3, Wei-Ming Chi3, Wen-Shyang Hsieh3,5,
Hsiao-Wei Wang3,4,5, Chi-Hung Lee1,2, Yung-Ching Liu1,2
Chen Ya Fen, Kuo Fu Mei, Chen Yi Jyun, Wu Yi Chi, Lee Shwu Yea, Chan Yu Jiun
Division of Microbiology, Department of Pathology Laboratory Medicine, Taipei Veterans General Hospital, Taiwan
1 Division
of Infectious Disease, Department of Internal Medicine, Taipei Medical University, Shuang Ho Hospital, New Taipei
city, Taiwan, Taipei Medical University, Shuang Ho Hospital, Taiwan
2 Division
of Infectious Diseases, Department of Medicine, School of Medicine, College of Medicine, Taipei Medical University,
Taipei, Taiwan
3 Department
of laboratory medicine; Taipei Medical University–Shuang Ho Hospital, New Taipei city, Taiwan
4 Department
of education, Taipei Medical University-Shuang Ho Hospital, New Taipei City, Taiwan
5 School
of Medical Laboratory Science and Biotchnology, Taipei Medical University, Taipei Medical University, Taipei, Taiwan
Invited Lecture
From 3 May to 30 June 2015, 31 patients in a hospital in Taipei were
culture-positive for Ralstonia pickettii from their blood specimens. They
had been exposed to 0.9% saline solution which was manufactured by
one company (YF Chemical Corp). Two of the 17 saline solution samples
collected at the hospital were found to contain Ralstonia pickettii. The
isolates were analyzed by MALDI-TOF MS for microbial identification. In
order to investigate the relatedness of the clinical isolates and the isolates
from saline solution, all isolates were genotypically related analyzed by
pulsed-field gel electrophoresis (PFGE) analysis. The PFGE (DNA-digested
with SpeI restriction endonuclease) result showed that the R.pickettii from
patients and the isolates from saline solution were related.
Special Lecture
Educational Lecture
Rhodotorula species infection, which was mainly presumed as contamination or colonization of specimen, is much less common than other yeast.
However, the number of this infection has clearly increased during the
last few years. Herein, we report two patients whose blood culture grew
Rhodotorula mucilaginosa. The first patient is a 38-year-old male with
obstructive sleep apnea under CPAP use and asthma. He presented with
fever, cough to ER with leukocytosis (12900/uL) and neutrophil predominant (94.2%). Meanwhile, R. mucilaginosa was cultured in one of the two
isolated blood bottles. No antifungal regimen was given and his symptom
relieved later. The second patient is an 80-year-old bedridden male due
to old CVA with CAD status post CABG and ESRD under hemodialysis. He
presented to out-patient department with right upper chest swelling for 1
week and was referred to ER later. Leukocytosis (29400/uL) was noted but
there was no left shift. The enlarging tumor was diagnosed as necrotizing
fasciitis by image and he received debridement + fasciectomy twice and
under Tygacil + Tapimycin use till the end. There were only negative finding in blood culture on admission day 1 and contamination in wound culture during admission day 1 and day 9. On admission day 18, the patient
presented with septic shock and he expired on the next day. Nevertheless,
two sets of blood culture from central vein catheter were performed on
the day before the patient expired. One of them grew nothing while the
other grew R. mucilaginosa. While the culture of Rhodotorula species from
a sterile site, such as blood, is usually indicative of infection. But in these
cases we should suspect contamination of Rhodotorula during the culture,
when it is not suggestive of infection.
Symposium
PA-07
A Nosocomial Outbreak of Ralstonia pickettii by
Contaminated Saline Solution in a Taipei Hospital.
Effect of Carbon Dioxide on Biochemical Phenotype of Capnophilic Proteus mirabilis
At a Regional Hospital
Effect of Carbon Dioxide on Biochemical Phenotype of Capnophilic Proteus mirabilis
PA-08
Case report-Diphyllobothriasis
Parasites infection of dietary
Case Conference
Oral Presentation
Poster Presentation
Chao-Tai Lee
HSINYI CHOU, CHUANCHI CHAN
Taiwan Society of Laboratory Medicine, Taiwan
Department of Laboratory Medicine, Taipei Tzu Chi Hospital, Buddhist Tzu Chi Medical Foundation, Taiwan
Purpose
A rare strain of carbon dioxide-dependent Proteus mirabilis was isolated
from urine specimen at a regional hospital, the first isolate of capnophilic
P. mirabilis at this hospital. The strain showed non-swarming and did not
produce H2S property. This study was conducted to study the biochemical
phenotype of the isolate in different CO2 atmosphere concentrations.
Methods
The capnophilic P. mirabilis as study strain (PM103), another patient in
this hospital for urine culture result non-capnophilic P. mirabilis as a control strain (PM104), and the P. mirabilis ATCC 7002 as a reference strain.
Three strains were inoculated on blood agar plate (BAP) and eosin-methylene blue agar (EMB), and the strains of biochemical test using conventional biochemical tests and API 20E. All tests were placed in ambient air, and
in 2.5%, 5%, 7.5% and 10% CO2 atmosphere, respectively to investigate
the effect of the biochemical characteristics in different CO2 conditions.
Results
The PM103 didn't grow in ambient air, but grew at CO2 environment
with non-swarming. In 5% and 7.5% CO2 atmosphere grow best, but the
CO2 concentration continues to increase to 10% content the isolate grew
poor. Regardless of the colonies grew on BAP and at any CO2 atmosphere
environment which had non-swarming. The PM104 and P. mirabilis ATCC
7002 could grow on BAP and EMB in any conditions with swarming.
The results of the PM103 of conventional biochemical tests and API 20E
showed no difference and without H2S property, wherever in 2.5%, 5%,
7.5% and 10% CO2 conditions. The PM104 and P. mirabilis ATCC 7002
showed same results of conventional biochemical tests and API 20E with
H2S property, wherever in 2.5%, 5%, 7.5% and 10% CO2 conditions.
Conclusion
A rare capnophilic P. mirabilis strain in different CO2 atmosphere does not
change the characteristics which non-swarming and doesn't produce H2S.
Along with the advancement and improvement of medical and health
habits,parasites infection is unusual in Taiwan. With the food culture,
parasitic infections are still inevitable. In 2016, a female patient can not
discharges parasite through stool solution, so have to pulls parasite out
manually. There are three parasites, the longest parasite is 43 cm, and
total weights are 3 gram of the three parasites. Parasitic segments are
broader than long, the genital pore above the midline of the parasitic segments, in according to parasites patterns determine that the pseudophyllidea preliminary. We use MIF stain to observe the eggs from the patients
feces, and the observation shows, the eggs are filled with yolk, double shell
and small nodules. However the existence of egg cover is not clear. We use
three staining methods including Papanicolaou stain, Trichrome stain and
Merthiolate-Iodine-Formalin stain to stain the parasitic segments. After
staining, we determine that the parasites are Diphyllobrothrium latum.
We ask the patient's dietary habits, and find out that she likes eat beef and
Japanese cuisine, therefone we sugges that she change the dietary habits
from raw food to cooked- food based, in order to avoid parasite infection
in the future.
46
Amino Acid Determination of the Dominant Protein Isolated
From Fertilized, Corticated Ascaris lumbricoides Egg
PA-10
Infection Rate Among Lymnaea spp. snails by
Cercarial Emergence in Barangay Cawongan Padre
Garcia, Batangas
Gregorio L. Martin I1, Carmela N. Bautista1, Adam Lesner A. Bernabe1, Ana Isabela C. De Guzman1,
Yves Aaron Julian Q. De Ocampo1, Alyssa Denise L. Sevilla1, Aedrick B. Yanga1, Leonardo A. Guevarra Jr.2
Gregorio L. Martin I, Glorianne Marie C. De Guzman, Emil Victor J. Lizarondo, Alyanna Marie B. Manego,
Yarah Jenna J. Paypa, Samantha C. Salaya, Tiffany Ella Rose DC. Say, Jess Mari H. Yara
1 Medical
Technology, University of Santo Tomas, Manila Phillipines
2 Biochemistry,
University of Santo Tomas, Manila, Philippines
Medical Technology, University of Santo Tomas, Manila, Phillipines
Keynote Speech
PA-09
Fasciola hepatica , commonly known as Sheep Liver Fluke, is the known
Special Lecture
cause of one of the global concerns in the health sector known as Fascioliasis. Its life cycle involves two hosts: fresh water snails belonging to the
Lymnaea spp., its intermediate host and herbivorous animals, including
humans, its definitive host. The study focused in determining the natural
infection rate of the Fasciola sp. in Lymnaea spp. through cercarial emergence. The snails, with an average weight of 0.090 g ( ± 0.16) and average height of 8.38 mm ( ± 3.20), from Barangay Cawongan, Municipality
of Padre Garcia, Province of Batangas were placed individually in a perforated wide-mouthed topped containers and were brought in a laboratory
controlled set-up consisting of adequate light and scheduled water changing. No cercariae emerged on a span of four weeks. However, the second
batch of snails (n=429) revealed the presence of free living protozoans,
operculated ova, Ascaris spp. egg, and cyst-like structures, and sporocystlike larval forms.
Invited Lecture
Ascariasis affects one billion people worldwide and this condition remains
asymptomatic in most cases. Despite having high prevalence, it is considered as one of the Neglected Tropical Diseases (NTDs).Even with technological advancements, the most common laboratory diagnosis is still
through the microscopic identification of Ascaris lumbricoides eggs from a
stool sample. Thus, this study was conducted to determine the amino acid
composition of the protein coat of fertilized, corticated A. lumbricoides
eggs. Fertilized, corticated eggs recovered from the stool sample were
concentrated and decorticated in order to separate the dominant protein
from the eggs. The concentrated eggs were decorticated using ascending
concentrations of Phosphate-Buffered Saline with Tween 20. Micro-Bradford assay of the different supernatants after egg decortication revealed
increasing absorbance suggestive of increasing protein content (r=0.7114).
Microscopic images of eggs before and after decortication were also noted.
High Performance Liquid Chromatography results exhibited peaks which
belong to arginine and tyrosine with concentrations of 1.627 micromol/
mL and 1.678 micromol/mL, respectively. Moreover, these two amino
acids are also present in proteins which are mammalian trypsin and chymotrypsin inhibitors. Information obtained from this study is significant in
developing Rapid Diagnostic Kits for parasitic identification.
Educational Lecture
Development of Ascaris lumbricoides in the Gut of
Periplaneta americana species
PA-12
In vitro Ovicidal Activity of Lansium domesticum (Meliaceae)
Bark Extracts on Ascaris lumbricoides
Medical Technology, University of Santo Tomas, Manila, Phillipines
1 Medical
Technology, University of Santo Tomas, Manila, Phillipines
2 Pharmacy,
University of Santo Tomas, Manila, Philippines
47
Poster Presentation
This study evaluated the ovicidal effect of Lansium domesticum through
in vitro testing against A. lumbricoides . The methanolic bark extract of L.
domesticum was obtained through percolation and the crude extract was
fractionated using chloroform, ethyl acetate, and n-butanol. These extracts
and fractions were evaluated through phytochemical testing and quantification of condensed tannin content (CTC) using gallic acid as standard.
The results shows that crude extract and ethyl acetate fraction yields the
highest condensed tannin content while the chloroform and n-butanol
(p=0.289) yields to almost the same condensed tannin content. The CTC
of the crude extract, chloroform, ethyl acetate, and n-butanol fraction are
1.110, 0.557, 0.871 and 0.434mg GAE/g sample, respectively. The Egg
Hatch Test was used to assess the effect of each crude extract and fractions on the viability of A. lumbricoides eggs. After 48 hours, Albendazole
(60mg/mL) and Crude extract (120mg/mL) showed similar ovicidal effect
(p=0.104). However, it was only after 96 hours when Ethyl acetate (60mg/
mL) and Chloroform (120mg/mL) exerted same activity as the standard
(p=0.470). All the other extractives and normal saline solution similarly
(p=0.143) did not show ovicidal effect during the observation period. The
Ova Analysis was used to assess the effect of the extractives on larvae
development after incubation in soil set-up. Albendazole (60mg/mL) is the
most effective test solution among the groups (p=0.124) followed by the
crude (120mg/mL), ethyl acetate (60 mg/mL) and chloroform (120mg/
mL) which shows similar effect to the development of eggs (p=0.052). All
the other extractives and the normal saline solution did not exhibit the desired effect (p=0.321). These in vitro studies revealed that the crude, ethyl
acetate and chloroform (120mg/mL) of L. domesticum exhibited ovicidal
effect against A. lumbricoides and therefore may be tapped as a potential
alternative anthelminthic drug.
Oral Presentation
Currently, intestinal helminthiasis affects an estimated 500 million to 1
billion people yearly, at least 400 million children of school age. Recent
studies show that the cockroaches are actual carriers of medically relevant
parasites causing serious health problems. They act as mechanical and
biological vectors as they carry parasites in their external and internal
parts, disseminating them in the environment from ingestion of contaminated human feces. Eighty-four cockroaches were collected for the study.
They were placed on an artificial environment containing bread with stool
positive for Ascaris lumbricoides . Any gross morphological changes that
occur to them were monitored in a four week interval after which the
gut was removed, dissected, and examined for the presence of various
developmental changes of the parasite. Based on the results, there was an
increased mortality rate observed in the experimental group over a three
week monitoring period from week 1 (42.86%) up to week 3 (76.19%).
All infected cockroaches showed a marked increase in their weight. Posteuthanized infected cockroaches revealed the presence of motile Ascaris
larva suggestive of continued viability of Ascaris lumbricoides. In conclusion, local cockroaches were capable of supporting the life of Ascaris lumbricoides . There was a positive correlation between the weight of the cockroach and the severity of the infection. Mortality rate of the cockroaches
increased possibly due to the lack of adequate resources inside an isolated
environment and succumb to infection. In the light of these findings the
researchers recommend the measurement of the larva developed within
the gut through ocular micrometry and associate it with its life span and
viability.
Case Conference
Gregorio L. Martin I1, Gina C. Castro2, Paola Louise R. Palma2, Fleur Jeizl P. Perez2,
Maria Godesa F. Refuerzo2, Michelle Nahty T. Reyes2, Kristiana Margarette C. Romina2,
Cyla Mariel C. Salvadora2
Gregorio L. Martin I, Renz R. Bautista, Katherine Ann T. Basco, Pio A. Capistrano II,
Aleja Grace S. Dungca, Stephanie Bernadette D. Ellao, Carlos L. Erquiaga
Symposium
PA-11
Keynote Speech
PA-13
Gram Staining Offers Simple and Real-Time Diagnosis of
Malassezia folliculitis: A Single-Center Series
Malassezia folliculitis, Gram stain, Real-Time diagnosis
PA-14
Meng-Huei Liang1, Ruo-Tzu Li1, Wei-Ming Chi1,3, Wen-Shyang Hsieh1,2,3, Hsueh-Hsia Wu3,
Woan-Ruoh Lee4,5,6, Wei-Ting Tu4, Yi-Hsien Shih4,5,6
Yu Jiun Chan1,2, Ping Feng Wu1, Su Pen Yang1, Mei Lin Lin1, Yi Chi Wu1, Fu Der Wang1, Yu Fan Juan1
1 Taipei
Veterans General Hospital, Taiwan
2 National
Yang Ming University
1 Department
of laboratory medicine, Taipei Medical University, Shuang Ho Hospital, Taiwan
2 Department
of education, Taipei Medical University-Shuang Ho Hospital, New Taipei City
3 School
of Medical Laboratory Science and Biotechnology, Taipei Medical University, Taipei
4 Department
of Dermatology, Taipei Medical University-Shuang Ho Hospital, New Taipei City
5 Department
of Dermatology, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan
6 Graduate
Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan
Invited Lecture
Background: Matrix-assisted laser desorption ionization-time of flight mass
spectrometry (MALDI-TOF MS) system may become a fast and reliable
tool for rapid fungal identification in routine clinical laboratories. The aim
of this study was to evaluate the efficacy of Vitek MS in the identification
of clinical isolates of Candida species cultured from blood or sterile body
sites from the patients in a major teaching hospital of northern Taiwan.
Special Lecture
Educational Lecture
Background: Malassezia folliculitis is a common skin disease caused by the
invasion and proliferation of the fungus, Malassezia spp. in the hair follicle.
It is usually diagnosed on clinical ground, but its distinction from common
mimickers, such as acne vulgaris and bacterial folliculitis, is crucial. Gram
staining visualizes both bacteria and the blastospores of Malassezia spp.;
therefore, it has the potential to resolve diagnostic difficulties. However, it
is only rarely used in the past for such purpose. Its accuracy in diagnosing
Malassezia folliculitis is also unknown based on past literature.
Patients and methods: To determine the diagnostic accuracy of gram
staining in Malassezia folliculitis, we retrospectively included 32 patients,
between June 2015 and January 2016, in whom gram staining was performed to diagnose Malassezia folliculitis. We quantified the amount of
Malassezia blastospores on gram-stained slides and compared the results
with final diagnosis, which was determined by skin biopsy (if available)
and positive treatment response (if a skin biopsy was not done). We also
calculated the sensitivity and specificity of both gram staining and skin
biopsy.
Results: In the 32 patients, 26 (81.3%) were diagnosed with Malassezia
folliculitis, 2 (6.3%) were diagnosed with bacterial folliculitis, and 4 (12.5%)
were diagnosed with mixed bacterial and Malassezia folliculitis. In 5 patients, gram staining results were different from the final diagnosis. The
sensitivity and specificity of gram staining in the diagnosis of Malassezia
folliculitis were 84.6% and 100%, respectively, compared to the 75% sensitivity and 100% specificity of skin biopsy.
Conclusion: Gram staining seems an easy and under-utilized method to diagnose Malassezia folliculitis. The presence of Malassezia blastospores but
not bacteria is highly suggestive of Malassezia folliculitis. We encourage
large-scale studies and comparison studies to consolidate our findings.
Symposium
PA-15
Comparison of the Accuracy of Vitek MS and Conventional Vitek 2 System with DNA Sequencing
Analysis in the Identification of Clinical Candida Isolates Collected from Sterile Body Sites
Material/methods: Fifty-nine clinical isolates were analyzed by Vitek-2,
Vitek MSTM and internal transcriber spacer 1 (ITS1) rDNA sequencing.
The efficacy of the Vitek MS was determined by comparison with Vitek 2
and ITS1 rDNA sequencing analyses.
Results: According to the results of ITS1 rDNA sequencing analyses, the 59
isolates were the followings: 11 isolates of C. albicans, 10 C. tropicalis, 12 C.
parapsilosis, 10 C. glabrata, 4 C. krusei, 5 C. guilliermondii, 2 C. lusitaniae,
2 C. intermedia, 1 C. rugosa, 1 C. pararugosa and 1 C. haemulonii. By using
the gene sequencing as the reference method, the correct species identification rates for Vitek MS and Vitek 2 were both 91.5% (54/59). The only
isolate of C. pararugosa was misidentified by Vitek 2 and unidentified by
Vitek MS system. The other 4 Vitek 2 misidentified isolates were correctly
identified by Vitek MS, and the other 4 Vitek MS unidentified isolates were
correctly identified by Vitek 2.
Conclusions: In conclusion, the Vitek MS system is comparable to Vitek 2
in the identification of clinically important isolates of Candida species.
Comparison of Routine Microbiological Methods for Identification of Candida spp.
Consistency between MALDI-TOF, Phoenix and Real-time PCR for Identification
of Candida spp
PA-16
Chuan-Ru Wang1, Yin-Tai Tsai1, Tzu-Ying Lee1, Wei-Ming Chi1,3, Wen-Shyang Hsieh1,2,3,4, Yu-Chun Sun4,
Shiao-ping Huang4
Laboratory investigation of suspected in-vivo acquisition of
antibiotic-resistant genes
In-vivo acquisition of antibiotic-resistant genes
Chao-Tai Lee
Clinical laboratory, Taiwan Society of Laboratory Medicine, Taiwan
Case Conference
1 Department
of laboratory medicine, Taipei Medical University, Shuang Ho Hospital, Taiwan
2 Department
of education, Taipei Medical University, Shuang Ho Hospital, New Taipei City, Taiwan
3 School
of Medical Laboratory Science and Biotechnology, Taipei Medical University, Taipei, Taiwan
4 Department
of Medical Laboratory Sciences and Biotechnology, Fooyin University, Kaohsiung, Taiwan
Oral Presentation
Poster Presentation
Purpose
At a regional hospital in southern Taiwan, on 16 March 2014, one isolate
of carbapenem-resistant Providencia stuartii (CRPS, P2) and one isolate
of carbapenem-resistant Klebsiella pneumoniae (CRKP, K1) were isolated
from the same urine sample from a patient came from nursing home. In
addition, another isolate of CRPS (P1) was isolated from this patient on
14 February 2014. Because two isolates of carbapenem-resistant Enterobacteriaceae coexisted were unusual, we investigated the relationship of
antibiotic-resistant mechanisms of P2 and K1.
Methods
Pulsed-field gel electrophoresis (PFGE) was used for bacterial genotyping
of P1 and P2. PFGE patterns were interpreted as same (no band difference), similar ( ＜ =three-band differences), or different ( ＞ =four-band differences) strains. Polymerase chain reaction (PCR) and sequence analysis
were used for detecting antibiotic-resistant mechanisms. Plasmid analysis
and Southern Hybridization were used for detecting the location of antibiotic-resistant genes. Replicon typing and plasmid multilocus sequence
typing were used for plamid typing. Restriction fragment length polymorphism (RFLP) was used for detecting the relationship of plasmids.
Results
PFGE pattern revealed P1 and P2 were the same strain. A 200 kb IncA/C
plasmid was present in both P2 and K1, which carried genes of blaCMY-2,
qnr, and aac (6')-Ib-cr. RFLP revealed that the 200 kb plasmids of P1, P2,
and K1 were almost same.
Discussion
As a result of this study, the antibiotic-resistant plasmid of K2 might come
from P2 by in-vivo acquisition. Although clonal spread was an important
cause of high rate of antibiotic-resistant organisms in long-term care facilities, this study revealed that the spread of antibiotic-resistant genes was
also an important cause.
Candida albicans and other Candida spp. are the most common fungal
pathogens causing fungal infections. Opportunistic fungal infections are a
major cause of morbidity and mortality in immunocompromised patients
such as transplant recipients. The clinical features of invasive candidiasis
are nonspecific, making early diagnosis of invasive candidiasis difficult.
Recently, the incidence of healthcare associated bloodstream infections
caused by Candida spp. has increased in immunocompromised patients.
Non-cultural techniques used previously have lacked sensitivity and specificity. New methods has been applied as a rapid, accurate and inexpensive
method for the detection of fungal infection early in the course of disease.
This study compared Phoenix™ Automated Microbiology System and
MALDI Biotyper™ System, and the real-time PCR methods with primers
designed for identification of Candida albicans, Candida tropicalis, Candida
glabrata, Candida parapsilosis, Candida krusei, and Candida lusitaniae.
There were 180 clinical fungal isolates applied in this study. The samples
were examined with the three methods for identification. The consistency
between the both Phoenix system and Real-time PCR for identification
of C.albicans, C.glabrata, C.krusei, C.lusitaniae, C.tropicalis, C.parasilosis,
were 89%, 94%, 100%, 100%, 89%, and 73%, respectively.The consistency between the both MALDI Biotyper™ System and Real-time PCR fo
identification of C.albicans, C.glabrata, C.krusei, C.lusitaniae, C.tropicalis,
C.parasilosis, were 74%, 96%, 100%, 100%, 85%, and 73%, respectively. In
addition, the mass spectra of MALDI Biotyper™ System were applied for
constructing dendrogram, which shows six cluster groups with a default
critical distance level of 700. In conclusion, this study provides us the efficacies of Automated Microbiology System, MALDI Biotyper™ System, and
the real-time PCR methods for identification of Candida spp. from clinical
samples.
48
To investigate an outbreak caused by carbapenem and tigecycline-resistant
Enterobacter cloacae
Outbreak caused by carbapenem and tigecycline-resistant Enterobacter cloacae
PA-18
Identification of bacterial flora in pediatric UTIs: a comparison of results between VITEK2 and VITEKMS
A comparison between different technologies for faster identification of bacterial flora in paediatric urine
samples
Purpose
At a regional hospital in southern Taiwan, 11 isolates, including 8 from
community and 3 from hospital, of carbapenems and tigecycline-resistant
Enterobacter cloacae were isolated from September 2010 to April 2012.
Because they were unusual, we investigate suspected an outbreak caused
by E. cloacae by molecular methods.
Methods
Antimicrobial susceptibility testing was performed by E-test. The antibiotics tested included imipenem, meropenem, doripenem, and tigecycline. All
results were interpreted according to the criteria suggested by the Clinical
Laboratory Standards Institute 2015, but tigecycline was according to
that by the European Committee on Antimicrobial Susceptibilities Testing
2015. Pulsed-field gel electrophoresis (PFGE) was used for bacterial genotyping. PFGE patterns were interpreted as same (no band difference), similar ( ＜ =three-band differences), or different ( ＞ =four-band differences)
strains.
Results
All of the 11 isolates of E. cloacae were nonsusceptible to at least one of
imipenem, meropenem, and doripenem as well as resistant to tigecycline.
PFGE patterns revealed 5 genotypes were present, including 1 A (E1), 5 B
(E2-E6), 3 C (E7-9), 1 D (E10), and 1 E (E11).
Discussion
As a result of this study, these were outbreaks caused by two E. cloacae
clones, including genotype B and C. Herein, we suggest that investigating
outbreaks by molecular methods should be done once encountering unusual increasing isolates of the same species with multidrug-resistance.
Few processes present greater difficulties in the way of diagnosis than
the identification of an infecting microorganism and the most effective
therapy for it. The Meyer Children Hospital is currently carrying out a
three-hour growth test in broth medium and analyzed in turbidimetry
(ALFRED60, ALIFAX) in order to distinguish the positive samples from the
negative ones: positive urines (30000 CFU/mL) are then plated and incubated for at least 24 hours. In addition, the identification and the resulting
antibiogram from VITEK2 (Biomerieux) last at least 9 to 15 hours. So, the
report will be available in 48/72 hours after the arrival of the samples
at the laboratory, increasing the waiting time to give the right therapy
to the child. This research has considered the possibility of making the
identification straight from the culture medium for screening with the subsequent analysis of pellet from centrifuged samples and, finally, analyzed
by VITEK-MS (MALDI-TOF Technology, Biomerieux). In two months, 236
samples have been analyzed for authenticity, 52 of them resulted positive
at ALFRED60. If compared, the results between the two instruments for
identification VITEK2 and VITEK-MS almost overlap: for example GRAM +
and GRAM - identification looks similar. VITEK-MS is more accurate than
VITEK2 in terms of giving results of GRAM +: 15 instead of 7 (T Test: p ＜
0.05). As far as GRAM - concern, E. coli is the most representative bacteria
having been found positive in 38 samples with VITEK2 and 40 in mass
spectrometry. What makes them different is the time of identification:
mass spectrometry takes only a few minutes, no longer than 5 hours after
the arrival of a sample at the laboratory. Taking everything into account,
VITEK-MS is significant as it offers an earlier identification of the bacteria
providing a faster therapy to the child.
Higher Results of Group B Streptococcus Screening among Pregnant Women.
GBS infection has evolved in the high risk of severe illnesses and death during
vaginal delivery. Especially in newborn infants.
PA-20
The mechanisms of levofloxacin-resistant Haemophilus
parainfluenzae
Levofloxacin-resistant Haemophilus parainfluenzae
Chin-Lu Chang
Department of Infectious Diseases, Tainan Municipal Hospital, Tainan, Taiwan
Introduction
At a regonal hospital in southern Taiwan, we found the prevalence of
levofloxacin-resistant Haemophilus parainfluenzae is increasing gradually.
Henc, this study was conducted to investigate the mechanisms of levofloxacin resistance.
Methods
Three isolates (namely HP1, HP2, and HP3) of levofloxacin-resistant H.
parainfluenzae were chosen for this study. The polymerase chain reaction
(PCR) and sequencing were used to detect the mutations of quinolone
resistance-detemining regions (QRDRs), including gyrA, gyrB, parC, and
parE. The H. parainfluenzae T3T1 was used as control strain to compare
amino acid substitution.
Discussion
As a result of this study, the mechanisms of levofloxacin-resistant H.
parainfluenzae were the mutations of gyrA and parC. Compared to other
studies, the mutations of Ser84 and Asp88 of gyrA as well as Ser84 of
parC were the important causes resulting in levofloxacin resistance. In addition, the role of mutation of Asp420 of parE contributing to levofloxacin
resistance need further investigation. By the way, this study cannot exclude other antibiotic-resistant mechanisms, such as permeability barriers
and efflux pumps.
49
Poster Presentation
Results
HP1 had the mutations of Ser84Tyr and Asp88Tyr of gyrA as well as
Ser84Phe, Ser138Thr, Met198Leu, and Asp209Glu of parC. HP2 had the
mutations of Ser84Phe, Asp88Tyr, and Gly178Glu of gyrA, Ser84Phe and
Met198Leu of parC, as well as Asp420Asn of parE. HP3 had the mutations of Ser84Phe and Asp88Tyr of gyrA, Ser84Phe of parC, as well as Asp420Asn of parE. All three isolates had the mutations of Ser84 and Asp88
of gyrA and Ser84 of parC.
Oral Presentation
GBS infection has significantly evolved in the high risk of severe illnesses
and death during vaginal delivery. Especially in newborn infants, which
can cause neonatal sepsis, neonatal meningitis, pneumonia and neonatal
death or neurological sequela.
To screen for GBS during the third trimester, which is the 35th to 37th
weeks of pregnant women, a swab is typically obtained from the lower
vagina and rectum. The identifications of GBS by appearing the &beta hemolysis on CNA agar, 3% Catalase negative result and the arrowhead area
of CAMP test.
If it shows &beta hemolysis with no arrowhead area or &gamma hemolysis on CAMP test, but still suspects might be Group B streptococcus. Then
inoculated on Bile Esculin Agar (BEA) slant and 6.5% NaCl agar slant. The
female vagina discharge, male urethral discharge and urine specimen are
selected as parities to compare with pregnant women.
As shown below, the latest estimates generated over the last two years
(2013 to 2015) report that approximately 1228 of the 5731 pregnant
women (21.43%) were examined with culture-positive for Group B Streptococcus by prenatal vagina swab specimen. The survey indicates that
GBS infection possibility in pregnant women is significantly higher than
females with vagina discharge (226/1478, 15.29 %), males with urethral
discharge (58/1478, 3.92 %), the urine and foley specimen (1355/58380,
2.32 %).
The antibiotic of Clindamycin and Erythromycin have the highly and increasingly resistant results (CM: 66.3%, EM: 59.5% ) during recent years.
Higher isolation frequency of Group B Streptococcus among pregnant
women is emerged as the leading cause of baby at risk of developing GBS
disease. However, the GBS diagnosis provided by healthcare can monitor a
healthy pregnancy with effective treatments to reduce the newborn morbidity and mortality.
Case Conference
Yung-Chen Chien1, Chung-Pin Kao2
1 Central
Bacteriology Lab, Taipei City Hospital,Renai Brach, Taiwan
2 Division
of Laboratory at Yangming Branch of Taipei City Hospital, Taiwan
Symposium
PA-19
Educational Lecture
ANTeL-Assiatel/AITIC, Italy
Special Lecture
Claudia Finocchi
Clinical laboratory, Taiwan Society of Laboratory Medicine, Taiwan
Invited Lecture
Chao-Tai Lee
Keynote Speech
PA-17
Keynote Speech
PA-21
The antimicrobial susceptibility patterns of community-acquired respiratory tract
organisms in southern Taiwan
Antimicrobial susceptibility patterns of community-acquired respiratory tract organisms
PA-22
Port-a-cath-related Bacillus cereus bacteremia: a case report
Port-a-cath-related Bacillus cereus bacteremia
Invited Lecture
Yi-Ling Tai
Pi-Hsiang Liu
Department of Internal Medicine, Tainan Municipal Hospital, Tainan, Taiwan
Department of Internal Medicine, Tainan Municipal Hospital, Tainan, Taiwan
Introduction
This study was conducted to re-investigate the antimicrobial susceptibility
patterns of community-acquired respiratory tract organisms in southern
Taiwan, guiding empiric antibiotics prescribed by clinicians.
Introduction
Once encountering blood cultures positive with Bacillus cereus, they are
generally regarded as contamination organisms. Hence, the case numbers
of Bacillus cereus bacteremia were rare in the literature. Herein, we reported a case of port-a-cath-related Bacillus cereus bacteremia.
Methods
At a regional hospital in southern Taiwan, all non-duplicate sputum and
throat swab cultures taken within 2 days of hospitalization positive with S.
pneumoniae, Haemophilus, and Moraxella catarrhalis were enrolled from
2011 to 2015. The most frequent organisms were calculated and those
results of antimicrobial susceptibility testing were analyzed.
Special Lecture
Case report
A 47-year-old female patient was admitted for chemotherapy on 8 March
2016. She had underdying diseases of colon and ovarian carcinoma. On
9 March, fever occurred and two sets of blood cultures, one from peripheral line and one from port-a-cath, were taken, and then piperacillintazobactam was given as empiric antibiotic. On 12 March, two blood
cultures were both positive with Bacillus cereus. On 15 March, port-acath was removed due to suspected port-a-cath-related bacteremia. On 17
March, piperacillin-tazobactam was shifted to vancomycin. On 18 March,
tip culture performed by semi-quantitative method of port-a-cath was
also positive with Bacillus cereus with more than 15 colony forming units
(CFU). Accordingly, port-a-cath-related Bacillus cereus bacteremia was
definitively diagnosed. Both transthoracic and transesophageal heart echo
were done on 14 March and 18 March, respectively, and revealed rule out
vegetations over aortic valve. On 28 March, vancomycin was shifted to
oral ciprofloxacin to complete treatment course.
Results
A total of 286 isolates were enrolled. The most frequent organisms were
H. influenzae (n = 106, 37.1%), H. parainfluenzae (n = 91, 31.8%) and S.
pneumoniae (n = 59, 20.6%). For H. influenzae, the susceptibility rates to
ampicillin, amoxicillin-clavulanate, cefuroxime, ceftriaxone, levofloxacin,
trimethoprim-sulfamethoxazole, and ertapenem were 34%, 72.6%, 78.3%,
99.1%, 45.3%, 25.5%, and 100 %, respectively. For H. parainfluenzae, those
were 84.6%, 94.5%, 94.5%, 100%, 41.8%, 51.6%, and 100 %, respectively.
For S. pneumoniae, the susceptibility rates to clindamycin, erythromycin,
penicillin G, amoxicillin, cefotaxime, levofloxacin, trimethoprim-sulfamethoxazole, and vancomycin were 30.5%, 6.8%, 62.7%, 72.7%, 72.7%, 69.5%,
37.3%, and 100%, respectively.
Educational Lecture
Discussion
According to the criteria to diagnose catheter-related bacteremia, the patient had two positive blood cultures with Bacillus cereus and one positive
tip culture with Bacillus cereus with more than 15 CFU performed by semiquantitative method; hence, port-a-cath-related Bacillus cereus bacteremia
could be definitively diagnosed. Herein, we suggest that two or more than
sets of blood cultures positive with Bacillus cereus should be further evaluation to prove true bacteremia. Moreover, heart echo may be performed to
rule out bacterial endocarditis.
Discussion
As a result of this study, for the three community-acquired respiratory
tract organisms, including H. influenzae, H. parainfluenzae, and S. pneumoniae, ceftriaxone had higher susceptibility rates than amoxicillin-clavulante
and levofloxacin. Which indicated that ceftriaxone was more appropriate
antibiotic to treat community-acquired respiratory tract infections than
amoxicillin-clavulanate and levofloxacin in southern Taiwan. In contrast,
levofloxain was less appropriate antibiotic due to lower susceptibility
rates.
Symposium
PA-23
The prevalence of beta-lactamase-negative ampicillin-resistant Haemophilus
influenzae/parainfluenzae in southern Taiwan
Beta-lactamase-negative ampicillin-resistant Haemophilus influenzae/parainfluenzae
PA-24
Hematological Disorders in patients on Antituberculosis Drugs in the South West Region of Cameroon
Drug induced Hematological Disorders in patients on Antituberculosis Drugs in the South West Region of
Cameroon
Benjamin T. Pokam1, Enoh J. Eeteneneng1, Aniekan O. Eyo2, Jules C. Assob1, Boris Forminyam1,
Marcelin N. Ngowe3
Pi-Hsiang Liu
Department of Internal Medicine, Tainan Municipal Hospital, Tainan, Taiwan
Case Conference
1 Department
of Medical Laboratory Science, University of Buea, Cameroon
2 Department
of Medical Laboratory Science, College of Medical Sciences, University of Calabar, Calabar, Nigeria
3 Faculty
of Health Sciences, University of Buea.
Oral Presentation
Poster Presentation
Introduction
The prevalence of beta-lactamase-negative ampicillin-resistant (BLNAR)
Haemophilus influenzae had been reported in Japan and China. To our
knowledge, they did not been reported in Taiwan. Hence, this study was
conducted to investigate the prevalence of BLNAR Haemophilus in southern Taiwan.
Methods
At a regional hospital in southern Taiwan, from July 2014 to March 2016,
all non-duplicate isolates of H. influenzae/parainfluenzae reported from
clinical laboratory were enrolled in this study. Disk diffusion method was
used for antimicrobial susceptibily testing. The antibiotics tested included
ampicillin, amoxicillin-clavulanate, cefuroxime, ceftriaxone, ertapenem,
levofloxacin, and trimethoprim- sulfamethoxazole. The results were interpreted according to the criteria recommended by Clinical and Laboratory
Standards Institutes 2014. Cefinase disk method (Becton Dickinson Microbiology System, Sparks, MD) was used to detect the production of betalactamase. If the isolates were negative for beta-lactamase but resistant to
ampicillin, they were regarded as BLNAR Haemophilus. If the isolates were
positive for beta-lactamase but resistant to amoxicillin-clavulanate, they
were regarded as beta-lactamase-positive amoxicillin-clavulanate-resistant
(BLPACR) Haemophilus.
Background: Antituberculosis drugs (ATD) have been known to efficiently
combat Mycobacterium tuberculosis either due to the active principle itself
or to its metabolites. The treatment regimen of tuberculosis (TB) patients
with first line drugs during the intensive and continuation phase have
been shown to have some adverse effects. This study was carried out in
the South West Region of Cameroon to determine the magnitude of drug
induced hematological disorder (DIHD) in TB patients under treatment.
Methods: Ninety six TB patients on ATD were enrolled and compared to 32
(control) individuals who were neither on ATD or any other treatment. In
this hospital based cross sectional study, consenting participants records
were reviewed for medical history and questionnaire issued. About 2ml of
venous blood was collected and analyzed using automatic hematological
analyzer.
Results: DIHD was observed in 62 (64.58%) of the 96 patients as compared to 5/32 (15.63%) of the control group (p= 0.00000157). In the 62
patients, a combination of drug induced hematological disorder was recorded: Thirty five (56.45%) had agranulocytosis, 24 (38.71%) leucopenia,
23 (37.1%) anemia and 17 (27.42%) thrombocytopenia. Gender (p=0.173),
age (p=0.461) and treatment duration (p=0.448) were not associated with
DIHD.
Conclusion: TB patients who receive standard ATD treatment do develop
drug induced hematological disorder, with agranulocytosis the most commonly observed. Close monitoring of such patients is advocated.
Keywords: Antituberculosis drugs, Hematological disorders, Agranulocytosis, South West Region, Cameroon
Results
A total of 111 isolates, including 54 H. influenzae and 57 H. parainfluenzae, were enrolled. Among H. influenzae isolates, 22.2% (12 of 54) and
11.1% (6 of 54) were BLNAR and BLPACR strains, respectively. Among H.
parainfluenzae, 3.5% (2 of 57) were BLNAR strains, and no BLPACR strain
was found.
Discussion
As a result of this study, the prevalence of BLNAR and BLPACR H. influenzae was 22.2 % and 11.1 %, respectively. Further, BLPACR strains might be
regarded as beta-lactamase-positive BLNAR strains because their resistant
mechanism to amoxicillin-clavulanate was both due to the mutations of
penicillin-binding protein 3. Consequently, the prevalence of BLNAR H. influenzae in southern Taiwan was 33.3% (18 of 54). In addition, the prevalence of BLNAR H. parainfluenzae in southern Taiwan was only 3.5%.
50
Flomoxef is an appropriate empiric antibiotic to treat urinary tract infections
in southern Taiwan
Flomoxef is an appropriate empiric antibiotic to treat urinary tract infections
PA-26
Doripenem has a higher susceptibility rate than imipenem against carbapenem-resistant
Enterobactericeae
Doripenem has a higher susceptibility rate against carbapenem-resistant Enterobactericeae
Chin-Lu Chang
Department of Infectious Diseases, Tainan Municipal Hospital, Tainan, Taiwan
Department of Infectious Diseases, Tainan Municipal Hospital, Tainan, Taiwan
Introduction
The common uropathogens include Escherichia coli, Klebsiella pneumoniae, and Proteus mirabilis. For these organisms, extended-spectrum betalactamase (ESBL)-producing strains are increasing worldwide. However,
flomoxef cannot be hydrolyzed by ESBL, and may remain susceptibility
against ESBL-producing strains. This study was conducted to investigate
that whether flomoxef was an appropriate empiric antibiotic to treat urinary tract infections in southern Taiwan.
Introduction
Carbapenem-resistant Enterobactericeae (CRE) are increasing worldwide,
resulting in treatment difficulty. However, CRE may be susceptible to each
individual carbapenems; hence, this study was conducted to compare the
in vitro susceptibility rates of doripenem and imipenem against CRE.
Methods
At a regional hospital in southern Taiwan, all isolates of E. coli, K. pneumoniae, and P. mirabilis reported by clinical laboratory in 2015 were enrolled. Disk diffusion method was used for antimicrobial susceptibility testing of flomoxef. The interpretive criteria of flomoxef recommended by the
manufacturer were as follows. The zone size of susceptibility was or more
than 18 mm, that of intermediate resistance was between 13 and 17 mm,
and that of resistance was or less than 12 mm. All intermediate-resistant
results were regarded as resistant results in this study.
Results
A total of 113 CRE isolates, including 56 Klebsiella pneumoniae, 21
Escherichia coli, 20 Enterobacter spp., 9 Citrobacter spp., 4 Providencia
spp., 2 Klebsiella ozaene, and 1 Morganella morganii, were enrolled. The
susceptibility rates of doripenem and imipenem against these CRE isolates
were 33.6% (38 of 113) and 13.3 % (15 of 113), respectively. The P value
was less than 0.05 calculated by chi-square test.
Discussion
As a result of this study, doripenem had a higher susceptibility rate than
imipenem against CRE isolates, indicating Enterobactericeae being less
easily resistant to doripenem than imipenem. Conseqeuntly, doripenem
may prefer to imipenem to treat infections caused by Enterobactericeae
due to less selection of CRE.
Discussion
As a result of this study, flomoxef had more than 80 % susceptibility rates
against the common uropathogens, including ESBL-producing strains. Consequently, flomoxef is an appropriate empiric antibiotic to treat urinary
tract infections in southern Taiwan.
Evaluation of a fast diagnostic chip method applied in
identification of Mycobacterium tuberculosis
The application of Biochip to identify Mycobacterium tuberculosis
PA-28
Hui-Jine Hsu1, Mei-Feng Lee2,3
rpoB Gene Analysis of Multidrug-resistant
Mycobacterium Tuberculosis Strains Isolated in
North Taiwan
Symposium
PA-27
Educational Lecture
Results
A total of 5584 isolates, including 3969 E. coli, 1199 K. pneumoniae, and
416 P. mirabilis were enrolled. Among these isolates, the rates of ESBLproducing strains of E. coli, K. pneumoniae, and P. mirabilis were 21.2%
(841 of 3969), 18% (216 of 1199), and 12.7 % (53 of 416), respectively.
The susceptibility rates of flomoxef against E. coli, K. pneumoniae, and P.
mirabilis were 83.7% (3322 of 3969), 81.4 % (976 of 1199), and 94.7%
(394 of 416), respectively.
Special Lecture
Methods
At a regional hospital in southern Taiwan, from June 2011 to March
2016, all isolates of CRE were enrolled. All isolates of Enterobactericeae
with nonsusceptibility to one or more of imipenem, meropenem, and
doripenem were defined as CRE. However, for isolates of Proteus spp.,
Providencia spp., and Morganella spp., if they were nonsusceptible to impenem, they should have to be nonsusceptible to any one or more of other
carbapenems because they had intrinsic resistance to imipenem. Disk
diffusion method was used for antimicrobial susceptibility testing. The interpretive creteria were according to the recommendation of Clinical and
Laboratory Standards Institutes. All intermediate-resistant results were
regarded as resistant results in this study.
Invited Lecture
Chin-Lu Chang
Keynote Speech
PA-25
Hsiu-Ru Huang1, Yi-Ling Liang1, Fang-Lan Yu1, Guei-Sin Huang1, Jau-Ching Lee1, Yi-Yuan Yang2,
Giueng-Chueng Wang1
1 Department
of Laboratory Medicine, Wan Fang Hospital, Taipei Medical University, Taipei, Taiwan
2 School
of Medical Laboratory Sciences and Biotechnology, Taipei Medical University, Taipei, Taiwan
Methods
This was a retrospective study at Chi Mei medical center in southern
Taiwan. From January to December 2013, all clinical samples sent for
identifying TB were enrolled in this study. Three methods were used for
identifying TB, including Biochip, TCM, and AFS.
Conclusion
As a result of this study, the detectable rate of TB was similar between
Biochip, a rapid method (less than 24 hours), and TCM, a time-consuming
method. In addition, Biochip had a higher detectable rate and accurate
rate than AFS (5.8% vs. 3.9% and 100 % vs. 58.5%, respectively). Hence, we
consider that Biochip should have an important role to identify TB, resulting in rapid diagnosis of tuberculosis and decreasing nosocomial spread.
51
Poster Presentation
Results
A total of 6819 samples, mostly obtained from sputum, were enrolled in
this study. The detectable rate of TB was 5.8 % (n = 396), 5.8 % (n = 394),
and 3.9% (n = 269) by using Biochip, TCM, and AFS, respectively. In addition, 510 (7.5%) and 473 (6.9%) NTM were detected by Biochip and TCM,
respectively. Of the all samples, 460 had positive result by AFS, indicating
that the accurate rate of identifying TB by AFS was 58.5% (269 of 460).
Oral Presentation
Rifampin is an antibiotic drug for treating tuberculosis treatment. Rifampin resistance is most frequently conferred by mutations in the 81 bp
of active center of RNA polymerase encoded by rpoB gene. In this study,
we intended to find the mutation patterns of rpoB gene in MDR-TB, and
the correlation of rpoB gene mutation patterns and the drug resistance.
Fragments in hot spot region spanning 1546-1593 nucleotides of RpoB
gene of 100 multiple drug-resistant Mycobacterium tuberculosis (MDRTB) strains isolated from Wan Fang Hospital in 2010 were amplified and
sequenced. MDR-TB strains were defined as mycobacteria are resistant to
0.2 ug/ml of INH and 1.0 ug/ml of rifampin. We also did second-line drug
susceptibility testing with the agar proportion method on 7H11 agar. The
mutations mainly occurred in codon 531 (59%), codon 516 (11%), and
codon 526 (9%). Only 1% mutation rate was in codons 511, 513, 522, and
533. MDR-TB strains were almost fully resistant to rifabutin when mutations occurred in rpoB hot spot region. It had chances for second-line drug
resistance when mutation occurred in codon 531. Moreover, mutations
occurred in codons 526 or 516 conferred the bacteria drug susceptibility
to amikacin, kanamycin, and capreomycin. Identifying gene mutation patterns is a critical point for rapid molecular diagnostic tests. Mutations in
rpoB 81 bp hot-spot region causing rifampin resistance are already known
for MTB therapy. Mutations in rpoB codons 531, 516, and 526 might be
useful diagnostic markers for rifampin resistance. The results would be
applicable when more samples are analyzed and compared with local epidemiology such as spoligotyping in future studies.
Background
Mycobacterium tuberculosis (TB) can result in nosocomial outbreak, especially delayed diagnosis. Traditionally, acid-fast stain (AFS) and traditional
culture method (TCM) are used to identify TB. However, AFS has a lower
detectable rate of TB and cannot differentiate between TB and non-tuberculosis Mycobacterium (NTM). Although TCM is regarded as the golden
standard of identifying TB, it is a time-consuming method. The aim of this
study was to evaluate the role of identifying TB by Biochip (CaptialBio, Beijing, China), a quickly diagnostic molecular method.
Case Conference
1 Division
of Microbiology, Department of Clinical Pathology Chi Mei Medical Center, Chi Mei Medical Center, Taiwan Society of
Laboratory Medicine , Taiwan
2 Modern
Clinical Laboratory, Tainan City, Taiwan
3 Department
of Nursing, Yuh-Ing Junior College of Health Care & Management, Kaohsiung City, Taiwan
Keynote Speech
PA-29
Analysis the Antimicrobial Resistance Rates and Contamination Rates of Bloodstream Infections (BSI).
BSI can cause important morbidity and mortality worldwide. Especially in delayed diagnostic and
treatment among bacteremia or sepsis patients.
PA-30
MOLECULAR CHARACTERIZATION OF VIBRIO
PARAHAEMOLYTICUS ISOLATED FROM SHELLFISH
FROM SUEZ CANAL AREA, Egypt
CHUNG-PIN KAO1, YUNG-CHEN CHIEN2
Ahmed I. Youssef1, Amani L. Farag2, Ehab M. Helal3
1 Division
of Laboratory, Yangming Branch of Taipei City Hospital, Taiwan
2 Division
of Bacteriology Laboratory at Renai Branch of Taipei City Hospital
1 Animal
hygiene and zoonoses, Division of Zoonoses, Faculty of Veterinary Medicine, Suez Canal University, Ismailia, Egypt
2 Food
Hygiene Department, Animal Health Research Institute, Dokki-Giza, Egypt
3 Bacteriology
Department, Animal Health Research Institute, Dokki-Giza, Egypt
Invited Lecture
Total blood culture bottles were detected between January 1st to December 31th in 2015 of Taipei City Hospital of Yangming Branch.
All bottles were placed into Automated BD BACTEC FX Blood Culture System. Then, the positive bottles were identified the isolates and its antibiotic
susceptibility by using BD PhoenixTM 100.
Special Lecture
Educational Lecture
Vibrio parahaemolyticus is considered a natural pathogen of the aquatic
environment. It is a leading cause of seafood-derived food poisoning
throughout the world. The main objectives of this study were to determine
the prevalence of contamination of shellfish by V. parahemolyticus in Suez
Canal area and to assess its molecular characteristics. The study included
205 samples of shellfish (82 clams, 43 mussles, and 80 shrimps) collected
from Suez Canal area (78 from Port Said, 82 from Ismailia and 45 from
Suez governorates). In addition, the harvesting water samples were collected from different sites. All samples were collected during the warmest
season (June, July and August) through two years research period. After
extraction of tissue and homogenization, enriched samples were identified by plating onto TCBS agar medium. Presumptive V. parahaemolyticus
blue to green colonies were selected and purified and further identified by
API20 and PCR techniques targeting toxR gene to be confirmed as V. parahaemolyticus. The pathogenicity of the isolates was examined by detection
of Thermostable Direct Haemolysin (TDH) and related Haemolysin (TRH)
genes. Results revealed that the overall prevalence of V. parahaemolyticus
in shellfish was 19/205 (9.27%) whereas in water was 6/24 (25%). Higher
contamination rate was detected in shrimp (15%), and the highest rate
was in Ismailia governorate (12.2%). The detection rate of TDH and TRH
genes was 15.78% and 8.33%, consequently. This study concluded that the
examined shellfish may have the potential human health risk associated
with the presence of pathogenic V. parahaemolyticus and preventative
measures should be considered.
First, 513 positive blood culture bottles were detected in 4915 samples
(513/4915, 10.5%). We yielded 550 clinically significant organisms which
including 28 patients who were infected with two or over two isolates
(28/513, 5.5%). The overall mortality was 19.3%. Second, 108 organisms
were resistant to antimicrobial agents (108/550, 19.6%) among ER (n=38,
6.9%), OPD (n=2, 0.4%), ICU (n=18, 3.3%) and ward (n=50, 9.1%). The
identified resistant-strains were MRSA (n=33, 6.0%), MRCoNS (n=20, 3.6%),
MRSE (n=4, 0.7%), ESBL-EC (n=37, 6.7%), ESBL-KP (n=10, 1.8%), ESBL- P.
mirabilis (n=3, 0.6%). Vancomycin resistance was seen in 0.2% of Enterococcus faecalis isolates. Finally, the patients with BSI from blood contamination was up to 20.4% (n=112) among ER (n=70, 12.7%), ICU (n=11, 2.0%),
ward (n=31, 5.6%). And there was no detection of contaminated blood bottle among OPD.
Comparing with the positive rate in blood culture between 2013 (14.3%)
to 2014 (12.5%). The BSI is decreased recently years. According to the
data, urged that the patients with BSI are very likely to be infected with
highly resistant isolates. Especially with MRSA or ESBL-EC. And we found
that ER had higher results of resistant-strains and contamination rates
rather than ICU and ward in blood culture. However, how to prevent the
blood contamination and provide the correct diagnosis can monitor with
effective treatments to reduce the patient morbidity and mortality.
Symposium
PA-31
High prevalense of Mycoplasma genitalium macrolide resistance in the Oslo area.
Detection of macrolide resistant Mycoplasma genitalium in Oslo, Norway.
PA-32
Salmonella identification by MALDI-TOF MS
Evaluation of MALDI-TOF MS as a replacement for
biochemical identification of Salmonella.
Case Conference
Marie E. Vad1, Kirsti Jakobsen1, Anne Marte Kran1,3, Anne Olaug Olsen2,3, Mona Holberg-Petersen1
Ina P. Haagensen, Irene Rauk, Trine M. Groenbeck, Nils O. Hermansen
1 Department
of microbiology, Oslo University Hospital, Norway
2 Department
of Rheumatology, Dermatology and Infectious Diseases, Oslo University Hospital
3 Institute
of Clinical Medicine, University of Oslo
Department og Foodborne Infections, Norwegian Institute og Public Health, Norway
Oral Presentation
Poster Presentation
Introduction:
Biochemical and serological methods are traditionally used to identify
Salmonella from clinical samples. The National Reference Laboratory for
Enteropathogenic Bacteria, Norwegian Institute of Public Health wanted
to examine if MALDI-TOF MS (Bruker Daltonik GmbH) could replace
biochemical tests in the identification of Salmonella to genus level. At the
same time, we wanted to examine whether various growth media would
influence the score obtained using this method.
Material and Methods:
A total of 101 Salmonella isolates from our laboratory collection were analysed: S. enterica (including all six subspecies: S. enterica (50), S. salamae
(11), S. arizonae (6), S. diarizonae (13), S. houtenae (10), S. indica (1)) and
species S. bongori (10). Morphological and phenotypical deviant strains
from S. enterica ssp. enterica were also included. Each isolate had previously been identified using standard biochemical and serological methods.
Bacteria were grown on lactose-, nutrient- and Columbia blood-agar and
subsequently analysed using Microflex LT MALDI-TOF MS with MALDI
biotyper 3.1 software. Each isolate was prepared in duplicate using the
smear method described by the manufacturer.
Results:
The results showed acceptable scores ( ≥ 2.0) for all Salmonella strains to
genus level except for S. enterica ssp. houtenae . This subspecies was incorrectly identified in some duplicate tests and generally showed lower scores
( ＜ 2.0). The score was ( ≥ 2.0) for 295 out of 303 analyses independent
of growth media.
Conclusion:
MALDI-TOF MS can replace biochemical methods for detection of Salmonella to genus level. The method did, however, have difficulties in identifying Salmonella houtenae , but infections caused by this microbe are very
rare in humans. Serological tests are still necessary to identify serotypes.
Additional biochemical testing must be performed if different subspecies
have equal antigenic formulae. The three growth media utilised are all
equally suitable for use in the identification of Salmonella with MALDI-TOF
MS.
Introduction
Mycoplasma genitalium is a sexually transmitted pathogen bacteria commonly treated with macrolides, ie azithromycin.
However, resistance to macrolides has been more frequently reported. The
aim of this retrospective study was to describe the prevalence of macrolide
resistance in M. genitalium from patient in Oslo, Norway.
Methods
M. genitalium PCR positive clinical samples obtained during a two months
period from patients visiting Olafiaklinikken, an open acess STI clinic in
Oslo, and general practitioners in the Oslo area, were included. Macrolide
resistance associated with point mutation in the 23S rRNA gene at positions 2058 and 2059 were analyzed by sequencing of PCR- products.
Results
A total of 3525 samples were analyzed by our in house M. genitalium realtime PCR, targeting MgPa adhesine gene, of which 155 (4.4 %) were positive. Ninety eight of the 155 M. genitalium positive samples were analyzed
for macrolide resistance-associated mutations, and mutations in position
2058 and 2059 were found in 63 samples (64 %). The predominant mutations identified were A2059G (36.7 %) and A2058G (23.5 %).
Conclusion
We have detected macrolide resistance-associated mutations in M. genitalium at a rate of 64 %.
This study shows that macrolide resistant M. genitalium isolates occur
with a high frequency in the Oslo area and confirms the needs for prospective detection of macrolide resistance prior to treating patients. Otherwise,
the importance of a test of cure to detect treatment failure to standard
treatment must be emphasized.
52
Seted up a new method for detection of Clostridium difficile Ribotypes in Stool
Samples
Extracted the DNA from stool specimen and direct PCR to analysis the Ribotyping
PA-34
Fu chieh Chang1, Tuan Jen Wang2
Molecular characteristics of Clostridium difficile isolated in
Kobe, Japan
Tomoko Hase1, Kayo Osawa1, Katsumi Shigemura2, Kenichiro Onuma3, Mari Kusuki3,
Tatsuya Nakamura3, Toshiro Shirakawa2,4, Masato Fujisawa2
1 Infection
control center, MacKay memorial hospital, Taiwan
2 Department
of Laboratory Medicine,MacKay memorial hospital
1 Department
of Biophysics, Kobe University Graduate School of Health Sciences, Japan
2 Department
of Urology, Kobe University Graduate School of Medicine
3 Department
of Clinical Laboratory, Kobe University Hospital
4 Department
of Advanced Medical Science, Kobe University Graduate School of Science, Technology and Innovation
Introduction: Clostridium difficile is a species of Gram-positive spore-forming bacteria.When stressed, the bacteria produce spores that are able to
tolerate extreme conditions that the active bacteria cannot tolerate. In this
study, we wanted to set up a new method that can direct PCR and ribotyping from stool sample.
Material and method : In this study , we setted up a new method that can
direct extracted DNA from stool sample. And the PCR ribotyping was modified to allow direct detection of Clostridium difficile from stool samples.
Result:Direct PCR ribotyping was possible in 60C. difficile-positive stool
samples(culture positive) , and in 54 cases (90%), the ribotype determined
directly from the stool sample was identical to the ribotype of the strain
isolated from the same stool sample. Finally, according to the sequencing
data showed that all of the C.difficile belong to 017.
Discussion: Direct PCR ribotyping gives the information on the presence and type of C. difficile within hours, in contrast to standard culturedependent methods where typing results can be obtained only after 3
days or more (48 h for culture and 1 day for PCR ribotyping). Direct PCR
ribotyping on DNA isolated from stool samples is convenient, rapid, and
useful for the detection of specific types of C. difficile in fecal samples.
Clostridium difficile infection is mainly a leading cause of diarrhea by toxins
producing C. difficile . C. difficile produces two main toxins, which are an
enterotoxin A (tcdA ) and a cytotoxin B (tcdB ). A third toxin, C. difficile toxin
(CDT) called binary toxin is composed of enzymatic (cdtA ) and cell binding/
translocation (cdtB ). CDT-producing C. difficile is frequently associated with an
increased risk for severe C. difficile infection. In those CDT-producing C. difficile
strains, the hyper-virulent strain, having deletions in tcdC as a putative negative
regulator of tcdA and tcdB , has caused outbreaks worldwide. Our purpose of
Invited Lecture
Materials and Methods
A total of 142 isolates of C. difficile were obtained from Kobe University Hospital between April 2012 and December 2014.We detected the presence of
tcdA , tcdB , tcdC , cdtA and cdtB by PCR and sequencing. The clonal relationship
of CDT-producing strains was explored by repetitive-sequence-based PCR (repPCR).
Special Lecture
Introduction
this study is to investigate the molecular characteristics of toxigenic (especially
CDT-producing) C. difficile strains isolated in Kobe, Japan.
Discussion
We found 4 CDT-producing strains in C. difficile isolates and one of them had
deletions in tcdC . The rep-PCR results revealed no clonal proximity in CDT-producing strains. Our findings suggest the trend of these CDT-producing strains
spreading in Kobe, Japan.
PA-36
Staphylococcus aureus
Hiromasa Fukatsu1, Takehisa Matsumoto2, Eriko Kasuga3, Tatsuya Natori3, Kenta Takehara3,
Kanae Mimura3, Mitsutoshi Sugano3, Takayuki Honda3
In-house Protocol for Clinical Sputum Specimen Inoculation
by Using Previ Isola Automated System
Symposium
Characterization and genetic analysis of a menadionedependent small-colony variant of methicillin-resistant
Educational Lecture
Results
Among the isolates, 99 (69.7%) were toxigenic. In the toxigenic strains, 4 (4.0%)
were CDT-producing strains (tcdA +/ tcdB +/ cdtA+ / cdtB +), and 95 (96.0%) were
non CDT-producing strains (81 strains: tcdA +/ tcdB +/ cdtA -/ cdtB -; 14 strains:
tcdA -/ tcdB +/ cdtA -/ cdtB -). One CDT-producing strain had a single base-pair
deletion at nucleotide position 117 and 18-bp deletion at positions 330-347 in
tcdC . The CDT-producing strains showed ＜ 90% similarity by rep-PCR.
PA-35
Keynote Speech
PA-33
Fang-Lan Yu, Guei-Sin Huang, Ting-Ya Yang, Chia-Ling Cheng, Jau-Ching Lee
Lab Medicine, Taipei Municipal Wanfang Hospital, Taiwan,
Introduction
Small-colony variants (SCVs) are mutant strains, which show atypical
colony morphology, and require specific nutrient factors for growth. These
strains sometimes cause persistent or recurrent infections. We report the
characteristics and genetic analysis of a menadione-dependent small-colony variant of methicillin-resistant Staphylococcus aureus (MRSA) isolated
from a clinical sample.
185 clinical sputum specimens were collected for the study (2015/08/052015/09/05). Specimens were first manually inoculated on to culture
media; the remained specimens were liquefied by equal volume of 5% Nacetyl-L-cysteine(NALC) in pH 7.4 PBS buffer. After vortex and 30 minutes
standing at room temperature, 10ul liquefied sample were applied for
Previ Isola® plate streaking. The variety of colonies, the quality of colony
growth, and the possibility to isolate single colony from the two inoculation methodologies were investigated and compared after 24 hours incubation.
The positive and negative specimen culture concordance for the two inoculation methodologies was 98.9 % (183 /185). 2 inconsistent results were
from trace sample: one positive in automated method and one in manual
method). In average, lab technician needs to re-isolate 5% of manual inoculated sputum specimens due to single colonies were hardly found after
incubation. In the study, the culture plate prepared by automated streaking system revealed single colonies in more specimens than manual procedure. The new protocol could help to speed up the time to result for those
samples for approximately 24 hours. This PREVI Isola® adapted streaking
procedure with sputum liquefying protocol is a good standardized tool
and time-saving method for clinical sputum specimen processing.
Results
The SCV was identified as S. aureus by the API Staph test, and it was resistant to cefoxitin. PCR demonstrated that the isolate possessed mecA , and
the auxotrophic test showed its dependence on menadione for growth.
This isolate, however, had no genetic mutation in eight genes including
menA , menB , menC , menD , menE , menF , menH , and ubiE .
Conclusion
The isolate in this study was a menadione-dependent SCV of MRSA. However, it had no mutation in the eight genes associated with menaquinone
biosynthesis. A mutation in the promoter region may cause loss of expression of these genes. Further genetic analysis is necessary to reveal the
cause of its dependence on menadione.
53
Poster Presentation
Methods
An SCV, isolated from a patient at the Shinshu University Hospital, was
used in this study. Identification and antimicrobial susceptibility tests were
performed by API Staph (Sysmex-biomerieux) and Microscan Pos MIC 3.3J
panel (Beckman coulter) tests, respectively. PCR for mecA was performed,
and an auxotrophic test was executed on BTB agar using three standard
disks impregnated with 50 µg of hemin, 10 µg of thymidine, and 1 µg of
menadione. DNA sequences of menA , menB , menC , menD , menE , menF ,
menH , and ubiE , all related to menaquinone biosynthesis, were examined
by the Applied Biosystems 3500 Genetic Analyzer (Applied Biosystems).
Oral Presentation
Lab Protocol Standardization is the approach to manage laboratory services to be more efficient and cost effective with reliable performance. With
the fashion invention, clinical labs now can apply automated specimen inoculating system to standardize lab routine inoculation protocol for liquidbase specimen. Sputum specimen, however, due to its nature is challenged
for establishing auto-inoculating protocol. In our hospital, sputum takes
50% of clinical microbiology lab sample size, and the sample size is increasing yearly. In this study, we evaluate Previ Isola® (bioM&eacute;rieux,
Craponne, France) with a special protocol for sputum specimen inoculation.
Case Conference
1 Department
of Health Sciences, Graduate School of Medicine, Shinshu University, Japan
2 School
of Health Sciences, Faculty of Medicine, Gunma University
3 Department
of Laboratory Medicine, Shinshu University Hospital
Keynote Speech
PA-37
The Role of Microbiology Laboratory in the Response to
Formosa Fun Coast Dust Explosion
powder explosion, burn-injury, microbiology laboratory
PA-38
Tzu-Ying Lee1, Yin-Tai Tsai1, Hsiao-Wei Wang2,3, Chi-Hung Lee2,3, Yung-Ching Liu2,3,4, Wei-Ming Chi1,6,
Hsueh-Hsia Wu6, Wen-Shyang Hsieh1,5,6
Tsulan Wu1, An-Jing Kuo1,2, Lin-Hui Su1,2, Jwu-Ching Shu2, Ju-Hsin Chia1
1 Laboratory
Medicine, Linko Chang Gung Memorial Hospital, Taiwan
2 Department
of Medical Biotechnology and Laboratory Science, College of Medicine, Chang Gung University
1 Department
of laboratory medicine, Taipei Medical University, Shuang Ho Hospital, Taiwan
2 Division
of Infectious Disease, Taipei Medical University; Shuang Ho Hospital, New Taipei city, Taiwan
3 Department
of Internal Medicine, Taipei Medical University-Shuang Ho Hospital, New Taipei City , Taiwan
4 Department
of Medicine, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan
5 Department
of education Taipei Medical University-Shuang Ho Hospital, New Taipei City, Taiwan
6 School
of Medical Laboratory Science and Biotechnology, Taipei Medical University, Taipei, Taiwan
Invited Lecture
Special Lecture
Objectives: Vancomycin resistance increased significantly to 35.4% among
Enterococcus faecium in 2006 and remained high thereafter at a university hospital in Taiwan. A longitudinal study was conducted to characterize
these vancomycin-resistant E. faecium
Methods: A total of 270 nonrepetitive vancomycin resistant E. faecium
blood isolates collected from 2002 to 2013 were studied. Multilocus sequence typing, PFGE , analysis of van genes and the Tn1546 structures
were performed.
Results: The majority (80%) of the isolates were associated with hospitalacquired infections. Molecular typing revealed 11 major pulsotypes and
five predominant sequence types (STs): ST17 (31.9%), ST414 (19.6%),
ST18 (14.1%), ST78 (11.1%), and ST203 (10.4%). Fluctuation of various
STs among the study years in association with some major pulsotypes was
noted. All isolates carried vanA genes, except that in four isolates vanB
genes were found. Among the vanA vancomycin resistant E. faecium carrying Tn1546 like elements, one predominant structure type (Type I, 68.4%)
was noted throughout the study years. Since 2009, another predominant
structure type (Type II, 29.3%) has emerged firstly in ST414 and gradually
spread to other STs in subsequent years.
Conclusion: Dissemination of some major STs and horizontal transfer
of two major types of vanA Vancomycin resistant E. faecium carrying
Tn1546 like elements may have together contributed to the increase of
VRE-fm infection.
Educational Lecture
On June 27, 2015, a tragic starch-based powder explosion and fire accident occurred at Formosa Water Park in the northern Taiwan, which
injuring 499 people with various degrees of thermal burn. Among the 13
patients seeking medical treatment in our hospital, 9 received complete
hospital courses. The experience of taking care of multiple massive deep
burn-injury patients generates some conclusions. First, cooling burn-injury
by dirty swimming pool water may facilitate environmental bacteria causing infections. Second, bacteria translocation from intestine and defected
skin may eventually cause bacteremia and septic shock. Third, medical
staff may cause bacteria inter-patient outbreak during daily care and
procedures. Fourth, construction works should keep away from these immunocompromised patients to avoid fungal, such as Aspergillosis, infections. Strict infection-control procedures should be performed to protect
these massive burn-injury patients. Furthermore, microbiology laboratory
professionals may take efforts to collect samples from accidental scene as
first-line database for burn-injury wound contaminations. Rapid test for
bacteria species, such as MALDI-TOF, should be used for early detection
and early intervention. Considering the feature of burn-injury wounds,
every wound culture should be isolated and cultures by detail, not only
the predominate pathogens. The species analyses and susceptibility test
should be performed on every yeast samples. By the cooperation between
laboratory experts and clinical doctors, better outcome of the massive
burn-injury victims can be expected.
Symposium
PA-39
Molecular epidemiology of vancomycin resistant Enterococcus
blood isolates in a Taiwan hospital over 12 years
changing epidemiology of VRE
PA-40
Prevalence of bacterial infections effects on semen of infertile men, at Mulago Hospital 2012-2005.
Evaluation of the Chromogenic Agar chromID C.
difficile
Rapid detection compared to traditional media
Case Conference
Mathias S Habalurema1,2, Gabriel S BIMENYA3
Fu chieh Chang1, Tuan Jen Wang2, Ming hsuan Li2
1 Clinical Laboratories, UMLTA, EAMLSA, Uganda
2 Department of Obstetrics and Gynaecology. Mulago Hospital
3 Department of Pathology, College of Health Sciences. Makerere University Kampala
1 Infection
control center, MacKay memorial hospital, Taiwan
2 Department
of Laboratory Medicine,MacKay memorial hospital
Oral Presentation
Poster Presentation
Introduction: C. difficile is gram-positive rod, spore forming, strict anaerobic bacillus and is part of the normal intestinal microbial in 1~3% of
healthy adults and 15~20% of infants. The mentioned statistics would be
increased considerably during long hospitalization and after surgery. The
important disorder caused by this bacterium is often termed C. difficileassociated diarrhea or C. difficile infection (CDI). CDI is one of the most
prevalent problems in hospitals and nursing homes where patients frequently receive antibiotics disease. With such a goal and such implications
however, the accuracy of the laboratory diagnosis is of crucial importance.
All strategies should aim at a same-day diagnosis in case of suspicion of
CDI. In case of a positive result, the immediate treatment of the patient will
improve his condition and limit the risk of room contamination. And the
rapid implementation of hygiene measures will prevent further spread of
the disease. With such a goal and such implications however, the accuracy
of the laboratory diagnosis is of crucial importance.
Material and method: All of the Stool were from inpatients ( ＞ 2y) suffering from antimicrobial- or chemotherapy-associated diarrhea. Between Jul
2015 and Dec 2015 retrospectively 24 positive stools.Two commercial
media, the CCFA agar and CHROMagar for C.difficile were compared in
a retrospective and prospective study.Colonies of C. difficile are black on
chromID, whereas they are fluorescent under UV light (360nm) on CHROMagar.
Result: The result showed that the positive ratio of CCFA and CHROMagar
were : 83.3%(20/24) and 91.6%(22/24).
Concussion: The new fluorescent CHROMagar for C.difficile is an excellent
medium for the detection of C.difficile in stool samples after 24h. Larger
colonies make identification easier.
Abstract: A retrospective analytical study to determine the prevalence of
bacterial infections, their effects on the semen quality, and the antibiogram of the bacterial isolates, in semen of infertile men referred to Mulago
Hospital Infertility clinic, Makerere University, Kampala-Uganda was done
from March 2012 to Aug 2005.
Methods: Clinical records of semen of infertile men attending the infertility clinic at Mulago Hospital were retrieved and analyzed. Only results of
samples analyzed according to World Health Organization guidelines of
(1999) were included in the study. The variables of interest were spermatozoa; concentration, motility, morphology and vitality ,bacterial isolates
and their antibiotic sensitivities; and effect of bacterial isolates on semen
quality and quantity. Sixty-six percent (n=57) of the semen specimen cultured had bacterial growth. Results:The most prevalent bacteria isolates
were Coagulase Negative Staphylococcus (40.35%), Staphylococcus aureus
(31.57%), and Enterococcus feacalis (19.29%. Others were Streptococcus
agalaceae (1.75%), Pseudomonas earuginosa (1.75%), Esch.coli (1.75%),
Ureaplasma urealyticum (1.75%) and Acitenobacter baumani (1.75%).
The semen abnomalities were Azoospermia (12.79%), Normozoospermia
(11.62%), Asthenoteratozoospermia (24.4%) and Oligoasthenoteratozoospermia (51.16%). Staph aureus was exhibiting 33.5% asthenoteratozoospermia, 33% oligoasthenoteratozospemia and 22 % Normozoospermia,
whereas CNStaphylococcus and Enterococcus feacalis exhibited 82% and
73% oligoasthenoteratozoospermia respectively. The rest of bacterial
infection exhibited astheno-teratozoospermia. The bacteria were mostly
sensitive gentamycin (90%) and augumentin (75%), as they were resistant
to Amoxyllin (100%) and cotrimaxazole at (100%).
In conclusion, it was found that bacterial infection in semen was quite
common in Uganda, leading to deterioration of semen quality resulting
in likely male infertility. Therefore bacteria culture and sensitivity should
routinely be done in semen analysis.
Key words: spermatozoa, semen quality, bacterial infection.
54
Application of MALDI-TOF MS and MALDI Biotyper for Analyzing the Susceptibility ofEnterococcus to Vancomycin
Analysis of vancomycin susceptibility ofEnterococcus with the dendrograms generated from main spectrum profiles of
MALDI-TOF MS with MALDI-Biotyper
PA-42
Effect of cultured metabolites on the pel genes expression ofPseudomonas aeruginosa
Application of cultured metabolites on genes expression of exopolysaccharides for interfering the
biofilm formation of Pseudomonas aeruginosa
The infection of Vancomycin-resistant Enterococci (VRE) is higher than
each year in the world. Matrix-assisted laser desorption ionization-time
of flight mass spectrometry (MALDI-TOF MS) has been applied as a rapid,
accurate, and inexpensive method for bacterial identification in clinical
laboratories. Although MALDI-TOF MS is a powerful tool for identification
of bacteria, the application of MALDI-TOF MS on antimicrobial susceptibility still needs to be further investigated. This study used MALDI Biotyper
to construct the dendrograms generated from the main spectrum profiles
of enterococcus either treated with vancomycin or without vancomycin.
After the acquisition of MALDI-TOF MS, the main spectrum profiles were
subjected to construct the dendrograms for analyzing the situations of
enterococci treated with vancomycin (6 ug/ml). The relative distance
levels of dendrograms were compared for analyzing the optimal bacterial
density and incubation time. The results indicated that distance level of
dendrograms were higher in the bacterial density of 6x108 CFU/ml and
the incubation time of 2-3 hours than those in the other bacterial density
and the other incubation time. Besides, the clinical isolates of enterococci
including E. faecalis , E. faecium , E.casseliflavus , and E. gallinarum were
applied in this study for the discrimination between VRE and Vancomycinsusceptible Enterococci (VSE). The results indicated that the method was
able to discriminate the VRE and VSE in different group. Eventually, the
combination of MALDI-TOF MS and MALDI Biotyper may be applied as a
potential tool for rapid detection of Vancomycin-resistant Enterococci .
The biofilm formation is a considerable problem in the treatment of P.
aeruginosa infection. Both adhesive factors and disruptive factors are criti-
cal during the process of biofilm formation. The quorum-sensing signal
is demonstrated to be associated with biofilm formation. Therefore the
secreted metabolites containing quorum-sensing signal were applied to
study the effect of cultured metabolites on the process of biofilm formation. Since three exopolysaccharides existed in P. aeruginosa biofilm
contain alginate, Psl, and Pel, the biosynthesis of exopolysaccharides were
explored with Congo red staining. The result of Congo red staining was
shown that the amounts of exopolysaccharides were increased as cultured
time. This study analyzed the amount of DNA, protein, and pyocyanin in
1-5 days of cultured metabolites and the results was obtained. First, the
cultured metabolites contained extracellular DNA. Second, the cultured
metabolites contained protein and the amounts of protein were increased
as cultured time (1-5 day). Third, the cultured metabolites possessed pyocyanin, and the amounts of pyocyanin were increased as cultured time
(1-5 day). Moreover, the effects of the cultured metabolites on the genes
expression of bacteria were analyzed. The results indicated that the mRNA
expressions of pel genes were shown to be decreased in medium by adding the cultured metabolites. In conclusion, this study revealed that the
cultured metabolites in medium may cause the changes in gene expression
and then interfere the biofilm formation of Pseudomonas aeruginosa .
Special Lecture
Ting-Yi Wang1, Tzu-Hui Wang2, Shih-Chieh Lo1, Shiao-ping Huang1
1 Department
of Medical Laboratory Science and Biotechnology, Fooyin University, Kaohsiung, Taiwan
2 Department
of Clinical Microbiology, Kaohsiung Veterans General Hospital, Kaohsiung, Taiwan
Invited Lecture
Li-Feng Bu1, Huey-Ling You2, Chun-Chih Chien2, Shu-Shen Cheng2, Shiao-ping Huang1
1 Department
of Medical Laboratory Science and Biotechnology, Fooyin University, Kaohsiung, Taiwan
2 Department
of Laboratory Medicine, Kaohsiung Chang Gung Memorial Hospital
Keynote Speech
PA-41
Educational Lecture
Alanine uptake effects on methicillin-resistant Staphylococcus aureus cell surface
charges and sensitivity to nisin
nisin, alanine, methicillin-resistant S. aureus, vancomycin, cytochrome c unbound test
PA-44
Evaluation of Drug Susceptibility Testing with the Broth Microdilution Method in Multidrug-resistant Mycobacterium
tuberculosis
The accuracy and turn-around-time of Drug Susceptibility Testing in Multidrug-resistant Mycobacterium tuberculosis
Multidrug-resistant (MDR) Mycobacterium tuberculosis (TB) has increased
in recent years. Drug susceptibility testing (DST) must be rapidly concluded for effective treatment the disease. Agar proportion method (APM)
is the gold standard assay of DST in TB, but it lacks of precision in some
drugs and MICs data to support chemotherapy. MYCOTBI(TREK, England)
is a new commercial MICs susceptibility plates for TB using broth microdilution method (BMM). The aim of this study was to compare BMM and
APM for DST in clinical MDRTB isolates, and combined with detection of
drug resistant genes, if necessary. A total of 60 clinical MDRTB isolates
were included and using H37Rv strain as control. Two of the sixty isolates
were excluded because of no enough control colonies grown. MYCOTBI
was performed in 96-well microtitre plates containing 12 drugs at appropriate dilutions. APM was performed as described in CLSI M24-A2. Results
could be read about 18 days after inoculation in BMM and 21 days of
APM. The consistent rate of BMM and APM in the first line drugs, Soniazid,
Rifampicin, Ethambutol, Streptomycin and the second line drugs, Ethionamide, Kanamycin, Ofloxacin, Para-aminosalicylic acid were 96.6%(56/58),
100%(58/58), 86.2%(50/58), 91.4%(53/58), 94.8%(55/58),
96.6%(56/58), 98.3%(57/58) and 96.6%(56/58), respectively. The drug resistant genes were examined in some inconsistent isolates and reanalyzed
the consistent rates. The corrected consistent rates of Soniazid, Rifampicin,
Ethambutol, Kanamycin and Ofloxacin were 98.3%(57/58), 100%(58/58),
92.5%(55/58), 100%(58/58) and 100%(58/58), respectively. MYCOTBI
had the advantages to shorten turn-around-time of DST and gave MICs
information. The concordance of BMM and APM was similar with previous
literatures, even most of the study materials were drug-resistant isolates
in this study but susceptible in previous literatures. The performance of
BMM was superior when adding the results in detection of drug resistant
genes. So BMM had the potential to replace APM in clinical laboratories.
55
Poster Presentation
Background: The drug sensitivity of methicillin-resistant S.aureus (MRSA)
to current antibiotics was found decreasing in these years.Looking for
an alternative antibacterial agent has become urgent. Objective: The goal
of this study is to analyze the antibacterial effects of nisin against various S.aureus strains,and,at the same time,to evaluate the effect of alanine
uptake on the change in cell surface charges,following which their drug
sensitivity may increase. Material and Method:In this study thirty one
MRSA strains and five methicillin-sensitive S.aureus (MSSA) strains,isolated
clinically,were utilized for assaying their sensitivities to nisin after uptake
of L-alanine. Result: Results demonstrated that the minimum inhibitory
concentrations (MICs) of vancomycin for these MRSA strains were ranged
between 0.5-1.0ug mL-1,and the MICs of nisin for these MRSA strains were
ranged between 1.25 -20 IU mL-1 with an average of 10 IU mL-1.Similar
MIC range,5-20 IU mL-1,with an average of 10 IU mL-1,was obtained for
MSSA stains.An increased sensitivity of most of the MRSA (77%) to nisin
was found,with average MICnisin reduced to 49 %,after being cultured
with L-alanine supplement (1%),it seemed that MRSA with MICVa of 1.0ug
mL-1 was affected most significant.The cytochrome c (cyt C) unbound test
results demonstrated the L-alanine uptake could decrease the bounding
capacity of cyt C to strains,which indicated that the negative charge on cell
surface was decrease. Conclusion: Furthermore,the study found that the
MRSA strain,with MICVa of 1.0ug mL-1,with less negative charge on cell
surface was more vulnerable to nisin.These finding will be help to establish the studies on MRSA for an alternative antibacterial agents.
Oral Presentation
Ya-Yen Yu, Yi-Chun Hung, Fu-Ai Chung, Szu-Yin Yu
Department of Laboratory Medicine, Chang-Hua Hospital, Ministry of Health and Welfare , Taiwan
Case Conference
Yuan-Ming Lee
Laboratory Medicine, No.152 Xin Min Rd, 26042, Taiwan
Symposium
PA-43
Keynote Speech
PA-45
Investigate Clinical Laboratory signs of patients with Mycoplasma pneumoniae community-acquired pneumonia in
Pingtung Taiwan
― retrospective study of The clinical laboratory profile in patients with suspected MP ― associated pneumonia
PA-46
Evaluating the quality of specimen before sputum culture to reduce inappropriate antibiotics prescription
Good quality of sputum culture specimen is a good way to reduce inappropriate antibiotic prescription
and labor staff workload
Wan-Wen Su, Liang-Lan Hsing, Chiung-Yu Wu, Ya-Fang Huang
Hsiu-Chen Lin1,2, Pei-Shan Wu2, Ching-Yu Chang2, Chun-hui Chung2
Department of Clinical Laboratory, Pingtung Christian Hospital, Pingtung City, Taiwan
1 Department
of Pediatrics, School of Medicine, College of Medicine, Taipei Medical University, Taiwan
2 Department
of Laboratory Medicine, Taipei Medical University Hospital
Invited Lecture
Special Lecture
Educational Lecture
Background: Mycoplasma pneumoniae (MP) is one of the main causes of
atypical pneumonia. This retrospective study is aimed to investigate the
clinical and laboratory profile in patients with suspected MP-associated
pneumonia.
Methods: 186 patients with clinical symptoms suggestive of pneumonia
during the period from October to December 2015 showed serum MPspecific IgM antibody positive. Exclusion criteria included patients with
gastroenteritis, influenza (cough, fever, rale), and chronic respiratory
disease (asthma, pulmonary tumor). The diagnosis of MP infection was
based on serological testing of antibodies by two MP-IgM ELISA systems
in Enzy-Well (Italy) or ImmunoWELL (USA). The patients were divided into
2 groups based on IgM analysis: MP negative pneumonia and MP positive
pneumonia. The cutoff criteria includes: an index value were defined as
optical density (OD) sample value/OD cutoff value ＞ 1.1 (Enzy-Well) and
serum MP-IgM levels ＞ 770 U/ml (ImmunoWELL).
Results: 63 of 186 (34%) patients with suspected pneumonia showed
serum MP-specific IgM antibody positive. The gender distribution of MPinfected patients was 35 (56%) males and 28 (44%) females. The mean +/standard deviation (SD) age of MP positive pneumonia was 6.38 +/- 3.99
years old. The patient aged from 1 to 5 years had the highest positive rate
of MP and ＞ =11 years with the lowest positive rate of MP (54% versus
14%). There were no significant differences in laboratory profile (AST, sodium, white blood cell, neutrophil, lymphocyte, and platelets, all p-values
＞ 0.05) between these groups.
Conclusion: 34% of suspected pneumonia during the study period showed
MP positive in Pingtung Taiwan. Most MP positive pneumonia was occurred in the aged group 1 to 5 years. There were no significant differences in laboratory profile between MP positive and negative pneumonia.
Objectives
The result of sputum culture is the gold standard of antibiotic treatment for
pneumonia. If sputum specimen is poor quality, such as contaminated with oropharyngeal colonizers, it may result in inappropriate antibiotic usage, prolonged
hospitalization, and increased antibiotic resistance. In addition, the contaminated
specimen also increase the overload of laboratory staff. In order to improve the
quality of sputum specimen, we introduced the Murray and Washington guideline, and spread to all medical practitioners. The guideline indicated that if the
epithelial cell ＜ 25 and neutrophil ＞ 25 at low-power field (LPF), then the specimen is good quality.
Methods:
From March 2014, each sputum culture specimen was accompanied with an
order of Gram stain in our hospital. According to the guideline, we rejected the
specimen for culture and report as mixed flora, then remark saliva contamination if the epithelial cell ＞ 25/LPF. We will not perform further isolate identification and antimicrobial susceptibility test. The positive rate of sputum culture in
two stages (Feb.2013-Feb.2014 and Mar.2014-Mar.2015) were analyzed by Ztest. The antibiotics prescription amount was presented by international scale
as defined daily dose (DDD)/1000 patient day (PD). The difference of antibiotics
prescription amount of these two stages was analyzed by Wilconxon Rank Sum
Test.
Results:
In Stage I (Feb.2013-Feb.2014), the positive rate of sputum culture was 64.1%
(2808/4381). In Stage II (Mar.2014-Mar.2015), the positive rate was 48.3%
(2232/4620). It had excluded poor sputum specimen (16.9%, 780/4620). The
difference of positive rates between these two stages was significant (p ＜ 0.01).
The third generation antibiotics prescription amounts were 130 and 117.8
DDD/1000PD in Stage I and II, respectively. The antibiotics usage in Stage I was
significant higher than Stage II (p=0.02).
Conclusion:
Evaluation of the quality of sputum culture specimen is a good way to reduce
inappropriate antibiotic prescription. It also reduce workload of microbiology
laboratory staff.
Symposium
PA-47
Application of 16S rRNA metagenomics to investigate the bacterial community
in northern Taiwan
16S rRNA metagenomics to evaluate the effectiveness of cleaning approaches
PA-48
Chi-Chao Tu1,2, Chang-Hua Chen3, Han-Yueh Kuo4, Hung-cheng Chou4, Ming-Li Liou2
Consistency of Vitek-2, MALDI-TOF, and chromogenic agar for differentiation of vancomycin-resistant Enterococcus faecium
Application of chromogenic agar for the differentiation ofEnterococcus in the inconsistent results between Vitek-2 method and
MALDI-TOF MS method
Chiao-tang Chang1, Tzu-Ling Yang1, Jao-Yu Chen1, Jia-Mei Lin1, Yu-Chiao Wang1, Shih-Chieh Lo2,
Shiao-ping Huang2
Case Conference
1 Department
of Medical Laboratory, Keelung Hospital Ministry of Health and Welfare, R.O.C, Taiwan
2 Department
of Medical Laboratory Science and Biotechnology, Yuanpei University, Hsin-Chu City, Taiwan
3 Division
of Infectious Diseases, Department of Internal Medicine, Changhua Christian Hospital Changhua City, Taiwan
4 Department
of Medicine, National Taiwan University Hospital Hsin-Chu Branch, Hsin-Chu City, Taiwan
1 Clinical
Laboratory, Yuans General Hospital, Taiwan
2 Deparment
of Medical Laboratory Science and Biotechnology, Fooyin University
Oral Presentation
Poster Presentation
The chromogenic agar contains both the isolation and differentiation purposes. The colony appearances help us in the differentiation of isolates.
The colony appearance on Chromagar VRE indicated that pink colonies
are either vancomycin-resistant Enterococcus faecium or vancomycinresistent Enterococcus fecalis , and blue colonies are either Enterococcus
gallinarum or Enterococcus casseliflavus . Both Vitek-2 automatic identification system and MALDI-TOF MS system are widely applied for the identification of Enterococcus spp. In this study, there were 22% (79/364) vancomycin-resistant Enterococcus and 21 isolates of vancomycin-resistant
enterococcus indicated the inconsistent results between Vitek-2 automatic
identification system and MALDI-TOF MS system. The result in Vitek-2 GP
card showed 13 cases of 50% E. gallinarum /50% E. casseliflavus , 6 cases
of 34% E. gallinarum /34% E. casseliflavus /33% E. faecium , 1 case of E. gallinarum and 1 case of E. casseliflavus . The results in MALDI-TOF MS system, all 21 isolates were revealed as the E. faecium . According to the color
of colony on agar, the Chromagar VRE is not merely applied for screening
of vancomycin-resistant Enterococcus but showed the results (18 cases of
E. faecium ) with high consistency to MALDI-TOF MS system (21 cases of E.
faecium ). In conclusion, the results indicated that application of Chroagar
VRE helps us to identify 86% (18/21) of inconsistent results of VRE. It is
contribute to the treatment of VRE infection.
Nowadays, transmission of healthcare-associated pathogens via surface
contamination is now a critical issue in healthcare-associated infections.
In this study, we applied a 16S ribosomal RNA (rRNA) metagenomics approach to investigate the bacterial community with two different cleaning
methods at one medical intensive care unit (MICU) and one respiratory
care center (RCC) at a hospital. MICU was performed terminal and daily
cleaning, whereas RCC was performed terminal cleaning. A total of 72
samples, including 9 samples per sampling time, were analyzed. A high
amount of microbial diversity was detected, with an average of 347 phylotypes per sample in MICU and an average of 328 phylotypes per sample in
RCC. Human skin microbiota (HSM) was predominant in both wards compared to hospital environmental microbiota (HEM). In addition, the colonization rate of HSM in the MICU is higher than that in the RCC, especially
Moraxellaceae. Net bacterial biomass was higher in non-cleaning areas
compared to cleaning areas. In addition, the lack of daily cleaning in the
RCC would contribute to the richness of bacterial communities. Comparing
five different healthcare associated pathogens in the same ward, a significant higher abundance among Acinetobacter sp., Streptococcus sp., and
Pseudomonas sp.. was shown in the RCC compared to the MICU by using
paired-t test. This is the first in Taiwan to apply the 16S rRNA metagenomics to evaluate the effectiveness of cleaning approaches.
56
Comparison of Pseudomonas aeruginosa biofilm on contact
lens; an effectiveness of contact lens care solutions
PA-50
khaemaporn boonbumrung1,2, Thanit Saeliw1, Ramin Mala-E1
Chiao-Ni Wen1, Hsiao-Chen Ning1,3, Tsu-Lan Wu2,3, Hsin-Yao Wang1, Yueh-Lin Chung1, Tsui-Ping Liu1,
Yu-Shan Wu1,3, Jang-Jih Lu1,3
1 Department
of Transfusion Medicine and Clinical Microbiology, Department of Transfusion Medicine and Clinical
Microbiology, Faculty of Allied Health Sciences, Chulalongkorn University, Thailand
2 Innovation
Center for Research and Development in Medical Diagnostic Technology, Faculty of Allied Health Sciences,
Chulalongkorn Univers
1 Department
of Laboratory Medicine, Chang Gung Memorial Hospital at Linkou, Taoyuan City, Taiwan
2 Department
of Medical Laboratories Administrative Center, Chang Gung Medical Foundation
3 Department
of Medical Biotechnology and Laboratory Science, Chang Gung University, Taoyuan, Taiwan
The clinical impact of all-the-time-processing positive blood
cultures
Symposium
PA-52
Educational Lecture
Effect of gram staining reports as soon as we get positive
signals in blood culture
Special Lecture
Blood cultures play a crucial role in the diagnosis of life-threatening bloodstream infection. Several studies have found the adequate volume of blood
drawn is the most important variable affecting positive rates of Blood
cultures. Therefore, CLSI and many guidelines recommend an optimal
volume of 20-30 mL for each set of blood bottles. However, many investigations based on cross-sectional studies just made before the existence of
automated blood culture systems. There is the necessity to reassess the relationship of blood volumes with culture rates, time to positivity, and other
clinical variables. Furthermore, in Taiwan, several hospitals could not meet
criteria of the appropriate volume duo to some difficulties in sampling process. In this way, we would like to evaluate the effectiveness of increasing
the blood volume to improve the performance of blood culture.
We conducted a systematic review and meta-analysis to reassess the
importance of blood volumes with positive rates. We searched MEDLINE
from inception to March 2016 and all identified titles and abstracts were
carefully examined by two independent reviewers. Of 531 studies, 7 papers were included and OR was the main outcome measure. The pooled
estimates under the random-effects model suggested the greater the blood
volume, the greater the culture rate in adults (10 ml blood volume versus
5 ml , pooled odds ratio [OR]=1.31, 95% confidence interval =1.03-1.67),
and an each additional 1 ml volume of blood lead to 0.8% increase in positive rates. However, there is true heterogeneity among these observational
studies ( I2=65.2%, p ＜ 0.001).
The inconsistency may arise through study populations with different
underlying diseases. In conclusion, to backup the lab practice, further
analysis will need to assess direct relationships between blood volumes
and positive rates and time to positivity.
Invited Lecture
The contact lens user has to be a significant risk factor for the development of microbial keratitis. The contact lens-related infections were led
by the improper use of lenses and their cleaning process. The contact lens
care solutions do not destroy all microbes, especially in the biofilm formation, even following instruction. The microbial communities in biofilm
are surrounded with polymeric substances which increased resistance to
antimicrobials and host immune responses. Pseudomonas aeruginosa is
frequently encountered microbes that associated with contact lens-related
infections. Such features make it resistant to some antiseptic including
contact lens solution. Some studies reported that P. aeruginosa can cause
keratitis in contact lens user due to the type of contact lens and poor quality of contact lens solution. This study compared the biofilm formation of P.
aeruginosa on contact lens in each brand, and an effectiveness of contact
lens solution on biofilm. The procedure started with culturing P. aeruginosa in Typtic soy broth in microplate at 37oC for 24 hours and then tested
with three brands of contact lens solution. This study also compared the
reduction of the living-cells in biofilm after tested with contact lens care
solution (CLCS) and control (PBS), in 2 to 3 log CFU reduction and 1 log
CFU reduction, respectively. The only contact lens solution can damage
biofilm structure but it could not against bacteria that living in biofilm
structure. The results similarly showed the biofilm formation of P. aeruginosa on all tested contact lens. This study has demonstrated that CLCS was
insufficient to prevent the contact lens-related infections on biofilm state.
PA-51
Effects of blood volume on blood culture results: a systematic
review and meta-analysis
Keynote Speech
PA-49
Junko Kawamura 1, Mitsutaka Iguchi2, Tetsuya Yagi2, Yukari Osada3, Hiroyuki Matsumoto3,
Tadashi Matsushita4
Yoshiko Shiraishi
Nagano Red Cross Hospital, Japan
Poster Presentation
57
Oral Presentation
Introduction: Processing positive blood cultures (PBCs) all the time are essential for providing early appropriate antimicrobial therapies, especially
in sepsis or bacteremia. However, 24-hour-open clinical microbiology laboratory is very rare in Japan unlike Europe or U. S. In our hospital, although
three-fourths of PBCs were detected on night shifts or holidays, further
processes were stopped until the next working day. We therefore started
to process them all the time since July 2006. Our purpose of this study is
to assess our activities toward PBCs. Our process flow: Once the automated blood culture system detects a signal, the ward with the corresponding
patient is noticed by FAX and the biomedical laboratory scientists are used
to process further steps immediately. Further steps include reporting a
result of Gram stain (GS), and processing subcultures and tentative drug
susceptibility testing by sampling from the positive blood culture bottle
according to the GS result. All the biomedical laboratory scientists participated in this process. Result: Following the procession at night shifts,
the microbiology biomedical scientists can observe colonies on the plate,
speculate bacterial/fungal species and get tentative susceptibility results,
and give doctors useful information to adjust antimicrobial therapy on
the next morning. Our process flow enabled to shorten 24-48 hours for
adjustment of inappropriate therapy. Conclusion: All the time processing
positive blood cultures had great impacts to early appropriate treatment
against sepsis or bacteremia. We’re planning to apply MALDI-TOF MS to
the work-flow for rapid identification as a next step.
Case Conference
1 Department
of clinical Laboratory, Nagoya University Hospital, Japan
2 Department
of Infectious Diseases, Nagoya Univ., Hosp., Japan
3 Department
of clinical Laboratory, Nagoya Univ., Hosp., Japan
4 Department
of Transfusion Medicine, Nagoya Univ., Hosp., Japan
Introduction : Blood culture is the gold standard for diagnosis of bloodstream infection. Many studies have shown that rapid isolation and identification of the microorganisms in blood culture are important to provide
appropriate antimicrobial therapies. It takes about half a day to obtain
these results. Therefore, we provide gram staining reports as soon as we
get positive signals on blood culture examinations as this contributes
to early reevaluation of antimicrobial therapies. This study showed that
many medical doctors require reports at all hours of the day and night, but
that this is not necessary in some cases.Methods : We conducted a survey
of more than 100 medical doctors working at our hospital. Moreover,
we investigated the medical records to determine whether the doctors
reevaluated antimicrobial therapy soon after receiving the report.Results :
The survey response rate was 65%. The doctors stated that the reports are
necessary. However, some doctors that did not respond seemed to feel that
the reports are not necessary in some cases when antimicrobial therapies
have already begun. Therefore, we developed a new gram staining report
system to resolve the differences on this issue. In the new system, doctors
must specify whether the reports will be needed at any hour of the day
and night or not when ordering blood cultures.Conclusion : We will present the results of the survey and the effects of the new system. We suggest that the system based on the doctors' instructions is useful for early
reevaluation of antimicobial therapy.
Keynote Speech
PA-53
Antimicrobial susceptibility and cfiA carbapenemase gene
positivity of Bacteroides fragilis group
PA-54
Manami Yamazaki 1, Ayako Nakamura1, Masayoshi Chonan1, Shigeki Misawa1, Takashi Horii1,
Yoko Tabe2, Akimichi Ohsaka1
Isao Nishi 1, Tomomi Mitsui1, Keigo Kimura1, Seishi Asari2, Yoh Hidaka1
1 Laboratory
for Clinical Investigation, Osaka University Hospital, Japan
2 Osaka
University Graduate School of Medicine
1 Juntendo University Hospital, Japan
2 Juntendo University School of Medicine
Invited Lecture
Special Lecture
Educational Lecture
Diagnostic procedures using endoscopy have markedly progressed, enabling precise diagnoses and minimally invasive treatments. The number
of endoscopic examinations and treatments consequently increases every
year. However, there are reports of infection via the endoscope; therefore,
a system of safe and advanced endoscopic procedures is required. At our
laboratory, we have been regularly monitoring the cleanliness of endoscopes managed by the endoscopy center since 2009 using microbial stain
and culture methods to control endoscopy-related infections. We selected
endoscopes that were frequently used in routine tests. Endoscopes that
were determined to be contaminated (bacterial growth) by the endoscope
cleanliness monitoring were retested after cleaning and disinfection. If an
endoscope was determined to be contaminated by the second test, we requested maintenance from the manufacturer and reported the case to the
nosocomial infection control committee. From 2009 to 2015, we examined 180 gastroscopes, 199 colonoscopes, and 253 bronchoscopes. Fortytwo gastroscopes and 29 colonoscopes tested culture-positive. The main
bacteria detected were Pseudomonas aeruginosa and Klebsiella pneumoniae for gastroscopes and Enterococcus spp. and Escherichia coli for colonoscopes. To further examine the cause of contamination, we examined the
cleaning and disinfection process for endoscopes, environment of the endoscope cleaning room, environment of the endoscopy center, endoscope
lumen, and surveyed the cleaning staff. Because endoscope contamination
was confirmed, despite using an automatic endoscope reprocessor, it is
important not to overestimate the cleaning and disinfection effectiveness
of the automatic endoscope reprocessor, and regular monitoring of endoscope cleanliness requires to be performed by a microbiology laboratory
to control infections associated with endoscopes. Monitoring of endoscope
cleanliness performed by the microbiology laboratory can prevent nosocomial endoscopy-related infections and be useful in determining the timing
of endoscope maintenance.
Background: Bacteroides fragilis group is responsible for the development of serious infections, such as intra-abdominal and bloodstream
infections. A variety of drug-resistant strains with metallo- Β -lactamase
production or metronidazole resistance have been reported, and the efficient monitoring of the drug-resistance, especially the resistance of carbapenems, is crucial for infection control in Japan. Materials and method:
Strains isolated from the clinical material in Juntendo university hospital
from January 2013 to October 2014, B. fragilis (83 strains), B. thetaiotaomicron (28 strains) and the other 9 species (39 strains), were used for
this study. Strains were isolated by Anaerocolumbia rabbit blood agar (BD),
then identified their species settlements by MALDI biotyper (Bruker). Drug
susceptibility testing has been performed using Kyokuto MIC plate (Kyokuto) following the CLSI performance standards (M100-S24). The cfiA gene
encoding carbapenemases was detected by PCR (Soki's method).Results:
Percent of drug-sensitive strains against major antimicrobials; B. fragilis
(%) /B. thetaiotaomicron (%) / others (%), ABPC/SBT, 77/68/69; PIPC/TAZ,
100/93/97; CMZ, 75/4/28; CLDM, 52/14/41; MFLX, 78/21/56. The MICs
of MEPM were 2-3 folds higher than those of IPM. No MNZ resistance was
found in all tested strains. Multidrug-resistance except metronidazole were
observed in 2 strains of B. thetaiotaomicron. cfiA gene has been detected
in 12 strains, and all of them were B. fragilis with MEPM MIC > 2µg/mL.
Conclusions: Because cfiA gene has been detected in carbapenem susceptable strains, the only drug susceptibility test likely to be missed was
suggested. For the screening test of carbapenemase-producing B. fragilis ,
using MEPM is more effective than IPM.
Symposium
PA-55
Endoscopy-related infection control using the microbiological
monitoring
Epidemiological chase of MRSA using POT method in NICU
and GCU for 20 months
PA-56
Tomomi Mochimaru 1, Yuiko Morokuma1, Makiko Kiyosuke1, Ruriko Nishida1, Hisanori Nishio2,
Nobuyuki Shimono2, Taeko Hotta1, Dongchon Kang1
Molecular epidemiological analysis of MRSA by PCR-based
ORF Typing(POT) method
Yumiko Kimura , Norihito Kaku, Kousuke Kosai, Yoshitomo Morinaga, Katsunori Yanagihara
Department of Laboratory Medicine, Nagasaki University Graduate School of Biomedical Sciences, Japan
Case Conference
1 Department
of Clinical Chemistry and Laboratory Medicine, Kyushu University Hospital, Fukuoka, Japan
2 Center
for the Study of Global Infection, Kyushu Univ., Hosp.
Oral Presentation
Poster Presentation
Background: Multilocus sequence typing(MLST) and Pulsed-field gel
electrophoresis(PFGE) have been widely used in epidemiological analysis
of MRSA. However, it is very difficult to perform them in the community
hospitals, because these methods are complicated and relatively expensive. Recently, a new epidemiological analysis of MRSA, polymerase
chain reaction(PCR)-based ORF Typing(POT) method, was developed.
The POT method could classify the MRSA strains by analyzing the pattern of detected amplified bands, and, based on the POT1-value, clonal
complex(CC) type and SCCmec type could be estimated. The operating
time is approximately 4 hours. In this study, we performed a molecular
epidemiological analysis of MRSA isolated from the skin and soft-tissue
using the POT method. In addition, we were also investigated possession
of Panton-Valentine leukocidin(lukS/F -PV) gene. Material/methods: We
used the MRSA 90 strains, which were isolated from the skin or softtissue at the National Hospital Organization Nagasaki Kawatana Medical
Center between 2005 and 2011. We analyzed the strains by using the
Cica Geneus Staph POT-kit(Kanto Chemical co., Tokyo, Japan). In addition,
we detected the Panton-Valentine leukocidin(lukS/F -PV) gene by real-time
PCR.Results:The POT-value of MRSA 90 strains were classified into 59
patterns. The most common POT-value(POT1:106, POT2:137, POT3:80,
106-137-80) was detected in 10 strains(11.1%), and the second most
common POT-value(98-155-79) was detected in 6 strains(6.7%). The lukS/
F -PV was detected in 4 strains. The POT-value of them were 106-127113(2 strains, CC8 and SCCmec type IV) and 110-4-37(2 strains, CC30
and SCCmec type IV). Based on the POT-value, they were differ from USA300 strain.Conclusions:The POT method could classify the MRSA strains
and estimate the MRSA clone. The POT method is a very useful molecular
epidemiological analysis of MRSA as well as the PFGE. Since the operating
time is short(approximately 4 hours), the POT method can contribute to
the quick determination of MRSA outbreak.
Introduction: Newly isolated MRSA strains in NICU and GCU were analysed by using the PCR based open reading frame typing (POT) method.
This method is one of the molecular epidemiological methods. The procedure of POT is quite easier than PFGE whereas its resolution power is
similar to that of PFGE. Here we report the POT analysis data of MRSA
in a long period of 20 months. Methods: A total of 82 MRSA strains
(NICU:67,GCU:15) were collected from May 2014 to December 2015, and
were analyzed by POT. A Cica Geneus Staph POT KIT (Kanto Kagaku) was
used for this analysis. Results: All isolates were classified into 20 POT
types. There were 2 major strains, typeA (POT number 106-183-34, 35%)
and typeB (POT number 106-9-80, 17%). TypeA strains were detected
from multiple patients in NICU and GCU at the same time, and continued
to be detected sporadically over following 6 months. TypeB strains were
detected in succession during four months in NICU. The TypeB strains
disappeared once, but were detected again 10 months later. The isolation
of both strains intermittently lasted for long periods. Some strains with
the same POT types were isolated from stethoscope earpieces of the hospital staff. Consequently, we started to disinfect the stethoscope earpieces
and strengthened the education of hand hygiene. The same POT type
strain has not been detected afterward. Disccusion: This 20-month-study
revealed that two strains were predominantly detected for the long term
from NICU in our hospital. The PCR based results are transformed into a
three-part POT number. So it is quite feasible to compare clinical strains
isolated over time and also to compare clinical and environmental strains.
The POT method is a useful epidemiological method for estimating the
transmission route and eventually for infection prevention.
58
Performance Evaluation of rep-PCR Analysis for MRSA
Clone Identification Application
PA-58
Yumiko Funashima 1, Zenzo Nagasawa1, Osamu Ueda2, Kyohei Kato3, Kenichi Sato1, Hideaki Hanaki4,
Hiroshi Miyamoto2
Chitoshi Sato 1, Keita Okamura2, Shiba Kumar Rai3, Paller VG4, Tadayoshi Hata5, Seiji Imamura5
1 Department
of Clinical Laboratory, Okazaki city Hospital, Japan
2 Division
of Central Clinical Laboratory, Mie University Hospital
3 Shi-Gan
Int'l College of Science and technology & Nat'l inst of Tropical Medicine and Public Health Reserch
4 Institute
of Biological Sciences, Univ. of the Philippines Los Banos
5 School
of Health Sciences, Fujita Health University
1 Department
of Medical Technology and Science, Fukuoka Health Care Faculty, International University of Health and Welfare,
Japan
2 Microbiology
Discipline, Pathology Course, Saga University Medical School
3 Department
of Clinical Labotatoriy,Medical Kouhoukai Takagi Hospital
4 Infection
Control Research Center, Kitazato University
Educational Lecture
Symposium
Ten-year observation of Blood Culture Results in a Japanese University Hospital
Impact of Improving Infection Control Activities over the Past Decade on Blood
Culture Results
Special Lecture
PA-60
Seasonal variation in blood stream infection in Japan
Invited Lecture
Introduction : Most of developing country has inferior public health environment. In this environment, some researchers reported a lot of people
are infected with diarrhea disease, especially enterocolitis cause high death
rate of infants and children. The aim of this study to investigate pathogenic
bacteria in school children, in Philippin and Nepal.Methods : Fecal samples
from 7 to 14 years school children were collected in these countries and
examined in Japan. There were 70 samples of Philippine school children
and 150 samples of Nepal school children examined. Investigation of bacteria in this study was Vibrio cholera, Vibrio parahaemolyticus, Salmonella
spp., Shigella spp., Staphylococcus aureus and diarrheagenic Escherichia
coli. In addition, feces in Nepal school children were examined for drugresistant (KPC, MRSA and ESBL). Results : Feces of Philippine school children were detected of Salmonella spp, MRSA and genes of diarrheagenic
Escherichia coli (EPEC eae, EAggEC aggR, EIEC invE). On the other hand,
feces of Nepal school children were detected of genes of diarrheagenic Escherichia coli (astA, aggR and LT) only. Conclusion : In developing country,
people easily get some antibiotic without medical prescription. Therefore,
those system cause drug-resistant bacteria increasing than Japan whose
system require medical prescription for getting antibiotic. However, we
demonstrated that a carrier rate of ESBL with drug-resistant are not difference between Nepal school children and Japanese. This study is contributed to be improvement of public health because those result appear
pathogenic bacteria in detail and be international cooperation between
Japan and Philippine and Nepal for human health.
Objectives: We have reported the epidemiological analysis procedure for
clone identification of methicillin-resistant Staphylococcus aureus (MRSA)
using matrix-assisted laser desorption/ionization-Time of flight mass spectrometry (MALDI-TOF MS;MALDI) , which had been preliminarily identified by Pulsed-field gel electrophoresis (PFGE) and Phage ORF Typing (POT)
techniques. The aim of this study was to evaluate the performance equivalency of repetitive extragenic palindromic PCR (rep-PCR) analysis for
MRSA clone identification compared to other three techniques; PFGE, POT
and MALDIMaterials and Methods: A total of twenty four MRSA clinical
isolates were collected from health care associated infection (HAI) sources.
Clinical isolates were treated with restriction enzymes Sma I for PFGE profiles and POT analysis was performed using a Cica Geneus® Staph POT
kit The DiversiLab® automated microbial typing systemwas used for repPCR analysis. PFGE, POT and rep-PCR analyses were performed according
to the manufacturer's IFUs. All specimen for MALDI measurement was
pre-processed under ethanol-formic acid extraction. MALDI Biotyper was
used for MALDI measurement. Results: All three analyses result of PFGE,
POT and clone-specific peaks of MALDI were completely matched each
other and were classified into six clone types for twenty four MRSA clinical
isolates. But multivariate analysis results and phylogenetic tree analyses
by MALDI Biotyper for all peaks of MALDI were classified into seven clone
types and eight clone types, respectively. A part of clone types were not
thoroughly matched with PFGE analyses. rep-PCR analysis results were
not thoroughly matched with PFGE analyses, neither and were classified
into fifteen, eleven, and nine clone types for XJ99%, KL99%, and PC99%,
respectively. Similar results were obtained from rep-PCR analysis for
XJ95%, KL95%, and PC95% but these were not completely matched with
PFGE profiles.
PA-59
Detection for pathogenic bacteria in feces of school children
in Philippine and Nepal
Keynote Speech
PA-57
Kazunari Yasuda 1, Masaki Tanabe2, Akiko Nakamura1, Toru Ogura3, Makoto Morimoto1,
Kazushi Sugimoto1, Kaname Nakatani1
Shinobu Ikegami , Hideki Nishyama, Teruaki Ohya, Makoto Minoshima, Hideki Kato, Yuasa Norihiro
Japanese Red Cross Nagoya Daiichi Hospital, Japan
Poster Presentation
59
Oral Presentation
Introduction：Infection control activities in Japan have been drastically
improved over time. Blood cultures (BCs) are important for diagnosis of
bacteremia and the number of BCs can be a marker of infection control activities. Our objective was to assess infection control activities through BCs
results over the past decade in our hospital.Methods：We retrospectively
analyzed all consecutive BCs performed in the Mie University Hospital,
Japan between January 2006 and December 2015. The blood culture
results were statistically analyzed by simple linear regression model. The
independent variable for this model was year, and the dependent variables
included annual incidence and multiple sets percentage of BCs, positive
BCs rates (excluding possible contaminant organisms), and the prevalence
of methicillin resistance among Staphylococcus aureus . P < 0.05 was
considered statistically significant.Results：A total of 23,271 BCs was
performed in 12,593 cases (11,584 inpatients and 1,009 outpatients), and
a total of 10,678 follow-up BC was performed in 4,136 patients. There
showed a significant increment in annual inpatient incidence of BCs per
1,000 new admissions (coefficient of year:1.656, 95% CI = 0.696 to 2.617;
p = 0.004) and in multiple sets of BCs (coefficient of year: 0.057, 95% CI
= 0.043 to 0.071; p < 0.001). There was no significant changes in positive
BC rates (coefficient of year: 0.052, 95% CI = -0.100 to 0.204; p = 0.450).
The prevalence of methicillin resistance among Staphylococcus aureus isolates decreased from 77.8% in 2006 to 37.1% in 2015 (coefficient of year:
-0.057, 95%CI = -0.083 to -0.032; p = 0.001).Conclusion：Frequencies of
BCs performed and their multiple sets percentages showed positive increment over the past decades without changes in positive BCs rates, in addition, percentage of methicillin resistance among Staphylococcus decreased
over time. These favorable trends may be related to continuing infection
control activities in recent years.
Case Conference
1 Department
of Central Laboratory, Mie University Hospital, Japan
2 Dept.
of Patient Safety and Infection Control, Mie Univ., Hosp.
3 Dept.
of Clinical Research Support Center, Mie Univ., Hosp.
Background and purpose: It is well known that several diseases have a
winter peak, however, seasonal variation of blood stream infection (BSI)
remains controversial. The aim of this study was to clarify seasonal variation of blood stream infection in Japan. Subject and methods: From
January 2005 to December 2014, the accumulated number of admissions
and positive blood cultures were 2,747,859 and 5,062, respectively. Only
the first blood culture for each patient/episode was included. The crude
incidence rates (IR) of BSI was calculated per 1000 hospital admissions.
Spring comprised March, April, and May; Summer-June, July, and August;
Autumn-September, October, and November; and Winter-December, January, and February. The association of IR of BSI with temperature was
evaluated by stratifying the data into each month.Results: The IR of BSI
was 1.84 during the study period. The IR of BSI from January to December
were 1.83, 1.63, 1.55, 1.57, 1.75, 1.90, 2.01, 2.22, 1.98, 1.90, 1.91, 1.85,
respectively, indicating monthly and seasonal variation (p<0.05). The most
common isolate was Gram-negative rod (GNR) followed by coagulase-negative staphylococci (CNS), Streptococcus sp ., Staphylococcus aureus , and
Bacillus sp . (The IR of BSI: 0.69, 0.38, 0.21, 0.20, and 0.14, respectively).
The IR of GNR-BSI from January to December were 0.65, 0.57, 0.55, 0.50,
0.57, 0.66, 0.91, 0.85, 0.80, 0.80, 0.73, and 0.67, indicating monthly and
seasonal variation (p<0.001). There was no significant monthly or seasonal variation in other bacterial species. The IR of BSI by GNR, CNS, and
Bacillus sp . were significantly correlated with temperature in Nagoya city
during the study period. Conclusions: The incidence of BSI had seasonal
variation, and was increased with temperature. This was primarily due to
seasonal variation of GNR-BSI.
Keynote Speech
PA-61
Dissemination of extended-spectrum beta-lactamase producing
PA-62
Escherichia coli in Indonesia
Epidemiolgy analysis of the Acinetobacter by the Phage Open
Reading Frame Typing method
Akiho Kanaida 1, Kayo Osawa2, Katsumi Shigemura2, Kuntaman3, Dadik Raharjo4, Eddy Bagus Wasito4,
Subijanto Marto Sudarmo5, Toshiro Shirakawa6
Suguru Hiramoto 1, Hiroki Machida1, Rumi Okazaki1, Azusa Uchida1, Miki Takahashi1, Tetsuo Machida1,
Haruyoshi Tomita2, Masami Murakami3
1 Department
of Biophysics, Kobe University Graduate School of Health Sciences, Japan
2 Department
of Biophysics, Kobe University Graduate School of Health Sciences
3 Department
of Clinical Microbiology, Dr. Soetomo Academic Hospital-School of Medicine, Airlangga University
4 Department
of Microbiology, Faculty of medicine, Airlangga University
5 Department
of Pediartrics, Faculty of Medicine, Airlangga University
6 Indonesia-Japan
Collaborative Research Center for Emerging and Re-emerging Infectious Diseases, Institute of Tropical
Disease, Airlangga University
1 Clinical
Laboratory Center, Gunma University Hospital, Japan
2 Depratment
of Bacteriology,Gunma University Graduate School of Medicine
3 Department
of Clinical Laboratory Medicine, Gunma University Graduate School of Medicine
Invited Lecture
Special Lecture
Educational Lecture
Introduction:The Acinetobacter is a gram-negative bacillus of the glucose
non-fermentation. Acinetobacter baumannii (A.baumannii ) is one of the
causative organisms of hospital infection. It is necessary to confirm the
presence or absence of blaoxa-51-like that A.baumannii contains on its
chromosome, and 16srRNA genetic analysis for accurate identification of
the strain. Therefore,it is difficult to identify A.baumannii in the routine
examination. Recently Phage Open Reading Frame Typing (POT) method
has been introduced to clinical laboratory. POT enables us to identify
several kinds of bacteria including A.baumannii , and to perform epidemical analysis among the different strains.Methods:We studied 13 strains
of Acinetobacter genus bacteria isolated in our hospital from January to
December in 2014.In addition to the POT method, we performed drug
sensitivity testing and pulsed-field electrophoresis. We performed POT
analysis and pulsed-field electrophoresis according to the manufactures
instructions. Results:Thirteen strains were classified into eight POT types
by the POT method. We were able to identify International clone II in one
strain. Conclusion:With the POT method, it took only four hours for us to
identify the bands from DNA extraction of the bacteria.The preparation of
the reagents are easy, and specific equipments are not necessary for POT
method. Therefore, POT method is possibly used in many clinical laboratories, and is a very useful tool to analyze strains for suspected nosocomial
infections in clinical settings.
Introduction: Extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli is one of the health problems worldwide. The ESBL-encoding
genes are known as blaCTX-M, blaTEM, and blaSHV, and the spread of blaCTX-M-15
cause great concern. Additionally, it has been reported the global spread of
the ESBL producing E. coli belonged to clonal complex (CC) 131 or CC10
by multilocus sequence typing (MLST). In Indonesia, bacterial diarrhea
caused by diarreagenic E. coli (DEC) is one of the most common causes
of mortality among children. There are few reports about antimicrobials
resistant DEC in Indonesia. The present study was aimed to investigate the
antimicrobial susceptibility and molecular characteristics of clinical E. coli
isolated from pediatric diarrhea patients in Indonesia. Methods: A total of
124 E. coli strains were isolated from pediatric diarrhea patients in Soetomo Academic Hospital, Surabaya, Indonesia between January 2012 and
December 2012. All isolates were tested 17 antimicrobial susceptibilities
and screened ESBL-producing. We detected the presence of blaCTX-M, blaTEM,
blaSHV and DEC-encoding genes by PCR and sequencing. ESBL-positive
isolates were analyzed for phylogenetic relationships using MLST. Fisher’s
exact tests were used to evaluate relationships between variables. Results:
ESBL-producing E. coli were detected in 25 (20.2%) of 124 strains. ESBLproducing strains had significant resistance to aztreonam**, piperacillin**,
gentamicin**, nalidixic acid** and ciprofloxacin** more than non ESBL-producing strains (**: p <0.01). Twenty-six (21.0%) strains including 2 (1.6%)
ESBL-producing strains had DEC-encoding genes. Among 25 ESBL-producing strains, 21 (84.0%) strains, containing 9 (36.0%) strains belonged
to CC10, were positive of blaCTX-M-15. Conclusion: ESBL blaCTX-M-15-producing
E. coli belonged to CC10 were isolated from pediatric diarrhea patients in
Indonesia, and had higher antibiotic resistance. The prevalence of these
strains were leading to a dissemination in Indonesia, since these genes are
major causes of antibiotic treatment failure.
Symposium
PA-63
Comparison of phage open reading flame typing and rep-PCR method
for genetic typing of multi-drug resistant Pseudomonas aeruginosa
strains
PA-64
Antimicrobial resistance pattern of Campylobacter coli
isolated from diarrheal patients and raw meat samples
Case Conference
Oral Presentation
Poster Presentation
Megumi Oho 1, Chinatsu Komatsu1, Yusuke Hashimoto1, Koji Kusaba1, Takanori Higashitani1,
Zenzo Nagasawa2, Hiroshi Miyamoto3, Eisaburo Sueoka1
Miyuki Fujioka 1, Takuya Sato1, Yurie Kudo2, Ryuna Sato1, Ayaka Ota2, Chikao Yoshida2,
Hiroyuki Nozaka1, Chowdhury Rafiqul Ahsan3
1 Department
of Laboratory Medicine, Saga University Hospital, Japan
2 Department
of Medical Technology and Sciences, International University of Health and Welfare, Fukuoka
3 Department
of Pathology and Microbiology, Faculty of Medicine, Saga University
1 Graduate
school of health sciences, Hirosaki univ., Japan
2 School
of health sciences, Hirosaki univ.
3 Dept.
of Microbiology, Univ. of Dhaka
IntroductionAlthough Pseudomonas aeruginosa usually rarely cause
infection in healthy people, it is important as bacteria causing healthcareacquired infections, such as opportunistic infections for the compromised
host in hospitalized patients. Among the Pseudomonas aeruginosa , the
multi-drug resistant Pseudomonas aeruginosa (MDRP) infections increase
mortality, morbidity, and hospital costs. We conducted a comparative
study of phage open reading flame typing (POT) and rep-PCR method for
genetic typing of multi-drug resistant Pseudomonas aeruginosa strains
in monitoring nosocomial infection.Subjects and Methods The subject
strains as MDRP used in this study were obtained from the stored strains
in Kitasato University Research Center for infections and antimicrobials, and those in Saga University Hospital. POT method was conducted a
multiplex PCR using a Cica Genius Pseudo POT kit® (Kanto Chemical Co.,
Inc.), following assessment and analyses of DNA band patterns. The repPCR method was carried out using a DiversiLab®fingerprinting kit (Sysmex
bioMerieux), and the obtained PCR products were analyzed by DiversiLab
System. Over 95% similarity by fingerprinting patterns of PCR products
was diagnosed as similar strains.ResultThe 48 strains examined were classified into 12 types by POT, and into 16 types by rep-PCR method, respectively. The results using both methods showed good correlations except for
a small discrepancy. DiscussionPulsed-field gel electrophoresis (PFGE) is a
standard method for molecular epidemiological study of microorganisms.
However, the method is expensive, time consuming, and requires specific
technique to conduct. Compared with PFGE, POT and rep-PCR methods
are simple and easy by using a kit, and it takes about only 4 hours for an
assay. Furthermore, it is also possible the comparison with the previous
data, and thus, both methods are useful for applying nosocomial infection
control.
Background: Campylobacter is widely known as one of the most common food borne pathogens and C. coli is a predominant causative agent
of Campylobacteriosis. In recent years, several studies have reported the
increasing levels of antimicrobial resistance in C. coli . The resistant strains
have been isolated from patients with diarrhea, who were not exposed to
any antimicrobials, and it is thought that the antimicrobial resistant strains
may have originated from domestic animals. In this study, a comparison
between fecal and raw meat originated C. coli was made, for better understanding of the present resistance pattern of this organism isolated from
local sources. Materials and Methods: Fecal samples from 100 diarrheal
patients and 132 raw meat samples (poultry: 66, swine: 43, and bovine:
23) were collected from retail stores located in Hirosaki city, Japan. C. coli
was isolated by the rinsed sample culture method and identified through
multiplex PCR. Antimicrobial resistance pattern was tested using the disk
diffusion assay using erythromycin (EM), tetracycline (TC), ciprofloxacin
(CPLX), ofloxacin (OFLX), and fosfomycin (FOM). Results: A total of 23 C.
coli (10 from fecal and 13 from raw meat samples) were isolated and the
results of the disk diffusion assays revealed that the isolates from feces
and raw meat samples had the following respective values of resistance to
antibiotics: EM (30.0%, 23.0%), TC (70.0%, 53.8%), CPLX (30.0%, 30.8%),
OFLX (30.0%, 30.8%), and FOM (0%, 0%). Discussion: FOM-resistance was
not observed in any of the C. coli isolates. On the other hand, TC-resistance
was found to be high (53.8-70.0 %), particularly, in strains of fecal origin
(70.0 %). Overall, the resistance pattern of C. coli was found to vary for
each antimicrobial and no relationship was observed between the antimicrobial resistance of C. coli isolated from fecal samples and that from raw
meat samples.
60
1
1
1
1
1
PA-66
1
Yuta Kamimura , Shinobu Ishigaki , Miwa Asahara , Yoshiko Atsukawa , Junpei Sasaki , Lisa Ichikawa ,
Yasuo Ono2, Taiji Furukawa1
- approach for establishing a standardized method -
Teruko Komoda, Shohei Maida, Miki Gen, Minami Sato, Ami Uchida, Hisaichi Bannai
Kyorin University, Japan
1 Department
of Cnetral Clinical Laboratory, Teikyo University Hospital, Japan
2 Department
of Microbiology & Immunology, Teikyo University School of Medicine
Symposium
Tatsuya Negishi 1, Takehisa Matsumoto2, Eriko Kasuga1, Kazuki Horiuchi1, Tatsuya Natori1,
Kenta Takehara1, Mitsutoshi Sugano1, Takayuki Honda1
The effect of farnesol on virulence factors of clinical isolates
of Pseudomonas aeruginosa
Educational Lecture
PA-68
Special Lecture
Analysis of thyA mutation in thymidine-dependent smallcolony variants of Escherichia coli
Invited Lecture
Introduction: A unified method is needed for routine works and epidemiological studies of Chlamydophila pneumoniae (C. pneumoniae ) infections. The study aims to establish a standardized method for isolation of
C. pneumoniae organisms.Methods: Three experiments were conducted
to determine the optimal conditions of C. pneumoniae isolation. 1) The effect of swabs of raw-materials (rayon for PL6S and BD BBL Culture Swab
PlusTM, polyester for BD-UVT) and transport medium (SPG and BD-UVT)
on the number of inclusions. 2) To determine the cell group with highest
sensitivity to C. pneumoniae , number of inclusions and content of C. pneumoniae specific DNA were compared among five different cell cultures: HL,
HeLa229, Hep#2, BICR18 and ChaGo-K-1, respectively. 3) HL cell cloning
was done to get the highest sensitive cell to C. pneumoniae by a limiting
dilution method.Results: Similar sizes and number of inclusions were obtained in both SPG and commercially available transport medium: BD-UVT.
A few inclusions were counted in the combination of PL6S swab (made of
rayon) and BD-UVT. HL cells were distinguished into 29 cell groups (HLKG1-KG29). The result was that the number of inclusions was BICR18 ＞
HL ＞ Hep#2 ＞ =HeLa229 ＞ ChaGo-K-1 ＞ HL-KG22. The sizes of inclusions were BICR18 ＞ HL ＞ =HL-KG1 and they were definitively larger
than that in HL-KG22 and ChaGo-K-1. Many large-sized inclusions were
observed in HL-KG1 cells and BICR18 cells. There were no discrepancies
between the number of inclusions and DNA content determined by realtime PCR.Conclusion: Results show that swabs of raw-materials were not
so effective on the number of inclusions formed when SPG was a transport
medium. Although BICR18 cells formed many inclusions, they were not
easy to make inti monolayers. We recommend the use of SPG (or BD-UVT)
for transport and stored mediums, and HL-KG1 cells for isolation.
Purpose:Extended spectrum-beta-lactamase (ESBL) producing bacteria
continue to be a major problem of healthcare-associated infection, and
also shows a change in drug susceptibility. Therefore, we investigated the
trend of ESBL producing strains at our hospital.Method:The subjects were
Escherichia coli , Klebsiella pneumoniae , and Proteus mirabilis of those
which drug susceptibility tests were conducted from 2000 to 2015. The
isolation frequency and trend of drug susceptibility of ESBL producing
bacteria were investigated using microscan panel (Beckman Coulter, Inc.)
Results:The occurrence of ESBL producing strain was 53 out of 1355 E.
coli (3.9%) in the first year (2000). That was 105 out of 699 K. pneumoniae (15.0%), and 0 out of 77 P. mirabilis . In the year 2015, the number
changed as follows: 258 out of 1174 E. coli (22.0%), 42 out of 500 K.
pneumoniae (8.4%), and 7 out of 80 P. mirabilis (8.8%). Among ESBL
producing bacteria, E. coli gradually shows an increase and surpass the
number of K. pneumoniae in 2006. K. pneumoniae shows a peak in 2000
and decreased to 3-9%. P. mirabilis were detected from 2004 and altered
(4-20%). The number of LVFX resistant strain in the year 2000 compared
to 2015 were as follows: E. coli 4 strains (8%) to 197 strains (76%), K.
pneumoniae 1 strain (1%) to 5 strains (12%), and P. mirabilis none to 2
strains (29%).Consideration:Out of ESBL producing bacteria detected at
our hospital, the results show that E. coli has gradually increased the number yearly and show a high rate of resistance to newquinolone other than
beta-lactam. Considering the results, further awareness will be needed
among isolation situations and the trend of drug susceptibility.
PA-67
Studies of isolation methods for diagnosis of Chlamydophila
pneumoniae infections
Keynote Speech
The isolation situation and drug susceptibility among
Extended spectrum-beta-lactamase(ESBL) procuding bacteria
at Teikyo University Hospital
PA-65
Shuhei Ishikawa 1, Ryoya Matsumoto2, Koichi Suzuki2, Yoko Mano2, Nobuhiko Furuya2
1 Department
of Laboratory Medicine, Shinshu University Hospital, Japan
2 School
of Health Sciences, Faculty of Medicine, Gunma University
Poster Presentation
61
Oral Presentation
Introduction: Pseudomonas aeruginosa is the most common Gramnegative pathogen causing nosocomial pneumonia, and it is frequently
implicated in hospital-acquired urinary tract and bloodstream infections.
It is commonly found in mixed infections with the yeast, Candida albicans .
The fungus produces farnesol activities as an effector in interspecies interaction. In this study, we investigated how the C. albicans farnesol might
affect clinical isolates of P. aeruginosa regarding virulence factors, such
as pyocyanin, elastase and total protease. Methods:The effects of farnesol
on virulence factors of P. aeruginosa clinical isolate were phenotypically
tested. Pyocyanin was extracted from culture supernatants and its absorption at 520nm was measured. Elastase and total protease activities were
determined using the Elastin Congo red (Sigma) assay and the Remazol
Brilliant Blue-Hide (Sigma) assay, respectively.Results:C. albicans culture
supernatants reduced the production of P. aeruginosa pyocyanin by
65.5%. Pyocyanin production levels in all P. aeruginosa culture supernatants in the presence of farnesol were lower than those in the absence of
farnesol. The reduction rates of elastase and total protease production in
the P. aeruginosa culture supernatants in the presence of farnesol had a
broad range (0-41%) and the mean elastase and total protease production
levels within P. aeruginosa culture supernatants with farnesol were tended
to produce lower than those in absence of farnesol, respectively.Conclusion: Exogenous addition of farnesol resulted in decreases of P. aeruginosa
exoproducts. These results suggest that farnesol may contribute to the
decrease of P. aeruginosa pathogenesis.
Introduction: Small-colony variants (SCVs) are mutant strains that grow
slowly and show atypical colony morphology. Thymidine-dependent SCVs
(TD-SCVs) are dependent on thymidine for their growth. In Staphylococcus
aureus , the occurrence of TD-SCVs is mainly due to a mutation in thyA ,
the structural gene of thymidylate synthase. The aim of this study was to
analyze the thyA sequences in TD-SCVs of Escherichia coli .Methods: In
this study, we used 5 strains of E. coli TD-SCVs, which were obtained from
clinical fecal samples. The revertants successfully grew on Mueller Hinton
medium and showed a normal phenotype. We sequenced the thyA of both
TD-SCVs and their revertants.Results: In three TD-SCVs, individual amino
acid mutations, p.Leu44Gln, p.Trp133Arg, and p.Ser180Arg, caused by
c.131T ＞ A, c.397T ＞ A, and c.540C ＞ A, respectively, were observed.
Another TD-SCV showed a nonsense mutation of p.Gln191X, produced by
c.571C ＞ T. The final strain contained a frameshift mutation caused by
c.143_144ins with a 1201-bp fragment. Four revertants of the TD-SCVs,
except for one with an insertion, were obtained. The revertant of TD-SCV
with p.Trp133Arg showed substitution of the same amino acid as the wildtype strain. The revertants of TD-SCVs with p.Ser180Arg and p.Gln191X
showed substitutions to different amino acids. The TD-SCV with p.Leu44Gln
showed the same thyA sequence to its revertant, which was thymidine-independent.Conclusion: In E. coli , thyA mutations are a major cause of the
occurrence of TD-SCVs, similar to that in S. aureus . However, other factors
in addition to thyA mutation may result in TD-SCVs because one isolate
showed no difference in the thyA sequence between the TD-SCV and the
revertant.
Case Conference
1 Graduate School of Health Care Science, Bunkyo Gakuin University, Japan
2 Bunkyo
Gakuen Univ.
Keynote Speech
PA-69
Identification of genes associated with penetration activity of
the human type of Edwardsiella tarda EdwGII
PA-70
The association between virulence factors and viscosity of
hypermucoviscosity phenotypes in Klebsiella pneumoniae
Semiquantitative evaluation of viscosity using venire calipers
Chigusa Suezawa 1, Masashi Yasuda1, Kiyoshi Negayama2, Taeko Kameyama3, Misato Hirauchi3,
Toshihiro Nakai4, Jun Okuda1
Norihito Morimoto1, Yoshie Nishida1, Miyuki Morimoto1, Saya Kamioka1, Tamae Morita1,
Katsumi Ogura1, Tetsuro Sugiura2, Yoshihisa Matsumura2
1 Dept.
of Microbiology, Kagawa Prefectural Univ., of Health Sciences, Japan
2 Dept.
of Clinical Laboratory, Kagawa Univ., Hosp.
3 Dept.
of Central Clinical Laboratory, Kagawa Prefectural Central Hosp.
4 Graduate
School of Biosphere Science, Hiroshima Univ.
1 Department
of Clinical Laboratory, Kochi Medical School Hospital, Kochi University, Japan
2 Dept.
Clin. Lab. Med. Kochi Med. Sch. Kochi Univ.
Invited Lecture
Special Lecture
Educational Lecture
Background: Klebsiella pneumoniae is normally found in the human intestines, and is the cause of respiratory tract and urinary tract infections. Hypermucoviscosity (HMV) phenotypes, particularly in serotypes K1 and K2,
of K. pneumoniae have been associated with invasiveness and pathogenicity. This study aimed to investigate the association between the presence
of virulence factors and viscosity in K. pneumoniae .Materials & methods:
We examined 47 K. pneumoniae clinical isolates. The isolates were mainly
obtained from sputum and bronchoalveolar lavage fluid, followed by
blood, pus, urine, vagina swab, pleural effusion, synovial fluid and feces.
HMV phenotypes were determined using the modified string test using
digital venire calipers (a string measuring 5 mm or longer was defined as
positive). Then, the viscous degree was classified as ＞ 10 mm (P2) or 5
- 10 mm (P1). Screening for capsular serotype (K1, K2 and K5) and virulence factor genes (magA , rmpA , uge and kfu ) was determined by PCR. Antibiotic susceptibility was assessed using the broth microdilution method.
Results: Sixteen of 47 isolates showed HMV phenotypes (P1: 5, P2: 11),
and 10 of these were serotype K1 or K2. Serotype K1 or K2 was detected
more frequently in P2 than P1 isolates. Regarding HMV phenotype, 15
(93.8%) and three (18.8%) isolates harbored rmpA and magA , respectively.
Only one rmpA -positive isolate exhibited a non-HMV phenotype. Overall,
no resistant strains were detected.Discussion: Nearly all HMV phenotypes
of K. pneumoniae harbored rmpA . However, more serotype K1 or K2 isolates exhibited high-degree viscosity in HMV phenotypes. These findings
suggest that high-degree viscosity in HMV phenotypes of K. pneumoniae
is more associated with capsular serotype K1 or K2 than virulence factor
genes.
Introduction: Edwardsiella tarda causes edwardsiellosis in fish. In human,
E. tarda causes gastrointestinal and extraintestinal infections. Several viru-
lence factors have been identified to date including T3SS and T6SS. However, the exact infection mechanism, especially that in humans, cannot be
explained based only on known virulence factors. We were interested in
determining whether differences existed in the pathogenicities of human
isolates and diseased fish isolates in humans. Methods: The penetration
activities of E. tarda , including human isolates and diseased fish isolates,
through Caco-2 cell monolayers were evaluated. To determine whether
T3SS and T6SS were associated with penetration activity, we analyzed the
distribution of the T3SS and T6SS-related genes by colony PCR. To identify
genes responsible for penetration activity, we screened transposon (Tn)
insertion mutants for reduced penetration activity. Motility assays were
performed to determine the effects of transposon insertion on motility.
We also carried out colony PCR to detect the wecC gene to differentiate
the strains showing penetration activity. Results: All the human isolates
exhibited penetration activity in contrast to the fish isolates, which did not.
There was no amplification for any of the tested T3SS and T6SS-related
genes in the human isolates examined. Two Tn mutants showed reduced
penetration activity, and we identified the wecC and fliF genes as Tn insertion sites. Motility by the wecC mutant was weaker than that by the
wild-type strain, while the fliF mutant was immotile. The human isolates
showed amplification of the wecC gene, whereas the fish isolates did not.
Conclusion: Motility mediated by the wecC and fliF genes appeared to be
essential for penetration activity of E. tarda through Caco-2 cell monolayers. Furthermore, it was possible to group E. tarda strains into two types
of human isolates and diseased fish isolates based on distribution of the
wecC gene, T3SS and T6SS-related genes.
Symposium
PA-71
Epithelial cell injury might differ according to the bacterial species
Cell viability in P. aeruginosa was reduced stronger than that in E.
coli and K. pneumoniae
PA-72
Misato Gorai 1, Natsumi Takeuchi2, Kazue Fujita3, Yoko Mano2, Takahiro Suzuki1, Yoshibumi Akatsu1,
Akihiko Gemma3, Nobuhiko Furuya2
Identification and drug susceptibility of Leptotrichia spp.
strains isolated from oral specimens
Hitoshi Miyamoto , Shinobu Murakami, Mina Fukuoka, Yuri Tanaka, Takuya Kondo, Tatsuya Nishimiya
Ehime University Hospital, Japan
Case Conference
1 Hitachi,Lid.Hitachi
General Hospital, Japan
2 Bunkyo
Gakuin University
3 Nippon
Medical School
Oral Presentation
Poster Presentation
Purpose: Leptotrichia spp. which are one of oral microflora, have similar
morphology to Fusobacterium nucleatum, but can grow under microaerophilic condition. Since it can cause sepsis in patients with blood disorders,
it is very important to understand identification and drug susceptibility
of clinical isolates. This study was conducted to examine identification
and drug susceptibility of Leptotrichia spp. strains isolated from oral
specimens in Ehime University Hospital.Methods: Between August 2015
to February 2016, seventeen strains were isolated from clinical oral specimens (11 bronchoalveolar lavage fluids and 6 oral abscess). Isolates were
identified using MALDI Biotyper and BLAST analysis of 16SrRNA DNA sequencing. Antimicrobial susceptibility of PCG, ABPC, S/A, T/P, CTRX, CMZ,
MEPM, CLDM, and LVFX was checked using dry plates with Brucella broth
after 46-48 hour incubation under anaerobic condition. All results were
interpreted for susceptible and resistance including intermediate.Results:
Using MALDI Biotyper, one strains was identified as Leptotrichia wadei,
but other 16 strains were not accurately determined. BLAST analysis of
16SrRNA DNA sequence demonstrated that one L. wadei strain identified
by MALDI Biotyper had 99.46 % sequence similarity to L. wadei. Since
sequence of other 16 strains showed less similarity than 98.7% (97.9598.23%) to L. wadei, isolates could be new species. All strains showed
susceptibility to of PCG, ABPC, S/A, T/P, CTRX, CMZ, MEPM, and CLDM,
and resistance to LVFX.Conclusion: MALDI Biotyper and 16SrRNA DNA
sequencing could not identify 16 of 17 isolates from oral specimens. Since
there is possibility for new species of Leptotrichia, further investigations
are needed. And we should be aware of LVFX resistant Leptotrichia spp.
strains.
Background: Pneumonia is the leading cause of premature death in the
world. The gram-negative bacilli, Pseudomonas aeruginosa and Klebsiella
pneumoniae , are important causes of pneumonia in hospitalized and immunocompromised patients. The lower respiratory tract is also a frequent
locus of emerging infections, such as severe acute respiratory syndrome.
Impaired epithelial barrier function correlate with disease severity of
acute respiratory syndrome. However, little is known about the association
between the extent of epithelial cell injury and the bacterial species. We
hypothesized the extent of epithelial cell injury may differ according to
the bacterial species. Here, we show in vitro study about the difference between the extent of epithelial cell injury and various bacterial species. Materials and Methods: P. aeruginosa , K. pneumoniae and Escherichia coli ,
which represent common gram-negative pathogens that may contribute to
nosocomial infection were evaluate in this study. A549 human airway epithelial cell viability assay, measurement of bacterial endotoxin and proteolytic activity of bacteria were performed to determine epithelial cell injury.
Resutls: Cell viability in P. aeruginosa was reduced stronger than that in
K. pneumoniae and E. coli . Cell viability in stationary-phase P. aeruginosa
was reduced stronger than that in log-phase P. aeruginosa . Proteolytic
activity of P. aeruginosa increased more than that of K. pneumoniae and E.
coli . However, levels of endotoxin did not significantly differ among these
pathogens. Conclusions: This study suggests that epithelial cell injury
might differ according to the bacterial species. Impaired epithelial cells
caused by P. aeruginosa are stronger than that by K. pneumoniae and E.
coli . Furthermore, this difference is due to proteolytic activity of bacteria.
Such information can support physicians' quick and accurate decision for
diagnosis and treatment. There is concern that the subtle communication
between the medical technologist microbiology and the infectious diseases
physician would impact good patient care outcomes.
62
Putative ligands for Dectin-2, a C-type lectin receptor in
PA-74
Cryptococcus neoformans
Daiki Tanno 1, Kazutaka Ohashi1, Keiko Ishii2, Noboru Ohana1, Hiroki Shimura1, Sho Yamasaki3,
Kazuyoshi Kawakami2
YUKI HARA , MAKOTO KAWASHIMA, SACHIE ASAI, NAOKI YAMADA, MAMORU ITO
Japanese Red Cross Society Nagoya Daini Hospital , Japan
1 Department
of Clinical Laboratory, Fukushima Medical University, Japan
2 Department
of Medical Microbiology, Mycology and Immunology, Tohoku University Graduate School of Medicine
3 Division
of Molecular Immunology, Medical Institute of Bioregulation, Kyushu University
Invited Lecture
Special Lecture
Introductions:Recently, the spread of extended spectrum Β -lactamase
(ESBL) producing bacteria and carbapenem resistant Enterobacteriaceae
(CRE) in the community has become a problem. A new commercial automated antimicrobial susceptibility test assay, called DPS192ix®(EIKEN
CHEMICAL CO., LTD., TOKYO, JAPAN), has kinetic system which records
bacterial growth every hour. So, DPS192iX®was expected to contribute
to rapid detection in multi-drug resistant Enterobacteriaceae (MDRE). We
evaluated the performance of DPS192iX® for rapid detection of MDRE.
Methods: The strains used for analysis were 63 isolates of gram negative Enterobacteriaceae including 35 ESBL producing bacteria, 10 AmpC
producing bacteria, 7 CRE and 11 negative controls. Negative controls do
not produce ESBL, AmpC and carbapenemazes. The drugs used included
cefpodoxime (CPDX), ceftazidime, ceftriaxone, cefepime, cefmetazole,
aztreonam, imipenem (IPM) and meropenem (MEPM). We recorded
bacterial growth hourly and calculated the resistant rates to these drugs.
Results:ESBL producing bacteria, AmpC producing bacteria and CRE presented a high resistant rates to CPDX in 6 hours (ESBL: 97.1%, AmpC: 70%,
CRE: 100% ). Whereas, all negative control strains presented susceptible to
CPDX. In addition, CRE showed a high resistant rates to IMP and MEPM in
6 hours (IPM: 66.7%, MEPM: 33.3%). On the other hands, all other strains
except CRE were susceptible to IPM and MEPM.Conclusion: In conclusion, we have shown that DPS192iX® may contribute to rapid detection of
MDRE in 6 hours.
Educational Lecture
Introduction: Cryptococcus neoformans , a yeast-form fungal pathogen,
is rich in polysaccharides in its cell wall and capsule. Dectin-2 is a C-type
lectin receptor (CLR) that recognizes high mannose polysaccharides.
Dectin-2 functions as a pattern recognition receptor (PRR) and plays a
central role in immune response to fungal pathogens. Recently, we have
analyzed the role of Dectin-2 in the host defense against C. neoformans
infection. However the precise structure of this fungus recognized by
Dectin-2 has yet to be defined. Our aim is to address the Dectin-2 ligand
in C. neoformans .Methods: 2B4-NFAT-GFP reporter cells expressing Dectin-2 were stimulated with B3501 (C. neoformans wild-type strain), Cap67
(acapsular mutant strain of B3501) or the disrupted lysates of these fungi
for 24 h. The expression of GFP was analyzed in the reporter cells by flow
cytometry. C. neoformans -derived mannoprotein with a molecular weight
of 98 kDa (MP98) was kindly provided by Dr. S.M. Levitz (University of
Massachusetts Medical School, Worcester, MA, USA).Results: Whole C.
neoformans yeast cells (B3501 and Cap67) did not induce GFP expression
by the reporter cells, whereas heat-killed Candida albicans (HKCA) using
as a positive control showed this activity. The disrupted lysates of B3501
and Cap67 induced GFP expression as highly as HKCA. This activity was
detected in the precipitates, but not in the supernatants of these lysates.
MP98 caused a low, but significant level of GFP expression.Discussion:
These data suggest the presence of some putative Dectin-2 ligand in C.
neoformans , which may not be expressed on the surface, but rather inside. Capsule may not be required for this activity, because the acapsular
mutant strain was as active as the capsular strain. Mannoprotein such as
MP98 could be a candidate of Dectin-2 ligand in C. neoformans . Further
investigation is necessary to define this structure.TEL: +81-24-547-1111
(Ext: 3551)
Comparison of screening methods for carbapenemase-producing Enterobacteriaceae
Comparison of Modified-Hodge test, Carba NP test, and carbapenem inactivation
method
PA-76
False-negative rate in VersaTREK blood culture system
Hanae Nakashima , Nobuki Miura, Takahiro Shimizu
Nagano Municipal Hospital, Japan
Background: The global emergence of carbapenemase-producing Enterobacteriaceae (CPE) has become a major concern. On the other hand,
63
Poster Presentation
several laboratory procedures have been developed to detect CPE in a
clinical laboratory. In this study, we evaluated the performance of three
screening methods, the Modified-Hodge test (MHT), Carba NP test (CNPt),
and the carbapenem inactivation method (CIM) for CPE. Materials and
Methods: Seventeen non-carbapenemase producing isolates (Extendedspectrum Β -lactamase producers and AmpC producers) and 33 CPE (IMP,
15; NDM, 8; OXA-48 like, 7; KPC, 3) isolates from Japanese hospitals were
used in this study. The MICs of imipenem, meropenem, and ertapenem
were determined by using the Etest. MHT, CNPt, and CIM were compared
as screening methods for CPE. CIM was performed with three carbapenem
disks (imipenem, meropenem, and ertapenem).Results: MIC ranges of
imipenem, meropenem, and ertapenem for CPE were 0.5– ＞ 32 µg/mL,
0.25– ＞ 32 µg/mL, and 0.25– ＞ 32 µg/mL, respectively. The sensitivities
of MHT (ertapenem), CNPt, and CIM (meropenem) were 90.9%, 84.8%, and
97.0%, respectively, and their specificities were 82.3%, 100%, and 94.0%,
respectively. Although false-negative results were observed in OXA-48
like producers and/or mucoid NDM producer by using MHT and/or CNPt,
CIM showed positive results in these isolates. When the imipenem disk
was used in CIM, false-positive results were particularly observed in AmpC
over-producers.Conclusion: Although CIM requires time for cultivation, it
showed excellent specificity, sensitivity, and cost-effectiveness. Therefore,
CIM is useful in general clinical laboratories. Furthermore, our results indicate that in CIM, meropenem or ertapenem disks should be used instead of
imipenem disks. Since CNPt results were obtained more rapidly compared
to the other two methods, CNPt might be useful for the rapid prevention
and control of infection in a hospital.
Oral Presentation
Introduction: The VersaTREK (VT) (Thermo Scientific) has been introduced as an automated blood culture system in our hospital. From February to April 2015, we noticed that three false-negative blood culture
results were obtained using the VT system. Therefore, this study was
conducted to analyze the false-negative rate of the VT blood culture
system.Methods: In total, 3228 blood culture bottles (1624 aerobic and
1604 anaerobic bottles) submitted to our laboratory from 15 June to 14
December 2015 were analyzed using the VT instrument (VTI). At the end
of 7 days, negative bottles were removed from the VTI and subcultured.
Drigalski agar, sheep blood agar, chocolate agar, Sabouraud dextrose agar,
and Anaero Columbia agar (two plates) were used as the subculture plates.
Drigalski and Sabouraud dextrose agar were incubated aerobically at 35°
C. Sheep blood and chocolate agar were incubated in carbon dioxide at 37
°C. One Anaero Columbia agar plate was incubated in microaerobic conditions at 42°C, and the other was incubated in anaerobic conditions at 37
°C. All agar plates were incubated for 7 days.Results: Among the aerobic
bottles, 1461 were determined to be negative by the VTI, and no microbial
growth occurred by subculture. Among the anaerobic bottles, 1488 were
determined to be negative by the VTI; of these, 11 bottles (0.74%) exhibited growth on subculture. The following microorganisms were detected
by subculture: Campylobacter jejuni (three bottles), Parvimonas micra (two
bottles), Staphylococcus epidermidis (one bottle), Pseudomonas aeruginosa (one bottle), Escherichia coli (one bottle), Propionibacterium acnes
(one bottle), Helicobacter cinaedi (one bottle), Aspergillus niger (one bottle).Conclusion: The false-negative rate in the anaerobic bottles was 0.74%
in this study. This rate is higher than the rate of 0.2% described in the VT
instructions.
Case Conference
Kageto Yamada 1, Katsumi Arai1, Noriyuki Nagano2, Ryoichi Saito3
1 Tokyo
Metropolitan Hlth. and Med. Treatment Corp. Toshima Hosp., Japan
2 Shinshu
University
3 Tokyo
Medical and Dental University
Symposium
PA-75
Rapid detection of Multi-Drug Resistant Enterobacteriaceae
using a new antimicrobial susceptibility test assay
Keynote Speech
PA-73
Keynote Speech
PA-77
Clinical utility of two-step culture method for detection of group B streptococcus in
pregnant women
GBS detection rate with the combination of selective separation and enrichment media
PA-78
Evaluation of urine flow cytometer Sysmex UF-5000:the
comparison with routine methods for identifying bacteria
Masahito Ohnishi , Sang-Tae Lee, kouji Ui, Akira Koizumi, Kayoko Toimoto, Hiroshi Yabuuchi,
Shinobu Tanaka, Yayoi Umeki
Kae Kawamura 1, Yoshitsugu Iinuma2, Daisuke Usuda2, Masami Matsumoto1, Yoshi Tanaka1,
Masanori Kawano3, Shota Tateyama3, Yuji Itose3
Nara Medical University Hospital, Japan
1 Department
of Clinical Laboratory,Kanazawa Medical University Hospital, Japan
2 Department
of Infectious Diseases, Kanazawa Medical University,Japan
3 Sysmex
Corporation,Japan
Invited Lecture
Special Lecture
Educational Lecture
Introduction:Group B streptococcus (GBS) is a causative bacterium of
meningitis and sepsis in neonates, and GBS screening of all pregnant
women at 35 to 37 weeks of gestation is recommended to prevent neonatal GBS infection. The aim of the present study was to evaluate the performance of a two-step culture method using the combination of selective
separation and enrichment media for GBS screening of pregnant women.
Methods:In our hospital, the culture method for GBS screening of pregnant women was changed in April 2015. Previously, vaginal and anorectal
clinical specimens from each pregnant woman were plated directly on
Try/Soy Blood Agar (Sheep) No.2 (Kyokuto Pharmaceutical Industrial Co.,
Ltd., Tokyo, Japan). In contrast, the altered culture method employs a selective enrichment medium (Bouillon Todd Hewitt + Antibiotiques, Sysmex
bioM&eacute;rieux Co., Ltd., Tokyo, Japan) with subculture to a selective
separation medium (chromID Strepto B, Sysmex bioM&eacute;rieux Co.,
Ltd., Tokyo, Japan). We compared the GBS detection rate between the
direct culture method and the two-step culture method.Results:The GBS
detection rate was 8.3% (75/903) with the direct culture method and
17.7% (130/736) with the two-step culture method. Identification of GBS
was clear and easy with the two-step culture method that distinguishes
pale pink to red GBS colonies. Also, a non- Β -hemolytic strain of GBS
was detected with the two-step culture method.Conclusion:These results
demonstrate that the combination method using selective separation and
enrichment media was more sensitive than the direct culture method for
GBS screening of pregnant women,indicating its usefulness in the prevention of neonatal GBS infection.
Symposium
PA-79
Purpose:A novel flow cytometer-based analyzer, Sysmex UF-5000 (UF5000) can classify Gram positive bacteria (GP) and Gram negative bacteria
(GN) from urine sample as Gram Positive?, Gram Negative?, Gram Pos/
Neg?, or Unclassified together with other urine particles within a few
minutes. The aim of this study was to evaluate the performance of classification by comparing with gram staining and culture identification
results.Materials and methods:188 urinary tract infection-suspected
urine samples in Kanazawa Medical University Hospital were used for this
study. These samples met the criteria as follows: GP or GN was confirmed
in Gram staining with direct smear examination, and ≥10 white blood
cells/ μ L and ≥100 bacteria/ μ L in UF-5000 measurement. In additional
to Gram staining, the samples were also tested by conventional culture
identification. The results of UF-5000 were compared with two methods
as mentioned earlier.Result:In the 188 samples, 49, 103, and 36 were determined as GP, GN, and both Gram positive and negative (GPN) by Gram
stain, respectively. Concordance rate (CR) and positive predictive value
(PPV) of the results of UF-5000 to Gram stain results were as follows:
75.5% and 80.4% in GPB, 70.9% and 88.0% in GNB, and 55.6% and 40.8%
in GPN, respectively. Meanwhile, 48, 102, and 38 were determined as GP,
GN, and GPN by culture identification, respectively. CR and PPV to culture
results were as follows: 77.1% and 80.4% in GP, 72.5% and 89.2% in GN,
and 55.3% and 42.9% in GPN, respectively.Conclusion:UF-5000 can potentially provide rapid classification of bacteria compared with the Gram stain
and culture identification in urine samples. UF-5000 is capable of updating analytical algorithm for classification of bacteria, which may improve
its performance.
Bacteremia Caused by Capnocytophaga Species in Kyushu
University Hospital, 2005-2015
PA-80
A case of infectious endocarditis due to Bartonella quintana
Case Conference
Makiko Kiyosuke 1, Tomomi Mochimaru1, Yuiko Morokuma1, Ruriko Nishida1, Hisanori Nishio2,
Nobuyuki Shimono2, Taeko Hotta1, Dongchon Kang1
Noriyuki Abe , Yuki Ono, Eriko Hashimoto, Hiroko Matsutani, Akihiro Nakamura, Saori Fukuda,
Hisashi Kouno, Fumihiko Nakamura
1 Department
of Clinical Chemistry and Laboratory Medicine, Kyushu University Hospital, Fukuoka, Japan
2 Center
for the Study of Global Infection, Kyushu University Hospital
Department of Clinical Laboratory, Tenri Hospital, Japan
We report a rare case of infectious endocarditis caused by Bartonella
quintana , in which patient's serum antibody titers led to a suspicion of B.
henselae infection but the correct diagnosis was obtained using genetic
Oral Presentation
Poster Presentation
Background The genus Capnocytophaga is a group of facultative, gram
negative rod-shaped anaerobe. They constitute a fraction of normal oral
flora in human and animals. Capnocytophaga canimorsus is known to
cause serious zoonotic infections in these years. Bacteremia caused by
Capnocytophaga species is found in many immunocompromised patients.
Materials and Methods Capnocytophaga species were detected in blood
cultures of seven cases from 2005 to 2015. We analysed the data of these
cases: underlying diseases, antimicrobials used before and after bacteremia, beta lactamase producing or not , results of microbial sensitivity testing, and prognosis, etc. Results We studied Capnocytophaga bacteremia
cases in Kyushu University Hospital, retrospectively. These patients had
been hospitalized in three clinical departments: four in pediatrics , one in
surgery, and two in hematology. All cases suffered bacteremia of Capnocytophaga species during chemotherapy for underlying diseases. The antimicrobials used before bacteremia were meropenem, cefazolin, cefpirom
and cefepime. Piperacillin/tazobactam, ciprofloxacin, doripenem and
meropenem were used for the treatment after diagnosis of bacteremia.
Capnocytophaga sputigena and Capnocytophaga gingivalis were isolated
from five and two patients, respectively. All the isolated strains produced
beta lactamase.Conclusions Oral mucosa is a frequent infection portal
in patients with Capnocytophaga species bacteremia. Capnocytophaga
bacteremia is common in immunocompromised hosts as the same in our
seven cases. One patient with febrile neutropenia (FN) was treated with
beta-lactams before bacteremia. Recently, there was a report on bacteremia caused by multidrug-resistant C. sputigena with ESBL-related cfxA3
gene. Fourth generation cephems which are used as first-line drugs for FN,
had no effect for that case. It is important to isolate and identify Capnocytophaga species as quick as possible for the appropriate treatment.
analysis. A 70-year-old male was hospitalized with frequent episodes of fever that had lasted for a year. Transthoracic and transesophageal echocardiography demonstrated vegetation in aortic valve, but it did not exhibit
marked oscillation. Six sets of blood cultures were all negative. Therefore,
blood culture-negative endocarditis caused by a low virulence bacillus was
most suspected and comprehensively examined the serum antibody titers.
As a result, we detected B. henselae IgG and IgM antibody titers of ＞ 1024
and 20, respectively. In contrast, B. quintana IgG antibody titer was 256,
and tests for IgM were negative. After treatment with ceftriaxone, gentamicin, and minocycline, the oscillation of cardiac vegetation increased,
then aortic valve replacement was performed.Bacterial cultures were obtained from aortic valve and associated pus around the valve, but after 4
weeks the cultures for Bartonella sp. were negative.Therefore, nucleotide
sequence analysis of the 16S rRNA and rpoB genes were performed after
extracting DNA from the aortic valve and pus. A BLAST search revealed
that the extracted DNA exhibited 99% homology with B. quintana and
94% homology with B. henselae .We suspected that cross-reactivity had occurred during the serum antibody titer tests and that the high B. henselae
antibody titer was a false-negative.[Conclusion] Although Bartonella infections are rare, they should be considered in blood culture-negative cases of
endocarditis. In cases of Bartonella infection-induced endocarditis in which
it is suspected that cross-reactivity might have occurred during serum antibody titer examinations, genetic analysis is important for discriminating
between Bartonella sp.
64
A case of pulmonary Nocardia cyriacigeorgica infection in a
patient with Mycobacterium intracellulare lung disease
PA-82
A case of Yersinia pseudotuberculosis infection
1 National
Hospital Organization Nagoya Medical Center, Japan
2 National
Hospital Organization Kanazawa Medical Center Dept. Medical Laboratory
3 National
Hospital Organization Kanazawa Medical Center Dept. Pediatrics
Introduction: Pulmonary nocardiosis is commonly recognized as an opportunistic infection in immunocompromised hosts. However, nocardiosis
can also affect immunocompetent hosts. In immunocompetent hosts,
pulmonary nocardiosis often co-occurs with an underlying disease such
as bronchiectasis or chronic obstructive pulmonary disease, or with treatment with steroids. In addition, a few recent reports have described pulmonary nocardiosis with nontuberculosis mycobacterial lung infection. Here
we report a case of pulmonary Nocardia cyriacigeorgica infection in a
patient with Mycobacterium intracellulare lung disease.Case presentation:
An immunocompetent 79-year-old Japanese woman with untreated M.
intracellulare lung disease for the past several years was admitted to our
hospital because of high fever and cough. Her sputum amount increased
and chest X-ray imaging and computed tomography revealed infiltrates in
the lower lobe of the right lung. Her white blood cell count increased to 20
× 103/mm3 with neutrophilic predominance (93.6%). Sputum Gram stain
showed gram-positive branching rods suspected as a Nocardia species,
and acid fast stain showed bacilli suspected as a Mycobacterium species.
Both a Nocardia species and M. intracellulare were isolated from her sputum. Antimicrobial treatment was initiated with piperacillin/tazobactam
and was changed to azithromycin on the 8th day of hospitalization, then
to doripenem on the 15th day. Her respiratory status improved, and antibiotics treatment was stopped on the 22nd day. The Nocardia species
was identified as N. cyriacigeorgica by 16S ribosomal RNA gene sequencing. Susceptibility testing of N. cyriacigeorgica was performed according
to Clinical and Laboratory Standards Institute (CLSI) document M24-A,
and it was regarded as sensitive to trimethoprim/sulfamethoxazole and
imipenem. Conclusion: Nocardia species may cause co-infection with
M.intracellulare , and we must be careful not to overlook this possibility.
Therefore, performing sputum Gram stain is important. Furthermore,
correct identification of the species is important for the treatment of pulmonary nocardiosis because antimicrobial susceptibility patterns differ
among Nocardia species.
((Introduction)) Yersinia pseudotuberculosis (YPT)infection usually is
a form of gastroenteritis, though various features including Kawasaki
disease-like symptoms and acute renal failure are known to manifest. Associations of exotoxin YPM (YPT-derived mitogen) with such symptoms
and super-antigen activity have recently attracted attention.((Case)) A
15-year-old boy presenting with fever in the morning was diagnosed with
upper respiratory tract inflammation at a nearby clinic, and CFDN was
prescribed. However, at night he was brought to our hospital emergency
outpatient department because of a higher fever (40 °C), chills, shuddering and vomiting, necessitating hospitalization. Follow-up with FOM
intravenous drip was based on a diagnosis of infectious gastroenteritis,
but sudden acute renal failure developed. The acute renal failure led to
septic shock and the antimicrobial was changed to MEPM. Furthermore,
the gamma globulin dosage, PMX endotoxin adsorption therapy, and continuous hemodiafiltration (CHDF) were applied more aggressively. Having
initially responded to treatment well, he was about to be discharged. However, the infectious gastroenteritis symptoms recurred and he was immediately re-hospitalized. In addition, the first fecal culture on admission was
negative, though V Β 3 was specifically increased based on T cell antigen
receptor repertoire analysis. Furthermore, his sequential symptoms raised
strong doubt about YPT infection.The bacteria were identified as YPT in
fecal culture at the time of reinfection in API20E with CIN nutrient medium (room temperature culture). A characteristic change in the anti-YPT
antibody titer and the anti-YPM antibody titer were confirmed, that is ,the
former increased faster than the latter. ((Conclusion)) A rare YPT infection
case was experienced and is reported herein. No other family members
developed symptoms and the source of infection could not be specified.
Primary lymphocutaneous nocardiosis caused by minocycline
non-susceptible Nocardia brasiliensis
PA-84
A case of Neisseria meningitidis infection, isolated from
blood cultures
Nana Nishida 1, Yasunao Wada1, Kumiko Yamada1, Atsuko Yamazaki1, Noriko Kitamura1, Mika Mori1,
Mitsuru Masaki2, Masahiro Koshiba2
1 Department
of Laboratory Medicine,Tohoku University Hospital , Japan
2 Department
of Infection Control and Laboratory Diagnostics, Internal Medicine, Tohoku University Graduate School of
Medicine
1 Hyogo
College Of Medicine Hospital, Japan
2 Hyogo
College Of Medicine
Poster Presentation
65
Oral Presentation
Introduction:N. meningitidis is a Gram-negative diplococcus, causes the invasive meningococcal diseases. When entered into the bloodstream, it can
spread to cerebrospinal fluid (meningococcemia) and induce meningitis.
Without the proper treatment it may progress rapidly, resulting in death
within a few hours. Reported here is a case of serious N. meningitidis infection that was isolated from blood cultures.Patient:A 48-year-old male
presented to another hospital with complaints of fever and respiratory discomfort. The patient developed to shock status with upper airway obstruction during the admission, thereby transported to our hospital. Findings on
admission: body temperature 38.9°C, WBC 14,200 / Μ L, CRP 7.55 mg/
dL. With the suspicion of the septic shock, two sets of blood cultures were
submitted.Bacterial isolation:Gram-negative diplococci were detected in
blood cultures in 48 hours, and phagocytosis by neutrophils was observed
on the staining. On the next day, the bacterium was identified as N. meningitidis by Microscan WalkAway (Beckman Coulter, Brea, CA). Susceptibility
test was carried out using MIC2000 Frozen Plate (Eiken Chemical, Tokyo,
Japan).Clinical Course:He had been administered meropenem for 3 days
from admission, and then antibiotic was changed to ampicillin according
to the susceptibility test result. He had been administered it for 2 weeks,
and his inflammation was improving. He is currently under the follow-up
observations.Conclusion:Immediate diagnosis and early treatment is important for the bacterial infections, especially for the meningococcal infection. In the case presented, we had received the information of the suspicious meningococcal infection before the culture started, which resulted in
the quick identification of the bacterial strain and led to the appropriated
selection of the anti-biotic reagent.
Nocardiosis is a rare infection and is clinically divided into cutaneous and
systemic type; the former presents either as part of a disseminated infection or as a primary infection resulting from the invasion of a pathogen
through an injury. It is well known that Nocardia brasiliensis is the most
predominant causative pathogen in primary cutaneous nocardiosis. It is
also known that some Nocardia spp. has typical antimicrobial susceptibility patterns, therefore CLSI (M24-A2) recommend to use susceptibility
patterns as a tool for predictive identification because of difficulty of
correct identification of Nocardia spp. (ex. N. brasiliensis : imipenem(R),
ciprofloxacin(R), minocyclin(S), sulfonamides(S), tobramycin(S),
clarithromycin(R), N. nova complex : imipenem (S), ciprofloxacin (R), minocyclin (variable), sulfonamides (S), clarithromycin (S), N. farcinica : imipenem (variable), ciprofloxacin (S), minocyclin (variable), sulfonamides (S), tobramycin (R), clarithromycin (R), N. abscessus : imipenem (R), ciprofloxacin
(R), tobramycin (variable), sulfonamides (S), clarithromycin (R)).Here, we
report a case of primary lymphocutaneous nocardiosis due to atypical (minocycline non-susceptible) N. brasiliensis in an immunocompromised patient who was successfully treated with oral trimethoprim-sulfametoxazole
(8 days) and minocycline (6 month). In addition, we also report antimicrobial susceptibility patterns analyzed 10 clinical N. brasiliensis isolates and
discuss current status of susceptibility patterns.This work was supported
by grants from Charitable Trust Laboratory Medicine Reserch Foundation
of Japan and Kurozumi Medical Foundation.
Case Conference
Mikiko Chiba 1, Masahiro Toyokawa1, Makoto Katsumi1, Takami Sato1, Junko Shoji1, Chiyuki Ishizawa1,
Mitsuaki Nagasawa1, Mitsuo Kaku2
Symposium
PA-83
Educational Lecture
1 Ina
Central Hospital, Japan
2 Shinshu
University Hospital
3 Graduate
School of Medicine, Shinshu University
Special Lecture
Toshiyuki Asaka 1, Shirou Nishio2, Mizue Mizuno2, Susumu Haneda2, Satomi Kasashima2,
Atsuhiro Kawashima2, Kazuhide Ohta3
Invited Lecture
Keisuke Soya 1, Takehisa Matsumoto2, Hiromasa Hukatsu3, Hiroyuki Yano1, Kiyoshi Kobayashi1,
Keiko Hirose1
Keynote Speech
PA-81
Keynote Speech
PA-85
A case of granulomatous mastitis caused by Corynebacterium
tuberculostearicum infection in a nulliparous young woman
PA-86
Daisuke Kitagawa 1, Miyako Oka1, Kazue Masuo1, Yutaka Yoshimura1, Akihiro Nakamura2
Yumiko Joichi 1, Shizuo Kayama2, Ikue Hayashi2, Makoto Onodera3, Maki Furushimo3,
Michiya Yokozaki4, Hiroki Ohge5, Motoyuki Sugai2
1 Nara
Prefecture General Medical Center, Japan
2 Tenri
Yorozu Hospital
1 Section
of Infection Diseases, Laboratory Division of Clinical Support, Hiroshima University Hospital, Japan
2 Project
Research Center for Nosocomial Infectious Disease, Hiroshima Univ.
3 Section
of Infection Diseases, Laboratory Div. of Clinical Support, Hiroshima Univ. Hosp.
4 Div.
of Laboratory Medicine, Hiroshima Univ., Hosp.
5 Dept.
of Infectious Diseases, Hiroshima Univ., Hosp.
Invited Lecture
[Introduction]Recently, infection with Lipophilic Corynebacterium such as
C. kroppenstedtii and C. tuberculostearicum has attracted attention. We
have experienced a case that was isolated C. tuberculostearicum which is
also lipophilic Corynebacterium from a case of mastitis.[Case]30s, woman.
Clostridium tertium is an endospore-forming anaerobic Gram-positive ba-
Special Lecture
Educational Lecture
The right mammary gland of patient had continued swelling and pain
from one month ago. After various inspections, breast cancer is negative.
Administration of antibiotics was started. However, admitted deterioration
and fever. In addition the patient was hospitalized. After the mammary
gland puncture, it was detected lipophilic Corynebacterium from aspirated
pus by bacteriological examination. The symptoms of a patient were improved by the drainage and antibiotics of levofloxacin.[Microbiological examination]Gram stain of the aspirated pus admitted many neutrophils and
a few Gram-positive bacilli, they grew slightly in 8 days after culture. Next
we used ApiCoryne identification kit (Bionumber: 2100105) and VITEK2
ANC (Bionumber: 6363100400405) could not identify bacterial species.
Their growth was encouraged using the lipidsupplemented medium.PCR
targeting C. kroppenstedtii 16S rRNA gene resulted in negative. At last, C.
tuberculostearicum was identified by mass spectrometry and 16S rRNA
broad-range PCR analysis.[Discussion]C. tuberculostearicum is likely to be
a pathogenic bacterium in this case. Because C. tuberculostearicum is not
included in the database of ApiCoryne and VITEK2, these can not identify
the speies accurately. In addition, it was reported that ApiCoryne (bionumber: 2040104) identified C. kroppenstedtii incorrectly as C. argentoratense . We consider that PCR and mass spectrometry are necesary in order
to distinguish the two species accurately. Mastitis caused by Lipophilic
Corynebacterium is frequently refractory to Β -lactam antibacterial agents
by its water-soluble nature. Therefore, reporting as Lipophilic Corynebacterium is a high clinical significance, although the species is not identified.
Symposium
PA-87
Two cases of Clostridium tertium infection identified by
MALDI-TOF mass and 16S rRNA sequencing analysis
cillus capable of growing in aerobic conditions that is often misidentified
as a Gram-negative organism. We report here two cases of C. tertium isolated from an aerobic blood culture. In both cases, aerobic blood cultures
became positive and Gram-negative (or variable) bacilli were recovered.
The organism grew in aerobic and anaerobic culture of blood, chocolate,
and Brucella HK agar plates showing similar colony morphology. In Case 1,
the organism was identified as C. tertium , but in Case 2 the organism was
reported as C. clostridioforme using VITEK2 compact and Rap ID ANAII
analyses. But we doubted this report due to several reasons: the organism
in case2 grew in aerobic condition but C. clostridioforme is highly anaerobic; the morphology of the organism was not like “foot-ball” form which
is typical morphology of C. clostridioforme ; the organisms possessed
endospore while C. clostridioforme does not form spore. When we further
performed an identification process using matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis
and subsequent 16S ribosomal DNA sequencing in case 2, the test strain
was identified as C. tertium . Bacterial identification based on MALDI-TOF
mass introduced a reliable alternative to provide rapid identification without the need for expert laboratory staff.
A case of successful isolation of Clostridium tetani from
drain fluid
PA-88
Mycobacterium abscessus in blood culture of a patient with a
long-term indwelling CVC (case report)
Case Conference
Kyohei Kato 1, Taeko Narita1, Kazuhiro Shinto1, Tetsuhiro Harada1, Yumiko Funashima2, Kenichi Sato2,
Zenzo Nagasawa2, Tsukuru Umemura2
Maki Furushimo1, Yumiko Koba1, Makoto Onodera1, Ikue Hayashi2, Shizuo Kayama2, Hiroki Ohge3,
Michiya Yokozaki4, Motoyuki Sugai2
1 Department
of Clinical Labotatoriy,Medical Kouhoukai Takagi Hospital, Japan
2 Department
of Medical Technology and Science, Fukuoka Health Care Faculty, International University of Health and Welfare
1 Section
of Infection Diseases, Laboratory Division of Clinical Support, Hiroshima University Hospital, Japan
2 Project
Research Center for Nosocomial Infectious Diseases, Hiroshima Univ.
3 Dept.
of Infectious Diseases, Hiroshima Univ., Hosp.
4 Div.
of Laboratory Medicine, Hiroshima Univ., Hosp.
Oral Presentation
Poster Presentation
Introduction: Clostridium tetani , an obligate anaerobic gram-positive
bacillus, is distributed in soil and produces neurotoxins including tetanospasmin. The tetanospasmin inhibits acetylcoline release at synapse and
leads to serious neurological symptoms like systemic muscle rigidity.
Although isolation of the microorganism is very rare in a clinical setting,
we report that we succeeded in shorten the time to prove the isolation of
Clostridium tetani from drain fluid with MALDI-TOF MS.Case presentation: An 83-year-old woman who had a history of abdominal distention
and melena, was emergently referred to an academic medical center for
the treatment of deteriorated symptoms. Computed tomography showed
intestinal gas and intestinal fluid retention. She was diagnosed with ileus
and emergent operation was performed. Postoperative drain fluid was examined for bacterial culture.Microbiological examination: The fluid was
anaerobically cultured onto "KBM" Anaerobic Rabbit Blood Agar medium.
A tiny amount of film-like colony appeared on 2nd day of culture. MALDITOF MS identified Clostridium tetani as the causative microorganism.
Besides, gram staining and spore staining of colonies revealed distinctive
drumstick appearance. Finally, Clostridium tetani was confirmed by 16S
rDNA gene analysis and toxin-producing gene was detected by PCR.Conclusion: This is a very rare case that Clostridium tetani , a highly anaerobic
microorganism, was isolated from drain fluid. Though this microorganism
requires time to grow in culture and is difficult to be identified, MALDITOF MS was helpful for rapid detection of Clostridium tetani in this case.
Case: A 23s man had history of primary pulmonary hypertension. He had
been placed a Hickman catheter for two years. He had episodes of catheter
insertion site infection and received several antimicrobials. He developed
fever with physical weariness, and was hospitalized. The organism wasn't
detected from the culture of catheter insertion site. On day 2 after hospitalization, the catheter was removed, and three sets of blood culture and
skin surface culture of insertion site were conducted. The skin surface
culture was negative but on day 4, three aerobic bottles of the blood culture became positive. Gram-staining failed to detect any microorganisms
but sub-culture on blood and chocolate agar plates yielded shiny white
colonies after two days. Gram-staining of the colonies showed Gram variable bacilli. We therefore performed acid-fast staining, and detected bacilli
with red suggesting acid fast bacteria. The organism was subsequently
identified as Mycobacterium abscessus by MALDI-TOF MS-Biotyper analysis, and sequencing analysis of 16S rRNA and hsp65 gene. M. abscessus
is a rapidly growing mycobacterium (RGM) existing in soil and water and
causes skin and lung infectious diseases in immunocompromised patients.
When blood culture becomes positive in a patient with hematological disease, skin or bone infection, or a long-term indwelling catheter, it is important to consider a possibility of acid fast bacterial infection and perform
acid fast staining and culture.In this case, MALDI-TOF MS was useful for
the identification of an acid fast bacteria isolated from blood culture, and
the doctor was able to choose an appropriate antimicrobial agent without
delay according to the bacterial information we submitted to the clinical
side.
66
A case of chromoblastomycosis we could identify based on
results of fungal cultute
PA-90
Tomohiro Ueno 1, Hideki Niimi1, Noriko Yoneda2, Satoshi Yoneda2, Masashi Mori3, Shigeru Saito2,
Isao Kitajima1
Chikako Ogushi , Yasuko Senda, Yukiko Takemori, Yoshio Sakai, Takashi Wada
Kanazawa University Hospital, Japan
1 Clinical
Laboratory Center, Toyama University Hospital, Japan
2 Department
of Obstetrics & Gynecology, Toyama University Hospital
3 Research
Institute for Bioresources and Biotechnology, Ishikawa Prefectural University
Evaluating the Usability of a New Rapid Immunoassay Test
Kit for Detecting Urinary Tract Infections
Mitsuru Matsumura 1, Yusuke Nomura1, Shinobu Ishigaki2, Yoshiko Atsukawa2, Kazunori Komatsu2,
Taiji Furukawa2
1 Clinical
Laboratory Center, Toyama University Hospital, Japan
2 Research
Institute for Bioresources and Biotechnology, Ishikawa Prefectural University
3 Department
of Clinical Infectious Diseases, Toyama University Hospital
4 Department
of Obstetrics & Gynecology, Toyama University Hospital
1 Department
of Clinical Laboratory Science,Teikyo University Faculty of Medical Technology, Japan
2 Department
of Central Laboratory Medicine, Teikyo University Hospital,
67
Poster Presentation
Introduction: Acquiring the earliest possible identification of pathogenic
microorganisms is critical for selecting the appropriate antimicrobial
therapy in infected patients. We herein report the novel “melting temperature (Tm) mapping method” for rapidly identifying the dominant
bacteria in a clinical sample from sterile sites.Methods: First, bacterial
DNA is extracted directly from a clinical sample. Then, nested PCR is
performed using the seven bacterial universal primer sets. Second, seven
Tm values are obtained by melting analysis of the amplicons. These seven
Tm values, when mapped on two dimensions, create a unique shape of
a specific bacteria. Finally, by comparing this Tm mapping shape to the
shapes in the database, the dominant bacteria in a patient sample can be
rapidly identified. Results: We tested the Tm mapping method using 200
whole blood samples obtained from patients with suspected sepsis, 85%
(171/200) of which matched the culture results based on the detection
level. A total of 130 samples were negative according to the Tm mapping
method, 98% (128/130) of which were also negative based on the culture
method. Meanwhile, 70 samples were positive according to the Tm mapping method, and of the 59 suitable for identification, 100% (59/59) exhibited a “match” or “broad match” with the culture or sequencing results.
These findings were obtained within three hours of whole blood collection.
Conclusion: The Tm mapping method enables the identification of the
dominant bacteria in a clinical sample within three hours of whole blood
collection. Moreover, this method can be used to rapidly diagnose the absence of bacteria in clinical samples. The Tm mapping method is especially
useful for detecting infectious diseases, such as sepsis, that require prompt
treatment, and is expected to contribute to the treatment of patients with
severe infections as well as reduce the rate of development of antibiotic
resistance.
Oral Presentation
Introduction:There are no rapid test kits that can detect urinary tract
infections (UTIs) for sale in Japan. A rapid immunoassay test for detecting
UTIs called RapidBac was developed by Silver Lake Research Corporation,
USA. We tested whether the RapidBac is effective to detect UTIs. Methods:
We tested what kind of bacteria is detectable with RapidBac using 23
ATCC standard strains and clinical isolated strains (Gram Negative Bacteria
n=14, Gram Positive Bacteria n=8, Yeast n=1). To determine the minimum
sensitivity, we used Escherichia coli (ATCC 35218) and tested RapidBac
with 5 different concentrations (105, 104, 103, 102, 10 cfu/mL) to see
which concentration would become positive. Also we tested RapidBac using 30 urine specimens submitted to Teikyo University Hospital for urine
culture and compared both results. Results: Yeast and Gram Positive bacteria were all negative. Among the 14 Gram Negative Bacteria, 9 strains
were positive and 5 strains were negative. The negative strains include
Morganella morganii , Proteus stuartii , Pseudomonas aeruginosa , Pseudomonas fluorescens , and Serratia liquefacians . The RapidBac kit on samples with ＞ 103 cfu/mL showed a pale test line, indicating a positive result
but weak. Samples with ＞ 104 cfu/mL the test result was positive and the
test line was clearly to be seen. The Sensitivity and Specificity were 76.5%
and 84.6% respectively. Among the 30 specimens, 2 specimens showed
a false negative (15.4%), both including Pseudomonas aeruginosa . The
false positive rate was 23.5%. Conclusion: The RapidBac test accurately
detected Gram Negative Bacteria in this study but some strains were undetectable, mostly strains from Pseudomonas species. We thought the low
detective rate of Pseudomonas species; the main pathogenic bacterium for
complicated UTIs is a problem and would need further investigation. We
thought RapidBac would be efficient for hospitals without bacteriological
tests or as a point of care testing kit.
Case Conference
Kazushige Sugie 1, Hideki Niimi1, Tomohiro Ueno1, Masashi Mori2, Yoshihiro Yamamoto3, Shigeru Saito4,
Isao Kitajima1
Symposium
PA-92
Educational Lecture
Melting Temperature (Tm) Mapping Method
A Novel Method for Rapid Identification of Unknown Pathogenic
Microorganisms within Three Hours of Sample Collection
Special Lecture
Introduction: Intra-amniotic infection has long been recognized as the
leading cause of preterm delivery. Microbial culture is the gold standard
for the detection of intra-amniotic infection, but several days are required,
and many bacterial species in the amniotic fluid are difficult to cultivate.
Methods: We developed a novel nested-PCR-based assay for detecting Mycoplasma, Ureaplasma, other bacteria and fungi in amniotic fluid samples
within three hours of sample collection. To detect prokaryotes, eukaryotemade thermostable DNA polymerase, which is free from bacterial DNA
contamination, is used in combination with bacterial universal primers. In
contrast, to detect eukaryotes, conventional bacterially-made thermostable
DNA polymerase is used in combination with fungal universal primers. To
assess the validity of the PCR assay, we compared the PCR and conventional culture results using 300 amniotic fluid samples.Results: Based on the
detection level (positive and negative), 93.3% (280/300) of Mycoplasma,
94.3% (283/300) of Ureaplasma, 89.3% (268/300) of other bacteria and
99.7% (299/300) of fungi matched the culture results. Meanwhile, concerning the detection of bacteria other than Mycoplasma and Ureaplasma,
228 samples were negative according to the PCR method, 98.2% (224/228)
of which were also negative based on the culture method. Employing the
devised primer sets, mixed amniotic fluid infections of Mycoplasma, Ureaplasma and/or other bacteria could be clearly distinguished. In addition,
we also attempted to compare the relative abundance in 28 amniotic fluid
samples with mixed infection, and judged dominance by comparing the
Ct values of quantitative real-time PCR.Conclusion: We developed a novel
PCR assay for the rapid detection of Mycoplasma, Ureaplasma, other bacteria and fungi in amniotic fluid samples. This assay can also be applied
to accurately diagnose the absence of bacteria in samples. We believe that
this assay will positively contribute to the treatment of intra-amniotic infection and the prevention of preterm delivery.
Invited Lecture
We report a case of chromoblastomycosis that developed on the back of
the right hand. The patient was a 77-year-old man and a farmer. He had
noticed a small growing hyperkeratotic nodule on the back of the right
hand 1 year previously. He has not been under an immunocompromised
condition. He visited another hospital and culture examination and biopsy
were performed, but diagnosis was difficult. The patient was referred to
Kanazawa University Hospital 2 months later. The nodule had grown to a
crusted erythematous plaque 35 × 50mm in size. Histopathological examination of the biopsied skin showed infiltration of leukocytes in the dermis
and lymphocytes, neutrophils, plasma cells, eosinophils, and multinucleated giant cells in the subepithelium. Further, there were sclerotic cells with
a blackish brown thick cell wall among the multinucleated giant cells and
chain fungal spores. Taken together, a diagnosis of chromoblastomycosis
was made. The isolated fungus produced black colonies on Sabouraud
dextrose agar at 30°C. The colony was 15mm in size, blackish brown. And
ash color aerial hyphae grew thickly on the surface. Slide culture showed
flask-shaped phialide and cup-shaped collarette. We identified the fungus
as Phialophora verrucosa based on these results of fungal culture. Later,
it was identified as the same fungus by gene analysis at Chiba University
Medical Mycology Research Center. The base sequence of the ITS region
was determined and analyzed using the BLAST research (http://blast.ncbi.
nlm.nih.gov/). It showed more than 98% homology with sequence data of
20 strains identified as P. verrucosa . Therefore, this strain was identified as
P. verrucosa on phylogeny. The patient was treated with oral itraconazole
(100 mg/day, for 5 months) and the lesion has healed. He is now an outpatient without recurrence 6 months after treatment.
+81-76-265-2000 (Ext.7156) Kanazawa University Hospital
PA-91
Eukaryote-Made Taq Polymerase Enables Reliable Detection of Pathogens in Amniotic Fluid of Preterm Labor Cases
Development of a Novel Nested-PCR-Based Assay for Detecting Mycoplasma, Ureaplasma, other Bacteria and Fungi
in Amniotic Fluid Samples
Keynote Speech
PA-89
Keynote Speech
PA-93
Identification and antimicrobial susceptibiities in non-beta
hemolytic streptococci causing bloodstream infections
PA-94
Yatin Mange1, Malina K Storer1, Kim Hibbard-Melles2, Brett M. Davis1, Jenny M. Scotter1
Sang-Tae Lee , Koji Ui, Sachiko Kiyomatsu, Akira Koizumi, Masahito Onishi, Kayoko Toimoto,
Hiroshi Yabuuchi, Yayoi Umeki
1 Syft
Technologies, New Zealand
2 Christchurch
Polytechnic Institute of Technology, Christchurch, New Zealand
Nara Medical University hospital, Japan
Invited Lecture
Special Lecture
Educational Lecture
Urinary tract infections (UTIs) are the most common kidney and urological
diseases in the United States and are among the most common bacterial
infections of any organ system. Determining the causative agent of a UTI
requires the isolation and quantitation of the pathogen. Despite the introduction of the more rapid molecular tests, microscopy and culture remain
the gold standard in everyday clinical practice. However these methods
may take 24 hours (or longer if additional information such as antibiotic
sensitivities is required). A rapid, automated monitoring system for urine
specimens, with results available within a few hours, which provides clinical information on the patient's bacteriuria, would speed up identification
of the infecting agent and could allow appropriate antibiotic therapy to
begin more quickly.
Volatile metabolites produced by infecting organisms may offer a faster
path to diagnosis of infection and the causative organism if different organisms produce unique profiles of metabolites. To be effective, a rapid,
selective, and very sensitive analyzer is required. In this study, we have applied Selected Ion Flow Tube Mass Spectrometry (SIFT-MS), which fulfills
these requirements and is an industry-proven analytical technique that
instantly and directly analyzes air to part-per-trillion-by-volume (pptv) concentrations.
Eight microbial species were investigated here (10 replicate samples of
each). The samples were prepared using thawed sterile urine (20 mL) from
healthy males inoculated to a concentration of between 10e+7 and 10e+9
cfu and incubated at 37 C for 6 h. The headspace was then analyzed using
SIFT-MS. When combined with discriminant analysis this technique may
be applicable to microbe identification, though from this preliminary study
it appears that additional VOCs may improve the discriminatory ability of
the assay. Improved discrimination coupled with shorter incubation time,
rapid analysis, and the newly available autosampler-SIFT-MS systems,
could yield a powerful early screening tool.
Identification and antimicrobial susceptibilities in non-beta hemolytic
streptococci causing bloodstream infectionsBackground/introduction:
Non-beta hemolytic streptococci (mostly viridans streptococci) are normal
flora in oral cavities (normal oral commensals). Although they have been
considered to be of low virulence, they can cause bacteremia and sometimes infective endocarditis in certain individuals with underlying diseases
such as cardiac valve disease. Identification of non-beta hemolytic streptococci has been challenging in routine clinical microbiology testing since
conventional biochemical methods do not accurately discriminate and
identify these species. In this study, we compared the identification results
of conventional biochemical tests with those of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and
evaluated the antimicrobial susceptibilities which is important in choosing antibiotics in severe infections. Materials and methods: The 48 blood
culture isolates analyzed in this study were detected as suspected non-beta
hemolytic streptococci (excluding Streptococcus pneumoniae ) by conventional methods (VITEK2 and Rapid ID32 Strep) in Nara Medical University
hospital between 2009 and 2015. Identification was performed with both
VITEK MS and MALDI Biotyper. Susceptibility tests were performed by
Dry-plate Eiken DP34 for 47 isolates identified as viridans streptococci
with MALDI-TOF. Results: The three methods showed full agreement for
30 isolates (55.6%). The agreement rate between VITEK MS and MALDI
Biotyper was 94.4%. Based on the identification with MALDI-TOF, Streptoccocus mitis / oralis were the most common species (37.0%) followed
by Streptococcus anginosus and Streptococcus constellatus (11.1%), and
Streptococcus parasanguinis and Streptococcus salivarius (7.4%). Susceptibilities to penicillin G, ceftriaxone, levofloxacin, vancomycin, imipenem
and meropenem were 98.1, 100, 78.8, 100, 100, and 100%, respectively.
Conclusion: Identification of non-beta hemolytic streptococci causing
bloodstream infections by MALDI-TOF is rapid, cost effective and accurate
and may be a better alternative to conventional methods. Resistance to
levofloxacin should be taken into account in selecting antibiotics.
Symposium
PA-95
Volatile Metabolites as Early Diagnostics for Urinary Tract Infections?
Detection of volatile compounds produced by microbial growth in urine by
selected ion flow tube mass spectrometry (SIFT-MS)
Evaluation of Laboratory Testing Strategies and Prevalence of HospitalAssociated Clostridium Difficile Infection
Two-Step Algorithm for Detection of Toxigenic Clostridium Difficile
PA-96
Isolation from soil and development of culture methods for
Balamuthia mandrillaris, a meningoencephalitis-causing
Kanako Yamanouchi 1, Hiroaki Arima2, Kosuke Kasai2, Takakiyo Tsujiguchi3, Ryo Saga3, Koichi Ito2,
Takashi Inaba2
Shyh-Ming Li
Hsin Chu Armed Force Hospital Laboratory Medicine, Taiwan
Case Conference
1 Division
of Bioscience and Laboratory Medicine, Hirosaki University Graduate School of Health Sciences, Japan
2 Div.
of Bioscience and Laboratory Medicine, Hirosaki Univ.
3 Dept.
of Radiological Life Sciences, Hirosaki Univ.
Background: Clostridium difficile (CD) is the major pathogen of nosocomial
diarrhea, but the conventional methods for the detection of CD is timeconsuming ( ＞ 3 days) and expensive. A two-step testing strategy includes
an initial negative screening via the detection of the glutamate dehydrogenase (GDH) antigen and GDH+ samples are then tested by a rapid toxin
test. This study evaluated the cost-effectiveness strategy and an overview
of baseline prevalence for CD in two regional hospitals in north Taiwan.
Oral Presentation
Poster Presentation
PURPOSE: Balamuthia mandrillaris is a free-living amoeba that in mammals can often cause Balamuthia amebic encephalitis (BAE). There have
been over 200 reports of Balamuthia infection worldwide, but the source
and route of infection remain largely unknown, with treatment not yet
established. Accordingly, infection has almost 100% mortality, and survivors are left with by severe neurological deficits. In this study, to clarify
the habitat of B. mandrillaris , we attempted to isolate the amoeba from soil
of Aomori Prefecture, Japan and simultaneously attempted to develop a
cultivation method for B. mandrillaris .METHODS AND MATERIALS:A
total of 13 soil samples were collected from arable plots of land and from
homes, cultured and used for soil DNA polymerase chain reaction analysis
at our laboratory. Samples suspected to contain Balamuthia amoebas were
amplified in Balamuthia-specific PCR, and PCR products were then used
for sequence analysis. B. mandrillaris derived from soil was stained with
propidium iodide and SYTO9 and observed with a confocal microscope
to confirm the presence of symbiotic bacteria. Previously reported B.
mandrillaris culture methods were used to promote growth of symbiotic
bacteria and we then investigated the effect of multiple botanical powder
formulations for limiting nutrient uptake in B. mandrillaris derived from
soil culture.RESULTS: PCR results indicated Balamuthia DNA in 4 samples. It was also isolate a B. mandrillaris from a soil sample. In the past, environmental isolation of B. mandrillaris had been disproportionately found
in regions with warm climate, such as South America and the Middle East.
However, as B. mandrillaris was isolated from soil from Aomori Prefecture,
which has a cold climate, our results suggest that this species can adapt to
other types of environment. Lastly, our findings that B. mandrillaris contained a large amount of live bacteria indicate the presence of symbiotic
bacteria in this species.
Methods: 95 stools were collected from the suspected CD infection cases
( ＞ 18 years of age) admitted in two regional hospitals in north Taiwan.
The samples were underwent anaerobic stool culture during the period
of January 2015 to January 2016. The cost-effectiveness analysis of GDH
and toxin combined assay (brand A) versus a two-step algorithm for optimal detection of toxigenic CD. The RIDAQUICK GDH and Toxin A/B were
used for algorithm assay.
Results: Reagent costs reduced 37% by using algorithm assay. A total of
95 suspected CDI patients, 27.4% (26/95) were GDH positive. Distribution
of gender with GDH+ samples was 16 males and 10 females. Most GDH+
samples occurred in aged ＞ =65 years (80.8%, 21/26). Eight (8.4%, 8/95)
were both positive of GDH and toxin. The risk of toxigenic CD (GDH+/Toxin
A/B+) infection between male and female was not significant (50% vs 50%;
p=0.887).
Conclusion: GDH served as an early diagnostic tool for outbreak control. Two-step algorithm for the detection of infection is a reliable, costeffective, and time-saving approach. The positive rate of toxicgenic CD
infection based on a two-step algorithm is similar to the previous literature
reports in Taiwan. Stool samples of GDH+/Toxin- are necessary to isolate
the strains and confirm by PCR.
68
Molecular typing of Acinetobacter baumannii clinical isolates
from a hospital in Nonthaburi Province, Thailand
PA-98
Improvement of Group B Streptococcus screening positive rate for pregnant women to reduce
neonatal infection
Increase of Group B Streptococcus screening positive rate for pregnant women more than 18%
Rachaneeporn Tiyawisutsri1, Tippawan Muennoo2, Ryoichi Saito3, Chihiro Tani4
Ching-Yu Chang1, Hsiu-Chen Lin1,2, Hung-Tu Lin1
1 Department
of Transfusion medicine and Clinical microbiology, Faculty of Allied Health Sciences, Chulalongkorn University,
Bangkok, 10330, Thailand
2 Graduate
Programme in Molecular Science of Medical Microbiology and Immunology, Faculty of Allied Health Sciences,
Chulalongkorn University, Bangkok, 10330, Thailand
3 Department
of Microbiology and Immunology, Graduate School of Health Care Sciences, Tokyo Medical and Dental University,
Tokyo, 113-8510, Japan
4 Department of Microbiology and Immunology, Tokyo Medical and Dental University (TMDU)
1 Department
of Laboratory Medicine , Taipei Medical University Hospital, Taiwan
2 Department
of Pediatrics, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan
Acinetobacter baumannii is opportunistic pathogen that becomes one of
PB-01
Sulforaphane promotes macrophage phagocytosis and natural killer cell
activities in a leukemia mouse model
sulforaphane, WEHI 3 induced leukemia mouse model, phagocytosis, macrophage
Yung Luen Shih1, LUNG YUAN WU2, HSU FENG LU3, JING GUNG CHUNG4
Dept. of Clinical Laboratory Science, Daejeon Health Institute of Technology, Korea
1 Department
of Pathology and Laboratory Medicine, Shin Kong Wu Ho Su Memorial Hospital, Taipei, Taiwan
2 School
of Chinese Medicine for Post Baccalaureate, I Shou University, Kaohsiung, Taiwan
3 Department
of Clinical Pathology, Cheng Hsin General Hospital, Taipei, Taiwan
4 Department
of Biological Science and Technology, China Medical University, Taichung, Taiwan
Scrophularia ningpoensis hemsl has been traditionally used in China and
69
Poster Presentation
Sulforaphane (SFN) is an isothiocyanate, inducing cytotoxic effects in various human cancer cells, including leukemia cells through cell cycle arrest
and apoptosis. However, the effect of SFN on the immune responses in a
leukemia mouse model remains to be investigated. The present study investigated whether SFN has an effect on the immune responses in a WEHI
3 induced leukemia mouse model in vivo. Normal BALB/c mice were injected with WEHI 3 cells to generate the leukemia mouse model, and were
subsequently treated with placebo or SFN (0, 285, 570 and 1,140 mg/kg)
for 3 weeks. Following treatment, all mice were weighted and blood samples were collected. In addition, liver and spleen samples were isolated to
determine cell markers, phagocytosis and natural killer (NK) cell activities,
and cell proliferation was examined using flow cytometry. The results indicated that SFN treatment had no significant effect on the spleen weight,
however it decreased liver and body weight. Furthermore, SFN treatment
increased the percentage levels of CD3 (T cells) and CD19 (B cell maker),
however had no effect on the levels of CD11b (monocytes) or Mac 3 (macrophages), compared with the WEHI 3 control groups. The administration
of SFN increased the phagocytosis of macrophages from peripheral blood
mononuclear cells and peritoneal cavity, and increased the activity of NK
cells from splenocytes. Administration of SFN promoted T and B cell proliferation following stimulation with concanavalin A and lipopolysaccharide,
respectively.
Oral Presentation
Vietnam for treatment of bacteria, atopy, pimple, tonsillitis, angina and
encephalitis for a long time. The main objectives of this study were to
evaluate the antibacterial activity of the Scrophularia ningpoensis hemsl
extract on biofilm formation of Klebsiella pneumoniae . Antibacterial activity was conducted using disc diffusion assay and minimum inhibitory
concentration (MIC), and minimum bactericidal concentration (MBC) were
determined using the broth micro dilution method in accordance to Clinical and Laboratory Standards Institute guidelines(CLSI). Furthermore,
cytotoxicity on L929 were assessed using animal cell culture for the proliferation test(MTT cell assay) and the biofilm forming capacity of the K.
pneumoniae were determined using the colony forming unit (CFU) assay.
The extract exhibited considerable antibacterial activity. K. pneumoniae
was susceptible to the extract with the MIC and MBC of 0.1875 and 1.5 ㎎
/㎖ respectively. Cytoxicity test in L929 showed no sign of toxicity at the
concentration of 0.75 ㎎ / ㎖ and at the same concentration the extract
caused inhibition of bacterial biofilm formation. The extract of Scrophularia ningpoensis hemsl possesses an in vitro antibacterial antibiofilm
activities against K. pneumoniae , with no sign of cytoxicity on L929.
Case Conference
Keun-Dol Yook, Na Young Ha
Symposium
Antimicrobial activity and cytotoxicity test of Scrophularia
ningpoensis hemsl extracts against Klebsiella pneumoniae
Educational Lecture
PA-99
Special Lecture
the most important nosocomial infections associated with elevated morbidity and mortality. Increasingly reports of multidrug-resistant (MDR) A.
baumannii are shown worldwide. The aim of this study is to investigate
molecular typing of A. baumannii clinical isolates using Pulsed-field gel
electrophoresis (PFGE) and PCR-based open reading frame typing method
(POT). In this study 62 non-duplicate isolates of A. baumannii were collected from a hospital in Nonthaburi Province, Thailand between 2012
and 2013. A total of 62 non-duplicate clinical isolates of A. baumannii
were tested for antimicrobial susceptibility (the disk diffusion method)
and molecular typing by PFGE and POT method. All of the isolates (100.0
%) were identified as MDR A. baumannii . Ten antibiotic drugs were tested,
only ampicillin / sulbactam revealed the lowest resistance rates, approximately 88.7 %. Eleven different PFGE patterns and 26 POT types were
identified by PFGE and POT method, respectively. One predominate PFGE
pattern was found; designed as PT1 (59.7 %), followed by PT2 (9.7 %),
PT3 (8.1 %), PT4 (6.5 %), PT5 (6.5 %) and 6 unique type. For POT method,
POT type IX was the largest cluster (27.4 %), followed by POT type V (12.9
%), POT type VI (11.3%), POT type XV (4.8%). The discriminatory power
using Simpson's Index of Diversity (SID) with 95% confidence intervals revealed that POT method had higher SID (0.898, range 0.845-0.951) than
PFGE (0.628, range 0.495-0.761). For genotyping of A. baumannii clinical
isolates, POT method can be a rapid, useful and has high discriminatory
power, among various method such as PFGE and MLST.
Invited Lecture
Objectives:
Group B Streptococcus (GBS) can result in severe neonatal infection, such
as meningitis, pneumonia, and sepsis. The mortality of these infections
was 10-15% and may result in neurological sequelae, these morbidity of
infected newborn needed long-term medical care. The perinatal infection
came from the GBS, which was colonizer in the vagina, anus, and transmitted to newborn via delivery. Therefore, improvement of screening positive
rate for pregnant women is a important clinical issue. According to the
surveillance data of carrier rate of GBS in pregnant women is 18% in Taiwan. In our laboratory, the mean positive rate of GBS in 2013 was 17.9%.
And the positive rate was less than 18% during seven months.
Methods:
We took action immediately to improve the positive rate. First, we replace
the enhanced broth (LIM broth) with chromogenic Strep B Carrot Broth;.
It can increase the sensitivity and specificity, and decrease the incubation
period. At the same time, we change the ordinary transtube to Amies transtube without charcoal, in order to decrease the interference by charcoal.
Furthermore, we used GBS DetectTM to replace the BAP agar. The agar
is supplied with selective nutrients to inhibit other bacterial growth and
enhanced beta-hemolysis of GBS. Finally, we performed the CAMP test to
confirm the isolates.
Results:
Following the improvement, we found the mean positive rate of GBS of
vaginal screening in 2014 and 2015 were 23.6% and 25.0%, respectively.
They significantly increased from 2013 (p ＜ 0.01). In addition, there was
not any monthly positive rate less than 18%.
Conclusion:
We performed the quality improvement of yield rate via the analysis, then
by change the transtube, culture broth, and detection agar. Finally, we
obtained the increased positive rate and maintained the high yield rate.
In conclusion, this improvement may decrease the neonatal infection, and
increase the patient safety.
Keynote Speech
PA-97
Keynote Speech
PB-02
Analysis of HLA allele in infertile patients from Taiwan
population
PB-03
HCV Related Cryoglobulinemia: A Case Report
An Experience of a Regional Teaching Hospital in Hsin-chu
of Taiwan
Ching Ju Chen
Yi Shun Chen1, Wei Yung Lo2, Ya I. Hsiao*1
Department of Clinical Pathology, Chi Mei Medical Center, Tainan, Taiwan
1 Laboratory
Medicine Department, National Taiwan University Hospital Hsin-Chu Branch, Taiwan
2 Division
of Rheumatology, Department of Internal Medicine, National Taiwan University Hospital Hsin-Chu Branch, Taiwan
Invited Lecture
Special Lecture
Educational Lecture
The reasons of infertility are very complex. Sperm immobilizing antibody
(SIA) in the sera of women cause low pregnancy rate. HLA alleles DR and
DQ may account for higher frequency in SIA groups. Our purpose was to
investigate the distributions of HLA alleles and estimate their associations
with infertile women in Taiwan population. We analyzed HLA database
of 100 infertile couples from Chi-Mei Hospital in southern Taiwan. HLA
typing was performed by polymerase chain reaction sequence specific
primer (PCR-SSP) method. The mean age of female infertile patients
was 37.8+/-7.5 years. There were 19 genes relative frequency for HLADRB1 and DQB1 in contrast to normal population. The frequency of HLA
DRB1*0301 (DR17) was 26.0% in infertile women, as compared to the
reference normal group (14.5%). The patients group was higher ratio
than normal group in 1.8X. A HLA-DQB1*0201 (DQ02) gene frequency of
26.0% was found in the infertile women in contrast to 19.0% in the normal
population. These results showed an increase in HLA DR17 and DQ02
alleles in the women patients compared with the control population. On
the other hand, we analyzed the HLA alleles sharing of 100 couples. There
was a highly significant similar of HLA sharing in the couple who have 7
HLA sharing antigens. There were 10 couples shared there HLA antigens,
15 couples shared four HLA antigens, and 5 couples shared five HLA antigens. The some cases of infertility might be caused by closed histocompatibility between partners, it can be performed desensitization therapy effectively. The high frequency of HLA-DRB1*0301 DQB1*0201 among the
patients may account for a higher frequency of SIA in the southern Taiwan
population. Since the causes of infertility are many, HLA typing used can
help clear reason of infertile couples.
Symposium
PB-04
Cryoglobulins, in three main types, are a group of immunoglobulins that
undergo reversible precipitation at low temperatures. Trace serum cryoglobulin levels , in the case of mixed type, could be identified in about
50% hepatitis C patients, usually together with appearance of rheumatoid
factors (RF) and antinuclear antibodies (ANA). Neglecting prior hepatitis
C history, clinicians might miss making pegylated interferon or anti-viral
medications to work for people who have associated cryoglobulinemia
due to misdiagnosed atypical rheumatoid arthritis. In clinical practice,
with higher awareness, cryoglobulin test would be ordered to perform for
chronic hepatitis C patients developing the following symptoms alone or
in combination: purpura, arthritis, glomerulonephritis, neuritis, etc.
This report here describes a case of highly suspected hepatitis C virus (HCV)
related secondary cryoglobulinemia. A 66-year-old man with HCV antibodies visited our hospital and presented with polyneuropathy on October 21,
2015. Laboratory data discovered trace-positive cryoglobulin, elevated
RF (35 IU/mL), positive ANA (titer of 1:40; speckled pattern), increased
erythrocyte sedimentation rate (1,2 hr: 17, 36mm ), elevated IgG ( 1730
IU/mL). Four weeks later, further analyzed data revealed the appearance
of HCV RNA ( genotype 1b), with HCV viral load up to 3.92 x10 7 IU/mL,
elevated AST/ALT ( 37/51 IU/mL), besides, polyclonal gammopathy could
be seen in serum protein electrophoresis.
In the laboratories, cryoglobulins are hard to accurately detect because of
temperature sensitive nature and generally trace amount in blood specimens. Therefore, a proper pre-analytical procedure of blood collecting is
the key point. Planning how to increase the detection rate and persistent
follow-up of laboratory data as well as therapeutic effects will be the goals
we are striving for in the future.
Seroprevalence and risk factors associated with Toxoplasmosis among HIV positive patients at TASO Entebbe, Uganda
Detection of Toxoplasma gondii using Rapid Testing Kits and ELISA Toxoplasma IgG and IgM Kits at TASO Entebbe,
Uganda
PB-05
THE USEFULNESS OF C-REACTIVE PROTEIN AND OTHER BIOMARKERS IN DIAGNOSING
TUBERCULOSIS/HIV-COINFECTION IN RESOURCE LIMITED SETTINGS
C-REACTIVE PROTEIN, ESR AND CD4 COUNT IN DIAGNOSING TUBERCULOSIS/HIV-COINFECTION
Case Conference
Oral Presentation
Poster Presentation
Joseph Ndarubweine, Polly Rwandekeye, Josephine Lunkuse
Anietie E. Moses, Veronica G. Bassey
Programs, TASO Uganda, Uganda Laboratory Technology Association (UMLTA), Uganda
Department of Medical Microbiology and Parasitology, University of Uyo, Nigeria
Introduction
Toxoplasmosis is an infection caused by a single celled parasite Toxoplasma gondii whose definitive host is the cat. It is estimated that 10 to 30%
of HIV 1 infected patients are seropositive for anti toxoplasma antibodies
and 25 to 50% patients experience symptomatic infections in the absence
of antimicrobial prophylaxis (Porter, 1992).
The AIDS Support Organization Entebbe in Uganda cares for 6,200 HIV
positive clients with 1 to 2 patients diagnosed with toxoplasmosis per
clinic irrespective being on AntiRetroviral Therapy and Cotrimoxazole
prophylaxis.
Objective
The objective of this study was to determine the seroprevalence of toxoplasmosis among HIV positive patients and the associated risk factors at
TASO Entebbe.
Methodology
The study employed a cross sectional design and random sampling method was used.
Results
The seroprevalence of Toxoplasmosis was 23.5% using Toxoplasma rapid
IgG and IgM test kits. Seroprevalence of Toxoplasmosis increased with age
ranging from 18.9% in the 18 to 25 years age group to 28.3% in greater
or equal to 45 years age group. Toxoplasma seroprevalence was higher in
male at 30.5% than in female at 20.2%. Toxoplasma seroprevalence was
higher among the fishermen at 53.8% compared to other occupations.
The toxoplasma seroprevalence was higher among the Muslims at 35.7%
compared to other religions at 15.0% to 23.8%. The toxoplasma seroprevalence among patients who have been on ART for 3 to 6 months was higher
at 41.7%. The seroprevalence for toxoplasmosis was high among clients
who have ever been treated for toxoplasmosis at 40.0%.
The results of this study were similar to studies conducted in South Africa
at 26%, (Sonneberg, Silber and Jentsch, 1998), and 24.6%, Milongo et al,
2000.
It is recommended that all newly diagnosed HIV patients be tested for
Toxoplasma gondii antibodies and confirmed with a molecular diagnostic
technique.
The challenge of tuberculosis diagnosis and disease progression monitoring in HIV coinfected persons is enormous especially in resource poor
countries with high burden of HIV and tuberculosis coupled with limited
diagnostic capacity. Assessment of biomarkers levels during active tuberculosis in HIV infected persons could be a veritable tool in diagnosis and
disease monitoring. This study investigates the association between C-reactive protein (CRP), erythrocyte sedimentation rates (ESR) and absolute CD4
counts in HIV patients coinfected with tuberculosis in Uyo-Nigeria. A total
of 72 HIV positive and 17 TB&HIV coinfected patients were recruited into
the study along with 27 apparently health blood donors as control. Serum
CRP levels, ESR and absolute CD4+ count were measured using sandwich
ELISA, Westergren and Flow cytometer techniques respectively. The mean
serum CRP level in TB&HIV coinfection (58.79 ± 38.2mgperdl) was significantly greater than HIV positive (20.45 ± 28.5mgperdl), TBpositive&HIV
negative (12.34 ± 20.9mgperdl) and apparently healthy blood donors
(0.44 ± 0.64mgperdl), except those with TB infection alone (29.83 ±
30.8mgperdl) (P<0.05). The mean serum CRP levels in TB positive alone
was significantly greater than the control group (P<0.05). The mean
ESR in TB&HIV coinfection (129.88 ± 15.5mmperhr) was significantly
greater than HIV positive only (106.15 ± 26.4mmperhr), TB positive only
(94.15 ± 30.9mmperhr) and HIVpositive&TBnegative groups (59.28 ±
36.6mmperhr) (P<0.05). There was no statistical significant difference between the mean ESR of TB and HIV positive groups but both groups were
significantly greater than HIVpositive&TB negative group (P<0.05). The
mean CD4+ count in TB&HIV coinfection (175.12 ± 85.79cellsperml) was
significantly lower than HIV positive pe(358.93 ± 240.1cellsperml), TB
positive (576.31 ± 326.3cellsperml) and HIVpositive&TBnegative groups
(1089.8 ± 331.3cellsperml). The study concludes that the use of serum
CRP levels alone or in combination with ESR and CD4+ count are promising indicators that could predicting active TB and disease progression in
TB&HIV coinfection especially in high disease burden areas.
70
PB-07
A case of prozone effect on PRA
Hyeyoung Kim, Younghee Im, Sukyung Kim, Heungbum Oh
PREVALANCE OF ROTA VIRUS AMONG CHILDREN UNDER
FIVE YEARS OLD ATTENDING PEDIATRIC CLINIC AT KNH.
ROTA VIRUS IN KENYA
Jeremiah o zablon, vincent gitau, bakari mwinyi, roselyne nyamwaka, flelisters obara, mary masakha,
PETER MATHU
Laboratory science, Asan Medical Center, Korea
Immunology, Kenyatta National Hospital, Kenya
Rotavirus is the most common cause of severe diarrhea among infants and
young children. It is a genus of double-stranded RNA virus in the family
Reoviridae. The virus is transmitted by the faecal-oral route. It infects and
damages the cells that line the small intestine and causes gastroenteritis.
The study objective was determine the prevalence of Rota virus in Kenyatta National hospital among children below 5 years attending pediatric
filter clinic. Sample size was 135 samples which was analyzed by ELISA
method. Data was analyzed by chi square statistical method.The study
showed a rotavirus prevalence of 32.163%. The prevalence of rotavirus
in relation to gastroenteritis among children under five years was 47.413
The prevalence was higher in children under 1 years old. The was no
difference on Rotavirus prevalence between male and female.This study
showed that the prevalence of rotavirus was high.
Special Lecture
Case: A 45-year old man had kidney transplantation at 2009. His creatinine increased to 1.49 mg/dl and PRA revealed DQ7 DSA (MFI 5,944) at
October 2014. Soon after, he had undergone kidney biopsy and ABMR
was diagnosed. He was treated with Rituximab, intravenous immunoglobulin and four sessions of plasma exchange. After desensitization, PRA was
retested at December 2015 and DQ7 DSA (MFI 19,807) was identified.
Because his DSA MFI increased significantly after desensitization, we suspected PRA result at October 2014 may be due to prozone effect. Ten-fold
dilution of the sample drawn at October were done to confirm the prozone
effect, and DQ7 DSA (MFI 18,084) was identified.
Educational Lecture
Conclusion: HLA molecules on cell surface are floating in the membrane.
This motility facilitates the binding of antibodies. However, HLA molecules
on beads are bound rigidly to the surface and cannot move. Therefore, the
density of HLA antigens on beads can influence the binding of antibodies.
The high HLA density on beads increases sensitivity, but it also increases
risk of prozone effect. If the antibody titer is very high in patient serum,
and HLA density on beads are high, antibodies can be packed around the
beads, resulting univalent binding. Because univalent binding is not stable,
it could be washed away through washing steps, resulting unexpected low
MFI. Most patients who receive desensitization have high titer of HLA antibodies. Therefore, it is recommended to test sample with 10 fold dilution
together in monitoring desensitization effect.
Immune responses of macrophage to the extracellular polysaccharide of Vibrio vulnificus
Effect of extracellular polysaccharide on the infection of Vibrio vulnificus induced
proinflammatory cytokines in RAW264.7 cell
PB-09
Comparison of Indirect Immunofluorescence Assay and EliA CTD Screen for
Measurement of Antinuclear Antibodies
Comparison of Indirect Immunofluorescence on HEp-2 Cells and EliA CTD Screen
Hao-Yun Chou1, Yen-Jung Lu1, Szu-Yu Lin1, Li-Ping Yuan1, Wei-Ming Chi1,3, Wen-Shyang Hsieh1,2,3
1 Department
of Laboratory Medicine, Taipei Medical University – Shuang Ho Hospital, New Taipei City, Taiwan
2 Department
of Education, Taipei Medical University – Shuang Ho Hospital, New Taipei City, Taiwan
3 School
of Medical Laboratory Science and Biotechnology, Taipei Medical University, Taipei, Taiwan
71
Poster Presentation
Our study aimed to evaluate the agreement of indirect immunofluorescence (IFA) on HEp-2 and EliA CTD screen (ECS) for ANA screening, and
to determine whether ECS can replace IFA. 711 serum samples were collected from the patients with suspected connective tissue disease, and
were examined simultaneously using IFA and ECS. Overall, 73 (10.3%)
were positive by IFA, 78 (11.0%) were positive by ECS, and 41 (5.8%) were
positive by both assays. After exclusion of all ECS-equivocal results, ECS
had sensitivity of 58.6%, specificity of 94.0%, positive predictive value
of 52.6%, negative predictive value of 95.3%, accuracy of 87.8%, positive likelihood ratio of 9.81, negative likelihood ratio of 0.44, and kappa
coefficient of 0.434, when IFA was used as the reference method for ANA
screening. Among the cases of common IFA patterns, 100% centromere,
73.3% speckled, 25.0% homogeneous, and 22.2% nucleolar pattern were
ECS-positive. The inconsistent results may be due to that antigen panel of
ECS not includes histone and nucleosome. IFA and ECS exhibited excellent agreement in the patients with anti-SSB/La, anti-SM, or anti-RNP. Of
34 patients with anti-SSA/Ro, 94.1% cases were ECS-positive and 47.1%
cases were IFA-positive. 92.9% cases were ECS-positive and 57.1% cases
were IFA-positive in 14 patients with anti-dsDNA. ECS had a better ability
to detect anti-SSA/Ro and anti-dsDNA than IFA. In conclusion, IFA and ECS
complement each other, so a combination of both assays can provide better performance of ANA testing.
Oral Presentation
As an essential component of innate immunity, macrophages have multiple functions in both inhibiting or promoting cell proliferation and tissue
repair. Diversity and plasticity are hallmarks of macrophages. When it
activated with different types of antigen, macrophages can acquire distinct
functional phenotypes via undergoing different phenotypic polarization,
including classically activated macrophages (M1) and alternatively activated macrophages (M2). The M1 macrophages are characterized by the
secretion of proinflammatory cytokines, including interleukin-1 beta, and
tumor necrosis factor-alpha. Conversely, the M2 macrophages induce a
weak immune response and reduce pro-inflammatory reactions with the
production of interleukin-10. However, when the bacteria lost it's virulence factor, but with the same infectivity, the innate immune reaction of
regulate macrophage may still remain unidentified. The EPS is demonstrated to be crucial in the induction of immune components which are
involved in the inflammatory response. This study examined the induction
of proinflammatory cytokines induced by the infection of wild type strain,
without the capsule of mutant strain (JF045), and the complementary
strain (JF049) of Vibrio vulnificus to understand whether the antigen contributes to the induction of proinflammatory cytokines and the mechanism
of pathogenesis. The bacteria strains including YJ016, JF045, and JF049
were applied in the infection of Vibrio vulnificus strains to RAW264.7 cell.
The extracted cellular RNA at the infection for 0.5, 1, 1.5, and 2 h were
applied in the reverse transcription and real-time PCR for measuring the
expression of cytokines including IL-1 beta, TRAF6, TLR-2, TLR-4, Arg-1,
TNF-alpha, and IL-4. The results help us to understand the effect of EPS on
the M1-M2 polarization of macrophage.
Case Conference
Shih-Chieh Lo, Ting-Yi Wang, Horn-Ren Lo, Shiao-ping Huang
Department of Medical Laboratory Science and Biotechnology, Fooyin Unviersity, Kaohsiung, Taiwan
Symposium
PB-08
Invited Lecture
Background: Donor-specific HLA antibody (DSA) can cause antibody-mediated rejection (ABMR). It is also a powerful markers to predict ABMR after
transplantation. Many laboratories use Luminex PRA to detect DSA. It is a
highly sensitive method, but it also has some limitations. We experienced a
case with prozone effect on PRA.
Keynote Speech
PB-06
Keynote Speech
Comparison of Indirect Immunofluorescence Assay and EliA CTD Screen for Measurement of
Antinuclear Antibodies
Advantages and Disadvantages of Two Commercial Assays for Antinuclear Antibodies Screening
PB-10
PB-11
Combining serum Procalcitonin with Lactic Acid as
biomarkers to elevate the diagnosis of bacteremia
Hao-Yun Chou1, Yen-Jung Lu1, Szu-Yu Lin1, Li-Ping Yuan1, Wei-Ming Chi1,3, Wen-Shyang Hsieh1,2,3
Chao-Wei Liu, Yu-Tin Cheng, Shi-Ying Huang
1 Department
of Laboratory Medicine, Taipei Medical University-Shuang Ho Hospital, New Taipei City, Taiwan
2 Department
of Education, Taipei Medical University-Shuang Ho Hospital, New Taipei City, Taiwan
3 School
of Medical Laboratory Science and Biotechnology, Taipei Medical University, Taipei, Taiwan
Department of Laboratory Medicine, National Taiwan University Hospital, Taiwan
Invited Lecture
Special Lecture
Bacteremia is the presence of bacteria in the blood which is always abnormal. The immune response to the bacteria can cause sepsis and septic
shock, which has a relatively high mortality rate. Therefore, it's a high
priority for the doctors to diagnose bacteremia. Bacteremia is most commonly diagnosed by blood culture. However, blood culture usually take
2-5 days for bacteria to multiply in order to be detected. We want to find
out whether there are other biomarkers that can help diagnose bacteremia
and can be detected in blood sample in just a few hours after blood collection. Measurement of procalcitonin can be used as a marker for infection caused by bacteria although levels of procalcitonin in the blood are
very low. In our study, we want to use another biomarker to increase the
diagnosis of bacteremia, ex: the level of lactic acid. Lactic acid gets higher
when strenuous exercise or severe infection.
We collected 185 bacteremia patients' samples from 2015/01/01 to
2015/12/31 in our hospital. Then we tested patients' procalcitonin ,lactic
acid level and also ran blood culture. All blood culture came out positive
with bacteria. The procalcitonin level range from 0.0602 to 266.8 ng/mL.
By using procalcitonin as a biomarker (cut off: 0.5 ng/mL), the sensitivity
was about 80%. If we combine lactic acid as a supplement biomarker (cut
off: 2.2 mmol/L), the sensitivity would elevate to 92%. In addition, out of
the 44 patients who had a higher procalcitonin level ( over 10 ng/mL), 30
patients' blood culture result in GNB growth. Furthermore, we want to test
the level of procalcitonin and lactic acid in blood culture results negative
bacteremia patients to ensure our finding.
Our study aimed to evaluate the agreement of indirect immunofluorescence (IFA) on HEp-2 and EliA CTD screen (ECS) for ANA screening, and
to determine whether ECS can replace IFA. 711 serum samples were collected from the patients with suspected connective tissue disease, and
were examined simultaneously using IFA and ECS. Overall, 73 (10.3%)
were positive by IFA, 78 (11.0%) were positive by ECS, and 41 (5.8%) were
positive by both assays. After exclusion of all ECS-equivocal results, ECS
had sensitivity of 58.6%, specificity of 94.0%, positive predictive value
of 52.6%, negative predictive value of 95.3%, accuracy of 87.8%, positive likelihood ratio of 9.81, negative likelihood ratio of 0.44, and kappa
coefficient of 0.434, when IFA was used as the reference method for ANA
screening. Among the cases of common IFA patterns, 100% centromere,
73.3% speckled, 25.0% homogeneous, and 22.2% nucleolar pattern were
ECS-positive. The inconsistent results may be due to that antigen panel of
ECS not includes histone and nucleosome. IFA and ECS exhibited excellent agreement in the patients with anti-SSB/La, anti-SM, or anti-RNP. Of
34 patients with anti-SSA/Ro, 94.1% cases were ECS-positive and 47.1%
cases were IFA-positive. 92.9% cases were ECS-positive and 57.1% cases
were IFA-positive in 14 patients with anti-dsDNA. ECS had a better ability
to detect anti-SSA/Ro and anti-dsDNA than IFA. In conclusion, IFA and ECS
complement each other, so a combination of both assays can provide better performance of ANA testing.
Educational Lecture
Symposium
Anorexia in the elderly and its association with latent inflammation:
the roles of inflammatory cytokines and appetite-related hormones in the
pathogenesis of anorexia of aging.
PB-12
1
1
1
2
PB-13
1
Differential Gene Experessions in Systemic Lupus Erythematosus (SLE), Sjogren's &
Sjorgen's Syndrome associated with SLE
Effects of BAFF and IFN-alpha on Expressions of Genes Invovled in Lipid Homeostasis
Case Conference
Fumika Hirano , Eri Nanizawa , Yumi Hayashi , Toshihiro Matsuura , Tetsuya Ishikawa
Su H Cho, Meredyth Wilkinson, Kirsty Waddington, Nicolyn Thompson, Elizabeth Jury
1 Department
of Radiological and Medical Laboratory Sciences, Nagoya University Graduate School of Medicine, Japan
2 Department
of Gastroenterology, National Center for Geriatrics and Gerontology
Division of Medicine, Faculty of Medical Sciences, University College London, United Kingdom
Oral Presentation
Poster Presentation
Introduction
B-cell activation factor (BAFF) is known to be associated with lipid raft
disruption in autoimmunity. The role of BAFF and its receptor on lipid
rafts of B lymphocytes leading to changes in downstream signalling pathways causing autoimmunity has been previously established, in SLE and
Sjogren's Syndrome. IFN-alpha is known to cause BAFF expression. In this
study, the genes associated with lipid raft signalling, and proteins involved
in lipid metabolism were studied to elucidate the relation between them
and the three autoimmune diseases, Systemic Lupus Erythematosus (SLE),
primary Sjogren's Syndrome (pSS) and Sjorgren's syndrome associated
with SLE.
Methods
Blood samples from patients and healthy controls, within the age-range
of (25-77), were used in this study. Patients were undergoing different
therapies - oral steroids, hydroxychloroquine, rituximab, azidothymidine/
mycophenolate mofetil/ methotraxate, and statins for cardiovascular
complications. Peripheral blood mononuclear cells (PBMCs) were isolated
using density gradient centrifugation. PBMCs were then cultured +/- BAFF
& IFN-alpha cytokines for 6, 24 and 72 hours to study their effects on B
lymphocytes and CD4+ immune cell lipid homeostasis. Plasma membrane
expression of cholesterol (filipin binding) and glycosphingolipids (using
cholera toxin-B subunit) were assessed by flow cytometry. The expresssional analysis of the genes influenced by these cytokines was studied using RT-qPCR.
Discussion
The expression of genes involved in metabolic pathways related to lipid
metabolism and homeostasis were studied, to test the effect of the two
cytokines BAFF & IFN-alpha. The genes included - SREBP1 & 2, HMGCR,
FASN, UGCG, SCD5, DHCR24, NPC1 & 2. After being confirmed, the differences in expression levels of genes in patients & healthy controls, between
these diseases, would be reported in the presentation.
Objectives: A certain proportion of the elderly people suffer from anorexia
without evident etiology. To understand the pathogenesis of anorexia in
these elderly people, we investigated the relation of appetite status to serum levels of appetite-related hormones and inflammatory cytokines. We
also investigated the influence of hunger hormone, ghrelin, on the inflammatory responses in the mouse liver injury models.
Methods: Twenty-four elderly people of 65 years and older were enrolled
in the study. Seven of them were the patients complaining of anorexia, and
17 of them were the controls received medical check-up. They all received
the assessment test of nutrition (MNA: Mini Nutrition Assessment-Short
Form, Nestlé Nutrition Institute), body measuring, and the blood tests for
hematology and biochemistry. The sera were also collected and stored
in -80C °, and served for the analysis of hormones ( ghrelin, leptin, and
adiponectin) and cytokines (TNF- α , et al.). To investigate the regulatory
function of ghrelin on inflammatory responses, we conducted mouse experiments in which we tested the effect of ghrelin administration on CCl4and concanavalin A-induced liver injury.
Results: BMI and MNA score were significantly lower in the patients compared to the controls (p<0.05). In the patients, serum TNF- α levels were
significantly higher than those in the controls (p<0.05). Serum levels of
ghrelin were tended to be high, while those of leptin were tended to be low
in the patients. In the mouse liver injury models, ghrelin administration
ameliorated liver inflammation and lowered mRNA expression of inflammatory cytokines, such as TNF- α .
Conclusions: The elevation of serum TNF-α levels in the patients suggested
that anorexia in the elderly was an inflammation-based clinical condition.
As ghrelin was proved to exert immune-regulatory action in the mouse
models, tendency of ghrelin elevation in the patients might be a compensatory reaction against the latent inflammation.
72
High-levels of CD8dimCD3-CD4-(NK8) cells is associated with undetectable viral load in HIV- infected
patients post-HAART
CD8dimCD3-CD4-cells associated with undetectable viral load in HIV- infected patients post-HAART
PB-15
Fernando Mendes1, Juan Jose Barcelo1,2, Olga Millan3, Alberto Crespo4, Artur Paiva1, Ana Valado1,
António Gabriel1, Manuel Juan2
Ching-Biau Liou, Sheng-Jun Lin, Chih-Chun Chang, Ming-Jang Su, Fang-Yeh Chu
Department of Clinical Pathology, Far Eastern Memorial Hospital, Taiwan
1 Polytechnic
Institute of Coimbra, ESTESC – Coimbra Health School, Department Biomedical Laboratory Sciences, Coimbra,
Portugal
2 Immunology
Service of the Hospital Clinic of Barcelona, Spain
3 Farmacologic
and Departamen of the Hospital Clinic of Barcelona, Barcelona, Spain
4 HIV
research group fundation of the Hospital Clinic of Barcelona, Barcelona, Spain
Szu-Yu Lin1, Hao-Yun Chou1, Yen-Jung Lu1, Li-Ping Yuan1, Wei-Ming Chi1,3, Wen-Shyang Hsieh1,2,3
Department of Laboratory Medicine, National Taiwan University Hospital, Taiwan
1 Department
of Laboratory Medicine, Taipei Medical University-Shuang Ho Hospital, New Taipei City, Taiwan
2 Department
of Education, Taipei Medical University-Shuang Ho Hospital, New Taipei City, Taiwan
3 School
of Medical Laboratory Science and Biotechnology, Taipei Medical University, Taipei, Taiwan
73
Poster Presentation
Mycoplasma pneumonia is the most common pathogen of atypical pneumonia, practically for school-age children and adolescents. People who
are sick with M. penumoniae infection usually get coughing but without
sneezing, and this symptom is be helpful for distinguished M. penumoniae
from other types of pneumonia. However, such symptom is not specific,
so we need a reliable test method to assist physicians to give accurate
diagnosis and proper treatment for patients to decrease antibiotic abuse.
M. pneumoniae is not easy to culture, so culture is not helpful for clinical
diagnosis. Now serologic test is the most important method in diagnosis
M. pneumonia. For now, M. pneumonia IgM is tested for determine if
pneumonia patients is infected by mycoplasma pneumoniae or not. But
for some patients, their IgM antibody cannot be detected and for some. M.
pneumonia IgM antibodies can persist for months, for all these situation
will be a problem for clinical diagnosis. However, the concentration and
lifespan are different in patients. Some literature point out M. pneumonia
IgA has high sensitivity than IgM in some acute M. pneumonia infection or
reinfected patient. Furthermore, some literature point out that high concentration M. pneumonia IgA in blood is the only indicator for M. pneumonia infection.
The advantage of detecting M. pneumonia IgA is that these antibodies usually appear in re-infection patients, and in this study, now we evaluate M.
pneumonia IgA diagnostic value for elder people reinfect with M. pneumonia.
The test is based on the ELISA principle (Enzyme linked Immunosorbent
Assay). Pipette the test serum 50 ul in well of the strip, and insert the strip
in Chorus Trio system (DIESSE Diagnostica Senese s.p.a.). Finally, the symptoms, age and results are analyzed to discuss the usefulness of
M. pneumonia IgA.
Oral Presentation
Introduction
Urinary tract infection (UTI) is one of the most common infectious diseases. Patients with febrile UTI generally present with mild illness in
primary care but may rapidly develop a life-threatening condition. When
a suspected UTI, generally use the urine routine test and urine culture to
assist with the diagnosis. However, urine routine are nonspecific and urine
culture needs at least 24 to 48 hours to attain the culture result.
Procalcitonin(PCT) are elevated among patients with severe infections
such as septic shock but are normal among patients with noninfectious inflammatory conditions. Plasma concentrations of PCT are very low among
healthy individuals and increase to very high values in response to bacterial endotoxins. Therefore, the purpose of this study was to determine the
ability of PCT measurements, compared with urine routine test and urine
culture in the diagnosis of urinary tract infection as an early diagnosis biomarker of UTI patients.
Methods
This retrospective study from January 2013 to December 2015, 62 patients with febrile urinary tract infection had their PCT levels measured
and all patients had their culture and urine routine test results.
Result
Of 62 patients with UTI, 42 is urine culture positive with PCT results range
from 0.06-1.25 ng/mL, mean 0.4 ± 0.1 ng/mL (Mean ± SD). Urine routine test all have higher sensitivity but lower specificity.
Conclusions
According to the study, PCT results in urine culture positive patients are
significantly higher than healthy people (<0.05 ng/mL). Although this retrospective study comprised a small number of patients, we found that PCT
was a useful marker for urinary tract infection and urine routine test may
play a screening role because their higher sensitivity and convenience.
Case Conference
Chao-Wei Liu, Shau-Han Wu, Ming-Yeh Lin
Symposium
Clinical Application and Diagnostic of Mycoplasma IgA in
Mycoplasma Pneumonia
The usefulness of Mycoplasma pneumonia IgA Assay
Educational Lecture
PB-17
Special Lecture
Procalcitonin as a biomarker in urine tract infection
diagnosis
Invited Lecture
Background: Ambient exposure to air pollutants, especially to PM2.5, was
reported to be linked to a spectrum of diseases. However, few studies estimated the influence of PM2.5 on the prevalence of influenza infection. The
aim of our study was to investigate the relationship of type A and type B
influenza infection to ambient air contamination in the haze weather.
Materials and Methods: The data of rapid diagnostic testing for type A and
type B influenza and monthly PM2.5 concentrations from 2013 to 2015
were obtained.
Results: From 2013 to 2015, the PM2.5 level in the haze weather was
significantly higher than that in the non-haze weather (27.64+/-4.96 vs.
19.76+/-4.22ug/m3, p ＜ 0.001). Additionally, the prevalence of type A
influenza infection was significantly increased in the haze weather when
compared with that in the non-haze weather (20.45+/-8.79 vs. 8.44+/5.58%, p ＜ 0.001). The prevalence of type B influenza infection was also
seemingly higher in the haze weather than that in the non-haze weather
(6.44+/-5.28 vs. 4.54+/-5.09%, p=0.075). Interestingly, the prevalence of
type B influenza infection was significantly increased in the haze weather
with a time lag of 2 months when compared with that in the non-haze
weather with the same time lag (8.25+/-5.93 vs. 3.07+/-4.15%, p=0.003).
Conclusion: Our study disclosed that ambient exposure of PM2.5 was
associated with the increased prevalence of type A and type B influenza
infection. The air contamination could partially attribute to the predicted
flu season. Besides, the lag effect of PM2.5 on the prevalence of type B influenza infection could be associated with alternate seasonal monsoon and
further research is needed.
Introduction: In post-HAART therapy the recuperation in HIV patients
depends on several factors, were the immune system has a key role, contributing for infectivity and capacity spread reduction. The CD3/CD4/CD8
cellular populations in HIV patients in some cases, show an increase of
CD8dim/CD3-/CD4- (NK8) cells above normal values.
Aim: Relate NK8 cells number in HIV patients regarding the evolution of
infection, taking in consideration the trinomial: viral load, TCD4 number
post-HAART and cell phenotype.
Material and methods: A total of 518 HIV patients participated, we determine viral load, TCD4 number in post- HAART therapy period, 15 healthy
controls to determine the cellular phenotype and functional activity. Stimulation of cytokine and cytometric assays were performed using murine
monoclonal antibodies conjugated with different fluorochromes.
Results: Of the 518 HIV patients, 339 show an NK8 expression between
2 and 8%, 124 express less than 2% and 55 expressed more than 8%.
We observed a correlation between negative viral load and NK8 cells values between 2-8 % and 8-25 %. The male patients group with NK8 cells
expression ＜ 2 % presented a higher viral load than the female group.
Patients with NK8 cells expression ( ＞ 2%) exhibit better quotient CD4/
CD8 than those that have expression NK8 ＜ 2%. The TCD8 cells superior
to 500 cells/mm3 and a NK8 cell population ＜ 2%, associates to increasing viral load. The stimulation with PMA plus ionophore was low for interferon-&#611; with higher expression of perforin in NK8 cells.
Conclusions: Values above 2 to 8% of NK8 cells in HIV patients show a
better correlation with undetectable viral load, favouring the CD4/CD8
quotient. Allowing the immune system regeneration for longer periods
without HAART treatment. Suggesting better prognosis in non-progression
of the virus in HIV patient's post-HAAT. The assessment NK8 cells should
be considered as another biomarker in HIV infection.
PB-16
The Association of Influenza Infection with Ambient Air Pollution in the Haze Weather
The Association of Type A and Type B Influenza Infection with Ambient Air Pollution
in the Haze Weather
Keynote Speech
PB-14
Keynote Speech
PB-18
Understanding the relationship between indoleamine
2,3-dioxygenase 1 (IDO1) and arginase 1 (ARG1)
IDO1 controls ARG1 expression in dendritic cells
PB-19
The frequency of the antibody for bovine protein in nonspecific reaction of HTLV-1 antibody
Linn Zimmerman1,2, Claudia Volpi2, Ursula Grohmann2, Giada Mondanelli2
Akira Miyano , Chikako Inaoka, Shinobu Morinaga, Futoshi Fujiwara, Makoto Takeuchi
1 Karlstad University, Sweden
2 Experimental Medicine, University of Perugia, Italy
Osaka medical Center and Research Institute for Maternal and Child Health, Japan
Invited Lecture
Special Lecture
Educational Lecture
Introduction :Antibody for bovine protein may cause the non-specific
reaction of HTLV-1 antibody. However, the frequency is unknown.
Objective:We demonstrate the non-specific reaction of HTLV-1 antibody
by the assay of bovine serum albumin (BSA) IgG antibodies in the patient's
serum, and the absorption test using adult bovine serum (ABS). Methods
:We measured IgG BSA antibodies with the ELISA method using a BSA
sensitization plate and peroxidase mark antihuman IgG antibody in the
sera of 50 patients. The HTLV-1 antibody was measured by "LUMIPULSE
G1200" (Fujirebio). The "SERODIA HTLV-1" (PA method : Fujirebio) were
used for the confirmation of the non-specific reaction of HTLV-1 antibody.
The absorption tests were as follows : 1) Dispense 0.1mL of ABS into a
test tube, 2) Add 0.1mL of the specimen, mix and incubate at 37 degrees
Celsius for 2 hours, 3) Centrifuge at 3000 rpm for 7 minutes, 4) Measure
the HTLV-1 antibody, 5) The phosphate-buffered saline was used instead
of the control to the ABS, 6) The non-specific reaction was used as the
absorption test less than half compared to the control.Results :We determined HTLV-1 antibodies in the 10,639 blood samples. PA method negative and LUMIPULSE positive (1 COI or more) were 102 cases (1%). The 50
blood samples of them were performed about absorption tests (age 0-38
years). Then 40 samples had a non-specific reaction in the ABS. It could
well absorb with ABS than BSA in the absorption tests. The BSA antibody
index of 40 samples as non-specific reaction (19.2 ± 9.0 index, 0-37 years
old) were significantly higher than those of 10 samples as specific reaction
(8.4 ± 8.4 index, 0-38 years old) (p = 0.0012). Conclusion :About 80% of
non-specific reaction of HTLV-1 antibodies had antibodies derived from
bovine protein in this study.
Indoleamine 2,3-dioxygenase 1 (IDO1) and arginase 1 (ARG1) are two
different enzymes that catabolize tryptophan and arginine, respectively.
Degradation of the amino acid tryptophan and arginine into downstream
metabolites (kynurenines and ornithine/urea, respectively) regulate both
innate and adaptive immunity over the short-term (mainly ARG1) and longterm (IDO1) prospective. As a result, altered activity of IDO1 and ARG1 are
often associated with pathology, i.e., neoplasia (where they would favor
tumor immune escape mechanisms), autoimmunity and chronic inflammation (where they are often defective). IDO1 is highly expressed in dendritic
cells (DCs) stimulated with the cytokine IFN-gamma (a T-helper 1 inducing cytokine) whereas ARG1 has the highest expression in macrophages
stimulated with IL-4 (Th2 inducing). All data accumulated so far seem thus
to indicate that the two immunosuppressive enzymes are at work in different cells and microenvironmental conditions. In the present study, we investigated whether IL-4 could also induce ARG1 in DCs and whether IDO1
could interfere (either potentiating or inhibiting) ARG1 expression. Results
showed that IL-4 represents a formidable inducer also in DCs and that lack
of IDO1 expression (evaluated by means of using DCs from Ido1 KO mice)
further increases ARG1 induction by the cytokine. Therefore, these data
would indicate, for the first time, that ARG1 expression also include DCs, in
which an important cross-talk appears to occur between ARG1 and IDO1.
In conclusion, a better understanding of the reciprocal influence between
arginine and tryptophan catabolism may pave the way to the development
of innovative therapeutic approaches in apparently very distinct disease
areas, such as neoplasia and autoimmunity.
Symposium
PB-20
Comparison of the risks of tuberculosis infection between Japanese
medical students and non-japanese international students
tuberculosis screening using a T cell interferon- Γ release assay
PB-21
Impact of a five-year measles-rubella vaccination catch-up campaign in Japan.
Remarkable increase in seroprevalence of measles-specific immunoglobulin G
among Japanese health care students.
Case Conference
Chiaki Suto 1, Toshiya Inoue1, Ai Takeichi1, Maika Sano1, Azusa Uchida1, Takao Kimura2,
Katsuhiko Tsunekawa2, Masami Murakami2
Ai Takeichi 1, Chiaki Suto1, Toshiya Inoue1, Maika Sano1, Azusa Uchida1, Takao Kimura2, Makoto Nara3,
Masami Murakami2
1 Clinical
Laboratory Center,Gunma University Hospital, Japan
2 Department
of Clinical Laboratory Medicine, Gunma University Graduate School of Medicine
1 Clinical
Laboratory Center, Gunma University Hosp, Japan
2 Department
of Clinical Laboratory Medicine, Gunma University Graduate School of Medicine
3 Infection
Control and Prevention Center, Gunma University Hosp.
Oral Presentation
Screening of medical students for tuberculosis (TB) at the time of admission is a key strategy to control and prevent the spread of infection on
university campus and teaching hospitals because of the high risk of exposure to TB patients. The Mycobacterium tuberculosis antigen-specific
interferon- release assays (IGRAs) are specific latent tuberculosis detection
methods used in such groups. In order to evaluate TB risk at the time of
admission in university campus and medical schools in Japan, a retrospective study was conducted. A total of 2377 students (1730 Japanese
students and 647 international students) were screened for TB using the
IGRAs at the time of admission. Only 0.3 % Japanese students were positive for IGRAs, and none were diagnosed with active TB at the follow-up. In
contrast, 9.1 % international students were positive for IGRAs, including 2
students diagnosed with active TB during follow up. Positive ratio of IGRAs
in international students was significantly higher than that of japanese
students at the time of admission. We propose a standard approach for TB
screening with IGRAs at the time of admission for medical students, especially international students in Japan.
Poster Presentation
In 2000, the United States declared that measles was eliminated from the
country. However, measles outbreak in 2014-2015 highlighted a growing
problem in the United States. In Japan, a measles epidemic occurred during the summer of 2007. In 2008, the Japanese government implemented
a 5-year measles – rubella vaccination catch-up campaign for individuals
aged 13 and 18 years old to eliminate measles until 2012. The present
study was intended to determine the impact of the 5-year vaccination campaign by monitoring the seroprevalence of measles-specific and rubellaspecific immunoglobulin (IgG) among Japanese healthcare students in
2007-2015. A total of 2613 Japanese healthcare students in the Gunma
University were serologically screened once a year at the time of admission. This study demonstrated that the vaccination campaign caused a
striking increase in the seroprevalence of measles-specific IgG, from 52.7%
in 2007 to 96.6% in 2013. Japanese government declared that measles
was eliminated from Japan in 2015. The seroprevalence remained ＞ 90%
during 2009 – 2015, except in 2014 (86.2%). On the other hand, the seroprevalence of rubella antibodies remained ＞ 90%, except in 2008 (85.6%).
In conclusion, the campaign raised the seroprevalence of measles-specific
IgG by ＞ 50% during the 2007 – 2015 study period. In contrast, the serprevalence of rubella-specific IgG did not significantly change during the
same period. Nonetheless, a substantial number of healthcare students
remain susceptible to vaccine-preventable infectious diseases. Because
their profession will involve frequent interactions with patients, including
immigrants and travelers, we propose a nationwide targeted immunization program for Japanese healthcare students using serological screening
prior to clinical training.
74
Novel base substitution in 5'UTR results in undetectable HCV
RNA quantification by CAP/CTM Real-time PCR.
PB-23
Haruna Hinohara 1, Kaneo Sato1, Makoto Osada2, Masumi Endo1, Mihoko Sakamoto1,
Norihiko Amemiya1, Katsue Suzuki-Inoue3
Yoko fukushima , Akemi onuki, Sachiko oya
Central Medicine inspection Laboratory, Japan
1 Division
of Laboratory Medicine, University of Yamanashi Hospital, Japan
2 School
of Medical Technology Faculty of Health Science, Gumma Paz College
3 Department
of laboratory and medicine, Faculty of Medicine, University of Yamanashi
Symposium
Kaori Ishihara 1, Tetsuya Usui1, Norihito Kaku2, Syouichi Fukui3, Souichirou Minami1, Hiroo Hasegawa2,
Atsushi Kawakami3, Katsunori Yanagihara2
The evaluation of fundamental performance of HISCL-5000
for the measurement of serum NT-proBNP.
Educational Lecture
PB-25
Special Lecture
Evaluation of serum MMP-3 in HTLV-1-positive patients with
rheumatoid arthritis
Invited Lecture
Introduction
Recently, the number of patients with I type allergic disease are increasing.
The specific IgE testing is a basic diagnostic method for I type allergic disease. However, some of specific IgE testing have to use many serum, it is
difficult to get serum from infants. In our laboratory, we utilize two analyzers for specific IgE, MAST3 and DiaPack3000. We can properly use these
methods depending on the situations. In DiaPack3000, its measurement
speed is 12minutes and the items are selectable on demand. In MAST3,it
can measure 33items at same time and it uses 200 μ L by a test, however
it takes about 6hours. We evaluated these analyzers, and compared each
performance.
Method
We researched the positive rate of each allergen in MAST3.We chose
5items in the order that had high positive rate, and evaluated a correlation between two methods. In addition, we evaluated quantity of serum,
between normal quantity and small quantity in DiaPack3000
Results
In MAST3, The positive rates were as follows: inhaled allergens: 1.4%58.1%, food allergens: 0.8%-14.7%, fruit allergens: 0.8%-1.7%. The correlations of the 5allergens that have highly positive rate ( Japanese cedar
pollen : T17, house dust mite : D2, cat dander : E1, egg white : F1, ovomucoid : F233) were 58.0 – 85.5% . We also evaluated serum quantity of DiaPack3000, The correlation coefficient between normal quantity and less
quantity samples were R=0.989-0.999.
Conclusion
We found it is important that the allergy testing methods can use properly
depending on the situations. We now understand purpose and characteristics of both testing method in specific IgE tests, and we are hopeful that
these methods will continue to be used in diagnosis and treatment.
Introduction: Accurate hepatitis C virus (HCV) RNA quantification is essential for the diagnosis and the management of chronic hepatitis C therapy. CAP/CTM HCV v1.0 and AccuGene m-HCV are highly sensitive assays
for HCV RNA quantification based on real-time polymerase chain reaction
(PCR) and widely used in many countries. Methods: Serum HCV RNA
quantification was performed by the Cobas AmpliPrep/Cobas TaqMan
HCV assay (CAP/CTM) (Roche Diagnostics, Inc) and Abbott RealTime HCV
assay AccuGene m-HCV (Abbott Japan Co.,Ltd). Two vectors expressing the
HCV 5' noncoding region between nt 68 and 307 were constructed, and
their sequences were identified by ABI-310.Results: We report a case of
58-year-old man with HCV genotype 2a. Serum HCV RNA was not detected
by CAP/CTM v1.0, although hepatitis C viremia was confirmed by the AccuGene m-HCV (5.71 logIU/ml) and two kinds of the HCV core antigen assay (Abbott: 2079.49 fmol/l, Ortho-Clinical Diagnostics, Inc: 2287.8 fmol/l).
We observed two substitutions at position 144 (T to C) and position 147
(T to C), which is the putative binding site of the TaqMan probe. Recently
CAP/CTM v2.0 with redesigned primers has been developed. HCV RNA
measured by this assay was 5.8 logIU/ml, which is consistent with that
measured by the AccuGene m-HCV. These findings suggest that CAP/CTM
v1.0 failed to detect HCV RNA in the patient due to the substitutions in the
binding site of the TaqMan probe. Discussion: Previous studies reported
that CAP/CTM HCV version1.0 failed to detect HCV RNA with substitutions
at positions 145, 158, 165 and 169. Here , we reported novel substitutions
at position 144 and 147. These substitutions probably causes mismatches
between the target sequences and the TaqMan probe. Clinicians need to be
aware that CAP/CTM HCV version1.0 occasionally fails to detect HCV RNA
genotype 2b and should consider alternative assays if there is discrepancy
between clinical findings and the laboratory result.
PB-24
Features and study on the use of Diapack3000 and MAST3
Keynote Speech
PB-22
Miki Zaima , Chiaki Iramina, Kazuki Isa, Megumi Yamauchi, Shiro Maeda
University of the Ryukyus Hospital, Japan
Poster Presentation
Introduction Matrix metalloproteinase-3 (MMP-3) is an enzyme, which
plays a role in the destruction of cartilage and bone in rheumatoid arthritis (RA). Several studies suggested serum MMP-3 reflects synovial inflammation. However, there was no study on serum MMP-3 levels in Human
T Lymphotropic Virus Type I (HTLV-1) antibody-positive patients with RA
(HTLV-1-pos-RA). In this study, we compared serum MMP-3 levels between
HTLV-1-pos-RA and HTLV-1 antibody-negative patients with RA (HTLV-1neg-RA), since serum C-reactive protein (CRP) levels and responses to the
treatment were different between two groups.MethodsTwenty patients
with RA (10 cases, HTLV-1-pos; 10 cases, HTLV-1-neg) who obtained the
informed consent about clinical research from 2009 June to 2015 June
were enrolled in this study. The serum MMP-3 levels, disease activity score
28 joints (DAS28)-CRP and DAS28 - ESR were evaluated at both pre- and
post-treatment.Results At the pre-treatment, the mean MMP-3 level in
HTLV-1-pos-RA were lower than that in HTLV-1-neg-RA (174.8 ng/ml ±
199 ng/ml vs. 234.4 ± 262, P=0.32). The mean CRP levels in HTLV-1-posRA were also lower than that in HTLV-1-neg-RA (1.15±0.94 mg/dl vs. 2.81
mg/dl ± 2.24, P=0.07). In HTLV-1-pos-RA, DAS 28 and MMP-3 levels at
post-treatment were significantly lower than those at pre-treatments (DAS28CRP, P<0.05; DAS28ESR, P<0.05; MMP-3, P<0.05). Although DAS28
at post-treatment were significantly lower than those at pre-treatment in
HTLV-1-neg-RA (DAS28CRP, P<0.05; DAS28ESR, P<0.05), there was no
significant change in MMP-3 level (P=0.06). In addition, there is no correlation between MMP-3 level and DAS28 in both HTLV-1-pos-RA and
HTLV-1-neg RA.Conclusion In HTLV-1-pos-RA, DAS28 as well as MMP3 were significantly decreased by the treatment. However, there were no
correlation between MMP-3 level and DAS28. Further study is needed to
determine the usefulness of serum MMP-3 in HTLV-1-pos-RA.
Oral Presentation
N-terminal pro B-type natriuretic peptide (NT-proBNP) is a useful biomarker for the diagnosis and/or management of chronic heart failure. Here, we
show the results of evaluation of a new instrument for serum NT-proBNP
assay, HISCL-5000(Sysmex). We performed fundamental evaluation of the
NT-proBNP assay using HISCL-5000, intra and inter-assay variance, linearity for the serial dilution samples and influence of several substances.
Coefficients of variance (CV) for intra and inter assays using control samples were between 0.9 and 2.4%.linearity was maintained in the range of
0-34,434pg/mL. There is no significant interference, less than 10%, by
existence of direct bilirubin, conjugated bilirubin, hemoglobin, chylous
fluids, and rheumatoid factor into the measurements of the NT-proBNP.
Furthermore, we evaluated a consistency of this assay with another electro-chemiluminescence immunoassay for NT-proBNP (ECLusys analyzer,
Roche Diagnostics KK, Tokyo, Japan) using 112 serum samples. Although
the absolute values for HISCL-5000 measurements tend to be lower compared with those for ECLusys measurements (the linear regression equation: y = 0.85x + 183.2), obtained absolute values were considered consistent with each other (correlation coefficient of 0.99).These results indicate
that HISCL-5000 can be applied for the serum NT-proBNP measurements
in routine use for the laboratory.
Case Conference
1 Dept.
of Laboratory Medicine, Nagasaki University Hosp. Japan
2 Laboratory
Medicine, Nagasaki University Graduate School of Biomedical Science, Nagasaki, Japan
3 Unit
of Translational Medicine, Department of Immunology and Rheumatology, Nagasaki University Graduate School of
Biomedical Sciences, Nagasaki, Nagasaki, Japan
75
Keynote Speech
PB-26
PB-27
Analytical Performance of the i-STAT cTnI assay
Detection of the anti Stress Granule antibody using U2OS
Yuki Fushimi 1, Yuko Okuda2, Mai Sakoya2, Daiki Kato2, Shinji Ujiie2, Masanori Sano2, Toshisuke Morita1
Takayuki MORIMOTO , Satoshi YAMASAKI, Yusuke YOSHIDA, Michiya YOKOZAKI, Eiji SUGIYAMA
1 Department
of Laboratory Medicine, Toho University School of Medicine, Japan
2 Dept.
of Clinical Laboratory, Toho Univ. Omori Medical Center
Hiroshima University Hospital, Japan
Invited Lecture
Special Lecture
Examination of autoantibody is important for making diagnosis or evaluating disease activity of autoimmune diseases. We succeeded in detecting
a novel autoantibody, anti stress granule antibody (anti SG ab). We used
U2OS cell, an osteosarcoma cell line, treated with sodium arsenite. Sodium arsenite is known to induce cytoplasmic formations of RNA granule
named stress granule by inducing oxidative stress in a cell. We applied an
indirect immunofluorescence technique by using antibody against eIF3
η , a marker protein for SG, to detect anti SG antibody in patient's sera.
Among 120 cases examined, which are randomly selected in our clinic, 17
samples(14.2%)were positive for anti SG antibody. None of those samples
were positive for anti cytoplasmic antibody by FLUORO HEPANA TEST or
U2OS cell without sodium arsenite treatment. The anti SG antibody positive cases include 6 cases of undifferentiated arthritis, 3 case of rheumatoid arthritis, 1 case of systemic lupus erythematosus, systemic sclerosis,
Sjögren's syndrome and others. All the cases except one had arthralgia as
a chief complaint when they visited our clinic. Among 17 cases positive
for anti SG antibody, rheumatoid factor or anti cyclic citrullinated peptide
antibody was positive only in 4 or 3 cases, respectively. It is of note that
anti SG antibody is positive in patients without these tests. Further studies
are required to prove a clinical relevance of this autoantiboy, however, it is
suggested that testing anti SG antibody contribute to making a diagnosis
of arthritis as an &uml;anti SG antibody positive arthritis” in undifferentiated arthritis cases.
Educational Lecture
The shortening of turn-around time (TAT) is always the problem, especially in emergency departments. Our laboratory requires 30 minutes to
report the result of cardiac troponin I (cTnI) from the arrival of the specimen. The i-STAT1 analyzer (Abbott Point-of-Care) can perform POC testing
of cTnI within 10 minutes. In this evaluation, we have checked analytical
performances such as its imprecision, inter-day CV(%), linearity, detection
limit, interferences, followed by comparison to the AIA900 (Tosoh).Imprecision was carried out by measuring three levels of specimens, pooled according to the cTnI concentration. The result of the total imprecision was
satisfactory. Furthermore, fifty of the randomly selected specimens tested
for two different lots, at concentrations ranging from 0.00 to 35.01 ng/
mL, showed no discordance. Inter-day CV(%) were carried out by measuring three levels of controls twice a day (10:00, 16:00). The total CV(%)
obtained for control 1 (0.24 – 0.44 ng/mL), control 2 (1.14 – 2.12 ng/mL),
and control 3 (11.98 – 31.58 ng/mL), were 6.8%, 3.6%, and 9.7%.The iSTAT cTnI assay demonstrated an upper limit of linearity at 47.5 ng/mL,
by serial dilutions of a high cTnI specimen. Limit of detection was at 0.03
ng/mL (mean ± 2.6SD of 10 replicates). These results were similar to
those of AIA900. The assay was not affected by common interferences.The
observed correlation coefficient between i-STAT and AIA900 was r=0.95,
y=1.509x+0.2. Out of 150 randomly selected patient specimens (Ethics
approval number 27-227), 1.3% were positive in AIA900, while the results
were negative in i-STAT.The i-STAT has the disadvantage of measuring
only one specimen at a time, and its running cost is four times higher than
that of AIA900. However, since i-STAT is portable and can perform rapid,
accurate tests, the usage of i-STAT would be suitable in emergency departments rather than central laboratories.
Symposium
PB-28
Comparison of PIVKA-II assay performances
CLEIA versus CLIA principles
PB-29
The comparison of highly sensitive HBsAg assay (HBsAg-HQ)
with HBVDNA assay
Case Conference
Eri Ishii , Tatsuo Satou, Hiroko Asanuma, Yasue Kurozumi, Ayako Ishikawa, Yumi Matsuo
Shingo Kujihashi 1, Hiroki Shirai1, Fumiko Ohshima1, Shigeki Ohnishi1, Yohji Urata2
Kurashiki Medical Center, Japan
1 Dept.
of Clinical Laboratory, Japanese Red Cross Kyoto Daiichi Hosp, Japan
2 Dept.
of Pathology, Japanese Red Cross Kyoto Daiichi Hosp.
Oral Presentation
Poster Presentation
We report here the analytical performances of recently developed Abbott Japan Architect PIVKA-II (hereinafter Architect) and EIDIA STACIA
PIVKA-II (hereinafter STACIA), the former of which basing on CLIA and the
latter of which basing on CLEIA principle. The study was performed upon
approval from ethics committee in our hospital. We evaluated the performances in terms of the following criteria and obtained the results as shown
below;1)Within-run reproducibilityWe evaluated the within-run reproducibility by running two levels of pooled sera ten times. The CV% of Architect
was in the range of 1.0-8.2%, while that of STACIA was in the range of 4.74.8%.2)Day-to-day reproducibilityWe evaluated the day-to-day reproducibility by running two levels of pooled sera ten days. The CV% of Architect
was in the range of 3.2-6.0%, while that of STACIA was in the range of 4.47.7%.3)Limit of detectionWe evaluated the limit of detection by calculating
the lowest concentration where mean-2SD is equal with blank+2SD. The
limit of detection of Architect was 4.0 mAU/mL,while that of STACIA was
5.0 mAU/mL.4)LinearityWe evaluated the linearity by measuring 2n dilution series with a high concentration sample and confirmed that the highest concentration where the linearity was retained was 27,700 mAU/mL
with Architect, while it was 33,000mAU/mL with STACIA.5)CorrelationWe
evaluated correlations of Architect, STACIA and EIDIA Picolumi PIVKAII MONO (hereinafterPicolumi) with 168 serum samples. The correlation
between Architect (Y-axis) and STACIA (X-axis) was y=0.938x-31.749 (
r=0.944), while that of Architect (Y-axis) and Picolumi (X-axis) and that of
STACIA (Y-axis) and Picolumi (X-axis) were y=0.759x+3.624 (r=0.941) and
y=0.825+32.599 (r=0.985),respectively. Here we evaluated the analytical
performances of Architect and STACIA and confirmed that the overall performances including reproducibility,sensitivity, linearity and correlation
were acceptable.
Introduction:HBVDNA assay has used as the standard method for detecting low viral loads, but the assay was too expensive and time-consuming.
Recently, a highly sensitive chemiluminescent enzyme immunoassay for
HBsAg (HBsAg-HQ®) was developed, which is as sensitive as HBVDNA assay. HBsAg-HQ is simple, less expensive, and expected to be a surrogate
marker of HBVDNA. In this study, we examined the relationship between
HBsAg-HQ and HBVDNA assay.Methods:Four hundred ninety-five serum
samples were collected from June 2014 to July 2014. HBVDNA assay was
measured by TaqMan PCR assay. HBsAg-HQ was measured by the twostep sandwich assay based on a chemiluminescent enzyme immunoassay.
In this study, negative of HBVDNA and HBsAg were defined as no detectable HBVDNA and HBsAg level <5.0 mIU/mL, respectively. We examined
concordance rate of HBsAg-HQ and HBVDNA assay. Moreover we studied
mismatch cases individually.Results:The concordance rate was 91.1%. In
44 mismatch cases, 28 have administrated nucleoside/nucleotide analogues (NAs), 7 were non-treated HBV carriers, 4 had immunosuppressive
therapy, 3 had under HBV reactivation, and the other 3 cases were treated
with antimetabolite, interferon, and antiviral drug for HIV infection.
Conclusion:Although 8.9% of cases showed mismatch results, HBsAg-HQ
was high concordance rate, and thought to be substituted for HBVDNA assay.
76
Significance of rapid assessment of aldosterone-renin ratio in
establishing a diagnosis of primary aldosteronism
PB-31
Chiaki Kobayashi , Yuko Nakanishi, Shinya Kato, Kazuya Murata, Kouji Murabayashi
Evaluation of free light chain assay in serum
Ai Okamoto , Yumi Taniguchi, Akiko Murakami, Takatsugu Honda, Nahoko Sumi, Mari Morimoto,
Tatsuya Nishimiya
Ise Red Cross Hospital, Japan
Department of Clinical Laboratory,Ehime University Hospital, Japan
Evaluation of the basic performance of BS-NIA IgG4
Introduction:IgG4-related disease (IgG4-RD) is a newly recognized disorder characterized by elevated serum IgG4 concentration and increased
IgG4-positive plasma cell infiltration in tissue. Since comprehensive
diagnostic criteria for IgG4-RD were proposed in 2011, the demand for
IgG4 level detection has increased. In this study, we evaluated the basic
performance of BS-NIA IgG4, which is widely used in Japan and also used
in the study for determining cutoff value of IgG4-RD diagnostic criteria.
Materials and Methods:IgG4 level was measured with a Siemens BN II
nephelometer using BS-NIA IgG4 (The Binding site). Precision was validated by ten replicate measurements for intra-day measurements and once a
day for 10 days for inter-day measurements, using pooled serum. In addition, linearity was determined by preparing serial dilutions of serum with
IgG4 concentration above the initial measuring range (0.13 -4.41 g/L) and
measuring each in triplicate. When IgG4 concentration is higher than 4.14
g/L, sample is automatically diluted and re-measured. The interferences of
conjugated and free bilirubin, hemoglobin, chyle, and rheumatoid factor
(RF) on IgG4 measurement were also evaluated using Interference Check
A Plus and Interference RF (Siemens). Results:The intra- and inter-day precisions, expressed as coefficient of variation (CV), ranged from 2.72% to
3.03% and 3.25% to 4.17%, respectively. Around 4.00 g/L IgG4, linearity
was poor. There was large discrepancy between initial and further dilution
measurement. However this discordance was not observed in measurements using two other lots of BS-NIA IgG4. The presence of conjugated
and free bilirubin, hemoglobin, and triglyceride did not influence the
measurement. Meanwhile, a 17% increase in IgG4 was observed at RF concentration of 500 IU/mL.Conclusion:Overall, the performance of BS-NIA
IgG4 for routine measurements was satisfactory, provided that its linearity
checked prior to use.
77
Poster Presentation
Background: Anti-phospholipid antibodies (aPLs) are heterogeneous
group of autoantibodies that appear in a variety of autoimmune diseases,
particularly systemic lupus erythematosus (SLE). The presence of aPLs is
associated with clinical events such as arterial and/or venous thrombosis
and recurrent fetal loss. We recently reported that the clinical picture
of SLE apparently depends on subclasses of aPLs in the patient’s sera,
but the contribution of each subclass remains uncertain.Methods: We
developed an up-to-date enzyme immunoassay (EIA) system using the
AcuStar®automated analyzer for parallel detection of seven subclasses
of aPLs: anti-cardiolipin antibodies (aCL) and anti- β 2-glycoprotein I antibodies (a β 2GPI), each of IgG, IgM, and IgA classes, plus newly introduced
anti- β 2-glycoprotein I-domain 1 antibodies (aDomain1) of IgG class.
They were measured in 276 normal healthy volunteers and 138 patients
with SLE: 29 patients wish arterial thrombosis, 16 with venous thrombosis, and 93 patients without thrombotic complications. aCL/ β 2 GPI was
measured with a standard ELISA kit commonly used in Japan. Results:
Multivariate logistic regression analysis revealed that the presence of
IgG-aDomain1 was most closely associated with arterial thromboembolic
complications. In contrast, the presence of IgG-a β 2GPI was most closely
related to venous thromboembolic complications. When the results of the
138 SLE patients measured by this EIA system were compared with those
measured by the most commonly used standard ELISA kit for aCL/ β 2GPI
in Japan, ROC analysis revealed that the accuracy of predicting thrombotic
complications based on the test results of the multiplex EIA system was
higher than that with the aCL/ β 2GPI-ELISA. Furthermore, both IgG-aDomain1 and IgG-a β 2GPI were independently associated with the presence
of lupus anticoagulant activity. Conclusions: Anti-phospholipid syndrome
in patients with SLE can be classified by antigenic specificities of their
aPLs as to susceptibility to either arterial or venous thromboembolic complications.
Oral Presentation
Yoko Usami 1, Nau Ishimine2, Kazuhiro Nagata2, Kenji Kawasaki2, Mitsutoshi Sugano2, Takayuki Honda2
1 Shinshu
University School of Medicine , Japan
2 Shinshu
University Hospital
Case Conference
Kazusa HARA 1, Yukari MOTOKI1, Toshiyuki SAKATA2, Junzo NOJIMA1
1 Department
of Laboratory Science, Faculty of Health Science Yamaguchi University Graduate School of Medicine, Japan
2 I.L.
Japan Co.,Ltd.
Symposium
PB-33
Educational Lecture
Novel Enzyme Immunoassay System for Simultaneous
Detection of Seven Subclass of Anti-Phosoholipid Antibodies
For Differential Diagnosis of Anti-Phospholipid Syndrome
Special Lecture
PB-32
Introduction: Monoclonal immunoglobulin free light chain(FLC) are
important tumor markers often present in serum of patients with plasmaproliferative disease disorders.Serum FLC were measured with two
commercial reagent kits.Objectives were to evaluate monoclonal antibodybased nephelometric N Latex FLC(Siemens).We assessd the analytical performance of the N Latex FLC assays and compared it with the
FreeliteTM(Binding Site) assays. Methods: Analytical precision was assessed
using control,linearity on FLC control and samples from monoclonal elevations of FLC.We demonstrated lot-to-lot consistency with variation of plural N Latex FLC reagent and calibrator.Serum samples submitted for routine FLC analysis(31 samples) were analyzed by both methods (FreeliteTM
and N Latex FLC) according to the manufacturers’ instructions,and in addition performed by Immunofixation(IF). Results: The intra- and inter assay
CVs of N Latex FLC κ and λ assays were 3.15-6.72%.The linearity of assay were excellent.The slopes of lot-to-lot consistensy with variation were
0.89-0.96 for κ and 1.02-1.17 for λ (n=13).For method comparison,the
slopes of regression line were 1.43(FLC- κ ),0.829(FLC- λ ) and 2.18(FLCκ / λ ratio),furthermore the between method variances were 9.7%(FLCκ ),38.7%(FLC- λ ) and 16.1%(FLC- κ / λ ratio) when comparing with
the FreeliteTM .Good agreement was observed for the κ / λ ratio(83.9%).
However,N Latex FLC assay was higher agreement rate of κ / λ ratio
against IF than Freelite TM (77.4% vs 67.7%). Conclusions: The N Latex FLC
assay good data performance,also good concordance and correlation in
comparison with two FLC methods.We want to present a usuful clinical
case of FLC analysis.
Invited Lecture
Introduction: Primary aldosteronism (PA) is known as a typical disease
for secondary hypertension and PA screening by Aldosterone Renin Ratio
(ARR) is recommend for hypertensive patients. ARR is calculated by measuring plasma aldosterone concentration (PAC) and active renin concentration (ARC) or plasma renin activity (PRA) simultaneously. PA measurement
by commercial labs requires 3-5 days due to radioimmunoassay (RIA).
Rapid PAC and ARC assays to be measured in approximately 10 minutes
have been recently developed on the fully automated immunoassay system, Accuraseed. We have evaluated the Accuraseed Aldosterone and Accuraseed renin(ARC) (Wako Pure Chemical Industries, Ltd.) in this study.
Subjects and Methods: PAC and ARC for PA diagnosis were measured in
138 subjects with possible secondary hypertension. The PAC and ARC assays were measured by using the Accuraseed and RIA assays. Results: In
terms of PAC and ARC, the results of assays done by Accuraseed showed
good correlation with those of the RIA assay. (PAC:y=0.846x+26.44 ,
r=0.88 , ARC v.s. PRA:y=12.95x-12.81 r=0.86) Compared with determination of PA screening, 8 outliers were detected and 2 were patients for
Captopril challenge test. Captopril challenge testing was performed 25
cases, and 8 were positive. Computed tomography revealed no adrenal
masses in the 8 cases, but 2 were diagnosed with PA after adrenal venous
sampling (AVS). Conclusion:ARR is important for definitive diagnosis of PA
based on the results of AVS because computed tomography cannot detect
small cancer smaller than 6 mm in diameter. ARR is strongly dependent
on ARC level because ARR is calculated by ARC as a factor of denominator.
Accuraseed renin and Accuraseed aldosterone were enabled to calculate
the ARR value with rapidity and high sensitivity. We concluded that those
rapid assays were useful for the PA screening.
Keynote Speech
PB-30
Keynote Speech
PB-34
Changes of M2BPGi levels in chronic hepatitis C patients
treated with interferon-based or interferon-free therapy
PB-35
The examination of ALP isozyme anomaly case profile in our
hospital
Hidekazu ISHIDA 1, Atsushi SUETSUGU2, Yuriko KATANO3, Rina TAUCHI3, Yukari OMORI3,
Nobuyuki FURUTA3, Hiroyasu ITO3, Mitsuru SEISHIMA3
Yachiyo Endo 1, Masanori Seimiya2, Mariko Watanabe1, Haruna Asano1, Toshihiko Yoshida1,
Sawabe Yuji1
1 Gifu
University Hospital, Japan
2 Division
of Gastroenterology/Internal Medicine, Gifu University Hospital
3 Division
of Clinical Laboratory, Gifu University Hospital
1 Chiba
University Hospital, Japan
2 International
University Of Hearth And Welfare
Invited Lecture
Special Lecture
Educational Lecture
The ALP isozyme profile provides an important clue to the diagnosis of
pathological conditions. However, an irregular electrophoretic pattern
(anomaly) may be accepted due to immunoglobulin-binding or a change of
transient carbohydrate chain structure. We examined ALP anomaly cases
in our hospital. Sera were from patients who visited our hospital. Written
informed consent was obtained from all participants prior to blood sampling, and the study protocol was approved by the Ethics Committee of
Chiba University Graduate School of Medicine. ALP activity was measured
with an automated biochemical analyzer equipped with the reagents used
in the JSCC-recommended method. ALP isozymes were analyzed using
Epalyzer II (Helena Laboratories). The immunoglobulin-binding ALP cases
were identified by countercurrent immunoelectrophoresis or immunofixation. Between 2007 and 2015, 4,055 patients developed ALP isozyme. A
total of 63 cases (1.6%) revealed an anomaly pattern. The most common
cause was transient hyperphosphatasemia in pediatric patients (THP, 43
cases). ALP electrophoresis revealed a pattern (a band in the fast a-2 region) characteristic of this condition. The other causes of anomaly cases
were immunoglobulin-binding ALP (10 cases), and others (10 cases). A
commonly suggested but unproven mechanism for THP or immunoglobulin-binding ALP is impaired clearance of ALP from the circulation. Once
patients have been diagnosed with isozyme profile, they do not need to undergo further evaluation, but may be followed-up regularly. It is particularly meaningful for children to avoid undergoing additional tests. Therefore,
the cause of anomaly isozyme profile should be diagnosed. In addition, if
the results of ALP isozyme analysis raise a suspicion of this condition, the
clinical laboratory scientist should provide the attending physician with
information and appropriate comments when reporting test results.
Aims and Background:It is well known that Hepatitis C virus (HCV) elimination improves liver fibrosis in chronic hepatitis C patients. Recently, the
direct-acting antiviral agents (DAAs) therapy has become a first choice for
the patients with HCV infection instead of interferon (IFN)-based therapy.
We report here the changes of the M2BPGi levels in chronic hepatitis C
patients treated with IFN-based therapy or DAAs therapies.Methods:We
measured serum M2BPGi levels in 64 chronic hepatitis C patients who
underwent at Gifu University Hospital before treatment, 2 and 4 weeks
after treatment, the end of treatment, and 12 or 24 weeks after the end of
treatment. Among 48 patients with genotype 1 HCV infection, 7 patients
were treated with pegylated-interferon (PEG-IFN), Ribavirin (RBV) and
Simeprevir (SMV) for 24 weeks, 30 patients were treated with Asunaprevir
(ASV) and Daclatasvir (DSV) for 24 weeks, and 11 patients with Sofosbuvir
(SOF) and Ledipasvir (LDV) for 12 weeks. Sixteen patients with genotype
2 HCV infection were treated with SOF and RBV for 12 weeks.Results:In
the patients treated with IFN-based therapy, M2BPGi levels during the
treatment were significantly higher than those before the treatment (P ＜
0.05), but M2BPGi levels at 24 weeks after the end of treatment were significantly lower compared with those of 4 weeks after treatment and the
end of treatment (P ＜ 0.05). In the patients treated with DAAs, including
ASV + DSV, SOF + LDV and SOF + RBV, M2BPGi levels at 2 and 4 weeks after treatment were decreased compared with those before treatment (P ＜
0.05). In the patients treated with any therapies, M2BPGi levels were positively correlated with serum ALT levels. Conclusion:We conclude that the
changes of M2BPGi levels by the treatment are associated with its degree
of liver injury and fibrosis.
Symposium
PB-36
The examination of LD isozyme anomaly case profile in our
hospital
PB-37
Mariko Watanabe 1, Seimiya Masanori2, Yachiyo Endo1, Haruna Asano1, Toshihiko Yoshida1,
Yuji Sawabe1
Identification of association of squamous cell carcinoma
antigen with IgA
Eriko Mori , Makoto Kurano, Akiko Tobita, Hironori Shimosaka, Shigeo Okubo, Yutaka Yatomi
University of Tokyo Hospital, Japan
Case Conference
1 Chiba
University Hospital, Japan
2 International
University of Health and Welfare
Oral Presentation
Poster Presentation
Background:Squamous cell carcinoma antigen (SCCA) is used as a tumor
marker for squamous cell carcinoma in uterine cervix, lung, pharynx, and
oral cavity. We examined SCCA levels with two different assays (chemiluminescent immunoassay utilizing ARCHITECTi2000 and fluorescent
enzyme immunoassay using AIA2000) and found that the SCCA values
were greatly deviated from each other in three of 123 cases. In this study,
we aimed to elucidate the mechanisms for the deviations. Methods and
Results: (1) To evaluate the reason for the difference in the SCCA level
between the two immunoassays, we conducted polyethylene glycol (PEG)
precipitation of these three deviated samples and found that the recovery rates after the treatment with PEG were extremely low, suggesting
the existence of larger molecular weight SCCA. (2) We next examined
the molecular weight of SCCA using size exclusion chromatography and
observed that SCCA was eluted not only in the original molecular weight
position but also in the fraction of a larger size than the IgG peak. (3) We
performed a spike recovery test and found that the recovery rates of these
three samples were low, suggesting the possible presence of autoantibody
against SCCA. (4) In all of the three cases, the absorption test with antihuman IgG antibody decreased SCCA values, while in one case, the absorption test with anti-human IgA antibody also decreased SCCA value. In the
latter case, the recovery rate was decreased to a greater degree when the
serum was incubated with both antibodies than when incubated with each
antibody alone. Conclusion:These results indicate the existence of large
molecular weight SCCA complexed with IgG (2 cases) and IgG plus IgA (1
case) in the sera showing the data deviation between the two immunoassays. To our knowledge, this is the first report for SCCA coupled with IgA.
In the analysis of LD isozymes, abnormal fractions may be detected at
unusual positions, which are called anomalies. In our hospital, we have
searched for causes of such anomalies wherever possible. In the present
study, we analyzed the trend of LD isozyme anomalies found in our hospital. Serum samples were collected from 4,691 patients for whom the
measurement of LD isozyme was requested to our clinical laboratory between April 2005 and September 2015. For the analysis of LD isozymes,
Epalyzer and dedicated reagents were used. For anomalies, LD isoenzymes
in hemolysates of patients erythrocytes were analyzed to examine gene
mutations. For cases without abnormal electrophoretic image with the
hemolysates, countercurrent immunoelectrophoresis or immunofixation
were conducted. In the past decade, 36 LD isozyme anomalies were detected, which accounted for 0.8% of all tested cases. They included genetic
abnormalities (3 cases), enzyme-linked immunoglobulin (14 cases), LD6
(ALDH) (1 case), and unknown cause (18 cases). Total activities were
abnormally high in all the 14 cases with immunoglobulin binding identified. This may have been caused by the extended turnover of plasma LD
through binding between LD and immunoglobulin. The 18 cases of unknown cause may have been due to binding to plasma components, such
as immunoglobulin, because anomalies occurred during and after their
release into the blood. In such cases, abnormally high LD values may not
reflect tissue injuries. Therefore, clinical laboratory scientist should clearly
report phenomena to facilitate their understanding. Although it is laborious to search for causes of anomalies, they should be identified for accurate diagnosis wherever possible.
78
A case report of retroperitoneal germ cell tumor with
chronic hepatitis C
an abnormal AFP fractionation was useful for diagnosis
PB-39
Kanae Maruyama , Masamune Aihara, Kazuhito Goto, Motoko Yamanaka, Miyuki Sakemoto,
Taeko Hotta, Dongchon Kang
Defective clot retraction by Bence Jones protein in a patient
with Multiple Myeloma
Yasushi Ueyanagi, Yoshimasa Aoki, Keiko Fujino, Shinya Matsumoto, Taeko Hotta, Dongchon Kang
Department of Clinical Chemistry and Laboratory Medicine, Kyushu University Hospital, Fukuoka, Japan
Department of Clinical Chemistry and Laboratory Medicine Kyushu University Hospital, Japan
Invited Lecture
Special Lecture
A patient with multiple myeloma has M-protein, monoclonal immunoglobulin made by increased malignant plasma cells. Bloods from patients
with multiple myeloma are known to show a defective clot retraction
though infrequently. These defective clot retractions are caused by Mprotein. The defective clot retraction is due to a lack of fibrin monomer
polymerization, but its mechanism remains to be fully elucidated. Here we
report a multiple myeloma patient with an impaired clot retraction, whose
M-protein is lambda type Bence Jones protein (BJP). In the most cases, Mprotein which impairs the clot retraction is intact immunoglobulin like IgG
and IgM. Thus it is extremely rare that the clot retraction is caused by BJP
consisting of only a light chain dimer. In the current study, the patient’s
BJP was composed of not only a light chain dimer type but also an unusual
high molecular weight type. The purified patient’s BJP inhibited thrombininduced fibrin polymerization, indicating that the BJP inhibited the fibrin
fiber formation. BJP usually does not work as antibody because it is not
intact immunoglobulin. The patient’s BJP also may inhibit fibrin fiber formation through a reaction other than antigen-antibody one although yet
confirmed. To our knowledge, this is the first case that BJP with a structural anomaly causes a defective clot retraction.
Most germ cell cancer occurs inside the gonads, however five % of malignant germ cell tumors appear outside their organs. Because extragonadal
germ cell tumors (EGGCTs) are found anywhere on the midline such as
mediastinum and retroperitoneum, the origin of this tumor remains controversial. EGGCTs are often seen between childhood and young adult;
an elderly patient with EGGCT is rarely met. Here we report a case that
an abnormal AFP fractionation pattern was helpful for early diagnosis
of retroperitoneal germ cell tumor.An elderly man with chronic hepatitis
C showed an intraperitoneal tumorous lesion on computed tomography
and thus hepatocellular carcinoma suspected. A serological test revealed
elevated total alpha-fetoprotein (AFP) level and AFP-L3 %. The latter is
the proportion of glycosylated AFP based on the lectin-affinity. Noticeably
the fractionation pattern of AFP of this patient was abnormal, suggesting
a diversity of lectin-affinity of AFP in germ cell tumors. This patient also
showed an atypical increase in beta human chorionic gonadotropin ( β
hCG). We propose the measurement of β hCG for early differential diagnosis of retroperitoneal germ cell tumor and hepatocellular carcinoma in
the case of detecting abnormal fractionation of AFP.
Keynote Speech
PB-38
Educational Lecture
Discrepancy between serum agarose gel electrophoresis and
immunofixation pattern
An analysis of a case of M-proteinemia
PC-01
Plasma Cell Leukemia
A Retrospective Analysis of 14 Cases from a Single Institute
in Taiwan
Plasma cell leukemia (PCL) is a rare and aggressive. It may present at the
time of diagnosis (primary PCL) or evolve as a late feature of plasma cell
myeloma (secondary PCL). PCL has not been reported from Taiwan yet.
We retrospectively searched for PCL in our institute from 2002 to 2015
and identified 14 cases including 8 cases of primary and 6 cases of secondary PCL, with the frequency of leukemic change at 2.5% (6/240) among
patients with plasma cell myeloma during this period. The 14 patients
were mostly males with a M:F ratio of 6:1. The median age was 70 (range,
43-90). Nearly all patients had anemia (100%; 14/14), thrombocytopenia
(100%; 14/14), and impaired renal function 93%; 13/14). We used threecolor flow cytometric analysis for immunophenotype of the neoplastic
plasma cells in PB. The CD45 non- or dim-expressors were gated. All cases
expressed surface (13/13) and cytoplasmic (14/14) CD38, and cytoplasmic CD138 (14/14). All cases were monotypic for light chain expression
with kappa light chain restriction more common (71% or 10/14). The only
immunophenotypic difference between primary and secondary PCL was
consistent CD56 expression in the secondary cases (50% or 4/8 in primary
vs. 100% or 6/6 in secondary cases; p= 0.04, Chi-square). Follow-up data
showed that all six patients with secondary PCL passed away in one month
after leukemic change. Of the 8 cases with primary PCL, two were alive (1
and 10 months, respectively) and the remaining 6 died of disease within
1 month (4 patients), 4 months (1), and 8 months (1), respectively. In conclusion, we presented the clinical and immunophenotypic features of PCL
in Taiwan. As compared to secondary PCL, primary cases seem to carry a
higher rate of CD56 expression and a better prognosis. A larger national
wide study is warranted.
79
Poster Presentation
The number of the M protein bands detected on immunofixation electrophoresis (IFE) generally shows high concordance with the number of "Mpeaks" on serum protein agarose gel electrophoresis (AGE). We herein
report a case of M-proteinemia which showed a different number of Mprotein bands between IFE and AGE and investigated the cause of this
discrepancy. The case was a 74-year-old woman diagnosed with solitary
plasmacyotma of the thoracic vertebra with mild hypercalcemia. A serum
analysis showed two "M-peak" bands in the Γ -globulin region on AGE
but only one M-protein (IgG- Λ ) band on IFE. All of the electrophoretic
analyses were performed using an Epalyzer 2TM automated system (Helena Laboratories, Tokyo, Japan) with either an AGE gel kit (Quick GelTM:
1% agarose gel with CAPSO buffer, pH 10.2; Helena Laboratories, Tokyo,
Japan) or an IFE gel kit (Quick Gel IFETM; 1% agarose gel with tris buffer,
pH 9.0, Helena Laboratories, Tokyo, Japan). To determine the cause of the
discrepancy, immunofixation electrophoresis using an AGE gel, instead
of an IFE gel, was performed. The findings of the fixation revealed the
presence of two IgG- Λ monoclonal proteins, which were consistent with
the AGE pattern. The pretreatment of the sera with neuraminidase, dithiothreitol (DTT), or 2-mercaptoethanol (2ME) did not affect the findings for
IFE. Based on these results, we concluded that the discrepancy was caused
by an interaction between the isoelectric point of the M-proteins and the
pH of the buffer. The AGE and IFE methods utilize the same gel material
(1% agarose) but different buffers, which may have caused the observed
discrepancies in the findings.
Oral Presentation
Yen-Chuan Hsieh1,2, Shih-Sung Chuang1, Ming-Chun Lee2
1 Department
of Pathology, Chi-Mei Medical Center, Taiwan
2 Department
of Cosmetic Applications & Management, Far East University
Case Conference
Tsunaki Sekita 1, Kazumi Kaihara1, Kyoko Komatsu1, Konosuke Nakayama1, Kazunori Miyake2
1 Japanese Foundation for Cancer Institute Ariake Hospital, Japan
2 Juntendo University,School of Medicine
Symposium
PB-40
Keynote Speech
PC-02
Leukemic Phase of Follicular Lymphoma
A Retrospective Study from a Single Institute in Taiwan
PC-03
How much sample is enough for complete blood count ?
Evaluation of the accuracy of complete blood count for
insufficient blood samples
Invited Lecture
Special Lecture
Educational Lecture
Po-Wen Cheng , Shih-Sung Chuang
Lin-Lin Pan, Chuang Fang-YI, Lee Chih-Yi, Shin Chia-Hsin
Department of Pathology, Chi-Mei Medical Center, Taiwan
Department of Laboratory Medicine Chung Gang Memorial Hospital, Chia-Yi ,Taiwan
Follicular lymphoma (FL) accounts for around 22% of lymphoma cases
in the West, while the frequency is much lower in the East. In Taiwan,
although the incidence is increasing in recent years, the frequency of FL
among all non-Hodgkin lymphoma is around 14%. FL usually involves
nodal tissue, leukemic change is rare and has not been reported from Taiwan yet. We retrospectively investigated leukemic FL in our institute from
January 2000 to December 2015. During this period a total of 168 cases
of FL was diagnosed, among them 10 (6.0%) cases experienced leukemic
change, either at presentation (6 cases) or at disease course (1, 3, 20, or 26
months after initial diagnosis). The 10 patients included four males and six
females with a median age of 47.5 (range, 30-75). The nodal lymphomas
were low-grade in eight cases (seven grade 1 and one grade 2) and highgrade in 2 cases (both grade 3A). Flow cytometric immunophenotyping
showed that the leukemic cells expressed CD10 (2/10, 20%), CD19 (10/10,
100%), CD23 (3/8, 33%), CD43 (1/7, 14%) CD5 (0/10, 0%). The patients
were treated with chemotherapy. Follow-up data showed that six patients
were alive including were were free of disease (median 51 months; range,
28-111) and one was alive with disease (19 months). The remaining 4
patients died of disease at 5, 23, 55, and 61 months, respectively, after
leukemic change. In this study, we reported the clinicopathological and
immunophenotypical features of FL with leukemic change in Taiwan. The
course could be aggressive, but a significant proportion of the patients
obtained complete response with chemotherapy. Studies on larger number
of patients are warranted for a better understanding of the impact of leukemic change in patients with FL.
Background:
Standard blood sample volume cannot usually be obtained for complete
blood count (CBC). This study aimed at investigating the impact of diminished blood sample volumes on accuracy of CBC analysis.
Methods:
Between August and December 2010, 332 blood samples from 332 subjects at a single medical institute were divided into nine groups according
to clinical settings, including new employees (n=39), individuals for physical check-up (n=39), patients scheduled for hospitalization (n=30), liver
transplant patients (n=48), patients undergoing hemodialysis
(n=39), patients in emergency department (n=35), adult inpatients (n=40),
adult outpatients (n=34), and pediatric outpatients (n=28). Ten milliliters
of peripheral blood sample from each subject was aliquoted into 3, 1, 0.6,
and 0.3 mL, each placed inside a separate K2 ethylenediaminetetraacetic
acid (EDTA)-containing collection tube. Parameters of CBC obtained from
1, 0.6, and 0.3 mL samples were compared with those acquired with 3 mL
sample that served as standards. Percentage bias for each parameter was
compared with the
criteria for within-subject biological variation (CVw).
Results:
Significant variations among samples from different clinical settings did
not notably interfere with data interpretation from small-quantity blood
samples. Percentage bias for most CBC indices significantly increased with
decreases in sample volume. White count was particularly unaffected by
diminished blood sample volume, whereas hematocrit (Hct) and
mean corpuscular volume (MCV) were vulnerable to sample volume reduction. Furthermore, decreases in median percentage biases with reduced
sample volumes were only observed in platelet count but did not affect
validity of data interpretation.
Conclusions:
Small volumes of blood samples conventionally considered insufficient
may
produce valid data for interpretation for most CBC indices. Dilution effect
occurred only in platelet count but did not affect data validity. The findings, therefore, may validate clinical use of small-quantity sample for CBC
analysis.
Symposium
PC-04
Cell Yield of Cerebrospinal Fluid Cell Count using Cytospin-3
and Cytopro-7620
Cell Yield of CSF Cell Count using Cytospin-3 and Cytopro-7620
PC-05
Reference interval of mean platelet volume and platelet mass
index in neonate from Taiwan
mean platelet volume, platelet mass index
Case Conference
Bon-Kyung Koo
Su-Mei Lin1, Yu-Hsuan Chien2
Laboratory Medicine, Samsung Medical Center, Korea
1 Department
of Pathology and Laboratory Medicine, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan
2 Department
of Pediatrics, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan
Oral Presentation
Poster Presentation
Background: The Cells are concentrated approximately 20-fold by cytocentrifugation. Even hypocellular samples with a hematocytometer cell
count of 0 cell/mcL can have a yield of approximately 35 cells on slide.
This study evaluated the nucleated cell number for cells recovered on slide
by using Cytospin-3 (Thermo Shandon Ltd., UK) and Cytopro-7620 (Wescor
Inc., USA) cytocentrifuges to hematocytometer cell count of 0-5 WBCs/
mcL of hematocytometer in the cerebrospinal fluid cell count. Next, this
study aimed to examine whether the cell yield was useful for slide preparation and retest criteria.
Methods: 148 samples of 0-5 WBCs/mcL on hematocytometer, were cytocentrifuged by Cytospin-3 and Cytopro-7620 instruments. The nucleated
cell number for cells recovered on slide was counted after Wright stain.
Results: The nucleated cell number for cells recovered on slide was 0-40
cells in the 44 samples of 0 WBC/mcL, and 3-95 cells in the 31 samples
of 1 WBC/mcL. It was observed that the nucleated cell number for cells
recovered on slide was 19-100 cells in the 42 samples of 2 WBCs/mcL,
and more than 100 cells in the 26 samples of 3-5 WBCs/mcL, respectively.
In addition, extremely normal lymphocyte, monocyte and polymorphonuclear neutrophil were observed in the 143 samples of 0-5 WBCs/mcL.
Macrophage and eosinophil were also rarely observed.
Conclusion: The nucleated cell number for cells recovered on slide was 20
cells, which were regarded as 1 WBC/mcL in body fluid cell count. However, in this study, we made alterations to report nucleated cell percentage
as 0% without preparing the cytocentrifuged slide at 0 WBC/mcL by using
the cell yield in a comparison between the value of 0-5 WBCs/mcL and
nucleated cell number for cells recovered on slide.
BACKGROUND: Normal Reference interval (RI) of mean platelet volume
(MPV) and platelet mass index (PMI) show variation from region to region
and between different ethnic groups. This descriptive cross sectional study
aimed at establishing the RI of MPV and PMI for preterm and term infants
from Taiwan.
METHODS: The study was conducted at our hospital from May 2015
to February 2016. We collected 205 neonates on post-natal day 1. A 0.5mL blood sample was collected in tube containing K3EDTA anticoagulant
and a complete blood count including MPV was performed by Sysmex XE5000 analyzer. Preterm group (gestational age (GA) < 36 weeks, n=81)
and term group (GA ＞ 37 weeks, n=124) were divided. We collected the
demographic data, including GA, birth body weight (BBW), and platelet
count. We analyzed the MPV and PMI between two groups.
RESULTS: The GA was 38.6 ± 1.1 weeks in term group and 33.4 ± 2.8
weeks in preterm group. The BBW was 3053.7 ± 439.5 gm in term group
and 2112.8 ± 579.5 gm in preterm group. The platelet count was 249 ±
51 x 109/L in term group and 244 ± 65 x 109/L in preterm group. The
RI of MPV and PMI in newborn are 9.8 ± 0.6 fL and 2474.7 ± 524.4,
respectively. There is a significant higher MPV in preterm group than
in term group (10.0 ± 0.6 fL and 9.7 ± 0.6 fL, respectively, p = 0.002).
However, there is no significant difference in PMI (2425.7 ± 477.8 and
2471.6 ± 552.9, respectively).
CONCLUSION: Our results provide RI for MPV and PMI in newborns
from Taiwan. There is a higher MPV in preterm then term group but no
difference about PMI between two groups.
80
Defining automated ESR analyser daily quality control
procedure by retained patient sample
QC setting procedure.
PC-07
Production of High Quality Specimen for Hematologic Diseases on Bone Marrow Examination
Production of Cytospin Slid use Clot Specimen for Hematologic Diseases on Bone Marrow
Aspiration
The erythrocyte sedimentation rate (ESR) is a non-specific marker of inflammation.
There is no adequate procedures for quality control monitoring because
the ESR measure a physical phenomenon that depends on many variables.
Therefore, our lab plan to establish control procedure using retained patient specimens. We use Roller 20PN automated analyzers (Alifax, Italy)
which capable of analyzing EDTA blood samples and reporting ESR result
in 20 seconds. First, twenty-two specimens were selected with ESR range
between 2 to 63 mm/hr, and then analyzed at 0, 2, 4, 8, 12, 24hrs after
sampling. The coefficient of determination and slope were calculated for
each time-series. We expected that retained patient samples can be used
as control materiel due to the ESR results remained stable within 24 hrs (r2
＞ 0.94, slope 0.99-1.12). Furthermore, ninety-five were obtained and analyze ESR at 0, 24hrs. The quadratic regression curve between was calculated (y = -0.002x2 + 1.0617x -0.932, r2=0.9297). Besides, we calculated
the 95% confidence intervals for intercept, primary constant and secondary constant from the formula above mentioned. The limits of agreement
for daily monitoring analytical system was based on calculation of the ESR
result at 0 hour. In conclusion, we had established appropriate quality assurance procedure by using retained sample as quality control material.
During medical procedures to acquire bone marrow, bone marrow is first
aspirated and then subsequently a biopsy is performed. However, during
bone aspiration, the syringe barrel is pulled strongly pulled to acquire a
robust sample often resulting in distortion of the bone architecture. Therefore, Fritsma et al. recommend that to conduct a precise assessment of tissue integrity, an initial biopsy must be performed. During this procedure,
mixture of sinusoidal blood is called 'dilution' which results in a diluted
sample rendering assessment of bone marrow cells difficult. According to
the accumulated literature, an aspirate volume of 1.0-1.5mL is considered
optimal and any increased amount leads to dilution. When a aspirate sample is diluted, it is difficult to examine cellular morphology and perform a
differential count of 300-1000(average 500) cells. However, base on our
clinical experience, a sudden pull of the syringe piston corresponding to a
three-fold more volume results in acquisition of 2.7mL of aspirate. Rapid
storage of the aspirate in a K3 EDTA tube results in prevention of dilution.
This diluted sample can now be used for pressure smear slide. In case of
failed aspiration, a touch print slide is an possible alternative. In addition,
a biopsy paraffin section slide stained with Wright's solution also can be
used for diagnosis. Furthermore, a coagulated aspirate sample can be used
to make a cytospin slide and then stained with Wright's solution for simple
diagnosis.
Keywords: Bone marrow, Aspiration, Clot, Cytospin, Wright stain
Special Lecture
Sangmuk Park
Biomedical Laboratory Science, Dongkang College University, Korea
Invited Lecture
MEI-CHIH Chen
Department of Laboratory Medicine, Taipei Tzu Chi hospital, Buddhist Tzu Chi medical Foundation, Taiwan
Keynote Speech
PC-06
Educational Lecture
Myeloperoxidase Isozyme Expression in Cases of Chronic Myelogenous Leukemic Cells
The Pattern of Myeloperoxidase Isozyme Expression in Chronic Myelocytic Leukemic
cell
PC-09
Hemostatic Potential of trunk extract of Gliricidia
sepium(Jacq.) Kunth ex Walp.
Rexy Cordial Alvarez1, Oliver Shane R. Dumaoal2
1 Department
of Laboratories, Bicol Medical Center, Philippines
2 Lyceum
of the Philippines University- Batangas
81
Poster Presentation
Plants used in traditional medicine provide an interesting and still largely
unexplored source for the development of new drugs. Gliricidia sepium
commonly known as Kakawate or Madre de Cacao is utilized for the
management of coagulation disorders in traditional medical practice but
there is no scientific study conducted yet to date that proves its hemostatic
potential. This study investigated the hemostatic effect of trunk extract
of Gliricidia sepium. Trunk extracts of Gliricidia sepium were obtained
through squeezing process and tested for its hemostatic potential in
citrated plasma through Prothrombin Time (PT), Activated Partial Thromboplastin Time (APTT) and Clotting Time (CT) using standard methods.
Phytochemical screening of the trunk extract revealed the presence of
sterols, alkaloids, saponins, and glycosides. The trunk extract significantly
decreased both PT and APTT (p ＜ 0.05) with an insignificant increase in
the CT (p ＜ 0.05). Results of the recent study indicated that there were
significant differences on each concentration of treated samples in the PT
and APTT when compared to the normal control. This was also observed
when treated per concentration on a dose-dependent manner. The 4% concentration of trunk extract optimally decreased the PT (p=0.0003) while
the 5.46% concentration of trunk extract optimally decreased the APTT
(p=0.0007). This implies the hemostatic property of the trunk extract of
Gliricidia sepium as well as its potential use and suitability for further development of the plant extract as a hemostatic agent.
Oral Presentation
Myeloperoxidase (MPO) is an antimicrobial enzyme in the primary granule
of neutrophils. Ion-exchange chromatography has determined that myeloperoxidase purified from human neutrophils takes three isozymal forms:
MPO-I, MPO-II, and MPO-III. We found that two isozymes, designated
MPO-1 and MPO-2, are expressed in crude MPO of neutrophils in normal
persons. In this study, we sought to determine whether the expression of
MPO-1 and MPO-2 is related to prognosis in patients with chronic myelogenous leukemia (CML). Expression patterns were tested in the peripheral
blood of three groups by means of native polyacrylamide gel electrophoresis (PAGE): normal persons (the normal group), patients with neutrophilia
(the reactive group) as the control group, and patients with CML (the leukemic group). The expression of MPO-1 and MPO-2 in the leukemic group
was more variable than in the normal group (p ＜ 0.000) but was similar
to that in the reactive group. In addition to these isozymes, an abnormal
band of MPO isozyme was found in 42% of the leukemic group but not in
the other two groups. In conclusion, the detection of this abnormal MPO
isozyme on electrophoresis could be a useful indicator of CML. Further
studies are needed to determine the sensitivity and specificity of this finding and whether it is a potentially acceptable clinical criterion for the diagnosis of CML. In addition, concurrent studies are in order to understand
the molecular biology of this abnormal MPO isozyme.
Keywords: Myeloperoxidase (MPO) isozymes, neutrophils, reactive neutrophils, chronic myelogenous leukemic cells
Case Conference
Sangmuk Park
Biomedical Laboratory Science, Dongkang College University, Korea
Symposium
PC-08
Keynote Speech
PC-10
Development of an assay for activated platelet-derived microparticles by ELISA
Fundamental examination for combinations of platelet-related antibodies with sensitivity
and specificity, reaction and conservation conditions, and instruments for sample collections
PC-11
The Effect of Specimen Hemolysis on PT and APTT test
The Interference in coagulation test
Kazuki SHITAMOTO 1, Kozue OKANO1, Minako ARAKI2, Kaori NAKANO3
Meichih Chen, Shuohao Chen, Yungshin Lin, Hsianglin Wan
1 Faculty
of Health Sciences, Yamaguchi University Graduate School of Medicine, Japan
2 Onoda
Red Cross Hospital
3 Yamaguchi
University Hospital
Department of Laboratory Medicine, Taipei Tzu Chi hospital, Buddhist Tzu Chi Medical Foundation, Taiwan
Invited Lecture
Special Lecture
Backgrounds: In the light of laboratory guideline, we reject PT and APTT
samples when it appears hemolysis. But in our laboratory, we found there
are no obvious differences between hemolysis or non-hemolysis sample
had changed on final data.
Method: A total of 159 patient samples with hemolysis and non- hemolysis
were collected from the clinic laboratory. These samples tested PT, APTT
or PT and APTT by clinicians' order. The hemolysis samples tested plasma
hemoglobin level to identify the degree of hemolysis.
Result: There is no statistically significant differences between hemolysis
or non-hemolysis samples in PT test (r2=0.972, p ＞ 0.01), but a little differences in APTT test (r2=0.8544, p ＞ 0.01). According to the CLIA' 88
guideline on test correlation limits, only one PT sample out of the 15% correlation limit (0.6%), but there are 15 APTT samples out of the limit (9.4%).
There is no evidence show the relation between the data correlation and
the degree of hemolysis.
Conclusion: These data show PT is more stable than APTT when in hemolysis sample. We recommend APTT should have a stricter rejected criterion
than PT when sample has hemolysis.
Educational Lecture
[Introduction]Platelet-derived microparticle (PDMP) has a strong influence on the blood clot formation and inflammatory, however, measuring
method and mechanism of clot formation have not been clarified. We
conducted a fundamental examination of an assay by ELISA using various
platelet-related antibodies to determine activated-PDMP (aPDMP).[Materials and Method]Platelet rich plasmas (PRP) were prepared from healthy
volunteers' citrated plasma. The platelet count of PRP was adjusted to
200,000/ μ l. Adjusted PRP were stimulated with 100 μ M ADP at 37
degrees Celsius for 10 minutes, and sonicated to define aPDMP to obtain
calibration curve. We collected blood with 21G type needles into two
vacuum collection tubes where citric acid was added, and only the second
blood tubes were employed. Blood samples were centrifuged at 1200g
for 10 minutes to extract supernatant. To confirm combination of various
platelet-related antibodies for aPDMP, we used anti-vWF/anti-GPIIb/III/
anti-AnnexinV antibodies for solid phases, and anti-GPIb/anti-CD40/antiCD36 antibodies for detectors and examined their sensitivity and specificity. Reaction temperatures were 4 degrees Celsius to 37 degrees Celsius
and reaction times were one hour to overnight. Specificity was confirmed
by the reactivity not only with platelet specimen but also with red blood
cells and neutrophils.[Result and Discussions]The sensitivity and specificity
of aPDMP varied according to the combination of platelet-related antibodies, however, the combination of anti-AnnexinV polyclonal antibody for
solid-phase and anti-GPIb monocleonal antibody for detector was the most
suitable for aPDMP. Freezing was suitable to preserve samples. The coefficient of variation showed a favorable reproducibility of 12.6%. Since different makers or types of needles and vacuum collection tubes and methods
for blood collections significantly influenced the results, making strict
conditions would be needed. Further research on the association between
aPDMP level and various diseases will be required in the future.
Symposium
PC-12
Case Report: A 9-year-old girl with Chediak-Higashi Syndrome
Case Report
PC-13
Research and development of portable device for hemoglobin
concentration test
The test performance for the Point of Care Testing
Hiroyuki Nozaka1, Masakatsu Ooura2,3, Aita Megumi1, Hiroki Sazawa1, Miyuki Fujioka1, Hideki Takami1,
Kazuyuki Kida1
Shuohao Chen , Meichih Chen, Yungshin Lin, Hsianglin Wan
Taipei Tzu Chi Hospital, Buddhist Tzu Chi Medicine Foundation, Department of Laboratory Medicine, Taiwan
Case Conference
1 Graduate
school of health sciences, Hirosaki university, Japan
2 Graduate
school of humanities and social sciences, Hirosaki university
3 CONSIS.
INC
Oral Presentation
Poster Presentation
Background: Chediak-Higashi Syndrome (CHS) is a rare autosomal recessive disorder that arises from a mutation of a lysosomal trafficking regulator protein, which leads to a decrease in phagocytosis. Patients with CHS
exhibit hypopigmentation of the skin, eyes, and hair. This disease damages
immune system cells, leaving them less able to fight off invaders such as
viruses and bacteria. As a result, most people with Chediak-Higashi Syndrome have repeated and persistent infections starting in infancy or early
childhood. These infections tend to be very serious or life-threatening.
Case Presentation: A 9-year-old female came to our hospital because of repeat fever. In the past, this patient also had history of regular visit the dermatologist because the skin spot lesions. When physical examination, she
had pale face and her height is lower than other same age children and
seems like a little sluggish in speech. When analysis the CBC and differential count, we found there are some giant pink granules in neutrophils and
lymphocytes cytoplasm. Combining her clinical symptoms and discussing
with clinicians and pathologist, we concluded these granules are ChediakHigashi anomaly. These granules are specific found in CHS patient's while
blood cell cytoplasm. With the careful surveillance by the clinical laboratory scientist, found the characteristic white cells inclusions and discussion with clinicians, the patient had the right time of diagnosis and proper
management.
1. INTRODUCTION
It is reported that nutritional management is strongly related to patient prognosis or recovery period. In addition, Nutritional management is also started in
home therapy. Therefore, the role of clinical examination in Nutrition Support
Team activities become more important. Hemoglobin, albumin and total protein value is used as a typical nutritional management indicator. However, Point
of care treatment (POCT) devices that nurses or public health nurses are able
to operate easily in the home therapy is few, it is required to develop new POCT
devices for medical activities in out-of-hospital. The aim of this work is development and evaluation of portable device for hemoglobin concentration test.
2. SYSTEM CONFIGURATION
The portable device was consisted of photo-transistor unit, LED unit, and data
logger system "NI-USB DAQ". Application software was developed by "NI-LabVIEW". Hemoglobin measurement principle was used SLS-hemoglobin method.
3.SYSTEM EVALUATION
Peripheral blood for this study were collected from 40 normal healthy persons.
We draw blood from fingertip by lancet, and collected into micro capillary. We
collected peripheral blood from brachial vein into blood collection tube with
EDTA-2Na. Hemoglobin value was measured by using our system, and the
reference hemoglobin value was measured by using Fuji dry-chemistry system.
We evaluated correlation and accuracy.
4.RESULTS
Two system showed good correlation. Analytical accuracy of our system
showed low compared to the Fuji dry-chemistry system, but accuracy in clinical
fields was sufficient.
5. CONCLUSION
Our system needs small amount of blood collection, not only nurse but also
patiant is able to drew blood easily. Operability is also simple and easy. It seems
this system is useful for medical activities in out-of-hospital. Our future work is
the evaluation of patients with anemia in the clinical field.
6. ACKNOWLEDGMENT
This work was supported by SCOPE of the Japan Ministry of Internal Affairs
and Communications.
82
Flow Cytometric Lymphocyte Screening with 14 Antibodies in 10 Colors
Experience in Clinical Flow Cytometry Laboratory at Fimlab
Laboratories in Finland
PC-15
Discrepancy between conventional cytogenetic analysis and
FISH signals for RUNX1/RUNX1T1
Insertion (21;8)(q22;q22q22); a cryptic t(8;21) in AML
Riitta Kemppi, Arja Alkula
Myoung Hee Ham1, Seok Young Hong1, Jung A Kim1, Dong Soon Lee1,2
Hematogy Department, Fimlab Laboratories, Finland
1 Laboratory
Medicine, Seoul National University Hospital, Seoul, Republic of Korea
2 Department
of Laboratory Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea
Clonality study of T-cell Lymphoproliferations in Taiwan
Review of 40 cases with comparison of different clonality
tests
Yung-Tung Hung, Yen-Chuan Hsieh
Department of Clinical Pathology, Chi-Mei Medical Center, Taiwan
Oral Presentation
Malignant lymphoma is a clonal proliferation of a malignantly transformed
lymphocyte of B-cell, T-cell , or natural killer (NK) cell. The clonal nature of
lymphoma cells can be elucidated by B-cell immunoglobulin gene or T-cell
receptor gene rearrangement study. Distinguishing malignant lymphoma
from a reactive process is sometimes a big challenge and this is the major
reason that molecular diagnostic assays for clonality have become increasingly important and popular in the clinical diagnostic laboratories. These
assays are performed to determine the clonality of suspected lymphoproliferations. In our previous study we have successfully used polymerase
chain reaction (PCR) method and conventional primers to detect the clonal
gene rearrangements of immunoglobulin heavy chain gene (IGH) and Tcell receptor Γ chain gene ( TCR- Γ ) for B-cell and T-cell clonality, respectively. A big study from 45 top European laboratories led to the launch of
BIOMED-2 multiplex primers in 2003. We retrospectively searched for Tcell lymphoma in our institute from 2001 to 2015 and identified 40 cases.
In this study, we compared the conventional and BIOMED-2 primers and
found that the latter was more sensitive and relatively easy to perform.
We confirmed that the BIOMED-2 primers were useful for clonality test on
paraffin sections. Further larger study is warranted for the comparison of
PCR-based study and GeneScan analysis.
Poster Presentation
Introduction
In Nigeria, there is a large number of sufferers of sickle cell anaemia, resulting from sickled red cells. As a result of this global problem of sickle
cell disease, scientists have been on deck in search of solutions to better
the lives of those involved.
Methodology
The effects of coconut water intake on haemorheology and glucose metabolism were studied in 35 subjects comprising 25 from the test groups
(SS, SC, and AS Genotypes) compared with 10 HbAA controls. Each subject
was investigated longitudinally after the intake of coconut water for a
period of five days. Parameters assessed include: relative plasma viscosity
(RPV), relative whole blood viscosity (RWBV), deformability index using
the transit time (DI), differential count, white blood cells (WBC), packed
cell volume (PCV), platelets count, fasting blood sugar (FSB) and two hours
post prandial Glucose (2hrPP) using standard methods.
Result
Relative whole blood viscosity was significantly lower in HbSS (4.35 ±
0.46) and HbSC (6.05 ± 0.56) on the first day of coconut water intake
compared with HbAA (6.47 ± 0.46) (P<0.05). There was a significant
reduction in whole blood viscosity in HbSS and HbSC on day five of coconut water intake compared with HbAA (P<0.05). No significant change
in blood sugar level was detected in all the subjects after administration
of coconut water until day 5, whereas FBS reduced significantly in HbSC
subjects compared with those in control group (HbAA) (P<0.05). The WBC,
PLT, neutrophil counts and percentage PCV significantly increased in HBSS
group compared with the HBAA group (P<0.05). From day 1 to day 5, PCV
concentration in the test groups (HBAS, HBSS, HBSC), remained relatively
unchanged (P ＞ 0.05).
Conclusion
In conclusion, coconut water intake markedly reduced the deformability
time in HbSS subjects, increased their haematological properties and did
not adversely increase the blood sugar level.
Case Conference
Florence I. Aboderin1, Ifedayo O. Ajayi2
1 Haemalogy
and Immunology, Obafemi Awolowo University, Nigeria
2 Department
of Physiology, University of Benin, Benin City, Nigeria
Symposium
PC-17
Educational Lecture
Effects of coconut water on haemorrheology and glucose
metabolism in sickle cell patients in Nigeria
Special Lecture
PC-16
Background
Translocation (8;21)(q22;q22) is one of the most recurrent cytogenetic
abnormalities in acute myeloid leukemia (AML). Typically, the RUNX1/
RUNX1T1 rearrangement shows balanced translocation between chromosomes 8 and 21. This case report is about variant forms of translocation
caused by cryptic insertion of RUNX1T1 (8q22) gene on RUNX1 (21q22)
gene. These forms of translocation are hardly detectable by conventional
cytogenetic studies. Fluorescence in situ hybridization (FISH) or reverse
transcriptase polymerase chain reaction (RT-PCR) techniques are needed
to reveal the rearrangements.
Case Report
A 47 year old woman referred to our hospital was diagnosed with AML
based on morphologic, cytochemical, and immunophenotypic findings on
bone marrow studies. Conventional cytogenetic analysis revealed 45,X,X[18]/46,XX[2]. But, interphase fluorescence in situ hybridization (FISH)
for the RUNX1/RUNX1T1 rearrangement showed chimeric fusion gene in
96% of bone marrow nucleated cells, one fusion, two orange, one green
signal pattern(1F2O1G) using RUNX1/RUNX1T1 dual-color dual fusion
probe[Vysis; Abbott Molecular, Downers Grove, IL, USA. RUNX1T1(orange),
RUNX1(green)]. Meanwhile, typical pattern of RUNX1/RUNX1T1 rearrangement is two fusion, one orange and one green signal. In this patient,
metaphase FISH revealed that one fusion signal was located on the long
arm of chromosome 21, but no fusion signal on derivative chromosome
8. Additionally, RUNX1/RUNX1T1 rearrangement was accompanied
by the loss of a whole X chromosome which is the most common additional cytogenetic aberration in AML with RUNX1/RUNX1T1. Molecular
genetic study by reverse transcriptase polymerase chain reaction (RTPCR) confirmed the presence of the chimeric transcript for RUNX1/
RUNX1T1. The final karyotype was 45,X,-X.ish ins(21;8)(q22;q22q22)
(RUNX1T1+;RUNX1+,RUNX1T1+)[18]/46,XX[2]
Discussion
We experienced the cryptic rearrangement that is hardly detected by conventional cytogenetic analysis. Molecular genetic studies such as FISH or
RT-PCR is very useful tool to detect the masked type of cryptic translocation.
Invited Lecture
Flow cytometric immunophenotyping is the method of choice to detect
abnormal lymphoid populations in blood and bone marrow samples. In
2009 EuroFlow consortium proposed 8-color and 12 antibody panel for
lymphocyte screening tube. A comprehensive panel to accurately classify
lymphoid neoplasms is selected based on the results of this screening tube.
In Fimlab Laboratories we have designed a lymphocyte screening tube
with 14 antibodies in 10 colors for Beckman Coulter Navios instruments
and applied this approach since January 2011. This screening tube is a
modification of Euroflow 8-color tube.
The proper screening tube is selected by the Biomedical Laboratory Scientist (BLS) based on clinical information according to written guidelines.
Lymphocyte screening tube is selected, if patient has lymphocytosis or
lymphatic infiltrates in bone marrow smears.
The composition of 14 antibody 10-color lymphocyte screening panel is:
CD8/lambda FITC, CD56/kappa PE, CD5 ECD, CD14 PC5.5, CD19/TCRgd
PC7, CD3 APC, CD7 APC-R700, CD38 APC-H7, CD20/CD4 BV421, CD45
KO. PB or BM sample is incubated with antibodies for 15 min in dark.
Red cells are lysed with BD FACS lysing solution for 10 min and cells are
washed with PBS-BSA and resuspended in 0.5 ml of buffer and acquired
on Navios flow cytometer.
A predefined strategy is applied for gating. BLS makes the primary analysis and in clear cases selects the confirmatory panel (T-cell, B-cell, NK-cell
or plasma cell panel). Final analysis and reports are given by Laboratory
Hematologist.
In the poster we illustrate the gating strategy and guidelines for selection of secondary panel. In conclusion, we find this strategy with primary
screening and targeted secondary panel cost effective and time saving.
With good training and written guidelines BLS can select the proper secondary panel. We find that the interpretation of results adds interest to the
whole procedure and is rewarding.
Keynote Speech
PC-14
83
Keynote Speech
PC-18
Ethnic Difference of Chromosome Aberrations and Genetic
Mutations in Chronic Lymphocytic Leukemia
Ethnic Difference in Chronic Lymphocytic Leukemia
PC-19
COMPARISON OF MEASUREMENTS IN THE TWO
POSITIONS AND SUITABILITY OF ERROR MESSAGE OF
THE PFA-100
Soonok Kim1, Jung-Ah Kim1, Si Nae Park2, Kyongok Im2, Seon Young Kim1, Dong Soon Lee1,2
Hyun-A LEE, Ji-Hye PARK, Myung-Hyun NAM, Soo-Young YOON
1 Department
of Laboratory Medicine, Seoul National University Hospital, Seoul, Republic of Korea
2 Cancer
Research Institute, Seoul National University College of Medicine, Seoul, Republic of Korea
Department of Laboratory Medicine, Korea University Medical Center, Korea
Invited Lecture
Background: The platelet function analyzer (PFA)-100 is an in vitro system for measuring platelet function in citrated whole blood. We have wondered whether retesting by error messages is suitable and experienced a
difference between measurements in the two positions of the instrument
since one follows the other.
Special Lecture
Educational Lecture
Background: Chronic lymphocytic leukemia (CLL) is considered as typical
malignancy with ethnic difference, which is one of the common leukemia
in western countries, while extremely rare in Asian countries. To investigate whether chromosome aberration and genetic mutation of Korean CLL
patients is different from that of Caucasian, we performed conventional
cytogenetic test, fluorescent in situ hybridization (FISH) and target capture
sequencing for 87 hematologic malignancy related genes using custom
designed capture panel.
Methods: A total of 71 CLL patients were enrolled. We performed Gbanding in 60 patients with CLL and fluorescent in situ hybridization (FISH)
for chromosome 12 enumeration, 13q14.3 deletion, and 17p13 deletions
in 51 patients with CLL. At the same time, target exome sequencing for an
87-gene panel using next-generation sequencing was performed in 41 patients with CLL. The sequencing reads were analyzed using a bioinformatics pipeline.
Results: Aberrant karyotypes were observed in 28.3% of patients by Gbanding and 66.7% of patients by FISH. Targeted exome-sequencing
analysis revealed 76 substitutions and insertion/deletions. The most common recurrent mutation ( ＞ 5% frequency) was ATM (20.8%), followed by
TP53 (14.6%), SF3B1 (10.4%), KLHL6 (8.3%), and BCOR (6.25%). The ATM
mut, KLHL6 mut and BCOR mut were more frequent in Korean notably
and the ATM mutation exhibited a 2-fold higher incidence (20.8% vs. 9%)
in Koreans compared to Caucasians.
Conclusions: The mutation profile of Korean CLL patients differed from
that of Caucasian CLL patients, but their cytogenetic aberration profiles
were similar. Novel mutations discovered in this study must be validated
through large cohort studies and may offer clues to the mechanisms underlying the ethnic difference in CLL incidence.
Symposium
PC-21
Objective: The purpose of this study was to investigate accordance rate of
repeated test results from abnormal values following simultaneous installation and to solve a difference in the test results of both positions.
Methods: A total of 10,956 samples were enrolled. If flagging error messages, the sample was duplicated. With prolonged closure times (CTs) in
both positions, we checked absence of previous history or laboratory results indicative of platelet dysfunction, and retested singly by transferring
the prolonged B position sample to the A position.
Results: 1,544 samples (14.1%) showed prolonged CTs. 97.8% among
those with error messages appeared any reportable message, showing
high relevance between initial testing and retesting. 17.2% of B position
samples with CT prolongation in both positions was changed to within the
reference range.
Conclusion: There was no need for routine duplicate testing for reportable
error messages. Checkup and retest for the prolonged B position sample
would be necessary in case of prolonged CTs in both positions simultaneous installation of PFA-100.
Refining reporting results of body fluid analysis for clinical purpose
Commenting summarization of significant results for reporting results
to highline
PC-22
Establishment of novel flow cytometric test for Adult T-cell leukemia (ATL) and its clinical application
- Precise quantification and evaluation of HTLV1 infected cells in ATL patients could support clinical
practice for 5 years -
Tomohiro Ishigaki1,2, Yuji Zaike1, Seiichiro Kobayashi3, Nobuhiro Ohno4, Naoyuki Isoo1,
Kaoru Uchimaru5, Arinobu Tojo3,4, Hiromitsu Nakauchi2
Hsiu-Chin Fan, Fang-Yu Wang, Ya-Ching Li, Pei-Chen Li, Chun-Chuan Lin
Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taipei, Taiwan
Case Conference
1 Department
of Laboratory Medicine, Research Hospital, The Institute of Medical Science, The University of Tokyo, Japan
2 Division
of Stem Cell Therapy, The Institute of Medical Science, The University of Tokyo
3 Division
of Molecular therapy, The Institute of Medical Science, The University of Tokyo
4 Department
of Hematology and Oncology, Research Hospital, The Institute of Medical Science, The University of Tokyo
5 Laboratory
of Tumor Cell Biology, The University of Tokyo
Oral Presentation
Poster Presentation
The intended purpose of body fluid cellular analysis is for the characterization of inflammatory, infectious, neoplastic, and immune alterations.
However the equivocal reference intervals of cellular constitute acquire
much more difficulty for clinicians to diagnosis. Despite limited clinical
usefulness, body fluid analysis brings about tedious workloads for testing
personnel in hematology laboratories. This study is aimed to develop solutions to enhance its clinical purpose and to ease off a little cumbrance for
testing personnel.
The 622 cases for body fluid analysis were collected in March 2016, of
which 65 cases were traumatic taps. 157 cases with nucleated cell count
beyond reference intervals comprised 13 CSF, 93 ascites, 23 Pleural fluids
and 28 synovial fluids. 29 cases with atypical cells (suspected neoplastic
cell) comprised 2 CSF, 7 ascites and 10 Pleural fluids. 13 cases with microorganisms consist of 10 ascites and 3 Pleural fluids. It shows that the most
frequency causes of ascites and Pleural fluids are infection and malignant
disease. Besides, it was found the cases with crystals and microorganisms
have strong associations with neutrophil predominant finding. To improve
clinical usefulness, we decide to amend the report format by adding texted
messages of commenting as necessity which summarize significant results
from each specimen, while the messages directly reveal the relevant characterization of pathology cause. Cell categories are redefined based on
the origin cells derived from and on their clear distinguish of morphology,
which make easier to perform cell differential. The criteria are set for each
type of specimen if using chamber differential is adequate to decrease reductant workloads.
Conclusion: Commenting summarization of significant results is helpful
to medical review, and for pleural fluids and ascites, when nucleated cell
numeration is under 0.5 × 109/L , the chamber counting for differential is
adequate for rule out bacterial infection.
Adult T-cell leukemia (ATL) is highly aggressive malignancy of mature Tcells caused by human T-cell lymphotropic virus type1 (HTLV1). A quantitative analysis of HTLV1-infected ATL cells in the peripheral blood of ATL
patients is important to assess disease progression and effectiveness of
therapy. Cells with abnormally hyperlobulated nuclei, which are termed
flower cells, are characteristic ATL cells, but ATL cells are not always typical flower cells. Discriminating HTLV1-infected ATL cells morphologically
from other lymphocytes, particularly from reactive atypical lymphocytes,
is really difficult. These difficulties therefore cause differences between
examiners and errors, and morphological quantification of ATL cells tends
to be inaccurate.
To settle these, we have been analyzing many clinical samples of acutetype ATL patients minutely with 12-color flow cytometry. We finally elucidated CD4-positive cells in HTLV1-infected ATL patients could be mainly
classified to three groups of CADM1-CD7positive (CD7P), CADM1+CD7dim
(CD7D), and CADM1+CD7negative (CD7N), which correspond to uninfected normal T-cells, HTLV1-infected cells, and ATL cells, respectively.
We applied these findings of basic research to a new clinical test which
can accurately quantify HTLV1-infected ATL cells using simple four-color
flow cytometry, and actually used the test in clinical practice. The test has
been supporting clinical practice for about 5 years. A strong correlation
was observed between the number of HTLV1-infected ATL cells measured
by flow cytometry and the number of abnormal lymphocytes measured
microscopically only by experienced technicians. We also found that the
frequency of CD7D and CD7N cells among CD4+ cells changed during
chemotherapy, which reflected differences between chemosensitive and
chemoresistant cases. Kaplan-Meier analysis with log-rank test showed
that this clinical laboratory test could be useful to evaluate effectiveness of
chemotherapy and predict prognosis.
84
Effect of pH levels on Blood Cell Morphology in vitro
The important maintainance of pH levels to play a key roles
in Blood Cells
PC-24
Usefulness utility of leukocyte and platelet Parameters for rapidly discriminating bacteremia and urinary
tract infections
Rapid Discrimination between bacteremia and urinary tract infections by Reliable complete Blood Cell Indice
Hyo Chan Kang1, Ki Ja Jung2
Ching-Mei Chen1, Hsiao-Yen Hung1, Kun-Pi Li1, Ming-Chung Wang2, Wan-Ting Huang3
1 Medical
Laboratory Science, Dongeui Institute of Technology, Korea
2 Laboratory
Medicine, Inje University Haeundae Paik Hospital
1 Department
of Laboratory medicine, Chang Gung Medical Foundation Kaohsiung Branch, Kaohsiung, Taiwan
2 Division
of Hematology and Oncology, Chang Gung Medical Foundation Kaohsiung Branch, Kaohsiung, Taiwan
3 Department
of Pathology, Chang Gung Medical Foundation Kaohsiung Branch, Kaohsiung, Taiwan
Back to the Basic : From Platelet to Hemostasis
The Role of Platelet in Hemostasis
The criteria for action following automated haematology analysis had
been suggested by experts for a decade, which help to improve uniformity
of automation applications among laboratories. However for those arrested results need slide reviews, there is no detailed instruction provided
for reviewers, laboratories are incline to mandate reviewers to report or
even revise instrument results based on their own experience. It makes the
slide review incapable to ensure the consistency of review reporting, neither guarantee for the more accurate. The study is aimed to enhance the
consistency of manual reporting by setting guides for reviewers.
The 6,938 cases of the arrested results were collected on day shifts in a
period of two weeks, of which it was found the results of 1,238 cases had
been revised. The 1,083 revised cases were inevitable because of interference, no DCs from instrument, and immature cells finding, however the
other 155 revised cases of DC resulted from incomparable DC between instrument count and reviewers' numeration. What make reviewers identify
incomparability? The answers from reviewers were diversity which really
needed algorithms to mitigate.
After analysis the algorithms for decision making process were drafted,
which triggered by serious of morphology flags. A newly purchased haematology analyser other than primary automation is also incorporate into
this process as a confirm system for blast, NRBC, PLT clumps. Random
revision of DC is not allowed owing to unreliability of variant smear preparations and limited cells counted. The protocol for manual numeration
of PLT is established through meticulous calculations and experiments,
although arbitrarily using manual numeration of PLT to replace the instrument results is not allowable, but using delta checking as criterion for replacement is encouraged. After all done, the reviewers are more clearly to
understand the limitation of slide review and either comfortable to define
the necessity for revisions.
Platelets are important in hemostasis. When blood vessel is injured, platelets attached there as monolayer. And then platelets come in contact with
each other by granules such as ADP, TXA2 etc from alpha granule and
dense core granule. Platelet is a small blood cell which is 1 to 4 micrometer and 5 to 10 days life span. It is found in bone marrow, tissue, blood
vessel and shapes small discoid in resting state but change it's shape as
round and further more looks like spurr cell when trigered by angonists
such as collagen, TXA2, thrombin, epinephrine, ristocetin. Platelet have
two types of functional granules which is alpha and dense core granules.
Alpha granule release fibrinogen, von willebrand's factor, platelet derived
growth factor(PDGW), platelet factor 4(PF4), and other proteins. Dense
core granule release ATP, ADP, serotonin, calcium ion and magnesium ion.
Platelets have three major functions which are adhesion, granules release
and aggregation. Hemostasis is divided by primary and secondary hemostasis. During the hemostasis, platelets are concerned as primary hemostasis. Disease associated with platelets are divided by genetic disorders, genetic defects, storage pool defects, acquired defects, quantitative disorders.
Bernard-Soulier disease and von willebrand's disease are genetic disorders.
Glanzmann's thrombasthenia is genetic defects. Gray-platelet syndrome,
Wiskott-Aldrich syndrome, Hermansky-Pudlak syndrome, Chediac-Higashi
anamaly are storage pool defects. Acquired defects are caused by drug and
diet. And last introduce the platelet function tests old and new.
85
Poster Presentation
Medical Laboratory Science, Dongeui Institute of Technology, Korea
Oral Presentation
Hyo Chan Kang
Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taipei, Taiwan
Case Conference
Chun-Chuan Lin, Ya-Ching Li, Fang-Yu Wang, Li-Ping Yin, Hsiu-Chin Fan
Symposium
PC-26
Educational Lecture
Guides for manual estimation of blood slide review
Developing algorithms to decide revising of automated CBC
and WBC differentia results based on manual review
Special Lecture
PC-25
Background: Urinary tract infections (UTIs) are a frequent problem worldwide which are caused by microbial invasion to different tissues of the
urinary tract. Bacteremia is the presence of viable bacteria in the circulating blood. Common signs and symptoms include fever, thrombocytopenia,
and neutrophilia. However, microbiological cultures; the conventional gold
standard diagnostic method, are often time consuming. It is important that
a rapid and sensitivity biomarker be discriminate patients with bacteremia
form UTIs patients. The complete blood count is one of the most common
blood tests in the laboratory. We decided to study the efficacy of new
hematologic parameters, such as immature platelet fraction (IPF%), mean
platelet volume and platelet distribution width, and high-fluorescent lymphocyte counts (HFLC%), and validated the application of discriminating
algorithms for screening subjects for bacteremia and urinary tract infections.
Methods: Our study included 3 groups, namely, 357 subjects with UTIs,
318 subjects with bacteremia, and apparently 298 healthy hospital employees. We used the Sysmex XE-5000 automated hematology analyzer
and analyzed with full hemocytometric parameter profile to ensure that
results of all parameters were available. For each sample, PCT, CRP, and
lactate were measured. The Pearson correlation and ANOVA were calculated; ROC curve analysis was used to determine their diagnostic performance.
Results: The patient with UTIs or bacteremia showed significantly higher
IPF%, HFLC%, PCT, CRP, and lactate than non-infection patients(P ＜
0.001). Additionally, IPF% and HFLC% value were significantly higher (P ＜
0.001) in samples with bacteremia than in UTIs samples. The performance
of IPF% and HFLC% (AUC=0.968) in the discrimination of bacteriemic
patients from UTI patients were significantly better than the PCT/CRP/
Lactate (AUC=0.910/0.870/0.811, respectively). Sensitivity (93.8%) and
accuracy (85.1%) of IPF were the best among all biomarkers.
Conclusion: We conclude that the advanced algorithms, derived from
extended CBC parameters, are rapid useful as laboratory devices for UTIs
screening.
Invited Lecture
Blood pH level is very important in the human body to maintain homeostasis. This study shows the effect of pH level on Blood Cell Morphology in
vitro. Blood sample were collected from healthy college students in early
20's. We studied blood cell morphology (esp. WBC and RBC) in each pH
levels from acid (pH 5.0) to alkali (pH 9.0) conditions and adjusted to pH
using 0.1N NaOH and 0.1N HCl in diluents. At first each samples were
analyzed by sysmex XS-1000i (Japan) and stained Wright's stain for microscopy. As a results at pH 8.0 RBC showed rouleau formation and at pH
9.0 RBC showed rouleau formation strongly, some WBC showed naked nucleus. While at pH 6.0 RBC changed from normal shape to tear drop and
burr cell types and WBC showed naked nucleus at the same pH 9.0. At pH
5.0 lots of RBCs showed schistocytes and spur cells. In conclusion Blood
cells are sensitive in pH variation and especially RBC was more sensitive
than WBC. This study shows how important pH conditions in our body
and blood cells to play s a key roles in their functions.
Keynote Speech
PC-23
Keynote Speech
PC-27
Effects of Improperly-Filling Dipotassiumethylenediaminetet
raacetic Acid (K2EDTA) with blood of Clinically Diagnosed
Dengue and Non-Dengue Individuals
PC-28
The Discussion of Intravascular Pattern
Plasma Hemoglobin versus Haptoglobin and The Others
Parameters
Gregorio L. Martin I, Carlos Jose I. Alves, Mikee Estella Gem P. Apostol, Belisarius Arandia,
Jejomar Emmanuel C. Caringal, Arveec C. Chiong, Karen Angelica R. Domalaon, Christine Mae E. Reyno
Yi-Yun Chen, Hui-Szu Tsai, Fang-Yu Wang, Chun-Chuan Lin, Chia-Lian Chung, Chuan-Po Lee,
Yen-Li Wang, Hsiu-Chin Fan
Department of Medical Technology, University of Santo Tomas, Philippines
Division of General Laboratory, Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taiwan,
R.O.C, Taiwan
Invited Lecture
Special Lecture
Under filling and overfilling K2EDTA tubes is an inevitable practice in hospitals especially in wards where patients have fragile veins, and are considered hard to extract patients. The researchers looked into the hematologic
effects of improperly filling K2EDTA tubes in the complete blood count of
clinically diagnosed dengue patients and healthy individuals. A total sample size of 48 consisting of 24 individuals with ages ranging from seven to
45 years old, diagnosed with dengue fever syndrome, and 24 healthy individuals with the same age range were selected for the study. Six milliliters
of venous blood were put on three tubes by under filling, properly filling,
and overfilling each tube; and ran in time intervals of 30 minutes, one, two,
and four hours. Using SPSS 20.0, the parameters affected by improperly
filling the tubes were RBC (p-value=0.027), Platelet (p-value=0.035), and
Segmenters (p-value =0.011); by time of feeding were Hemoglobin (p-value
= ＜ 0.001), Hematocrit (p-value =0.034), Segmenters (p-value= ＜ 0.001),
Lymphocytes (p-value= ＜ 0.001), Monocytes (p-value=0.001), and Eosinophils (p-value=0.006); by control group were WBC (p-value= ＜ 0.001),
Hemoglobin (p-value=0.018), Hematocrit (p-value=0.008), and Platelet (pvalue= ＜ 0.001); by volume and time was Hematocrit (p-value=0.042); by
control group and time were Monocytes (p-value=0.0030), and Basophils
(p-value=0.001).
While increasing destruction of RBC in intravascular hemolysis, the level
of free hemoglobin in plasma (plasma-Hb, ΑΒ dimers) shall be elevated
and integrate with haptoglobin. Then the hemoglobin-haptoglobin complexes were removed by macrophages in the reticuloendothelial system
like spleen, liver, lymph nodes. Besides, plasma-Hb would filter out by
glomerular and shall be reabsorbed by renal tubular epithelial cells, if
not, the urine hemosiderin would present. The reference range of plasma
hemoglobin tested by tetramethylbenzidine (TMB)-colorimetry is ＜ 5 mg/
dL and the clinical determined concentration is ＞ 20mg/dL. There are
not many tests directly relative to detect intravascular hemolysis, so we
were interested in the pattern of it and conducted a survey of 289 cases to
discussion the relations between plasma hemoglobin, haptoglobin, LDH,
bilirubin, hemoglobin, reticulocyte and urine hemosiderin. The concentration of plasma hemoglobin was divided into three groups: ＜ 5 mg/dL
(group I), 5~20 mg/dL (group II), ＞ 20 mg/dL (group III). The results of
haptoglobin were low while results of LDH, bilirubin and reticulocyte were
high in group II and group III. Also, the statistical results of haptoglobin,
bilirubin and reticulocyte all showed significant difference (p ＜ 0.05) in
group I, group II and group II, group III. When the results of plasma hemoglobin were lower than 5 mg/dL and haptoglobin test, it's likely to reduce
the possibility intravascular hemolysis.
Educational Lecture
Symposium
PC-29
Haematological Reference Ranges among Health Adult Cameroonians.
Haemotological Reference Ranges For Healthy Cameroonians in the
Bamenda City.
PC-30
Victor Nnibah FONDOH1, Teresa Woyen N2, Nakeli Nakeli B3, Mokom Blessing F3, Kinge Thompson N3,
Ndassi Julinna PhD3, Njukeng Patrick A3, Shang Judith D3
How to deal with platelet aggregation in EDTA-blood
Else Marie Jørdre, Yvonne Dikkanen
Oslo University Hospital, Oslo, Norway
Case Conference
1 Laboratory,
Regional Hospial Bamenda, Cameroon
2 Global
Health Systems Solution, Limbe, Cameroon
3 Center
for Disease control and Prevention (CDC), Camroon
Oral Presentation
Poster Presentation
Introduction:In our laboratory we sometimes discover samples with platelet clumping in Edta blood. We want to show different methods we use to
solve the problem. The 3 methods we use are:
1.Kanamycin method
2.Blood smear and microscopy
3.Phase contrast and microscopy
4.Citrate blood.
Material and methods:
1.Kanamycin method: As soon as we discovered samples from EDTA-tubes
with platelet aggregation we treated them with the aminoglycoside Kanamycin , and then reanalyzed. We are compering pictures of scatter plot
before and after adding Kanamycin to the samples.
2.Blood smear and microscopy: After making a blood smear we coloring
the smear with May-Gr&uuml;nwald and Giemsa.
3.Phase contrast: A drop of EDTA-blood on a slide covered by a glass microscoped by Phase Contrast Optical Microscop
4.Citrate blood: We have to take an additional sample from our patient using the citrate tube. The tube has to be analyzed within a few minutes.
Result:
1.Kanamycin method: We discovered the results of platelets being improved in most cases. As an additional advantage of this method our White
blood differential plot was improved, a great advantage.
2.Blood smear and microscopy: After making a blood smear we coloring
the smear with May-Gr&uuml;nwald and Giemsa. We discover if there are
platelets aggregation or not.
3.Phase contrast: We will discover if the platelets are aggregated. Easy to
see if the Platelets are free ore clumped.
4.Citrate blood: This is time consuming and a problem for our patients.
Conclusion: The advantage of Kanamycin method is that we get the correct result of the platelets and we even get the White blood differential
counting correct as well. We use the EDTA-tube we already have and a
new sampling is not required. We were able to produce results faster and
better.
The other methods helps us to verify if there are platelet clumping or not.
Haematological reference ranges are essential for interpretation of data
for diagnostic orientation, treatment decision and research studies. The
use of reference ranges from manufacturers' kits and text books has been
an issue in clinical Laboratories in Cameroon. These values might affect
the clinical decision of the patients. The aim of this study was to determine
the mean, median, 2.5th and 95.5th percentile of reference interval for
healthy adults in the Bamenda City, Cameroon. This was a retrospective
study. Secondary data was explored from method verification records of
Complete blood count auto analyzer (Mindray BC 2800) was explored.
Precision and accuracy of the machine were assured by running internal
quality control daily and external quality control (RIQAS of RANDOX)
every two week respectively. Data for 175 healthy voluntary blood donors
were considered in the study. Only 150 subjects aged 17 to 60 years were
negative for HIV, HBsAg, HCV, syphilis and without Haemoglubin abnormalities in Hb-Electrophoresis. The mean: Leukocyte (WBC) count was
lower in males than in females (5.8X 103/ul versus 6.6X103/ul, p=0.004),
Erythrocyte (RRC) count was higher in males than in females (5.2X106/ul
versus 4.6X106/ul, p=0.000), Haemoglobin level was higher in males than
in females (14.9g/dl versus 12.9 g/dl, p=0.000), Haematocrit was higher
in male than in females (44.1% versus 39.1%, p=0.000) and Platelet count
was 205X103/ul in male and 229X103/ul in female. We propose that, in
the absence of previously established hematological reference interval in
Cameroon we offer to use these results for clinical management of patients
and interpretation of laboratory data for research purpose.
86
Expression and localization of RNase and RNase inhibitor in
blood cells and vascular endothelial cells
Expression of RNase involved in blood coagulation
PC-32
Laboratory evaluation of coagulation activity change by
antineoplastic drugs for the treatment of hematological
tumors
Background: Idarubicin (IDR), doxorubicin (Dox), and vorinostat (Vor) are
effective for treatment of myeloid and lymphoid tumors. During the initial
course of induction therapies activation of intravascular coagulant activity
is induced, which leads to disseminated intravascular coagulation or deep
vein thrombosis.Aims: We examined the procoagulant effects of the antineoplastic drugs for myeloid and lymphoid tumors, especially focusing on
tissue factor (TF) and anionic phospholipid phosphatidylserine (PS).Methods: We examined the procoagulant effects of pharmacological concentration of IDR, Dox, and Vor using a human vascular endothelial cell line
EAhy926, and several myeloid and lymphoid tumor cell lines, respectively,
as laboratory cell models. We further investigated the effects on expression on cell surface TF or PS by using flow cytometer. We investigated the
effects of antineoplastic drugs on whole blood clotting using thromboelastometry.Results: These regimens induced cell surface procoagulant activity (PCA) by exposure of PS on tumor cells and vascular endothelial cells.
IDR and Dox also induced the expression of TF antigen on the surface of
each type of cells. By using thromboelastometry, we observed that clotting
time after IDR treatment for whole blood was shorter than that of nontreatment.Conclusion: These data suggest that these regimens may induce
PCA in vessels through PS exposure associated with cellular apoptosis
and/or increased TF expression on tumor and vascular endothelial cells.
The laboratory methods described in the present study may be useful for
evaluation of the procoagulant effects of antineoplastic drugs.keywords:
chemotherapy, deep vein thrombosis, phosphatidylserine, procoagulant
activity, tissue factor
Classification of acute promyelocytic leukemia by setting high
myeloperoxidase activity area in ADVIA cytogram
PC-34
Significance of the MPXI in the differential diagnosis of APL
and other subtypes of AML
1 Hyogo
Cancer Center, Japan
2 Hyogo
Cancer Center/Kobe Univ.Greduate School of Health Sciences
3 Kobe
Univ.Greduate School of Health Sciences
Introduction:Several studies on the classification of acute myeloid leukemia (AML) using hematological analyzers have been performed. The
mean myeloperoxidase index (MPXI) is rapidly calculated as part of
routine complete blood counts using the hematological analyzer ADVIA
(Siemens). We previously reported that the MPXI values of acute promyelocytic leukemia (APL) patients were higher than those of patients with
other AML subtypes. Strong positivity of myeloperoxidase represents the
typical cytochemical pattern of APL cells, and it is empirically known that
APL cases show a typical cytogram pattern in ADVIA PEROX channel. The
aims of this study were to distinguish APL from other AML subtypes and
to detect morbid neutrophils remaining during the routine classification
of blood cells by setting the high myeloperoxidase activity area in PEROX
cytogram. Methods:We studied medical records of 5 patients with APL
and 14 patients with other AML subtypes and analyzed the frequency
of cells on the high myeloperoxidase activity area using the FCS system
(Siemens). Healthy volunteers were enrolled as controls. We also investigated changes in the number of cells in the setting area for APL patients
during their progress to complete remission.Results:The frequencies of
the cells in the setting area for APL patients were significantly higher than
those for patients with other AML subtypes and for the healthy controls.
In addition, significant reduction in the number of cells in the setting area
for APL patients was observed during their progress to complete remission. Conclusion:These data suggest that information from the cytogram
obtained using the hematological analyzer ADVIA could be useful for the
classification of APL and for the detection of morbid cells remaining during the treatment period.
Introduction: The mean myeloperoxidase index (MPXI) is calculated
during the routine complete blood count (CBC) performed using the autoanalyzer ADVIA2120i. We previously reported that the MPXI in acute promyelocytic leukemia (APL) patients was higher than that in patients with
the other types of acute myeloid leukemia(AML). Furthermore, MPXI was
useful in the follow-up of APL. In this study, we studied a larger samples
size and provided statistical reports showing that the significance of the
MPXI in more detail. Materials and Methods: We measured the MPXI in
26 patients with AML treated at the Hyogo Cancer Center using an appropriately controlled ADVIA2120i. Statistical analyses were performed using
the ANOVA and Tukey’s tests. The significance level for each test was set
at P values less than 0.01. Results: The averages of the MPXI values in the
different groups were as follows: APL group, 33.12 (minimum: 24.1, maximum: 44.8, SD: 7.47); M2 group, 3.48 (minimum: -21.4, maximum: 14.1,
SD: 10.75); M4 group, -2.55 (minimum: -8.5, maximum: 2.9, SD; 5.74); M5
group, 5.03 (minimum: 0.2, maximum: 7.8, SD: 3.62); and M6 group, -0.13
(minimum: -9.4, maximum: 9.2, SD: 9.19). The difference in the average
MPXI values between the APL group and the other AML groups was statistically significant. Discussion: These data suggest that MPXI values are
useful for the diagnosis of APL or the other AML subtypes. Interestingly,
MPXI is effective in the differential diagnosis in the case that is difficult to
diagnose morphologically including APL variant and the M2.
87
Poster Presentation
1 Hyogo
Cancer Center, Japan
2 Hyogo
Cancer Center / Kobe Univ. Graduate School of Health Sciences
3 Kobe
Univ. Graduate School of Health Sciences
Oral Presentation
Takako Tenjin1, Kenji Yonezawa2, Chika Omoto1, Tomoe Yagi1, Aya Adachi1, Masanori Nakamura1,
Toru Murayama1, Mitsuhiro Ito3
Case Conference
Chika Omoto 1, Kenji Yonezawa2, Takako Tenjin1, Asami Kawai1, Azusa Mizuta1, Masanori Nakamura1,
Toru Murayama1, Mitsuhiro Ito3
Symposium
PC-33
Educational Lecture
Extracellular RNA may be released from cells in cases of injury and vascular disease. It has been recognized as a novel procoagulatory and permeability-increasing factor and is counteracted by RNase. RNase family consists of eight members. However, only a certain RNases degrades RNA. We
focused on RNase 1 which exists in plasma and can degrade RNA strongly.
No detailed characterization of the expression and localization of RNase 1
and its inhibitor in the vasculature has been carried out so far. We aimed
to investigate the expression and localization of RNase 1 and RNase inhibitor (RI) in cells which come in contact with blood, such as platelets, mononuclear cells (MNCs), polymorphonuclear cells (PMNs), and red blood cells
(RBCs) compared with human umbilical vein endothelial cell line EAhy926
cells. We further investigated the effect of thrombin on the expression of
RNase 1 and RI in platelets.RNase activity in supernatant derived from
EAhy926 was 50 times higher than blood cells (after 60 min). In cell
lysates, blood cells had RNase activity but EAhy926 had it by far. RNase 1
mRNA derived from EAhy926 was highest at 2.6 times of blood cells, however, RI mRNA was similarly expressed in all cells. We could see RNase 1
and von Willebrand factor (VWF) were partly colocalized in EAhy926 and
platelets. RNase activity derived from lysate of platelets decreased after
thrombin was added, but activity derived from supernatant did not change.
So it seemed that RNase was inhibited by RI combining electrically.We
suggest that at injured vascular site vascular endothelial cells with high
RNase activity are damaged and platelets and leukocytes in the thrombus
show little RNase activity, which may support hemostatic thrombus formation. However, activated platelets and leukocytes, for example at the site of
inflammation, may accelerate pathologic thrombus formation.
Special Lecture
Misae Tsunaka , Reina Arai, Haruka Shinki, Takatoshi Koyama
Laboratory Molecular Genetics of Hematology, Field of Applied Laboratory Science, Graduate School of Health Care Sciences
Tokyo Medical and Dental University, Japan
Invited Lecture
Ayaka Ohashi , Yuichiro Cho, Shizuko Ichinose, Osamu Hoshi, Takatoshi Koyama
Laboratory Molecular Genetics of Hematology, Graduate School of Health Care Sciences, Tokyo Medical and Dental University,
Japan
Keynote Speech
PC-31
Keynote Speech
PC-35
FIR haploinsufficiency switches PKM1 to PKM2 in mice
thymic lymphoma revealed by quantitative proteomic analysis
PC-36
The significance of medical technologists (MTs) participating
in conferences on bone marrow (BM)
Asako Kimura 1, Kouichi Kitamura2, Guzhalinue Arkin2, Mamoru Satoh3, Nobuko Tanaka2,
Fumio Nomura3, Kazuyuki Matsushita2
Nozomi Aoyama , Noriko Fujimaki, Rika Fujii, Yuduru Ito, Miyuki Kikuchi, Fumihiko Goto,
Masakazu Arai, Hajime Horiuchi
1 Dept.
of Medical Science Technology, Narita School of Health Sciences, International Univ. of Health and Welfare, Japan
2 Dept.
of Molecular Diagnosis, Graduate School of Medicine, Chiba Univ.
3 Clinical
Proteomics Research Center, Chiba Univ. Hosp.
NTT Medical Center Tokyo, Japan
Invited Lecture
Special Lecture
Educational Lecture
PurposeIn Japan there are few reports concerning hospitals regularly
holding a conference on BM and where MTs participate. In our hospital,
in addition to hematologists and pathologists, MTs participate and give
explanations on myelogram and morphological findings. In this instance,
we'd like to report on the benefits and significance of MTs participating in
conferences on BM with representative case reports. Example casesCase
1: A 70's man was found to have pancytopenia in a preoperative examination. A BM examination showed there were dysplastic changes for each
lineage from smears, so we calculated the percentage of dysplastic cells of
each lineage with our own table for assessing morphologic dysplasia. The
result showed nuclear hyposegmentation comprised of 13.0% neutrophils,
megaloblastoid changes comprised of 98.0% erythroid precursors, ring
sideroblasts comprised of 45.0% erythroid precursors, and micromegakaryocytes or non-lobated megakaryocytes comprised of 97.0% megakaryocytes. Thus, he was diagnosed as myelodysplastic syndrome refractory
cytopenia with multilineage dysplasia.Case 2: A 60's man, developed Follicular Lymphoma (FL) earlier in his life, was suspected as transformation to
Diffuse Large B-cell Lymphoma. A BM examination showed atypical cells
were not detected in smears, but aggregations of small lymphocyte-like
cells were slightly detected in clot sections and biopsies. Combining these
with the result of immunostaining, he was diagnosed as infiltration of FL
into the BM. DiscussionBM smears are superior in rapidly grasping morphological findings of each cell as Case 1 shows. On the other hand, clot
sections and biopsies are superior in evaluating cellularity and proving
lymphoma cell infiltration as Case 2 shows. Through these cases we could
clearly recognize the merits of each method of BM examination. ConclusionIt's suggested participating in this conference leads MTs to keep motivated and improves the clinicopathological knowledge. Thus, we'd like to
recommend our activities to other institutions.
Introduction:FUSE-binding protein (FBP)-interacting repressor, FIR, is a
transcriptional repressor of c-myc gene. An alternatively spliced form of
FIR is activated as a dominant negative in several cancers. Previously, FIR
hetero knockout mice (FIR+/- ) was prepared, as a dominant negative model of FIR, and showed c-Myc activation in peripheral lymphocytes. Moreover, FIR+/-TP53-/- mice frequently developed thymic lymphoma and/or T
cell type acute lymphoblastic lymphoma (T-ALL). Methods:To examine the
mechanism of FIR’s role in tumor development, the quantitative protein
profile was revealed by six-plex tandem mass tags (TMT) in the above mice
thymic lymphoma model. Furthermore, for potential marker among identified proteins in this study, the protein level was confirmed by western
blotting and the mRNA by qRT-PCR analysis as well.Results:TMT indicated
that 648 proteins were up- or down regulated in mice thymic lymphoma
tissues including transcriptional factors, DNA damage repair proteins, DNA
replication, T-cell activation/proliferation, apoptosis and so on. Notably,
pyruvate kinase subtype M2 (PKM2) protein, but not PKM1, was activated
two times more in mice thymic lymphoma than that of thymus in wild type
mice. qRT-PCR analysis revealed that PKM2 mRNAs of thymic lymphoma
or T-ALL cells in FIR+/-TP53-/- mice were also activated than those of control lymphocytes, sorted by flow cytometory analysis. Notably, knockdown
of FIR by siRNA suppressed hnRNPA1 expression that regulates splicing of
PKM2/PKM1 switch in HeLa cells. Conclusion:These results indicated that
FIR haplo-insufficiency switches alternative splicing of PKM1 to PKM2 in
thymic lymphoma cells prior to T-ALL cells potentially via at least partly
affecting hnRNPA1 expression. Since PKM2 activation is already observed
even in thymic lymphoma tissues, alternative splicing switch from PKM1
to PKM2 is required but not sufficient for T-ALL progression in our mice
model. Together, FIR and its related spliceosomes are vulnerable therapeutic targets for T-ALL and cancers.
Symposium
PC-37
Screening of somatic mutations in JAK2 V617F-negative
myeloproliferative neoplasms using High-Resolusion Meltingcurve analysis
PC-38
Effecacy of CellaVision@ Competency Software as
educational tool for learning blood cell morphology
Asami Naito 1, Etsu Suzuki2, Michikuni Ishijima1, Takayuki Yamamoto1, Ryou Sunaga3, Kazumasa Isobe4
Shumpei Mizuta , Tomoya Minami, Hiroshi Kawabata, Atsuko Ohtani, Momoko Tanaka,
Makirou Ishibashi, Takao Komai
Case Conference
1 Tsukuba
i-Laboratory LLP, Japan
2 Medical
Laboratory of Education and Research
3 LSI
Medience Corporation
4 Department
of Laboratory Medicine, University of Tsukuba
Hyogo Prefectural Amagasaki General Medical Center, Japan
Oral Presentation
Poster Presentation
Investigation of somatic mutations is useful in diagnosis of BCR/ABL-negative myeloproliferative neoplasms (MPN) which includes polycythemia
vera (PV), essential thrombocythaemia (ET), and primary myelofibrosis
(PMF). Especially, JAK2 V617F mutation that causes autonomous proliferation in hematopoietic stem cells exists in 95% of patients in PV, 60%
of patients in ET and PMF. Recently, MPN specific somatic mutation was
reported in JAK2 V617F non-mutated MPNs, CALR exon 9, MPL exon 10
and JAK2 exon 12 mutation. CALR mutation is detected in 25% of ET and
35% of PMF patients, and MPL mutation is 5% of ET and 10% of PMF patients. JAK2 exon 12 mutation is detected in 5% of PV patients. These mutations are exclusive, and the discrimination of them by DNA sequencing at
diagnosis is important for predicting the risk of thrombosis and prognosis.
For example, it was reported that triple negative or CALR mutation in ET
patients were associated with favorable prognosis and lower risk of thrombosis. Therefore, simplified methods are desired for screening of these mutations, using the allele specific PCR is sometimes difficult because these
mutations have various patterns. Herein, we show the method to detect
MPN specific somatic mutations in JAK2 V617F non-mutated MPNs efficiently and rapidly using High-Resolution Melting-curve (HRM) analysis.
The sufficient sensitivity and the specificity of HRM analysis enable us to
detect CALR, MPL and JAK2 exon12 mutations. Hence, HRM analysis is
suitable technic for mutation screening in JAK2 V617F- negative MPNs.
IntroductionContinuous education and cross training are required for
developing and mainitaining laboratory hematology proficiency. CellaVision® Competency Software (CCS) is a program for education and competency testing of manual blood cell differentials in the laboratory. In this
study we evaluated the efficacy of this program.Materials and MethodsWe
recruited 13 medical technologists who had no experience of blood cell
morphology as study participants. The participants performed the skill
test once a week for 4 times. The skill tests were designed to classify 300
kinds of cells, including abnormal and normal cells, into 13 groups. After
testing, the participants were shown their results by the examiner, including a cell by cell comparison and their attainment compared to the other
participants.ResultsAll of the participants improved in their correct answer
rate, with an average increase rate of 16%. At first students incorrectly
classified the cells, but by the 4th week the classification had become more
consistently correct.ConclusionUsing CCS helped inexperienced clinical
laboratory technicians to develop high levels of proficiency in their blood
cell morphological examination accuracy. At our institute we have held a
foreign trainee invitation program since 2011 and have used this software
to train medical technologist from Cameroon, Nigeria, and the Philippines.
They have also appreciated the game-like, active self-learning style of CCS.
88
Assessment of cilostazol inhibition using whole blood platelet
function tests: a translational study
PC-40
Performance of prothrombin time for monitoring of patients
treated with rivaroxaban
Kaneo Satoh 1, Masato Ohta1, Isao Fukasawa2, Kazuya Hosokawa3, Tomoko Oonishi3, Yukio Ozaki4
Daiki Shimomura , Aya Fukuda, Tsuda Katsuyo, Yukinari Okayama, Fumihiko Nakamura
1 University
of Yamanashi Hospital, Japan
2 Kofu
Jounan Hospital
3 Fujimori
Kogyo Co.
4 Fuefuki
Chuo Hospital
Department of Laboratory Medicine, Tenri Hospital, Japan
Special Lecture
Introduction: Cilostazol inhibits phosphodiesterase, increases intracellular
cyclic AMP and inhibits platelet aggregation, particularly in the presence of prostaglandin E1 (PGE1). This study aimed to establish a cilostazol
monitoring assay based on whole blood samples.Methods: Whole blood
samples without or with several PGE1 were assessed using VerifyNow®,
Multiplate® and Total Thrombus-formation Analysis System (T-TAS®).
Results: Cilostazol was no inhibition without PGE1 without PGE1 measured
by the three tests in in vitro and ex vivo studies. In vitro platelet aggregation, in the presence of cilostazol and PGE1, measured by the aspirin, IIb/
IIIa and P2Y12 tests, decreased by 19%, 44% and 9%, respectively. Ex
vivo platelet aggregation, measured by these tests, decreased by 8%, 46%
and 23%, respectively. However, platelet aggregation inhibition was not
assessed by multiplate, thrombus formation or T-TAS both in vitro and
ex vivo. Compared with pre-dosing blood samples, blood samples from
cerebral infarction patients after dosing showed significant platelet aggregation inhibition in the presence of 3 nM (36%) and 10 nM PGE1 (75%).
Conclusion: Platelet aggregation measured by VerifyNow using the IIb/IIIa
test in the presence of 10 nM PGE1 is the most suitable tool for assessing
the inhibitory effects of cilostazol.
Invited Lecture
Background: Prothrombin time (PT) can provide a qualitative assessment
of the relative intensity of anticoagulation by rivaroxaban. More than ten
types of assay are available for the measurement of PT in clinical settings,
but it is not yet fully understood whether their interactions with rivaroxaban are uniform or inconsistent. Furthermore, the lot-to-lot variation of the
assays has not been reported.Methods: We examined 139 blood samples
from patients taking rivaroxaban. We measured PT using five different
commercially available assays. We also evaluated the estimated rivaroxaban concentration using a chromogenic anti-factor Xa assay. We also
examined whether there was lot-to-lot variation for each of the five assays,
using two lots of each assay to measure PT in 34 samples.Results: The median estimated concentration of rivaroxaban was 192 ng/ml (interquartile
range 85 – 284 ng/ml). The correlation coefficient (r) between PT and the
estimated concentrations of rivaroxaban was as follows: Thromborel S, r =
0.768; Thrombocheck PT, r = 0.861; Coagpia PT-N, r = 0.909; Neoplastin
Plus, r = 0.882; and Triniclot PT Excel S, r = 0.870. The gradients of the
regression plots differed more than fourfold, and the standard deviation of
the regression line ranged from 1.001 to 2.980, which tended to be higher
for the assays with the higher regression slope gradients. The existence of
lot-to-lot variation measured using two different lots of each PT assay was
observed with most assays.Conclusion: The estimated concentration of
rivaroxaban varied greatly depending on the PT assay, so the PT measured
in patients treated with rivaroxaban should be interpreted with caution.
Keynote Speech
PC-39
Educational Lecture
Effects of direct oral anticoagulants on general clotting time
assay results
PC-42
Laboratory monitoring of oral anti-factor Xa anticoagulants:
rivaroxaban, apixaban and edoxaban
Yuya Masuda 1, Kazuhiko Matsuno1, Masanao Hatase1, Takayuki Usami1, Hitoshi Shibuya1, Mari Emmi2,
Kaoru Kahata1, Chikara Shimizu1
1 Department
of Clinical Laboratory, Health Sciences Univ., of Hokkaido Hosp., Japan
2 Department
of Internal Medicine, Health Sciences Univ., of Hokkaido
3 Department
of Internal Medicine, Health Sciences Univ., of Hokkaido Hosp.
4 Department
of Microbiology and Immunochemistry, Asahikawa Medical Univ.
1 Hokkaido
University Hospital, Japan
2 Kyowa
Medex Co.,Ltd.
89
Poster Presentation
Introduction: Direct oral anticoagulants (DOACs) have been developed for
prophylaxis and treatment of thromboembolic disorders. When utilized,
laboratory monitoring is not necessary, though their effects on clotting
assay results are not well understood. We investigated the effects of dabigatran, rivaroxaban, and apixaban on the results of clotting assays used in
general examinations.Methods: Each of examined DOAC was separately
added to 4 normal plasma samples (2 commercially available normal
plasma samples, homemade pooled normal plasma from 6 healthy subjects, plasma from a healthy donor) at concentrations of 100 and 400 ng/
ml. Using those samples, we measured activated partial thromboplastin
time (APTT), prothrombin time (PT), fibrinogen (FBG), antithrombin (AT),
protein C (PC), and protein S (PS), as well as the activities of coagulation
factor II, V, VII, VIII, and IX using a clotting time assay.Results: PT and
APTT were prolonged by the addition of dabigatran and rivaroxaban in
a concentration-dependent manner, while there was no such effect with
apixaban. FBG was not influenced by any of the DOACs. PC and PS were
significantly affected by dabigatran in a concentration-dependent manner, and AT was affected by rivaroxaban. The influence of apixaban on
PC, PS, and AT was lower as compared to dabigatran and rivaroxaban.
Finally, each of the tested DOACs exerted influence on the activities of all
examined coagulation factors, except for factor II. Conclusion: We found
that the tested DOACs have varied influence in a concentration-dependent
manner on the results of clotting assays used in general examinations.
Apixaban tended to exert an overall lower level of influence as compared
to dabigatran and rivaroxaban.
Oral Presentation
Introduction: Direct oral anticoagulants (DOACs) have recently become
available for prevention of ischemic stroke in non-valvular atrial fibrillation (NVAF). Sensitive monitoring methods for DOAC therapy are required
in some situations. Rivaroxaban, apixaban and edoxaban have been approved as direct factor Xa anticoagulants. Here, we investigated the utility
of chromogenic anti-Xa assay and standard clotting time assays (PT and
APTT) for monitoring of anticoagulant therapy with each Xa inhibitor using plasma samples from NVAF patients. Materials and Methods: Blood
samples were collected from NVAF patients treated with rivaroxaban,
apixaban or edoxaban. We performed calibrated chromogenic anti-Xa assay using S-2222 (chromogenic substrate for Xa) to estimate plasma drug
concentration and clotting time assays (PT and APTT) to evaluate blood
coagulability. Results: Basic performance of calibrated chromogenic assay using S-2222 was sufficient to determine plasma drug concentration.
Rivaroxaban and edoxaban induced prolongation of PT in a concentrationdependent manner. PT was significantly and strongly correlated with plasma drug concentration; however, sensitivity for drug concentration varied
widely with PT reagents. On the other hand, correlations between plasma
concentrations of rivaroxaban or edoxaban and APTT were weaker than
those for PT. With regard to apixaban, PT and APTT was inadequate for
monitoring of apixaban therapy because of its low sensitivity for plasma
apixaban concentrations. Conclusion: Calibrated chromogenic anti-Xa assay was useful for monitoring of anticoagulation therapy with each Xa anticoagulants. PT, which is a conventional and widely available clotting test,
was also useful for evaluating the efficacy of rivaroxaban and edoxaban.
Case Conference
Sumiyoshi Naito 1, Masahiro Ieko2, Takeshi Suzuki2, Mika Yoshida1, Osamu Kumano2,
Nobuhiko Takahashi2, Mitsuru Moriya3, Nobutaka Wakamiya4
Symposium
PC-41
Keynote Speech
PC-43
Usefulness of clot waveform analysis on the identification of
cause for APTT-prolongation
PC-44
Naoki Tokunaga 1, Chihiro Inoue1, Yusuke Inoue1, Saki Akaiwa1, Hiroko Yoshida1, Takayuki Nakao1,
Toshio Doi2
Tomoko Matsumoto 1, Keiji Nogami2, Yuka Tabuchi3, Koji Kurono3, Nobuo Arai3, Midori Shima2
1 The
Course of Hemophilia Treatment & Pathology, Nara Medical Univ., Japan
2 Dept.
of Pediatrics, Nara Medikal Univ.
3 Sysmex
Corp.
1 Division
of Medical Technology, Tokushima University Hospital, Japan
2 Division
of Clinical Laboratry, Tokushima University Hospital
Invited Lecture
Special Lecture
Educational Lecture
Hemophilia A (HA) results from a deficiency of the procoagulant proteins
factor VIII (FVIII) and is the most common of the severe, inherited bleeding disorders. A newly developed comprehensive coagulation function test,
clot waveform analysis (CWA), can evaluate presence of low level ( ＜ 1
IU/dl) of factor VIII activity (FVIII:C) in patients with hemophilia A (HA-pts).
There is a room to improve for evaluating potential to detect FVIII level
ranging from very low to absent levels, however. To establish the assessment of very low levels of FVIII:C for severe HA-pts by the CWA using a
coagulation analyzer CS-2000iTM , aPTT-based clot waveforms in samples
mixed with various amounts of recombinant FVIII and FVIII-deficient
plasma were monitored. The clot times (CT) were shortened and maximum
coagulation velocity (|min1|) and acceleration (|min2|) were increased
in FVIII dose-dependent manners, ranging from 0.25-100 IU/dl, suggestive of the lowest level of FVIII:C (0.25 IU/dl) for detection. Plasmas with
modestly severe HA-pts (MS-HA; FVIII:C 0.4 ± 0.1 IU/dl; n=5), very severe
HA-pts (VS-HA; FVIII:C ＜ 0.2 IU/dl; n=5), HA-pts with inhibitors (HA-inh;
FVIII:C ＜ 0.2 IU/dl , 51.5 ± 43.7 BU/ml; n=10) were monitored. The CT
was markedly prolonged but showed little significant differences in all
groups. The |min1| were showed of MS-HA (0.66 ± 0.09), VS-HA (0.7 ±
30.12) and HA-inh (0.48 ± 0.06), respectively. The |min2| of HA-inh was
detected in significant low levels with HA-inh (0.01 ± 50.005), compared
with MS-HA (0.022 ± 0.005) and VS-HA (0.02 ± 10.007). The |min1| and
|min2| of HA-inh were lower than those of other groups (p ＜ 0.05). CWA
could provide the evaluation in potential true absence of FVIII and useful
information of prediction the development for FVIII inhibitor in HA-pts.
Introduction: Prolongation of the activated partial thromboplastin time
(APTT) may be caused by a factor deficiency, inhibitor of coagulation factor, lupus anticoagulant, or anticoagulants use. However it is difficult to
identify its cause by using the mixing test. In this study, we tried to identify the cause of APTT-prolongation by using a clot waveform analysis of
the 2nd derivative curve derived from APTT assay on ACL-TOP® system.
Methods: We measured an APTT by using APTT-SP® reagent and ACLTOP 500® (Instrumentation Laboratory) in 24 patient with coagulation
factor deficiency (FD), 6 patients with inhibitor of coagulation factor (FI),
61 patients with lupus anticoagulant (LA), 226 patients treated with anticoagulants including unfractionated heparin (n=75), dabigatran (n=71)
and rivaroxaban (n=80). For the identification of the cause of APTTprolongation, we firstly checked the presence of abnormal waveform
including biphasic or shoulder curve on the 2nd derivative curve. After that,
we calculated other parameters including acceleration time (AT) is the
time from baseline to peak, deceleration time (DT) is the time from peak to
bottom, and DT/AT ratio (D/A ratio). These cut-off values were determined
using ROC curve. Furthermore, we compared the peak value on the 2nd
derivative curve (2DP). Results: We detected abnormal curves in 95.8% of
FD group, 83.3% of FI group and 95.1% of LA group. However, we didn’
t find any abnormal curve in the group treated with anticoagulants. Also,
AT could discriminate FD group from LA and FI group by using the cut-off
value of 7.8 second. Furthermore, D/A ratio could discriminate LA group
from FI group by using the cut-off value of 4.4. Finally, 2DP of FI group
were lower than FD and LA group (P ＜ 0.01). Conclusion: Clot waveform
analysis of the 2nd derivative curve derived from an APTT assay might be
useful for identification of the cause of APTT-prolongation.
Symposium
PC-45
Clot waveform analysis detects very low levels of factor VIII
with severe hemophilia A
A novel diagnostic method APTT clot waveform analysis of
coagulation in patients with lupus anticoagulant
PC-46
Dilute prothrombin time assay to efficiently detect lupus
anticoagulants
Case Conference
Kazunori Kanouchi 1, Hiroto Narimatsu2, Reiko Ohta1, Makiko Sato1, Tosio Watanabe1, Naoki Tokunaga3,
Keita Morikane1
Masato Matsuda 1, Takayuki Matsuto2, Masato Moriyama3, Misao Takano1, Yoshiki Hoshiyama1,
Hirohito Sone4
1 Laboratory
Center for Clinical Investigation, Yamagata University Hospital, Yamagata, Japan
2 Cancer
Prevention and Control Division, Kanagawa Cancer Center Research Institute, Yokohama, Japan
3 Division
of Medical Technology, Tokusima University Hospital Tokushima,Japan
1 Medical
Laboratory Division, Niigata University Medical and Dental Hospital, Japan
2 Division
of Clinical Preventive Medicine, Niigata University Graduate School of Medical and Dental Sciences
3 Division
of Medical Oncology, Niigata University Graduate School of Medical and Dental Sciences
4 Division
of Hematology, Endocrinology and Metabolism, Niigata University Graduate School of Medical and Dental Sciences
Oral Presentation
Poster Presentation
Introduction: Patients with extended activated partial thromboplastin time
(APTT) usually receive the screening tests for lupus anticoagulant (LA),
however; such screening tests needs higher cost with the special equipment. We usually experience the abnormal change in the waveform, as
which the absorbency change during time course was plotted in the test
for APTT. Recently, wave pattern analytical method clot waveform analysis (CWA) was developed, which can be conduct without additional costs.
Method: A total of 85 samples from patients in whom the presence of LA
was measured at Yamagata University Hospital between April 2014 and
March 2015. 85 samples were measured by CWA with APTT reagents on
the ACL TOP® . Positive cut-off level( ＞ 2.10)was determined by using
samples from the healthy donors. The presence of LA was confirmed by
a positive result in STACLOT LA ® or the dilute Russell viper venom time
(DRVVT), according to the diagnostic criteria for anti-phospholipid antibody syndrome (APS) which is considered as the gold standard. Results:
Positive-LA was presented in 35 of the 85 samples (41.2%). The average
APTT values in LA 35 positive cases and 50 LA negative cases were 62.5
± 25.4 seconds and 33.3 ± 3.7 seconds, respectively. The mean values
and standard deviations of the time ratio of acceleration in LA 35 positive cases and 50 LA negative cases were 2.32 ± 0.51 and 1.95 ± 0.27,
respectively. The sensitivity and specificity of APTT method were 91.4%
and 45.0%, respectively. In those 85 patients, 37 were diagnosed with LA
by CWA. The sensitivity and specificity of CWA method were 94.3% and
92.0%, respectively. Conclusion: The sensitivity and the specificity of CWA
to LA diagnosis was higher than those of APTT. LA diagnosis was possible
by the CWA of APTT, with lower cost, which can be conducted in many
laboratories.
Background and Purpose: Phospholipid-dependent coagulation tests are
employed to detect lupus anticoagulants (LA) in clinical laboratories, especially activated partial thromboplastin time (APTT) and dilute Russell viper
venom time (dRVVT) are recommended. These two tests, however, show
considerable variability in diagnostic performance depending on the reagents used. A dilute prothrombin time (dPT) test is also expected to support diagnosis of LA theoretically, but its method has not been established
yet due to variations of analytical techniques and reagent's properties. This
study aims to establish an efficient procedure of the dPT test for LA detection using Thromborel® S reagent which is widely used in Japan. Methods: Lyophilized Thromborel® S was reconstituted with distilled water,
and then diluted doubling from 1:10 to 1:320 with 20mmol/L CaCl2. Employing standard PT assay program, the optimal dilution ratio for the dPT
test was determined for various samples; coagulation factor(s) deficient,
LA positive, heparin- and warfarin-containing plasmas. The procedure
using diluted Thromborel® S was applied to samples with or without LA
obtained from patients with various diseases including antiphospholipid
syndrome (APS). Results: The dPT was prolonged depending on the dilution ratio for all samples. The optimal dilution ratio to distinguish LA from
factor deficient was 1:160. Heparin-containing and warfarin-containing
samples also showed extreme dPT prolongation. But the prolongation by
heparin could be corrected by addition of protamine. The prolongation by
warfarin was clearly discriminated from LA using the 'dPT-ratio' which is
the ratio of dPT value at 1:160 dilution to that without dilution. The dPTratio of LA was higher than that of warfarin samples. Moreover, the dPTratio increased in APS patients whose APTT values were not prolonged.
Conclusion: The dPT-ratio obtained from the assay procedure using
diluted Thromborel® S might be applicable to patient's samples showing
prolonged dPT and useful to detect LA.
90
Usefulness of the platelet parameter
Mean platelet volume and Mean platelet component obtained
by ADVIA2120i as convenient indicator for MDS diagnosis
PC-48
Influence of heat shock protein 72 on platelet aggregation in rodents
HSP72 promotes platelet aggregation in the presence of various
platelet activators
Ryota Masutani 1, Toshiyuki Ikemoto1, Ayako Maki1, Hiroko Tanada1, Yoshinori Iwatani2,
Yasuhito Okada3
Hideaki Suzuki 1, Yuuko Kosuge1, Koji Kobayashi1, Naohito Ishii2, Naoyoshi Aoyama3,
Kazuhiko Ishihara1, Takafumi Ichikawa2
1 Dept.
of Central Laboratory, Osaka Medical College Hosp., Japan
2 Dept.
of Biomedical Informatics, Div.of Health Sciences, Osaka Univ. Graduate School of Medicine
1 kitasato
Junior College of Health and Hygienic Sciences, Japan
2 Kitasato
University Graduate School of Medical Sciences
3 Kitasato
University School of Medicine
Atsushi Ogasawara 1, Yumiko Tanaka1, Yukari Shirasugi2, Kiyoshi Ando2, Satomi Asai3,
Hiromichi Matsushita3, Hayato Miyachi3
Mean Platelet Volume is a beneficial prognostic biomarker in
patients with type 2 diabetes
Symposium
PC-50
Educational Lecture
A simple method based on peripheral blood parameters for
early diagnosis of chronic myelogenous leukemia
Special Lecture
PC-49
Background: Heat shock protein 72 (HSP72) blood levels positively correlate with increased numbers of clots in coronary arteries of patients
with acute myocardial infarction (MI). The clinical significance of elevated
HSP72 levels is not well understood; however, platelet aggregation is
known to contribute to the formation of thrombi in ruptured plaques
during MI. Aim: This study aimed to understand the role of HSP72 in
thrombosis by analyzing the effect of HSP72 on platelet aggregation.
Method: Platelet-rich plasma (PRP) was prepared from the blood of
13-week-old male SD rats. PRP platelet aggregation activity was measured
in the presence of platelet activators: ADP, collagen, ristocetin, or TRAP6. Changes in aggregation were estimated by simultaneous addition of
recombinant HSP72 and anti-HSP72 antibody. Serum of ApoE-deficient
12-week-old male mice fed a high-fat diet was collected and HSP72 was
quantified. Heart coronary arteries were immunostained with an antiHSP72 antibody. Statistical significance was evaluated using Dunnett's
multiple comparison test and JMP12 software.Results: Addition of HSP72
increased platelet aggregation in a dose-dependent manner. The surge in
platelet aggregation was observed in the presence of low amounts of ADP,
collagen, ristocetin, and TRAP-6, but was reduced by addition of the antiHSP72 antibody. Furthermore, hyperlipidemic mice presented elevated
serum HSP72 levels and stained positive for HSP72 during thrombosis.
Conclusion: Platelet aggregation was found to increase in the presence
of HSP72 and low amounts of platelet activators in rat PRP. In addition,
HSP72 was present during thrombosis in hyperlipidemic mice. We suggest
that increased HSP72 in the blood promotes platelet aggregation during
formation of thrombi on failed plaques. Thus, HSP72 blood levels may
help predict MI.
Invited Lecture
INTRODUCTIONIt is well known that degranulated platelets or giant platelets may appear in the peripheral blood of the patent with myelodysplastic
syndrome (MDS). In general, diagnostic criteria based on morphological characteristics have a defect in its reproducibility because of a wide
variability among observers.Thus, those morphological abnormalities of
platelets may be difficult to use as indicators to discriminate MDS from
other hematological abnormalities. Here, we report that the mean platelet
component concentration (MPC) and the mean platelet volume (MPV),
which can be obtained by the ADVIA2120i (Siemens Inc.), are useful and
convenient indicators for MDS diagnosis.MATERIALS AND METHODSMPC
and MPV of platelets in the peripheral blood obtained from 38 cases with
MDS, as well as those obtained from 1256 healthy controls, were analyzed,
and the cut-off value to discriminate the MDS group from the healthy
control was determined.The association of the MPC and MPV values with
the morphological abnormalities of platelets was confirmed by the microscopic observations of smear specimens. In addition, degranulation of
platelets in patients with MDS was observed using an electron microscope.
RESULTThe MPC of MDS patients (median 23.0g/dL) was significantly
decreased compared with that of healthy controls (median 26.5g/dL). In
smear specimens of the peripheral blood, degranulation of platelets was
detected in MDS patients. The MPV of MDS patients (median 10.7fL) was
significantly elevated compared with that of healthy controls (median
7.8fL). In smear specimens, giant platelets were detected in MDS patients.
When the cut-off value was determined as MPC ＜ 25.3 g/dL as well as
MPV ＞ 10.0fL, each value of the area under curve, sensitivity, specificity, positive predictive value and negative predictive value showed 0.920,
78.9%, 99.9%, 96.8% and 99.4%, respectively.CONCLUSIONThe MPC and
MPV obtained by the ADVIA 2120i are considered as useful and convenient indicators for MDS diagnosis.
Keynote Speech
PC-47
Inoue Hiroyuki , Iio Hiroki, Takeno Kengo, Saito Mayumi, Kouchi Kumiko, Yoshimura Yutaka
Dept.of Central Clinical Labolatory, Nara Prefecture General Medical Center, Nara, Japan
91
Poster Presentation
Introduction:Chronic myelogenous leukemia (CML) is a representative disease of myeloproliferative neoplasms (MPN), featured by increase of WBC
and platelet counts at the time of onset. The diagnosis of CML essentially
requires detection of BCR-ABL1. However, its result is not always promptly
available. In order to develop a simple and rapid method for the early diagnosis of CML, we analyzed the peripheral blood parameters of the patients.
Methods:A total of 115 adult CML patients diagnosed at Tokai University
Hospital between 2004 and 2016 were enrolled. As a control, 541 adult
patients with leucocytosis ＞ 8.0 × 109 /L and 193 patients with other
types of MPN were included. Peripheral blood parameters at the first visit
were studid. A reciever operating characteristic (ROC) curve for each parameter was constructed, and the optimal cut-off value was detemined. For
NAP score, statistical difference between CML and MPN, and the cut-off
value were determined. Result:The median (range) of peripheral blood parameters of CML were 38.0 (6.0-435.3) × 109/L for WBC, 479 (93-2395)
× 109/L for platelet, 5.6 (0-174.1) × 109/L for immature granulocytes
(IG), 2.56 (0-25.85) × 109/L for basophil and 66(10-214) for NAP score.
The area under curve of ROC curve was the highest in basophil at 0.981,
followed by IG at 0.975, WBC at 0.912 and platelet at 0.747. The cut-off
value was 0.43 × 109/L, 0.48 × 109/L, 20.95 × 109/L and 326.0 × 109/L,
respectively. With the cut-off value of basophil, 93.0% (107/115) of CML
was screened. NAP score was statistically different between CML and MPN
(p ＜ 0.001). The cut-off value was 127.5.Conclusion:In screening of CML,
the absolute number of basophil was the most suitable marker. When the
BCR-ABL1 is not promtly available, early screening of CML and its differentiation from other types of MPN can be made simply by a stepwise use
of the absolute number of basophil, followed by NAP score.
Oral Presentation
Background and ObjectiveMean Platelet Volume (MPV), the most commonly used measure of platelet size, has recently been reported to be
associated with a risk for cardiovascular disease. Then, MPV is thought to
be potentially useful as a predictor of cardiovascular risk. Cardiovascular
events in patients with type 2 diabetes mellitus may also be associated
with accelerated platelet activity shown in high MPV. So, we hypothesized
that increased MPV may be related to the pathogenesis of type 2 diabetes and can be a beneficial biomarker for reflecting platelet activity in
the setting of diabetic vascular complications.Materials and MethodsWe
classified the patients into three groups with diagnostic criteria (fasting
glucose level and HbA1c); non-diabetic group (Control group), patients
with impaired glucose tolerance (pre-DM group), patients with type 2 diabetes (T2DM group). MPV was measured by SIEMENS' hematology analyzer ADVIA2120i or ADVIA120 whose measurement principle was flow
cytometry-based analysis of blood.ResultMPV was significantly increased
in T2DM group as compared to Control group. In pre-DM group, MPV
was already significantly increased. We also found a tendency that fasting
glucose level and HbA1c was elevated as MPV was increased. To determine whether an association between MPV and arteriosclerosis in type 2
diabetes, we next measured Cardio Ankle Vascular Index (CAVI) by using
pulse wave velocity and Intima-Media Thickness (IMT) by performing carotid artery echo. Then, T2DM group showed significant increase of CAVI
and IMT as compared to Control group. Moreover, MPV showed a positive
correlation with CAVI and IMT. ConclusionIt was suggested that platelet
activity of diabetic patients become more reactive due to increased MPV.
Furthermore, enhanced cardiovascular risk in type 2 diabetes may be a
result of high MPV. Therefore, MPV can be a potentially beneficial prognostic biomarker of cardiovascular complications in patients with type 2
diabetes.
Case Conference
1 Clinical
Laboratory Center, Tokai University Hospital, Japan
2 Division
of Hematology/Oncology, Department of Medicine, Tokai University School of Medicine
3 Department
of Laboratory Medicine, Tokai University School of Medicine
Keynote Speech
PC-51
Immature reticulocyte fraction may predict the hematopoietic
recovery after hematopoietic stem cell transplantation
PC-52
The serum survivin levels and pathological findings of diffuse
large B-cell lymphoma
Noriyuki Okubo 1, Aya Sato1, Shingo Sugawara1, Asami Sasaki1, Mitsuaki Nagasawa1, Hideo Harigae2,
Mitsuo Kaku2
Yumiko Taguchi 1, Machiko Kawamura2, Junko Okabe- Kado3, Haruna Tanaka1, Atsuko Kawarai1,
Yu Nishimura4, Masafumi Kurosumi4, Toshihiro Iwata1
1 Tohoku
University Hospital, Japan
2 Tohoku
University Graduate School of Medicine
1 Clinical
laboratory, Saitama Cancer Center, Japan
2 Hematology,
Saitama Cancer Center
3 Research
Institute for Clinical Oncology, Saitama Cancer Center
4 Pathology,
Saitama Cancer Center
Invited Lecture
Special Lecture
Educational Lecture
Introduction: Hematopoietic stem cell transplantation is one of the important therapies in hematopoietic malignancies. It is important to assess
engraftment and hematopoietic recovery after hematopoietic stem cell
transplantation. Since immature reticulocyte fraction (IRF) and immature
platelet fraction (IPF) indicate hematopoietic recovery, it is possible that
IRF and IPF are useful predictors for hematopoietic recovery after hematopoietic stem cell transplantation.Methods: Subjects were 15 patients who
underwent hematopoietic stem cell transplantation in Tohoku University
Hospital from June 2015 to January 2016. The median age of subjects
was 50 years old (range, 32-66 years old). We measured neutrophil, reticulocyte, platelet, IRF and IPF in peripheral blood sample by automated
hematology analyzer XN-1000 (Sysmex). Engraftment was defined as the
first day of neutrophil ＞ 0.5 × 109/L, reticulocyte ＞ 1.0%, and an unsupported platelet count ＞ 20 × 109/L or 50 × 109/L for three consecutive
days. Wilcoxon signed rank test was used to determine the difference in
day of neutrophil engraftment, reticulocyte engraftment, platelet engraftment, IRF ＞ 10%, IPF ＞ 2.0% and IPF ＞ 5.0%. Also, we analyzed variation
pattern of each parameter by Friedman’s test and multiple comparison
using Bonferroni correction. Results: The median day of neutrophil, reticulocyte and platelet engraftment after hematopoietic stem cell transplantation was 15.0 (interquartile range, 12.5-17.0), 19.5 (15.3-27.8) and 28.0
(20.8-72.8), respectively. The median day of IRF ＞ 10%, IPF ＞ 2.0% and
IPF ＞ 5.0% was 15.0 (13.0-19.0), 18.0 (15.0-21.0) and 21.5 (18.5-35.8),
respectively. The median day of neutrophil engraftment, IRF ＞ 10% and
IPF ＞ 2.0% was shorter than that of reticulocyte and platelet engraftment
(p ＜ 0.05, respectively). IRF was significantly increased on day 15 after
hematopoietic stem cell transplantation (p ＜ 0.01). Neutrophil and IPF
were not significantly increased on day 15 after hematopoietic stem cell
transplantation.Conclusion: Our results indicated that IRF may be a useful
predictor for hematopoietic recovery after hematopoietic stem cell transplantation.
Symposium
PC-53
BACKGROUND: Survivin is a member of the inhibitor of apoptosis (IAP)
family, and thought to contribute to cancer cell survival. Over expression
of survivin was found to correlate with poor prognosis in patients with
various tumors. There are two subtypes of diffuse large B-cell lymphoma
(DLBCL); germinal center B-cell (GCB) and activated B-cell (ABC) subtypes.
ABC subtype is associated with worse outcomes when treated with standard therapy. The aim of this study is to investigate serum survivin levels
and their correlation with other biomarkers or pathological findings in patients with DLBCL, and to demonstrate their clinical significance. METHODS: 26 patients diagnosed as having DLBCL aged from 15 to 84 years,
were investigated. Serum samples were obtained at diagnosis and relapse.
Serum survivin levels were determined using ELISA (R&D Systems), and ＞
20 pg/ml were defined as positive. RESULTS: Positive patients were 0/3
at stage I, 1/6 at stage II, 0/4 at stage III, 7/10 stage at stage IV, and 3/3 at
relapse. The levels were not correlated with other serum biomarker levels;
such as LDH or sIL-2R and DLBCL subtypes. Serum survivin levels were
higher in stage IV patients at diagnosis and in patients at relapse. GCB
or ABC subtype was not correlated with serum survivin levels. However,
there is one case with CD5+ ABC lymphoma at stage II showing a higher
level of survivin. CD5+ DLBCL is a distinct subgroup of DLBCL with poor
outcomes. CONCLUSIONS: The results of this study suggested that serum
survivin levels might be useful for the early detection of relapse and bone
marrow involvement; i.e., stage IV. Serum survivin levels were not correlated with other biomarker levels or pathological subtypes with an exception of survivin-positive CD5+ ABC lymphoma. Further studies are needed
to identify the clinical significance of serum survivin levels.
Basic evaluation of platelet aggregation measuring system
with the fully automated coagulation analyzer
PC-54
Evaluation of the coagulation factor VIII and IX activity
measurement using the chromogenic reagent
Case Conference
Yukari OMORI , Hidekazu ISHIDA, Yuriko KATANO, Rina TAUCHI, Nobuyuki FURUTA, Hiroyasu ITO,
Mitsuru SEISHIMA
Madoka Inoue 1, Reiko Shizuka1, Ayako Sato1, Masaki Hayakawa1, Tetsuo Machida1,
Katsuhiko Tsunekawa2, Hiromi Koiso3, Masami Murakami2
Div. of Clinical Laboratory, Gifu University Hosp., Japan
1 Clinical
laboratory center, Gunma University Hospital, Japan
2 Department
of Clinical Laboratory Medicine, Gunma University Graduate School of Medicine
3 Infection
Control and Prevention Center, Gunma University Hospital
Oral Presentation
Poster Presentation
Introduction:Platelet aggregation test is useful for the diagnosis of platelet
function and the efficacy assessment of antiplatelet drug. Recently, the fully automated coagulation analyzer with light transmission aggregometry
has been launched. In this study, we evaluated the basic performance of
this equipment system.Methods:We used pooled plasma and plasma of patients treated with antiplatelet agents in the current study. Adjusted platelet-rich plasma (PRP) with a platelet count of 200 x 109/L was obtained by
diluting PRP with the platelet-poor plasma.Platelet aggregation test was
performed on coagulation analyzer CS-2500 (Sysmex Corporation, Japan)
and MCM HEMA TRACER 712 (MC Medical Inc., Japan; H-TRACER) was
used as a reference method. Revohem ADP and Collagen (Sysmex Corporation, Japan), MCM ADP and MCM Collagen H (Laboratory & Medical
Supplies) were used as agonists in CS-2500 and H-TRACER, respectively.
Within-run precision (n=10) was performed in CS-2500 with each agonist.
Interference Check A Plus (SYSMEX Corporation, Japan) was added to
PRP.Results:The coefficient of variations in within-run imprecision by ADP
3.0 µM and collagen 0.5 µg/mL showed less than 7% in CS-2500. In comparison study (n=32), the regression equation and correlation coefficient
in ADP 1.0 µM and 10.0 µM were Y = 1.11X+2.86 and r = 0.865, and Y =
0.93X+8.61 and r = 0.812, respectively. Those of collagen 2.0 µg/mL and
5.0 µg/mL were Y = 0.85X+11.94 and r = 0.929 and Y = 0.81X+15.3 and
r = 0.896, respectively. Only hemolysis affected the aggregation test in the
interference study. Conclusion:Our present study showed that the ability
of platelet aggregation measurement on CS-2500 was satisfactory. Therefore, it was suggested that this automated technique would contribute to
the standardization of platelet aggregation test.
Introduction: Recently, the use of the recombinant agent for hemophilia
treatment with extended half-life was approved in Japan. The coagulation factor activity of patients treated with the drug is reported to show
different results between one-stage clotting assay and the chromogenic
assay. We report the performance evaluation of coagulation factor VIII (F8)
and factor IX (F9) using the chromogenic assay reagent. Materials and
Methods: We evaluated 1) Repeatability, 2) Reproducibility, 3) Linearity, 4)
Minimum detectable sensitivity and 5) Effect of the interfering substance.
The activity of F8 and F9 in commercial plasma was measured using
Biophen FVIII:C and Biophen Factor IX as a reagent on the automated
coagulation analyser CS-5100 (Sysmex). Results: 1) F8 was 0.3-3.8%, F9
was 0.6-1.1%. 2) F8 was 4.5-4.6%, F9 was 4.1-4.6%. 3) Both F8 and F9 low
range were good to around 0.0%. 4) F8 was 0.8%, F9 was 0.3%. 5) Both F8
and F9 were hardly affected to 680mg/dL hemoglobin, 40mg/dL bilirubin
and 400mg/dL chyle. Conclusion: Each basic examination about the performance of the F8 and F9 activity measurement based on chromogenic
assay showed good results, and the reagents will be useful in routine clinical testing.
92
Prediction of Myeloid Engraftment after Hematopoietic
Stem Cell Transplantation Using an ADVIA2120i-derived
parameter, BP ratio
PC-56
Usefulness of RET channel in XN-Series automated hematology analyzers
RBC-O, another RBC count parameter, by RET channel provides accurate RBC
count for blood specimens with cold hemagglutination
Tenri Yorozusoudansho Hospital, Japan
Background:- ＞ In hematopoietic stem cell transplantation (HSCT), since
graft rejection lead to a fatal clinical course, early and objective prediction
of engraftment is desired. ADVIA2120i, one of the blood cell analyzer used
in Japan, automatically calculates parameters, %Blast suspects and %PMN.
We defined a new parameter BP ratio as %Blast suspects divided by
%PMN, and verify whether it can predict the engraftment after HSCT. Furthermore, we investigated the relationship of BP ratio and morphological
characteristics of the cells appearing on and after the day of peak BP ratio.
Method:- ＞ Blood samples of 34 engrafted and one rejected patients after
HSCT were analyzed using ADVIA2120i. BP ratio was calculated and monitored. The cut-off value of BP ratio was determined using ROC curve. Morphological and surface marker-oriented characterization was performed
using flow cytometric analysis on CD45 gating, and May Giemsa and Esterase staining on the cyto-spin samples.Results and Discussion:- ＞ The
cut-off value of BP ratio for predicting the engraftment was determined as
0.20%. To rule out the false positive due to low cell number, Blast suspects
＞ 4/µL was defined as a prerequisite. Whereas BP ratio in all engrafted
patients exceeded the cut-off value seven days before the engraftment,
BP ratio did not reached the cut-off value in the rejected patient. CD45dim
CD11b- CD36- HLA-DR+ blasts were dominant at the day of peak BP ratio.
After that, in accordance with the decrease of BP ratio, CD15+ myeloid progenitors and CD36+ promonocytes increased. These results suggests that
BP ratio reflects the kinetics of CD45dim CD11b- CD36- HLA-DR+ blasts, and
the transient increase of this cell population in early phase may be related
to the engraftment.
Introduction: The XN-Series automated hematology analyzers (Sysmex)
have two red blood cell (RBC)-measuring channels; one is an impedancebased RBC/platelet (PLT) channel used to examine RBC count (RBC-I) and
the other is a flow cytometric reticulocyte (RET) channel mainly used for
reticulocyte measurements. Although the RBC/PLT channel provides accurate RBC counts in most cases, analyzers show false low RBC counts when
analyzing agglutinated blood. Samples are immediately re-examined after
warming. However, another RBC count parameter, RBC-O, is measured
using the RET channel and can be used as a reference RBC count for specimens with cold hemagglutination.Materials and Methods: The correlation
between RBC-I and RBC-O was examined using 56 blood specimens with
no agglutination (normal specimens). These parameters and mean corpuscular volume (MCV) and mean corpuscular hemoglobin concentration
(MCHC) were then compared in four specimens with cold hemagglutination between two conditions: at room temperature (RT) or immediately after warming at approximately 40℃ .Results: An excellent correlation (y =
0.9959x – 0.0075, R2 =0.9954) within ± 0.15 × 106/µL was demonstrated between RBC-I and RBC-O in normal specimens. In the four specimens
with cold hemagglutination, RBC-I and hematocrit were significantly lower
at RT than at 40℃ (– 0.49 to – 0.91 × 106/µL and – 4.2 to – 7.5% respectively). On the other hand, RBC-O was similar (– 0.04 to +0.15 × 106/µL)
under both conditions. MCV and MCHC were falsely elevated (+1.7 to +9.0
fL and 37.0 – 55.1 g/dL) at RT, whereas MCHC recovered to within normal
range (32.3 – 34.9 g/dL) after warming.Discussion: The robustness (accuracy) of RBC-O may be because of the reaction temperature, 41℃ , in the
RET channel. Therefore, if RBC-I is significantly lower than RBC-O at RT,
the specimen will have cold hemagglutination. If there is no discrepancy
between RBC-I and RBC-O after warming, the specimen will not be affected
by cold hemagglutination.
A case of TTP
ELISA assay of ADAMTS13 and inhibitor were useful for the
early treatment
PC-58
Toshiyuki Niiya 1, Tatsuya Nishimiya1, Kazushi Tanimoto2, Taichi Azuma2, Takaaki Hato2
A rare case of acquired von Willebrand syndrome associated
with systemic lupus erythematosus
Symposium
PC-57
Educational Lecture
Tokushima University Hospital, Japan
Special Lecture
Naomichi Tsuchiya , Kimiko Hioki, Katsuyo Tsuda, Masashi Shimada, Yukinari Okayama,
Fumihiko Nakamura
Invited Lecture
Akishige Ikegame , Yusuke Inoue, Hiroshi Kanamori, Chihiro Inoue, Satiko Ogasa, Takayuki Nakao,
Toshio Doi
Keynote Speech
PC-55
Taku Nonaka , Michiko Yamazaki, Emi Suzuki, Naoko Maruyama, Yumiko Sakai, Ayano Yoshihara,
Yuuki Tanaka, Takashi Yamada
Department of Medical Technology, Nagaoka Red Cross Hospital, Japan
93
Poster Presentation
Acquired von Willebrand syndrome (AVWS) is a rare acquired bleeding disorder that occurs in association of various underlying diseases. There have
been few reports about this syndrome and systemic lupus erythematosus
(SLE) is an infrequent cause of AVWS. Here, we report on our experience
of a rare case of AVWS associated with SLE. The patient was a 67-year-old
woman on dialysis for chronic renal failure. She was referred to a department of gastroenterology because upper gastrointestinal bleeding was
suspected. Although she underwent upper gastrointestinal endoscopy, it
led to hemorrhage from upper pharynx and caused uncontrolled bleeding.
Therefore, she was eventually referred to a department of hematology due
to bleeding tendency. There was no family history of bleeding diathesis.
Initial laboratory examinations revealed a prolonged activated partial
thromboplastin time at 63.5sec while prothrombin time was within reference range. Subsequent examinations demonstrated that factor VIII activity was markedly decreased at 6.2% and von Willebrand factor ristocetin
cofactor activity (VWF:RCo) was severely decreased at less than 0.77%.
From the above results, we performed mixing study, so that the patient’s
plasma inhibited VWF:RCo in the presence of normal plasma, suggesting
the presence of an inhibitor against VWF in the patient’s plasma. In fact,
the presence of antibody against VWF was detected by enzyme linked immunosorbent assay and electrophoretic analysis revealed low levels of the
high-molecular weight VWF multimer. In addition, the patient met criteria
for SLE due to biopsy-proven lupus nephritis, lymphopenia, positive antinuclear antibody, positive anti-Smith antibody and hypocomplementemia.
Therefore, these findings resulted in the diagnosis of AVWS associated
with SLE. Although rare, if we encounter patients with bleeding tendency,
we should always keep in mind the possibility of AVWS.
Oral Presentation
ADAMTS13 is a von Willebrand Factor (vWF) cleaving enzyme, and the
patient, with congenital defects of ADAMTS13 or aquired antibodies
against it, develops TTP. Here, we report a representative case of TTP
patient to whom ELISA assay of ADAMTS13 and inhibitor were useful for
the early treatment of TTP. The patient is an 80 year old man, he had discomfort of the chest and appetite loss. He was undergoing treatment for
hypertension. He had prostatic cancer and appendicitis in his past history.
In Blood cell count, 11,600/ Μ L of WBC, anemia and thrombocytopenia
were recorded. He had slight consciousness disorder but could converse
without dysarthria. And petechiae were evident on the body and extremities. Head CT examination showed slightly brain atrophy. The data at
admission showed anemia, thrombocytopenia and hyperplasia of reticulocytes. LD was elevated and haptoglobin was not detectable. Schizocytes
were seen on smear slides. We measured his activation of ADAMTS13
with ELISA kit and found the activation was less than 1% compared with
normal control. Next, the activation of ADAMTS13 inhibitor was measured,
and the data showed 3.2 Bethesda Units. After treatment with plasma
exchange and PSL, laboratory data was improved except of Schizocytes,
which were detected until day40. But in this patient, TTP relapsed on
day230. At that time, ADAMTS13 activity was less than 1% and Inhibitor
was 19.4 Bethesda Unit, it was higher than day1. Relapse TTP was refractory, so he was treated with PSL, plasma exchange plus CD20 antibody,
“Rituximab”. These treatment appear to be effective for this patient, so far.
Case Conference
1 Department
of Clinical Laboratory, Ehime University Hospital, Japan
2 The
first department of Internal Medicine, Ehime University Hospital
Keynote Speech
PC-59
PC-60
A case report of acquired factor V inhibitor.
Takeshi Osawa 1, Kazuo Kawasugi2, Kimiko Nogi1, Mayumi Matsuzawa1, Taiji Furukawa2
Kazuhiro Itishita , Tiduko Nonaka, Erina Sibata, Yasuyo Hirakawa, Kaoru Urakawa, Kazuma Taniguti,
Yasusi Kawabuti, Katuyuki Nagatoya
1 Teikyo
University Hospital, Japan
2 Teikyo
University School of Medicine
JOHAS Osaka Rosai Hospital, Japan
Invited Lecture
Special Lecture
Educational Lecture
We report a case of a patient of factor V inhibitor. The patient is a male in
his seventies with a history of cerebral infarction and of oral anti-platelet
drug. Because of a coxalgia on his right side and swelling with subcutaneous hemorrhage, he became unable to walk. Then he was sent to our
hospital.Findings on admission went as follows : WBC 12.1 × 109/L, Hb 6.4
g/dL, PLT 321 × 109/L, prolonged prothrombin time (PT) 56.2 sec, PTINR 4.08, activated partial thromboplastin time (APTT) 177.8 sec, fibrinogen 457 mg/dL. Infusion of fresh-frozen plasma and administration of
vitamin K could’t correct coagulopathy. Prothrombin time mixing studies
suggested presence of an inhibitor. Assays for specific coagulation factors
and factor-inhibitors were performed, which confirmed the presence of
a factor V inhibitor (8.9 BU). Oral prednisolon was administered on 12th
day, and the coagulation parameters improved gradually. Consequently,
these parameters fell within reference interval on 39th day.According to
a systematic review (Franchini and Lippi, 2011), 159 cases of factor V
inhibitor were emerged on literature, and most of them (78 cases) were
related to bovine thrombin usage. However, other causes were increasing
due to the increased usage of recombinant thrombin. These include autoimmune affliction, cancer, tuberculosis or human immunodeficiency virus
(HIV) infection, blood transfusion, and antimicrobial drugs. In our case,
bovine thrombin usage, cancer, infection and autoimmune affliction were
deniable. Meanwhile, antimicrobial drugs were used and blood transfusion was performed. In addition, the patient had taken an oral anti-platelet
drug (clopidogrel) which is reported to be one of the cause of acquired
hemophilia A. Clinical course and drug usage history suggested the antiplatelet drug would be a one of the most possible cause of acquired factor
V inhibitor in this case.
Symposium
PC-61
One case of the articular rheumatism that developed a
hemophagocytic syndrome
IntroductionHemophagocytic syndrome (HS) is occasionally associated with autoimmune diseases, including juvenile rheumatoid arthritis
(RA) and Still’s disease. However, HS related to adult RA has been rarely
reported. CaseA male patient in his 70s who had developed RA 4 years
before underwent methotrexate therapy for RA deterioration in June. The
initial dose was 6 mg/week, which was increased to 8 mg/week in July. He
was diagnosed with pancytopenia in October. On admission to the Osaka
Rosai Hospital, white blood cell (WBC) count was 0.5 × 109/L, red blood
cell (RBC) count was 215 × 1012/L, and platelet count was 46 × 109/L.
Because pancytopenia was assumed to be induced by methotrexate, it was
discontinued and instead folate- and granulocyte colony-stimulating factor,
antibiotics, and gamma globulin preparations were administered. He also
underwent RBC and platelet transfusions. Nonetheless, WBC count did not
increase, and he developed thrombocytopenia. Bone marrow aspiration
was performed on day 5. Total nucleated cell count abnormally decreased
to 0.7 × 109/L, and 17.6% of them were macrophages. Hemophagocytosis
was diffusely observed by microscopy. The patient was diagnosed with
HS, received intravenous methylprednisolone, and gradually recovered.
DiscussionHS associated with viral infections can occur among adult RA
patients. However, a viral infection appeared unlikely in this case. Methotrexate was frequently selected before the onset of an HS coincident with
RA, meaning that RA activity is exacerbated in these patients. In this case,
HS appears to have been caused by severe RA or a viral infection (except
Epstein – Barr virus or cytomegalovirus during pancytopenia) because of
methotrexate. Drug-induced pancytopenia associated with HS can become
quite severe and can be lethal with late diagnosis.ConclusionHS coincident
with RA was evaluated. In pancytopenia patients, bone marrow aspiration
should be promptly considered for early HS diagnosis.
A case of sicklecell disease with abnormally low level of
HbA1c by the HPLC method
PC-62
Hisami Baba 1, Takayoshi Tokutake2, Youhei Kitaya2, Kozue Tokita2, Michie Nozaki2, Hisae Asati2,
Shouhei Nakata3, Hikaru Kobayashi4
Development of an internal quality control application for
immature granulocyte differential
Naoya Ichimura , Ayako Itoi, Yuuki Ohkubo, Yuuki Koda, Michio Hagihara, Shuji Tohda
Clinical Laboratory Dept, Medical Hosp, Tokyo Medical and Dental Univ., Japan
Case Conference
1 Nagano
redcross Hospital, Japan
2 Dept.of
Clinical Labo.Nagano Redcross Hosp.
3 Dept.of
Blood Transfusion.Nagano Redcross Hosp.
4 Div.of
Hematology.Nagano Redcross Hosp.
Oral Presentation
Poster Presentation
Objectives: Thus far, there are no internal quality control (IQC) procedure for counting the percentage of immature granulocytes (IG), which
consist of myeloblast, promyelocyte, myelocyte, and metamyelocyte, on
blood smears. We developed an IQC application to assess the examiner's
performance and to standardize criteria for IG classification. The aim of
this study is to evaluate the inter-examiner agreement rates of correctly
recognizing IG.Methods: The IQC application was developed using Microsoft Access. Photo images of leukocytes including IG were downloaded
from website of the Japanese Society for Laboratory Hematology (JSLH,
http://www.jslh.com), which are opened to standardize the differential of
leukocyte count. The application automatically selects 50 images daily and
randomly, which include at least five images of myeloblast, promyelocyte,
myelocyte, metamyelocyte, atypical lymphocyte, and normal lymphocyte.
Three examiners observed those images on computer monitors and decided the cell type. In early period (until 14 days after introducing this application) and late period (56 days after early period), the agreement rates
of each cell type chosen by each examiner were assessed using Cohen's
κ coefficient.Results: The agreement rate of the cell type chosen by each
examiner increased with trial frequencies, which was finally more than 90
% except myelocytes. In early period, examiners incorrectly classified some
images into more immature or mature cell type. However, such misclassification reduced in late period. Although κ coefficient for examiner-JSLH
classification agreement ranged from 0.89 to 0.96 in early period, its values
increased from 0.93 to 0.98 in late period.Conclusions: Our application was
easy and repeatable for IQC. We showed that the application is useful for
standardizing criteria of IG classification and reducing the inter-examiner
variation of differential leukocyte count.
Introduction: We experienced a case of sickle cell disease with abnormally
low level of HbA1c by the HPLC method.Also another HPLC chromatograph
showed the presence of HbA2 and HbS in the patient,but we observed only
targetcells in peripheral blood smear.We report the morphological change
of RBC to sickle cell using simple method in this case.Patient:6-year-old
female with back pain and stomachache.Parvovirus B19 IgM antibody
positive.Father is a half of Brazilian and African. Mother is Japanese.Methods:1) We observed peripheral blood smears of the patient and mother. 2)
Their HbA1c were measured using HLC-723G9. 3)Their hemoglobin were
analysed using ß-thalassemia mode of HLC-723G8 at Tosoh Corporation.
4) We put 0.5 drops of their blood on each slide glass and put cover glass
on their blood to produce anaerobic condition,and we observed morphological change after 2 hours (sickle cell forming test).Results:1) Targetcells
were observed on their peripheral blood smears. 2) HbA1c of the patient
was 1.7%, which was considerably lower than the reference values(4.6 6.2%).3) In ß-thalassemia mode, HbA2 and HbS of the patient were 5.6% (2.0
- 3.5%)and 54.2% (0.0%). 4)The patient's erythrocytes changed their shape
to sicklelike in anaerobic condition,but mother's didn't.Conclusion:Since
targetcells were observed in peripheral blood of the patient and mother,
we suspected thalassemia,but HPLC analysis showed that the patient
had HbS.The reason of abnormally low level of HbA1c in the patient was
considered to be caused by high level of HbS. The patient was finally diagnosed of having compound heterozygote of HbS and ß-thalassemia.On this
case,pain from vaso-occlusive episode and bone marrow necrosis triggered
by parvovirus B19 infection were suspected.In this study,we succeeded in
making sickle cell with simple method.Sickle cell forming test was useful
for diagnosis of sickle cell disease.
94
Evaluation of an internal QC application for correctly
recognizing immature granulocytes on blood smears
PC-64
Validation of inter-laboratory correlation using clinical
specimens
Naoya Ichimura , Ayako Itoi, Yuuki Ohkubo, Yuuki Kouda, Michio Hagihara, Shuji Tohda
Noriko Muraoka 1, Saori Iemata1, Junko Imai2, Yuriko Kamimura1, Hiroshi Kondo3
Clinical Laboratory Dept., Medical Hosp., Tokyo Medical and Dental Univ., Japan
1 Sanki
Medical Laboratory, Japan
2 Wakakusa-Daiichi
Hospital
3 Kansai
University of Health Sciences
Rie Yamamoto 1, Shigeharu Ueki2, Yuki Moritoki2, Masahide Takeda2, Tomoo Saga2, Ayumi Omokawa2,
Noriko Kobayashi1, Makoto Hirokawa2
Improving Laboratory Quality for Single Hematocrit Order on Current Dialysis
Anemia Monitoring in Taiwan
― By a case of cold agglutinin induced mismatch between hematocrit and hemoglobin
Symposium
PC-66
Educational Lecture
The effect of adiponectin on human eosinophil functions:
a possible mechanism for relationship between obesity
and asthma
Special Lecture
PC-65
Introduction:The Wakakoukai group has two hospitals and a registered clinical laboratory. They are Wakakusa-Daiichi Hospital(DHosp),Wakakusa-Tatsuma Rehabilitation Hospital(T-Hosp),and Sanki
Medical Laboratory(SM-Lab). The 2 hospitals'clinical laboratories usually
analyze samples of patients in urgent need of treatment.The SM-Lab analyzes other samples.All clinical laboratory results are displayed as timeseries data on the network-linked clinical laboratory information system.
Doctors can check all laboratory results at the three facilities in our group.
Therefore, inter-laboratory standardization in common laboratory testing
is indispensable.In this study,we validated the inter-laboratory correlation using the laboratory results of the complete blood count(CBC) and
leukocyte 5-differential count(5-diff). Methods:Sysmex XS1000i hematology analyzers were used in the two hospitals,and the Sysmex XT1800i
hematology analyzer was used in the SM-Lab.Peripheral blood samples
were drawn from out-patients from 8:30 am to 10:00 am.Five hundredmicroliter anti-coagulated blood samples were dispensed into 3 microsample tubes,respectively.The sample tubes were stored in a shipping
container with refrigerant and a self-recording thermometer.The shipping
containers were sent from the SM-Lab to the two hospitals.Sample analysis
was duplicated in the manual mode within 4 hours after blood drawing.
Results:Inter-laboratory coefficient of correlation values of CBC were
from 0.993 to 0.999(n=107),and coefficient of correlation values of 5-diff
were from 0.529 to 0.998(n=106).The mean hemoglobin concentration
of T-Hosp was higher than in the other two facilities.Mean hemoglobin
concentrations of SM-Lab,D-Hosp,and T-Hosp were 12.4,12.5,and 12.8 g/
dL(n=36),respectively.Hemoglobin concentrations of the three facilities
converged between 12.9 and 13.0 g/dL(n=45) by adjustment of the automated hematology analyzer.Conclusion:Validation of the inter-laboratory
correlation could be carried out smoothly using clinical specimens.The
results of correlation analysis proved useful to improve the deviation of
hemoglobin concentrations.
Invited Lecture
Objectives: We developed an internal quality control (IQC) application for
recognizing immature granulocytes (IG) consisting of myeloblast, promyelocyte, myelocyte, and metamyelocyte, as shown in our other presentation
in this congress. Here, we examined whether our application improves
inter-examiner variation of the recognition on patients' blood smears.
Methods: In differential leukocyte count, patients' data visually counted by
three examiners were used from June/14 to May/15 (period-1: n=21,451),
before and after introducing the application (from June/15 to Aug/15, period-2: n=3,125; from Sep/15 to Nov/15, period-3: n=3,639). The numbers
of samples in which IG were detected by each examiner (O), the numbers
of samples ordered by each clinical department (e.g. hematology, pediatrics, etc) (C), and the percentage of samples in which IG were detected in
each clinical department (P) were calculated. Weighted expected value (E)
was defined as following equation [ ∑ (Pi × Ci); i:clinical departments].
The deviation between O and E was shown using the value of [(O-E)2/E].
Statistical analysis for comparing between the data in each period was performed using χ 2 test.Results: The percentages of samples in which each
cell type of IG were detected were statistical different between period-1
and period-3, and between period-1 and period-3 except myeloblasts, but
not between period-1 and period-2. The value of (O-E)2/E significantly
decreased from period-2 to period-3 except metamyelocyte (myeloblast:
from 1.9 to 1.6, promyelocyte: from 5.1 to 1.9, myelocyte: from 12.3 to 1.4,
and metamyelocyte: from 4.8 to 6.6, on average). The goodness of fit between O and E was statistically different except myeloblast in period-2, but
not in period-3 except metamyelocyte.Conclusions: The decrease of the
deviation between O and E results from the increase of the inter-examiner
agreement. IQC using the application improved the inter-examiner agreement of IG differential.
Keynote Speech
PC-63
Chi-Chao Tu, Meng-Ting Wu, Chih-Hao Hsu, Pei-Ning Lee, Ai Wang, Chien-Yuan Lan
Department of Medical Laboratory, Keelung Hospital Ministry of Health and Welfare, R.O.C, Taiwan
95
Poster Presentation
Objective: Obesity is associated with asthma, in terms of increased prevalence, reduction in lung functions, and reduced response to medication.
Adiponectin, an adipocyte-derived cytokine, is known to have anti-inflammatory effects with reduced concentrations in obese subjects. Recent findings raised the intriguing possibility that adiponectin might play a role in
allergic inflammation, although the mechanistic basis for their relationship
remains unclear. The aim of our study was to examine whether adiponectin might affect functions and intracellular signaling of eosinophils, which
play an important role in the pathogenesis of asthma.Methods: Purified
human peripheral blood eosinophils were used. RT-PCR and flow cytometry were used to study expression of adiponectin receptors AdipoR1 and
AdipoR2. The effect of adiponectin on eosinophil survival was investigated
using annexin V and propidium iodide staining. Eotaxin-induced cell
adhesion was investigated using ICAM-1-coated plates. Boyden chamber
and real-time horizontal migration system were used for eotaxin-directed
chemotaxis assay. Calcium signaling, phosphorylated ERK (p-ERK), CCR3,
and CD11b expression were studied using flow cytometry.Results: Both
AdipoR1 and AdipoR2 were expressed in human eosinophils. Adiponectin
did not affect eosinophil survival, CCR3, or CD11b expression; however,
eotaxin-enhanced adhesion was inhibited by pretreatment with adiponectin. Adiponectin also diminished eotaxin-directed chemotactic responses
by disturbing both velocity and directionality. Calcium influx in response
to eotaxin was attenuated by adiponectin, although p-ERK expression was
not.Conclusions: The series of our study indicated that adiponectin attenuated the eosinophil chemotaxis and adhesion induced by eotaxin through
modification of calcium signaling. These findings provide the evidence for
the previously unrecognized mechanisms of direct interaction between
adipocytokine and eosinophil functions.018-834-1111(ext.2447)
Oral Presentation
Purpose: In Taiwan dialysis treatment followed DRG system. Clinician followed the practices from Ministry of Health and Welfare, Taiwan Society
of Nephrology, they ordered single HCT (Hematocrit) out of full CBC for
anemia monitoring. Single HCT could lead misjudgments of anemia on a
case. We made corrective actions.
Material: A 59 y/o woman underwent dialysis treatment. On 2015-Sep9, HCT32.7%. Sep-22, HCT28.7%. On Oct-8, her HCT dropped to 23%. We
doubted and rechecked, and found MCHC53.9g/dL.
Method: With unreasonable MCHC, blood appearance looked like sands,
these tallied with cold agglutination and falsely low HCT. Under microscope RBCs agglutinated, cold agglutination test 1:128 (1+). By using
plasma replacement and water bath (37oC/30 min), no more agglutination
under microscope, then examined CBC immediately on the sample from
bath.
Result: On Oct-8 before pre-treatment Hb12.4g/dL, HCT23% and
MCHC53.9g/dL. After pre-treatment Hb12.8g/dL, HCT37.6% and MCHC34g/dL. Cold agglutination was confirmed.
Conclusion: MCHC (Hb/HCT ratio) is commonly used to judge erroneous
results. But if the order is only HCT, LIS only collects HCT. With insufficient
information, MTs could report HCT wrongly. After discussion we made 2
actions. The first, combine Hb and HCT for the order in our hospital. This
will help clinician to evaluate the anemia condition. There's no great repulsion for the other examinations under the DRG system. The second, LIS
transforms the one-item to full CBC order for analyzer. This will help MTs
to judge the results. There's no cost difference between single item and full
CBC on our analyzer.
Case Conference
1 Central
Clinical Laboratory, Akita University Hospital, Japan
2 Department
of Infection, Allergy, Clinical Immunology and Laboratory Medicine, Akita University Graduate School of Medicine
Keynote Speech
PC-67
BPC 157 AND ITS EFFECTS ON HEMOSTATIC
PARAMETERS IN RATS TREATED WITH WARFARIN,
L-NAME AND L-ARGININE
PD-01
Mirjana Stupnisek1, Antonio Kokot1, Sanja Konosic2, Ivan Grzibovski2, Aleksandar Vcev1, Sven Seiwerth2,
Predrag Sikiric2
Makoto Yamaura, Sunao Morita, Atsushi Hori, Hiroya Hidaka
Department of Biomedical Laboratory Sciences, Shinshu University School of Medicine , Japan
1 Faculty of Medicine, J.J. Strossmayer University of Osijek, Osijek, Croatia
2 School of Medicine, University of Zagreb, Zagreb, Croatia
Invited Lecture
Special Lecture
Educational Lecture
[Background and aim]
Lysophosphatidic acid (LPA) consists of a glycerophosphoric acid backbone and an acyl base and is a metabolic product of lysophosphatidylcholine or phosphatidic acid. LPA is a lipid mediator with potent bioactivity,
and its binding with specific receptors is influenced by the identity of its
fatty acid side chain. LPA has been found in serum, and its level in the
blood has been found to correlate with the occurrence of various diseases.
However, there is no appropriate analytical method available for quantifying LPA molecular species in the clinical laboratory. In this study, we examined LPA molecular species in serum and plasma using matrix-assisted
laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF
MS) with mini-column chromatography.
[Methods]
Serum and plasma were collected from healthy young volunteers. Sample
lipids were extracted with butanol solution (pH4). The LPA-rich fraction
was partially purified by DEAE-Sepharose column chromatography and
analyzed by MALDI-TOF MS.
[Results]
The detection limit of this method was 100 nmol/ml. For serum, eight
peaks were detected at m/z 391.3 - 481.3 by MALDI-TOF MS and were attributed to LPA by the product-ion analysis. The eight peaks were assigned
to LPA [C16:0], LPA [C18:0], LPA [C18:0], LPA [C18:1], LPA [C18:2], LPA
[C18:3], LPA [C20:4], LPA [C20:5], LPA [C22:6]. Serum LPA was increased
by exposure to room temperature, and the plasma LPA level was below the
sensitivity of this measurement.
[Conclusions]
This study revealed that LPA can be simply and sensitively measured by
partial purification via column chromatography followed by MALDI-TOF
MS.
Serum LPA may be derived from peripheral blood cells and may be produced by autotaxin.
Background. BPC 157 is a stable gastric pentadecapeptide (GEPPPGKPADDAGLV, M.W. 1419) described in numerous studies that proved its cytoprotective effect, and recently implicated with a role in hemostasis. While NO is
largely implicated in hemostatic mechanisms, in tail-amputation models under
warfarin administration, both the NO-synthase (NOS)-blocker L-NAME (prothrombotic) and the NOS-substrate L-arginine (antithrombotic), have been little
investigated.
Objective. This study determines the effect of the pentadecapeptide BPC157
on the values of hemostatic parameters in rats after administration of anticoagulant warfarin. Also, this study extends and clarifies the effect of BPC 157 on
hemostasis with addition of N(G)-nitro-L-arginine methylester (L-NAME) and Larginine in warfarin treated rats.
Materials and Methods. Male Albino Wistar rats were used in all of the experiments (approved by the Local Ethics Committee). Tail amputation, and/or i.g.warfarin (1.5 mg/kg/day for 3 consecutive days) were used in rats. Treatment
includes BPC 157, L-NAME, L-arginine, applied alone and/or their combination. Laboratory tests (hematological and coagulation) were performed according to manufacturer’s instructions.
Results. After (tail) amputation, with or without i.g.-warfarin, BPC 157 (10 μ
g/kg, i.p./ i.g.) reduced bleeding time and/or haemorrhage and counteracted
thrombocytopenia. As for L-NAME and/or L-arginine, we noted: L-arginine (100
mg/kg i.p.)-rats: more bleeding, less/no thrombocytopenia; L-NAME (5 mg/
kg i.p.)-rats: less bleeding (amputation only), but present thrombocytopenia; LNAME+L-arginine-rats also exhibited thrombocytopenia: L-NAME counteracted
L-arginine-increased bleeding, L-arginine did not counteract L-NAME-thrombocytopenia. All of the animals receiving BPC 157 in addition (BPC 157 μ g+LNAME; BPC 157 μ g+L-arginine, BPC 157 μ g+L-NAME+L-arginine), exhibited
decreased haemorrhage and markedly counteracted thrombocytopenia.
Conclusions. BPC 157 has an effect on hemostatic parameters, especially on
bleeding and platelet count, in treated rats. L-NAME (thrombocytopenia), L-arginine (increased haemorrhage) counteraction and BPC 157 (decreased haemorrhage, counteracted thrombocytopenia) with rescue against anticoagulant
(warfarin), implicate a BPC 157 modulatory and balancing role with rescued
NO-hemostatic mechanisms.
Keywords: BPC 157, bleeding, platelet count, warfarin, NO system, rats.
Symposium
PD-02
Serum and plasma lysophosphatidic acid analysis using column chromatography and MALDI-TOF
mass spectrometry
Evaluation of the lysophosphatidic acid species analysis in blood using MALDI-TOF mass spectrometry
Analysis of human serum sphingomyelin species by MALDI-TOF mass
spectrometry in negative ion mode
Application to a clinical laboratory study of sphingomyelin species measurement
PD-03
To Integrate Point-of-Care and Laboratory Automation
System of BNP testing
― integrate POCT and automation system in the hospital
Case Conference
Oral Presentation
Poster Presentation
Sunao Morita, Makoto Yamaura, Atsushi Hori, Hiroya Hidaka
Mei-E Yang, Ching-Yi Lin
Department of Biomedical Laboratory Sciences, Shinshu University School of Medicine , Japan
Department of Clinical Laboratory, Mackay Memorial Hospital, Taipei, Taiwan
Background:
Sphingomyelin (SM) is an important component of biological membranes,
lipoproteins, and the myelin sheath and plays important roles in biological
membrane maintenance and cell signaling. SM metabolic abnormality is
associated with numerous diseases such as arteriosclerosis and adrenoleukodystrophy. Previously, we reported an analytical procedure for detecting serum SM species by matrix-assisted laser desorption/ionization timeof-flight mass spectrometry (MALDI-TOF MS) in the positive ion mode. Ion
peaks corresponding to both proton and sodium adducts were detected
in the positive ion mode, making the calculation of the peak composition
cumbersome. In the negative ion mode, only the demethylated species was
observed in the mass spectrum.
Aim:
We develop a simple method for profiling human serum SM in the negative ion mode.
Methods:
Sera were collected from healthy young volunteers. Serum lipids were
treated with phospholipase A2 and extracted using the Folch method. Lipids were mixed with a matrix and then analyzed by MALDI-TOF MS.
Results:
We detected 16 peaks at m/z 700 - 850, which were identified as SM species by the analysis of product ions. The correlation coefficient between
values obtained in the negative and positive ion modes was r = 0.9962.
The concentration and dilution rate were linearly related. The reproducibility for major SM species was CV = 4.1% - 13.9%.
Conclusions:
A simple and rapid procedure for the analysis of human serum SM species by MALDI-TOF MS in the negative ion mode was developed and may
be applied in clinical laboratories in the future.
Congestive heart failure (CHF) is a complex syndrome occurs when the
heart is unable to pump sufficiently to maintain blood flow to meet the
body's need, and may cause dysfunction of heart and lead the fluid retention in peripheral vascular and lung. When under the stress, the heart
secret a 108-amino acid pro form of natriuretic peptide (proBNP), then
proBNP will be cleaved into two molecular: a 32-amino acid B-type natriuretic peptide (BNP) and a 76-amino acid N-terminal pro B-type natriuretic
peptide (NT-proBNP). According to 2013 ACCF/AHA Heart Failure Guideline, both BNP and NT-proBNP have good correlation with heart failure
severity, and both of them can be used to heart failure diagnosis and prognosis.
There are multiple commercial BNP and NT-proBNP testing in the market,
include automation system and point-of-care testing (POCT) system, each
of them has its own characteristic. POCT is a global trend to obtain patient
data in 15-20 minutes at the bed site, it is a suitable tool for physicians to
diagnose emergency cases immediately. However, for central laboratory,
how to control the data quality and consistency is the most important
objective. Therefore, how to integrate POCT and automation system in the
hospital is imperative. Triage BNP perform good correlation between Triage MeterPro (POCT system) and Beckman DxI 600 (Beckman BNP = 1.105
x Triage BNP - 20.6, R=0994). It means that we can diagnose emergency
patients by Triage BNP in a short TAT, then monitoring CHF patients by
Beckman system. In addition, we also investigate the clinical performance
of BNP and NT-proBNP on Triage system, to find out which biomarker is
more suitable for the group difference in our hospital.
96
Highsensitivity double-kinetic assay of creatinine in serum
with enzyme cycling method
Preliminary study part 1
PD-05
Measure the effects of temperature on blood ketone stability
Measuring creatinine has important functional renal evaluation. Some
methods for assaying creatinine in serum samples, for suitable with
automated analyzer, are reported. However, they are generally used for
measuring reagents in the laboratory, and don't have enough sensitivity to
measure creatinine correctly. We aim at development a high sensitivity assay method, which can discriminate difference just 0.01 mg/dL creatinine.
We developed enzyme cycling method for measurement of creatinine.
The procedure is sensitive and precise for NAD+ concentrations as low as
0.01mg/dL, and linear to 2.3 mg/dL. Reagent 1, for the first step, contains
a decomposition reaction of creatinine system by creatinine deiminase
and NAD synthetase to produce NAD+ and elimination of endogenous NH3
system by glutamine synthetase. Reagent 2, for the second step, contains
starting enzyme cycling system by L-lactic acid lithium salt, which is substrate of lactate dehydrogenase. Creatinine in the specimen is changed into
NAD+, which is reduced by dehydrogenase, and it becomes NADH. This
NADH reduces tetrazolium salt (WST) through the medium of 1-methoxy
PMS. The NADH lost reduction power back to NAD+ but it is to be reduced
by dehydrogenase and generate NADH. Reproduced NADH is to reduce
WST again. We report just the condition of enzyme cycling method.
Background: Pre-analytical handling of blood samples can influence the
laboratory results. With the increases in lab tests, sample collection time
and various transported conditions can adversely affect the sample quality
and accuracy of test results. Blood ketone traditionally requires to be collected separately from ammonia and lactate. In this study, we examine the
possibility of combining blood ketone test with ammonia and lactate by
measuring the stability of ketone under iced and room temperature conditions.
Methods: Blood samples were collected in heparinized tubes and analyzed
by electrochemical method using the MediSense Optium Xceed. Each sample was divided into 2 aliquot and stored at room or refrigerated ( ice bath
) temperature until time of test. To determine the effect of temperature on
ketone stability, the heparinized whole blood was analyzed at 0, 30, 60,
120 and 180 min incubation time points.
Results: In the concentration of 0.6 - 3.0 mmol/L, the measurement difference was within 10% ( 7.0 ± 6.6 ) for 120min. However, in high level (
more than 3.0 mmol/L ), the difference of blood ketone was less than 5%
( 2.7 ± 2.0 ) within 120min. The statistics results displayed significantly
decreased about 20% in 180 min. These results show that temperature has
minimal effect on the blood ketone stability up to 120 min after specimen
collection.
Conclusion: Given that ammonia and lactate levels are known to be stable
on iced condition, our results suggest it is possible to include blood ketone
with ammonia and lactate tests using heparinized tubes transferred on ice
condition and finished as soon as possible.
From Diagnostic biobank to research biobank, a pilot study
PD-07
Are patients willing to give consent to transfer surplus samples
to a research biobank and how many samples this generates.
Lifestyle impact on serum level of vitamin D and PTH, in
elderly
Level of vitamin D and PTH in elderly
Symposium
PD-06
Educational Lecture
Department of Laboratory Medicine, National Taiwan University Hospital, Taipei, Taiwan
Special Lecture
Chao-Wei Liu, Chih-Hsuan Kuo, Tzu-I Chien, Mo Siu-Mei Lee
Invited Lecture
Akito Mominoki, Eri Ohta, Miki Kawano, Takiko Tateishi, Eisaku Hokazono, Yuzo Kayamori
Division of Medical Technology, Department of Health Siences, Graduate school of Medicine, Kyushu University, Japan
Keynote Speech
PD-04
Fernando Mendes1, Marta Ribeiro1, Joao Paulo Figueiredo2, Ana Valado1, Armando Caseiro1,
Nadia Osorio1, Antonio Gabriel1
Kai Guttulsrod, Mette Bang Nyholm
Clinical Chemestry department, Diakonhjemmet Hospital AS, Norway
Poster Presentation
97
Oral Presentation
Introduction: The aging process is responsible for health decline and may
lead to the dependence and consequent institutionalization. Bone metabolism involves serum calcium regulators, such as vitamin D and PTH. The
following study evaluated and compared serum concentrations of 25(OH)
D and iPTH in elderly people living in institutions and living in their homes
(free-living), with active and independent life.
Methods: We evaluated 50 elderly (25 institutionalized and 25 not institutionalized). First of all, we made an individual questionnaire about lifestyle,
general health and autonomy. Then, we collected blood to measured the
serum concentrations of iPTH and 25(OH)D using immunochemical methods.
Results: Not institutionalized elderly showed higher 25(OH)D serum levels,
comparing with institutionalized elderly (p-value ＜ 0,05). The serum concentration of 25(OH)D was inversely correlated with iPTH. Furthermore
calcium supplementation correlated with higher serum levels of 25(OH)
D (p-value ＜ 0,05) and lower concentrations of iPTH (p-value ＜ 0,05).
However, the free-living elderly who didn't use to take any type of vitamin
supplementation, showed higher 25(OH)D (p-value ＜ 0,05) and lower
iPTH levels (p-value ＜ 0,05), comparing with the institutionalized group.
The free-living elderly who practice three or more activities per day, had
higher concentrations of 25(OH)D (p-value ＜ 0,05) and lower concentrations of iPTH (p-value ＜ 0,05), compared to the institutionalized elderly.
Outdoor activities showed also correlation with serum concentrations of
both hormones (p-value ＜ 0,05).
Conclusion: The adoption of an active lifestyle and the contact with nature,
carry profit to a better aging process. It is essential to recognize vitamin D
deficiency as a public health problem and implement policies of vitamin D
fortification. The incentive of sun exposure, vitamin supplementation for
calcium metabolism, food enrichment and the adoption of an active lifestyle, should be taken into account, in our country, especially targeted to
high-risk groups.
Case Conference
1 Biomedical
Laboratory Sciences, Polytechnic Institute of Coimbra, ESTESC – Coimbra Health School, Department Biomedical
Laboratory Sciences, Coimbra, Portugal
2 Complementary
Science Department, Coimbra Health School, Polytechnic Institute of Coimbra, Portugal
According to the Health Research Act the definition of a biobank is «A collection of human biological material»
Normally biobanks are divided in three main categories: Diagnostic
biobank, therapeutic biobank and research biobank.
A diagnostic biobank contains a lot of samples, which are normally stored
for a week. Turned into a research biobank, it would become valuable. To
store these samples, we need a written consent from the patients.
Regional committees for medical and health research ethics has approved
a consent form allowing transferal of surplus biological material to a research biobank and collecting information from the hospital journal and
other health registries. Further use of the biobank requires an up-front approval from REC.
The study period was three months. We intended to ask all patients hospitalized at a Medical department at Diakonhjemmet Hospital to sign the
consent form after given oral information by the nurse and the physician.
During the study period 736 patients were hospitalized at the ward. 34,1%
of these patients were actually asked to sign the form. Of the collected
forms 69,7% gave their permission, 4,4% declined and 25,9% of the forms
were not valid due to formal errors.
From the 175 patients, 924 blood collections were performed. This generated 870 5ml serum-gel tubes, 765 4ml EDTA-tubes and 361 2,7ml
Citrate-tubes.
We have calculated the theoretical surplus volume in our diagnostic
biobank based on the amount of collected blood and use of sample volume
on our analysers. The potential volume for the research biobank, when divided into 0,5ml aliquots is 1740 serum aliquots, 3060 EDTA aliquots and
722 Citrate plasma aliquots.
The pilot shows that patients are positive contributing to a research
biobank and it would rapidly contain a large volume of patients and samples. This gives great opportunities for a variety of future research projects.
Keynote Speech
PD-08
Clinical Chemistry
Establishment of QMS Everolimus Assay on Abbott Architech
c8000 Automatic Chemistry Analyzer
PD-09
Chia-Jung Tsai1, Hsiang-Lan Chen2, Da-Jhih Jian3, Hsiu-Chen Teng3, Chung-Chi Chen3, Chih-Wei Chen3,
Yu-Chia Chih4, Kung-Sheng Tang3
Yu-Hsuan Yang, Ya-Wen Tsai, Hsin-Yin Chou, Li-Chuan Wang, Li-Ching Wu
Department of Pathology, Chi Mei Medical Center, Tainan, Taiwan
1 Department
of Laboratory Medicine, Chang Gung Memorial Hospital Kaohsiung Medical Center, Taiwan
2 Department
of Laboratory ,Kaohsiung Municipal of Kai-Syuan Psychiatric Hospital, Taiwan
3 Department
of Medical Laboratory Science and Biotecnology,Fooyin University, Taiwan
4 Department
of Biomedical Science,Fooyin University, Taiwan
Invited Lecture
Special Lecture
Educational Lecture
Background:
The Thermo Scientific QMS Everolimus is the newest addition to a full
menu of immunosuppressant drug monitoring immunoassays. There are
applications for a variety of clinical chemistry analyzers; however, studies
on this assay adapted to the Abbott Architech c8000 chemistry analyzer
have not been published. This study evaluated the analytical performance
of Abbott Architech c8000 clinical chemistry analyzers for Everolimus.
Methods:
The analysis was performed according to the QMS assay package insert.
Analytical performances (imprecision, linearity, limit of detection, and limit
of quantification) of this new immunoassay were evaluated. The analyzers
was compared with an HITACHI 7600 Analyzers and, which was recommend by the manufacture.
Results:
The assay was linear in the range of 0.0-20.0 ng/mL. Limit of detection
was 1.5 ng/mL and lower limit of quantitation was 1.3 ng/mL. Within-day
and between-day (20 days) coefficients of variation were between 3.1%
and 8.76% at mean levels of 3.6, 8.0, and 15.2 ng/mL, respectively. We
obtained a Deming regression of y = 0.9061x+0.4714 (r = 0.9752) when
comparing with the HITACHI 7600 Analyzers.
Conclusion:
The results demonstrated acceptable performance, validating
the use of the QMS Everolimus Assay on the Abbott Architech c8000analyzer, and will provide an effective monitoring system for patients receiving Everolimus therapy.
Symposium
PD-10
Association of the DNA Repair Gene hOGG1 Polymorphisms with Nasopharyngeal Carcinoma
hOGG1Ser 326Cys might have potential as a genetic marker for nasopharyngeal carcinoma
susceptibility
The DNA repair enzyme OGG1 is a DNA glycosylase/AP lyase that has
been hypothesized to play an important role in preventing carcinogenesis
by repairing oxidative damage to DNA . Specifically, glycosylase/AP lyase
can efficiently repair 8-OH-G a major base lesion produced by ROS, formed
as a byproduct of endogenous metabolism or exposure to environmental
oxidizing agents, such as ionizing radiation or chemical genotoxic compounds. 8-OH-G is highly mutagenic and, if not excised on DNA replication, can cause GC to TA transversions, which occur frequently in several
oncogenes and tumor suppressor genes.Ser326Cys polymorphism in the
hOGG1 gene is involved in the repair of 8-hydroxyguanine in oxidatively
damaged DNA. Nasopharyngeal carcinoma has a striking geographic and
ethnic distribution, with particularly high rates observed among southeast
Chinese and other individuals of Chinese descent . Nasopharyngeal carcinoma is linked to EBV infection . In addition to EBV, numerous other environmental and host factors have been shown to be associated with the development of nasopharyngeal carcinoma. In particular, longterm cigarette
smoking, consumption of salted fish and foods containing nitrosamine
or nitrosamine precursors at an early age, and occupational exposure to
wood dust have been shown to be consistently associated with this disease. In this study, hOGG1 genotyping was performed by PCR-restriction
fragment length polymorphism analysis of genomic DNA isolated from 84
Taiwanese nasopharyngeal carcinoma cases and 345 individual healthy
donors . We found that distribution of hOGG1 Ser326Cys genotype among
controls (Ser/Ser,18.6%; Ser/Cys, 46.9%; and Cys/Cys 34.5%) was significantly different from that among nasopharyngeal carcinoma cases
(9.5%, 60.7% and 29.8%,respectively )(p ＜ 0.05) .Significantly increase
risk for nasopharyngeal carcinoma was observed. These results suggest
that hOGG1Ser 326Cys might have potential as a genetic marker for nasopharyngeal carcinoma susceptibility.
Association of the Multiple Drug Resistance 1(MDR1) gene 3435 Polymorphisms and bladder
cancer
MDR1 3435 gene might have potential as a genetic marker for bladder cancer susceptibility
PD-11
Hsiang-Lan Chen1, Hao-Hsuan Tang2, I-Han Cheng3, Hsiu-Chen Teng3, Chung-Chi Chen3, Chih-Wei Chen4,
Yu-Chia Chih3, Kung-Sheng Tang3
Electrochemiluminescence immunoassay for cyclosporine and
tacrolimus using Elecsys®Cyclosporine and Elecsys®Tacrolimus
assays with cobas e411 analyzer
Maki Sasano1, Shigeki Kimura1, Ikuhiro Maeda1, Yoh Hidaka2
Case Conference
1 Department
of Medical Technology, Osaka University Hospital, Japan
2 Laboratory
for Clinical Investigation, Osaka University Hospital
1 Department
of Laboratory ,Kaohsiung Municipal of Kai-Syuan Psychiatric Hospital, Taiwan
2 Department
of Pharmacy ,Tajen University, Taiwan
3 Department
of Medical Laboratory Science and Biotecnology,Fooyin University, Taiwan
4 Department
of Biomedical Science,Fooyin University, Taiwan
Oral Presentation
Introduction: Cyclosporine (CsA) and tacrolimus (TAC) are immunosuppressant drugs that are often used to treat autoimmune diseases and
as transplantation therapy; therefore, their concentrations need to be
monitored carefully. Because, Whole blood is recommended for measurements of CsA and TAC concentrations. The samples pretreatments
are needed to measure CsA and TAC concentrations. We have studied
pretreatment conditions and evaluated the analytical performance of the
Elecsys®Cyclosporine and Elecsys®Tacrolimus assay kits, which have
been developed to measure CsA and TAC concentrations.
Poster Presentation
The P-glycoprotein, a product of multiple drug resistance 1 (MDR1) gene,
is a membrane efflux pump localized in epithelial cells in the small and
large intestine, a part of the gastrointestinal barrier that protects cells
against xenobiotics from our diet, bacterial toxins, drugs and other biologically active compounds, possibly carcinogens.Bladder cancer is strongly
associated with smoking and occupational and environmental exposures
to carcinogens.Tobacco smoking is estimated to be the main identifying
risk factor for this cancer, which is responsible for 40 – 50% and 30% of all
bladder cancer cases.The study population consisted of 108 subjects with
pancreatic cancer and 227 healthy controls.The C3435T MDR1 gene polymorphisms was identified using the polymerase chain reaction- restriction
fragment length polymorphism method.We found that distribution of
MDR1 genotype among controls (CC, 41.1%; CT, 44.0%; and TT 14.7%)
was significantly different from that among bladder cancer cases (24.0%,
53.7% and 22.3%, respectively) (p ＜ 0.05). Significantly increase risk for
bladder cancer was observed.These results suggest that MDR1 3435 gene
might have potential as a genetic marker for bladder cancer susceptibility.
Methods: We used residual whole blood samples from autoimmune disease and transplantation patients who were being treated with CsA or TAC.
For sample pretreatment conditions, the mixing time was evaluated at 10,
60, and 180 seconds using pooled whole blood samples. CsA concentrations were measured using an affinity chrome-mediated immunoassay
(ACMIA) and an electrochemiluminescence immunoassay (ECLIA). TAC
concentrations were measured using a chemiluminescence immunoassay
(CLIA) and ECLIA.
Results: CsA samples that were mixed for 10 and 60 seconds revealed
lower results than those obtained from samples mixed for 180 seconds
(* p ＜ 0.05). Within-assay coefficients of variation were 1.8 - 3.6% (CsA:
94.1 - 1237.6 ng/mL) and 2.1 - 3.9% (TAC: 2.1 - 17.8 ng/mL), whereas
day-to-day coefficients of variation ranged between 3.0 - 4.1% (CsA: 91.5 –
1239.8 ng/mL) and 2.8 - 3.9% (TAC: 2.0 – 17.6ng/mL). The limits of quantitative value were 15.5 ng/mL (CsA) and 0.95 ng/mL (TAC). A method
comparison using a standardized major axis regression analysis of ACMIA
and ECLIA was r=0.995, y=0.924x-1.175, n=200 (CsA), while that of CLIA
and ECLIA was r=0.994, y=1.080x-0.197, n=200 (TAC).
Conclusion: The analytical performances of the Elecsys®Cyclosporine and
Elecsys®Tacrolimus assays were good. Furthermore, CyA and TAC concentrations may be simultaneously measured using a single pretreatment.
Thus, the Elecsys®Cyclosporine and Elecsys®Tacrolimus assays may be
suitable for routine therapeutic drug monitoring.
98
Examination of purification of Tamm-Horsfall Protein in urine
Improvement of recovered amount and working efficiency in
purification of Tamm-Horsfall Protein in urine
PD-13
Trace element levels in type 1 diabetes patients
Objective: Several trace elements are involved in insulin signal transduction and glucose metabolism. Present study aimed determine the levels
of three important elements – magnesium, chromium, and zinc – as well
as one oxidative stress marker – malondialdehyde (MDA) – in young type
1 diabetic patients at different periods of their growth, and to realize the
relationships between trace elements, oxidative stress, and growth stages.
Methods: A total of 88 patients with type 1 diabetes mellitus in different
growth stages and 76 gender- and age-matched healthy subjects were
included in this study. The levels of MDA were measured through HPLC
using a C-18 column. Zinc, magnesium, and chromium concentrations in
serum were assessed using atomic absorption spectrophotometry.
Results: We found higher levels of blood malondialdehyde (MDA; p ＜
0.001), significantly lower levels of magnesium (p ＜ 0.001), and no differences in zinc and chromium levels (p = 0.153 and 0.515, respectively) in
younger type 1 diabetic subjects relative to those of control subjects. Only
3.4% (3/88) of younger diabetic subjects exhibited hypomagnesemia; similar results were obtained when comparing different subgroups: children,
adolescents, and adults. We also observed no differences in the levels of
the three elements between the genders and among the growth stages (p
＞ 0.05) of the diabetic subjects. There were no correlations between the
three trace elements and HbA1C, diabetes duration, and insulin dose/BMI
(all p ＞ 0.05), but there was a significant difference between zinc levels
and insulin dose/BMI (p = 0.043) in the diabetic patients.
Conclusions: We found elevated blood MDA, decreased magnesium, and no
changes in zinc and chromium levels in younger type 1 diabetic subjects
relative to those of control subjects. Only 3.4% of younger diabetic subjects exhibited hypomagnesemia. Whether magnesium supplementation
is suitable for improving insulin sensitivity and decreasing oxidative stress
and inflammation will require confirmation through additional studies.
Method: We tried to arrange the method with the diatomaceous earth
filtration by Franca et al. to purify from pooled urine of healthy volunteers.
We evaluated our present method with imitated urine as the purity and
quantity of separation (The evaluation of purity used sodium dodecyl
sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and THP concentration was measured by absorbance in 277 nm.).
Results: Compared with the conventional method, our method was able to
obtain 1.4 times the amount of THP. And the purified component did not
include most proteins in urine except THP. In addition, the present method
was able to shorten the operation time.
Conclusion: The present method can purify more THP in urine more easily
in a shorter time. Not only are more problems avoided in THP measurement, but also our simple protocol will facilitate further research on the
physiologic role of THP.
A microtubule inhibitor, ABT-751, induces autophagy through
inhibition of the AKT/MTOR pathway in Huh-7 cells
PD-15
Ren-Jie Wei1,2, Su-Shuan Lin1, Shang-Tao Chien1, Yow-Ling Shiue2
The effects of varying intensities of light in serum bilirubin
Symposium
PD-14
Educational Lecture
Aim: Tamm-Horsfall Protein (THP) is synthesized in the thick ascending
limb of the loop of Henle and is the most abundant protein in human
urine. The increase and decrease of the quantity of this urinary excretion
are related to the various clinical conditions and the course of disease,
especially renal calculus. This protein has the characteristic that is easy to
aggregate by various factors (salt concentration, osmotic pressure in urine
etc.). This brings about some problems in purification for measuring THP,
especially enzyme-linked immunosorbent assay (ELISA)-based methods.
Therefore, we aimed to establish an efficient protocol for purification of
THP in urine for a standard, which is used on analyzing THP.
Special Lecture
Ching-Chiang Lin1,2, Guey-Ju Tsweng3, Yeou-Lih Huang3,4
1 Laboratory
Medicine, Fooyin University Hospital, Taiwan
2 Department
of Medical Laboratory Science and Biotechnology, Fooyin University
3 Department
of Medical Laboratory Science and biotechnology, Kaohsiung Medical University
4 Department
of Laboratory Medicine, Kaohsiung Medical University Hospital
Invited Lecture
Kouki Hosaka1, Miki Kawano1,2, Eri Ohta1, Takiko Tateishi1,3, Eisaku Hokazono1
1 Division
of Medical Technology, Department of Health Siences, Graduate school of Medicine, Kyushu University, Japan
2 International
University of Health and Welfare School of Health and Sciences at Narita Department of Medical Technology and
Sciences
3 Department
of Medical Technology, Faculty of Health Siences, Junshin Gakuen University
Keynote Speech
PD-12
Gregorio L. Martin I, Allbvi Enric E. Basa, Carmina Alissa G. Baquiran, Arvin N. Constantino,
Angeline Joy C. Garcia, Jill Cristine G. Genuino, Michael James O. Lazatin, Ma. Dianne C. Quevedo
Medical Technology, University of Santo Tomas, Philippines
99
Poster Presentation
Bilirubin is a photosensitive bile pigment derived from red cells. Specimens
submitted for bilirubin determination must always be protected from light
so as not to affect the results. In this study, the researchers determined
how the intensity of light affects the degradation of serum bilirubin levels.
The samples used for the study were pooled serum specimens and these
were classified based on a color chart in order to determine whether it
was normal or icteric serum. Thirty-four (34) pooled serum samples were
collected, seventeen (17) being normal samples and seventeen (17) being
icteric samples. The specimens were divided for exposure in an enclosed
set-up at 100 lux, 150 lux, and 200 lux and had a fix distance of 1.5 meters. The bilirubin levels were measured using Prestige ChemAnalyzer
at different time interval (1 hour, 2 hours and 3 hours) with baseline
values taken prior to exposure. The results of the experiment showed
that light intensity in relation to time did not affect the bilirubin values
since p-values of percentage change for Total Bilirubin [F4,128=0.265,
p=0.900], Direct Bilirubin [F4,128=0.074, p=0.990], and Indirect Bilirubin
[F4,128=0.607, p=0.658] were greater than 0.05. Further, the joint effect
of intensity to total, direct, and indirect bilirubin [F4,128=0.140, p=0.967]
was not significant. However, the number of samples (normal & icteric)
showed significant difference when values were compared with the baseline. In conclusion, the study shows that as the light intensity increases,
the greater the effect on the bilirubin concentration regardless whether it
is normal or icteric samples. Thus, the researchers recommend to consider
longer time interval in the exposure of serum bilirubin in LED light and
other types of light such as the incandescent. The researchers also recommend determination of the maximum light intensity necessary in order to
have a significant decrease of bilirubin level.
Oral Presentation
Hepatocellular carcinoma (HCC) is the most common primary malignant
neoplasm of the liver worldwide, accounting for approximately 6% of all
human cancers annually.
ABT-751 is an oral antimitotic sulfonamide that binds to the colchicine
binding site Beta-tubulin and inhibits polymerization of microtubules. As
a single agent and in combination with standard cytotoxics and radiation,
ABT-751 is active in multiple human tumor xenograft models, including
fibrosarcoma, leukemia, breast, gastric, non-small cell lung and colon cancers. We therefore aimed to study its apoptotic and autophagic effects on
HCC-derived cell lines, Huh-7 with distinct genetic background.
We observed the IC50 of ABT-751 in Huh-7cells were ＞ 50 uM for 24
h; 1.5 uM for 48 h; 1 uM for 72 h. ABT-751 destabilized tubulin in HCCderived cells. DiOC6(3) can be applied to monitor the mitochondrial
membrane potential were higher in HCC-derived cells treat with ABT-751
than control. ABT-751 induced G2/M cell cycle arrest through ATM-CHK1CDC25C checkpoint pathway in Huh -7 cells. ABT-751 induced autophagy
in Huh-7, upregulated MAP1LC3B-II/-I/ACTB folds and downregulated
of SQSTM1 protein levels, through inhibition of the AKT/MTOR pathway.
Monitoring autophagic flux by Cyto-ID Autophagy Detection Kit showed
that ABT-751 treatments for 24 and 48 h induced autophagy in Huh-7
cells, compared to the control (DMSO); rapamycin (500 nM) was served
as a positive control. Inhibition of autophagy enhanced ABT-751-induced
apoptosis. Above results indicated that ABT-751 could dysregulated microtubules and induces autophagy through inhibition of the AKT/MTOR
pathway. Inhibition of autophagy significantly enhanced apoptotic cells.
Case Conference
1 Pathology,
Kaohsiung Armed Forces General Hospital, Taiwan
2 Institute
of Biomedical Sciences, National Sun Yat-Sen University
Keynote Speech
PD-16
Effects of the Use of Different Covering Materials on Serum
Samples for Bilirubin Determination
PD-17
Gregorio L. Martin I, Nielle Jesnin S. Bansil, Karl Daniel S. Bautista, Ma. Clarice DC. Cruz,
Eileen Joie Nina C. Eltanal, Leandro P. Maralit, Rachel Marie C. Taton, Ma. Janina R. Trinidad
Assessing the diagnostic characteristics of procalcitonin assay
in patients suspected bacterial sepsis
PCT assay for bacterial sepsis
Chiou-Huey Wang, Pei-Ling Lai, Jun-he Chen, Suan-Hung Yu
Department of Clinical Pathology, Tungs' Taichung MetroHarbor Hospital, Taiwan
Medical Technology, University of Santo Tomas, Manila, Phillipines
Invited Lecture
Background: The procalcitonin (PCT) assay is a useful screening test for
diagnosis of bacterial sepsis. The expected value of PCT ( ＜ 0.1 ng/mL)
in reagent instruction (Siemen) was obtained from 465 normal subjects.
However, the diagnostic characteristics of PCT assay in patients suspected
bacterial sepsis were not clear.
Special Lecture
The effects of the use of different covering materials on serum bilirubin
were determined. Seventeen pooled sera each of icteric and normal samples were covered using bond, carbon, and aluminum foil. Aliquots of
the collected sera were exposed to fluorescent light at a predetermined
distance of 2.1 meters for 3 time intervals of 1-hour, 2-hour, and 3-hour.
Baseline values of the sera were measured prior to exposure. Values of the
total, direct, and indirect bilirubin after exposure were compared to the
established baseline values. Results for both icteric and normal samples
showed that joint effect of covering material and time of total bilirubin is
not significant with (p=0.417 and p=0.400) values respectively. Similarly,
the mean direct bilirubin using the four covering material for both icteric
and normal samples do not show any significance with (p=0.500 and
p=0.900) values respectively. Moreover, the mean bilirubin do not depend
on the covering material. This indicates that the use of different covering
materials on both normal and icteric samples do not have significant effect
with (p=0.227 and p=0.555) values respectively. These imply that any of
the four covering materials in the study may be used for serum bilirubin
determination.
Materials and Methods: Two hundred and sixteen patients suspected with
bacterial sepsis were recruited in the study from October to December in
2015. Blood from the patients was collected to conduct both of PCT test
and blood culture. Positive results of blood culture were considered as
standard method for diagnosis of bacterial sepsis. Sensitivity, specificity,
positive predictive value (PPV), and negative predicative value (NPV) with
95% confidence intervals were obtained.
Results: Blood from the patients was performed for PCT test in Advia centur CP system (Siemen) and the results of blood culture were monitored
by BACTEC™ Instrumented Blood Culture Systems (BD). Of the 216 blood
sample, positive results and negative results in blood culture were 48
specimens and 168 specimens, respectively. The cut-off value of blood
PCT was set in 0.1 ng/ml according to reagent instruction. We found that
the sensitivity, specificity, PPV, and NPV of PCT assay were 97.9%, 8.3%,
23.4%, and 93.3%, respectively.
Educational Lecture
Conclusion: The PCT level at 0.1 ng/mL performed useful sensitivity and
NPV as a screening test for diagnosis of bacterial sepsis.
Symposium
PD-18
PD-19
IgA/ λ Plasma Cell Myeloma with λ Light Chain Amyloidosis : A Case Report
Lactate to Promote the Patients for Early Sepsis Diagnosis and Survival
PCT and Lactate biomarkers combin measurements for in early
diagnosis sepsis
Case Conference
Kai Pei Huang, Ting Chung Hung, Li Ching Wu
Tzu Ling Chen1, Yu Fen Chen2, Wan Yen Hu1, Chieh Tien Wang1
Department of Pathology, Chi-Mei Medical Center, Tainan, Taiwan
1 Department
of Clinical Pathology,Chi Mei Medical Center, Liouying1 Taiwan
2 Department
of Medial Management Chia Nan University of Pharmacy and Science
Oral Presentation
Poster Presentation
Amyloidosis refers to a variety of conditions wherein amyloid proteins are
abnormally deposited in organ or tissues and cause harm. Among the several forms of amyloidosis, the principal types of that in inpatient medical
services are the AL amyloidosis (primary) and AA amyloidois (secondary).
AL Amyloidois is due to deposition of protein derived from overproduction
of immunoglobulin light chain in plasma cell myeloma. Furthermore, it is
a systemic disorder that can present with a variety of symptoms, including
heavy proteinemia and edema, heptosplenomegaly, otherwise unexplained
heart failure. We reported a 78-year-old female presenting dysuria, oliguria and leg edema for several months. Laboratory data showed proteinuria
(UPCR:1679.8) , leukocytosis (WBC:16.2 x 103/uL), results of serum urea
nitrogen (39mg/dL), creatinine (0.76 mg/dL), IgG(748 mg/dL.), IgA(635
mg/dL), IgM(63 mg/dL), kappa light chain(18.8 mg/dL), lambda light
chain(110.0 mg/dL) and kappa/lambda ratio(0.17). Renal biopsy found
amyloid fibrils in glomerular mesangial area and Congo red stain highlights amyloid deposition in glomeruli. Additional lab studies included
serum protein electrophoresis, which show a major monoclonal peak in
beta region and minor small peak in gamma region, and the immunotyping studies for serum showed two IgA/ λ type. We treated sample with
beta-mercaptoethanol which reducing the polymerized immunoglobulin to
clarify two IgA/ λ are secreted from the same plasma cell clone in bone
marrow. Later examination confirmed it existed plasma cell infiltration in
bone marrow and the immunohistochemical staining showed monotypic
for λ light chain and are positive for IgA. All findings mentioned above
reveal it is a case of plasma cell myeloma with λ Light Chain Amyloidosis.
Recently, the incidence of sepsis continues to increase tendency. Sepsis is
used to define as systemic inflammatory response syndrome (SIRS) and
causes many symptoms. Furthermore, severe septic shock is in a high risk
of mortality. It has been known that lactate increases during apoptosis, but
the link between lactate and sepsis is uncertainty. Nevertheless, procalcitonin (PCT) is an important biomarker for detection of sepsis and septic
shock. In this study, we aimed to investigate if the combination of PCT and
lactate helps to improve the sensitivity for the diagnosis of sepsis, achieving the purpose of early treatment.
The study comprised 261 patients who had already been diagnosed with
sepsis in 2015. The serum samples were tested for PCT and NaF plasma
samples were tested for lactate respectively. Pearson correlation statistics
and binary regression statistics method was used to analyze significant
changes between PCT and lactate.
The result showed that both PCT and lactate level elevated in 261 sepsisdiagnosed patients in early-stage of disease. As Pearson correlation coefficients, positively statistical correlation was found between PCT and lactate
(r=0.439, r2=0.190).Binary regression statistics gave p values ＜ 0.01,
indicating significant positive association between the increase of both
PCT and lactate level and sepsis. In addition, by using the combined measurement of PCT and lactate, patient who was early diagnosis of sepsis was
received appropriate management timely. There were 94 of 117 patients
surviving from sepsis by early investigation and then receiving early treatment. The survival rate was 80.34%.
In the present study, it was found that PCT level rise in early stage of sepsis accompanied with elevation of lactate level. In summary, when patient
is with suspected sepsis, PCT and lactate-combined measurement provides
a tool for early diagnosis of sepsis and beneficial to timely and appropriate
treatment, even to reduce mortality from sepsis.
100
CA125 AS SPECIFIC BIOMARKER FOR THE DETECTION
OF OVARIAN CANCER IN WOMEN PATIENTS
PD-21
The Relationship of Phosphorus, Parathyroid Hormone and Alkaline Phosphatase in Hemodialysis Patients
The serum phosphorus level can be used as an index for monitoring the long-term hemodialysis patient's
condition.
Ridvana * Mediu1, ELDA MARKU2, NAIM MEDIU3
Ming Hui Chen, Xian Lang Fang, Hen Li Chao, Shun Liang Chen
1 Department
of "Senior technical in medical Laboratories", Logos University Tirana, Albania
2 Faculty
of Natural Sciences, Chemistry Department, Tirana, Albania
3 Surgery
Department, General Hospital, Durres, Albania
Tungs' Taichung MetroHarbor Hospital, Taiwan
MMP-10 levels in serum and saliva of patients with different
body-mass index
PD-23
MMP-7 LEVELS IN SERUM AND SALIVA OF OBESE
PATIENTS
1 Polytechnic
Institute of Coimbra, ESTESC – Coimbra Health School, Department Biomedical Laboratory Sciences, Coimbra,
Portugal
2 Polytechnic
Institute of Coimbra, ESTESC – Coimbra Health School, Department Physiotherapy, Coimbra, Portugal
Obesity is a multifactorial chronic disease and its incidence has been
increasing dramatically, being considered a global epidemic by World
Health Organization. It's characterized by increased adipose tissue that
results from hyperplasia and hypertrophy. Their development is associated with extensive modifications in adipose tissue involving adipogenesis,
angiogenesis, and extracellular matrix (ECM) remodelling. The Matrix Metalloproteinases (MMPs) are proteolytic enzymes that are collectively able
to cleave all of the ECM components as well as several non-ECM proteins,
such as cytokines and other MMPs. MMPs which are involved in inflammatory process may contribute to the development of new perspectives
in terms of disease monitoring. MMP-10 is activated by serine proteases
such as plasmin and neutrophil elastase and its active form is able to
activate other MMPs (MMP-1 and MMP-8). MMP-10 has been studied in
other inflammatory diseases such as diabetes mellitus, however, studies of
MMP-10 in obesity don't exist. Therefore, it becomes important to prematurely understand the potential consequences and damage of the obesity,
through the evaluation of biomarkers that can reflect the individual inflammatory state.
Aims: Assess the levels of MMP-10 in the serum and saliva, and it's correlation of MMP-10 in individuals with different body-mass index.
Material and Methods: The semi-quantification of MMP-10 in serum and
saliva was performed by slot blot technique.
Results: The individuals with the highest percentage of body fat (overweight and obese) showed higher MMP-10 serum levels. The individuals
with normal weight presented higher MMP-10 saliva levels. A statistically
significant correlations between the levels achieved in serum and in saliva
was observed.
Conclusion: MMP-10 is altered in obesity and appear to be implicated in
ECM remodeling. Saliva showed to be a fluid with a greater ability to monitor the remodeling of extracellular matrix and with a better diagnostic
potential than serum.
Keywords: Obesity; MMP-10; Saliva.
INTRODUCTION: Obesity is a chronic disease, resulting in expansion of
adipocytes that involves extracellular matrix remodelling, performed by
matrix metalloproteinases (MMPs) and its tissue inhibitors (TIMPs). Some
secondary injuries are caused by chronic inflammation related to obesity,
through the break of the homeostasis between MMPs and TIMPs. Several
studies found altered levels of MMPs in obesity. Recently, it was shown
that MMP-7 and MMP-8 may reflect anti-inflammatory or defensive potential in human inflammatory disease. There are few investigations about
TIMP-3 in obesity, to understand if the inhibitors have different affinities
to MMPs. Furthermore, saliva is very poorly studied in obese young adults.
AIMS: Determine the levels of MMP-7, MMP-8 and tissue inhibitor TIMP-3
and their ratio in serum and saliva samples in normal weight, overweight
and obese young adults. Additionally, correlate both biological fluids.
MATERIAL & METHODS: The levels of MMP-7, MMP-8 and TIMP-3 in serum and saliva samples were evaluated using slot blot technique.
RESULTS: The individuals with normal weight and overweight showed
higher levels of TIMP-3 in saliva samples comparing with the group of
obese young adults. The ratio of MMP-7/TIMP-3, in saliva and serum
samples, were superior in the group of normal weight, comparing with the
obese adults.
DISCUSSION & CONCLUSION: The high levels of TIMP-3 in obese adults
compared to normal weight adults, suggest that plays an important role
in the pathophysiology of obesity. The high ratio levels of MMP-7/TIMP3 in the normal weight adults compared with the obese adults, in both
biological fluids, suggest that MMP-7 shows an anti-inflammatory pattern
through its down-regulation. The correlation between biological fluids allows us to propose the saliva sample as an alternative to serum, having a
great potential in early diagnosis.
Keywords: Obesity, inflammation, MMP-7, MMP-8, TIMP-3, saliva.
101
Poster Presentation
1 Polytechnic
Institute of Coimbra, ESTESC – Coimbra Health School, Department Biomedical Laboratory Sciences, Coimbra,
Portugal
2 Polytechnic
Institute of Coimbra, ESTESC-Coimbra Health School, Department Physiotherapy, Coimbra, Portugal
Oral Presentation
Fernando Mendes1, Frederic Mota1, Ana Freitas1, Lisa Antunes1, Carlos Tavares2, Rui Goncalves2,
Antonio Gabriel1, Armando Caseiro1
Case Conference
Fernando Mendes1, Ana Freitas1, Frederic Mota1, Lisa Antunes1, Cristina Tavares2, Rui Gonçalves2,
Anonio Gabriel1, Armando Caseiro1
Symposium
PD-22
Educational Lecture
Keywords: CA125, tumor marker, ovarian cancer, premenopausal, postmenopausal
Special Lecture
Carbohydrate antigen 125 (CA125) is the established biomarker for detecting OC recurrence and monitoring therapeutic response. In addition,
recent guidelines recommend its measurement in the primary care setting
in women with suggestive symptoms or at high risk for OC, in combination
with pelvic ultrasound. Different methods of quality control are available
and routinely used nowadays. It is important that the methods used for assessment of analytical performance are suitable for the intended purpose
and for the specific conditions of our laboratories.
The purpose of this study was the evaluation of CA125 values and their
statistical analysis, according the age of patients. These patients were
selected when gynaecological disease or, more specifically, a pelvic mass
suspected for OC was present. The serum CA125 levels were measured in
409 outside women patients with a mean age of 42 years. CA125 plasma
concentrations were determined using automated analyzer Cobas®6000
(Rosche Diagnostics) using a chemiluminescent immunometric assay. All
the analytical data were statistically treated by using Descriptive Statistics.
The mean CA-125 level was 55.53 U/ml, the cut-off value for CA-125 was
determined ＜ 35 U/ml. In different age groups, the mean values were
different according to the women status (premenopausal or postmenopausal). According to this study, we suggest that CA125 biomarker should be
included in a prospective study of patients diagnosed with an ovarian cyst
scheduled for surgery.
Invited Lecture
As long-term hemodialysis patients glomerular filtration rate drops to 30
mL per min, the concentration of phosphorus in the blood will increase
leading to a status of hyperphosphatemia and hypocalcemia. The status of
hyperphosphatemia and hypocalcemia stimulates parathyroid hormone
increase, resulting in secondary hyperparathyroidism and many complications such as renal osteodystrophy, with subsequent rise of alkaline phosphatase in the blood.
It has been suggested that the serum phosphorus in these patients should
be controlled at 5.5 mg per dL or less. This study investigated the correlation among the values of serum phosphorus, parathyroid hormone, and
alkaline phosphatase in patients who visited hospital for regular hemodialysis. The results showed that for patients with serum phosphorus equal
or higher than 5.5 mg per dL, their average alkaline phosphatase was
126.4 IU per Liter, with a mean parathyroid hormone at 841.1 pg per mL.
In contrast, the patients with serum phosphorus lower than 5.5 mg per
dL, their alkaline phosphatase average was at 80.3 IU per Liter and mean
parathyroid hormone at 107.7 pg per mL.
Our data showed that the phosphorus lower than 5.5 mg per dL group
had significantly lower alkaline phosphatase and parathyroid hormone
levels than the equal or higher 5.5 mg per dL group (p lower than 0.05).
Our data indicated that when the serum phosphorus was well-controlled
to be lower than 5.5 mg per dL, the secondary hyperparathyroidism and
renal osteodystrophy might be well prevented. Further surgical removal
of the parathyroid gland will not be needed. Thus, the serum phosphorus
level can be used as an index for monitoring the patient's condition as well
as a guideline for patient outcome improvement.
Keynote Speech
PD-20
Keynote Speech
PD-24
Analysis for Stat Biochemistry Laboratory Specimen
Rejection Rate
Improvement of Patient Safety and Patient Care Quality
PD-25
Quantitative analysis of free/total carnitine in plasma by
Liquid Chromatography/tandem mass spectrometry assay
A method detect carnitine deficiency
Invited Lecture
Special Lecture
Educational Lecture
Shu-Wen Lin, Ya-Ching Li, Fang-Yu Wang
Sung Fa Huang, Tuen Jen Wang, Chi Kuan Chen, Chih Kuang Chuang, Tzu Lin Chen
Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taiwan, R.O.C, Taiwan
Mackay Memorial Hospital, Taiwan
Physicians rely on accurate laboratory test results for proper disease diagnosis and for guiding therapy. During laboratory practice, error rates for
preanalytic perforamance measures were about 46 to 68.2%. When the
quality of a blood specimen is poor, it cannot be processed by the laboratory. This leads to a second request for blood specimen and therefore lead
to an increased turnaround time for the laboratory, which is positively
correlated with the delay in diagnosis. Specimen rejection rate is one of
the important quality indicators for laboratory quality and patient safety.
Programs to track laboratory quality have reported aggregated specimen
rejection rates ranging from 0.30% to 0.83%. However, specimen rejection
rate of stat biochemistry lab in our hospital was 1.06%, 7 times higher
than general biochemistry workstation which was 0.15%. We based on
reason records of specimen rejection in LIS system, to analyze specimen
rejection rate and reasons from January to December in 2015. The result
shows rejection volume was 4,227, total sample volume was 398,241, and
the average rejection rate was 1.06% (minimum 0.88%; maximun 1.35%).
Top three reasons for specimen rejection rate were severe hemolysis (74%),
insufficient specimen (13%), and specimen contamination (4%). Statistics
differences of hemolysis interpretation between personnel of stat lab, we
found that total volume of hemolysis was 2,475, the minimum volume was
432 (17.5%), the highest volume was 552 (22.3%). We also analyzed sites
of specimen collection, our data indicated that specimen collected by nursing staffs in ED had highest volume of hemolysis specimen. According to
our findings, we can further communicate with nursing staff in ED, provide
specimen collection notices, and improve consistency of colleague's interpretation of hemolysis, in order to reduce specimen rejection rate of stat
biochemistry lab.
Background: Carnitine is a key substance for transportation of long-chain
acyl-CoA esters across mitochondria membrane for subsequent fatty acid
oxidation. Carnitine can be obtained from exogenous meat supply and
endogenous synthesis in liver. Defects of carnitine may eventually result in
carnitine deficiency. The conventional method for quantitative analysis of
plasma free/total carnitine is carnitine acetyltransferase (CAT) spectrophotometric assay. However, the CAT method is highly affected by hemolysis
and any compound in physiological fluids containing SH functional group.
In addition, a larger volume of blood sample is required which makes the
measurement more unfeasible. In this study, we intended to establish a
liquid chromatography/tandem mass spectrometry method for clinical
diagnostic purpose.
Methods: By using multiple reaction mornitoring (MRM) of tandem mass
analysis, the mass-to-charge (m/z) of the precursor and the product ions of
carnitine was 162.2/85.0. The internal standard, a stable radio-isotope D3carnitine, was used for the calibration and basis of quantitative measurement. The m/z of D3-carnitine was 165.2/103.0.
Results: According to the preliminary results, the within-run and betweenrun precisions of the method were 4.1% and 5.5%, respectively.The recovery of this tandem mass assay was 94.5%, and the subsequent regression
analysis showed good correction with the CAT method (r2=0.957). The reference values of free and total carnitine in normal control were were 42.3
and 52.4 μ mol/L, respectively, with a mean standard deviation of 42.3 ±
8.01(Free carnitine, n=130);52.4 ± 8.80(Total carnitine, n=130), and the
samples showed a normal distribution.
Conclusions: The developed LC-MS/MS method for plasma free/total
carnitine determination is specific, sensitive, validated, and accurate. The
method is applicable and a great benefit particularly to the patients from
pediatric department when a small volume of plasma sample is avaiable.
This method is feasible and can be applied for clinical diagnostic services.
Symposium
PD-26
Type 2 diabetes mellitus with dabetic autonomic neuropathy and brachial-ankle pulse wave velocity
Globulin levels and hypertension are related with brachial-ankle pulse wave velocity in type 2
diabetic patients with dabetic autonomic neuropathy
PD-27
Effects of potassium in the platelets on serum levels of potassium
Change value of serum potassium in the case of adding the
frozen platelets in whole blood
Mutsuko Hosoya 1, Hiroshi Fujita2, Yoko Shiotani1, Yuko Takada2, Tomoko Igarashi2, Shizuka Hasegawa1,
Ayane Kato1, Yoshio Sato1
Yi-Mei Wang
Department of Neurology, National Taiwan University Hospital, Yun-Lin Branch, Taiwan
Case Conference
1 Clinical
Laboratory,Tokyo Metropolitan Bokutoh Hospital, Japan
2 Dept.
of Transfusion Medicine,Tokyo Metropolitan Bokutoh Hosp.
Oral Presentation
Poster Presentation
Background: Dabetic autonomic neuropathy (DAN) is a common complication of type 2 diabetes mellitus (T2DM), and represents a significant cause
of cardiovascular morbidity and mortality in diabetic patients.
Objective: Brachial-ankle pulse wave velocity (baPWV) is known to be a
good surrogate marker of vascular damages. The goal of this study was to
investigate the relationship between baPWV in T2DM with DAN.
Methods: This was a cross-sectional study. A total 93 T2DM patients (42
men and 51 women, age 46-76 years) without history of cardiovascular
disease (CVD) who visited our hospital from March 2011 to April 2014
were assessed. After accurate clinical examinations and biochemical evaluations, the enrolled subjects underwent automatic waveform analyzer to
assess baPWV. DAN was assessed by the electrocardiograms to assess
coefficient of variation in the R-R intervals (CVR-R), the CVR-R related indexes, including CVR-R at rest (CVR-R rest), CVR-R with deep breaths (CVR-R
breath) and their difference (CVR-R breath minus CVR-R rest: CVR-R dif (reference
range: ＞ 2%)), were defined.
Results: There were 15 subjects (33.0% men, mean age 62.0 ± 10.9 years)
with and 78 subjects (47.0% men, mean age 61.4 ± 7.1 years) without
DAN. Subjects with DAN had higher baPWV values than subjects without
(16.6 ± 2.6 vs. 15.2 ± 1.8 m/s, p=0.01). Subjects with DAN had higher
systolic blood pressure, diasolic blood pressure, mean arterial Pressure
(MAP), and pulse pressure elevated red blood cell count, hemoglobin, hematocrit, Glycated hemoglobin, albumin, globulin, high sensitivity C- reactive protein, D-dimer, fibrinogen, and urine albumin-creatinine ratio than
subjects without DAN. Multivariate analysis revealed a significant correlation of baPWV with MAP ( β value:0.151, 95% CI:0.030-0.272, p=0.03)
and globulin levels ( β value:9.195, 95% CI:0.644-17.746, p=0.04).
Conclusions: Independently of other cardiovascular risk factors, serum
globulin levels and hypertension are associated with baPWV even in
T2DM patients with DAN, factors known to increase the risk of CVD.
Introduction: Patients with essential thrombocytosis have been reported
to reveal pseudo-hyperkalemia. The mechanism on pseudo-hyperkalemia
has been known that the increased platelets release potassium from platelets themselves during the clotting in the venipuncture tubes. Recently, we
studied that Japanese patients with polycythemia vera (PV) also revealed
pseudo-hyperkalemia. In this study, we speculated a role of erythrocytes in
pseudo-hyperkalemia and examined the effects of potassium from destructive platelets on serum levels of potassium in detail.Materials and Methods:
Platelet concentrates (PC) from Japan Red Cross used in the experiments
were expired in our hospital (day 4). Whole blood was obtained from normal volunteers after informed consents. First, plasma levels potassium in
eight PC were measured by EA07U (A&T Co. Ltd.), and the same PC were
frozen for 8 hours at -20 degrees (destructive platelets). After dissolving,
plasma levels of potassium in PC were measured to evaluate the potassium level presence in the platelets. Second, destructive platelets from PC
(platelets numbers calculated by Coultae ACT) exogenously added into the
whole blood. Serum levels of potassium were measured in the whole blood
exogenously added the various doses of destructive platelets, using saline
as control.Results: Plasma potassium in PC through freezing procedure
increased (destructive platelets: 0.3~0.8 mmol/L). Next, serum levels of
potassium increased in the whole blood added the destructive platelets
in a dose dependent manner (experiment group: 3.0~6.3 mmol/L, control
group: 0~2.4 mmol/L).Discussion: Increased serum levels of potassium
from the whole blood added the destructive platelets were higher than
those from the destructive platelets themselves. These data suggested that
the increased serum levels of potassium were due to potassium derived
from the whole blood affected by the destructive platelets. We speculate
that pseudo-hyperkalemia in the patients with PV may be due to the result
of erythrocytes-platelets communication in the venipuncture tubes.
102
Renal function is the major influencing factor for the highsensitivity cardiac troponin measurements
PD-29
ITO ATSUSHI 1, Niizeki Noriyasu2, Tomoda Yutaka2, Akasaka Kazumi2, Fujii Satoshi2
Takeo Yuno 1, Tomoko Takayama1, Michiko Nagano1, Ohe Yukiko1, Misuzu Sakai1, Mikio Nagahara1,
Yoshio Sakai1, Takashi Wada2
1 ASAHIKAWA
MEDICAL UNIVERSITY HOSPITAL, Japan
2 Asahikawa
Univ. Hosp.
1 Clinical
Laboratory, Kanazawa University Hospital, Japan
2 Department
of Nephrology and Laboratory Medicine, Kanazawa University
Special Lecture
Introduction : Although cardiac troponin T (TnT) is considered to be a
specific biomarker of a myocardial injury, it is not fully investigated whether there are any factors influencing on TnT in patients without cardiac
diseases. In this context, the aim of this study was to investigate factors
related to the value of TnT in blood of patients without cardiac diseases.
Methods：The data of TnT and several clinical indices were assessed using database in our laboratory from 1 January 2015 through 31 January
2016. Single and multiple linear regression analysis were used to examine
relationships between TnT density and clinical indices (Age, Sex, Diabetes,
TP (Total protein), CRP (C-reactive protein), eGFR ( creatinine-based estimated glomerular filtration rate), CK-MB (Creatine phosphokinase-MB),
LDH (Lactate dehydrogenase)). Results：A total of 184 patients without
cardiac diseases were enrolled. Univariate analysis showed that TnT was
correlated with CRP (R= 0.33, P = 0.01), BUN (R= 0.43, P <0.01), TP (R=
-0.37, P <0.01), LDH (R= 0.29, P <0.01) and eGFR (R= -0.37, P <0.01). In
multiple linear regression analysis, multivariate analysis indicated that TnT
was associated with TP ( β =-0.288, P <0.01), LDH ( β =0.293, P <0.01)
and eGFR ( β =-0.351, P <0.01).Conclusion：This study suggests that TnT
is associated with clinical index of kidney function and lipid-related markers in patients without cardiac diseases.
Invited Lecture
Introduction Recently, highly sensitive assays for cardiac troponin I (cTnI)
and troponin T (cTnT) are available for early diagnosis of acute coronary
syndrome. Because cardiac troponin levels are affected by factors such as
renal dysfunction, anemia, diabetes mellitus, and heart diseases, we analyzed the factors affecting high-sensitivity cardiac troponin I and T (hs-cTnI
and hs-cTnT) levels using multivariate analysis. Materials and methods
Serum samples in patients at Department of Medical Laboratory and Blood
Center, Asahikawa Medical University hospital with the consent of the
secondary use were obtained (n=244). These samples were classified into
6 stages (G1: 42, G2: 45, G3a: 48, G3b: 40, G4: 35, G5: 34) based on CKD
stages of eGFRcreat (mL/min/1.73m2). The average values of hs-cTnI (cutoff value: 26 pg/mL) and hs-cTnT (cut-off value: 14 pg/mL) were compared
between each stage. Multivariate analysis was performed to determine
the factors affecting the values of hs-cTnI and hs-cTnT using eGFRcreat,
diabetes mellitus, heart disease, urinary protein, blood hemoglobin concentration, and NT-proBNP as determinants. Results Values of hs-cTnI and
hs-cTnT tended to increase depending on eGFRcreat stage on CKD. The
average values of hs-cTnT in G3a-G5 were higher than the cut-off value of
hs-cTnT. The average value of hs-cTnI in only G5 was higher than the cutoff value of hs-cTnI. In multivariate analysis, NT-proBNP, myocardial stress
marker, and eGFRcreat remained as the factors that define the value of hscTnT. Only NT-proBNP remained as the factor that defines the value of hscTnI. ConclusionThis study convincingly showed that (1) high-sensitivity
cardiac troponin levels are affected by renal function, and that (2) hs-cTnI
is less sensitive than hs-cTnT to renal function.
Educational Lecture
Usefulness of abnormal reaction data-detecting function of
automated biochemical analyzer
PD-31
saori honda 1, masanori seimiya2, yoshitake suzuki1, yuji sawabe1, toshihiko yoshida1
Comparison of fetal hemoglobin levels in infants by days
after conception and days after birth
Symposium
PD-30
Factors affecting blood high sensitive troponin T
concentrations in patients without cardiac disease
Keynote Speech
PD-28
Yuki Watanabe 1, Kayo Osawa2, Itsuko Sato1, Ruri Kono1, Ikuyo Hayakawa1, Nobuhide Hayashi1,
Ichiro Morioka3, Jun Saegusa1
1 Department
of Clinical Laboratory, Kobe University Hospital, Japan
2 Department
of Biophysics, Kobe University Graduate School of Health Sciences
3 Department
of Pediatrics, Kobe University Graduate School of Medicine
103
Poster Presentation
Introduction: Fetal hemoglobin (HbF) to adult hemoglobin switching
is necessary to facilitate trans-placental oxygen exchange in the blood.
The switching was noted to take place after birth, but the mechanism is
unclear. This study aimed to evaluate whether there was an association
between the HbF levels and days after birth, or days after conception according to gestational age (GA). Methods: The blood samples (n=1,095)
were obtained from 407 infant patients at birth to 364 days excluding 18
patients with hereditary disease or post-transfusion. The HbF levels were
measured by HPLC method. The samples divided into 3 groups as follows: preterm (<34 weeks' GA, n=262), late preterm (34 to 36 weeks' GA,
n=229) and term (37 to 41 weeks' GA, n=604). We performed to compare
the HbF levels of the samples among 3 groups by days after conception
(7 categories) or days after birth (7 categories). Results: In days after conception, the medians of HbF levels among 3 groups were a difference in
a category (under 280 days, p<0.001), while those were closely matched
in 6 categories (281 to 310 days, 311 to 340 days, 341 to 370 days,
371 to 400 days, 401 to 430 days, or more than 431 days). In days after
birth, the medians of HbF levels among 3 groups were not matched in 7
categories (at birth to 6 days**, 7 to 30 days**, 31 to 60 days**, 61 to 90
days**, 91 to 120 days**, 121 to 150 days* or more than 151 days**)(*:
p<0.05, **: p<0.01). Conclusion: The HbF levels were well matched with
the days after conception rather than the days after birth regardless of GA.
Our results supported to recognize the Hb switching and might be used as
reference values of HbF.
Oral Presentation
In laboratory tests using an automated biochemical analyzer, unexpected
problems lead to erroneous test values. Recently, to detect abnormal reactions and failures of the device, a reaction data monitoring system has
been provided for analyzers. In this study, we aimed to investigate the
usefulness of this system to prevent errors in routine tests. Sera were from
patients who visited our hospital. Written informed consent was obtained
from all participants prior to blood sampling, and the study protocol was
approved by the Ethics Committee of Chiba University Graduate School of
Medicine. The absorbance of various test items measured during a 5-day
period from July 13, 2010 (7,283 samples in total) was summed, and the
following items were calculated: 1) variances of operated absorbance at
photometric time-points to calculate the measurement result, 2) variances
of differences in absorbance after mixing the sample and reagent between
adjacent photometric points. When an abnormality was detected by the
system in a routine test, the person in charge confirmed the reaction data
in the reaction monitor of the analyzer. We identified cross contamination
with other test reagents, abnormal test values due to clouding caused by
mixing a sample and reagent, and variation in absorbance due to deterioration of the halogen lamp. These were detected by the reaction data
monitoring system, and the reporting of erroneous test results could be
prevented. When samples in which clouding was caused were subjected
to protein electrophoresis, 22 cases of monoclonal gammopathy were
detected in 20 months. The reaction data monitoring system of the automated biochemical analyzers was useful to prevent false reports due to unexpected problems. It was also suggested that, when a false reaction with
a reagent is detected, a new pathology, such as monoclonal gammopathy,
may be identified by close and careful examination of the sample.
Case Conference
1 chiba
university hospital, Japan
2 internastional
university of health and welfare
Keynote Speech
PD-32
Monthly variation of free testosterone and growth hormone
levels in men and women
PD-33
Development of equation to calculate true kalium levels in
hemolyzed blood samples
Chie Hirayama-Negishi 1, Kazumasa Isobe2, Keiko inoue3, Michikuni Ishijima3, Hitoshi Oikawa1,
Toru Nanmoku1, Yasushi Kawakami2
Noritaka HANDA 1, Kesae HATAGUCHI2, Emi YAGASAKI2, Youki NAKAJIMA2, Waki KANAI2,
Toshiyuki OZAWA2
1 Division
of Laboratory,University of Tsukuba Hospital, Japan
2 Dept.
of Laboratory Medicine,Univ.of Tsukuba
3 i-Laboratory
LLP
1 Department
of Clinical Laboratory, Saku Central Hospital Advanced Care Center , Japan
2 Dept.
of Clinical Laboratory, Saku Central Hosp. Advanced Care Center
Invited Lecture
Special Lecture
BackgroundHemolysis increases serum kalium (K) levels. Our purpose
is to develop a K-compensating equation to estimate the true K levels using hemolyzed blood samples.Mterials and MethodsHeparinized blood
samples from fifty patients were used. K levels and hemolysis index were
determined using BM6050 (JEOL).1. Washed the heparinized blood three
times and diluted with saline. Adjusted hemoglobin levels to 2,500 mg/
dL. Hemolyzed by freezing at -30 °C. This solution was referred to as the
hemolysis reproduction reagent.2. Mixed hemolysis reproduction reagent
with saline and pooled serum and adjusted hemoglobin levels to 0, 100,
200, 300, 400 and 500 mg/dL. Determined K levels and hemolysis index
and developed a K-compensating equation.3. Serum samples from forty
patients whose blood was collected twice because the first sample was
hemolyzed were used to verify the accuracy of the equation. Compare K
levels of non-hemolyzed samples with compensated K levels of hemolyzed
samples.ResultsCorrelation between hemolysis index (x) and increase in
K levels (y) is shown as 1 y = 0.125x + 0.0027. Correlation between hemolysis index (x) and increase rate (y) is shown as 2 y = 3.623x + 0.0767.
We developed two compensation equations: 3 y = A - (0.125x + 0.0027)
and 4 y = A × 100/(100 + (3.623x + 0.0767)); where hemolysis index (x),
compensated K levels (y), and K levels of hemolyzed samples (A). Using
equation 3, the differences in K levels of non-hemolyzed samples and compensated K levels of hemolyzed samples are shown as 0.60 ± 0.44 mEq/
L. Using equation 4, these differences are shown as 0.34 ± 0.36 mEq/
L.ConclusionWe found a correlation between the hemolysis index and
increase in K levels. If we use equation 4, we can compensate K levels and
estimate the true K levels in hemolyzed blood samples.
AbstractIntroduction: A variety of factors, such as sex, age, diurnal variation, exercise, and diet, influence laboratory determinations of hormones.
Growth hormone and testosterone are known to produce sex difference
and are age-dependent. The purpose of this study was to assess monthly
variation and other factors influencing free testosterone and growth hormone levels. Methods: We recruited 9 volunteers (4 men and 5 women,
aged 20-59 years). A 5-mL blood sample was obtained at 5 pm every
week for a month from each participant to measure their free testosterone and growth hormone. LH, FSH, estradiol, and progesterone were also
measured to determine ovulatory stages. Amino acids supplement or zinc
supplement were administered for 7 days. The Pearson moment correlation coefficient was used to determine correlations among the quantitative
variables.Results: In women and men, a monthly variation of free testosterone and growth hormone were observed. In women, these hormones were
increased at the pre-ovulatory stage, and a strong linear correlation between them was found (r=0.672, p=0.0474). In men, muscle training and
zinc supplement enhanced free testosterone level and growth hormone
level.Keywords: Monthly variation, free testosterone, growth hormone
Educational Lecture
Symposium
PD-34
Influence of red blood cells on the pseudo-hyperkalemia
Involvements of kinds of blood collecting tubes, the number
of red blood cells and disease.
PD-35
Yoko Shiotani 1, Hiroshi Fujita1, Mutsuko Hosoya1, Miyoko Katou1, Shizuka Hasegawa1, Ayane Katou1,
Hiroe Oriuchi1, Yoshio Sato2
Influence of In Vitro Hemolysis on 80 Laboratory Tests
Yasuhiro Yanagisawa 1, Kazumasa Isobe2, Etsu Suzuki3, Takayuki Yamamoto4, Michikuni Ishijima4
Case Conference
1 Tsukuba
Medical Laboratory of Education and Research Tsukuba i-Laboratory LLP, Japan
2 University
of Tsukuba
3 TMER
4 Tsukuba
i-Laboratory LLP
1 Tokyo
Metropolitan Bokutoh Hospital, Japan
2 Tokyo
Metropolitan Otsuka Hospital
Oral Presentation
Poster Presentation
Introduction: Either thrombocytosis or leukocytosis induce spurious high
serum potassium, because potassium is released from platelets and white
blood cells during the clotting in the venipuncture tube (VT).In addition,
patients with polycythemia vera reveal higher serum potassium. Therefore, we examined the effects of RBC on the spurious hyperkalemia from
the viewpoints of hemolysis due to techniques on blood sampling. Method:
Clinical settings: Serum, plasma potassium were measured in the patients
with myeloproliferative neoplasm (MPN, N=23) or non-hematological
diseases (NHD, N=42) in our hospital from 2013 to 2015. We firstly compared the differences of serum, plasma potassium ( Δ K=serum potassiumplasma potassium) between the MPN and NHD. Secondly, we made samples of various hematocrits using phlebotomied blood (Hct: 30 to 70%).
Potassium were measured in full volume of blood or only 1mL of blood
sampled into the various VT. Results: While serum potassium, plasma K
and Δ K in MPN were 5.09mEq/L, 4.03mEq/L, and 0.80 mEq/L (range:
0-3.09), those in NHD were 4.22mEq/L, 4.00mEq/L, and 0.22 mEq/L
(range: 0-0.5), respectively. In MPN, Δ K in polycythemia vera (0.71 mEq/
L) was lower than one in essential thrombocythemia (1.30 mEq/L).In the
experiments using samples from phlebotomied blood, hemolysis in 1ml of
blood was almost stronger than one in the full volume of blood. Potassium
levels increased in a hematocrit-dependent manner from 50% to 70%.
Discussion: In MPN, the cause of increased serum potassium was not
certain. However, the serum potassium might increase dependently on the
numbers of the red blood cells in NHD. Moreover, we checked the differences of VT, blood volume of samples and venipuncture techniques in the
experiments described as above. These data suggested that the pseudohyperkalemia might be due to the effects of separation gels and accelerator for clotting on RBC in a hematocrit dependent manner.
Introduction： In vitro hemolysis is probably the most common preanalytic problem in laboratory medicine. It influences many laboratory
tests in many ways.The purpose of this study was to assess the quantitative effects of hemolysis on 80 routine laboratory tests.Methods： We
examined the ratio of hemolysis in our hospital from January 1 to March
31,2015.Next,to study the effect of in vitro hemolysis of whole blood, we
added lysed erythrocytes to pooled specimens of serum to give hemoglobin concentrations of 90 to 810 mg/dl and a rating by colorimetry of 0
to 3+ hemolyzed . Then, 80 laboratory tests were determined with Hitachi
7700 for biochemical tests and with AIA2000,Cobas 6000,and Lumipulse
G1200 for other tests. Results： Hemolysis occurred in a total in 8.6% of
the specimens in our hospital.At level 2 hemolysis ,11 test levels increased
and 7 test levels decreased due to hemolysis.Among the 11 tests, potassium, asparate aminotransferase, lactate dehydrogenase,TTT,ZTT and
hyaluronic acid tests showed proportional increases due to hemolysis. We
could estimate the quantitative effects of hemolysis on these tests using
a factor.Conclusion： Hemolysis is a common problem for accuracy in
laboratory medicine. We can only roughly estimate the quantitative effects
of hemolysis on some tests. But it is still useful to know the approximate
extent of the influence of hemolysis on routine laboratory tests.
104
The effect of water quality degradation for automatic analyzer
Mixture mechanism of calcium and consideration of the
influential amount
PD-37
Shigeo Sueyoshi 1, Masashi Amemiya2, Yoshihito Ito3, Yasuhiro Yoshikawa4, Masanori Kanazawa5
Koyo Shirai1, Chitoshi Sato1, Kosuke Amano1, Kazuhiro Hayashi1, Akira Ishih2, Osamu Yamada1,
Mitsuhiro Hori1
1 Chiba
Cancer Center, Japan
2 Chiba
Children's Hospital
3 Sanritsu
Co.,Ltd.
4 Kameda
Medical Center
5 Merck
Ltd.
1 Okazaki
city Hospital, Japan
2 Dept.
of Infectious Diseases, Hamamatsu Univ. School of Medicine
Invited Lecture
Special Lecture
Recently, point-of-care test using a portable analyzer has been dramatically developed in world. The i-STAT1 portable point-of-care analyzer (Abott,
USA) is cartridge-typed measuring device which has an advantage of
examining various kinds of tests by changing a suitable cartridge for test
item, such as blood gas. Operating temperature of i-STAT1 is recommended between 16 and 30 degrees Celsius, and thus it seems to be difficult to
use this equipment under severe environment in the field of tropical and
cold area in the world as well as severe seasons in Japan.We have already
reported some device keeping the optimum measurement environment for
working i-STAT1 analyzer elsewhere. Then we compared laboratory data
acquired by i-STAT1 analyzer kept under three different environments of
measurement. Three i-STAT1 analyzers introduced for a disaster and/or
emergency in our hospital were used. Each analyzer was put the inside of
the air-conditioning room, the outside of the building, or the inside of the
polystyrene foam box containing warm and cold agents in the outside. To
examine the influence of measurement environment, clinical laboratory
data of the same sample were acquired monthly from each machine. The
data of the same sample were simultaneously examined by Rapidlab 1265
(Siemens, Germany) which is used as routine work in our laboratory, and
then these acquired data were compared. We would like to report results
and discuss the possibility of introduction of i-STAT1 portable point-ofcare analyzer at the time of disaster.
Educational Lecture
Introduction: We have observed that the results of 7 items out of 31 are
affected if unpurified municipal water is used to feed automatic analyzers.
Ca and Mg were the most affected items. We investigate the data shift and
wider dispersion mechanism of 2 electrolytes, it proved water mixed into
the reaction cuvette through the sample pipette.Methods: In this study,
the impact of different water qualities on the biochemistry analysis of 17
items in blind samples was measured with Hitachi P7700analyzer.Results:
Significant differences were observed for 7 items with municipal water.
However, the values observed were below the detection limits, except for
Ca and Mg. As the conductivity of water went up to 100 and 200µ S/cm,
the blind sample values shifted to proportionally higher values. To investigate the contamination, Orange G (absorbance=1.02) was used as sample
and reagent. The solutions were manually sampled from the reaction
cuvette at each step of the operation: sample injection, reagent injection
and mixing. These solutions were measured with a spectrophotometer. At
the reagent injection step, the absorbance changed from 0.948 (92.3%) to
0.953 (92.8%). There were no significant absorbance changes at the other
steps. The reagent pipette is where the water mix into the reaction cuvette
and the volume is around 10µ L. It is 3 to 5 times of sample volume of
Ca.Conclusion: Although Clinical Laboratory Reagent Water (CLRW), as
defined by the CLSI, should have a conductivity below 0.1µ S/cm, some
analyzer manufacturers still recommend less than 1 µ S/cm. 1 µ S/cm is
equivalent to a maximum of 0.082mg/dL Ca. Assuming a target sample
contains 10mg/dL, Maximum 4% Ca is added into a reaction cuvette. In
conclusion, water quality is an important parameter in the measurement
of electrolytes. 1µ S/cm water is too high conductivity for reliable results.
Possibility of introduction of i-STAT1 portable point-of-care analyzer for
acquiring clinical laboratory data at disaster
First report: Optimum measurement environment for using i-STAT1 analyzer
PD-39
Evalution of Immunobiochemical Hybrid-type Automatic
Analyzer cobas8000
Junya Takahashi , Noriyasu Niizeki, Hitomi Kurose, Keisuke Nozawa, Naomi Onodera, Mineji Tachibana,
Yutaka Tomoda, Satoshi Fujii
1 Okazaki
City Hosp, Japan
2 Department
of Infectious Diseases, Hamamatsu University School of Medicine
Dept. Of Medical Laboratory and Blood Center, Asahikawa Medical Univ. Hosp, Japan
Poster Presentation
105
Oral Presentation
Introduction: The basic performance of cobas8000 composed of photometric measuring unit c702 and Electrochemiluminescence technology
measuring unit e602 (Roche) was evaluated.Methods: Sera from patients
with consent for secondary use were utilized to evaluate 21 biochemical
items and 3 electrolyte items. The following parameters were assessed.
(1) Repeatability: 2 concentrations of control sera and pooled sera.(2)Accuracy: calibrators of each item. (3) Linearity: each concentrated sample
was diluted in stages. (4) Effects of interfering substances: Interference
Check A Plus (Sysmex) was used. (5) Correlation: correlation with contrastive reagents with LAbOSPECT008 (HITACHI) and GA09 (A&T). (6)Limit
of detection: limit of quantitation (LoQ) by measuring low level samples.
(7) Effect of carry-over: carry-over from high-level HBs antigen samples on
antigen-negative samples using 3 patterns of the probe cleaning system in
c702.Results: Coefficient of variance (CV) of within-run reproducibility is
0.16-21.30%. CV of between-run reproducibility is 0.29-28.48%. (2) Difference from target value is –3.07-2.80%. (3) The linearity of up to 1390U/L
in AST, 1286U/L in ALT, 127g/L in ALB, 1398.8mg/L in CRE, 11800mg/
L in GLU and 204.15mmol/L in Na were confirmed. (4) Hemolysis hemoglobin affords positive error in TP, AST, LD, IP and negative error in CKMB
and DBIL. Free bilirubin affords positive error in CKMB. Chyle and ascorbic acid afford no effects on all items. (5) Correlation coefficient between
contrastive reagents is 0.910-0.999. (6) LoQ is 5.5U/L in AST, 3.6U/L in
ALT, 0.7g/L in ALB, 0.48mg/L in CRE, and 4mg/L in GLU. (7) With probe
cleaning system carry-over is absent. Discussion and conclusion: Basic performance of cobas8000 and correlation with the contrastive reagents are
satisfactory. Probe cleaning system is considered to prevent carry-over.
Cobas8000 may reduce reporting time by selecting shortest measure time
of immune and biochemistry items and added measure of immune items.
The Japanese Government has dispatched a Japan Disaster Relief Team
for assistance in the event of natural disasters in the world. In the case of
disaster in Nepal, Japan Medical Team for Disaster Relief (JMTDR) activity
especially equipped both ward function and operation equivalent to “Type
2 medical team” of the FMT standard, WHO, and afterward expanded medical functions at the disaster are achieved.At the situation of disaster, we
are often forced to do laboratory testing under severe environment such
as high temperature or high humidity – for example, the temporary tent
at the temperature more than 45 degrees Celsius without air conditioner
in JMTDR Pakistan mission, and to keep measuring equipment working
from the rain caused by the squall in JMTDR Philippine mission. Although
quality of the environment surround laboratory equipment and reagents
is very important to offer a high precise test result to medical settings, it
has not been clarified whether or not laboratory equipment and reagents
brought from Japan showed the precise laboratory data under the severe
environment such as the disaster. The aims of this study are following; i) to
clarify the precision changes of laboratory equipment under severe environment, ii) to consider practical invention to keep optimum environment
being necessary for laboratory equipment and regents in the disaster spot.
In this project, effects of temperature and/or humidity changes on laboratory data were examined inside the air-conditional room producing severe
conditions, using i-STAT1 portable point-of-care analyzer introduced for a
disaster and/or emergency in our hospital. The measurement environment
for working i-STAT1 analyzer was produced by polystyrene foam box and
warm and cold agents being available in Japan and other countries. We
would like to report results and discuss the optimum environment.
Case Conference
Kosuke Amano 1, Chitoshi Sato1, Koyo Shirai1, Kazuhiro Hayashi1, Akira Ishih2, Osamu Yamada1,
Mitsuhiro Hori1
Symposium
PD-38
Study for possibility of medical technology at disaster using
the i-STAT1 portable point-of-care analyzer
Second report measurement in the field
Keynote Speech
PD-36
Keynote Speech
PD-40
Evaluation of Free and Total Prostate-specific Antigen Assay
on the AIA-CL2400 Analyzer
PD-41
Evaluation of an L-FABP assay using LTIA and stability of
urinary L-FABP at different temperatures
Mika Iwai 1, Kiyomi Nakajima1, Keita Kamiyama1, Sakie Fujii1, Tetsuo Machida1, Osamu Araki2,
Kazuto Ito3, Masami Murakami2
Iiduka Takaki 1, Tomoaki Tsukushi1, Hitomi Yamaguchi1, Hiroko Nakazaki1, Kazuhiro Uchida1,
Shin-ichi Munekata1, Yuhsaku Kanoh2
1 Clinical
Laboratory Center, Gunma University Hospital, Japan
2 Department
of Clinical Laboratory Medicine, Gunma University Graduate School of Medicine.
3 Department
of Urology, Gunma University Graduate School of Medicine,
1 Department
of Clinical Laboratory, Kitasato University Hospital, Japan
2 Department
of Laboratory Medicine, School of Medicine, Kitasato University
Invited Lecture
Special Lecture
Educational Lecture
Introduction:Liver-type fatty acid binding protein (L-FABP) is a 14-kDa
protein expressed in proximal tubular epithelial cells. Urinary L-FABP as
associated with urinary protein levels and severity of tubulointerstitial
injury. In the present study, we evaluated the performance of an L-FABP
kit, "Nordia L-FABP" (Sekisui Medical Co., Ltd.) using a latex turbidimetric
immunoassay (LTIA). We also investigated the stability of L-FABP in urine
samples.Methods:The analytical parameters assessed included within-run
and between-day precisions, limit of detection, dilution linearity, interference, and the correlation between LTIA and the enzyme-linked immunosorbent assay (ELISA; CIMIC Co., Ltd., Tokyo, Japan). To evaluate the stability of urinary L-FABP, we observed its changes when stored at room (25°
C), refrigeration (4 °C), and freezing (-80 °C) temperatures for 24 hours
each.Results:The within-run and between-day precisions ranged from
2.83-7.96% and 2.19-2.41%, respectively. Limit of detection and dilution
linearity were 1.42 ng/mL and 240 ng/mL, respectively. The interference
was good, except for that of glucose. As urinary glucose concentration
increased above 1,000 mg/dL, urinary L-FABP concentration decreased by
6-10%. The correlation analysis compared it with the ELISA kit using spot
urinary samples from 48 patients. The Passing-Bablok analysis revealed y
= 0.7755x + 0.2483, r = 0.9374. Urinary L-FABP greatly increased in samples stored at room temperature for 24 hours, whereas it did not change in
samples stored at refrigeration and freezing temperatures.Conclusion:The
performance of "Nordia L-FABP" was excellent, and the test using LTIA was
quick. However, urinary L-FABP decreases if the sample contains glucose
levels above 1,000 mg/dL. In addition, the sample must either be assessed
or stored at refrigeration or freezing temperatures immediately after collection, because urinary L-FABP is unstable at room temperature.
Introduction:Prostate-specific antigen (PSA) is a protein that is produced
from epithelium of prostate gland and is widely used as a marker of prostate cancer. We evaluated the performance of the newly developed free
and total PSA reagents based on chemiluminescence enzyme immunoassay (CLEIA).Methods:Both free and total PSA were measured with a twostep sandwich CLEIA on the fully automated chemiluminescence enzyme
immunoassay analyzer AIA-CL2400 (TOSOH Corporation). Specimens
included residual serum were submitted after routine testing and control
samples. Correlation studies were performed with ST AIA-PACK free PSA
and ST AIA-PACK PSA II on the AIA-2000 analyzer (TOSOH Corporation),
respectively.Results: Both free and total PSA assay showed good performance in imprecision, dilution linearity, interference and correlation study.
Based on the imprecision profile, functional sensitivity at CV20% of the
CLEIA PSA was 0.0013 ng / mL.The total PSA assay on the AIA-CL2400
was confirmed to have an adequate equimolar-response to report precise
PSA concentration.A comparison of free/total PSA ratio was performed
between both systems to evaluate their ability to detect prostate cancer in
34 patients with prostate cancer and 61 patients without prostate cancer
in men with PSA levels of 4 to 10 ng / mL. At 85 % sensitivity, free/total
PSA ratio obtained by CLEIA showed greater specificity for prostate cancer as compared with that obtained by AIA-2000.Conclusion: The newly
developed free and total PSA reagents based on CLEIA showed good performance. Clinical specificity of AIA-CL2400 was greater than that of AIA2000. In addition, the new assays on the AIA-CL2400 are able to report in
15 minutes, show high sensitivity, and need lower sample volume. Thus,
these assays are useful for diagnosis of prostate cancer, monitoring the
course of treatment, and detection of prostate cancer reoccurrence in clinical practice.
Symposium
PD-42
The Interference of Hydroxyurea in the Measurement for Plasma Glucose.
The Interference of Hydroxyurea in the Measurement for Plasma Glucose
by Glucose Oxidase Hydrogen Peroxide Electrode Method.
PD-43
Evaluation of the improved Dimension-TAC assay for the
determination of tacrolimus concentrations in whole blood
Case Conference
Yukio Kume 1, Megumi Takahashi1, Yoshikazu Ono1, Masahiro Jona1, Shigeo Okubo1, Makoto Kurano2,
Yutaka Yatomi2
Tomoaki Tsukushi 1, Minami Katsumata2, Hitomi Yamaguchi2, Hiroko Nakazaki2, Kazuhiro Uchida2,
Shin-ichi Munekata2, Yuhsaku Kanoh3
1 Department
of Clinical Laboratory Medicine, The University of Tokyo, Japan
2 Department
of Clinical Laboratory Medicine, Graduate School of Medicine,The University of Tokyo
1 Kitasato
University Hospital, Japan
2 Department
of Clinical Laboratory, Kitasato University Hospital
3 Department
of Laboratory Medicine, School of Medicine, Kitasato University
Oral Presentation
Poster Presentation
Bacground: Hydroxyure (HU) is one of the common clinical medicines for
the treatment of essential thrombocythaemia (ET). It is well known that
some reagents, such as ascorbic acid, interfere with laboratory testing,
however the interference of HU with laboratory testing is not common.
We have experienced a case of a possible positive interference of HU with
glucose oxidase (GOD) hydrogen peroxide electrode method. Therefore, in
this study, we investigated whether HU interferes this method.Methods:
We measured plasma glucose with two glucose assay; GOD immobilized
enzyme membrane/ hydrogen peroxide electrode method (GODE-method,
chemical reagents and ADAMS Glucose GA-1171 form ARKRAY Inc,
Kyoto, Japan) and Hexokinase Assay method (HK-method, Quick Auto Neo
GLU-HK from SHINO-TEST CORPORATION , Kanagawa, Japan). (1) We
measured glucose concentrations of the plasma samples obtained from 4
subjects taking HU with these methods. (2) We investigated the interference of HU with glucose values determined with each method by adding
HU to plasma samples. (3) We determined how much HU passed through
the GOD immobilized enzyme membrane with newly developed HPLC-UV
detection assay. Results: (1) the positive deviation of glucose values determined with GODE-method from those determined with HK-method was
5~8%. (2) The addition of HU did not affect the glucose values determined
with HK-method, while the addition of HU elevated those determined with
GODE-method. (3) When we challenged the membrane with 10 mg/ml HU
solution, we observed that 60 % of the HU had passed through the membrane.Conclusions : The medication with HU influences the glucose concentrations determined with GODE-method, but not those determined with
HK-method. When we measured glucose with GODE-method, we should
consider the interference of HU with glucose concentration.
Introduction: Tacrolimus is macrolide immunosuppressant derived from
the actinomycetes. Because of its narrow therapeutic index, large interand intrapatient variability in pharmacokinetics, and poor correlation
between dose and trough blood concentrations, tacrolimus concentrations
in the blood must be monitored regularly. The Dimension tacrolimus assay
(TACR), using the antibody-conjugated magnetic immunoassay was reported to show a relatively large variation in the low concentration range.
In present study, we evaluated the analytical performance of the improved
Dimension tacrolimus assay (TAC).Methods: The analytical parameters
assessed included within-run and between-day precision, limit of quantitation, dilution linearity, interference studies, and assay kit comparisons TAC
and TACR. Results: The within-run and between-day precision were obtained with 5.2-7.4% and 3.2-5.7%, respectively. The limit of quantitation
with between TAC and TACR assay were determined to be 1.1ng/mL and
2.6ng/mL, respectively. Dilution linearity was found up to nearly 30.0ng/
mL. No interference was observed in the testing of specimens containing
potential interfering substances such as free bilirubin, conjugated bilirubin,
chyle, rheumatoid factor and hematocrit. However in the EDTA interference study, tacrolimus concentrations was decreased 57.7% at EDTA-2Na
3.0mg/mL and the influence was observed in a concentration-dependent
manner. In comparison study, the correlations between the Dimension
TAC assay and the Dimension TACR assay were as follows: n=79, r=0.959,
y=1.06x - 0.1.Conclusion: The Dimension TAC assay showed high analytical performance. In limit of quantitation study, it admitted to improve of
performance with low concentration range in comparison with the Dimension TACR. However, it is necessary to pay attention to the amount of collected blood, because of influence of EDTA concentrations on tacrolimus.
106
Usefulness of eGFRcre corrected by muscle-mass-volume
creatinine in the bedridden patients
PD-45
Youhei Takahashi 1, Minoru Ukida2
Shinobu Doi 1, Tomoko Kobayashi1, Izumi Mori1, Masafumi Koga2, Takuji Kohzuma3, Midori Ishibashi4,
Ikki Shimizu5, Kazuyuki Akesaka1
1 National
Organization for Automotive Safety & Victims'Aid Okayama Medi-Care Center, Okayama, Japan
2 Okayama
Saiseikai General Hospital
1 Ehime
Prefectural Central Hospital, Japan
2 Hakuhokai
Central Hospital
3 Asahikasei
Phama Corporation
4 New
Tokyo Hospital
5 The
Sakakibara Heart Institute of Okayama
Atsushi Hori 1, Makoto Yamaura2, Sunao Morita2, Kenji Kawasaki1, Mitsutoshi Sugano1,
Takayuki Honda1, Hiroya Hidaka3
Nara Prefecture General Medical Center, Japan
1 Department
of Laboratory Medicine, Shinshu University Hospital, Japan
2 Health
Sciences, Graduate School of Medicine, Shinshu University
3 Laboratory
Science, School of Health Sciences, Shinshu University
107
Poster Presentation
[Background and Aim]Galactosylsulfatide (SM4s) is widely distributed
in various tissues, especially in the brain, nervous system, kidney, lung,
and gastrointestinal tract. SM4s is an important molecule that is involved
in processes such as myelination and insulin secretion and serves as a
ligand for microorganisms. SM4s is composed of a galactosyl sulfate and
a ceramide, which is in turn composed of a sphingoid framework and a
fatty acid. Previously, it was reported that blood SM4s levels are altered
in kidney disease, type-2 diabetes mellitus, and arteriosclerosis. However,
most studies have evaluated SM4s in terms of its total level rather than the
levels of its molecular species. In this study, we aimed at identification of
human serum SM4s and assignment of the structure of SM4s molecular
species.[Methods]Serum was collected from healthy young volunteers,
and 500- μ L samples were treated with lipid hydrolase. Lipids were extracted after adding an internal standard (SM4s [d18:1 C12:0]). Next, the
SM4s-rich fraction was partially purified by phenyl sepharose gel column
chromatography and mixed with a matrix (9-AA). SM4s was analyzed by
matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and TOF/TOF-MS (AB SCIEX) in negative ion mode.
[Results and discussion]Mass ion peaks were measured at m/z 750 – 937,
with 64 ion peaks being attributed to SM4s and assigned to its molecular
frameworks and fatty acids. This method was used to estimate the main
peak profiles of SM4s in serum from healthy young subjects. SM4s molecular species derived from human organ tissues exhibited certain profiles, and the variation of the SM4s total level in blood was associated with
specific diseases. This method may be applied to clarify sulfatide species
metabolism.[Conclusion]This study presented a simple, sensitive method
for analyzing sulfatide species in human serum using MALDI-TOF-MS.
Oral Presentation
[Objective] HbA1c, which is saccharification hemoglobin, reflects blood
sugar level in 1 to 2 month. We measure it by HPLC which is the standard
method. I experienced an abnormal hemoglobin blood symptom estranged
from conversion it using GA , and report it.[Case] 44years old woman.
Weight loss(5kg in 6 months). [Anamnesis and family career] Type 2
diabetes, diabetes retinopathy, Doubt of hemolytic anemia(in other hospital). Her grandmother has diabetes.[Hospitalization views] Blood sugar
level380mg/dl, HbA1c6.9, HbF2.5 , GA42.4, Reduced value(HbA1c-2) ×
30=147mg/dl, Reduced value(GA ÷ 4)+1.73=12.3[Close inspection views]
Examination of outside orderGLT50 was not detected, HbH inclusion body
component red blood cell was not detected, Gene analysis did not lead
abnormality[Chart of the HPLC] Abnormal fraction was not detected.[The
other measurement]HbA1c of enzymatic test4.1[progress] Blood sugar
control with insulin started after education hospitalization. The level decreased, and the control was good. I doubted hemolytic anemia and Hb abnormality, and did close inspection and gene analysis. These did not lead
abnormality. I measured HPLC and enzyme methods, because HbA1c(3.8)
is estranged from conversion one(5.7). This case was followed up as an
unstable Hb symptom.[consideration] Thought from the weight loss and
blood sugar level, this patient was supposed to be chronicity sustained
hyperglycosemia. The desirable HbA1c was more than 10percentage.
HbA1c is estranged from conversion one(12.3). At outpatient time, HbA1c
is estranged from conversion one(5.7). I could not detect the abnormal
fraction and the cause of decreased HbA1c. Datas were thought as proper.
This time, the conversion HbA1c was also reported to clinic.[conclusion]
I experienced HbA1c abnormality which is not detected in HPLC. Conversion HbA1c using GA, is available for a case like this time.
Case Conference
Ai Mutou , Hiroki Yanagida, Shinsuke Ueno, Kazuo Nakamoto, Kumiko Kouchi, Yutaka Yoshimura,
Kana Nishimoto, Hiroki Iio
Symposium
Simple and sensitive identification of molecular species of
human serum sulfatide using MALDI-TOF mass spectrometry
Educational Lecture
PD-47
The case that a result of HbA1c and GA became estranged
Special Lecture
Background: Pancreatectomy is associated with pancreatic exocrine
and endocrine hypofunction, which may cause in decreased metabolism
of glucose, lipid, and protein. We encountered a patient with a markedly
high glycated albumin (GA) level at 97.8%, who had a subtotal pancreatectomy. To investigate cause of high GA, we analyzed the glycation sites of
albumin, self-monitoring of blood glucose (SMBG), HbA1c and GA.Patient:
A 77-year-old male patient who had developed type 2 diabetes 17 years
ago, and treated with sulfonylurea and biguanide. In March 2013, he was
operated on subtotal pancreatectomy against caput pancreatic cancer and
then treated with DPP4 inhibitor. After 6 months, he was admitted to our
hospital because of deterioration of glycemic control in the short-term.
Plasma glucose (PG), HbA1c, and GA were 589 mg/dL, 15.8%, and 97.8%,
respectively. Immediately he was treated with intensive insulin therapy.
Method: To determine the glycation sites of albumin, samples were reduced and S-carboxymethylated and digested by Glu-C endoprotease, and
peptides were analyzed using liquid chromatography/mass spectrometry.
Results: PG was decreased without delay after the start of intensive insulin
therapy. HbA1c and GA levels also decreased almost linearly. However, the
GA/HbA1c ratios were still high (≥4.0) after glycemic control improved.
Whereas measured HbA1c was consistent with estimated HbA1c obtained
from the calculation using six points SMBG measurements, measured GA
showed falsely high compared to estimated GA. The glycation sites of albumin of the pre-treatment patient were extended to various sites in addition
to the major glycation site of Lys-525. The glycation sites were decreased
according to the decrease of GA level, but no abnormal glycation site was
found compare to previous report.Conclusions: These results suggest that
the markedly high levels of GA in this case might reflect deterioration of
glycemic control in the short-term and hypo-catabolism of albumin.
Invited Lecture
BUCKGROUND:For the most of bedridden patients, muscle-mass-volume
(mv) and serum creatinine value (CRE) decrease because of reduction of
the exercise or the activity. It is hard to use estimated glomerular filtration rate (eGFR) using CRE (eGFRcre) for the evaluation of renal function
in such condition. So, Cystatin-C (Cys-C) which is not affected by mv has
been used usually in the suspicious case of the renal impairment.The
aim of this study is to investigate the usefulness of eGFR corrected by mv
CRE (eGFRmvcre) compared to eGFR using Cys-C (eGFRcys).SUBJECTS
AND METHODS:The subjects were 42 patients (male 27, female 15
and average age 42.3 ± 20.5) of persistent vegetative state admitted to
our hospital. We measured mv of each patient using body composition
analyzer (InBody S10), and calculated the correction value from standard
mv. The CRE corrected by mv was calculated by CRE and the correction
value (CRE corrected by mv = CRE × (standard mv ÷ measured mv)).
We compared eGFRmvcre with eGFRcys using regression equation and
correlation.RESULTS:The average mv of the patient was 32.8 ± 6.4kg
compared with the standard mv of 44.6 ± 8.7kg adjusted by height and
sex. The average correction value was +26.2%. Each of eGFR value were
eGFRcys = 94.0 ± 23.5, eGFRcre = 139.5 ± 38.5 and eGFRmvcre = 98.8
± 22.9 (Units:mL/min./1.73m2). The regression equation of eGFRcre and
eGFRcys was y=1.63x-14.2, and the correlation coefficient was r=0.268.
The regression equation of eGFRmvcre and eGFRcys was y=0.97x+7.2,
and the correlation coefficient was r=0.367.CONCLUSIONS:The eGFRcre
value was significantly higher than eGFRcys. There was no significant difference between eGFRmvcre value and eGFRcys value. And thus correction
by mv was improved both the regression and the correlation. The results
suggested that eGFRmvcre was useful for the evaluation of renal function
in bedridden patients.
PD-46
Markedly high levels of glycated albumin in a case with
subtotal pancreatectomy
Keynote Speech
PD-44
Keynote Speech
PD-48
Development of a novel measurement method for
L-hydroxyproline using a new enzyme
PD-49
Setting referrence intervals of serum analytes in Japanese
elderly populations
Invited Lecture
Special Lecture
Educational Lecture
Kazuaki Yamamoto 1, Seiya Watanabe2, Michio Hagihara1, Shuji Tohda1
Noriaki Harada 1, Juon Lee1, Kiyoshi Ichihara2, Yasuo Yano1, Nobuko Ikeda1, Yoshihisa Itoh1
1 Department
of Clinical Laboratory, University Hospital of Medicine, Tokyo Medical and Dental University, Japan
2 Graduate
School of Agriculture, Ehime University
1 Eiju
General Hospital, Japan
2 Yamaguchi
University
Background: L-hydroxyproline (L-Hyp) is one of the amino acid included
in the collagen. It is considered that L-Hyp can prevent the binding of LDLcholesterol to lipoprotein(a) deposited in the vascular wall. Currently, LHyp concentration is measured by HPLC, but this method takes time for
measurement. Therefore, we developed the quick and simple enzymatic
measurement method for L-Hyp using a new enzyme, and evaluated its basic performance.Method: L-Hyp concentration was measured using an automated analyzer LABOSPECT 008 (HITACHI). The reagent compositions
were as follows: the first reagent was pH 8.0, 0.1 mol/L HEPES buffer (including 1 mmol/L EDTA), 0.18 U/mL L-hydroxyproline epimerase and 5.5
μ mol/L 1-methoxy PMS, and the second reagent was pH 8.0, 0.1 mol/L
HEPES buffer (including 1 mmol/L EDTA), 0.074 U/mL D-hydroxyproline
dehydrogenase and 55 μ mol/L WST-3. 1) We measured L-Hyp standard
solution (25 μ mol/L and 50 μ mol/L), and evaluated the within-run
reproducibility (n=20). 2) We made the dilution series of L-Hyp standard
solution, and obtained the linearity by measuring each three times. 3)
We made the dilution series of L-Hyp standard solution, and obtained the
detection limit by measuring each five times.Results: 1) The coefficient of
variation (CV%) was 0.96 % for 25 μ mol/L L-Hyp, and 1.33 % for 50 μ
mol/L L-Hyp. 2) In the linearity of dilution, it was linear up to 500 μ mol/
L. 3) The detection limit was 2 μ mol/L. 4) This method measured L-Hyp
within 10 minutes. Conclusion: We showed that our novel method was
able to measure L-Hyp concentration quickly. Its within-run reproducibility, linearity and the detection limit were good enough. We are planning
to measure L-Hyp concentration in healthy and patient specimens, and
investigate the relevance of L-Hyp, triglyceride, LDL-cholesterol, and HDLcholesterol.
Setting reference intervals (RIs) started from serum proteins when its international reference material, CRM470, was introduced to Japan. While
progressing standardization of every analytes in the laboratory, Ichihara
et al. continuously has developed the internationally-recognized statistical
procedure setting for RI. This is an initial report to set RI in Japanese elderly populations in Taito Ward Medical Checkup. After informed consent,
total 2459 sera were collected from November 2013 to January 2015 at
Taito Ward health checkup. It consisted of 924 males and 1565 females
aged 39 to 94. RI was determined by the established method: an initial
selection of apparently normal reference individuals without remarkable
present/past history of diseases, followed by a LAVE method for selecting
them statistically based on analytes, setting by Box-Cox transformation.
CBC was determined on Ruby (Abott), while 40 biochemical and immunochemical analytes were determined by turbidimetry on Hitachi Analyzer
(LAbOSPECT 006). Total, male and female RIs were set for each analytes in
the age cut off as 65 years old. RIs below 65 years old were almost same
with that in previous reports. Cystatin C constantly increased as ages rise.
It probably reflects physiologically and pathologically decreased number
of nephron and function. Same may be true for UN. Of noteworthy are C3
and CRP which proved to be BMI-dependent elevation regardless of the
age. The upper limit of RI of CRP in elderly populations was, however, in
the pathologic state. Decrease of zinc and its ligand albumin becomes decreased. This could be explained as an aging process as well as effects of
inflammation. Definition of reference individuals is difficult. They more or
less have mental and physical alternations including pathologic conditions.
While seeking proper reference parameter like RIs, we recognized an importance of follow up individually by deep insights on analytes incorporating all possible information.
Symposium
PD-50
Extent of hemoglobin interference during serum total protein
measurements is affected by biuret reagent used
PD-51
Ken-ichi Nagai 1, Makoto Matsushita2, Tomoko Arai2, Noriko Ihara2, Yoshimi Muramoto3,
Jyun-ya Yamaguchi1, Ryuuji Miki4, Kiyoshi Kamiyama5
Detection and Regulation of Apolipoprotein M in Central
Nervous System
Naoko Hisasue 1, Makoto Kurano1, Yuki Morimoto2, Koichi Tsuneyama2, Kubota Tetsuo3, Yutaka Yatomi1
Case Conference
1 Department
of Clinical Laboratory, The University of Tokyo Hospital, Japan
2 Department
of Pathology and Laboratory Medicine, Institute of Biomedical Sciences, Tokushima University Graduate School
3 Department
of Microbiology and Immunology, Tokyo Medical and Dental University, Graduate School of Health Care Sciences
1 Department
of Clinical Laboratory Saiseikai kawaguchi General Hospital, Japan
2 Saitama
Prefectural University Graduate School
3 Faculty
of Health & Medical Care, Saitama Medical University
4 Dokkyou
Medical University Koshigaya Hospital
5 Medical
Diagnostic Center of URAWA Medical Association
[Background] Apolipoprotein M (apoM) is a minor apolipoprotein rid-
Oral Presentation
Poster Presentation
ing on HDL. Recently, apoM is elucidated to be a carrier of sphingosine
1-phosphate, a bioactive lipid mediator, which contributes to several
pleiotropic effects of HDL, including anti-apoptosis, anti-inflammation, and
vaso-relaxation. Although it is well known that HDL-like lipoproteins exist
in central nervous system (CNS) and play important roles in the pathogenesis of several diseases, pleiotropic effects of those in CNS are yet to be
known. In this study, we aimed to examine the existence and regulation of
apoM in CNS.[Methods] (1) We examined the existence of apoM in CNS by
western blot analysis with cerebrospinal fluids, immunostaining with an
autopsy brain, and RT-PCR from U-251 MG cells, an astrocytoma cell line,
and SH-SY5Y cells, a neuroblastoma cell line.(2) To elucidate the kinetics of
apoM in CNS, we investigated the modulation of apoM by apolipoprotein
E (apoE) and LDL receptor, which are involved in the homeostasis of apoM
in circulation, with adenoviral overexpression in U-251 MG cells.[Results]
(1) ApoM was detected in cerebrospinal fluids at 1 ~ 10 % of sera. The immunostaining analysis revealed that apoM was expressed mainly in glial
cells. ApoM was detected by the RT-PCR in both cells, together with S1P
receptors.(2) In both ApoE- and LDL receptor-overexpressed U-251 MG
cells, supernatant apoM levels were decreased.[Conclusion] The present
findings indicate that apoM exists and may contribute to the pleiotropic
properties of HDL-like lipoproteins in CNS and that apoE and LDL receptor are involved in the homeostasis of apoM in CNS. Considering apoE and
LDL receptor have important roles in the pathogenesis of neurological diseases, such as Alzheimer disease, apoM in CNS might possibly guide a way
for the development of novel laboratory testing and therapy for neurological diseases.
Introduction: The biuret method is widely used as a reference procedure
for determining serum total protein concentrations in clinical laboratories
because equal amounts of color are produced per gram of protein regardless of the type of protein. However, the reagents and analysis conditions
employed can differ markedly between the various commercially available biuret assays. Methods: In the present study, we compared the extent
of hemoglobin interference between six types of commercially available
biuret assays, A (SEKISUI MEDICAL, a 1-reagent system), B (SHINO-TEST, a
1-reagent system), C (Wako Pure Chemical, a 1-reagent system), D (KAINOS,
a 2-reagent system), E (SHINO-TEST, a 2-reagent system), and F (Wako
Pure Chemical, a 2-reagent system). All analyses were performed using the
FURUNO CA-270 plus autoanalyzer, but assays A, B, and C were carried
out using the one point/dual wavelength method, whereas assays D, E, and
F were performed using the two points/dual wavelength method. Results:
The total protein levels of serum samples that had been spiked with 1.0 g/
dL of hemoglobin were compared with those of untreated serum samples.
In assays A, B, C, D, E, and F 1.0 g/dL of hemoglobin was equivalent to total
protein levels of 1.26, 1.08, 1.26, 0.70, 0.55, and -0.03 g/dL, respectively.
Conclusions: In the 1-reagent systems, (assays A, B, and C), the effect of
hemoglobin interference on serum total protein measurements was equivalent to the sum of the error due to the absorption of hemoglobin and the
error caused by hemoglobin participating in the biuret reaction. In contrast, in assays D and E (2-reagent systems), the effect of hemoglobin interference was dependent on the hemoglobin concentration. Furthermore,
in assay F (a 2-reagent system) the two different hemoglobin interference
mechanisms were considered to have canceled each other out.
108
PD-53
Role of adiponectin in chronic kidney disease.
RYOSUKE KIKUCHI 1, Yasuda Yoshinori2, Hiroyuki Matsumoto1, Toyoaki Murohara3,
Tadashi Matsushita4
Masashi Miyoshi , Takayuki Nakao, Toshio Doi
Tokushima University Hospital, Japan
1 Department
of Medical Technique, Nagoya University Hospital, Japan
2 Department
of CKD Initiatives/nephrology, Nagoya University Graduate School of Medicine
3 Department
of Cardiology, Nagoya University Graduate School of Medicine
4 Department
of Clinical Laboratory Medicine / Transfusion Medicine, Nagoya University Hospital,
WAN-LING CHIU1,2, YU-HSUAN SHAO1, PEI-WEN LEE2, CHIA-LING CHEN3
Serum CETP status is independently associated with reduction
rates in LDL-C in pitavastatin-treated diabetic patients
possible involvement of LXR in its association
Symposium
PD-55
Educational Lecture
The Maintenance of the Accuracy of Point-of-care testing (POCT) for Glucometer
The Evaluation Study of Verification for FEGO Glucometer and Roche Accu-Check
Active
Special Lecture
Introduction: Chronic kidney disease(CKD)is a public health concern in
the worldwide. Vascular endothelial growth factor A(VEGF-A)is constitutively expressed by epithelial cells of nephron from embryonic to adult
kidneys. Chronic kidney disease is associated with reduced VEGF-A expression. To date, essentially nothing is known about the role VEGF-A splice
isoforms, so called VEGF-A165b, play in kidney physiology and pathology
is still unclear. Previous studies suggest that VEGF-A165b expression is
protective in renal function in diabetic rodent models. This study aimed
to investigate whether urinary VEGF-A165b concentrations are able to
estimate kidney function.Methods and Results: A total number of 157
patients were enrolled from in and outpatient clinics at Nagoya University
Hospital. Patients were divided according to G1(estimated glomerular filtration rate(eGFR≥90), G2(90 ＞ eGFR≥60), G3a(60 ＞ eGFR≥45), G3b (45
＞ eGFR≥30), G4(30 ＞ eGFR≥15) and G5 (30 ＞ eGFR≥15) groups. Urinary
total VEGF-A and VEGF-A165b levels were determined by enzyme-linked
immunosorbent assay. Statistical analyses were completed with Graphpad
Prism6. Urinary levels of total VEGF-A were no significant difference in
each groups. However, as eGFR declined, urinary levels of VEGF-A165b were
significantly decreased(G2: p<0.1267, G3a:p<0.0001, G3b: p<0.0001, G4:
p<0.0001, G5: p<0.0001 compared with G1 subjects). Urinary VEGF-A165b
levels were also correlated positively with eGFR, eGFRcys and inulin clearance. Conclusion:Our data indicate the close association of low urinary
VEGF-A165b levels with kidney function, suggesting that urinary VEGFA165b represents a novel biomarker for kidney dysfunction.
Invited Lecture
Adiponectin is one of the bioactive substances secreted by an adipocyte. It
may play an important role in the progression of chronic kidney disease
(CKD); however, there is limited information regarding the underlying
protective mechanism.In the present study, we measured the level of adiponectin in patients with CKD and correlated the data with the functional
index and risk stage. In addition, we examined the relationship between
serum adiponectin levels and organ dysfunction. In this study, patients
examined at the Tokushima University Hospital with CKD (n=228) were included. Additionally, healthy participants were enrolled as controls (n=45).
We analyzed the patients according to their sex.Our findings suggest that
serum adiponectin levels were significantly correlated with creatinine (men:
r=0.559, women: r=0.449, P ＜ 0.01), estimated GFR (men: r=0.484, women: r=0.430, P ＜ 0.01) and urinary protein levels (men: r=0.555, women:
r=0.463, P ＜ 0.01). We also found that patients with renal failure have
elevated serum levels of adiponectin (P ＜ 0.01). We categorized the patients with CKD into 4 stages based on the risk of mortality or the stage of
organ dysfunction, and compared the adiponectin levels among these risk
stages. We found that serum adiponectin levels increased with advanced
risk stages (P ＜ 0.01).Urinary adiponectin levels were significantly correlated with urinary protein (P ＜ 0.01) and urinary albumin levels (P ＜
0.01). Although urinary adiponectin levels were not correlated with serum
adiponectin levels, we considered that high serum levels in patients with
renal failure were not associated with an increase excretion in urinary excretion.In conclusion, serum adiponectin levels were increased in patients
with progressive renal dysfunction. However, this result is in contrast
with the protective role of adiponectin.Although the pathways involved in
increased adiponectin secretion are not well understood, these results suggest that serum adiponectin levels are associated with renal dysfunction
and that adiponectin has protective roles in the kidney.
PD-54
Urinary VEGF-A165b and estimated glomerular filtration
rate
- Does urinary VEGF-A165b estimate kidney function ? -
Keynote Speech
PD-52
Akihiro Shimada 1, Hideki Kimura1, Koji Oida2, Hideo Kanehara3, Yasuki Ito4, Masayuki Iwano5,
Tsutomu Hirano6, Haruyoshi Yoshida7
1 Department
of Clinical Laboratory and Nephrology, University of Fukui Hospital, Japan
2 Division
of Internal Medicine, Fukui Chuo Clinic
3 Division
of Endocrinology, Fukui-ken Saiseikai Hospital
4 Research
and Development Department, Denka Seiken Co. Ltd.
5 Division
of Nephrology, Department of Medicine, School of Medicine, Faculty of Medical Sciences, University of Fukui
6 Department
of Medicine, Division of Diabetes, Metabolism, and Endocrinology, Showa University School of Medicine
7 Division
of Nephrology, Obama Municipal Hospital
109
Poster Presentation
Background: Statins decrease cholesteryl ester transfer protein (CETP)
levels, which have been positively associated with hepatic lipid content as
well as serum low density lipoproteins-cholesterol (LDL-C) levels. However,
the relationship between the CETP status and statin-induced reductions in
LDL-C levels has not yet been elucidated in detail. We herein examined the
influence of the CETP status on the lipid-reducing effects of pitavastatin
in hypercholesterolemic patients with type 2 diabetes mellitus as well
as the molecular mechanism underlying pitavastatin-induced modifications in CETP levels. Methods: Fifty-three patients were treated with 2
mg of pitavastatin for 3 months. Serum levels of LDL-C, small dense (sd)
LDL-C, and CETP were measured before and after the pitavastatin treatment. The effects of pitavastatin, T0901317, a specific agonist for liver X
receptor (LXR) that reflects hepatic cholesterol contents, and LXR silencing on CETP mRNA expression in HepG2 cells were also examined by a
real-time PCR assay.Results: The pitavastatin treatment decreased LDLC, sdLDL-C, and CETP levels by 39%, 42%, and 23%, respectively. Despite
the absence of a significant association between CETP and LDL-C levels
at baseline, baseline CETP levels and its percentage change were an independent positive determinant for the changes observed in LDL-C and
sdLDL-C levels. The LXR activation with T0901317 (0.5 μ M), an in vitro
condition analogous to hepatic cholesterol accumulation, increased CETP
mRNA levels in HepG2 cells by approximately 220%, while LXR silencing
markedly diminished the increased expression of CETP. Pitavastatin (5 μ
M) decreased basal CETP mRNA levels by 21%, and this was completely
reversed by T0901317. Conclusion: Baseline CETP levels may predict the
lipid-reducing effects of pitavastatin. Pitavastatin-induced CETP reductions
may be partially attributed to decreased LXR activity, predictable by the
ensuing decline in hepatic cholesterol synthesis. Trial registration: UMIN
Clinical Trials Registry ID UMIN000019020
Oral Presentation
The progress of technology brings the convenience life, then the testing
method is using small and portable examine equipment within minutes
has test result. But these results of the accuracy, quality control, calibration, responsibility belongs are challenge clinical laboratory.
According to IDF worldwide total of 4.15 million people suffer from diabetes, and an average of every 11 persons is diabetics, so glucometer plays
an important roles in the diabetics.
FEGO Pedometer and Blood Glucose Monitoring System is EC Certificate
approved, wearable devices, the movement is important for diabetics.
Pedometer function let diabetics to easy control their health is very prospective in the future.
The other Roche Accu-Check Active glucometer is the highest market
share product in glucometers so we doing contrastive test.
The standard test result is use BECKMAN DxC 600 biochemical machine.
According to ISO 15197: 2013 new standard we doing evaluation study
to ensure that glucometer result and patient local clinical laboratory result
will consistent and enhance reliability in POCT.
The evaluation study of Verification is using 40 people collection specimen
to contrastive test, this result using the Intraclass Correlation Coefficient
(ICC) analyze statistics, and ICC is a measure of the reliability of measurements or ratings.
For the purpose of assessing inter-rater reliability and the ICC, two or preferably more raters rate a number of study subjects.
The results showed no significant difference between the different analytical principles of glucometer, the contrastive test result are within ISO
15197: 2013 new standard specifications.
However, validation and evaluation studies only in clinical laboratories,
and experienced medical laboratory scientist operation can enhance accuracy.
In POCT a medical laboratory scientist is irreplaceable.
Case Conference
1 Graduate
Institute of Biomedical Informatics, Taipei Medical University, Taiwan
2 Department
of Clinical Laboratory, Taipei City Hospital Yangming Branch, Taiwan
3 Nursing
Department, Far Eastern Memorial Hospital, Taiwan
Keynote Speech
PD-56
Evaluation of RG II POCT Devices for Glucose Testing
Point-of-Care Testing
PD-57
Development of ICG measurement (long-wavelength control)
using an automatic biochemical analyzer
Invited Lecture
Special Lecture
Educational Lecture
Hye Jung Kim1, Hye Lim Kim1, Yong Lim2
Yuka Sato 1, Masanori Seimiya2, Toshihiko Yoshida1, Yuji Sawabe1
1 Bongseng
Memorial Hospital, Korea
2 Dong
Eui Univerisity
1 Chiba
University Hospital, Japan
2 International
University Of Health And Welfare
Point-of-care testing (POCT) is an increasingly popular means of providing laboratory testing at or near to the site of patient care. POCT provides
rapid results and has the potential to improve patient outcome from
earlier treatment. However, a faster result is not necessarily an equivalent
result to traditional, core laboratory testing. Glucose meters are used
routinely in hospital wards to manage blood glucose levels in patients
requiring frequent monitoring of blood glucose. The primary objective of
our study was to investigate whether all these RG II glucose meters (Sejong
Biotechnology, Suwon, South Korea) results in hospitalized patients during routine clinical care jointly satisfy the specified quality specifications,
as defined by Clinical and Laboratory Standards Institute (CLSI) guideline
POCT12-A3. The records of hospitalized patients who underwent simultaneous measures of glucose levels with both glucose meters and a central
laboratory analyser were retrospectively analysed. We also performed a
prospective evaluation of the accuracy of the RG II glucose Strip. Glucose
concentrations measured in 80 patients ranged from 2.01 to 32.17 mmol/
L The Bland-Altman difference plot between the auto analyser and RG
II glucose meters revealed a mean bias of -3.1%, with analytical biases
for the two methods varying from -38.6% to 32.4%. The glucose meter's
values were within ± 14.3% for values 6.28 mmol/L of the comparative
laboratory glucose values and 89% of the results were within 24% of the
reference for glucose＞ 4.5 mmol/L and 65% of the results were within 1.3
mmol/L for glucose<4.5 mmol/L. RG II glucose meter readings in hospital
settings, especially in hypoglycaemic patients, should be confirmed by
central laboratory analysers whenever possible.
Corresponding author: Yong Lim
ICG plasma disappearance rate (ICG-PDR) is important for understanding
liver functions. We development the new autoanalyzer methods (longwavelength control method) for ICG plasma disappearance rate (ICG-PDR).
Serum samples were collected from 150 patients undergoing the ICG-PDR
in our hospital. Written informed consent was obtained from all participants prior to blood sampling, and the study protocol was approved by
the Ethics Committee of Chiba University Graduate School of Medicine. In
the manual method (standard method), the absorbance of serum samples
before and after ICG loading was measured at 805 nm using a spectrophotometer. Our method is based on the fact that ICG has little absorption
at 885 nm. Absorbance of serum at 805 nm without ICG loading is estimated from that at 885 nm with ICG loading. Subsequently, the estimate
is subtracted from the absorbance of serum after ICG loading. For the
analysis, JCA-BM2250 automatic biochemical analyzer (JOEL) was used.
Samples submitted to our laboratory for ICG loading tests were evaluated
for fundamental performance. The within-run or between-run reproducibility was ＜ 2% CV. The linearity range of concentration was 0-1mg/dL
ICG. The reagent was stable for three weeks or longer after they were set
in an automatic analyzer. The correlation between this (y) and standard
(x) methods was examined in 150 patients: y = 1.04x + 0.006, r = 0.998.
However, patients whose serum samples before ICG loading were turbid
showed some deviation. This method may provide more accurate results
for turbid samples. In conclusion, the long-wavelength control method
does not require blood sampling before ICG injection, and enables the automatic calculation of the retention rate, making it practicable.
Symposium
PE-01
Improve HE staining quality helper for detection of Helicobacter pylori in Gastrointestinal tract biopsy.
Helicobacter pylori, Hematoxylin and Eosin staining, Gastrointestinal tract, surgical pathology, Gastric
cancer
PE-02
Detection Proliferation markers in prostate cancer using
Bioinformatics tools as methodology
Vslidation prolifrration new biomarkers
Case Conference
Oral Presentation
Poster Presentation
Hsing Cheng Tseng, Jia-Bin Liao, Jyh-Seng Wang, Herng-Sheng Lee
Sarkaft Gh Omer, Graham Ball
Department of Pathology and Laboratory Medicine, Kaohsiung Veterans General Hospital, Taiwan
Biology bsc and biomedical science m.sc, Iraq
Background:
Gastric cancer is the third leading cause of cancer related death worldwide
and H. pylori infection is strongly related with many gastroduodenal diseases. Histology is usually considered to be the gold standard in the direct
detection of H. pylori infection and the Hematoxylin and Eosin staining
is usually sufficient for diagnosis of H. pylori infection in routine clinical
histological exam[2]. However, several factors influence the diagnostic
accuracy of histology, such as site, size and number of biopsies, staining
quality and experience of the examining pathologist. The purpose of this
study were to improve H&E stains quality that to identifying H. pylori in
turnaround time.
Prostate cancer is most common in men in the United Kingdome and it
exhibits a huge variability in clinical behavior from indolent condition to
aggressive cancer condition. The current
proliferation-related genes are emerged in tumour cells. The over expression of proliferation genes has been observed in tumour microarray dataset and most of them related to poor prognosis and aggressive cancer [25].
Proliferation-related biomarkers can be used as prognosis and predictor
biomarkers to differentiate aggressive prostate cancer(tiger phenotypes)
to non- aggressive Prostate cancer ( pussy cat phenotype) as well as clinically, proliferation markers can be used to avoid patients from overtreatment, and help physicians to give patients a right therapy. Ki-67-marker
of proliferation- is a surrogate endpoint biomarkers and an independent
predicator of disease recurrence and progression in patients and survival,
especially, who are treated with transurethral resection, radiotherapy or
radical prostatectomy [30, 31, and 71]. Here, in this study, it was attempt
to find new proliferation-related biomarkers and identify a correlation
between proliferation markers which are in some way are associated with
tiger phenotype and pussy phenotype in gene expression dataset by using Neural Artificial Network as a method, which is an bioinformatics tool
used to find interaction between transcripts.
As result, we were identified the best 20 genes related to proliferation
prostate cancer which they will be needed to be validated their function
by RNA interference and , verify its presence in tissue by Immunhistochemstry.
Material and Methods:
First, we're choose some organ tissue to assess quality of H&E stain.
Second, we tried a series temperature and time of heating. And then, we
observation effects of stain that whether Hematoxylin reagent to filter.
Results:
A contaminants ramdan observated in some organs, include lung, stomach,
skin, myoma and oral tissue in microscopy through hematoxylin and eosin
staining. The quality of staining that seems to be no relationship between
the time and temperature of heating in our lab. By careful quality control,
xylene may be a key point in H&E staining procedure.
Discussion:
There is a trend of increasing in the number of the pathological specimens
year by year in our hospital. How to enhance the effectiveness of the work
and to improve the quality and turn around time of the Hematoxylin and
Eosin staining procedure becomes an important issue, and we have succeeded finding out a simple staining procedure that can help pathologist
make a detection of H. pylori within a short time.
110
Clinico-pathological analysis of HER family expression in human colorectal cancer
Co-expression of HER family is clinico-pathological biomarker which relates to
progression in human colorectal cancer
PE-04
Hiroyuki Nozaka1,5, Miku Togashi2, Sayaka Kurosawa3, Ai Igarashi4, Noriyuki Yamada5,
Yayoi Takahashi5, Kazuyuki Ishida5, Tamotsu Sugai5
Chae Jong-hyuck, Hong Sungchul, Ma Sangchul, Kim Kihyun, O Jongwon, Lee Munjung, Park Wookjae
KAMT, Korea,
1 Graduate
school of health sciences, Hirosaki university, Japan
2 Clinical
laboratory, Sendai Medical Center
3 Clinical
laboratory, Yamagata National Hospital
4 Clinical
laboratory, Hachinohe JRC Hospital
5 Iwate
Medical University School of Medicine
Invited Lecture
Special Lecture
Background: As one of the methods of measuring the accuracy of staining in special staining, there is a method of evaluating the accuracy of
staining using positive control. The positive control is carried out in order
to see the structure of a tissue and to check the microorganism infected
functionally or externally, and in AFB staining, it is necessary to evaluate
the accuracy of staining by staining a positive control group, together,
in order to evaluate that. The existing method used positive tissues into
which tubercular bacilli invaded as a positive control, but it became more
difficult to supply this, constantly, as the rate of the initial discovery of
tuberculosis increased. Thus, if the control block is produced, integrated
with an AFB stain by the process of producing a cell block used in a cellular pathology lab, the quantity or distribution of bacteria is constant in the
section because of the characteristic of the cell block, so such a block was
produced as it was expected that it could serve as an AFB positive control
sufficiently.
Materials and methods: The types of the culturing medium include Ogawa
medium, a solid medium and Middlebrook 7H9, a liquid medium. A control
block was produced through the following procedures: First, scoop out the
colony of bacteria cultured on the solid medium, Ogawa medium and mix
physiological saline for vortex mix. Put egg albumin in it like producing a
cell block. Then, spin it in a centrifuge and process it with an automatic tissue processor. On the liquid medium, Middlebrook 7H9, it was produced
in the same way. First, spin the cultured bacteria in a centrifuge. Put them
in a cassette and process them with an automatic tissue processor.
Educational Lecture
1.Background
HER family is composed of four members, and Epidermal Growth Factor
Receptor (EGFR) is a member of HER family. EGFR is expressed in the case of
60-80% of colorectal cancer, and it is reported that the expression of EGFR is
strongly involved in the invasion and metastasis. Meanwhile, it is known that
HER family member formed dimer structure with other HER family member,
but a few reports shows co-expression of HER family and clinicopathological significance in human colorectal cancer. In this study, we examined coexpression of HER family mRNA and protein in human colorectal cancer, and
evaluated significance as the clinicopathological biomarker.
2.Design
Tumors for this study were collected from 60 patients diagnosed with primary advanced colorectal cancer. After the surgery extraction, we collected
a part of both normal and tumor tissues from fresh colon or rectum. The half
of tissue was used for extraction of total RNA with TRIzol reagent, and the
left-behind tissue was fixed in the 10% formalin and embedded in the paraffin (FFPE). EGFR, HER2 and HER3 mRNA expression were analyzed by qRTPCR. EGFR, HER2 and HER3 protein was detected by immunohistochemistry
on FFPE tissue sections.
3.Results
HER2 and HER3 mRNA showed over expression in 38% and 48% of all cases
respectively, and co-expression cases of EGFR/HER3 mRNA was 80% of EGFR
mRNA over expressed cases. Meanwhile EGFR and HER3 showed high expression, and HER2 showed low expression by IHC. The co-expression cases
of EGFR/HER3 was half of EGFR-positive cases. In addition, the co-expression
cases of EGFR/HER3 showed significant difference in the depth of invasion,
but it didn't show significant difference in lymph node metastasis.
4.Conclusion
It was suggested that co-expression of EGFR/HER3 associated with the
progression of the tumor. In our future work, we will study the effect of the
EGFR / HER3 inhibition.
FAST PML/RARA FISH at the suspicion of Acute Promyelocyte leukemia (APL)
Biomedical Laboratory Scientist Vinni Bredahl, Zealand University Hospital,
Department of Surgical Pathology, Denmark
PE-06
Preparation of tissue sections of nail specimens
How to create a high-quality slide of nail tissue.
Introduction:
If a patient morphological or clinical features of acute promyelocyte leukemia (APL) has a translocation of PML/RARA, t (15;17) the patient has APL
and can start treatment right away. It is therefore essential for the diagnosis to perform a rapid FISH PML/RARA.
Normally we perform a quick FISH with a conventional FISH probe, which
can be finished in 5 hours. With the FAST PML/RARA probe from Cytocell,
we can perform the FISH in 2 hours.
Quicker and more accurate diagnosis of nail disease is possible through
histological examination, so it is most important to prepare and provide
quality slides.
The nature of nail tissues in which 50% of the components are keratin
component, it is difficult to cut them and prepare a quality slide, so it is
a burden to a pathologist, and there is a difficulty in diagnosis as well. In
addition, even if they are cut uniformly, the impact of the reagent in the
process of tissue softening may be an inhibitory factor in special staining,
so the process of softening nail specimens requires careful attention.
In the generalized process of preparing nail specimens, it is common to
soften tissues in 4% NAOH solution and cut them into pieces sized 3 to
4micro but this study prepared tissue sections and carried out PAS staining by changing the process as follows: surface softening in 10% HCL solution for 30 min and cutting 1 micro cutting.
As a result of an experiment, there was a falling off of the fungal colony on
the slide in the existing process of softening using NAOH solution; however, the fungal colony remained intact on the slide in the process of softening using HCL solution. In addition, in preparing sections, there were wide
differences in the falling off of the tissues on the slide according to their
thickness.
Thus, the production of tissue sections of nail specimens requires quite
difficult conditions, unlike the existing softening process does, so observations by histopathological examination are needed, but in reality, the
credibility of the histological examination itself is not high. Therefore, it is
judged that it is important for a medical laboratory technologist to prepare
good tissue sections by repeatedly improving research and experiment to
have a good effect on diagnosis and treatment.
Materials and method:
Blood- or bone marrow smears from 15 different patients, 5 suspected of
APL and 10 controls, were analyzed for t(15;17) using the quick FISH and
the FAST FISH method and the agreement between the two methods were
assessed.
The only difference in protocol are the hybridization time for the probe.
The quick FISH hybridizes for 4 hours and the FAST FISH 1 hour. All other
parameters in the protocol are the same.
Results:
There were bright and clear signals with both probes. In 13 cases, the result from the two FISH methods showed no translocation and in two cases,
was a translocation detected. There was complete agreement between the
two methods.
Discussion and conclusion:
There were no difference in the signals, whether it was blood- or bone
marrow smears.
The FAST PML/RARA probe gives a valid result, equal to conventional
PML/RARA probe. Cytocell has produced a probe, that is easy to work with
and the signals obtained with the two probes are comparable.
We now routinely use the FAST PML/RARA probe.
111
Poster Presentation
samsung medical center, Korea
Oral Presentation
sungeui kim, Sungchul HONG, Seungwoo HAN
Case Conference
Vinni Bredahl
Pathology, Denmark
Symposium
PE-05
Producing an AFB Control Block
Keynote Speech
PE-03
Keynote Speech
PE-07
Biobanking of prostate cancer.
A novel technique for harvesting of fresh tissue from radical
prostatectomy specimens.
PE-08
Th2-type cytokines (IL-4 and IL-13) upregulation in patients with immunoglobulin G4-related aortic
aneurysm
Serum and local pathological cytokine valance in patients with immunoglobulin G4-related aortic aneurysm
Jona Gudjonsdottir, Claes Lindh, Lars Egevad
Satomi Kasashima1, Atsuhiro Kawashima1, Zen Yoh2, Satoru Ozaki3, Fuminori Kasashima4
Dept of oncology-pathology, Karolinska Institutet, Sweden
1 Department
of Pathology and Clinical Laboratory, National Hospital Organization, Kanazawa Medical Center, Japan
2 Department
of Diagnostic Pathology, Kobe University Graduate School of Medicine
3 Department
of Clinical Laboratory Science, Kanazawa University
4 Department
of Cardiovascular Surgery, National Hospital Organization, Kanazawa Medical Center
Invited Lecture
Introduction:
Harvesting of representative fresh tumor tissue for cancer research provides better access to genetic information than formalin fixed paraffinembedded tissue. Biobanking of prostate cancer is particularly challenging
because of difficulties in identifying tumor macroscopically. We aimed to
develop a method that allows biobanking of fresh prostate tissue without
hampering morphological assessment.
Special Lecture
Educational Lecture
Objective: IgG4-related aortic aneurysms (IgG4-AA) are one of the most
common lesions of IgG4-related diseases. The pathogenesis of IgG4-related
disease was recently associated with the upregulation of T-helper-2 cytokines (Th2) and regulatory T cells (Treg). In this study, we describe the
cytokine profile of the blood and tissues of patients with IgG4-AA.
Methods: This study included four groups of patients: IgG4-AA (n = 10),
non-IgG4-related inflammatory abdominal aortic aneurysm (non-IgG4IAAA) (n = 5), atherosclerotic AAA (aAAA) (n = 10), and almost normal
aorta without dilatation (n = 10). They were examined using (1) serum
interleukin IL-4, IL-10, IL-13, and interferon (IFN)-g levels and (2) resected
aortic tissues, immunohistochemical and mRNA in situ hybridization (ISH),
local cytokines expression, CD34 (endothelium and mesenchymal cells),
and CD163 (macrophage, histiocytes).
Results: Serum IL-10 levels were significantly higher in IgG4-AA (median
1.3 pg/mL, range 0.5- 27.5 pg/mL) than in the other atherosclerotic arteriopathies. IL-13 levels (median10.3 pg/mL, range 3.3-18.5 pg/mL) were
elevated in two patients with IgG4-AA. The median number of IL-4, IL-10,
and IL-13 immunopositive cells (45.9, 13.0, 102.6 / high power field, respectively) was significantly higher in the adventitia of the IgG4-AA group,
compared with the aAAA and autopsy controls. ISH confirmed the IL-4, IL10 and IL-13 mRNA expression in the adventitia of IgG4-AA. Co-expression
of IL-13-mRNA and CD34-mRNA or CD168-mRNA was detected in IgG4AA.
Conclusion: These results suggested that up-regulation of IL-10 and IL13 in the adventitia of IgG4-AA is related with Th2 and Treg predominant
cytokine balance and pathogenesis of IgG4-AA.
Methods:
A custom-made double-bladed knife was developed for harvesting of tissue from unfixed radical prostatectomy specimens. A 4 mm thick slice was
cut by a horizontal section through the prostate and then split in smaller
segments that were frozen in OCT gel at -80 °C. The rest of the prostate
was pinned to cork, formalin-fixed, sliced at 4 mm, paraffin-embedded and
whole-mounted. This technique was used for biobanking of 20 prostatectomy specimens. Frozen sections were cut from all blocks and cutting time
monitored. A pathologist marked out tumor areas, slides were scanned
and the slice reconstructed.
Results:
A total of 155 frozen blocks were cut (4-9 per case). Average cutting time
per case was 162 minutes (range 79-253 minutes). Tumor was found in
frozen sections of 85% (17/20) of cases and in 46% (72/155) of blocks.
None of the radical prostatectomy specimens were over-sampled (pT0).
Gleason scores were 6, 7 (3+4), 7 (4+3), 8 and 9 in 4, 7, 6, 2 and 1 of the
cases, respectively.
Discussion and conclusions:
This novel method for biobanking of prostatectomy specimens provides
high quality frozen tissue for research purposes while it also allows mapping of cancer distribution for accurate reporting. Cancer is harvested in
most cases and the technique allows identification of tumor location in
multifocal cancer. Disadvantages with this method is that it is labor intensive and requires considerable freezer space, but these problems can be
minimized by cutting only blocks that are likely to contain tumor.
Symposium
PE-09
Case Report: Empyema Caused by Trichomonas tenax in an
Elderly Female Taiwanese
A rare case of empyema caused by Trichomonas tenax.
PE-10
Text-Mining Application in Clinical Document Record of Lung Cancer Pathological Staging
The Influence Factors of Overall Survival for Early Stage Pulmonary Mucoepidermoid
Carcinoma
Case Conference
Hsinchieh Lu1, Hsianglin Wan1, Chiakwung Fan2
Yung-Han Sun1,3, Shih-Wei Lin2,4, Chih-Cheng Hsieh1,5
1 Department
of Laboratory Medicine, Taipei Tzu Chi Hospital , Buddhist Tzu Chi Medicine Foundation, Taiwan
2 Department
of Molecular Parasitology and Tropical Diseases, School of Medicine, College of Medicine, Taipei Medical
University, Taipei, Taiwan
1 Division
of Thoracic Surgery, Taipei Veterans General Hospital, Taiwan
2 Department
of Information Management, Chang Gung University, Taoyuan, Taiwan
3 Graduate
Institute of Business and Management, Chang Gung University, Taoyuan, Taiwan
4 Stroke
Center, Linkou Chang Gung Memorial Hospital, Taoyuan, Taiwan
5 School
of Medicine, National Yang-Ming University, Taipei
Oral Presentation
Poster Presentation
Empyema is one of the potential complications of lower respiratory tract
infections or pneumonia. Infections of the pleural space are usually bacterial in origin; however, in predisposed individuals empyema can be the
result of microorganisms other than bacteria. Here, we present a rare case
of empyema caused by Trichomonas tenax.
A case of a 83-year-old female patient with more than 20 years history of
type II diabetes mellitus had symptoms of cough with sputum and dyspnea for days. At ER , physical examination showed tachypnea respiration
rate, decrease breath sound, wheezing and lower leg pitting edema (3+).
Vital sign was normal. Blood analysis did not show anemia (Hgb: 10.2g/
dl); in contrast, the data showed leukocytosis (WBC count: 20890/ul) with
left shift and impaired renal function (BUN/CRE:37/1.5 mg/dl). Chest xray showed right pleural effusion and chest CT showed RLL pulmonary
abscess and right empyema. The pleural fluid from right pigtail was
examined and showed red-brown color and WBC count reached 83308/
ul. A wet mount of the pleural fluid showed many neutrophils and several
flagellated organisms demonstrating tumbling motility. According to the
size and morphology, we determine that Trichomonas species primary,
and then determine that Trichomonas tenax by using PCR. She denied any
history of smoking, alchol, and betel nut chewing. She was admitted to
ordinary ward for further evaluation and management due to suspected of
pneumonia and acute respiratory failure. The bacterial staining of pleural
fluid was identified to be G (+) Streptococcus sanguinis then antibiotics
of Cravit was given. Pulmonary infections with Trichomonads might be
underestimated because of diagnostic difficulties. The utility of molecular
biology for species identification is underlined and the pathogenicity of
Trichomonad parasites in human lungs should not be ignored.
Introduction
Pulmonary mucoepidermoid carcinoma (PMEC) is a rare tumor of all pulmonary cancers. In previous studies, PMEC was mostly reported in small series
or described in case reports. The aim of this study was to compare the clinical characteristic by text mining in pathological reports, and identify factors
influencing survival of early stage PMEC.
Methods
Pathology reports of 4567 patients undergoing surgery of lung cancer
were collected. Text mining techniques were used to analyze cancer staging
related keywords in pathology reports. The medical records of patients with
a diagnosis of PMEC from January 1991 to July 2015 in pathology reports
were retrospectively reviewed with text mining. The overall survival (OS)
was calculated from the day of surgery to the time of the first relapse (recurrence or metastasis) and day of surgery to the date of death from any cause.
All the statistical analyses were performed using SPSS 16.0.
Results
From 4567 pathology reports, 34 patients were diagnosed as early stage
of PMEC. Most patients were male (70.6%). The mean age of the patients
was 62.1 years and the median follow-up time of the 34 surviving patients
was 47.3 months. In pathological stage, there were 23 (67.6%) patients
with stage I and 11 patients with stage II. The 5-year OS and DFS rates were
74.8% and 72.5%. In univariate of OS, there were significant difference in
age ＞ 65 years (51.5%, P =0.001) and intermediate cell types of high-grade
(68.4%, P ＜ 0.001), there were no significant difference in man (66.7%, P
=0.14) and stage I (76.9%, P =0.749).
Conclusion
The percentage of age less than 65 years and intermediate cell types of
low-grade treated were greater than the percentage of age ＞ 65 years and
intermediate cell types of low-grade for early stage of PMEC in 5 years survival.\
112
MiRNA-21 in Tumor-Derived Exosome as a Novel Diagnostic Biomarker for Oral Squamous
Cell carcinomas.
Exosomal miRNA-21, the Potential as a Novel Biomarker of Oral Squamous Cell carcinomas.
PE-12
Immunohistochemical analyses of human atrioventricular
node using paraffin sections
Objective: Normal cardiac contraction critically depends on electrical impulses generated and conducted by the cardiac conduction system (CCS),
which consists of sinus node, atrioventricular node, His bundle and bundle
branches. CCS ensures coordinated contraction of atria, then ventricles,
to optimize blood delivery and return. As it is often difficult to identify
the CCS, especially atrioventricular node, on usual HE sections, we try to
search for useful immunohistochemical markers applicable to paraffin sections.
Materials and method: The subjects were ten normal human hearts taken
from autopsy cases at Tokyo Medical and Dental University Medical Hospital. Immunohistochemical study was performed on paraffin sections using antibodies against CCS-specific maker proteins, including Connexin 40,
Connexin 43, HCN4 and Tbx3.
Results: The positive membranous expression of Connexin 40 in CCS cells
was observed in AV node (6/9 cases; 66.6%) and in His bundle (8/10;
80%). The negative membranous expression of Connexin 43 was present
in AV node (7/9; 77.7%) and in His bundle (5/10; 50%). The Connexin43
was abundantly expressed in the working myocardial cells. The positive
cytoplasmic expression of HCN4 was found in AV node (10/10; 100%) and
in His bundle (7/10; 70%). Similarly, the positive expression of Tbx3 was
observed in AV node (4/8; 50%) and in His bundle (5/9; 55.5%).
Conclusion: We succeeded in identifying CCS-specific immunohistochemical markers applicable to paraffin sections. Some negative (imperfect)
results warrants further study to search for more specific markers. Furthermore, we will try to study the CCS of the patients with cardiac conduction disturbances to analyze the immunohistochemical relationship to the
arrythmia.
Methods: We have isolate exosomes from primary epithelial cell culture of
both neoplastic and keratinized gingival tissue of 3 oral cancer patients.
We examined the expression of exosomal miRNAs using microarray and
quantitative real-time reverse transcription PCR. MicroRNA-21 was the
most markedly increased in OSCCs patients. Therefore, miR-21 was selected as a candidate for further functional analysis.
Results: In microarray study for the exosomal miRNA expression, there
were 52 miRNAs downregulated or up-regulated in the exosomes of the
cancer cells as compared with those of the keratinized gingival tissue. The
exosomes of the oral cancer cells had significantly higher expression of
miR-21, miR-205 and miR-155. Exosomal miR-21 alone yielded an receiver-operating characteristic curve area of 0.856, with 85.2% sensitivity and
79.1% specificity for distinguishing OSCCs patients from healthy controls.
Exosomal miR-21 was significantly correlated with the TNM stage (r=
0.932; P=0.017). Furthermore, transfection with an miR-21 mimic into
normal gingival cells markedly induced cell proliferation, cell cycle progression, cell invasion and colony formation.
Conclusion: The biomarkers detected in tumor-derived exosomes imply a
potential for exosomes in cancer diagnosis. These data demonstrated that
miR-21 is considered to be a typical 'onco-miR', which limit the activity of
signalling pathways such as AKT and MAPK. Thus, miR-21 may serve as a
promising candidate for the early diagnosis of OSCCs.
Microenvironmental factors for neurofibroma promote differentiation and granule maturation of mast cells.
Accelerated growth of neurofibroma by tumor-induced maturation of mast cells in close association between
tumor cells.
PE-14
Transmission Electron Microscopy contribution in paediatric tumours
diagnosis using residual material from fine needle aspiration
Own method of collecting the specimens for TEM
Introduction:
Small round blue cells tumours in children are often undifferentiated or
poorly differentiated, making difficult to provide a definite diagnosis. In
this retrospective study we assessed, the transmission electron microscopy (TEM) contribution in the diagnosis of tumours in which fine needle
aspiration biopsy (FNAB) had been performed on patients from Paediatric
Oncology, Universitas Hospital, Bloemfontein, South Africa, between 2005
and 2015.
Material and methods:
The FNAB specimens were collected using a syringe with a 22 gauge needle. After collecting the material for cytology, the syringe and the needle
are rinsed with 3 % glutaraldehyde. The specimens obtained are suitable
for TEM, and are processed applying the routine procedures of the laboratory.
Results:
From 196 biopsies, 147 (75%) were submitted for TEM analysis. From
147 cases for TEM, 72 (49%) were contributory to the diagnosis, 33 (23%)
made the diagnosis, 17 (11%) non-contributory and 25 (17%) were inadequate. From 122 specimens suitable for TEM, the most common tumours
were nephroblastomas of 68%. Neuroblastoma represented 14% and other
tumours represented 18%. From 17 cases of neuroblastoma, TEM made
the diagnosis in 14 cases (82.4%) and in three cases TEM was contributory. In 83 nephroblastoma cases, TEM confirmed the cytology diagnosis
in 73%, and made the diagnosis in 15%. The TEM diagnostic contribution
has been 86%.
Conclusion:
A total of 75% of biopsies have been submitted to TEM. The TEM can
support or confirm cytology results and provide a differential diagnosis
through accurate morphological findings, especially in neuroblastomas,
very important for the patient therapy and prognosis. For TEM sampling,
no extra cost or procedures are involved. In conclusion through this study
we confirm that TEM is a valuable tool with a high contributory rate (86%)
in diagnosis of paediatric tumours, using residual material from the FNAB.
113
Poster Presentation
Von Recklinghausen Disease neurofibroma type I(NF1) is autosomal-dominant inherited disease, featuring development of numerous neurofibromas
in the dermis since puberty. Histologically, neurofibroma is a benign tumor
mainly consisting of Schwann cells and fibroblasts. It is well documented
that mast cells scattered in the tumor promote the proliferation of tumors
via soluble factors such as TGF-beta1, basic FGF and chemical mediator,
but details are not disclose. Recent studies suggested that mast cells present mature by interaction with fibroblasts in normal skin. Based on these
knowledge, we assume that fibroblasts and Schwann cells in the tumor
interact with surrounding mast cells and promote their maturation with
increased granules. In turn, the mast cells accelerate the proliferation of
neurofibroma.
In order to clarify, we produced environment of neurofibroma by remodeling tissue with NF1 tumor cells and mast cells in vitro. We employed
NF1 tumor cells obtained from neurofibromas and precursor mast cells
differentiated from human bone marrow monocyte cells (BMMC). We
observed the process of mast cell maturation within "spheroid" which was
built by densely co-culturing NF1 cells with precursor mast cells on 96
low-adhesive plate.
Maturation of mast cell granules was evaluated by their metachromasy using toluidine blue staining. The size and number of mast cell granules were
measured by Transmission Electron Microscope (TEM). As a result, TEM
showed time-dependent increase in mast cell granules in the spheroid. The
distribution of mast cells in the spheroid was analyzed by immunofluorescent staining. Confocal laser microscope revealed that mast cells formed
clusters in the spheroid. We also recognized that spheroids created by coculturing NF1 tumor cells with mast cells was bigger than NF1 tumor cells
only.
These results suggest that neurofibroma promotes differentiation and
granule maturation of mast cells, which in turn accelerate the growth of
neurofibroma.
Oral Presentation
Valerica Necsulescu, Ilse E van der Westhuisen, David K Stone
Anatomical Pathology, SMLTSA, South Africa
Case Conference
Takuya Murakami, Takuya Sakomura, Natsumi Yamashita, Misa Yamamoto, Hiroo Kawano
Yamaguchi University graduate schools medicine and helth science, Japan
Symposium
PE-13
Educational Lecture
Background: Oral cancers account for over 7,000 cases of cancer per
year in Taiwan. Despite advances in treatment, the mortality rate of oral
squamous cell carcinomas (OSCCs) has not changed markedly over the last
few decades. Recent studies have demonstrated that microRNAs in tumorderived exosomes are stably detectable and can serve as useful biomarkers
for cancer. This study was proposed to analyze the exosomal miRNAs released from the oral cancer cells to find their potential for early diagnosis
of oral cancer.
Special Lecture
Muyasar Abdusalam1, Yurie Soejima2, Masanobu Kitagawa1, Motoji Sawabe2
1 Department
of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental
University, Japan
2 Department
of Molecular Pathology, Graduate School of Health Care Sciences, Tokyo Medical and Dental University
Invited Lecture
Ching-Mei Chen1, Chih-Yen Chien2, Chao-Cheng Huang3
1 Department
of Laboratory medicine, Chang Gung Medical Foundation Kaohsiung Branch, Kaohsiung, Taiwan
2 Department
of Otolaryngology, Chang Gung Medical Foundation Kaohsiung Branch, Kaohsiung, Taiwan
3 Department
of Pathology, Chang Gung Medical Foundation Kaohsiung Branch, Kaohsiung, Taiwan
Keynote Speech
PE-11
Keynote Speech
PE-15
Clinical impact on HER2-FISH results by the 2013 ASCO/CAP
Guideline
PE-16
HIDEHIRO IWATA , AYA UMEMURA, YOSHIAKI MIZUNO, KENJI NITTA, HIROE MIZUSHIMA,
HIROYUKI OSADA, SHUKO SEKO, TOYONORI TUZUKI
Secretagogin, a newly found Ca2+ binding protein, containing
neurons in the striatum
Seiko Yasuda
Dept. of Medical Technology and Sciences, school of Health Sciences at Fukuoka, International Univ. of health and welfare, Japan
Japanese Red Cross Nagoya Daini Hospital, Japan
Invited Lecture
Special Lecture
Introduction:Secretagogin (Scgn), a member of the EF hand calciumbinding protein (CaBP) superfamily, was recently found in subsets of
developing and adult neurons. In this study, we examined the expression
of Scgn in neurons of the striatum of rats and mice. We also assessed the
co-localization of Scgn with 4 chemical markers of striatal interneurons,
choline acetyltransferase (ChAT), NOS, parvalbumin (PV) and calretinin
(CR).Results: In rats Scgn-positive neurons were scattered throughout the
whole striatum, which were nearly comparable with PV-positive neurons
in number. They were heterogeneous in their structural features; one
type was relatively large, and the other was relatively small and mainly
located in the peripheral portion of the striatum. The co-localization analyses revealed that Scgn-positive neurons were apparently different from
chemically-defined 4 types of interneurons previously reported, although
they overlapped PV-, ChAT- and CR-positive ones to some extent. In the
mouse striatum, Scgn-positive neurons were far smaller in number and
they appeared to correspond to the smaller type of neurons in the rat
striatum. Scgn-positive neurons in the mouse striatum overlapped ChATand CR-positive ones to some extent, but were distinct from PV- and NOSpositive ones. Conclusion:In this study, we showed that Scgn neurons were
rather numerous in the rat striatum and presumed to be the fifth group
of interneurons. Furthermore they showed prominent species differences
between rats and mice. Future studies should aim to study the structural
features and functions of striatal Scgn neurons in detail.
Educational Lecture
＜ Objectives ＞ The guidelines of American Society of Clinical Oncology/
College of American Pathologists were revised in 2013 (2013ASCO/CAP
guideline). The criteria of immunohistochemistry and FISH to evaluate
HER2 states in breast cancer have been changed. Major changes are as
follows: Positive threshold of HER2 signal to HER2/CEP17 ratio become
more than 2.0, which was 2.2 previously. Average HER2 copy number is
more than 6.0 also become positive even if HER2/CEP17 ratio less than 2.0.
The new criteria for equivocal is HER2 copy number is 4.0 to less than 6.0
in case with less than 2.0 HER2/CEP17 ratio. The aim of this study is to
elucidate how 2013ASCO/CAP guideline influences on clinical breast cancer practice. ＜ Methods ＞ From April 2014 to December 2015, we corrected 307 HER2-FISH tests for breast cancers, including both needle core
biopsy and mastectomy specimen. According to 2013ASCO/CAP guideline,
we reevaluated the results of HER2-FISH tests from their original data. We
also investigated the result of estrogen receptor (ER), progesterone receptor (PGR), HER2 test by immunohistochemistry. ＜ Results ＞ Of 307 cases, 47 (15.4%) cases were positive. Two of 47 positive cases were HER2/
CEP17 ratios were less than 2.0, but HER2 copy numbers were more than
6.0, which had been negative, previously. Thirty-five (11.4%) case were
equivocal. Among these cases, 6 cases were triple negative (ER (-), PGR
(-), HER2 (-)) breast cancers. ＜ Conclusions ＞ 2013ASCO/CAP guideline
increased additional positive and equivocal cases in HER2-FISH testing,
which had been diagnosed as negative by the previous guideline. Not negligible number of cases became equivocal by 2013ASCO/CAP guideline.
In addition, 17.1% cases which were diagnosed as "Triple negative" by the
previous criteria might be candidates for HER2-targeted therapy if diagnosed by 2013ASCO/CAP guideline. Our data suggest that 2013ASCO/
CAP guideline expands candidates for patients receiving HER2-targeted
therapy.
Symposium
PE-18
Developing techniques for differentiating between squamous
cell carcinoma and keratoacanthoma by using iCCD.
PE-19
Evaluation of High-Resolution-Melting-Analysis(HRMA) as a diagnostic tool to detect KRAS/
NRAS(All RAS) and BRAF mutations
with DNA extracted from formalin-fixed paraffin-embedded(FFPE) specimens of colorectal cancer
Case Conference
Yanagita Emmy 1, Matsuoka Ryosuke2, Itoh Tomoo1
Tomoya Minami , Shinichiro Matsuki, Shumpei Mizuta, Takao Komai, Makiro Ishibashi
1 Kobe
University Hospital, Japan
2 Kobe
City Medical Center General Hospital
Hyogo Prefectural Amagasaki General Medical Center, Japan
Oral Presentation
Poster Presentation
Introduction: Anti-EGFR monoclonal antibodies such as cetuximab and
panitumumab inhibit cell growth and proliferation in colorectal cancer.
Recent studies showed that somatic mutations in KRAS/NRAS(All RAS)
and BRAF were predictors of resistance to the anti-EGFR monoclonal
antibodies, because these genes participated in downstream of EGFR in
the signal transduction cascade. Additionally, it has been reported that
patients harboring these mutations not only do not benefit from but may
be harmed by anti-EGFR therapy. Thus the rapid and accurate screening
of All RAS and BRAF mutations is valuable, therefore it has been required
to develop the simple and cost-effective method for these mutations detection.Methods: We described a High-Resolution-Melting-Analysis(HRMA)
for mutation screening of All RAS (codon12/13, codon59/61, codon117,
codon146) and BRAF(V600E) with DNA extracted from formalin-fixed
paraffin-embedded(FFPE) specimens of colorectal cancers, and analyzed
the results of mutation screening by HRMA compared with the results by
PCR-rSSO or direct sequecnce.Results: To date, we tested 33 diagnostic
specimens from patients with colorectal cancer. All analysis results by
HRMA corresponded with the results by PCR-rSSO or direct sequence, and
the sensitivity tests revealed our HRMA protocol to be able to detect 5% of
mutant allele in wild-type DNA. By a coming exhibition at the meeting, we
are going to detect All RAS and BRAF mutations of more specimens using
HRMA.Conclusion: Our study reveals that HRMA is an extremely useful
method for mutation screening in genes with various mutation patterns
such as All RAS and BRAF. In addition, HRMA enables us to detect somatic
mutations in these genes more simply and efficiently.
[Background]Keratoacanthoma and squamous cell carcinoma are characterized by similar clinical presentations that are hard to differentiate.
Because these two diseases require distinct regiments of therapy despite
their similarity, differentiation between them is crucial. To achieve this, we
developed a multiplex immunohistochemical method that can simultaneously stain cells in the G1 and S/G2/M phases and undergoing apoptosis
using 3 markers cdt1, geminin, and H2AX.[Method]The tissue speciments
of keratoacanthoma, squamous cell carcinoma and normal skin cells were
fixed in 10% buffered neutral formalin and embedded in paraffin. We
used immunohistochemistry-based cell cycle detection (iCCD) to compare
the staining patterns of each specimen. (A Novel System to Visualize Cell
Kinetics on Formalin-fixed Paraffin-embedded Tissue. Am J Surg Pathol
2012;36:796-773)[Results]Specimens from normal skin and keratoacanthoma showed organized staining patterns of cells positive for these cell
cycle markers unlike those of squamous cell carcinoma. As compared to
normal skin and keratoacanthoma, squamous cell carcinoma showed an
increased proportion of geminin-positive cells and decreased proportion of
cdt1-positive cells. The result shows that iCCD is superior to conventional
single-color immunostaining, it allows examination for multi-cell populations at one time. In addition, unlike multicolor immunofluorescence
techniques, which requires the use of fluorescence microscopes, iCCD only
requires light microscopes. Western blotting quantitatively examines the
expression of proteins but it cannot determine the cellular origin.[Conclusion]In conclusion, the multicolor immunostaining method such as iCCD
enables the differentiation between two distinct pathologies with similar
the differentiations, such as keratoacanthoma and squamous cell carcinoma.
114
Development of the efficient FISH signal detection by the
FISH method using the immunostaining specimen.
PE-21
Study of rapid FISH method using the IQ buffer.
Introduction:In late years various molecular target drugs are developed,
and the FISH method is enforced with immunohistochemistry staining for
decision for the dosage. The FISH method often hesitates about a judgment to observe it under dark field using a fluorescent microscope whether it is the cell which we should count. We usually judge the cell which we
should count in reference to an H&E staining specimen. However, we do
not necessarily serve as a reference because a slice side is different from
an H&E staining specimen with the FISH specimen. When there are few
neoplastic cells in particular, it is concerned about being judged to be with
a normal epithelium cell or stroma cell by mistake. Therefore the count of
the FISH method is difficult and must perform it carefully.Objective:We can
judge the cell which we should count from only a fluorescent microscope
without an H&E staining specimen exactly. And we examined it to find the
method that could enforce the FISH method where had high precision than
before.Method:It enforced an FISH method to an immunohistochemistry
staining specimen developed by DAB. We immunohistochemistry stained
it with an auto immunity staining device.Results:DAB did not have the selffluorescence under the fluorescent microscope and was able to observe
the FISH signal clearly. Fading of DAB was not seen by the operation of the
FISH method, and the cell fusion was not seen, too. The judgment of the
cell which we should count was easy.Conclusion:An immunohistochemistry stained specimen became able to judge the cell which we should count
only in a fluorescent microscope by enforcing an FISH method. We think
that the difference of the count by the observer must become small if we
use this method.
Introduction:We spend overnight on Hybridization by the conventional
FISH method, and the FISH method is performed for two days. Then we
count FISH signal with a fluorescent microscope. However, we often
reexamine the FISH method by various reasons such as the nonspecific
reaction of the background being too strong or a signal is weak or it is not
detected. Therefore we need two days for inspection more, and a result
report becomes slower and slower. In addition, it is bad with the human
body and environment because formamide is included in conventional Hybridization Buffer. In late years Hybridization Buffer which hybridization
finished in 60-120 minutes was released, and the FISH method became
able to be finished in three hours. A result report was enabled on the day
even if we carried out reexamination. Besides, the human body and environment do not have adverse effects because formamide is not included in
this Hybridization Buffer.Objective:We diluted other companies probe using IQ FISH FAST Hybridization Buffer (IQ Buffer) and performed the FISH
method according to the FISH method for three hours protocol and performed the examination that a signal could detect.Method:Using several
kinds of other companies probes, we diluted it in Hybridization Buffer to
recommend each and carried out the FISH method according to a protocol
to recommend. We performed the FISH method according to the protocol
of the FISH method for three hours when the same probe was recommended in diluted form using IQ Buffer. Other reagents used the reagent
which the company of each probe recommended.Results:The FISH method
was possible for three hours even if we diluted other companies probe using IQ Buffer.
Study of the efficient FISH signal detection and rapid FISH
method.
PE-23
Three dimensional imaging analysis of the promotion process
from BCAC to colonic adenomas
Symposium
PE-22
Educational Lecture
Kobe university hospital, Japan
Special Lecture
Endo Akikazu , Yanagita Emmy, Yamada Hiroshi, Tsukamoto Ryuko, Itoh Tomoo
Kobe university hospital, Japan
Invited Lecture
Endo Akikazu , Yanagita Emmy, Yamada Hiroshi, Tsukamoto Ryuko, Itoh Tomoo
Keynote Speech
PE-20
Kazuo kase 1, Nobuko Saito1, Yuji Sato1, Kenzou Hirosima2, Yukari Okita3, Yukihide Watanabe3,
Hiroyuki Suzuki3, Mituyasu Kato3
Endo Akikazu , Yanagita Emmy, Yamada Hiroshi, Tsukamoto Ryuko, Itoh Tomoo
Kobe university hospital, Japan
Poster Presentation
115
Oral Presentation
Introduction : Early process of colon adenoma formation is not fully understood, though cancer cells as well as tumor microenvironment are thought
to have important roles. In this study, we examined cell proliferation status
of neoplastic cells and microvascular structures during the process of early
adenoma development from s-catenin accumulated crypts(BCAC) in mice
models.Methods : Dextran sodium sulfate(DSS) was given to ApcMin/+mice at
5 weeks of age in drinking water for 7 days to induce colonic inflammation
and adenoma formation. Serial sections of whole BCAC were stained for Ki67 and podocalyxin, and reconstructed the 3 dimensional images using VG
studio Max 2.0. Ki-67 positive and negative cell numbers and the vascular
structures were quantitatively analyzed during the early developmental
process of colon adenoma.Results : ApcMin/+mice has multiple BCACs in the
colon but BCACs rarely develop to colon adenoma. DSS initiated adenoma
formation and visible adenomas with multiple branching structures were
develped within 3-4 weeks. Increase of Ki-67 positive cells in BCACs was
observed as early as 5thday during DSS treatment and reached to 90% of
the total cell numbers in 3-4 weeks. Then, it was finding of shape rise from
at 1 to 4 weeks after DSS. At the same time, in the surrounding microenvironments, neovascular formation and increase of spindle shaped stromal
cells were also detected during the early time points of adenoma formation.Conclusion : DSS treatment induced developmental transition of BCAC
to adenoma in the colon of ApcMin/+mice. During this process, neovascular
formation and increase of spindle shaped stromal cells accompany to adenoma formation from the early time points.
Case Conference
1 Edogawa
Medical Laboratory Center, Japan
2 Tokyo
Womens Medical Univ.Yachiyo Medical Center
3 Dept.of
Experimental Pathology,Faculty of Medical,Univ.of Tsukuba
Introduction:In late years various molecular target drugs are developed,
immunohistochemistry staining and the FISH method are indispensable
to the adaptation. However, we need two days before a result is given by
the conventional FISH method. In addition, we must carry out reexamination because a signal is not often seen depending on a specimen. The fine
adjustment of the preprocessing process is necessary for this reexamination and needs two days extra. Furthermore, the FISH method is observation under the dark field, and it is difficult to distinguish the cell which
we should count.Therefore we spend time with great labor to look for the
signal of the objective cell and must count it.Object: We shortened time to
spend it for the FISH method and performed the examination that could
not distinguish the cell which we should count more under dark field.
Methods:Because the FISH method of three hours was possible, by a method using IQ FISH FAST Hybridization Buffer (IQ Buffer), even the probe of
other companies diluted a probe using IQ Buffer. The FISH method obeyed
the FISH method of three hours of IQ Buffer. In addition, we used the
specimen which carried out immunohistochemistry staining because the
effective detection of the FISH signal was enabled by performing the FISH
method using an immunostaining specimen developed by DAB.Results:We
could distinguish only the signal of the objective cell under dark field, and
a report was enabled as a result of FISH method in half day.Conclusion:By
this technique, we can give molecular pathological diagnosis and treatment quickly.In addition, we think that we can contribute to the life convalescence of the patient very much.
Keynote Speech
PE-24
Assessment of leucine-rich repeat-containing G-protein-coupled
receptor 5 expression in colorectal neuroendocrine tumor
PE-25
Tomoyuki Nakajima1, Takeshi Uehara1, Yuko Otani2, Yasuhiro Maruyama1, Yukihiro kobayashi1,
Takayuki Honda3
Yurie Soejima 1, Motoji Sawabe1, Takumi Akashi2, Yoshinobu Eishi2, Toshio Fukusato3
1 Department
of Molecular Pathology, Graduate School of Health Care Sciences, Tokyo Medical and Dental University, Japan
2 Dept.
Human Pathology, Tokyo Medical and Dental Univ. Graduate School
3 General
Medical Education and Research Center, Teikyo Univ.
1 Shinshu
University Hospital, Japan
2 Niigata
Prefectural Central Hosp.
3 Shinshu
Univ.
Invited Lecture
Special Lecture
Educational Lecture
Introduction: We have reported that the expression levels of Α 6 Β 4( Β
4) and Α v Β 6( Β 6) integrins was significantly higher in intrahepatic
cholangiocarcinoma (ICC) than in cholangiolocellular carcinoma. Because
ICCs have various biological behavior and prognosis, we analyzed the
expression of Β 4, Β 6 integrins in ICC immunohistochemically and compared the results with the clinicopathlogical characteristics of subtypes of
ICC.Materials and Methods: After alcian-blue mucin stain, and CK7 and
HepPar1 immunostain for the diagnosis of ICC, 48 surgical cases of ICCs
were examined by immunohistochemistry using anti- Β 4, Β 6 integrin
antibodies on formalin-fixed paraffin-embedded tissue sections to evaluate the relationship between integrin expressions and clinicopathological
parameters.Results: The expression of Β 4 and Β 6 was demonstrated in
46 (96%) and 35 (73%) ICC cases, respectively. We divided ICCs into two
groups, negative or low expression groups ( Β 4:29 cases, Β 6:36 cases)
and high expression groups ( Β 4:19 cases, Β 6:12 cases). The integrin expression levels were higher in perihilar localization type than in peripheral
localization type ( Β 4:70% vs 32%, p<0.05, Β 6:50% vs 18%, p=0.054),
higher in periductal infiltrating gross type or intraductal growth type than
in mass-forming gross type ( Β 4:100% vs 31%, p<0.005), lower in poorly
differentiated type than in well or moderately differentiated type ( Β 4:8%
vs 51%, p<0.01, Β 6:0% vs 34%, p<0.05), and was higher in infiltrative
growth type than in expansive growth type ( Β 4:56% vs 22%, p<0.05, Β
6:36% vs 13%, p=0.065). In addition, the integrin expression was related
to bile duct involvement ( Β 4:58% vs 18%, p<0.01, Β 6:35% vs 14%,
p=0.089). Conclusion: These results might indicate that the expression of
Β 4 and Β 6 integrins are associated with invasion and progression of
ICC. Ligands for integrins will be investigated in future studies.
Introduction: Leucine-rich repeat-containing G-protein-coupled receptor
5 (Lgr5) has been identified as a putative intestinal stem cell marker. Lgr5
is also expressed in various tumors. In this study, we investigated Lgr5
expression in colorectal neuroendocrine tumor (NET) and analyzed its
pathological characteristics.Methods: We evaluated the clinicopathological features of 8 NET grade 1 (G1), 4 NET grade 2 (G2), and 4 NET grade
3(G3) (termed neuroendocrine carcinoma (NEC)) cases and Lgr5 expression using an RNAscope, a newly developed RNA in situ hybridization
technique, with a tissue microarray of the NET samples. Lgr5 staining in
individual tumor cells was scored semi-quantitatively using an H-score
scale. Ki-67, ß-catenin, and synaptophysin were also examined in all cases
by immunohistochemistry. We performed a combination of Lgr5 RNA
in situ hybridization and synaptophysin immunohistochemistry.Results:
All cases contained tumor cells with some Lgr5 -positive dots. Both Lgr5 positive and synaptophysin positive cells were observed in most cases. In
all cases, although rather weakly, H-scores showed a positive correlation
with nuclear ß-catenin expression (r= 0.5004, p=0.0484). There was no
correlation between H-score and Ki-67 expression. In the NET G1 and
NET G2 groups, there was also no correlation between H-score and Ki-67
expression, or between H-score and ß-catenin expression. In NEC there
was strong negative correlation between H-score and Ki-67 expression (r=
– 1, p ＜ 0.0001). In NEC there was a strong positive correlation between
H-score and ß-catenin expression (r= 1, p ＜ 0.0001).Conclusion: Lgr5 expression might be affected by ß-catenin expression in NET and especially
in NEC. The mitosis ability of Lgr5 -positive cells might be low in NEC.
These characteristics suggest that Lgr5 -positive cells may be stem cells in
NEC. A further study with a larger number of NEC cases is warranted to
confirm the conclusions.
Symposium
PE-26
Immunohistochemical studies of beta4 and beta6 integrins in
intrahepatic cholangiocarcinoma
Improved Azan staining protocol and the utility
-clear color contrast and short staining time-
PE-27
Shigenobu Tatsumi , Takeshi Nishikawa, Hisae Suzuki, Mao Takeuchi, Yoshimasa Fukui, Kyouko Tanaka,
Yayoi Umeki
Glyoxal fixation is useful for histological evaluation.
A comparative analysis of tissue fixation with formaldehyde
and glyoxal.
Fuminori Sakanashi , Mami Mizuno, Misaki Uehara, Koji Tateishi, Masazumi Tsuneyoshi
Fukuoka Sanno Hospital, Japan
Case Conference
Hospital Pathology, Nara Medical University Hospital , Japan
Oral Presentation
Poster Presentation
Introduction:Formalin has been the leading fixative in histopathological
examination. However, carcinogenic of formaldehyde was pointed out by
World Health Organization(WHO) in 2004, and now the use and disposal
of formalin are monitored and regulated strictly in Japan.Glyoxal, a nonformaldehyde-containing histological fixative, is one of the most popular
substitutes for formalin, and said to have the advantage of being less
hazardous than formaldehyde.Our purpose of this study is to verify the
usefulness of glyoxal fixed specimens, especially focusing on histological
staining.Materials and Methods:We assessed a plurality of biopsy specimens obtained from the same organs of the same patient (47 cases).Some
of them were fixed with formalin, and the others glyoxal-based fixative
for 18-24 hours.Hematoxylin and Eosin stain, special stains (Elastica van
Gieson stain, Masson’s trichrome stain, Fontana Maason stain, and Alcian
blue – PAS stain), and immunohistochemical stains (3 kinds of cytokeratin,
E-cadherin, smooth muscle actin, HER2 for cytoplasm or cell membrane
and TTF-1, p63, Estrogen receptor, Progesterone receptor, Ki-67 for nucleus) were performed in the same protocol to determine the difference in
dyeability.Results:There wasn’t a significant difference between the two
fixation methods in almost all staining.For immunohistochemical nuclear
staining, both the intensity of staining and the frequency of stained cells
were attenuated or disappeared with glyoxal fixed specimens as compared
to formalin-fixed, but we have succeeded in recovering the similar staining
by trying several different conditions.Conclusion:Considering the environmental and health concerns, the non-formaldehyde-containing tissue
fixative “glyoxal” is a useful substitute fixative for histological evaluation in
pathology laboratories.
[Objectives] We have reported about a new pre-treatment method and
mechanism of Azan staining in years. In this report, we introduce improved Azan staining which has a clearer color contrast and shorter staining time.[Materials and Methods] Liver fixed with 10% neutral buffered
formalin were adopted. After deparaffinizing and rinsing with running
tap water, the sections were incubated in our developing saturated aqueous solution of picric acid including 0.1% azocarmin G at 60 °C for 30
min and rinsed briefly in distilled water. Next, the sections were replaced
in 5% phosphotungstic acid aqueous solution at room temperature (RT)
for 30 min and rinsed briefly in distilled water. After rinsing, the sections
were soaked in aniline blue/orange G staining solution at RT for 15 min.
And then, the sections were rinsed and dehydrated through pure alcohol,
changed in xylene, and finally coverslipped (improved Azan staining). Also,
in case of adding elastic fibers staining, after deparaffinizing, the sections
were soaked in resorcin fuchsin solution at RT for 50 min and dipped at
ten times in 70% alcohol including 1% hydrochloric acid(Elastica improved
Azan staining).[Result] Improved Azan staining gained the clear color
contrast between collagen fibers and cellular components because the
staining to cytoplasm of aniline blue was prevented. Collagen fibers were
blue by aniline blue, cytoplasm was red by azocarmin G, nuclei were deep
red by azocarmin G, and erythrocytes were yellow or orange by picric acid
or orange G. This improved Azan staining required about 1.5 hours totally,
at least about more than an hour shorter than conventional Azan staining.
Similarly, Elastica improved Azan staining acquired the clear color contrast.[Conclusion] We acquired the higher tissue selectivity and established
a new staining method which has shorter time and less process than a
conventional method.
116
PE-29
Development and utility of improved silver protein
Using ultrasonic waves to infiltrate resin into electron
microscopic specimens
Nobuo Kuninaka 1, Yuko Sato2, Yoshiyuki Umeto2, Masanobu Higo1, Shigeo Murayama3, Yuko Saito2
Yamada Hiroshi 1, Yanagita Emmy2, Morito Satoshi2, Endo Akikazu2, Tsukamoto Ryuko2, Ito Tomoo2
1 National
Hospital Organization Yokohama Medical Center, Japan
2 National
Center of Neurology and Psychiatry
3 Tokyo
Metropolitan Geriatric Hospital & Institute of Gerontology
1 Kobe
University Hospital, Japan
2 The
Department of Diagnostic Pathology of Kobe University Hospital
Yuichiro Cho , Kenji Sato, Osamu Hoshi
Immunohistochemical study of PD-L1 expression and its
clinicopathological correlation in invasive non-small cell lung
cancers
Symposium
PE-31
Educational Lecture
Reconstruction of the canine pelvic nerve and its application
to substitute bladder
Special Lecture
PE-30
Invited Lecture
[Introduction]For creating transmission electron microscopic specimens,
resin (plastic-like material) is used in the final embedding to withstand the
heat of electron beams. Compared with paraffin, resin is relatively viscous
and low in permeability, often scattering when sliced into ultra-thin pieces,
which is due to poor permeability even if many processes and much time
are used. Resin also often shrinks or breaks when photographing. We
examined using an ultrasonic wave device to promote infiltration when
embedding resin.[Method]In our hospital, we usually use a vacuum pump
to promote infiltration when embedding a specimen in resin. A specimen
is placed in pure resin (Epon812) and a vacuum pump is used to withdraw
it for approximately 1 hour. The vacuum state is maintained for one night.
It is put into a beam and polymerized on the next day.We examined using
ultrasonic waves with the Histra-DC (manufactured by JOKOH CO., LTD.),
which is a device for securing, degreasing and decalcifying that uses ultrasonic waves instead of a vacuum pump. We used [skin] as a specimen, and
we found that the infiltration is not enough because of poor permeability.
[Results]Compared with specimens that were treated with vacuum pumps,
specimens treated with Histra-DC were less damaged when they were
sliced and photographed.[Conclusion]Using ultrasonic waves promotes
infiltration of reagent into tissues because of cavitation. However, using
a regular ultrasonic generator causes resin polymerization due to heat
generated by the ultrasonic waves. The Histra-DC has a cooling apparatus,
therefore preventing increased temperatures; favorable resin filtration can
be performed. After the vacuum pump stopped, the vacuum state could be
maintained for a few hours. After that, however, the air pressure returned
to atmospheric pressure. If the operation continues for a night, oil flows
backward. Continuous operation was not appropriate. We report these results along with images.The Department of Diagnostic Pathology of Kobe
University HospitalHiroshi YamadaTEL：+081-078-382-6474e-mail：
r50bmw1@medo.kobe-u.ac.jp
Introduction:Since the discontinuation of the silver protein
product(MERCK) in 2009, there has been no silver protein suitable for use
with Bodian staining. Thus, it was inevitable to either abandon this staining procedure or use immunostaining instead. In this situation, Bodian
staining may be in danger of disappearance. However, it is an indispensable, critical staining method for facilities specialized in neuropathology.
Therefore, we prepared our own silver protein referring to the report of
Hattori (Rinsho byori.1971) and were able to obtain stainability which
can be substituted for that of MERCK’s silver protein product(Kuninaka,
Neuropathol. 2013). However, the previous protocol was complicated as
silver nitrate oxide (AgNO3) had to be prepared for each use, and axons
were unstably stained. Purpose:To study whether more stable stainability
can be obtained by simplifying the silver protein preparation via replacement of silver nitrate oxide, requiring preparation at each use, with a commercially available product. Matrials & Methods:We employed paraffinembedded sections of central and peripheral nerves fixed with acid and
neutral buffered formalin. As for diseases, Alzheimer's disease, Lewy body
disease and multiple sclerosis were selected. The MERCK staining results
were compared with the stainability of silver protein either prepared by
using AgNO3 prepared at the time of use or commercially available silver
nitrate oxide. Results:Stainability results similar to those obtained with
the MERCK product were achieved with minor adjustment of the amount
of commercially available AgNO3 added. The unstable staining associated
with the preparation required for each use also disappeared. As for fixing
liquid, acid formalin fixation was optimal, though neutral buffered formalin was also sufficient.Conclusion:As more stable stainability was more easily obtained than with the previous method, we demonstrated that Bodian
staining can still be used.However, the problem with this silver protein is
that it cannot be stored due to its liquid form.
Keynote Speech
PE-28
Yuji Uno , Maiko Taira, Kanna Kishimoto, Naoko Akiyama, Masatoshi Sado, Naoyuki Miyokawa,
Hidehiro Takei
Graduate School of Health Care Sciences, Tokyo Medical and Dental University, Tokyo, Japan
Poster Presentation
117
Oral Presentation
Introduction: Recently, immune checkpoint inhibitory approaches have
shown considerable promise as innovative effective therapies for cancer
patients. For lung cancers, especially non-small cell lung cancers (NSCLCs),
two immune checkpoints, PD-1 and PD-L1, have emerged as important
targets for immunotherapy. Our aim is to correlate immunohistochemical
(IHC) expression of PD-L1 with clinicopathological parameters in NSCLC.
Methods: We examined 99 cases (63 men and 36 women) of invasive
NSCLC (Stage IA to IIIA), including 41 squamous cell carcinomas (SCCs)
and 58 adenocarcinomas (ADCs), resected in our institution for a recent
2-year period. One representative histologic section of each case was immunostained with anti-PD-L1 antibody. The extent of immunoreactive
tumor cells and infiltrating intratumoral lymphocytes was respectively
scored in a semiquantitative fashion. The proportion score (PS) was statistically correlated with multiple clinicopathological parameters. In addition,
for the purpose of dichotomization, immunoreactivity seen in ＞ 1% of
tumor cells was considered IHC "positive". Results: PD-L1 PS was significantly higher in SCCs than in ADCs (p=.0154), and 56.1 and 32.8% IHC
"positive" cases were found, respectively. In addition, the same was true
in intratumoral lymphocytes (p=.0125). There was a positive correlation
between the pathologic stage and IHC PS (p=.0131). No correlation was
found between PS and gender, PS and age, PS and tumor size, or PS and
lymphovascular invasion status (i.e., present or absent). Conclusion: PDL1 expression was more significantly seen in higher pathologic stages of
NSCLC cases, which might be associated with tumor immune evasion. This
finding further supports the fact that the anti PD-1/ PD-L1 therapy could
be of potential use in immunotherapy for patients with advanced NSCLC.
Case Conference
Asahikawa Medical University Hospital, Japan
The pelvic nerve (PN) and the hypogastric nerve (HN) play crucial roles in
the control of bladder function, and the colonic nerve (CN) controls only
colon function. This study was undertaken to explore the possibilities of
restoring and preserving PN function after PN-HN-CN reattachment in
transplantation of the colon to bladder as a substitute bladder. PN regeneration and functional reconstruction of substitute bladder were investigated in five dogs. Responses of substitute bladder to electrical stimulation
in the segments of PN and HN were examined in the dogs that had undergone autotransplantation of the colon to bladder as a substitute bladder
with PN-HN-CN reattachment. Eighteen months after unilateral PN-HNCN reattachment, the dogs were anesthetized, and electrical stimulation
in the segments of PN and HN elicited the elevation of substitute bladder
pressure in all five dogs examined. The constriction pressure of substitute
bladder was between 4 mmHg minimum and 15 mmHg maximum with an
average of 8.3 mmHg. Regeneration and ultrastructural alteration of sutured PN-HN-CN were investigated using hematoxylin and eosin staining,
silver impregnation staining, immunohistochemical staining, and electron
microscopy method. Regenerated nerve fibers were existed in all segments
of sutured PN-HN-CN tissue. In the segment of PN-HN, the presence of
increased numbers of connective tissue and disordered arrangement of
small axons were observed. There were more deformed nerve fibers and
scattered small axons which make Schwann cell units into the increased
connective tissue in the distal segment of PN-HN-CN than in the proximal.
Thus, regeneration of PN in sutured PN-HN-CN was confirmed. The above
results indicate that the function of PN can be restored to the colon as a
substitute bladder after PN-HN-CN reattachment, and that the transplanted
colon to bladder can become functionally advanced substitute bladder.
Keynote Speech
PE-32
Immunohistological analysis of hmlh1 expression in
carcinogenesis of endometrioid adenocarcinoma grade3
PE-33
Sayaka Kobayashi 1, Otona Oikawa2, Mayu Kikuchi2, Tomomi Yoshida1, Toshio Fukuda1
Rie Nakata 1, Takeshi Uehara1, Yui Nakashima2, Tomoyuki Nakajima1, Yasuhiro Kinugawa3,
Yasuhiro Maruyama1, Yukihiro Kobayashi1, Takayuki Honda3
1 Dept.
of Histopathology and Cytopathology, Graduate School of Health Sciences, Gunma Univ, Japan
2 School
of Health Sciences, Faculty of Medicine, Gunma Univ, Japan
1 Shinshu
University Hospital, Japan
2 Shonankamakura
General Hosp.
3 Shinshu
University
Invited Lecture
Special Lecture
Educational Lecture
Introduction Type I endometrial carcinoma (Type-I) shows endometrioid
adenocarcinoma Grade 1 (G1) or 2 (G2). Type II endometrial carcinoma
(Type-II) shows serous, clear cell and endometrioid adenocarcinoma
Grade 3 (G3). We previously reported that hMLH1 expression loses at an
early of Type-I carcinogenesis. Occurrence of serous and clear cell adenocarcinoma of Type-II is reported to be caused by other cancer-causing
mechanisms. However, G3 carcinogenesis is not understood. And there are
coexistent G3 cases in G1, G2. Therefore, it is necessary to analyze separately Type- I, II. We assumed that G3 cases coexisted hyperplasia, G1 and
G2 was Type-I (I-G3), and G3 cases not coexisted them was Type-II (II-G3).
And we analyzed hMLH1 expression by immunohistochemistry in G1, G2
and G3 to analyze relationship between hMLH1 expression and I-G3 or IIG3 carcinogenesis. Material and Methods From 82 cases of endometrioid
adenocarcinoma, formalin-fixed paraffin-embedded sections were stained
with anti hMLH1 antibody. We evaluated percentage of positive tumor
cells, and staining intensity, and classified into three groups: negative, low
and high expression.Results hMLH1 lost in 7/14 (50%) cases of I-G3, 2/6
(33%) cases of II-G3. hMLH1 negative were observed in both I-G3 and IIG3. In the study of non-cancer lesion (NCL) and cancer lesion (CL), degree
of hMLH1 negative in CL and NCL of G1 and G2 were almost the same.
However, degree of NCL were lower than CL of I-G3 and II-G3, it was 0%.
We suggested that hMLH1 expression loses before glands shows morphological changes of atypia in G1, G2, whereas hMLH1 expression loses after
glands shows morphological changes to carcinoma in G3. ConclusionRegardless of Type I, II, time of hMLH1 loss-expression in G3 were later than
G1, G2. And G3 may have different cancer-causing mechanism.
Symposium
PE-34
Use of immunohistochemistry with antibody cocktail of IgG
subclasses instead of IgG in IgG4-related diseases
Introduction: Immunoglobulin G4 (IgG4)-related diseases (RD) are systemic diseases that frequently show elevated serum IgG4 levels and tumor-like
masses with infiltration of IgG4-bearing plasma cells. Although diagnostic
criteria of IgG4-RD indicate an IgG4/IgG ratio ＞ 40%, it is often difficult to
count IgG-positive cells because of the weakness of IgG staining intensity.
Because use of an antibody cocktail containing mixed IgG1, IgG2, IgG3,
and IgG4 might have similar results to IgG immunohistochemistry in IgG4RD organs, we compared antibody cocktail reactivity with the expression
of IgG in autoimmune pancreatitis (AIP), a representative IgG4-RD.Methods: Five AIP cases were selected using medical records. To determine the
usefulness of the antibody cocktail, immunohistochemistry was performed
and compared with IgG alone. The coefficient of variation was used to
analyze differences between antibody cocktail and IgG in AIP by 5 boardcertified pathologists.Results: There was no difference in cytoplasmic
intensity of cells classified as positive between the antibody cocktail [3(2
– 3)] and IgG [3(2.5 – 3)] (p=0.8130). There was a significant difference in
background intensity between the antibody cocktail [1(1 – 2.5)] and IgG
[1(0.5 – 1)] (p=0.0497). Although there was no difference in mean values
of IgG-positive plasma cells between the antibody cocktail [46.0(24.3 –
57.7)] and IgG [51.3(23.3 – 74.7)] (p=1.000), the coefficient of variation
value was lower in the antibody cocktail (27.8%) than in IgG (35.2%).Conclusions: The antibody cocktail might be used to count IgG-positive cells in
place of IgG because of the lower coefficient of variation value. A further
study with a larger number of AIP cases is warranted to confirm the conclusions.
Using ultrasonic waves to promote delipidation and
decalcification
PE-35
A case report : Myeloid sarcoma with chromosomal
aberration
Observation of tumor cells using the chromosome analysis
Case Conference
Yamada Hiroshi 1, Yanagita Emmy2, Endo Akikazu2, Tsukamoto Ryuko2, Ito Tomoo2
Masaru Nakamura 1, Yoshiya Goto1, Masayo Shuto2, Kouji Nagata1, Masanori Yasuda1, Takao Tashiro3
1 Kobe
University Hospital, Japan
2 The
Department of Diagnostic Pathology of Kobe University Hospital
1 Dep.
Pathology, Saitama medical Univ. International Medical Center, Japan
2 Faculty
of Health and Medical Care, Saitama Medical Univ.
3 Graduate
School of Arts and Science, The Open University of Japan
Oral Presentation
Poster Presentation
[Introduction]Pathological sample creation has many processes such as fixing, cutting, delipidating decalcifying, paraffin infiltrating, block creating,
slicing and dyeing. Reporting pathological diagnosis takes much time. The
delipidation and decalcification operations are the most time-consuming
processes. If these processes can be shortened, the turn-around time (TAT)
can be speeded up. We examined shortening the delipidation and decalcification processes with Histra-DC (manufactured by JOKOH CO., LTD.)
that uses ultrasonic waves to rapidly fix, delipidate and decalcify samples.
[Method]1) Delipidation: Compare the time to complete delipidation between the regular method (ethanol delipidation method) and ultrasonic
wave method (US method) with Histra-DC for mammary gland2) Decalcification: Compare the time to complete decalcification between the regular
method (still-standing method) and US method with decalcifying reagent
(Plank-Rychlo, 10% formic acid, EDAT, etc.) for bone tissue[Results]1)
Delipidation: The time to complete delipidation using the US method was a
fifth of the regular method. The dyeability of HE dyeing and immunological dyeing stood comparison with the regular method.2) Decalcification:
The time to complete decalcifying using the US method was a third of the
regular method. The dyeability of HE dyeing and immunological dyeing
stood comparison with the regular method. For immunological dyeing, the
dyeability of the US method seemed to be better than that of the regular
method because the time exposed to decalcifying liquid was shorter.[Conclusion]Using ultrasonic waves promotes infiltration of reagent into tissues
because of cavitation. This effect can also be observed in delipidation reagent and decalcifying reagent; the heat generated by ultrasonic waves also
helps promote this reaction. Accordingly, the use of Histra-DC with ultrasonic waves is effective for shortening TAT and improving the dyeability in
decalcification.The Department of Diagnostic Pathology of Kobe University
HospitalHiroshi YamadaTEL：078-382-6474e-mail：r50bmw1@medo.
kobe-u.ac.jp
[Introduction]According to WHO classification, MS is defined as a tumor
mass consisting of myeloid blasts with or without maturation occurring
at an anatomical site other than the bone marrow. It is important to distinguish from lymphoma / leukemia in the pathological diagnosis, to see
the eosinophilic myelocyte to HE specimen in high differentiation MS is an
indicator of the differential diagnosis.[Case]A 10 years old boy. Tummor
lesions in the anterior mediastinum on chest CT was observed and pointed
out the heart effusion. Clinically was implemented doubt thoracotomy biopsy and pericardial inspection the T lymphoblastic lymphoma / leukemia
.[Histological findings]Atypical cells with high N / C ratio with irregular
nuclei are monotonically growth. IHC stainig were performed on paraffin sections. MPO weakly positive cells was observed few to scattered.
Eosinophilic myelocyte was found very few.[Findings of pericardial cell
block]High N / C ratio of atypical cells were mainly. MPO-positive cells war
observed. It was also observed scattered and partial leaves eosinophils, eosinophilic myelocyte in the background.[Cytology]Atypical cells with high
N / C ratio is partially karyotype irregular in somewhat large compared
to the mature lymphocytes, nucleoli were seen. [Chromosome inspection]
Tumor culture: 46, XY, inv (9) (p12q13), t (10; 11) (p13; q14)Pericardial
culture: 48, XY, inv (9) (p12q13), t (10; 11) (p13; q14), + 4, + 19[Consideration]This case is observed monotonically lymphoblastic cells in the
biopsy tumor tissue, in the pericardial cell block from the fact that showed
the differentiation trend. Consider the differentiation trend of tumor cells
to add the IHC.In addition, t(10; 11) detected by chromosome analysis and
compares the biopsy tumor tissue and pericardial specimens using FISH
method .
118
To make multiplex immunohistochemistry more efficient and
quicker in Electric Field Non-Contact Techniques.
PE-37
Developing techniques for differentiating between clear cell
adenocarcinoma and serous adenocarcinoma in ovary.
[background]1.Therefore, accurate differentiation between clear cell adenocarcinoma and serous adenocarcinoma is essential for the selection of
appropriate therapies. 2.We have recently developed a new IHC method
based on an alternating current electric field to facilitate the antigen-antibody reaction (R-IHC). This technique was developed for non catalytically
mixing microdroplets. In this method, an alternating-current (AC) electric
field is used to promote the antigen antibody reaction within the microdroplet. According to this technique, it is possible to immunostaining only
30min. [purpose]We demonstrate the ability to develop techniques for
differentiating between clear cell adenocarcinoma and serous adenocarcinoma in ovary.[experimental]We have employed antibody cocktail and RIHC to study differentiating between clear cell adenocarcinoma and serous
adenocarcinoma in ovary.We designed a rapid multiplex immunostaining
method using a novel 2 antibody cocktail. This antibody cocktail consists
of the following antibodies: rabbit for HNF-1beta, mouse for WT-1. All procedures can be completed within 3 hours. This method labels the nuclei
of clear cell adenocarcinomas as blue with HNF-1beta. Serous adenocarcinomas could be differentiated from clear cell adenocarcinomas with an
inverse staining pattern : brown nuclei with WT-1. The staining procedure
was performed using R-IHC. The sections were incubated with antibody
for 7 minutes followed by the secondary antibody for 7 minutes.Comparing the staining[results]According to this method, it is possible to staining
only 30minutes. And this method have proven that two antigens on the
one glass. This was easily achieved by using the R-IHC. The results showed
the effectiveness of the proposed method.
PE-40
Improved efficiency for an auto slide preparation system
Pathologic features of desmoplastic malignant mesothelioma
Symposium
PE-39
Educational Lecture
[Background]1. Immunohistochemical (IHC) examination plays an important
role in differentiating various tumors. Although multiplex immunohistochemistry only requires a single slide glass in detecting antigens, it is timeconsuming, especially in the staining process.2. We have recently developed
a new IHC (called R-IHC) method in which alternating current electric field is
applied to mix microdroplets on the slide and thus to facilitate the antigenantibody reaction. In this method, it only takes 30 minutes for immunostaining.[Objective]To seek a quicker procedure in multiplex immunohistochemistry through R-IHC.[Method]1. 4-μ m-thick sections of tissue biopsied from
normal pancreas and pituitary gland were fixed in 10% buffered neutral
formalin and embedded in paraffin.2. The slides were categorized into three
subgroups in terms of the number of staining procedures to undergo (i.e. (I)
single-staining, (II) double-staining, and (III) triple-staining). The antibodies
used for each sub-group are as follows:I. anti-somatostatin for pancreas, and
anti-human growth hormone (hGH) for pituitary gland.II. anti-somatostatin
and anti-insulinin for pancreas, and anti-hGH and anti-adrenocorticotropin
(anti-ACTH) for pituitary gland.III. anti-somatostatin, anti-insulin and antiglucagon for pancreas, and anti-hGH, anti-ACTH, and anti-prolactin for pituitary gland.3. The staining process is 5 minutes for each section. The sections
in the double-staining and triple-staining sub-groups were re-stained once
and twice, respectively, with the respective antibodies mentioned above.[results]This results show the shortened duration of time in the R-IHC method;
60 minutes for double staining and 80 minutes for triple staining.
Special Lecture
Yanagita Emmy , Endo Akikazu, Yamada Hiroshi, Tsukamoto Ryuko, Itoh Tomoo
Kobe University Hospital, Japan
Invited Lecture
Yanagita Emmy, Endo Akikazu, Yamada Hirishi, Itoh Tomoo
Kobe University Hospital, Japan
Keynote Speech
PE-36
Sadayuki Hiroi1, Susumu Tominaga2, Tatsuya Yamazaki3, Tomoko Yokoo1, Mari Takashima1,
Satoru Nakano1, Tetsuro Seita1, Ayumi Sasaki1
Miho Yoshida , Arisa Kan, Naoko Yaumura, Hiroki Fujisawa, Hiroshi Oonishi, Hideki Nakano
NHO Kure Medical Center Chugoku Cancer Center, Japan
Poster Presentation
119
Oral Presentation
Background : Desmoplastic malignant mesothelioma (DMM) is a rare neoplasm that is proposed as a subtype of malignant pleural mesothelioma. It
is difficult to distinguish DMM from reactive pleural fibrosis. Histologically,
DMM has abundant collagenous tissue, forming sarcomatous, storiform or
patternless pattern.
Aim: We examined the cytopathological and immunohistochemical features of DMM.
Case : 77-years-old man admitted to a hospital for chest pain and dyspenia. He had a 5-year history of occupational asbestoes exposure. He had
worked for a roof industrial company between his ages 35 to 40. In the
detail examination, CT (computed tomography scan) showed a pleural
thickening, and a pleural effusion. In PET-CT, focally strong uptake was
recognized. In addition to these findings, thoracoscopic lung biopsy was
performed and was suggesting of DMM. The patient had chemotherapy,
but unfortunately he died about 4 months after admission due to respiratory failure and cardiac depression by tumor invasion.
Cytological findings: In pleural effusion cytology, papanicolau stain
showed large tumor cells with high N/C ratio and ireggular nuclei, and
showed some nucleoli. In imprint cytology, many spindle and polygonal
tumor cells with abundant cytoplasm were found. The nuclei were atypical
and with large nucleoli. Immunocytochemistry of this case, Tumor cells
showed positive reaction of calretinin, D2-40 and cytokeratin19.
Histopathological findings: Histologically, the tumor was composed of
spindle and polygonal cell prolifelation in dense areas of collagenous
tissue. And high density of the tumor cells at part of tumor tissues. Immunohistochemistry of tissue specimens, tumor cells were positive for
calretinin, D2-40, WT-1 and cytokeratin 19. These results were the same
as cytology specimens.
Conclusion: DMM usually shows cytologic atypia but with a poor cellular
component. So it may not be difficult to diagnose 'malignancy', but in case
of poor cell.
Case Conference
1 Department
of Clinical Laboratory Sciences, Nitobebunka college, Japan
2 Department
of pathology and Laboratory Medicine, National Defense Medical College
3 Department
of Human Pathology, Graduate School of Medicine Gunma University
[Background]An auto slide preparation system (AS-400M; DAINIPPON
SEIKI CO., LTD.) was introduced to our center in September 2013. The AS400M had a specimen manufacture rate of 68.5% from introduction until
April 2014. [Objects]We processed 21,323 paraffin blocks (alimentary
canal, uterus, prostate, placenta, lymph nodes, etc.) of materials surgically
resected from May to December 2014 and May 2015 to March 2016.
Excessive calcification and very small specimens were handled manually.
[Methods]We investigated causes of poor specimens for automatic slice
conditions, and reviewed preliminary procedures. Poor specimens were
confirmed by pathologists. ＜ 1 ＞ The main causes for poor specimens
were surgical lint and calcification. Adjusting slice speed and steam
humidification time was done for four kinds of tissue by automatically
setting the slice condition as: "alimentary canal tissue," "hard tissue," "considerable blood tissue," and "small tissue." ＜ 2 ＞ Additionally, to reduce
defective slides due to a cracked specimen, operators were coached to: (1)
decalcify the surface using KCX (1:1) for two to three hours, (2) optimize
the slide sequence, (3) shave off paraffin around the block, and (4) position
the slides correctly. [Results] 1. The specimen manufacture rate improved
to 76.8% (7,423/9,667 pieces) from May to December 2014.2. After investigation and changes in procedures for the above-mentioned device, the
specimen manufacture rate from May in 2015 to March in 2016 improved
to 94.0% (10,951/11,656 pieces), and the re-slice rate decreased.[Conclusions]We investigated use of an auto slide preparation system, procedures,
and various metrics. After implementing several changes, the efficiency
of tissue sections by the automatic slice apparatus improved, which also
facilitated improved utilization for pathology.
Keynote Speech
PE-41
Tumor complexity index is significantly associated with
metastasis in patients diagnosed with colon carcinoma
PE-42
Cytotoxic activity of crude extract from selected Philippine seaweeds against human lung adenocarcinoma cell line
Cytotoxic activity of crude extract from Sargassum polycystum and selected Philippine seaweeds against A549
human lung adenocarcinoma cell line
Victoria Hahn-Strömberg
Krisel R Sandoval1,2, Oliver Shane R Dumaoal1,2, Anacleta P Valdez1,2
Department of Medical Cellbiology Uppsala university, Sweden
1 Graduate
School, Lyceum of the Philippines University - Batangas, Philippines
2 College
of Allied Medical Professions, Lyceum of the Philippines University - Batangas
Invited Lecture
Special Lecture
Educational Lecture
Background and Aim: Growth pattern of the tumor has been studied for
its association with survival in colorectal cancer. Among different growth
pattern evaluating techniques, very little is known about prognostic significance of complexity index. Aim of this study was to develop a prognostic
model, which could be used to predict survival as well as tumor metastasis
in the patients diagnosed with colon carcinoma.
Material and methods: Formalin fixed paraffin embedded tissues samples
from 316 patients' who had undergone surgery after being diagnosed with
colon carcinoma, were used to prepare immunohistochemical slides. Slides
were stained for cytokeratin-8 and images were captured at invasive front
of the tumor. Images were threshold to get tumor area black and tumor
outline as a single pixel line to get fractal dimension and number of cells.
These two features were then used to calculate the complexity index by
performing tree diagram analysis. Complexity index was correlated with
5-years survival and other clinicopathological data of the patients.
Results: Five years survival of the patients was not influenced by complexity index (P ＞ 0.05) but the clinicopathological parameters like tumor metastasis, localization, gender and differentiation were significantly associated with complexity index with p=0.000, p=0.002, p=0.024 and p=0.000
respectively. A positive trend was also observed between complexity index
of tumor and age variable (p=0.051).
Conclusion: We conclude that complexity index is associated with systemic metastasis and differentiation of tumor but is not a predictive biomarker of survival in patients diagnosed with colon carcinoma. However,
complexity index is a reliable technique to analyse tumor characteristics
and further investigations with follow-up periods are required to reveal
the potential targets for therapeutic intervention.
Key words: prognosis, immunohistochemical slides, biomarker.
Symposium
PF-01
Cancer is one of the most dreaded diseases worldwide with limited treatment and management strategies often accompanied by serious side effects. Previous studies on marine products particularly seaweeds pose a
significant avenue for alternative cancer treatment. This study determined
the cytotoxic activity of selected Philippine seaweeds namely Caulerpa
lentillifera (CLDE), Eucheuma denticulatum (EDDE), Kappaphycus alvarezii (KADE) and Sargassum polycystum (SPDE). Dichloromethane crude
seaweed extracts were tested against A549 human lung adenocarcinoma
cell line using 3-(4,5-dimethylethylthiazol-2-yl)-2,5-diphenyltetrazolium
bromide (MTT) assay. Results indicate that SPDE exerts the highest cytotoxic activity against the cancer cell line with IC50 of 6.00 + 0.19 ug/mL
as compared to other seaweeds tested (CLDE = 49.39 + 0.61 ug/mL; EDDE
= ＞ 50 ug/mL; KADE = 45.44 + 4.51 ug/mL). Phytosterol is the common
phytochemical among the seaweed extracts tested using standard phytochemical analysis and Fourier transform infrared spectroscopy (FTIR).
SPDE shows potential for the treatment of lung adenocarcinoma and warrants further studies for the isolation of its bioactive compounds.
Fixation of serous effusions.
– or not.
PF-02
A case of primary prostate neuroendocrine tumors appeared
in the urine
case report
Yosuke Tajika1,2, Yuuki Nakajima1, Megumi Orita1, Yukie Asakura1, Mikie Takahasi1, Youko Sakai3,
Taizou Kazama4, Johji Imura2
Lisbeth Gregersen, Susanne Nielsen
Department of Surgical Pathology, Zealand University Hospital, Denmark
Case Conference
1 Anatomic
and Clinical Pathology, Social Welfare Organaization Saiseikai Imperial Gift Foundation, Inc. Saiseikai Toyama
Hospital, Japan
2 Diagnostic
Pathology, Graduate School of Medicine and Pharmaceutical Sciences University of Toyama
3 Clinical
Laboratory, Kamiichi General Hospital
4 Urology,
Social Welfare Organaization Saiseikai Imperial Gift Foundation, Inc. Saiseikai Toyama Hospital
Oral Presentation
Poster Presentation
Background:
Three pathology departments are to be united in a few years. Currently, serous
effusions are received non-fixated in one department and fixated in two of the
departments.
Purpose:
To find the optimal morphology of tumor cells in order to use either fixated or
non-fixated specimens. The unification of fixation would be an advantage for the
clinicians, who send material to the three different departments.
Material:
10 residual non-fixated material is split in 5 different tubes each:
1. Non-fixated, prepared immediately
2. Non-fixated, prepared after 3 days (kept refrigerated)
3. Fixated in 70 % ethanol
4. Fixated in Sure Path fixative
5. Fixated in CytoRich Red fixative.
3-5 are fixated immediately and prepared in 1-3 days.
Method:
All specimens are centrifuged for 5 min. From the cell pellet, 3-5 drops are applied to a Sure Path vial for Liquid Based Cytology (LBC) and set for preparation
at the Multiprocessor. Afterwards, SurePath Papanicolau (PAP) is performed at a
Slide Prep Processor. The remaining cell pellet is either prepared by the PlasmaThrombin method (non-fixated specimens) or will be centrifuged in tubes with a
formaldehyde-ethanol mixture for a clot to embedding (fixated specimens).
The PAP stained specimens and the Hematoxylin/eosin-stained sections from the
cell pellet will be blinded, and the quality and morphological assessment (primarily chromatin structure) performed by two experienced cytotechnologists.
Preliminary results:
The background in non-fixated specimens seem bright in LBC, while the fixated
specimens has many precipitations. Morphology is similar for all. The cell pellets
from the non-fixated specimens are more compact, but the tumor cells have a
better morphology in the fixated specimens. No difference is seen between the
fresh and the 3 days old material, and the three different fixatives shows similar
morphology.
Conclusion:
Not yet possible. The project is ongoing
The origins of neuroendocrine prostate tumor are rare, the cells which
is unusual to be seen in the diagnosis appear in the urine cells of human
waste. Our primary report about neuroendocrine prostate tumor in our
hospital which have experienced a case that has already been seen in the
urine cytology. The case is pointed out earlier from urine cytology class
at another hospital; it's recognized as an non-uniform swelling of the
prostate at CT, which suspected as malignancy from a 82-year-old man.
History had been underwent, when a 72-year-old has been suspected with
gastric cancer hysterectomy and angina. For the definitive diagnosis, it
will be referred to our hospital urology, which enacted from prostate TUR
and biopsy.Inspection findings is TP, Alb and Ch-E having a low value, we
acknowledged an increasing in anemia, LDH and creatinine, with high
sensitivity,occult blood (3 +), with a leukocyte reaction (2+) urinary protein
(3+).It's have been admitted if the entire circumference of thickening hypertrophy and bladder wall of CT in prostate. Histological findings the appearance is flat, which showed the atypical cells with high N / C ratio kind
circular. Cells are relatively large, chromatin has been admitted there's
1-2 in the nucleolus by thin granular. In addition, it shows if the separated
small cells appeared in the glandular cavity-like sequence in small clumps.
Histological findings is tumor cells is slightly in ductal formation, it's been
recognized as the comedones necrosis in solid alveolar, showed a fencelike sequences while in marginal obscure. Furthermore, a slight cytoplasm
was undifferentiated cells with a class round nuclei. Immunohistochemical
PSA slightly Synaptophysin is positive, which were hardly to recognized as
the positive findings, P504S were positive.We have reported an example
of prostate nerve primary neuroendocrine tumors appeared in the urine .
120
Liquid based cytology (LBC) preparation method in Routine
Work at our Laboratory
PF-04
A cytological study of ALK-positive lung cancer
Introduction:- ＞ Since characteristic histological findings of ALK-positive
lung cancer have been reported, we investigated cytological findings.
Methods:- ＞ Using 49samples from 42patients with pulmonary adenocarcinoma in whom the ALK fusion gene was searched for and the investigation of cytology preparations was possible, cytological findings were
compared between ALK-positive and - negative groups.Results:- ＞ In the
ALK-positive group, cells were present in aggregates in impression preparations, and cells containing mucus in the cytoplasm and signet-ring cells
were observed. The nuclei were mostly small, and irregularity of the nuclear shape was mild to moderate, but a small number of large cells were
mixed. In celomic fluid preparations, many cells formed aggregates and
the aggregates were large in the ALK-positive group.Conclusion:- ＞ It is
difficult to judge the presence or absence of the ALK fusion gene based on
cytological findings alone, but when small atypic cells containing a clear
nucleolus are mainly present in addition to mucus-producing cells and
signet-ring cells in impression preparations with mixed large atypical cells,
or when large cell aggregates are present in a celomic fluid preparation, it
may be necessary to consider the possibility of ALK-positive lung cancer.
The utility of touch smear cytology of an ovarian tumor
during an operation.
PF-06
Shiho Azami1, Yuji Aoki2, Mizuki Iino 2, Asumi Sakaguchi2, Kanako Ogura2, Toshiharu Matsumoto2
Endometrial carcinoma associated with endometrial polyp.
Clinicopathological and cytological analysis.
Symposium
PF-05
Educational Lecture
Introduction:Our laboratory changed the way to make fluid specimen
preparation from the conventional method to the LBC method two years
ago. Now the fraction of gynecological specimens treated with the LBC
method has increased from 1/5 to 4/5. Cytological preparations have
been made more efficient through the introduction of the LBC method.
The preparation technique before and after a start of LBC method will be
compared.Effusions specimens:The number of preparation slides for fluid
specimens has decreased from 3 to 1 with this method and has led to saving time for cytotechnologists to observe them. To add fixative solution to
sediment can not only keep cellular morphology but also reduce a fixation
work at night at our attached satellite laboratory. This method is unsuitable for Giemsa’s stain but applicable to mucinous stain or immunocytological stain instead.Aspiration and curetting specimens:Medical doctors put
the specimen sampling brush and puncture needle into the container filled
with fixative solution. Thus combination of conventional preparation method and LBC can collect more amounts of cells.Modified slide rack:We use
a modified slide rack with which two spots of smear can be placed on one
slide. This helps to collect more amounts of cells, too. Specifically, we can
prepare endocervix and endometrium specimens on one slide. The equipment is useful for immunocytological stain in which two or more types of
antibodies are used. Conclusion:LBC method is effective to keep cellular
morphology or cellular condition. Also, the method improves efficiency at
work, and shortens time for examination with an optical microscope and
describing medical reports. Furthermore our full-time staff’s skill for LBC
work reaches an almost required level, so that a cytotechnologist can have
enough time for examination of prepared slides.
Special Lecture
Harumi Kamiyama , Shigeru Tsuchida, Takuya Fusegawa, Chizuko Tomioka
Gunma Prefectural Cancer center, Japan
Invited Lecture
Tsuyoshi Ikezawa, Megumi Satou, Yukiko Sutou, Rumiko Araya, Hiroaki Kikuchi
Central Medicine Inspection Laboratory Co, Ltd, Japan
Keynote Speech
PF-03
Tomomi Kato 1, Saze tomoko1, Yasuo Kamakura1, Naoki Oogane2, Eito Kozawa3, Kosei Hasegawa4,
Masanori Yasuda1
1 Dept.of
Pathol, Saitama Med Univ. Int Med Center, Japan
2 Div.
of Pathol, Ashigarakami Hosp.
3 Dept.of
Imag Diag, Saitama Med Univ. Int Med Center
4 Dept.of
Gyne Oncol, Saitama Med Univ. Int Med Center
121
Poster Presentation
Introduction: Endometrial (EM) polyp harbors becomes a condition where
carcinoma tends to arise, especially in elderly women. Many of these carcinomas are serous carcinoma, including serous endometrial intraepithelial
carcinoma (SEIC).Materials: The examined endometrial carcinoma specimens were taken from the 21 patients who underwent total hysterectomy
with/without adnexectomy and lymph node dissection. The majority of endometrial carcinoma is found in EM in these cases. The 3 patients have a
history of breast cancer with tamoxyphen administration.Results: (1) Clinicopathological features: [Age] postmenopausal, range 49-78, mean 62;
[Positive ratio of EM cytology] 95% (20/21); Positive ratio of EM biopsy,
58% (11/19); [EM polyp size] range 8 to 50 mm , mean 19 mm; [Histological type] serous for 15 cases (including 6 cases with SEIC alone), endometrioid for 5 cases, clear cell for 1 case; [Location] limited to EMP for 15
cases, both EMP and endometrium for 6 cases; Stage (FIGO classification),
IA for 20 cases, II for ---, III for ---, IVA for cases.(2) EM cytological findings:
clear background with little or no necrosis; accompanied by atrophic endometrial glands; micropapillary pattern, tubular structure or flat sheet-like
arrangement; with/without variable metaplastic changes.Conclusion: EM
cytology is more helpful in detection of the carcinomas at early stage, i.e.,
limited to EM and/or endometrium in the background than biopsy since
the carcinoma cells are able to be easily designated as malignancy on the
comparison with small-sized atrophic benign endometrial cells. In spite of
lack of the deep myometrial invasion, these EM carcinomas may be developing extra-uterine extent, which is detected by peritoneal cytology.
Oral Presentation
Introduction:A frozen section diagnosis during an operation is important
to make a decision of an operative method in the case of ovarian tumor.
However, there are many hospitals which are not able to perform frozen
section diagnosis because of human factors and/or poor facilities. So we
report the utility of touch smear cytology during an operation, which is
more popular and technically simple, compared with that of a frozen section diagnosis.Methods:We studied 40 cases of ovarian tumors (malignant;
23 cases, borderline; 7 cases and benign; 10 cases) in which permanent
section diagnoses had been conducted in our hospital for three years. The
samples of touch smear cytology were performed from several parts of a
tumor which were macroscopically different. We investigated them blindly
in clinical information, and the results were compared with those of the
frozen and permanent section diagnoses.Results:All cases of benign and
malignant tumors could be diagnosed by touch smear cytology. However,
it was impossible to make diagnoses of all 7 cases of borderline tumor. As
compared to frozen section diagnoses, we could more reproducibly observe the histological subtypes in the 4 cases of malignant tumor by touch
smear cytology.Conclusion:We could recognize the benign and malignant
tumors accurately by touch smear cytology. On the other hand, it was difficult to make correct diagnoses in the cases of borderline tumors. So, gross
findings and close communication with clinicians can help improving the
diagnostic accuracy.
Case Conference
1 Department
of Diagnostic Pathology, Juntendo University Nerima Hospital, Japan
2 Department
of Diagnostic Pathology, Juntendo University Nerima
Keynote Speech
PF-07
The basic examination of washing fluid, washing cytology of
urinary tract
PF-08
Study on the effectiveness of bedside diagnosis and sample
production in EUS-FNA
Yuji Aoki 1, Shiho Azami2, Mizuki Iino2, Asumi Sakaguchi2, Kanako Ogura2, Toshiharu Matsumoto2
Misao Yoneda1, Kazuki Kanayama1, Chise Matsuda2, Koji Yamamoto3, Taizo Shiraishi2
1 Department
of Diagnostic Pathology, Juntendo University Nerima Hospital, Japan
2 Department
of Diagnostic Pathology, Juntendo University Nerima
1 Suzuka University of Medical Science, Japan
2 Mie
University Graduate School of Medicine
3 Saiseikai
Matsusaka General Hospital
Invited Lecture
Special Lecture
Educational Lecture
Introduction:The washing cytology from urinary tract is useful method for
diagnosis of tumor localization and spread.However, the cell morphology
was degenerated because of washing by physiological saline.Therefore it is
difficult to observe the cell morphology, and there are a lot of cases to rack
its brains about differentiation of benign or malignancy for microscopic
examination.We report that performed the examination about washing fluid. in washing cytology from urinary tract.Materials and Methods:With the
cell obtained from operation extraction materials that urithelial carcinoma
of urinary tract and the lymph node, it was washed by physiological saline
and infusion preparation such as an acetic acid Ringer's solution, a lactic
acid Ringer's solution and manufactured a cytodiagnosis specimen from a
suspension.I observed a cytodiagnosis specimen and weighed the cell morphology such as nuclear finding, cytoplasm finding, the nuclear area.Result
and Conclusion:With the physiological saline, nuclear chromatin structure
became indistinct, and the nuclear area was large.The cell degeneration
was remarkable, and the cell morphological observation was difficult.In
the acetic acid Ringer's solution, nuclear chromatin structure was clear,
and there was little degeneration, and the cell morphological observation
was easy.With a lactic acid Ringer's solution, the nuclear chromatin structure was a letter of concentration, and the nuclear area was small. The cell
degeneration was remarkable, and the cell morphological observation was
difficult.Infusion preparation is preparation of the electrolyte compositions
like a cell external solution, and there is little influence on living body, too.
The lactic acid Ringer's solution has little cell degeneration and is washings superior to physiological saline and other infusion preparation.It
leads to the preparation of a good cytodiagnosis specimen by applying a
lactic acid Ringer's solution to urinary tract washing fluid and thinks that,
besides, I contribute to improvement of the diagnosis precision.
Symposium
PF-09
Endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) was applied to the diagnosis for confirming a duodenal or pancreatic tumor, and
it now takes a central role in this field. In this study, we discussed the cell
forms of a pancreatic tumor and the sample production methods for DiffQuik stained samples and immunohistochemically stained samples. We
studied 10 cases of pancreatic duct cancer, 4 cases of pancreatic endocrine tumor, 1 case of acinar cell cancer, 2 cases of STPT, and 1 case of
anaplastic pancreatic cancer for which EUS-FNA was conducted in a period
from January 2013 to December 2015. Glandular cavity formation was
detected in 5 cases of pancreatic duct cancer, 4 cases of mucus production,
4 cases of poor background, and 1 case of the formation of multiple nuclei. For anaplastic pancreatic cancer, all of them were observed. Salt-andpepper-like chromatin, acinar structure, and spindle cells were found in all
cases of pancreatic endocrine tumor, STPT, and acinar cell cancer. The vessel axis was detected in the cases of pancreatic endocrine tumor and STPT.
If a patient is suspected of having a disease other than pancreatic duct
cancer as a result of the bedside Diff-Quik staining, it will be necessary to
conduct immunohistochemical staining to identify the disease. Therefore,
it was considered necessary to produce LBC and cell blocks. The bedside
Diff-Quik staining diagnosis by cytotechnologists in EUS-FNA is expected
to enable sample production suited for each disease and reduce the number of times of puncture and the burden on patients.
A case of Mammary Analogue Secretory Carcinoma which
Diagnosis was Supported by Cell Block Preparation.
PF-10
Yumeko Matsunaga 1, Nobuhisa Yajima1, Makoto Abe2, Takeshi Aida1, Etsuko Okusawa1,
Yasuhumi Sudo1, Hitomi Itakura1, Hidetomo Takahata1
Diagnosis of metastatic pancreatic leiomyosarcoma by EUSFNA; a case report
Kazuki Kanayama1, Chise Matusda2, Misao Yoneda1, Taizo Shiraishi2
Case Conference
1 Department
of Clinical nutrition, Suzuka University of Medical Science, Japan
2 Department
of Oncologic Pathology, Mie University Graduate School of Medicine
1 Hachinohe
City Hospital, Japan
2 Hirosaki
University, Hachinohe City Hospital
Oral Presentation
Poster Presentation
Primary and metastatic malignant mesenchymal tumors of the pancreas
are rare, and a few reports about the metastatic pancreatic leiomyosarcoma diagnosed by EUS-FNA have been published. Herein, we present a
case of metastatic pancreatic leiomyosarcoma diagnosed by EUS-FNA.
A 70-year-old woman was admitted to our hospital for evaluation of the
pulmonary multiple nodular lesions. Computed tomography (CT) -guided
needle biopsy of the pulmonary tumoral nodules were performed, and
diagnosed with the leiomyosarcoma. The patient received segmentectomy
and radiofrequency ablation (RFA). During follow up, the lesion of pancreas tail was detected by CT. EUS-FNA was performed. A cytotechnologist
was present in the endoscopy room. The sample preparation was carried
out a cytotechnologist. Cytologic specimens showed clustered or scattered
spindle cells. Nuclei were large and irregular shaped. Their chromatin pattern was fine granular, and nucleoli were small conspicuous. Histologic
specimens were prepared from tiny pieces simultaneously obtained at
EUS-FNA, which revealed malignant spindle cell tumor. Immunohistology
showed positivity for a-SMA and desmin. MIB-1 index was indicated 20%.
The lesion was diagnosed as metastatic leiomyosarcoma. EUS-FNA allowed
the differential diagnosis with other mesenchymal tumors, such as gastrointestinal stromal tumor, schwannoma. EUS-FNA suggests that it is useful
for accurate diagnosis of mesenchymal tumor. In addition, adequate material can be obtained via EUS-FNA to allow for processing into smears and
cell block, including immunohistochemical stain. The sample preparation
at bedside by cytotechnologist might contribute to the appropriate specimen processing and improvement of diagnostic yield.
Background: Mammary analogue secretory carcinoma (MASC) is a rare
salivary gland neoplasm which possess unique fusion gene ETV6-NTRK3.
Histology of the tumor shares features with the secretory carcinoma of
the breast. Here, we report a case of MASC of the minor salivary gland of
which the diagnosis was supported by cell block preparation.Case Report:
A 76-years-old woman who had a history of invasive carcinoma of the
right breast noticed a mass on the left upper lip. Fine needle aspiration cytology was performed. The specimen was viscous liquid. Conventional cytological smear and cell block were prepared. HE stain from the cell block
showed papillary and microcystic tumor cell clusters, and signet ring cell
like cells. These findings were common to smear preparation. Immunohistochemistry of the cell block revealed diffuse positive staining of S-100
protein and mammaglobin. From these results, possibility of metastatic
breast cancer as well as MASC were suspected. The patient underwent
surgical resection of the tumor. Histology revealed infiltrating tumor cells
with identical morphological and immunohistochemical features with the
cell block. Diagnosis of MASC was finally established by RT-PCR from FFPE
tissue, which detected characteristic ETV6-NTRK3 fusion gene.Conclusion:
Cell block preparation could strongly support the diagnosis of salivary
gland tumor including MASC by allowing multiple immunohistochemistry.
122
The breast pleomorphic lobular carcinoma with eosinophilic
cytoplasm: A case report
PF-12
Small Cell Carcinoma Combined with Urothelial Carcinoma
and Adenocarcinoma of the Urinary Bladder
Background: Small cell carcinoma of the bladder (SCCB) is very rare and
occupies less than 0.7% of all cancers arising from the urinary bladder.
SCCB also has been reported admixing with other types of carcinomas.
Case: A 73-year-old man was admitted to the Okayama Saiseikai General
Hospital with asymptomatic gross hematuria. An urethrocystoscopy revealed a broad-based papillary mass, 3 cm in diameter on the right wall
of bladder.The cytology of voided urine specimen showed cell debris,
pleomorphic with coarsely granular chromatin, and thick chromophilic
light-green cytoplasm, characteristic for UC. However, the histology of the
specimen by trans-urethral resection showed high grade UC, together with
small cell carcinoma, and also adenocarcinoma. Immunohistochemical
study of the resected material showed that small cells were CD56 (+), synaptophysin (+), chromogranin A (partially +), and Ki-67 labeling index was
over 80 %. In retrospect, the cytology of the urine specimen revealed the
presence of atypical small cells. Adenocarcinoma could not be identified in
cytology, possibly because of the morphological resemblance to UC cells.
The pathogenesis of such combined carcinoma is probably due to divergent differentiation of immature tumor cells.Conclusions: A case of SCCB
combined with UC and adenocarcinoma in the urinary bladder is reported
in this paper. In cytology smears of voided urine, the presence of atypical
small cells with other types of tumor requires careful consideration, because the incidence of combined SCCB with UC is higher than pure SCCB.
Usefulness of intraoperative rapid immunocytochemistry
A case of pineal germinoma diagnosed by intraoperative
histology and cytology
PF-14
Yumi Yanagida 1, Masaru Hosone1, Satoru Arai1, Hironori Katayama1, Zenya Naito2
A Study on Pregnancy and Miscarriage Rate According to
Morphology of Transferred Blastcysts in SBT
Symposium
PF-13
Educational Lecture
Background:Pleomorphic lobular carcinoma of a breast has a more poor
prognosis than classical invasive lobular carcinoma. Therefore, early
diagnosis is required. We report the case with difficulty for cytological
diagnosis of pleomorphic lobular carcinoma, because of eosinophilic cytoplasm like apocrine metaplasia on fine needle aspiration biopsy of the
breast.Case:A 50-year-old woman noticed the mass on her right breast .
She consulted the hospital nearby and she had been under the follow-up
until she stopped admitting to the hospital. 3 years later, she found that
the mass was enlarged, so she consulted another doctor. A core needle
biopsy was performed to detect invasive lobular carcinoma. Incidentally,
another mass was found on her left breast during the examination of the
right breast tumor. Then, she consulted our hospital for a treatment. To
confirm diagnosis, she underwent fine needle aspiration biopsy and core
needle biopsy of her left breast.Cytological findings:The tumor cells were
round shaped with eosinophilic cytoplasm like apocrine metaplasia on fine
needle aspiration biopsy. Because of the mild cellular atypia and apocrine
differentiation, benign tumors were suspected for the diagnosis. However,
it was difficult to completely deny apocrine carcinoma or other malignant
tumors with apocrine metaplasia because they had prominent nucleoli
and a loose cohesive pattern.Pathological findings:The tumor cells with
eosinophilic cytoplasm like apocrine metaplasia proliferated intraductally
on core needle biopsy. The tumor was diagnosed as pleomorphic type of
non-invasive lobular carcinoma for these reasons: 1) the tumor cells did
not show immunoreactivity for E-cadherin, 2) it has cytological atypia.
Conclusion:We report a detail of features of pleomorphic lobular carcinoma with eosinophilic cytoplasm including the differentiation points from
other tumors with apocrine metaplasia.
Special Lecture
Hiroki Yamamoto , Akiko Kawata, Tetuya Simizu, Masae Yabuki, Soichiro Nose, Kazuo Hamaya
Okayama Saiseikai General Hospital, Japan
Invited Lecture
Mizuki Iino , Yuji Aoki, Shiho Azami, Asumi Sakaguchi, Kanako Ogura, Toshiharu Matsumoto
Department of Diagnostic Pathology, Juntendo University Nerima Hospital, Japan
Keynote Speech
PF-11
Ayaka Murata 1, Ai Nakagawa1, Takaaki Suzuki1, Koji Yamamoto2, Hiroshi Nakano2, Shigeto Takeuchi1,
Ken Sugaya1
1 ART
Center, Saiseikai Matsusaka General Hosp., Japan
2 Dept.
of Clinical Laboratory, Saiseikai Matsusaka General Hosp, Japan
123
Poster Presentation
We examined the pregnancy rate(PR) and the miscarriage rate(MCR) in
single frozen blastocyst transfer according to the morphology of the transferred blastocysts.Blastocysts were graded before transfer according to
Gardner’ s grading criteria; developmental stage of the blastocysts(Grade16),grade of inner cell mass(ICM) and trophectderm cell(TE)(A,B,C).With
regard to developmental stage of the blastocysts(Grade3-6), the PR were
35.4% for Grade3, 50.0% for Grade4, 56.3% for Grade5 and 69.0% for
Grade6. The MCR were 20.6% for Grade3, 20.6% for Grade4, 20.0% for
Grade5 and 35.0% for Grade6. The PR was significantly higher in Grade
4-6 than in Grade3 (P< 0.05). For the MCR, there were no significant differences among each developmental stage. With regard to ICM grade of the
blastocysts, the PR were 66.0% for ICM grade A, 49.7% for ICM grade B
and 44.6% for ICM grade C. The MCR were 11.4% for ICM grade A, 24.7%
for ICM grade B and 16.2% for ICM grade C. The PR was significantly
higher in ICM grade A than in grade B and C (P< 0.05). For the MCR, there
were no significant differences among each ICM grade. With regard to TE
grade of the blastocysts, the PR were 69.8% for TE grade A, 54.8% for TE
grade B and 35.1% for TE grade C. The MCR were 10.0% for TE grade A,
24.8% for TE grade B and 17.0% for TE grade C. The PR was significantly
higher in TE grade A and B than in grade C (P< 0.05). For the MCR, there
were no significant differences among each TE grade.The results show
that the PR are closely related to developmental stage, grade of ICM and
TE.And, the results suggest that morphology of the transferred blastocysts
bear no relation to pregnancy prognosis.
Oral Presentation
Introduction Intraoperative rapid diagnosis of various brain tumors has
become a routine practice in pathology departments and often proves to
be challenging when only a frozen histological specimen is available. On
the contrary, cytology specimens are basically free from freezing artifacts,
enabling detailed observation of individual cells.We herein report a case of
pineal gland germinoma diagnosed by intraoperative combined analysis
on histology and cytology with rapid immunocytochemistry and introduce
briefly our intraoperative immunocytochemical staining system. CaseAn
18-year-old Japanese male visited a local hospital with chief complaints of
headache and anorexia. Brain CT revealed a pineal tumor with a diameter
of 4 cm causing severe obstructive hydrocephalus when he was referred
to our hospital. Intraoperative histo-cytological findings A rapid H-E
histology specimen of frozen section revealed a sheet-like proliferation of
atypical polygonal cells with mature small lymphocytes forming a “twocell” pattern. Intraoperative cytology exhibited clusters of ovoid cells with
prominent nucleoli in the background of mature small lymphocytes. Intraoperative rapid immunocytochemistry successfully demonstrated LCA(-)
and PLAP(+). Based on these characteristic morphology and a list of differential diagnoses enumerated in our pre-operative clinico-pathological conference with the neurosurgeons, our intraoperative diagnosis was pineal
germinoma. Postoperative permanent pathological diagnosis In addition
to above-mentioned histo-cytological findings, epithelioid cell granulomas
were also observed in the vicinity of the tumor cells. With a diagnostic
finding of Oct-4 positivity in formalin-fixed permanent section, our final diagnosis was pineal germinoma. Conclusions We have presented a case of
pineal germinoma diagnosed by both histology and cytology using intraoperative rapid immunocytochemistry. Immunocytochemistry has usually
a higher sensitivity than that of immunohistochemistry and often provides
valuable information required for a correct diagnosis. Intraoperative combined analysis on histology and cytology with rapid immunocytochemistry, therefore, is regarded as an effective and practical tool for a routine
intraoperative rapid diagnosis.
Case Conference
1 Nippon
Medical School- Tama Nagayama Hospital, Japan
2 Nippon
Medical School
Keynote Speech
PF-15
Sperm cryopreservation for patient with malignant or nonmalignant diseases in ART center
PG-01
Takaaki Suzuki 1, Ai Nakagawa1, Ayaka Murata1, Koji Yamamoto2, Hiroshi Nakano2, Shigeto Takeuchi1,
Ken Sugaya1
Tuan-Jen Wang1, Chih Kuang Chuang2, Sung Fa Huang1, Tzu Lin Chen1, Chi-Kuan Chen1
1 Laboratory
Medicine, MacKay Memorial Hospital , Taiwan
2 Medical
Research, MacKay Memorial Hospital
1 ART
Center, Saiseikai Matsusaka General Hosp., Japan
2 Dept.
of Clinical Laboratory, Saiseikai Matsusaka General Hosp., Japan
Invited Lecture
Special Lecture
Educational Lecture
Cerebrovascular and cardiovascular disease are the second and the forth
most common causes of death in Taiwan, and both result in serious
health injure and high mortality. The principle etiology of above diseases
is arteriosclerosis which is caused by prolonged and slowly progressive
inflammation on the vascular epithelium cells. Arachidonic acid (AA), Eicosapentaenoic acid (EPA) and their ratio in plasma are thought to be an
accurate indication of the level of inflammation occurring within the body.
In this study, a unique tandem mass spectrometry method that measures
the ratio of Arachidonic acid (AA) to Eicosapentaenoic acid (EPA) in serum
is proposed.
Blood samples were collected from 50 normal adults after 12-hour fasting, and from 40 patients with increased C-Reactive Protein (CRP; normal
reference range: <0.8mg/dl) of suffering atherosclerotic event. Serum samples were pretreated by organic solvent and hexane extraction, and the
extract was ready for LC/MS/MS analysis. Derivative was not necessary in
this method. An AB 4000 Q TRAP LC-MS/MS system with multiple reaction monitoring (MRM) mode was applied.
The within-run and between-run precisions (CV %) and the linearity of AA
and EPA based on the IS were good (both less than 13.6%). The recovery
of AA and EPA by using LC-MS/MS method (n=6) was 78.9% and 66.8% in
average, respectively. The mean AA and EPA was 6.19 ± 2.31 and 2.77
± 1.25) µg/mL in normal control (n=50) and 4.31 ( ± 2.80), and 0.25
( ± 0.22) µg/mL in CRP increased patients (n=40), respectively. The average AA/EPA ratio was 2.21 in normal control and 16.6 in patients with
increased CRP.
The AA/EPA ratio is significantly elevated 7.5-fold in patients with high
CRP value than that in normal control (p value <0.001). Our results show
that the AA and EPA quantitative analyses may provide valuable information for the monitoring chronic inflammation, such as arteriosclerosis.
We reviewed cases of sperm cryopreservation in malignant and nonmalignant diseases. 76 cases from 2000 January to 2015 June were
included in this study. The clinical recodes were reviewed retrospectively.
The age at cryopreservation, marriage statue, original diseases, the timing
of cryopreservation, semen quality, and result were analyzed.The age at
cryopreservation ranged from 13-64 years old (median 29). 17 patients
were married, 59 were single at the time of cryopreservation. The original
diseases were leukemia in 45 patients, testicular cancers in 16 patients,
other diseases in 15 patients. 66 patients froze, but 10 patients couldn't
freeze for azoospermia. The average of total sperm number after chemotherapy was 21.8 x 106 , and the average of total sperm number before
treatment was 123.6 x 106. The patients after chemotherapy decreased
the average of total sperm number significantly more than the patients
before treatment (p<0.05).7 patients used frozen/thawed sperm in ART, 5
among them achieved pregnancies and 4 among them were successful.A
treatment results of malignant diseases improves by progress of medical
technology, and sperm cryopreservation is advanced for QOL after treatment. After 2000, sperm cryopreservation to a patient with a possibility of
the decline of the spermatogenic function and the disappearance is being
conducted in ART center aggressively. The number of sperm decreases by
a patient after chemotherapy and the patient who becomes an azoospermia exists. It is important to freeze a spermatozoon before chemotherapy for
fertility preservation.
Symposium
PG-02
Establishment of the LC-MS/MS quantification method for serum free arachidonic and
eicosapentaenoic acid
Developmental and clinical practice of the tandem mass analytical method for free fatty acid
Pre-operative language mapping with MEG in patients with
temporal lobe epilepsy
PG-03
Effect of hypoxic training on renal function
A cross-over study in healthy subjects
Case Conference
Makoto Ishida1,2, Masaki Iwasaki3, Akitake Kanno4, Kazutaka Jin2, Suguru Asagi1, Takashi Miki1,
Ryuta Kawashima4, Nobukazu Nakasato1,2
Tsuneo Watanabe 1, Juri Nakayama1, Hazuki Ohashi1, Koichi Shinoda1, Yuzuru Nohisa1,
Nobuyuki Furuta1, Toshio Matsuoka2, Mitsuru Seishima1
1 Clinical
Physiology Center, Tohoku University Hospital, Japan
2 Department
of Epileptology, Tohoku University Graduate School of Medicine
3 Department
of Neurosurgery, Tohoku University Graduate School of Medicine
4 Department
of Electromagnetic Neurophysiology, Smart Aging International Research Center, Institute of Development, Aging
and Cancer, Tohoku University
1 Division
of Clinical Laboratory, Gifu University Hospital, Japan
2 Department
of Sports Medicine and Sports Science, Gifu University Graduate School of Medicine
Objective : The purpose of this study was to investigate the influence of hy-
Oral Presentation
Poster Presentation
poxic physical exercise on renal function and to compare its effects on several parameters related to renal function to those of a control group who
with training under normoxic conditions. Methods : Nine healthy men were
examined. Participants performed treadmill exercise under either normobaric hypoxic or normobaric normoxic conditions for 40 min (including a
5-min warm-up and 5-min cool-down) after a 15-min rest period. Exercise
was performed at the target heart rate (HR), which was calculated as following formula: (220 - each individual's resting HR) × 0.7 + each individual's resting HR. Training under the different environmental conditions was
performed 3 months apart to ensure a sufficient wash-out period. During
the exercise session, HR was monitored not to exceed the target HR. Both
blood and urine examinations related to renal function were determined
before and after exercise.Results : In the serum biochemical examinations,
urea nitrogen (UN), creatinine (Cr), cystatin C (cysC), sodium (Na), potassium (K), and serum osmolality were significantly higher after exercise than
before exercise in both hypoxic and normoxic groups. Meanwhile, UN,
Na and pH of the urine after exercise were significantly lower than that
before exercise. Concerning urinary sediment examinations, hyaline casts,
epithelial casts, and tubular epithelial cells were significantly higher after
exercise in both hypoxic and normoxic groups than before exercise. Furthermore, significant main effect according to the exercise conditions was
observed for urine protein [F(1,8) = 5.6, P = 0.033]. Subsequently, urine
protein after exercise was significantly higher in the hypoxic group than
that before exercise (3.8 ± 1.1 mg/dL vs. 5.4 ± 1.9 mg/dL, P = 0.038).
Conclusion : Our results suggest that hypoxic training may generate more
load on the renal function than a similar exercise intensity under normoxic
conditions.
Purpose: To investigate clinical utility of non-invasive language mapping
with magnetoencephalography (MEG).
Methods: This study included 25 right-handed patients (14 females; a
mean age of 30.1 years) with drug-resistant temporal lobe epilepsy who
underwent MEG language mapping with auditory word recognition task
as a part of pre-surgical evaluation. To determine language dominant
hemisphere, late MEG responses between 200 to 2000ms after stimulus
onset were investigated by two methods; equivalent current dipole (ECD)
modeling analysis and statistical analysis of event-related desynchronization (ERD)/ event-related synchronization (ERS). In ECD analysis, laterality
index (LI) was calculated from the number of ECDs localized on the posterior language area in each hemisphere. LI values greater than 0.5 and
less than -0.5 were considered as indicative of left and right hemispheric
dominance, respectively, and values between -0.5 and 0.5 were indicative
of bilateral activation. In ERD/ERS analysis, language dominance was determined on the presence of statistically significant changes in ERD or ERS.
The MEG result was also compared with functional magnetic resonance
imaging (fMRI) under verb generation tasks.
Results: In the ECD analysis, 11 patients (44.0%) were judged as left hemispheric dominance, 4 (16.0%) were right dominance, and 10 (40%) were
bilateral activation. ECD analysis was concordant with ERD/ERS analysis and fMRI in 90.9% and 90.0% of the patients with left hemispheric
dominance, respectively. However, the concordance was only 50% in the
patients with right hemispheric dominance. In the patients with bilateral
activation, 6 patients (60.0%) were judged as left dominance by ERD/ERS
analysis and by fMRI.
Conclusion: Language lateralization can differ between different analytical
methods, especially in patients without clear dominance to the left side. It
is necessary to elucidate different functional aspects of language detected
by MEG analysis for accurate determination of language dominance.
124
Vascular damage associated with CKD and laboratory medicine
A decline in kidney function closely is associated with
progression in atherosclerosis
PG-05
Usefulness of Virtual Touch Quantification for the diagnosis
of pancreatic solid lesions
Yusuke Nakade , Tadashi Toyama, Kengo Furuichi, Yoshiyasu Miyajima, Hiroyasu Oe, Mikio Nagahara,
Yoshio Sakai, Takashi Wada
Yusuke Kudo 1, Mutsumi Nishida1, Satomi Omotehara1, Takahito Iwai1, Taisei Mikami2, Hitoshi Shibuya1,
Kaoru Kahata1, Chikara Shimizu1
Kanazawa University Hospital, Japan
1 Division
of Laboratory and Transfusion Medicine, Hokkaido University Hospital, Japan
2 Faculty
of Health Sciences, Hokkaido University
Naohiro Ichino 1, Keisuke Osakabe1, Toru Nishikawa2, Hiroko Sugiyama2, Tadayoshi Hata1,
Naoto Kawabe3, Senju Hashimoto3, Kentaro Yoshioka3
A case of sigmoid colon cancer with tumorous embolism
Utility of sonography
Symposium
PG-07
Educational Lecture
Controlled attenuation parameter for non-invasive assessment
of hepatic steatosis in chronic hepatitis C
Special Lecture
PG-06
PURPOSEVirtual Touch Quantification (VTQ) is a novel ultrasound technique that evaluates tissue stiffness by Shear Waves Velocity (SWV) quantification. Clinical use of VTQ for the liver fibrosis has been established,
however, few studies demonstrated its usefulness for the pancreas. The
aim of this study was to assesse the diagnostic usefulness of VTQ method
in pancreatic solid lesions.SUBJECTS and METHODSSWV of pancreatic
solid lesions and pancreatic parenchyma in 30 healthy volunteers were
measured by VTQ method. The examination was performed with the Siemens Acuson S2000, using the 4C1 convex prove. SWV were measured
10 times in each of lesions and parenchyma. Median SWV values were
compared with Kruskal-Wallis test. Differences were considered significant
at P ＜ 0.05. RESULTS31 patients were included, in these, 17 had pancreatic adenocarcinoma, 10 had neuroendocrine neoplasm (NEN), 4 had
metastasis from renal cell carcinoma. Median SWV values (range) were
2.62 m/s (1.72m/s-4.52m/s), 2.42m/s (0.94m/s-4.35m/s), and 1.40m/s
(0.72m/s-2.85m/s), respectively. In the healthy volunteers group the median SWV values and range were 1.01m/s (0.71m/s-2.08m/s). Significant
difference between SWV median values of pancreatic adenocarcinoma and
metastasis, normal pancreatic parenchyma was found (P ＜ 0.001, respectively). SWV median values of pancreatic adenocarcinoma tend to be higher compared with that of NEN, they did not reach statistical significance.
CONCLUSIONVTQ method would be useful in the presence diagnosis of
pancreatic adenocarcinoma, and differential diagnosis of pancreatic solid
lesions non-invasively.
Invited Lecture
Introduction: Carotid echo indexes [intima-media thickness (IMT)] are
commonly used surrogate markers for cardiovascular disease; However,
the impacts of chronic kidney disease (CKD) on changes in IMT are unclear. We examined associations between CKD and IMT in participants
with and without type 2 diabetes through longitudinal analysis.Methods:
In total, 424 subjects were enrolled in this study. IMT was measured as per
carotid echo indexes. Relationships between IMT and risk factors were analyzed using multiple linear regression analysis, in which we de?ned IMT
as the dependent variable and atherosclerosis related factors (age, sex,
blood pressure, total cholesterol, body mass index, estimated glomerular
filtration rate (eGFR), uric acid, smoking index, number of antihypertensive
drugs, statin use, urinary protein levels, past cardiovascular event, glycated
hemoglobin, and diabetes duration) as independent variables.Results: The
study population was composed of 70.3 % male subjects. Participants with
diabetes accounted for 64.4 % of the total population. The mean followup duration was 2.2 ± 1.5 years. Mean frequency of examination during
the study period was 2.6 times. There was a negative correlation between
eGFR and changes in IMT for participants without diabetes. After adjusting for multiple risk factors, there was a tendency for increased IMT with
lower eGFR ( Β = -0.0091, p = 0.06) in all participants. In participants
without diabetes, eGFR (+10ml/min/1.73m2) ( Β = -0.022, p = 0.04) and
diastolic blood pressure ( ＞ 85mmHg) ( Β = -0.149, p = 0.01) were significantly associated with increased IMT even after adjusting for confounding
factors. In contrast, no trend was observed in participants with diabetes.
Moreover, an interaction term between eGFR and urinary protein in participants without diabetes was not significant (p = 0.792).Conclusion: Low
eGFR was associated with progression of carotid thickness independent of
common cardiovascular risk factors in non-diabetic participants.
Keynote Speech
PG-04
Kenta Muto , Tetsuya Nishiura, Emina Katsurada, Shigeki Oda, Shinji Naito
National Hospital Organization Ureshino Medical Center, Japan
125
Poster Presentation
Aim: Hepatic steatosis can be a co-factor in many chronic liver diseases
that can lead to liver fibrosis and cirrhosis. A novel non-invasive tool
based on ultrasound attenuation, called controlled attenuation parameter (CAP), was attached with FibroScan for assessment of liver steatosis
quantitatively. The aim of this study was to evaluate the performance of
CAP for assessment of hepatic steatosis in chronic hepatitis C. Methods:
In a total of 113 patients, 69 men and 44 women with chronic hepatitis
C, CAP values were measured, and liver biopsies were performed. The
measurement of CAP was done ten times on right lobe of the liver from
the right intercostal space, and the median values were adopted for CAP
values. Steatosis of liver specimen was categorized as S0: <5%; S1: 5-33%;
S2: 34-66%; or S3: ＞ =67%. The CAP values were compared with steatosis
grade and also with the ratio of hepatic steatosis area that was calculated
by digital image analysis liver specimen. Results: The CAP values of the
patients with steatosis grade of S0 (n =53), S1 (n =32), S2 (n =18) and S3
(n =10) were 193.7 ± 40.7 dB/m, 206.5 ± 27.8 dB/m, 229.4 ± 38.4 dB/
m and 256.5 ± 45.0 dB/m, respectively. The CAP values of those with S2
and S3 were significantly higher than in those with S0 (P =0.0035 and P
=0.0005). Furthermore, the CAP values of those with S3 were significantly
higher than those with S1 (P =0.0012). The relationship between the CAP
values and ratio of steatosis area was assessed by simple linear regression
analysis. There was a significant positive moderate correlation between
the CAP value and ratio of steatosis area (r =0.43, P <0.0001). Conclusion:
This study suggested that CAP is a promising tool for the non-invasive assessment and quantification of hepatic steatosis in chronic hepatitis C.
Oral Presentation
Introduction: We report that we experienced a case of sigmoid colon cancer with tumor embolus in mesenteric vein and sonographic examination
was so useful for that detection. Case: A 97-year-old woman complaining
the difficulty of body movement due to bone fracture was admitted. In further examination, abdominal CT indicated the dilatation of the pancreatic
duct of the body and tail of pancreas and the abdominal ultrasonography
showed the 30 mm in size mass lesion in sigmoid colon with low brightness, irregular shape, internal heterogenicity and pseudo-kidney sign. In
addition, it also showed low bright echoic lesion, suggestive of embolus, in
from portal to splenic vein and inferior mesenteric vein. As that embolus
was continuous with tumorous lesion of sigmoid colon and color Doppler
indicated the blood stream in it, it was thought to be tumorous embolus.
Colonscopy was performed and it was diagnosed as moderately-differentiated adenocarcinoma by bioptic examination for the tumorous lesion of
the sigmoid colon. Study: Colon cancer invades a mesenteric vein through
the vessels invasion in the intestinal wall and has metastasis to liver via
portal vein. Then, it is very rare to be observed as the identifiable tumor
embolus in a mesenteric vein without metastatic liver mass by sonography.
Conclusion: In the present case, the simple CT just indicated the dilatation
of pancreatic duct. However, the abdominal ultrasonographic examination
contributed to detect sigmoid colon cancer and its embolus in the mesenteric vein. In this way, the sonography was thought to be extremely useful
for the intravenous examination such as characteristics in embolus and
the observation of bloodstream signal.
Case Conference
1 Faculty
of Medical Technology, School of Health Sciences, Fujita Health University, Japan
2 Center
of Ultrasound Diagnosis, Fujita Health University Hosp.
3 Dept.
of Liver, Biliary Tract and Pancreas Diseases, School of Medicine, Fujita Health Univ.
Keynote Speech
PG-08
Evaluation of the cross-sectional area by ultrasound in
peripheral neuropathy
PG-09
Evaluation of peripheral neuropathy by sensory nerve action
potentials comparison.
Ako Ito1, Tsuneo Watanabe1, Megumi Yamada2, Koichi Shinoda1, Yuzuru Nohisa1, Nobuyuki Furuta1,
Hiroyasu Ito1, Mitsuru Seishima1
Masafumi Katayama 1, Yasuyuki Teramato2, Fumitomo Iwanaga2, Kohei Nishimura2,
Takuya Matsunaga2, Kaoru Matsunaga3, Ryoji Nakanishi2
1 Division
of Clinical Laboratory, Gifu University Hospital, Japan
2 Dept.
of Neurology and Geriatrics, Gifu Univ. Graduate School Medicine
1 International
university of health and welfare, Japan
2 Kumamoto
Kinoh Hospital
3 Kumamoto
Onjaku Hospital
Invited Lecture
Special Lecture
Educational Lecture
Purpose: High-resolution sonography is a novel method that provides
morphological information for peripheral nerves. The purpose of this study
was to evaluate the cross-sectional area (CSA) of the median nerve using
ultrasonography (US).Methods: Twelve hands of 6 patients with carpal
tunnel syndrome (CTS) group (mean age, 53.8 ± 11.6 years), 16 hands
of 8 patients with type 2 diabetes mellitus (DM) group (mean age, 61.1 ±
6.1 years), 10 hands of 6 patients with other peripheral neuropathy (OPN)
group (mean age, 59.3 ± 20.0 years): 1 patient with chronic inflammatory
demyelinating polyneuropathy, 1 patient with multifocal motor neuropathy,
2 patients with amyotrophic lateral sclerosis, 1 patient with hereditary motor sensory neuropathy and 1 patient with Guillain-Barre syndrome. Twelve
hands of 6 healthy volunteers (controls) (mean age, 53.3 ± 5.3 years) were
also included in the study. The CSA was measured by US at defined sites
(CT, carpal tunnel inlet; FA, midpoint of the forearm; AM, midpoint of the
arm), and one way analysis of variance (ANOVA) was carried out among
the groups.Result: There was no significant differences in age among four
groups including controls. Significant differences for groups were observed
in CT [F(3, 45) =6.1, p = 0.002] and AM [F(3, 45) =4.9, p = 0.005]. The CSA
in the CT was significantly higher in CTS group (12.2 ± 4.6 mm2) than in
controls (8.5 ± 1.8 mm2) (p ＜ 0.001). The CSA in the AM was significantly
higher in OPN group (16.1 ± 10.2 mm2) than in other three groups: vs.
controls 9.7 ± 3.4 mm2(p = 0.024), vs. CTS 8.3 ± 2.0 mm2 (p = 0.004), vs.
DM 9.9 ± 1.7 mm2( p = 0.021).Conclusion: Our study suggests that nerve
US is useful for diagnosis of peripheral neuropathy.
Symposium
PG-10
Introduction: We investigated the Sensory nerve action potential (SNAP)
recorded from different two positions and compared the data from normal
subjects and patients with peripheral neuropathy.Subjects and methods:
Twenty nine healthy volunteers, 18 cases of carpal tunnel syndrome (CTS)
and 23 cases of diabetic peripheral neuropathy (DPN) participated in the
present study. SNAP were recorded after median nerve (MN) stimulation
from the wrist, at proximal and distal position in the middle finger. We
investigated SNAP recorded from proximal position (P-SNAP) and SNAP
recorded from distal position (D-SNAP). The latency, amplitude and duration were measured at the two positions. Furthermore, the sensory nerve
conduction velocity (SCV) in wrist-finger (W-F SCV) and finger-finger (F-F
SCV) were calculated. The distance between the wrist and proximal recording electrode was 140mm. The two electrodes on the middle finger were
set 25mm apart.Results: The results of the P-SNAP and D-SNAP measurements in the normal subjects were as follows, latency: 2.35 ± 0.19ms, 2.79
± 0.21ms, amplitude: 74.59 ± 29.60uV, 56.27 ± 24.26uV, duration:
1.37 ± 0.21ms, 1.51 ± 0.28ms. W-F SCV was 61.3 ± 3.2m/s and F-F SCV
was 58.2 ± 3.6m/s. We used a distribution map to examine correlation
among the latency, amplitude, duration and SCV in P-SNAP and D-SNAP.
In the CTS cases significant increase in the rate of amplitude (amplituderatio) was observed in P-SNAP and D-SNAP. In the DPN cases the SCV-ratio
significantly decreased (p ＜ 0.01: ANOVA).Conclusions: Terminal segment
neuropathy such as in DPN cases was evaluated easily by this method. The
method may be applied to detailed diagnoses of other neuropathy cases.
Evaluation of Peripheral Sensory Perception Pre- and PostRevascularization
PG-11
Kaori Sugawara1, Mika Miki1, Takashi Miki1, Daijirou Akamatsu2, Hitoshi Goto2
Novel Echocardiographic Method to Assess Left Ventricular Chamber Stiffness and End-Diastolic
Pressure
Usefulness of Time-Velocity Integral Measurements of Pulmonary Venous and Transmitral Flows
Kazunori Okada 1, Rika Abiko2, Sanae Kaga1, Masahiro Nakabachi3, Hisao Nishino3, Shinobu Yokoyama3,
Mutsumi Nishida3, Taisei Mikami1
Case Conference
1 Clinical
Physiology Center, Tohoku University Hospital, Japan
2 Department
of Advanced Surgical Science and Technology, Tohoku University Hospital
1 Fuculty
of Health Sciences, Hokkaido University, Japan
2 Department
of Health Sciences, School of Medicine, Hokkaido University
3 Division
of Laboratory and Transfusion Medicine, Hokkaido University Hospital
Oral Presentation
Poster Presentation
Introduction: Data regarding ischemia-related sensory perception of the
fingertip are limited. We examined factors related to reduced functional
sensory perception of the lower limbs after revascularization.Methods:
This is a retrospective review of 17 patients (19 limbs; mean age, 71.0
years; range, 47 – 82 years; 15 males [78.9%]) who underwent revascularization and quantitative analysis of sensory perception before and
after revascularization from April 2015 to February 2016. Before and
after revascularization, sensory perception intensity was evaluated in the
medial forearm and halluxes using a painless electrical stimulation system, PainVisionTM (NIPRO Co., Ltd, Japan). Minimum perceived current
was defined as the minimum electrical stimulation sensed by the subject.
Normal range of peripheral sensory perception has not been established.
Based on our findings, a hallux – forearm ratio (HFR) of ＜ 1.99 was defined as normal sensory perception. Cases in which HFR decreased after
revascularization were considered to have improved sensory perception.
Nine of 19 limbs had dysesthesia; therefore, the relation between sensory
recovery and smoking status, underlying disease, and biochemistry data
was also examined. Results: Mean ( ± standard deviation) HFR was 3.07
± 2.76 before surgery and 2.97 ± 2.17 after surgery. Five of the nine
limbs showed improvement (pre-HFR, 5.66 ± 3.15 vs. post-HFR, 3.31 ±
2.46), whereas the remaining four did not (pre-HFR, 3.97 ± 3.24 vs. postHFR, 4.56 ± 3.31). All limbs with preoperative dysesthesia had improved
microvascular blood flow after surgery. There was a significant difference
in smoking status between the sensory recovery and non-recovery groups
(p = 0.0164).Conclusion: Smoking may result in lower sensory recovery
after revascularization. Smoking leads to endothelial dysfunction; capillary
arteriosclerosis reduces the material exchange function of microcirculation and may prevent improvements in peripheral sensory perception.
Regarding total foot care, the assessment of sensory function to recognize
external stimulation leads to wound prevention.
Background: Difference between the atrial systolic pulmonary venous (PV)
and transmitral flow durations have been reported to be useful to estimate
the left ventricular (LV) end-diastolic pressure (LVEDP). We aimed to examine the usefulness of our novel parameters based on the time-velocity
integral (TVI) measurements of the PV and transmitral flows for assessing
the LV chamber stiffness and LVEDP.
Methods: In consecutive 50 cardiac patients who underwent cardiac catheterization, the LVEDP and pressure increase at atrial contraction ( Δ Pa)
were measured from the LV pressure waveform. LV volume change during
atrial contraction ( Δ Va) was measured using echocardiographic biplane
method of disks and was corrected for body surface area, and the Δ Pa/
Δ Va was calculated as an index of LV chamber stiffness. Using transthoracic pulsed Doppler echocardiography, we measured the duration and
TVI of the backward PV flow during atrial contraction (DPVA and IPVA, respectively) and the ratio of IPVA to the PV flow TVI through a cardiac cycle
(FPVA). Also the duration and TVI of the atrial systolic forward transmitral
flow (DA and IA, respectively), and the ratio of the IA to the transmitral TVI
during a cardiac cycle (FA) were measured to calculate DPVA – DA, IPVA/IA and
FPVA/FA.
Results: DPVA – DA significantly but weakly correlated with Δ Pa/ Δ Va
(r=0.51) and LVEDP (r=0.56). IPVA/IA and FPVA/FA were significantly and well
correlated with Δ Pa/ Δ Va (r=0.77 and r=0.81) and LVEDP (r=0.79 for
both). The area under the ROC curve to predict LVEDP ＞ 18 mmHg was
0.87 for DPVA – DA, 0.93 for IPVA/IA and 0.96 for FPVA/FA.
Conclusion: Our novel parameters based on the TVI measurements of the
backward PV and forward transmitral flows during atrial contraction are
useful for the noninvasive assessment of LV chamber stiffness and LVEDP.
126
Changes in gender preference of female patients for repeated
transthoracic echocardiography
PG-13
Accuracy of the Estimation of Left Ventricular Relaxation and Filling Pressure by Using Echocardiography
A Comparison among Single Doppler Parameters, a Simple Algorithm, and Comprehensive Evaluation
1 Division
of Laboratory and Transfusion Medicine, Hokkaido University Hospital, Japan
2 Department
of Cardiovascular Medicine, Hokkaido University Graduate School of Medicine
Introduction:It is reasonable to presume that some female patients, especially young ones, would prefer female sonographers for transthoracic
echocardiography(TTE) because of the need to expose the chest. This
would interfere the logistics of echo labs. Because not all the patients are
familiar with what will happen during TTE exam, little is known about
how they really feel about it and especially if they change their mind for
the second time. We, thus, examined whether their preference for the
gender of sonographers would change.Methods:Since October 2013,
female patients referred to TTE underwent the following questionnaire
before the examination. ‘Would you prefer female sonographers, or do not
care? ‘ The TTE was performed according to their preference. Those who
underwent TTE twice were included in this analysis. Results:Among 891
female patient who underwent TTE between October 2013 and February
2015, four hundred(44.9%) preferred female sonographers and 68 had
two TTEs. They were 60 ± 16 years old. Twenty-seven(39.7%) patients
preferred female sonographers at the initial examination, forty(59.7%)
did not care sonographers’ gender. Actually, one(0.1%) patient preferred a
male sonographer. In their second TTE, fifty-four(79%) patients expressed
the same preference as the initial examination, however 14(21%) changed
their preference for sonographers’ sex between the initial TTE and the second. Their breakdown was that nine preferred sonographers with either of
sex for the first TTE then female ones for the second, and the rest five vise
vasa. Among the nine cases, three underwent TTE with male sonographers
and the rest six with female ones for the first TTE.Conclusion: About half
of the female patients preferred female sonographers but not all did. Majority of them did not change their preference for the second time. However, some changed their needs. Repeated questionnaire would be one of
the options for patient satisfactions.
Background: The comprehensive echocardiographic evaluation of left
ventricular (LV) diastolic function is recommended; however, its accuracy
has not yet been established.Methods: The study data cited the multicenter
study of strain/strain rate versus myocardial velocity for assessing left
ventricular relaxation and filling pressure (SMAP study). In 77 patients,
peak early- (E) and late-diastolic (A) LV inflow velocities and early-diastolic
mitral annular velocity (e') were measured using Doppler methods, and
the ratios of E to A (E/A) and E to e' (E/e') were calculated. Accuracy for
predicting invasively defined abnormal LV relaxation and elevated filling
pressure (FP) was compared among 1) e' and E/e', 2) a simple algorithm
based on E/A and E/e', and 3) the comprehensive evaluation based on all
conventional echocardiographic parameters by an expert. In a simple algorithmic approach, E/A ＜ 0.75, or 0.75<E/A ＜ 1.5 and E/e' ＞ 10, or E/A
＞ 1.5 were classified as abnormal relaxation, and 0.75<E/A ＜ 1.5 and E/
e' ＞ 10, or E/A ＞ 1.5 were classified as elevated FP.Results: The e' and E/
e' only weakly correlated with tau (r=-0.32) and LVMDP (r=0.50), respectively. In the estimation of LV relaxation, comprehensive evaluation had
the highest accuracy (accuracies of 3 methods: 28%, 60%, 74%). In the estimation of LVFP, E/e' had the highest sensitivity (sensitivities of 3 methods:
82%, 64%, 64%) and negative predict value (negative predict values of 3
methods: 96%, 93%, 94%), but low positive predict value (positive predict
values of 3 methods: 39%, 41%, 64%). Comprehensive evaluation had the
highest specificity (specificities of 3 methods: 79%, 85%, 94%) and positive
predict value, although sensitivity of comprehensive evaluation was low.
Conclusions: Comprehensive evaluation of LV diastolic function based on
many conventional echocardiographic parameters should be combined
with several quantitative echocardiographic parameters.
Factors related to residual shunt after transcatheter closure
of atrial septal defect
PG-15
Electrocardiographic changes in patients with chronic
obstructive pulmonary disease
Atsushi Ichikawa 1, Tetsuro Sugiura1, Hiroshi Ohnishi2, Hiromi Kataoka3, Katsumi Ogura4,
Akihito Yokoyama2, Yoshihisa Matsumura1
Okayama University Hospital, Japan
1 Department
of Laboratory Medicine, Kochi Medical School, Kochi University, Japan
2 Department
of Hematology and Respiratory Medicine, Kochi Medical School, Kochi University
3 Kawasaki
University of Medical Welfare
4 Clinical
Laboratory, Kochi Medical School, Kochi University
127
Poster Presentation
Background: Chronic obstructive pulmonary disease (COPD), which is
characterized by airflow limitation that is not fully reversible, is one of
the leading causes of morbidity and mortality in both industrialized and
developing countries because COPD primarily affects the lungs but also
produces significant cardiac consequences. Previous studies reported
characteristic electrocardiogram (ECG) changes in patients with COPD.
Accordingly, we elucidated the important ECG indices to detect COPD.
Methods: Association between respiratory function test and ECG indices
was retrospectively analyzed in 45 patients with COPD and 100 patients
free of COPD (controls). ECG indices including P axis, P interval, P amplitude, QRS axis, QRS interval, QRS amplitude in V1, QRS amplitude in lead I,
R amplitude in V1, R amplitude in V5, and QTc interval were automatically
measured by the ECG device. Respiratory function test indices included
forced expiratory volume in one second/forced vital capacity (FEV1/FVC)
and percent predicted value of FEV1 (%FEV1) after bronchodilator inhalation.Results: There were significant differences in 6 ECG indices (P axis,
P interval, P amplitude, QRS axis, QRS amplitude in lead I, R amplitude in
V5) between the 2 groups. To determine the important ECG variables present in patients with COPD, 5 variables were used for the multiple logistic
regression analysis. QRS amplitude in lead I emerged as a significant ECG
variable related to COPD (partial regression coefficient = -4.208, p = 0.002).
ROC curve analysis showed that the cut-off value of QRS amplitude in lead
I to detect COPD was less than 0.54 mV (sensitivity: 71%, specificity: 76%,
area under the curve: 0.78 [95% confidence interval: 0.69 – 0.86], p ＜
0.001).Conclusion: Low voltage in lead I was an independent predictor of
COPD and QRS amplitude less than 0.54 mV in lead I was an important
ECG criterion to detect COPD.
Oral Presentation
Purpose: To identify factors related to residual shunt after transcatheter
closure with the Amplatzer septal occluder. Methods: Two hundred and
fifty three patients (median 49.0 years, range from 6 to 83 years) underwent transcatheter closure of ASD with the Amplatzer septal occluder in
our institution. Follow up transthoracic echocardiography was performed
at 24 hours and 6 months after ASD closure. Influence of maximal ASD diameter and deficient rims on existence of residual shunts by color Doppler
imaging were evaluated.Results: Mean maximal ASD diameter was 18.0 ±
6.7 mm. One hundred and eighty five patients (73%) had a deficient rim
( ＜ 5mm; aortic rim deficiency=179, other rim deficiency=6). Echocardiography at 24 hours and 6 months after the procedure showed residual
shunts in 123 (49%) patients and 54 (21%) patients, respectively. Patients
with residual shunt had higher frequency of deficient rim than patients
with complete closure both at 24 hours and 6 months examination (87%
vs. 60%; p ＜ 0.01, 89% vs. 69%; p ＜ 0.01, respectively). Besides, patients
with residual shunt had larger maximal ASD diameter than patients with
complete closure both at 24 hours and 6 months (20.0 ± 5.9mm vs. 16.1
± 6.8mm; p ＜ 0.01, 20.4 ± 7.0mm vs. 17.4 ± 6.5mm; p ＜ 0.01, respectively). There was no significant difference in device to defect ratio between patients with residual shunt and complete closure both at 24 hours
and 6 months (1.21 ± 0.21 vs. 1.27 ± 0.34; p=0.10, 1.24 ± 0.29 vs. 1.24
± 0.28; p=0.94, respectively).Conclusions: Deficient surrounding rims and
larger defect diameter may be related to residual shunt after transcatheter
closure of ASD.
Case Conference
Madoka Ikeda , Hiroki Oe, Nobuhisa Watanabe, Yasufumi Kijima, Youichi Takaya, Ken Okada,
Hiroshi Ito
Symposium
PG-14
Educational Lecture
1 Clinical
Laboratory Center,Gunma University Hospital, Japan
2 Department
of Clinical Laboratory Medicine, Gunma University Graduate School of Medicine.
Special Lecture
Masahiro Nakabachi 1, Satoshi Yamada2, Taichi Hayashi2, Hiroyuki Iwano2, Hitoshi Shibuya1,
Kaoru Kahata1, Chikara Shimizu1, Hiroyuki Tsutsui2
Invited Lecture
Kenya Okada 1, Kouji Kurosawa2, Takao Kimura2, Kanako Niwa1, Takahiro Ikoma1, Keiko Morita1,
Tetuo Machoda1, Masami Murakami2
Keynote Speech
PG-12
Keynote Speech
PG-16
Value of Left Atrial Function in Patients with Aortic Stenosis
Assesment of Using Speckle-tracking Echocardiography
PG-17
Kazuto Yamaguchi 1, Hiroyuki Yoshitomi1, Eri Nitta1, Seiji Mishima1, Kazuaki Tanabe2, Atushi Nagai1
Yuichi Maruta , Masami Fujii, Hirochika Imoto, Hisaharu Gotou, Hiroyasu Koizumi, Hideyuki Ishihara,
Sadahiro Nomura, Michiyasu Suzuki
1 Shimane
University Hospital, Laboratory Medicine, Japan
2 Shimane
University Hospital, Department of Cardiology
Department of Neurosurgery, Yamaguchi University Graduate School of Medicine, Ube, Japan
Invited Lecture
Special Lecture
Educational Lecture
BackgroundThe chronically increased afterload is accompanied by several
structural and functional changes as progressive left atrial (LA) enlargement and dysfunction. In severe AS, both LA dilatation and dysfunction
have been shown to adversely affect the outcome. Assessing the relationship between LA size and function is thus of clinical importance.The aim
of the present study was to assess the LA function in patients with AS and
to evaluate its impact on the symptoms and AS progression.MethodsThe
study consisted of 25 consecutive patients (mean age 76 ± 9 years) with
moderate to severe AS. Patients were divided into 3 groups; moderate AS
(aortic valve area 1.0-1.5 cm2), severe AS (aortic valve area ＜ 1.0 cm2)
without symptoms, and severe AS with subsequent aortic valve replacement (AVR). All patients underwent comprehensive echocardiography. The
LA global longitudinal strain LA-GLS) curve was assessed in all patients. 3
aspects of LA-GLS were recorded: contractile, conduit and reservoir strain.
ResultsLAVI was increased in severe AS (without symptom; 66 ± 22 mL/
m2, AVR; 64 ± 22 mL/m2) compared with moderate AS (40 ± 14 mL/m2).
There was no significant difference in LAVI between severe AS groups.
LAEF (moderate AS; 51 ± 6%, AS without symptoms; 43 ± 10%, AVR; 31
± 13%, ANOVA=0.006) were significantly reduced in severe AS and were
correlated with AS progression. Of all indices of LA strain, LA reservoir
strain had the significant difference to identify patients with cardiac symptoms (moderate AS; 13.8 ± 2.8%, AS without symptoms;12.8 ± 3.3%, and
AVR; 8.8 ± 1.7 %, ANOVA=0.008).ConclusionsImpaired LA reservoir strain
in patients with AS relates to AS progression, independently of the increase in LA volume. Increased LA stiffness may be associated with cardiac
symptoms in patients with AS.
Symposium
PG-18
Strategies and Pitfalls of MEP Monitoring during
Supratentorial Aneurysm Surgery
Background The aim of this study was to reveal the strategies and pitfalls
of motor evoked potential (MEP) monitoring methods during supratentorial aneurysm surgery and discuss the drawbacks and advantages of
each method by reviewing our experiences. Methods Intraoperative MEP
monitoring was performed in 250 patients. Results from four monitoring
techniques using combinations of two stimulation sites and two recording
sites were analyzed retrospectively.Results MEP was recorded successfully
in 243 patients (97.2%). Direct cortical stimulation (DCS)-spinal recorded
MEP (sMEP) was used in 134 patients, DCS-muscle recorded MEP (mMEP)
in 97, transcranial electrical stimulation (TES)-mMEP in 11 and TES-sMEP
in one. TES-mMEP during closure of the skull was used in 21 patients.
DCS-mMEP was able to detect waveforms from upper and/or lower limb
muscles. Alternatively, DCS-sMEP (D-wave) could accurately estimate
amplitude changes. A novel "early warning sign" indicating ischemia was
found in 21 patients, which started with a transiently increased amplitude
of D-wave and then decreased after proximal interruption of major arteries. False-negative findings in MEP monitoring in two patients were caused
by a blood insufficiency in the lenticulostriate artery and by a TES-sMEP
recording, respectively. Conclusions The results of this study suggest that
to perform accurate MEP monitoring, DCS-mMEP or DCS-sMEP recording
should be used as the situation demands, with combined use of TES-mMEP
recording during closure of the skull. DCS-sMEP is recommended for accurate analysis of waveforms. We also propose a novel "early warning sign"
of blood insufficiency in the D-wave.
Effect of sleep stages on distribution of interictal fast ripples
in intractable focal epilepsy
PG-19
Rie Sakuraba 1, Masaki Iwasaki2, Suguru Asagi1, Takashi Miki1, Nobukazu Nakasato3
Progression of Left Ventricular Diastolic Dysfunction in
Patients with CKD.
Yoshiyasu Miyajima 1, Tadashi Toyama2, Hiroyasu Ohe1, Mikio Nagahara1, Kengo Furuichi2,
Yoshio Sakai3, Takashi Wada3
Case Conference
1 Clinical
Physiology Center, Tohoku University Hospital, Japan
2 Department
of Neurosurgery, Tohoku University Graduate School of Medicine
3 Department
of Epileptology, Tohoku University Graduate School of Medicine
1 Dept.
of Clinical Laboratory, Kanazawa Univ. Hosp., Japan
2 Div.
of Nephrology, Kanazawa Univ. Hosp.
3 Dept.
of Nephrology and Laboratory Medicine, Kanazawa Univ. Hosp.
Oral Presentation
Poster Presentation
Rationale: High-frequency oscillations (HFOs) are EEG markers of epileptogenicity. Removal of the brain region hosting high-rate interictal HFOs
is related to good seizure outcome after surgery. However, for the accurate diagnosis of epileptogenicity, pathological HFOs must be carefully
distinguished from physiological HFOs. Occurrence of HFOs is strongly
influenced by sleep stages. Recently, we reported that interictal ripples (80
– 200Hz) may provide a specific marker of epileptogenicity during REM
sleep (Sakuraba et al., 2015). In this study, we investigated that effect of
sleep stages on distribution of interictal fast ripples (200 – 500Hz, FRs)
and correlation to epileptogenic area.Methods: The subjects comprised
7 patients with drug-resistant epilepsy who underwent extraoperative
intracranial EEG monitoring and became seizure freedom after surgery.
Interictal FRs were automatically detected from different sleep stages. The
relationship of high-rate FR electrodes to the area of surgical resection
was compared between REM and NREM sleeps. Upon the result, further
analysis was performed by dividing the FR into two frequency ranges;
200 – 299Hz and 300 – 399Hz. Then, the relationship of FR occurrence
to the area of surgical resection was compared between REM and NREM
sleeps for each frequency range.Results: High-rate FR were identified in
20 (15.9%) and 4(1.9%) electrodes inside and outside the resection during
NREM sleep, respectively, and in 12 (9.5%) and 0 (0%) electrodes inside
and outside the resection during REM sleep, respectively. The relationship
of the high-rate FR electrodes to the area of surgical resection was not
different between NREM and REM sleeps. The occurrence of FRs was associated with the area of resection during REM sleep at the 200 – 299Hz
(P ＜ 0.0001), but not at the 300 – 399Hz frequency range.Conclusions:
Influence of sleep stages is probably smaller on the FR than on ripples.
Introduction: Recent studies revealed that left ventricular diastolic dysfunction is associated with development of heart failure. Advanced age and
high blood pressure have been reported as risk factors for progression of
left ventricular diastolic dysfunction, but relationships of chronic kidney
disease (CKD) are not well considered.Aim: To investigate the relationships
between CKD and progression of left ventricular diastolic dysfunction.
Methods: A historical cohort study was performed. We included patients
who were inpatient or outpatient of Kanazawa University Hospital and
received echocardiography examination for more than once with intervals
of more than one year. We excluded patients with organic heart disease,
such as valvular disorder or clinically diagnosed coronary artery disease.
Patients were examined their left ventricular peak velocity of blood flow
across the mitral valve (E) and their diastolic peak velocities of mitral annulus (e´). We calculated the ratio (E/e´) as an index of left ventricular
diastolic function. Low glomerular filtration rate (GFR) was defined as
estimated GFR (eGFR) less than 60 ml/min/1.73 m2. Relationship between
changes of E/e´ and status of CKD were examined using linear regression
model.Results: A total of 705 patients met the eligibility criteria. The mean
follow-up period was 3.3 years. Patients with low GFR showed significant
increase in E/e´ compared to patients without low GFR (adjusted mean
+0.37/year and +0.06/year, respectively; p=0.01). Analysis in patients
with urinary examination revealed that either proteinuria or low GFR were
significant risk factor for increase in E/e´; moreover, their combination
showed a marked progression (adjusted mean +0.56/year).Conclusion:
CKD appears to be a risk factor for the progression of left ventricular diastolic dysfunction progress.
128
Experience of the first certification of ISO15189:2012 in
physiological examination in Japan
PG-21
Hidemasa Matsuo , Kanako Suzuki, Kuniko Iwata, Tomoya Yoneda, Yuko Nakayama, Takeshi Higuchi,
Shuichi Shiga, Satoshi Ichiyama
Non-Invasive evaluation method of the liver fibrosis using
ELF score and shear wave elastography
Koji Yamamoto, Yoshiteru Fukumoto, Hiroko Ushiba, Hiroshi Nakano, Atsuya Shimizu
Saiseikai Matsusaka General Hospital, Japan
Department of Clinical Laboratory, Kyoto University Hospital, Kyoto, Japan
Educational Lecture
We have reported the process to acquire ISO15189 in physiological examination. We are planning to confirm the effect of ISO15189 on education of staff, their way of thinking, and the number of incidents/accidents
in the future. At Kyoto University Hospital, multinational clinical trials
including iPS cell research will be promoted as a medical institution with a
physiological laboratory certified by ISO15189.
Utility of monitoring spinal cord function during cervical
cord surgery
PG-23
Intraoperative motor evoked potential monitoring method utilizing cross-correlation coefficient
Utility of intraoperative motor evoked potential (MEP) monitoring method that applies crosscorrelation coefficient
Hiroyasu Oe 1, Yusuke Nakade1, Yuko Manbu1, Mikio Nagahara1, Mika Mori2, Kenshi Hayashi2,
Yoshio Sakai2, Takashi Wada2
Handa City Hospital, Japan
1 Department
of Clinical Laboratory, Kanazawa University Hospital, Japan
2 Department
of Nephrology and Laboratory Medicine, Graduate School of Medicine, Kanazawa University
129
Poster Presentation
Introduction: Nerve monitoring during an operation is assessed by the
change of the amplitude in the motor evoked potential (MEP). However,
the operative field environment is frequently influenced by the operative
procedure, which makes assessment difficult. In the present study, the
utility of an MEP monitoring method with the use of the cross-correlation
function was examined.Methods: Twenty examples of surgical cases in
the brain and spine areas were investigated. The analyses included comparisons of the shapes of the preoperative control waves with those of the
recorded intraoperative waves to calculate the cross-correlation coefficient
(CR).Results: The cross-correlation function of the monitored wave shape
presented a damped oscillation pattern. The dissociation was observed by
the wave recovery process, although the CR usually corresponded to the
maximum value (MAX CR) and zero circulation point values (τ= 0 CR). This
dissociation is explained by the change in the latency during the recovery
process. The MEP monitoring technique with the use of the CR was able
to detect the change in the entire monitor waveform by the transition
of the CR; furthermore, the partial waveform change that was unable to
be detected by the latency method and the amplitude measurement was
caught by the CR technique. Moreover, the CR was also considered to
be advantageous for measurements in a noisy environment, because the
random noise was removed and only the periodic element was used for
calculation.Conclusion: The MEP monitoring technique with the use of the
CR possessed an anti-noise characteristic, enabling the detection of slight
changes in the evoked potential waveform. Therefore, it is expected to be
useful as an intraoperative monitoring technique overcoming the issues of
conventional methods.
Oral Presentation
Background: Spinal cord function monitored is used to evaluate the
neurological function of anesthetized patients during surgery, including
detecting perioperative neuropathy and intervening to improve outcomes.
In this patient study, the association between wave pattern change and
prognostic evaluation was monitored during cervical cord surgeries.
Materials: Forty-two cervical vertebrae surgeries of the middle low rank
domain were performed, with monitoring. Participants were 30 men, and
12 women (24_82 years of age at the time of surgery; mean =62.6 year).
Methods: Transcranial motor evoked potentials were used for monitoring.
The stimulus conditions had a current intensity of 180-200mA, duration
of 0.3ms, and 5 stimulus trains. Complete vein anesthesia was achieved
by using propofol and remifentanil, with a muscle relaxant during intubation. Control waveforms were recorded after development after train of
four was confirmed as ＞ 75%.Study: This study considered the amplitude
and latencies of wave pattern from the deltoid, biceps, triceps, abductor
pollicis brevis(APB), and abductor hallucis(AH).Results: AH latencies decreased by an average of 0.9ms by the end of surgery, and APB latencies
decreased by anvaverage of 1.1ms. Amplitude increases were observed
in deltoids(114%), biceps(116%), and Triceps(119%). As disease duration
is shorter cases, recovery of the waveform was remarkable.Conclusion:
Monitoring by sensing pressure on the nerve may not be required when
spinal cord function is monitored during cervical spine surgery. Increased
amplitudes of evoked wave patterns reflected recovery of the marrow
clause obstacle, and were indicated by recovery latencies for the strand
road obstacle.
Case Conference
Keita Nishiwaki , Hiroka Tamura, Natsumi Aoki, Daichi Kiyohara, Kazuya Saito, Toshihisa Funabashi,
Koichi Sugiura
Symposium
PG-22
Special Lecture
In March 2013, we started to prepare standard operating procedures
(SOPs) of each examination including EKG, EEG, USG, and spirometry.
SOPs had to be written to satisfy 20 requirements defined as ISO15189,
however, the requirements were originally defined for examination of
specimens, not for physiological examinations and it was difficult to
meet the requirements. The criteria for emergency calls in each examination, including abnormal EKG and epileptic seizure in EEG, were unclear.
Therefore, we discussed and defined the criteria with clinicians. Moreover,
we made manuals (laboratory infection control manual etc.) to ensure
safety for patients and medical staff. We also made the daily check-lists
containing the name of the person in charge, machines' conditions, room
temperature and humidity, and maintenance history to ensure highquality examinations. A cause and effect diagram, called a "fishbone" was
made for each examination, which enables us to find the cause of unstable
results efficiently. Through the internal audit, main examination by Japan
Accreditation Board (JAB), and correction of problems, we finally acquired
the certification in May 2015.
Invited Lecture
At present transdermal liver biopsy is still gold standard method as an index of liver fibrosis in a chronic liver disease. But it is invasive and involves
danger of a complication, and operation of continuous observation by this
method is also difficult. So, we evaluated liver cirrhosis patients using two
non-invasive testing methods (Enhanced Liver Fibrosis score and shear
wave elastography) reported overseas.Enhanced Liver Fibrosis (ELF) test is
an in vitro diagnostic multivariate index assay intended to provide a single
ELF score by combining in an algorithm the quantitative measurements
of hyaluronic acid (HA), amino-terminal propeptide of type III procollagen
(PIIINP) and tissue inhibitor of metalloproteinase 1 (TIMP-1) in human
serum. Shear Wave Elastography (SWE) measures the speed propagated
in the organization using a shearing wave.We made the cutoff value of the
liver cirrhosis 10.65 of ELF score value and 2.11 m/s of SWE value this
time. The sensitivity, specificity and positive predictive value(PPV) of ELF
score and SWE of this study, were 80.0%,77.5%,69.2%,96.4%,62.2%,65.9%
respectively. ELF score and SWE are useful as one of non- invasive method
of liver fibrosis.
ISO 15189 is a well-known international standard for medical laboratories. More and more medical laboratories acquire the certification in
Japan. However, until now, there have been no physiological laboratories
certified by ISO15189, because the physiological examination is a unique
task for medical technologists, and the accreditation system had not
been established. Here, we report the first case of acquiring ISO15189
(ISO15189:2012) in physiological examination.
Keynote Speech
PG-20
Keynote Speech
PG-24
Nocturnal sleep and respiration in pregnant women with and
without obesity and non-pregnant women
PG-25
Influence of changes of head position on balance assessed by the
Gravicorder
Consideration of the output test and Frankfort horizontal plane
Midori Ura 1, Keisaku Fujimoto2, Haruna Yamazaki3, Yuka Teramae4
Sasahara Kinuyo , Triumi Yukiko, Oda Yasuko, Tanaka Yuuko
1 Shinshu
University Hospital, Graduate school of Medicine, Shinshu University, Japan
2 Shinshu
University
3 Shinshu
University Hospital
4 Tokyo
Metropolitan Ohtsuka Hospital
Kanagawa Dental Universty Yokohama Clinic, Japan
Invited Lecture
Special Lecture
Introduction: It has been reported that head position (submaxillary position) is important for balance.We studied the relation between balance
and head position, as well as the relations between balance and the output
test or the Frankfort horizontal plane.Subjects and Methods: The subjects
were 49 persons (28 males and 21 females with a mean age of 25.4 years
or 25.1years, respectively). They underwent assessment of balance by
Gravicorder with head position changes. They also underwent the Mann
test, one-leg test, blindfolded vertical writing test, stepping test, hearing
test, and assessment of the Frankfort horizontal plane.Results: Most of the
subjects showed no problems in the hearing test, Mann test, one-leg test,
and blindfolded vertical writing test. When the influence of head position
changes on balance was investigated, there was a significant difference of
the deflection envelope area between the 90 degrees head position and
the 45 degrees head position (t-test). With regard to the Frankfort horizontal plane and the stepping test, a significant correlation between balance
(deflection envelope area) and the Frankfort horizontal plane or stepping
test was found (Pearson's test).Conclusion: These findings suggest that
balance (deflection area) is influenced by head position and the Frankfort
horizontal plane and stepping test.
Educational Lecture
Introduction: Obesity is a potential risk factor for the onset of gestational
diabetes mellitus (GDM) in pregnant women. We examined the nocturnal
sleep and respiration of pregnant women with and without obesity and
non-pregnant women to investigate the relationships between obesity
and sleep disorders in pregnant women.Methods: Nine pregnant women
aged 24-40 years (mean ± standard deviation: 33.8 ± 4.9) and 12 nonpregnant women aged 20-24 years (22.3 ± 1.2) participated in the study
after their written informed consent was obtained. The pregnant women
were divided into two groups depending on their body mass index ([BMI]:
kg/m2) before pregnancy: five women with obesity (BMI of 30 or over) and
four women of normal weight (BMI <25). The following data were collected simultaneously during the night from the pregnant women at the 37th
week of pregnancy: respiratory disturbance index (RDI), oxygen saturation,
electroencephalograms, and autonomic nerve activity. Data for the above
measures were also collected from non-pregnant women, as comparison
controls. A Kruskal-Wallis H test was used to compare the data among the
groups.Results:Significant differences (all p ＜ 0.05) were observed among
the groups for the RDI (obese: 10.3 ± 2.7, non-obese: 5.7 ± 2.5, control:
2.3 ± 1.2). Compared to the other groups, significantly decreased oxygen
saturation was observed in the women with obesity (94.3 ± 1.3, 96.6 ±
0.1, 96.4 ± 1.1). Decreased deep sleep (non-REM3) and sleep efficiency
were observed in both groups of pregnant women compared to in nonpregnant women. Moreover, four women with obesity developed GDM at
various stages of pregnancy, whereas no pregnancy-induced complications occurred in the pregnant women without obesity.Conclusions: The
pregnant women with obesity experienced more complications and serious sleep-disordered breathing during pregnancy. Worsened sleep quality
might contribute to the development of complications among pregnant
women.
Symposium
PG-26
The estimated pulmonary artery systolic pressure by
echocardiography to grade the severity of heart failure
PG-27
Novel Echocardiographic Method to Estimate Pulmonary
Vascular Resistance Based on Measurements of Pulmonary
Regurgitant velocities
Case Conference
Shunsuke Suzuki 1, Maki Naitou1, Naoki Hiramatsu1, Akihiro Sonoda1, Hiroki Sakamoto2,
Genichi Sakaguchi3, Toshio Shimada4
Sanae Kaga1, Kazunori Okada1, Nobuo Masauzi1, Masahiro Nakabachi2, Hisao Nishino2,
Shinobu Yokoyama2, Mutsumi Nishida2, Taisei Mikami1
1 Department
of Clinical Laboratory Medicine, Shizuoka Prefectural Hospital Organization, Shizuoka General Hospital, Japan
2 Cardiovascular
Medicine, Shizuoka Prefectural Hospital Organization, Shizuoka General Hospital
3 Cardiovascular
Surgery, Shizuoka Prefectural Hospital Organization, Shizuoka General Hospital
4 Clinical
Research Center, Shizuoka Prefectural Hospital Organization, Shizuoka General Hospital
1 Faculty
of Health Sciences, Hokkaido Univ., Japan
2 Div.
of Laboratory and Transfusion Medicine, Hokkaido Univ. Hosp.
Oral Presentation
Poster Presentation
Introduction: Pulmonary vascular resistance (PVR) is an important
hemodynamic parameter in patients with heart failure, especially when
pulmonary arterial pressure is reduced owing to decreased stroke volume.
Although several echocardiographic methods to estimate PVR have been
proposed, their applications in patients with left-sided heart diseases have
been limited. The aim of the present study was to examine the usefulness
of our new method to estimate PVR (PVRPR) based on the continuous-wave
Doppler velocity measurements of pulmonary regurgitation in these patients. Methods: We studied 43 consecutive patients who underwent right
heart catheterization and echocardiography within one day. PVRPR was
calculated as the difference between the Doppler-derived early- and enddiastolic pulmonary artery (PA)-right ventricular (RV) pressure gradients
divided by the cardiac output measured in the left ventricular outflow tract
by echocardiography.Results: The PVRPR better correlated with invasive
PVR (PVRCATH) (r=0.81, p ＜ 0.001) than any of the conventional echocardiographic PVRs reported by Scapellato et al. (r=0.49), Abbas et al. in
2003 (r=0.54), Dahiya et al. (r=0.54), Lindqvist et al. (r=0.76), Abbas et al.
in 2013 (r=0.66) and Kanda et al. (r=0.76). In the receiver operating characteristic analyses to determine the patients with abnormal elevation of
PVRCATH ( ＞ 3 Wood units, WU), the area under the curve was greater for
PVRPR (0.985) than the conventional PVRs (0.705-0.839). PVRPR had 100%
sensitivity and 97% specificity at the optimal cut-off value of 2.95 WU in
identifying patients with PVRCATH ＞ 3 WU. Conclusion: Our new method
based on the continuous-wave Doppler measurements of early- and enddiastolic PA-RV pressure gradients is useful for the noninvasive estimation
of PVR in patients with left-sided heart diseases.
Introduction:Natriuretic peptide family (NPF: ANP, BNP and NT-proBNP)
concentration is highly reliable for objectively grading the severity of
heart failure (SHF) and has been widely used as biomarkers to grade and
monitor SHF both at rest and during exercise. In contrast, transthoracic
echocardiography (TTE) has been widely used for noninvasive assessment
of a hemodynamical SHF. The aim of this study is to clarify if NPF concentration measured on the same day as TTE is helpful for grading SHF
in comparison with echo parameters, including the estimated pulmonary
artery systolic pressure (PASP).Methods:218 consecutive patients with
chronic HF who underwent TTE and blood sampling at the same time
were recruited. Several variables were extracted using a multivariate logistic regression model with the dependent variable, PASP divided up and
down by the median. One-way analysis of variance (ANOVA) was executed
on the extracted significant variables for PASP category.Results:NPF was
extracted as a significant variable.PASP was classified in ascending order
to make up quartile groups (the 1st quartile group (A): 5.13-20.98mmHg,
the 2nd quartile group (B): 20.98-24.16mmHg, the 3rd quartile group(C):
24.16-29.21mmHg, the 4th quartile group (D): 29.42-65.69mmHg). As
each NPF concentration was increasing, PASP increased proportionally,
and the result of ANOVA showed that each NPF concentration was much
higher in the other groups than in control group (the 1st quartile group
(A)).Conclusion:PASP is proportionally and concomitantly related to NPF
concentration, which reflects SHF objectively. Therefore, we conclude that
PASP is a potent factor for grading and monitoring SHF.
130
Diagnosis of Multiple Atrial Septal Defects by Transthoracic
Echocardiography
PG-29
Nobuhisa Watanabe 1, Hiroki Oe1, Teiji Akagi1, Youichi Takaya2, Ken Okada1, Hiroshi Ito2
Rika Takemoto 1, Hiroki Oe1, Nobuhisa Watanabe1, Kazufumi Nakamura2, Hiroshi Morita2, Ken Okada1,
Fumio Ootsuka1, Hiroshi Ito2
1 Okayama
University Hospital, Japan
2 Okayama
University
1 Okayama
University Hospital, Japan
2 Okayama
University
Yuri Mizutani 1, Yuka Takeuchi2, Hirofumi Kusuki3, Keiko Sugimoto1, Keisuke Osakabe1, Naohiro Ichino1,
Tadayoshi Hata1
Diabetic patient with hypertensive response to light exercise
is related to poor exercise tolerance
Symposium
PG-31
Educational Lecture
The Dynamics of Repolarization Interval in Children with
Ventricular Septal Defect
Special Lecture
Introduction: Fractional area change(FAC) has been shown to correlate
well with right ventricular(RV) ejection fraction(EF) by cardiac magnetic
resonance(CMR). The guideline recommends that multiple echocardiographic views should be obtained to evaluate RV function. However, there
is no clear recommendation which view should be used to evaluate RV
FAC.Methods: CMR and transthoracic echocardiography(TTE) were performed in 90 consecutive patients for evaluation of RV function. We measured tricuspid annular plain systolic excursion (TAPSE), FAC on apical four
chamber view (A4CV)(A-FAC) and that of RV focused A4CV (F-FAC) and
modified A4CV (M-FAC) and pulsed Doppler peak velocity at the tricuspid
annulus (s’) as an assessment of RV systolic function. The association
between echocardiography-derived parameters of RV systolic function
and CMR-derived measurement of RVEF and RV volume were evaluated.
Results: Both A-FAC and F-FAC measurement were fesible in 90 patients
(100%) and M-FAC measurement was fesible in 84 patients (93%). End
diastolic area (Area ED)(cm2) of A-FAC and F-FAC showed correlation with
CMR-derived end diastolic volume, respectively (p< 0.0001, r=0.768 vs
p< 0.0001,r=0.784 ). End systolic area (Area ES)(cm2) of A-FAC and F-FAC
showed correlation with CMR-derived end systolic volume, respectively (p<
0.0001, r=0.791 vs p< 0.0001, r=0.820) A-FAC and F-FAC has good correlation with CMR-derived RVEF(p< 0.0001, r=0.638 vs P< 0.0001,r=0.663,
respectively). M-FAC has no correlation with CMR-derived RVEF(p=N.
S., r=0.438). There were no significant agreement between TAPSE, s’ and
CMR-derived RVEF in this study.Conclusion: Both A-FAC and F-FAC had
good correlation with CMR-derived RVEF, and M-FAC didn’ t show correlation with CMR-derived RVEF. We should use F-FAC to evaluate FAC.
Invited Lecture
Background: The prevalence of multiple ASDs is approximately 8-10% of
all ASD and transcatheter closure for multiple ASDs is still challenging.
However, accurate diagnosis of multiple ASD using transthoracic echocardiography (TTE) is always difficult even in experienced sonographer. In
this study, we prospectively investigated the diagnostic ability of TTE for
patients with multiple ASDs in our institution. Methods: We enrolled 374
patients with secundum ASD referred to our institute for transcatheter
closure. All patients underwent transthoracic echocardiography (TTE) first
and then trasnsesophageal echocardiography (TEE) before the procedure.
All TTEs were performed by well-trained sonographer. We checked the
timing when the accurate diagnosis was made.Results: Thirty-seven patients (9.9%) were diagnosed with multiple ASDs. Twenty-seven patients
(73.0%) were diagnosed by TTE and 8 patients were diagnosed by subsequent TEE. Two patients (5.4%) were identified multiple defects during the
sizing balloon procedure.Conclusion: Even in TTE evaluation, more than
70% of multiple ASDs can be diagnosed before the catheter intervention
if it was performed by well-trained sonographer. Such pre-interventional
information can be contributed to the valuable information for the establishment of therapeutic strategy.
PG-30
Right ventricular FAC obtained in different echocardiographic
views. Comparison with RVEF by cardiac-MRI.
Keynote Speech
PG-28
Motoki Otsuji , Ayaka Matsumoto, Katsunori Bettou
Ise Red Cross Hospital, Japan
131
Poster Presentation
Introduction: In patients with ventricular septal defect (VSD), the left to
right shunting increases the left ventricular preload. This pathological
dynamics modulates the myocardial depolarization and repolarization
processes and causes arrhythmogenic substrates. Variability in the repolarization interval of children requiring surgery was compared with that
of healthy children to elucidate the effect of VSD on myocardial repolarization. Methods: The subjects were 25 children with VSD who required
surgical closure (mean left-to-right shunt ratio: 2.60 ± 0.55). Subintervals
(QT, JT, J point to T peak; JTp, T peak to T end; Tp-e) were determined
from preoperative ECG, and the corrected heart rate and variability ratio in
the repolarization (variability index, VI) were estimated. The corrected repolarization interval and VI were compared between the group requiring
surgery and matched healthy group (25 children) of age. Results: Significant differences in corrected QT, JTp, and Tp-e intervals between the two
groups were found. The VI of subintervals (QTVI, JTVI, JTpVI and Tp-eVI)
also showed significant differences. However, using a linear regression
analysis no correlation was found in the QTVI and QTc. Conclusion: Children requiring surgery were confirmed as having higher myocardial repolarization variability due to enhanced autonomic nervous activity based on
changes in hemodynamics.
Oral Presentation
Introduction: Diabetic patients for education hospitalization in our hospital has implemented a treadmill stress test (TMT) for both detection of
asymptomatic myocardial ischemia and evaluation of exercise tolerance.
In TMT, hypertensive response with the short time of the load was often
shown. However, it was fully evaluated whether hypertensive response
to exercise (HRE) was related to exercise habits and exercise tolerance.
Methods: From September 2011 to September 2014, type 2 diabetic
patients without myocardial ischemia were enrolled in our study. Systolic
blood pressure at early exercise (e-SBP) was measured at 1.5METs at
Bruce protocol. One hundred sixty diabetic patients were divided into 3
groups, Control: resting systolic blood pressure (r-SBP) <130mmHg and
e-SBP <160mmHg, HRE: r-SBP <130mmHg and e-SBP ＞ 160mmHg, and
Hypertension (HTN): r-SBP ＞ 130mmHg. The history taking of exercise
habits, blood sampling, pulse wave velocity (PWV), and transthoracic
echocardiography at rest were examined, and the association with blood
pressure response to exercise was evaluated.Results: Fifty two patients
(35%) had exercise habits. There were no significant differences about age,
body mass index, HbA1c, and LVEF among 3 groups. The duration of exercise was significantly shorter, and PWV and E/&Eacute; were significantly
higher in HRE and HTN groups compared with Control groups (Control:
520 ± 242, HRE 356 ± 140, HTN 399 ± 148 (sec), 1396 ± 424, 1595
± 356, 1679 ± 340 (cm/sec), and 8.7 ± 2.5, 9.7 ± 2.2, 10.8 ± 3.0,
p<0.05, respectively). There were no significant differences about exercise
habits among 3 groups, however, both HRE and HTN groups tended to
have poor exercise habits compared to Control groups (Control: 44%, HRE:
33%, HTN: 21%, p=ns). Multivariable regression analysis with clinical variables demonstrated that e-SBP was an independent determinant of exercise duration with standardized coefficient of -0.454. Conclusion: Diabetic
patients with hypertensive response to light exercise had poor exercise
tolerance, regardless of resting systolic blood pressure.
Case Conference
1 Graduate
School of Health Sciences, Fujita Health University, Japan
2 Divi.
of Clinical Laboratory, Ise Red Cross Hospital
3 Divi.
of Laboratory, Chukyo Hospital, Japan Community Health Care Organization
Keynote Speech
PG-32
Usefulness of the facial nerve motor evoked potentials in skull base surgery.
Evolution of transcranial facial motor evoked potentials using supra threshold
level stimulation method.
PG-33
Relationship between olfactory function and gustatory
function and pathophysiology in Alzheimer's disease
Invited Lecture
Special Lecture
Educational Lecture
Tsunenori Takatani , Sayomi Yamamoto, Hideko Yoshida, Yayoi Umeki
Minoru Kouzuki, Syouta Nakamura, Yuto Katsumata, Yuki Fujihara, Ayumi Takamura, Katsuya Urakami
Division of Central Clinical Laboratory Nara Medical University, Japan
Department of Biological Regulation, School of Health Science, Faculty of Medicine, Tottori University, Japan
Background: The preservation of facial nerve function is one of the primary objectives in skull base surgery. Tc-FNMEPs has been recognized as
a good method for quantitative monitoring of facial nerve function in skull
base surgery. Its function can be continuously monitored by Tc-FNMEPs in
facial nerve target muscles. While most authors use a 50% reduction in FNMEP response amplitudes as a warning criterion,in this paper the authors
approach was to keep the response amplitude constant by increasing the
stimulation intensity and to establish a warning criterion based on the "supra threshold-level" method.Methods: 38patients undergone elected skull
base surgery using Tc-FNMEP monitoring were studied. Supra threshold
level transcranial stimulation of Tc-FNMEP was established with minimum
intensity to elicit the waveform from recording muscles. A train of 4 pulses
was delivered through corkscrew electrodes at C3/C4. Subdermal needle
electrodes placed in the orbicularis oculi and oris muscle for recording.
Significant change of amplitude was defined as more than 50% decrease
compared with baseline amplitude. Facial nerve function was evaluated
preoperatively and postoperatively using the House & Brackmann grading system. The reliability of Tc-FNMEP was assessed by sensitivity and
specificity to detect postoperative facial nerve dysfunction.Results: Control
Tc-FNMEP waveforms were successfully recorded in all patients. Of 38
patients, significant sustained decreases of Tc-FNMEP until the end of
surgery were observed in 9 patients. Postoperative new facial nerve dysfunction or worsen facial nerve function were observe in 7 patients. Of the
7 patients with postoperative deterioration of facial nerve function, 4 patients had intraoperative significant decline of Tc-FNMEP. The sensitivity
and specificity of intraoperative Tc-FNMEP to detect postoperative facial
nerve dysfunction were 85% and 100%, respectively.Conclusions: The
results in this study indicated the feasibility of intraoperative Tc-FNMEP
using supra threshold level transcranial stimulation during skull base surgery.
IntroductionPatients with Alzheimer's disease (AD) are expected to develop olfactory dysfunction in the early stage by senile plaques and neurofibrillary tangles in olfactory-related domain. In addition, patients with
dementia may cause taste disorder by cerebral degeneration. However,
no study investigates olfactory and gustatory function for mild cognitive impairment (MCI) which is a pre-AD state, and it is not clear about
relationship between pathology. The aim of this study is to investigate
olfactory and gustatory function in AD and MCI compared with healthy
elderly subjects, and analyze correlation with those functions and pathophysiology.Methods31 AD patients (79.1 ± 10.2 years), 9 MCI patients
(77.9 ± 3.1 years) and 15 healthy elderly subjects (71.9 ± 11.8 years)
were tested Odor Stick Identification Test for Japanese (OSIT-J), intraoral
dropping method using taste solutions, Touch Panel-type Dementia Assessment Scale (TDAS) and beta-amyloid (A Β ) 42 and phosphorylated
tau (p-tau) 181 in cerebrospinal fluid (CSF) by ELISA.ResultsOSIT-J scores
decreased significantly in AD group compared with other two groups. In
addition, specific correlation was provided with A Β 42, p-tau181 and
TDAS scores. On the other hand, taste test scores showed no significant
difference among the three groups. In comparison with other tests, there
was significantly only correlation with TDAS scores. ConclusionOlfactory
function was related to CSF biomarkers and cognitive disorders. Therefore,
it is suggested that olfactory function likely to be impaired in the early
stage of pathology. However, it was not able to distinguish between MCI
and healthy elderly subjects. It is necessary to develop a more sensitive
olfactory test. In gustatory function, there was specific correlation with
TDAS scores. But, there was not associated with CSF biomarkers and no
significant difference among three groups. Therefore, this study showed
that gustatory function may not impaired in the early stage of disease.
Symposium
PG-34
Detection of left ventricular hypertrophy using
electrocardiography in cases pre-identified by
echocardiography
PG-35
Evaluation of effects of introducing exercise therapy in
cardiac rehabilitation
Case Conference
Maya Ishiguma, Toshiharu Umeki, Ichirou Tanabe, Taemi Akiyoshi, Takanori Higashitani,
Eisaburo Sueoka
Takeshi TERAJIMA 1, Takashi OHORI2, Masashi SORIMACHI1, Katsumi KANEKO3, Mio ONDA3,
Kyoko TAGUCHI3, Motohiko UEKI4, Yoshiko ABE3
Saga University Hospital, Japan
1 Niigata
Koseiren Uonuma Hospital, Japan
2 Niigata
Koseiren Joetsu General Hosp.
3 Niigata
Koseiren Itoigawa General Hosp.
4 Niigata
Koseiren Keinan General Hosp.
Oral Presentation
Poster Presentation
SummaryUltrasound cardiography (UCG), is a reliable method to detect
left ventricular hypertrophy (LVH). The 12-lead electrocardiogram (ECG)
is often used for LVH screening as well. However, several different sets of
criteria exist for diagnosing LVH by ECG, and their sensitivity and specificity differ by report depending on differences in race and body type. Therefore, we assessed the reliability of two sets of ECG criteria against that of
UCG in the detection of LVH.Subjects and MethodsEighty-eight patients
who underwent UCG and were diagnosed with LVH at Saga University
Hospital between January and December 2015 were included in our
study. All subjects received ECGs; the percentage of correctly positive LVH
diagnoses from ECGs was assessed using the UCG findings. Two sets of
criteria for detecting LVH with ECGs were used: the Sokolow-Lyon criteria
(S standard) and the Cornell voltage criteria (C standard). ResultsAmong
88 patients, 67 subjects were male and 21 were female. Using the S criteria, 16 (18.2%) cases were incorrectly considered LVH-negative; using
the C criteria, 19 (21.6 %) cases tested negative. Using both sets of criteria
together, 25 (28.4%) cases tested negative. The average UCG-measured
wall thicknesses in cases that could not be detected by the combined ECG
criteria were 13.5 ± 0.34mm (max 19mm) for ventricular septa and 12.4
± 0.37mm (max 16mm) for posterior walls. Among 25 cases classified as
LVH according to UCG, one case was assessed as normal, 12 were concentric remodeling, and 12 were concentric hypertrophy.ConclusionWe found
that approximately 28% of LVH cases diagnosed by UCG were not detected
by the combined set of ECG criteria for diagnosing LVH. In our study, the
detection rate of LVH by ECG did not differ according to which set of criteria was used to diagnose LVH.
Introduction: Cardiac rehabilitation (CR) is a comprehensive program
focused on exercise therapy and includes lifestyle guidance, patient education, and counseling. Its goal is to improve motility, prevent disease
relapse, and increase quality of life in patients with heart diseases such
as myocardial infarction, angina pectoris, and heart failure, and in those
with PAD, after cardiovascular surgery, and with DM. Target, Methods:
We included 52 subjects (32 men, 20 women) with a mean age of 70.5
± 8.5 years. CPX(peakVO2, AT, VE/VCO2), 6MWD, UCG(EF, LVDd), blood
pressure-pulse wave tests (CAVI, ABI),and blood tests (BNP, hs-TnT) were
performed before and after CR.Results: Significant improvements were
observed for peakVO2, from 900.3 ± 214.7 mL/min to 1006.6 ± 272.3
mL/min (p ＜ 0.001); AT, from 597.7 ± 145.0 mL/min to 673.5 ± 138.9
mL/min (p ＜ 0.001); VE/VCO2, from 34.2 ± 9.5 to 31.3 ± 6.5 (p ＜ 0.01);
6MWD, from 432 ± 76 m to 495 ± 89 m (p ＜ 0.001); EF, from 58.6 ±
15.2 to 61.5 ± 12.2 (p ＜ 0.03); BNP, which went from 117.1 ± 115.3
pg/mL to 91.4 ± 89.8 pg/mL (p ＜ 0.03); and hs-TnT, to 0.021 ± 0.001
ng/mL (p ＜ 0.001). Significant improvements weren’t observed for LVDd,
from 49.3 ± 7.5 to 48.1 ± 7.7(n.s.); CAVI, from 9.1 ± 1.4 to 9.0 ± 1.3(n.
s.); ABI, from 1.08 ± 0.12 to 1.10 ± 0.13(n.s.).Discussion: The 150 days
of CR caused skeletal and respiratory muscle to develop, vascular endothelial functions to recover, cardiac and peripheral circulating blood volume
to increase, and exercise tolerance and motility to improve significantly.
For cardiac functions, LV remodeling did not occur, and cardiac systolic
and diastolic functions improved. Further, the hs-TNT results indicated
exercise increased cardiac volume, attenuated sympathetic nerve activity,
and suppressed myocardiopathy. Improvements were also seen in peripheral arterial hardness and blockage. Conclusion: CR can improve exercise
tolerance, which is directly linked to daily activities; suppress diseases or
prevent their recurrence; and improve quality of life.
132
Assessment of fever for infection control using thermography
Facial thermography in patients with fever
PG-37
Osamu Horie 1, Hiromi Shibata2, Gou Tsuji3, Shunichi Kumagai3, Masahiro Koshiba4
Rie Mitsui 1, Yosuke Sugioka1, Nobuki Fukuhara1, Michitaka Kato2, Fumi Nihei1, Akira Kubo1,
Yoshihiko takeda1
1 Tenri
Health Care University, Japan
2 Hyogo
University of Health Sciences
3 Shinko
Hospital
4 Hyogo
College of Medicine
1 Ginza
Hospital, Japan
2 Tokoha
University
Symposium
Yuka SHIBATA 1, Taeko KAMIYA1, Ryuhei KANDA1, Hiroya TANI1, Takahiko KISHI1, Minehiro GOTOU1,
Hideki KAMIYA2, Jiro NAKAMURA2
About ABI value and pulse waveform numbers in ASO screening
Cases UT of blood pressure pulse waveform numerical value was
useful
Educational Lecture
PG-39
Special Lecture
A Point-of-care Sural Nerve Conduction Device (DPN
CheckTM )for the Identification of Diabetic Polyneuropathy
Invited Lecture
BACKGROUND:Blood tests and ultrasound are common ways to assess
liver function. Recently, the Controlled Attenuation Parameter (CAP), a
new tool of quantifying the degree of hepatic steatosis using the FibroScanR, has been developed to allow simultaneous measurement of hepatic
steatosis and liver stiffness. This study assessed the correlation between
the degree of hepatic steatosis measured by the CAP and various test
results to evaluate the usefulness of the CAP in assessing disease risks.
SUBJECTS:This study included patients who went through a medical
checkup at Ginza Hospital between June 2014 and March 2016 and had
their CAP and liver stiffness measured by the FibroScanR. Patients with
known liver diseases, including those with positive HCV or HBV results,
were excluded. A total of 192 patients (113 male and 79 female, mean
age 51.8 years old) were included. METHODS::We evaluated the association between the CAP and 227 measurements from the health checkup,
including blood tests, physiological tests, radiological tests, and patient
interviews. RESULTS:Among the 227 measurements, the CAP showed
significant correlation with BMI (r=0.570 P ＜ 0.001), waist circumference (r=0.562 P ＜ 0.001), total body fat (r=0.560 P ＜ 0.001), visceral
fat (r=0.545 P ＜ 0.001), total-PAI-1 (r=0.510 P ＜ 0.001), cholinesterase
(r=0.450 P ＜ 0.001), insulin (r=0.410 P ＜ 0.001), small-dense LDL
(r=0.384 P ＜ 0.002), ALT (r=0.384 P ＜ 0.001), free testosterone (r=-380
P ＜ 0.042), triglycerides (r=0.351 P ＜ 0.001), RLP-cholesterol (r=0.330 P
＜ 0.004), and high-molecular weight adiponectin (r= － 312 P ＜ 0.013).
DISCUSSION:The CAP correlated well with several diagnostic factors of
metabolic syndrome, and unlike waist circumference, it could be useful in
evaluating metabolic syndrome associated with visceral fat. The CAP could
also be valuable in assessing the risks of thrombotic and arteriosclerotic
diseases as it showed significant correlation with the total PAI-1 and other
atherosclerotic markers. CONCLUSION:The CAP is a noninvasive measurement and may be useful in assessing and managing the risk of various
diseases.
Introduction: To control infections such as Ebola haemorrhagic fever,
thermography can be used to monitor patients with fever resulting from
infection. However, evidence-based cut-off levels for the thermographic
index to reasonably discriminate patients with fever from healthy individuals have yet to be established. We therefore compared facial temperatures
between patients with fever and healthy volunteers, and reconsidered
assessment for infectious control using thermography. Methods: Thermographic examination was performed in 50 healthy volunteers, 48 patients
with influenza A, 13 patients with influenza B and 65 patients with the
other pyrogenetic disease. Results: Facial temperatures were significantly
higher for patients with fever than for healthy volunteers. We compared
facial thermography with axillary temperature as measured using a clinical
thermometer in patients with fever. Significant correlations with axillary
temperature were observed for all parts. Facial temperatures of patients
with fever were higher than those of healthy volunteers. However, some
patients showed lower temperatures than some healthy volunteers. Furthermore, the correlation coefficient was lower than previously report, and
the reliability of the data was considered low. The correlations are not sufficiently strong to clearly detect patients with fever.Conclusion: Thermography not only measure temperature, but also can provide the imaging. In
our efforts to develop a universally accepted method for infection control,
it would seem that thermography offers one of the more promising techniques. A new evidenced based standard of thermography for detection of
patients with fever is necessary to avoid spreading pyrogenetic disease.
PG-38
Potential use of measuring Controlled Attenuation Parameter
with the Fibroscan during health checkups
Keynote Speech
PG-36
Michiko Enshu, Yutaka Yoshimura, Sachiko Nakamura, Emiko Nakata, Keiko Yamaguchi
Nara Prefecture General Medical Center, Japan
133
Poster Presentation
“Background and Aims” For the diagnosis of diabetes polyneuropathy
using objective electrophysiological tests (nerve conduction study: NCS)
is used as golden standard method. But the use of NCS examination technique is limited to the specialized laboratories and technicians. A point-ofcare device , DPN CheckTM (HDN-1000: OMRON COLIN Co. Japan) can
quantitatively measure sural sensory nerve action potential in less than 5
minutes with no special techniques. In this study, sensory nerve conduction velocity and action potential amplitude were measured bilaterally to
know the usefulness of DPN CheckTM ,it compared with Neuropack X1
(NIHON KOHDEN CO. Japan) for the electromyography standard method.
"Methods" Fifty-seven diabetes patients (28 males) were enrolled with
informed concents on the DPN CheckTM examination. Mean age, mean
diabetes duration, mean HbA1c values, and mean BMI were 58.1 years, 8.9
years, 9.6 %, and 26.6 kg/m2, respectively. After measuring both legs with
Neuropack X1, DPN CheckTM was used to examine both legs. The results
using Neuropack X1 were compared with those using DPN CheckTM. The
two styles of using DPN CheckTM one is twice examinations by same technician, and the other single examination separately by two technicians,
were checked for reproducibility. "Results" Pearson correlations were performed to determine the association between DPN and electromyography
standard method. The correlation coefficient (R) for velocity and amplitude
were R=0.774 and 0.616 respectively.The reproducibility of the data by
DPN CheckTM was admissible. "Conclusions" There is difference about
principal s method, DPN CheckTM is orthodromic conduction method, and
the other is antidromic conduction method. But however both examination
results showed admissible statistical agreement correlation. DPN CheckTM
is designed for ease-of-use and probably useful device for diagnosis of diabetes polyneuropathy.E-mail address: yukau@aichi-med-u.ac.jp
Oral Presentation
Introduction: Vascular disorder has increased the blood vassel due to
stenosis, clogging arteriosclerosis.Among them, there is the Ankle Brachial
Index (ABI) as a screening test for occlusive sclerosis that occurs lower extremities. It can measure the presence or absence of ASO by measuring the
ABI in a simple.Methods: Obstruction, the index of stenosis is Mean Artery
Pressure (%MAP),Upstroke time (UT). ABI is a was the normal value ,ASO
in lower limb arteries echography and an ABI value of the patients was observed ,were examined pulse waveform numerical value (%MAP ,UT) for in
the course ofobservation. It reports some of the views were obtained. Use
equipmen, was used VaSera VS-1500A (Fukuda Denshi Co.,Ltd). Case:age
74-year-male.Chief complaint ,appearance pain sense of left and right foot.
Elapsed ,diabetes from 38 years of age. Angina in 70 years ,percutaneous
transluminal coronary angioplasty(PCI) enforcement.Result: Although ABI
value immediately after complaining of pain and %MAP was normal ,UT of
the reference value 180 the it was over. It was carried out follow up there
times in the subscription training test in ,but it was almost normal expect
UT. However ,about 40-50% of stenosis in both side the common femoral
artery was observed in the subsequent lower extremityartery echography.
Summary: Present case the atenosis 70% or less , lower limb blood pressure lowering because blood flow had been maintained were almost no.
However,at it had exceeded the reference value at a relatively early stage.
Occlusion and stenosis is considered to pulse waveform numerical value
than ABI value is preceded. Futher report repeated studies.
Case Conference
1 Department
of Clinical Laboratory Aichi Medical University Hospital, Japan
2 Division
of Diabetes, Department of Internal Medicine,Aichi Medical University
Keynote Speech
PG-40
Comparison between FibroScan and virtual touch
quantification for liver stiffness measurement in HCV patients
PG-41
A case of congenital biliary dilatation associated with
gallbladder and common bile duct cancers
Keisuke Osakabe1, Naohiro Ichino1, Toru Nishikawa2, Hiroko Sugiyama2, Tadayoshi Hata1,
Naoto Kawabe3, Senju Hashimoto3, Kentaro Yoshioka3
Rika Shimizu , Takako Oura, Harumi Fukuda, Yuko Sakurai, Makoto Morimoto, Kazushi Sugimoto,
Kaname Nakatani
1 Faculty
of Medical Technology, School of Health Sciences, Fujita Health University, Japan
2 Center
of Ultrasound Diagnosis, Fujita Health University Hospital, Aichi, Japan
3 Department
of Liver, Biliary Tract and Pancreas Diseases, School of Medicine, Fujita Health University, Aichi, Japan
Div. of clinical laboratory, Mie university hospital, Japan
Invited Lecture
Special Lecture
Educational Lecture
Case presentation: We herein report a case of 76-year-old female who
had congenital biliary dilatation (CBD) associated with gallbladder and
common bile duct cancers. Three years before, she underwent imaging
studies for elevation of CA19-9 and was diagnosed as having adenomyomatosis and gallbladder polyp. Two years before, the follow-up studies
showed the shape of gallbladder polyp becoming irregular and she was
referred to our hospital. Laboratory studies revealed no abnormal values
except for slight elevation of CA19-9 (79.5 U/ml). A diagnosis of adenomyomatosis, gallbladder polyp and gallbladder stone were simultaneously
made based on the findings of computed tomography (CT), endoscopy
ultrasound (EUS), and magnetic resonance imaging (MRI). Six months
later, the following CT showed gallbladder polyp was becoming enlarged,
raising the suspicion of malignancy. However, there were no remarkable
changes in the thickened gallbladder wall from body to fundus. One month
later, abdominal ultrasonography (AUS) was performed and showed a
cystic dilatation of the extrahepatic bile duct and a wide flat lesion on the
wall of the dilated common bile duct. A blood flow signal in the tumor
was also detected. The wall of the fundus of gallbladder was diffusely
thickened and hyper echoic lesion with acoustic shadow was seen in it.
MRI was performed again and it is suspected that she had the bile duct
tumor associated with CBD (Type Ia) and pancreaticobiliary maljunction.
An elevated lesion which was not enhanced at the fundus of gallbladder
was also detected. Surgical operation was performed and histological examination showed bile duct adenocarcinoma spreading to a lymph node
and gallbladder adenocarcinoma. Conclusion: In this case, the patient was
initially suspected as having gallbladder polyp which was later diagnosed
as adenocarcinoma in CBD. AUS revealed the patient had CBD, which led to
accurate diagnosis.
Aim: Recently, various apparatuses for measurement of liver stiffness using the ultrasound have been developed. The liver stiffness values (LS) by
FibroScan (FS) and the shear wave velocity (Vs) by virtual touch quantification (VTQ) was compared.Subjects: LS and Vs were measured in 112 patients with chronic hepatitis C virus infection who underwent liver biopsy
consecutively in Fujita Health University Hospital from November 2010 to
December 2014. Fibrosis stage by Metavir score wereF0-1 in 26 patients,
F2 in 25, F3 in 25 and F4 in 25.Methods: LS values (kPa) and Vs values
(m/sec) were measured ten times at the right intercostal space, and the
median value was adopted.Results: LS values were significantly correlated
with fibrosis stage of the Metavir score (r =0.642, P ＜ 0.0001). LS values
significantly differed between F0 – 1 and F2 (P =0.0044); between F2 and
F3 (P =0.0149). LS values significantly differed between F0 – 1 and F2 (P
=0.0044); between F2 and F3 (P =0.0149). The optimal cut-off value of LS
value was 6.0 kPa for F ＞ or = 2, 9.3 kPa for F ＞ or = 3 and 14.6 kPa for
F4.Vs value were significantly correlated with fibrosis stage of the Metavir
score (r =0.600, P ＜ 0.0001). Vs values significantly differed between F0
– 1 and F2 (P =0.0042); between F2 and F3 (P =0.0278). The optimal cutoff value of Vs value was 1.31 m/sec for F ＞ or = 2, 1.51 m/secfor F ＞ or
= 3 and 1.81 m/sec for F4.LS values was significantly correlated with Vs
values (r=0.762, P ＜ 0.0001).Conclusion: LS values and Vs values were
confirmed to be correlated with each other and also with fibrosis stage
of the Metavir score. The ability to diagnose each fibrosis stage of FS and
VTQ is equivalent with each other.
Symposium
PG-42
FCU recordings in the nerve conduction study for evaluation
of ulnar neuropathy at the elbow.
PG-43
Iwanaga Fumitomo 1, Matunaga Takuya1, Nishimura Kohei1, Taramoto Yasuyuki1, Miyamoto Utako2,
Nakanishi Ryoji3, Matsunaga Kaoru4
APNEA DUE TO GASTROESOPHAGEAL REFLUX
DISEASE IN A CASE OF NEWBORN BABIES
Keiko Ishigo , Naomi Nakashima, Seiko Sawamura, Manayo Hattori, Akari Tabata
Ogaki Municipal Hospital, Japan
Case Conference
1 Clinical
Neuro-physiological Loboratory, Kumamoto Kinoh Hospital, Japan
2 The
Department of Neurology, Kumamoto Kinoh Hospital
3 The
Department of Rehabilitation Medicine, Kumamoto Kinoh Hospital
4 The
Department of Neurology, Kumamoto Onjaku Hospital
Oral Presentation
Poster Presentation
Introduction Gastroesophageal reflux disease (GERD) is common in newborn babies or infants and can be physically recognized. When newborn
babies or infants have GERD, usually the symptoms resolve when they are
12 to 18 months old. However, severe cases of infant GERD can be associated with various symptoms. And respiratory symptoms such as chronic
coughing, wheezing, repeated respiratory infections, and apnea. Here, I
would like to report a case of a newborn baby who was not suspected of
having GERD and underwent a polysomnography check.Case Newborn
twin boys were delivered by cesarean operation at 32 weeks and 5 days
post-conception. The first baby boy weighed 1975 g with an Apgar score
of 4/7. The second baby boy weighed 1483 g with an Apgar score of 8/9.
They were referred to our hospital to be evaluated for apnea and arrhythmia when they were 42 weeks and 5 days old.Methods PSG and Halter
electrocardiography were performed simultaneously. The brain waves
during sleeping were used for the PSG. Breathing and an abdomen sensor tests were performed using respiratory inductance plethysmography
(RIP), in addition to a SUM analysis. Results The Halter electrocardiogram
showed no abnormalities. The first baby’s apnea hypopnea index showed
23.6 events/hour (central sleep apnea 201, obstructive sleep apnea 20,
mixed sleep apnea 10 and hyponea 231); the 2nd baby’s AHI was 14.0/
hour (CSA 82, OSA 7, and hypopnea 145).Conclusion The apnea may have
been due to the babies’ feeding or sleeping, which caused choking, thereby
reducing the heart rate and oxygen saturation. Although GERD was not
thought to be present, an upper gastrointestinal series was performed,
which revealed no abnormalities. The babies’ diet was changed from
breast-feeding to AR milk, after which, the incidences of choking and apnea were reduced.
Objective:The aim of this study was to investigate the utility of flexor carpi
ulnaris (FCU) recordings in the nerve conduction study for evaluation of
ulnar neuropathy at the elbow (UNE). Methods:Twenty-eight hands of 16
healthy volunteers and 26 hands of 19 patients with UNE were examined.
Compound muscle action potentials (CMAPs) were recorded from Abductor Digiti Minimi(ADM)and FCU muscles after electrical stimulation of
the ulnar nerve. We measured two following parameters; (1) motor nerve
conduction velocities (MCVs) obtained using both FCU and ADM recordings. (2) distal motor latencies of FCU-CMAPs recorded from stimulation of
ulnar nerve above the elbow (17cm proximal from the electrode position
on the FCU).Results:The optimal recording electrode position on the FCU
was approximately 10cm (about 2/5 of the length of the forearm) distal
from the elbow. Significant correlation(P<0.001) was found between ADMMCV and FCU-MCV. Distal motor latencies of the FCU were 3.60 ± 0.23ms
in normal subjects and 4.92 ± 1.24ms in patients. FCU-CMAPs could be
evoked even when ADM-CMAPs could not be in 3 patients. In addition,
ADM-MCVs could not be obtained in 2 patients since ADM-CMAPs could
not be evoked from stimulation of ulnar nerve below the elbow due to the
high stimulation threshold. In these patients, distal motor latencies of FCUCMAPs were significantly prolonged than the normal value. Conclusion:
FCU recordings were useful in the diagnosis of UNE, especially when ADMCMAPs cannot be evoked due to the severe intrinsic muscle atrophy or the
high stimulation threshold below the elbow.
134
A Case of Intestinal Angina with Obstructive Hypertrophic
Cardiomyopathy
PG-45
Toshihiko Hamada 1, Hiroyasu Uzui2, Yuka Otake1, Yumiko Tsuda1, Norikazu Hashimoto1,
Kenichirou Arakawa2, Hiroshi Tada2, Hideki Kimura1
Aya Ikeda , Takuto Hamaoka, Noriko Kawai, Wataru Omi, Yoshiteru Sekiguchi, Masako Kobayashi
Kanazawa Municipal Hosp., Japan
1 Department
of Clinical Laboratory Science, University of Fukui, Japan
2 Department
of Cardiovascular Medicine, University of Fukui
An 83-year-old asymptomatic female patient of
Atrioventricular Septal Defect
Symposium
PG-47
Educational Lecture
A case of the atrium thrombus was difficult to visualize in
transthoracic echocardiography
Special Lecture
Background:The pathology of cardiac Fabry disease primarily involves
left ventricular hypertrophy, and reductions in cardiac function and progression of fibrosis are observed. Recently, cases have been diagnosed
before reductions in cardiac function are seen based on factors such
as family history and measurement of alfa-galactosidase A activity. We
herein initiated enzyme replacement therapy for two patients with normal
cardiac function, monitored the course of cardiac function over time, and
investigated the usefulness of myocardial strain assessment by two dimensional (2D) speckle tracking echocardiography.Case presentation:We
performed enzyme replacement therapy for a 35-year-old man (Case 1)
and a 27-year-old man (Case 2) with Fabry disease, and subsequently performed conventional echocardiography over time. Over approximately the
same period, gadolinium-enhanced magnetic resonance imaging (MRI) for
assessing myocardial fibrosis and 2D myocardial strain assessment were
performed. At the time of initiation of enzyme replacement therapy, both
patients had no abnormalities on any of the tests. In the sixth year, MRI
showed no fibrosis, and conventional echocardiography showed no reduction in wall motion or left ventricular wall thickening. On the other hand,
2D myocardial strain assessment showed a reduction in the longitudinal
strain of the base-lateral wall ( ＜ -15%). In addition, in Case 1 a reduction
in the post-systolic strain in the base-lateral wall region was observed.
Conclusion:Measurement of the longitudinal strain using 2D speckle
tracking enabled early detection of cardiac function abnormalities in patients with Fabry disease who had not yet developed cardiac hypertrophy
or fibrosis. These findings suggest the importance of adding 2D speckle
tracking to conventional echocardiography in monitoring the course of
cardiac function in patients with Fabry disease.
Invited Lecture
An 86 years old man with obstructive hypertrophic cardiomyopathy
(HOCM) was admitted to our hospital with postprandial abdominal pain
and anorexia. We suspected intestinal angina. Computed tomography
showed mild stenosis of the celiac artery and severe stenosis of the inferior mesenteric artery (IMA). An ECG revealed bradycardia (40 bpm) due
to advanced AV block and left high voltage with ST-T change. Transthoracic echocardiography showed obstruction of the left ventricular outflow
tract (LVOT) and severe mitral regurgitation with systolic anterior motion
(SAM) of the anterior mitral leaflet. After pacemaker implantation (right
ventricular apical pacing), the pacing AV delay was optimized for minimum LVOT pressure gradient under the echocardiographic estimation (AV
delay 100ms for LVOT, 14.6mmHg). The patient became free of abdominal symptoms after these procedures. This case suggests that systemic
hemodynamic change could contribute significantly to intestinal perfusion,
despite mild stenosis of the celiac and mesenteric arteries.
PG-46
Early detection of a latent cardiac dysfunction of Fabry
disease by 2D speckle tracking
Keynote Speech
PG-44
Yuki Higuchi 1, Hiromi Umeda1, Tamami Kudo1, Kuninori Sugita1, Nobuyuki Sadasue1, Takako Karube1,
Akihiro Isotani2, Harushi Niu1
Yurina Kato , Akira Yabuki, Saori Abe
South Miyagi Medical Center, Japan
Poster Presentation
135
Oral Presentation
Endocardial cushion defect(ECD), also called atrioventricular septal
defect(AVSD), has common features; absence of muscular AV septum;
inlet/outlet disproportion; abnormal lateral rotation of the posteromedial
papillary muscle; and the abnormal configuration of the AV valves. AVSD is
diagnosed in 1.5 to 4.0% of patients with congenital cardiac anomaly. Patients may have no symptoms until their third or fourth decade, however,
almost all of them are followed by progressive symptoms of congestive
heart failure, atrial arrhythmias, complete heart block and pulmonary hypertension after fifth decade.We report a rare case of a high aged woman
with incomplete AVSD who had only a very mild symptom. An 83 year-oldwoman was diagnosed to have asymptomatic atrial fibrillation and cardiomegaly in a primary care hospital in 2011 and was referred to our hospital for further workup.Transthoracic echocardiography(TTE) visualized a
huge ostium primum atrial septal defect(ASD) of 43mm, membranous ventricular septal defect of 3mm, mital cleft with moderate to severe mitral
regurgitation and severe tricuspid regurgitation with severe pulmonaly
hypertension(PH) of 86mmHg. Estimated Qp/Qs was 3.6. Right atrium and
ventricle were markedly enlarged. Left ventricle(LV) had normal systolic
function, however, the LV showed D-shape deformity during a whole cardiac cycle.She had no dyspnea on effort, no shortness of breath and no palpitation regardless of huge ostium primum ASD with significantly elevated
Qp/Qs and moderate to severe AV valve insufficiency with significant PH.
Despite severe cardiac condition, she lives in the eighth decade with no or
very mild symptom. She didn't agree with surgical repair because those
cardiac abnormalities didn't restrict her daily work in her farm. Instead,
she comes to our hospital for annual follow up. TTE was effective for not
only the diagnosis but also for the regular follow up of this complex congenital heart disease.
Case Conference
1 Kokura
Memorial Hospital, Japan
2 Kokura
Memorial Hospital Cardiovascular Medicine
A man in his sixties with a chronic heart failure caused by atrium fibrillation was admitted to our hospital owing to a diagnosis of cerebral
infarction by brain MRI at another hospital. He received a diagnosis of
cardioembolic ischemic stroke because of atrial fibrillation. Transthoracic
echocardiography (TTE) showed the left atrial dilatation and revealed that
the inferior wall of the apex is moderate hypokinesis, but did not show
thrombus and any embolus source. Transesophageal echocardiography
(TEE) revealed a lesion like tumor (34.8 × 23.3mm) on esophagus side of
the left atrium, but we could not diagnose it as myxoma or thrombus. It
was thought an operation is necessary in either case. He was operated at
another hospital. There was a 4cm size of white thrombus on esophagus
side of the left atrium. It was a thrombus in pathological findings too.It
was difficult to find the thrombus in TTE, because it is the farthest from
chest wall. We became able to visualize a lesion because observed it by
TEE nearby.TTE has limitations, so we should carry out TEE. Therefore we
can find an embolus source easily.
Keynote Speech
PG-48
A case of shosin beriberi whole hemodynamics could be
assessed by echocardiography
Case Report
PG-49
The pericarditis patient who showed exacerbation and
remission in a short period: a case report
Natsuki Nakagawa 1, Ayano Hashizume1, Ayaka Araki1, Hiroomi Shimotsuka1, Tsutomu Sakamoto1,
Hiroyuki Ihori2, Tomoki Kameyama2, Hiroshi Inoue2
Tatsuya Yoshinouchi 1, Kanako Imamura1, Satoko Anai1, Yuki Goto1, Hisayo Yasuda1,
Nobuhiro Nakanishi2, Katsuyoshi Ikeda1, Hirotaka Matsui1
1 Dept
of Clinical Laboratory, Social Welfare Organaization Saiseikai Imperial Gift Foundation,Inc.Saiseikai Toyama Hosp.,
Toyama, Japan
2 Dept
of Internal Medicine, Social Welfare Organaization Saiseikai Imperial Gift Foundation, Inc. Saiseikai Toyama Hosp.,
Toyama, Japan.
1 Dept
of central clinical laboratory medicine, Kumamoto univ. hosp., Japan
2 Dept
of cardiovascular medicine, Kumamoto univ. hosp.
Invited Lecture
Special Lecture
Educational Lecture
Background: We have recently encountered a patient with pericarditis who
showed repeated exacerbations and remissions within one month, who
was followed-up by echocardiography.Case report: A 62-year-old woman
who had cough, a difficulty in breathing and a chest pain at inspiration
had been referred to our hospital. The patient showed a pericardial hypertrophy and brightness and respiratory fluctuations of ventricular inflow
in the echocardiogram, which initially led us to suspect of the constrictive pericarditis. However, the patient also showed higher inflammatory
markers such as CRP, WBC and BNP in the blood testing in addition to the
pericardial hypertrophy and effusion by CT and MRI examinations, thus
leading us to make a diagnosis of acute pericarditis. Actually her chest
pain was relieved together with the improvement of inflammatory markers
after the initiation of the treatment. However, thereafter the symptoms developed again and fluctuated during a short period. Discussion: While the
case was initially suspected of acute pericarditis from the clinical symptoms and signs of acute inflammation, it could not be ruled out of constrictive pericarditis from the findings in the echocardiogram examinations,
in which the patient showed pericardial hypertrophy and brightness from
the beginning. Overall, it was finally concluded that the patient had been
suffering from recurrent pericarditis accompanied by signs of constructive pericarditis. Intriguingly, the pericardial thickness was well correlated
with the clinical symptoms and inflammatory markers during the clinical
course. We learned from this case that there might be recurrent pericarditis patients with the features of constrictive pericarditis who could be improved by proper treatments. Thus careful inspections and monitoring by
echocardiography are clearly necessary in all the patients who are initially
diagnosed with constrictive pericarditis.
A sixty-five-year-old man was admitted to another hospital, because he
fell down after drinking the previous day. In the next morning, he became
shock (systolic blood pressure 70 mmHg), and then was transferred to our
hospital. He had suffered alcoholic liver disease. On admission, his consciousness was alert, and he had hypotension (70/44 mmHg), anemia (Hb
6 g/dl), hyperkalemia (6.26 mEq/L), and metabolic acidosis (pH 6.762, PCO2
27.7 mmHg, HCO3 4 mmol/L, anion gap 28). Echocardiography showed
increased left ventricular contraction (ejection fraction 88.8%). Initially, we
suspected he had hemorrhagic shock, but any source of bleeding was not
detected. After moved to the ward, he developed cardiopulmonary arrest.
Cardiopulmonary resuscitation (CPR) with chest compression, intravenous
adrenaline injection and blood transfusion were started. CPR was temporarily effective, but, cardiac arrest recurred repeatedly. His hemodynamics
did not improve with blood transfusion or catecholamine infusion. After
reviewing his history of alcohol drinking, his shock was attributed to shoshin beriberi, a fluminant form of cardiovascular beriberi, caused by vitamin B1 deficiency due to alcohol abuse. After starting a bolus intravenous
injection of thiamine, his hemodynamics was improved immediately and
dramatically. Serial echocardiography showed that ejection fraction, cardiac output, and estimated right ventricular systolic pressure decreased.
Acidosis and hyperkalemia were also improved. We report a rare case of
shosin beriberi whose hemodynamics could be assessed serially by echocardiography.
Symposium
PG-50
A case of Giant coronary artery aneurysms exceeding 5 cm
in size
PG-51
A case of AA amyloidosis with Castlemans disease that
showed reversible ventricular hypertrophy
Case Conference
Chiho Ogawa, Hiroki Usuku, Hisayo Yasuda, Kanako Imamura, Yuki Goto, Satoko Anai,
Katsuyoshi Ikeda, Hirotaka Matsui
Imamura Kanako 1, Yasuda Hisayo1, Horibata Yoko2, Kojima Sunao1, Goto Yuki1, Anai Satoko1,
Ikeda Katuyoshi1, Matsui Hiotaka1
Kumamoto University Hospital, Japan
1 Kumamoto
University Hospital, Japan
2 Saiseikai
Kumamoto Hospital
Oral Presentation
Poster Presentation
Introduction: Giant coronary artery aneurysms (gCAAs) with a diameter
exceeding 50mm are extremely rare and there had been few such reports.
Here we report an extremely rare case of gCAA that was evaluated by the
echocardiogram.Case Report: A 74-year-old man, who had a past history
of hypertension and dyslipidemia, was admitted to a primary hospital
because of a chest pain, and was diagnosed as myocardial infarction with
ST elevation (STEMI) at the inferior wall, based on biochemical blood examination and ECG. As the coronary angiography and other examinations
revealed a complicated coronary lesions and a severe myocardial dysfunction in the patient, he was referred to our hospital for further interventions.The echocardiography in our hospital showed that his cardiac function was severely impaired (Ejection Fraction = 20%), and that there was a
mass lesion (74 × 79mm) excluding right atrium, right ventricle and the
tricuspid valve. In addition, the CT examinations revealed that the mass
was ruptured and perforated into the right atrium.These findings led us to
diagnose that the patient had a gCAA that gave rise to the arteriorrhexis
and STEMI. The gCAA, which was excised by the emergency surgery, was
rich with organized thrombi.Discussion: While there were no past history
of cardiac trauma, collagen diseases, coronary artery dissociation, or coronary artery fistula, it might be possible that the gCAA in this patient was
caused by Kawasaki disease, because it has been reported that the disease
promotes arteriosclerosis. Actually, the blood vessels close to the gCAA in
the patient was full of such lesions.Conclusion: The echocardiography is
highly useful for non-invasive morphological and functional evaluation,
and enables us to make a proper decision of treatment strategy.
Here we report case of AA amyloidosis who showed a good progress
through an early diagnosis of the left ventricular wall thickening under
the echocardiography.BackgroundCastleman’s disease is known as not
only one of the lymphoproliferative diseases characterized by hyperplasia of lymphoid follicles but also as an underlying disease of secondary
amyloidosis: AA amyloidosis.We report an AA amyloidosis patient with
Castleman’s disease who was followed up for a long period, mainly by
the echocardiography. Case reportA 53 year old man, who had an attack
of unconsciousness, was diagnosed as sick sinus syndrome. He had been
pointed out by echocardiography that he had a left ventricular hypertrophy (17mm), and showed a markedly higher level of serum amyloid A
(SAA) protein (429µg/mL). In addition, the patient also had retroperitoneal
masses and depositions of AA amyloid in the duodenum. Pathological
examinations of the excised retroperitoneal tumor revealed AA amyloidosis complicated by Castleman’s disease in the patient. After a resection
of the retroperitoneal mass, SAA value was reduced markedly to 4.7µg/
mL, and left ventricular wall thickening was also improved from 17mm to
12mm.DiscussionIn AA amyloidosis with Castleman’s disease, it has been
reported that large amount of IL-6 produced by the swollen lymph nodes
promote SAA production in the liver. Actually, the left ventricular hypertrophy in this patient was markedly reduced after a resection of the enlarged
lymph nodes. Although this is a very rare case, it is important not to miss
reversible ventricular hypertrophy and to diagnose Castleman’s disease
correctly.
136
Usefulness of measuring fractional flow reserve in
determining renal artery stenosis due to fibromuscular
dysplasia
PG-53
Hiroki Kono 1, Naomi Bou1, Naoki Kawabata1, Kazuaki Shimizu2, Kanichi Otowa3, Kouichi Kifune4
Yuya Onozawa 1, Susumu Obata1, Shinichi Munekata1, Taira Toki2, Yutaka Nonoda2, Toshiyuki Iwasaki2,
Takahiro Iizuka3, Yuhsaku Kanoh4
1 Municipal
Tsuruga Hospital Medical Technology Department of laboratory, Japan
2 Municipal
Tsuruga Hosp Dept of kidney internal medicine
3 Municipal
Tsuruga Hosp Dept of Cardiology
4 Municipal
Tsuruga Hosp Dept of Radiology
1 Department
of Clinical Laboratory, Kitasato University Hospital, Japan
2 Department
of Pediatrics, Kitasato University School of Medicine
3 Department
of Neurology, Kitasato University School of Medicine
4 Department
of Laboratory Medicine, Kitasato University School of Medicine
Agreement rate of the sleep stage scoring in the PSG analysis
Sachiko Kurosaki, Yukio Yamadera, Ayumi Kikuchi, Chika Yasuda, Chiaki Suzuki, Naoko Sakurai,
Kyouko Kaneta
1 Hyogo
College Of Medicine Hospital, Japan
2 Hyogo
College Of Medicine
Ohta General Hospital Foundation Ohta Nishinouchi Hospital, Japan
Poster Presentation
137
Oral Presentation
Introduction:The sleep stage scoring of PSG becomes an important indicator to use for diagnosing sleep disorders and judging the course of treatment. However, as scoring is performed visually, the inter-scorer difference
may occur. This time we will report on our investigation of the agreement
rate and the factor that causes the inter-scorer difference by scoring the
arousal and sleep staging on a same cases by multiple technologists.
Subject･Methods:Six Medical technologists scored arousals and sleep
stages on five PSG data (A-E) and made a comparative review of the following viewpoints with reference to the leading technologist’s results;(1)
Agreement rate of the arousal scoring(2)Agreement rate of the sleep stage
scoring(3)Agreement rate by each sleep stagesResults:(1)The agreement
rate of the arousal scoring: Patient A=87.5%, B=95.5%, C=89.7%, D=91.2%,
E=89.5%, average=90.7 ± 3.0%.(2)The agreement rate of the sleep stage
scoring: Patient A=88.4%, B=84.0%, C=90.4%, D=95.4%, E=83.3%, average=88.3 ± 5.0%.(3)The agreement rate of the scoring by sleep stages:
Stage W=95.1 ± 4.8%, Stage N1=82.6 ± 2.1%, Stage N2=87.6 ± 6.4%,
Stage N3=70.4 ± 21.0%, Stage R=89.4 ± 8.8%.Conclusion:Agreement
rate were high in both arousal and sleep stage scorings. Regarding the
agreement rate by each sleep stages, they were high in the order of Stage
W ＞ R ＞ N2 ＞ N1 ＞ N3.Regarding Stage N3 that was the lowest agreement rate, mix of artifact due to the sweating was particularly the factor
that caused disagreement.We will push forward the approximation of the
disagreement part within the scorers and, at the same time, we think it is
important to set a rule for the bad quality signal recording cases.
Introduction: Electroencephalogram (EEG) is not routinely performed on
the patients with possible dementia, although this test is not invasive and
relatively easy to perform. Since it is recently suggested that the epilepsy
patients with memory disturbance and amnesia be misdiagnosed as dementia, we conducted a study to evaluate the validity of EEG toward the
patients with possible dementia.Materials and Results: The study was
conducted on EEGs of 70 patients (29 males and 41 females, age 53-94)
who visited our hospital or Medical Center for Dementia from January,
2013 to December, 2015, having some episodes of memory disturbance
and amnesia. Abnormal findings were detected in 26 cases (37 %), most of
which were slightly low background activities, slow Α , and/or θ waves.
It should be noted that one patient with spike was diagnosed as having
localization-related epilepsy, and another case with periodic synchronous
discharges (PSD) was diagnosed as Creutzfeldt-Jakob disease. Discussion:
Low background activity, slow Α and θ waves represent the reduction in
general brain function, which can be found on the condition of dementia.
In our study, however, 3 % of the patients (= 2/70) suffered from the diseases other than dementia. Focused on our Medical Center for Dementia,
approximately 420 patients visited the Center during the study period, and
only 47 cases had their EEG examined. Our study revealed that the conditions other than dementia such as epilepsy and encephalopathy might
be overlooked when patients have symptoms that are consistent with the
diagnosis of dementia. Therefore it is concluded that the EEG examination is useful and should be performed for the detection and differential
diagnosis of the diseases on patients showing the symptoms suggestive of
dementia.
Case Conference
Hiromi Minato 1, Saori Shibayama1, Noriko Hatakeda1, Ayumi Igaki1, Yasunao Wada1, Koji Inuzumi1,
Masanaka Takeda2, Masahiro Kosiba2
Symposium
PG-55
Educational Lecture
Evaluation of the validity of EEG toward the patients with
possible dementia
Special Lecture
Objective: To report a case of acute necrotizing encephalopathy (ANE), in
which evoked potentials studies were useful in the prediction of functional
outcome. Backgrounds: ANE is a rare severe form of acute encephalopathy
characterized by bilateral thalamic lesions. ANE mainly affects children
in Asia and Western countries, with an estimated mortality rate of 30%.
Methods: A case report. Results: A 7-year-old boy was admitted to our
hospital in Dec. 2013 with status epilepticus due to influenza B infection. He was initially admitted to another hospital, and was treated with
intravenous peramivir hydrate. However, he had convulsive seizures in
the next day, and he was transferred to our hospital. On arrival (day 1),
he was in coma; with a temperature of 40.6°C, blood pressure at 68/30
mmHg, a pulse of 198 bpm, and the oxygen saturation at 70%. Blood-test
results showed metabolic acidosis, leukocytosis, DIC, increased CK, mild
hypoglycemia, and renal dysfunction. CSF examination showed a few cells
(WBC 7/ Μ L), with mildly elevated protein (51 mg/dl), and normal glucose. Brain CT showed bilateral thalamic lesions. The patient was treated
with therapeutic hypothermia and CHDF on mechanical ventilatory, and
pulse therapy of methylprednisolone from day 1. Brain MRI on day 4
showed multifocal T2/FLAIR high signals. After gradual recovery from hypothermia, he remained in coma. An EEG obtained on day 8 showed burstsuppression pattern. However, evoked potential studies with ABR on day 8,
VEP on day 17, and SSEP on day 18 showed no abnormalities. Following
the treatment he improved gradually and was transferred to a rehabilitation hospital on day 48. He returned to school at 4 months after presentation. At the last follow-up (26 months after presentation), the pediatric
cerebral performance category (PCPC) was scored 2.Conclusion: Evoked
potentials studies are useful in the prediction of functional outcome in
ANE.
Invited Lecture
Background: Fractional flow reserve (FFR) determination is a technique
used in coronary catheterization to measure pressure differences across a
stenotic coronary artery, to determine the likelihood of the stenosis impeding oxygen delivery to the heart muscle. Fibromuscular dysplasia (FMD) is
a non-atherosclerotic, non-inflammatory disease of the blood vessels that
causes abnormal growth within the walls of an artery. FMD is frequent
in middle-aged women where it affects any arterial structure in the body.
I applied FFR to a case of renal artery stenosis that assumed FMD as the
diagnosis.Case presentation: The patient was a 49-year-old woman with
a history of hypertension. Sonography revealed a fast PSV in both renal
arteries (PSV232cm/s,PG22mmHg; left:PSV171cm/s,PG12mmHg). Angiography revealed a coffee bean-like appearance on both sides of the intermediate portion of the renal artery.The pressure of the surrounding renal
vessels was measured after expansion with hydrochloric acid papaverine
using a pressure wire (left ventral branch: FFR 0.86/PG30mmHg; left dorsal branch: FFR 0.84/PG30mmHg; right dorsal branch: FFR 0.70/PG55mmHg; right ventral branch upper pole: FFR 0.88/PG15mmHg; lower pole:
FFR 0.86/PG25mmHg).Pressures obtained were mostly in the upper limit,
with the right dorsal branch having the highest measurement, more than
55mmHg (PG comparison sonography/FFR=22mmHg/55mmHg; Stenosis
degree comparison: angiography/FFR=50 – 75%/0.70).Conclusion: The severity of a lesion was altitude FFR when I evaluated the stenosis degree in
each parameter. Like the disorder that included stenosis at multiple points,
with FFR, I could evaluate the pressure range of the culprit lesion selectively. Fixed quantity can evaluate lesion disease severity for cases in which
visual stenosis degree scoring by angiography is difficult, like the disorder
by using an FFR level. Using pressure wire FFR of the fixed-quantity rating
system, the disease severity of stenotic lesions can be judged precisely.
PG-54
Evoked potentials may predict of functional outcome in a case of acute necrotizing encephalopathy
Clinical utility of evoked potentials to predict of functional outcome in a case of acute necrotizing
encephalopathy
Keynote Speech
PG-52
Keynote Speech
PG-56
Cancer assosiated thrombosis
The incidence of complication of DVT according to a type of
cancer
PG-57
Electrocardiography scoring is useful in predicting left
ventricular wall motion abnormality after subarachnoid
hemorrhage
Harumi Ueda , Chikako Ogino, Hiroko Noguchi, Satomi Isshiki, Hidemi Yasugi, Masanori Nakamura,
Akiko Nonaka
Keiko Sugimoto 1, Akira Yamada2, Risako Tanaka3, Ayako Takahashi3, Kazuhiro Nakamura3,
Kunihiko Sugimoto3, Joji Inamasu4, Tadayoshi Hata1
Hyogo Cancer Center, Japan
1 Fujita
Health University School of Health Scineces, Japan
2 Fujita
Health University School of Medicine Div. of Cardiology
3 Fujita
Health University Hospital Clinial Laboratory
4 Fujita
Health University School of Medicine Div. of Neurosurgery
Invited Lecture
Special Lecture
Educational Lecture
IntroductionDeep vein thrombosis (DVT) has been found to be an important and crititical complication of cancer patients. However ,there is scarce
data on the incidence of DVT. So we investigate the incidence of complication of DVT according to a type of cancer.Methods We investigated 423
patients who have a symptom likely to DVT, such as leg edema ,and/ or
D-dimer 1.0 Μ g/ml using by lower extremity ultrasound sonography
among 3427 cancer patients who were consulted to our center from November 2014 to October 2015. ResultsIncidences of DVT complication
according to cancer type were the peritoneal cancer (16.7%), soft tissue
tumor (14.3%), ovarian cancer (11.8%), malignant melanoma (7.8%),
multiple myeloma (7.1%), pancreatic cancer (6.1%), gastric cancer(5.6%),
colon and small intestine cancer (5.1%), bladder cancer (3.5%), lung cancer
(3%), prostate cancer (2.9%), endometrial cancer (2.4%), malignant lymphoma (1.7%), Pagets disease and Bowen's disease(1.5%), breast cancer
(1.5%), kidney cancer (1.3%), esophageal cancer (0.6%) , pharynx cancer
and larynx cancer (0.6%).The positive rate of DVT according to cancer
type were soft tissue tumors(50%),pancreatic cancer (42.9%), malignant
melanoma (40%), ovarian cancer (34.4%),multiple myeloma (33.3%), lung
cancer (33.3%), stomach cancer (31.7%), colon and small intestine cancer
(27.9 %), kidney cancer.Conclusion1 Peritoneal cancer, soft tissue tumors,
ovarian cancer have higher incidence of DVT complication, respectively
16.7%,14.3%,11.8%.2 Soft tissue tumors, pancreatic cancer, malignant
melanoma, ovarian cancer have higher positive rate of DVT, respectively
50%,42.9%,40%,34.4%.
Symposium
PG-58
Purpose: Cardiac wall motion abnormality (WMA) is a common complication in patients with subarachnoid hemorrhage (SAH), which is one of the
determinants of their prognosis. The aim of this study was to examine
whether electrocardiography (ECG) findings could predict WMA after SAH.
Subjects: We studied 212 patients with SAH who were hospitalized in our
institution between April 2007 and November 2010. We performed bedside 2-dimensional transthoracic echocardiography and 12-lead surface
ECG within 24 hours of hospital admission. Each of ECG changes as follows was scored 1 point: ST elevation, ST depression, T wave inversion, QT
prolongation. We summed up the points in every patient and compared
with WMA evaluated by echocardiography.Results: The study subjects
were classified into 2 groups based on the occurrence of WMA. Multivariate analysis revealed that ST elevation, ST depression and T wave inversion
were strong independent predictors of WMA, while QT prolongation was
a weak independent predictor. Receiver operating characteristics analysis
determined that the threshold value to predict WMA was 4 points (p ＜
0.0001, sensitivity 86.5%, specificity 83.1%, AUC 0.94).Conclusion: ECG
scoring was very helpful in predicting WMA after SAH. ECG score over 4
could predict the occurrence of WMA.
Prognostic Significance of Right/Left Atrial Volume Ratio by
Echocardiography
in Interstitial Lung Disease Patients with Pulmonary Hypertension
PG-59
Three Dimensional Left Atrial Speckle Tracking may detect
Cardiac Changes due to Chemotherapy by Trastuzumab
Case Conference
Naka Saito 1, Shingo Kato2, Ayumi Tanaka1, Takako Ishikawa1, Kozue Nakagomi1, Noritaka Saito1,
Mamiko Nakamura1, Kazuki Fukui3
Naoki Kimura , Yuji Masaki, Masashi Nagatomo, Minako Furukawa, Yasushi Kawabuchi,
Katsuyuki Nagatoya
1 Department
of Clinical Laboratory, Kanagawa Cardiovascular and Respiratory Center, Japan
2 Department
of Medical Science and Cardiorenal Medicine, Yokohama City University Hospital
3 Department
of Cardiology, Kanagawa Cardiovascular and Respiratory Center, Japan
Osaka Rosai Hospital, Japan
Oral Presentation
Poster Presentation
[Background]Drug-induced cardiomyopathy often occurs in patients
who undergo trastuzumab chemotherapy. Thus, it is important for these
patients to undergo periodic echocardiography. In the early phase of the
drug-induced cardiomyopathy, a diastolic disorder appears first, which
may be detected by a three dimensional left atrial speckle tracking echo.
[Methods]Using three dimensional echocardiography, we assessed 140
patients between January 2014 and June 2015. Patients were divided into
two groups: a post chemotherapy trastuzumab group (12 patients, age:
65 ± 5.8 years) and a control group without heart disease (21 patients,
age: 73 ± 6.5 years). We analyzed the three dimensional left atrial speckle
tracking: left atrial global longitudinal strain(LA GLS), left atrial global
circumferential strain (LA GCS), and the left atrial global radial strain (LA
GRS). Using these three indices with the diastolic index (E/A) and systolic
index (LVEF), we were able to compare the differences between the two
groups.[Results]Significant differences were observed for LA GLS (chemo.
vs. cont. 21.8 ± 6.1 vs. 16.8 ± 4.7, p ＜ 0.05) and E/A (0.84 ± 0.21 vs.
0.68 ± 0.19, p ＜ 0.05). No significant differences were observed for LA
GCS (27.32 ± 21.6 vs. 17.9 ± 10.9, p = 0.08), LA GRS ( – 23.9 ± 12.7 vs.
?19.2 ± 7.4, p = 0.14), and LVEF (70.1 ± 3.2 vs. 69.1 ± 5.5, p = 0.26).
[Conclusion]The post chemotherapy group showed changes in the three
dimensional LA GLS and E/A. This suggests that LA GLS may be useful in
detecting the cardiac changes due to chemotherapy by trastuzumab.
Intoduction Pulmonary hypertension (PH) is a well-known prognostic
factor for interstitial lung disease (ILD) patients. Primary cause of PH is
usually pre-capillary depending on pulmonary vascular disease, resulting
in increased right atrial (RA) dilatation. On the other hand, post-capillary
PH is associated with left heart disease and accompanied with left atrial
(LA) dilatation. Recent studies have shown that right/left atrial volume
ratio (AVR) is useful for differentiate pre- and post-capillary PH. The aim
of this study was to assess the prognostic value of echocardiography derived AVR in ILD patients with PH. Methods We performed a retrospective
cohort study of 103 patients with ILD, who underwent echocardiography
and right heart catheterization. By echocardiography, AVR was calculated
by using a formula that RA volume divided by LA volume. We defined PH
as mean pulmonary artery pressure ≥ 25mmHg by right heart catheterization. Patients were allocated to ILD with PH group and those without (nonPH group). Furthermore, ILD with PH group divided into PH-AVR ≥ 1.0
group and PH-AVR ＜ 1.0 group. Results Of 103 ILD patients, 79 (77%)
patients were without PH, 14 (14%) patients were PH-AVR ≥ 1.0, and 10
(9%) patients were PH-AVR ＜ 1.0. At a mean follow-up of 731 days, 9 of
24 (38%) ILD patients with PH died. In ILD patients with PH, AVR ＜ 1.0
was associated with a ＞ 6-fold hazards increase for death (hazard ratio:
6.57; p=0.02). Kaplan-Meier curves showed that death rate was significantly increased in PH-AVR ＜ 1.0 group compared with non-PH group (p
＜ 0.001) and PH-AVR ≥ 1.0 group (p ＜ 0.01). Conclusion In ILD patients
with PH, lower right/left atrial volume ratio was associated with higher
risk of death. The results indicated that assessment of post-capillary PH is
important for the risk stratification in ILD patients with PH.
138
Phagocytosis Phenomenon in Urinary Tract of a PoorGlycemic Control Patient
A Case Report and Survey
PH-02
Quality Improvement of Image-Based Automatic Urinalysis
A Two-PDCA-Cycle Experience
A 73-year-old man with previous medical history of hypertension, hyperlipidemia, type II DM sent to the emergency department due to general
muscle weakness and drowsy consciousness for one day after head collided in bicycle accident. After examinations, he was finally diagnosed as
acute right centrum semionale infarction, urinary tract infection (UTI) and
chronic kidney disease (CKD). The biochemistry data were glucose 434
mg/dl, BUN 19 mg/dl, creatinine 1.53 mg/dl, eGFR 45 ml/min/1.73 m2,
Na 138 mmole/L, K 3.4 mmole/L and CRP 3.01 mg/dl without any viral
infection tests. The urinalysis data were glucose 1.0 g/dl, OB 3+, protein
30 mg/dl, nitrite +, leukocyte esterase 2+, RBC 11~20/HPF, WBC 1+/HPF,
atypical cell 0~2/HPF, renal tubular cell 0~2/HPF and bacteria many as
normal floral cultured. Those atypical cells refered to cytoplasmic inclusion body bearing cells or phagocytosing cells that were seen in almost
every HPF. Meanwhile there were 219 cases of cytoplasmic inclusion
body in 11,958 urinalysis specimens (OPD 6,741, IPD 3,454, ER 1,763)
we reviewed within 30 days. The average present rate is 1.83% which is
higher in ER specimens 2.78% and that is highly relative to protein, OB/
RBC, nitrite, leukocyte esterase/WBC, bacteria/yeast (P ＜ 0.001). Analyzing the 219 cases versus the results of urinalysis, positive rate of OB/RBC,
leukocyte esterase/WBC, bacteria are 79.5%, 80.8% and 63.5%. The survey
reveals the presence of cytoplasmic inclusion body is relative to urinary
tract bleeding or immune activity though viral infection is thought as the
main reason. DM and CKD patients are immunocompromised and have
high risk to infection. The muddling immune status in urinary tract of this
subject patient formed numerous cytoplasmic inclusion body bearing cells
or phagocytosing cells. We present this case to highlight that the presence
of cytoplasmic inclusion body is much useful in diagnosis of urinary tract
diseases.
Since microscopic examination of urine sediment is labor-intensive, timeconsuming, and has wide inter-observer variability, two image-based
automatic analyzers [USCANNER(E), TOYOBO] and auto-verification using
8 rules were introduced to our laboratory in February 2014. The 8 intercept rules were OB ＞ 1+, LEU ＞ 1+, cast ＞ 1-2/LPF, crystal ＞ 0-5/LPF,
renal tubular epithelial cell ＞ 1/HPF, urothelial cell ＞ 1/HPF, OB -/RBC
＞ 3-5/HPF, and LEU -/WBC ＞ 6-10/HPF. In this phase, we performed
199,532 specimens (OPD 135,159, IPD 64,373) with 2.5 staff (3 ones
in 2013). The complete rate of OPD/IPD specimens in 30 min-TAT, 60
min-TAT, 120 min-TAT were 70.1%/76.5%, 96.3%/96.7%, 99.7%/99.8%
that were comparable to the performance of 2013. But the reports didn't
include the findings of renal tubular cell (RTE), urothelial cell (URO), inclusion body or oval fat body (OFB) until January 2015 formally. After oneyear-training, the second phase, those cells were reported with manual
microscope confirmed and meanwhile the specimens from emergency
department (ED) were transferred to the identical system. In 2015, we had
201,735 specimens (OPD 115,268, IPD 59,941, ED 26,526) and reported
RTE 16,330 cases (8.09%), URO 6,377cases (3.16%), inclusion body 3,695
cases (1.83%), OFB 1,097 cases (0.54%). Besides, the complete rate of
OPD/IPD/ER specimens in 30 min-TAT, 60-min TAT, 120 min-TAT were
80.2%/85.7%/97.8%, 99.4%/98.5%/100%, 100%/100%/100% that were
even better than those in 2014 and in accordance with the CLSI GP16-A3
or European Urinalysis Guideline. The auto-verification rate in specimens
of OPD, IPD, ED are 54.4%, 52.1%, 32.8%. We also found the sensitivity,
specificity, positive predict value, negative predict value of the analyzer
in RTE are 96.4%, 81.0%, 30.9%, 99.6% that RTE is closely relative to the
renal function. With such examination system and improvements, we not
only save the manpower but also have enough time to identify more cells
in order to achieve better quality and provide more valuable clinical information.
Urinary protein analysis using cellulose acetate electrophoresis
in the case of a negative or (+/-) result of the method of urinary
protein test paper
PH-04
Classification of uropathogens using Sysmex UF-1000i contributes to empiric therapy of urinary tract infection
Application of bacteria morphology parameters on rapid diagnosis and antibiotic choose prior to urine culture
The method of urinary protein test paper has been used as a screening
test for kidney disease. There may be an early stage of kidney disease
and mild kidney injury even in the case of a negative or (+/-) result of this
assay. We aimed to analyze urinary protein in cases of negative or (+/-) results on urinary protein test paper using cellulose acetate electrophoresis.
Urine samples were obtained from 40 patients who were being monitored in the Department of Pediatrics, Nippon Medical School Hospital.
The results of analysis of urinary protein test paper in these patients were
negative or (+/-). Analysis of urinary proteins was based on cellulose acetate electrophoresis. After electrophoresis, the proteins were stained with
a silver staining solution. The protein fractions from these patients were
analyzed in urinary protein analysis software. This software enables classification of urinary protein patterns into glomerular, tubular, and mixed using relative mobility of each band during cellulose acetate electrophoresis
of urine samples from patients.
The glomerular pattern was observed in 9 (22.5%) cases, tubular pattern
in 8 (20.0%), mixed pattern in 1 (2.5%), and other patterns in 22 cases
(55.0%). This urinary protein analysis software allowed us to identify the
type of kidney injury even in patients with a negative or (+/-) result on urinary protein test paper.
Analysis of urinary proteins using cellulose acetate electrophoresis may
be useful for early detection of damage sites in kidneys.
Background: Urinary tract infection (UTI) is a common infection disease
in Taiwan. Urine cultures are still considered as gold standard for clinical
diagnosis of UTI patients. However, culturing of the samples is both time
and labor consuming, with most of samples yielding no growth. Besides,
clinicians usually start empiric therapy with antibiotic agents mainly
against Gram negative bacteria, but they are not very effective against
Gram negative bacteria. A reliable screening tool would be very important
for clinicians and laboratories.
Material and methods: We collected 1053 urine samples in this study.
Bacteria amount (/uL) and morphology information (Rods? or cocci/
mixed?) were obtained from a Sysmex UF-1000i analyzer. These data was
compared to the results of urine culture, containing bacteria quantification
and identification (ID). Antibiotics susceptibility tests (AST) were further
collected for understanding the effect of antibiotic drugs in UTI patients.
Results: The criteria of urine culture is 10^5 CFU/mL. In our data, a cutoff point of 100 bacteria/uL from Sysmex UF-1000i analyzer was used,
in which the sensitivity and NPV were 90.9% and 93.8%. Comparison of
bacteria morphology information from UF-1000i with ID results, the specificity and PPV were both 100% for identification of rods-group uropathogens. According to the AST data, three first-line antibiotic drugs, cefazolin,
gentamycin and colistin, had higher susceptible for patients with rods
groups, but only colistin can be used for the patients with non-rods group.
Conclusion: Sysmex UF-1000i analyzer is a reliable screening tool for ruleout of UTI. It also can specifically identify the bacteria with rods morphology (mainly gram negative bacilli) to provide advanced information for
empiric therapy before complete of urine culture reports.
139
Poster Presentation
Department of Pathology, National Defense Medical Center, Division of Clinical Pathology,
Tri-Service General Hospital, Taipei, Taiwan
Oral Presentation
Ling-Chiao Tsai, Pin-Ching Pan, Jin-Biou Chang, Tzong-Shi Chiueh
Case Conference
Mihoko Nishizawa1, Ryo Kubota1, Toru Igarashi2, Nobue Sakai1
1 Graduate
Course of Health and Social Services, Saitama Prefectural University, Japan
2 Department
of Pediatrics, Nippon Medical School Hospital
Symposium
PH-03
Educational Lecture
Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taiwan
Special Lecture
Hui-Szu Tsai, Sheng-Hsiu Wei, Chuan-Po Lee, Ya-Chin Li
Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taiwan
Invited Lecture
Hui-Szu Tsai, Chuan-Po Lee, Ya-Chin Li, Hsiu-Chin Fan
Keynote Speech
PH-01
Keynote Speech
PH-05
Development of cellulose acetate membrane electrophoresis
based urinary proteomics
Combination of old and new methodology
PH-06
The Coexisting of Renal Tubular Epithelial Cells and Cystine
Crystals in Acetylcysteine Dysmetabolism Case
A Case Report and Survey
Aki Nakayama Howley, Mai Kimino, Kiyoko Shiba, Shiro Iijima
Yen-Li Wang, Chuan-Po Lee, Ya-Chin Li, Hui-Szu Tsai, Hsiu-Chin Fan
Bunkyo Gakuin University, Japan
Division of General Laboratory, Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taiwan,
R.O.C.
Invited Lecture
Introduction: Development of non-invasive urine based tests would be
a tremendous benefit to patients with renal diseases. A urinary protein
panel, which is a renal disease assessment involving several novel protein
markers increases the diagnostic accuracy. Cellulose acetate membrane
electrophoresis (CAME) coupled with highly sensitive colloidal silver staining is one of the useful methods in analyzing an entire scope of urinary
protein abnormalities. In this study, we developed a CAME-based proteome
analysis strategy to increase utility urinary protein fraction on CAME.
Special Lecture
Educational Lecture
The 86-year-old male, who had history of liver cirrhosis, irritable bowel
syndrome and gastric ulcer, was brought to our emergency room (ER).
Urinalysis and biochemistry were normal except CRP evaluated (4.05 mg/
dl); renal tubular epithelial cells (6-10/HPF) and cystine crystals were
presented in urine sediment. Urinary cystine crystals occur rarely, usually
found in cystinosis, cystinuria, Fanconi syndrome and other congenital disorders of metabolism. In this case, the patient used acetylcysteine (NAC) as
a mucolytic agent led to cystine crystals occurrence in urine. High concentrated cystine in urine may deposit as crystals in renal tubular epithelial
cells because of most of the cystine metabolic insufficiency or glomerular
malabsorption of cystine, and then causing cystine stones. Therefore, we
could find many cystine crystals accompanied by renal tubular epithelial
cells falling off in urine sediment. It could be used as a reference to acetylcysteine dosage. We should take renal malfunction and kidney stones into
consideration. Renal tubular epithelial cells are extremely sensitive to high
nephrotoxic drugs. Those will be impaired after exposing to high nephrotoxic renal filtrate and shed within 2 hours, then return to normal after 24
hours. In our hospital, there are over 150,000 urinalysis specimens a year
in average. We analyzed 11,958 specimens of 2016/02 and 968 renal
tubular epithelial cells cases (8.09%) were noted. We found renal tubular
epithelial cells were related to OB, PRO, LEU, RBC and WBC in urine (p ＜
0.05). In addition, the present rate of specimens in OPD, IPD and ER were
6.19% (417/6,741), 9.53% (329/3,454) and 12.59% (222/1,763). It was
very interesting that the highest present rate was in ER specimens and it
was because of higher pressure. Before abnormal warnings of blood biochemical examination, renal tubular epithelial cells that are shedding in
urine can be used as an important indicator of early kidney damage.
Methods: A urine sample loaded with ten lanes on CAME. After electrophoresis, both ends were cut and stained with colloidal silver. The unstained
lane number two to nine were then cut out around the stained area. The
membrane strips were further cut into smaller segments and then incubated with a SDS PAGE sample preparation buffer to extract proteins in each
fraction. Protein identification was performed by SDS PAGE combined with
in-gel digestion and mass spectrometry.
Results: Total 31 proteins were identified in the patients with tubulointerstitial nephritis, including 20 urinary proteins that were newly identified
in the present CAME-based proteome analysis. This included beta-2-glycoprotein and alpha-1-B-glycoprotein, candidate markers of exacerbation of
renal function.
Conclusion: Combining the conventional method of CAME and the relatively new methodology of proteome analysis enabled us to identify the
biomarkers of renal disease. These results increase the utility of urinary
protein fraction using CAME as a diagnostic tool in clinical laboratory.
Symposium
PH-07
Influence of Vitamin C on Urine Dipstick Test Results
The utility of a urine strip with a vitamin C indicator
PH-08
Kyoung A Kim1, Yong Lim2
Evaluation of the association between Light's criteria and the
microscopic test in pleural effusion
Hizuru Hoshina 1, Chika Miyasaka2, Miki Hayashi2, Miho Kasai2, Toshiyuki Habara1, Noriyuki Ozeki1,
Masaru Ozawa1, Shigeharu Okada1
Case Conference
1 Haedong
Hospital, Korea
2 Dong
Eui Univerisity
1 The
JAMT committee for standardization of Body Fluid Analysis, Japan
2 Suwa
Central Hosp.
Oral Presentation
Poster Presentation
Vitamin C is a strong reducing agent found at high levels in various foods,
and it may influence the results of urine strip tests even at an ordinary
consumption levels. After oral administration, we measured urine vitamin
C levels using urine strips and evaluated whether vitamin C interfered with
various test items. The utility of a urine strip with a vitamin C indicator
was assessed. Twenty-five healthy volunteers each ingested 1,000 mg of
vitamin C. Their urine samples were tested for vitamin C using a URiSCAN
11 strip (YD Diagnostics, Korea) before and after administration of vitamin C. Standard materials were added to normal pooled urine to generate
urine samples with various concentrations of the analytes tested (blood,
bilirubin, nitrite, leukocytes, and glucose), and vitamin C was spiked to
predetermined levels. These samples were then tested using two urine
strips - URiSCAN and Chemstrip test strip (Roche Diagnostics, Germany) to evaluate interference from vitamin C. In clinical samples with positive
vitamin C results, microscopic and chemical analyses were also conducted
to examine the differences. Nine urine samples from the 25 volunteers
were positive for vitamin C before ingestion, and all subjects were positive
after ingestion. Vitamin C spiking of urine demonstrated false-negative
results at various concentrations. Vitamin C in urine can cause significant
interference with urine strip tests. A urine strip with a vitamin C indicator
is useful to reduce the risk of incorrect results in regard to disease states.
Background: As a clinical test for pleural effusion (PE), the microscopic
cell counting and chemical composition (e.g. protein and LD) are usually
examined. Light's criteria are frequently used for distinguishing exudate
from transudate, and the discrimination is significant for the treatment of
the underlying diseases. Our the JAMT committee for standardization of
Body Fluid Analysis are considering the standardized method of chamber
count and differentiation and evaluated the association between the biochemical data including Light's criteria and the microscopic test in exudate
and transudate PE. Method: We examined 346 PE and the underlying diseases include 78 heart failure, 86 tumor, 117 bacterial inflammation, 65
other conditions. To discriminate exudate from transudate, Light's criteria
and Serum-effusion albumin gradient (SEAG criterion; albumin is higher
than 1.2g/dL in exudate) were demonstrated as biochemical examination.
On the other hand, we examined numerical and differential cell counting
as cytological test, and defined the specimen which is higher than 1.0x103
cells/µL as exudate. Result: In 252 cases (78%), the results of exudate and
transudate by the cytological criterion was consistent with the biochemical one. Regarding the discrepancies, the differential counting tended to be
dominated by lymphocytes. In tumor cases, there were more discrepancies
between both methods, compared to other diseases. These tended to be
exudate in Light's criteria and transudate in cell counting, which was also
dominated by lymphocytes. In the fluids from the pneumonia and after
empyema treatment, the absolute cell counts tend to be low in spite of
exudate in biochemical criteria. Conclusion: The numerical cell counts in
PE were well correlated with Light's criteria and may be also useful for distinguishing exudate from transudate. Furthermore, we considered that the
differential counts might be notable information for chronic inflammation
and the treatment progress.
140
Analysis of urinary protein components in individuals with
orthostatic proteinuria
PH-10
The examination for outpatients of the formula (Tanaka-formula)
The examination of the formula that uses the sodium and creatinine
concentrations in spot urine specimens to estimate salt intake.
Ryo Kubota 1, Mihoko Nishizawa1, Toru Igarashi2, Kiyoko Kanamori3, Kiyoko Shiba3, Nobue Sakai1
Erina Kobayashi , Yoshiaki Uchida, Kaori Abe, Nanae Takano
1 Graduate
Course of Health and Social Services, Saitama Prefectural University, Japan
2 Department
of Pediatrics, Nippon Medical School Hospital
3 Faculty
of Health Science Technology, Bunkyo Gakuin University
Ibaraki Prefectural Central Hospital, Japan
PH-12
Fabry disease can be found by urinary sediments
1 Department
of Medical Technology, Osaka University Hospital, Japan
2 Laboratory
for Clinical Investigation, Osaka University Hospital
3 Division
of Health Sciences, Osaka University Graduate School of Medicine
Introduction: Chronic kidney disease (CKD) significantly contributes to
the increased number of dialysis patients with end stage renal disease.A
Japanese CKD risk classification established in 2012, which is defined by
albuminuria, eGFR values and underlying disease,demonstrates the relative risks of CKD in great detail. Although CKD with albuminuria stage can
be detected by using urine test paper, the screening test for CKD with nonalbuminuria stage is not established. In this study, we evaluated the clinical significance of urinary hyaline cast (HC) as a biomarker in CKD with
non-albuminuria stage. Materials and Methods: We have categorized 187
non-albuminuria patients into 2 groups (reference and CKD), and we have
calculated ROC curves, AUC of the ROC, sensitivity, specificity for diagnosis
of CKD, as well as odds ratios by using various kidney function markers, including urinary sediment. Further, we have demonstrated the relationship
between the number of HC and various kidney function markers. Results:
AUC of HC (AUC = 0.687) was larger than that of the other biomarkers,
excluding NT-proBNP (AUC = 0.770), and cutoff value for the number of
HC in CKD diagnosis was 10-29 /WF (sensitivity 70.8%, specificity 60.0%,
odds ratio 3.64). Moreover, the eGFR value was significantly lower in the
group with ＞ 10 HC /WF, particularly in hypertensive patients but not in
diabetes mellitus, than that in the group with ＜ 10 HC /WF. Conversely,
the Cys-c value was significantly higher in the group with ＞ 10 HC/WF
than that in the group with ＜ 10 HC /WF. The other biomarkers had no
relevance to the number of HC.Conclusion: Our present study suggests
that the presence of ＞ 10 HC /WF indicates decreased eGFR, and the HC
counting may be important and useful for the screening and early detection of CKD with non-albuminuria stage.
Fabry disease is an X-linked inborn error in metabolism characterized by
the lack of lysosomal hydrolase a-galactosidase A activity. Its pathological
feature is the accumulation of globotriaosylceramide (GL-3) in lysosomes,
particularly in the vascular endothelium of the kidney, heart and brain. It
is a relatively common disease among the lysosomal diseases. Since it can
be found in varied age groups and it presents various symptoms, several
departments such as pediatrics, internal medicine, cardiology, neurology,
dermatology, otolaryngology, and ophthalmology should be engaged in
its diagnosis. Screening of Fabry disease is difficult and time-consuming,
since its diagnosis usually accompanies same specialized examinations.
Recently, some studies reported that the detecting mulberry bodies in
urinary sediments of the patients of Fabry disease is useful in diagnosis.
Majority of mulberry bodies are in the size around 6-10µm and transparent. Their surface structure is coil-like or spring-like, and they look like a
mulberry when they are gathered. A mulberry body looks similar to a fat
globule, but it is easy to distinguish these two by the difference in their
surface structures.Mulberry bodies appear in the urine from the most of
the patients of Fabry disease. Thus, detecting mulberry bodies in the urinary sediments is the simplest way to screen Fabry disease.
141
Poster Presentation
1 Faculty
of Medical Science, Suzuka University of Medical Science, Japan
2 Department
of Clinical Laboratory, Gifu University Hospital
3 Department
of Informative Clinical Medicine, Gifu University Graduate School of Medicine
Oral Presentation
Masaki Hotta 1, Wataru Kobayashi1, Aya Iwata1, Kaori Doi1, Ikuhiro Maeda1, Toru Takano2, Yoh Hidaka2,
Norio Sakai3
Case Conference
Masato Hoshi 1, Mariko Ishida2, Hidekazu Ishida2, Seiko Ushimaru2, Isao Inagaki3, Nobuyuki Furuta2,
Hiroyasu Ito3, Mitsuru Seishima3
Symposium
Is Urinary hyaline cast a new biomarker for CKD with nonalbuminuria stage?
Educational Lecture
PH-11
Special Lecture
Orthostatic proteinuria is diagnosed through a lordotic load test. When
only orthostatic proteinuria is present, the disorder is thought to be benign
with benign causes that will likely disappear over time. We suggested that
close observation is necessary in such cases because the proteins excreted
in the urine of individuals with orthostatic proteinuria are the same as
those excreted in the urine of individuals who have proteinuria with renal
disease. Therefore, we analyzed the urinary protein components after the
lordotic load test to detect specific urinary proteins, in individuals with
orthostatic proteinuria The urine samples after the lordotic load test were
analyzed using cellulose acetate membrane electrophoresis, SDS-PAGE,
and 2D electrophoresis. When the urine samples from before the lordotic
load test and 30, 60, and 90 min after the lordotic load test were analyzed
using cellulose acetate membrane electrophoresis, the urine sample from
30 min after the lordotic load test showed a similar pattern to the serum
protein fraction. However, the urine samples from 60 and 90 min after the
lordotic load test showed protein patterns similar to those observed before
the lordotic load test. Urine samples after the lordotic load test showed
specific bands at 27.7 and 39.2 kDa, based on the SDS-PAGE results. When
the specific bands at 27.7 kDa and 39.2 kDa were analyzed by 2D electrophoresis, spots for apolipoprotein A1 at 27.7 kDa and haptoglobin at 39.2
kDa were detected using the SWISS-2D PAGE database. The detection of
urinary apolipoprotein A1 and haptoglobin may be useful for diagnosing
orthostatic proteinuria.
Invited Lecture
Introduction:Stroke, high blood pressure, and heart disease can be attributed to high salt intake. Tanaka et al, developed a formula (T-formula) that
uses the sodium and creatinine concentrations in spot urine specimens to
estimate salt intake. However, the formula is not accurate enough for clinical use. In this study, to improve accuracy of the T-formula, we tested different factors that could cause a discrepancy between estimated and measured sodium intake and can affect the formula’s accuracy.Methods:269
spot urine samples used in this study were collected June 15th to July 28th
in 2015 in Ibaraki Prefectural Central Hospital. All samples were collected
in accordance with the ethical guidelines and this study was approved
by the Ethics Committee. The patients’ dietary intake was measured and
recorded electronically for 44 days. These measured electronic record
values were compared to the estimated sodium intake values calculated by
the T-formula. We focused on 13 independent factors: gender, hypertensives, BMI, obesity, thin, GFR ＜ 60mL/minute, GFR ＜ 90mL/minute, urine
sugar, urine protein, infusion, high sodium included infusion, diuretic medicine, and high sodium included medicine. The Mann-Whitney U test was
used for statistical analysis.Results:High blood pressure, thin, urine sugar,
urine protein, high sodium included infusion, diuretic medicine, and high
sodium included medicine were shown to have a significant difference between T-formula values and electronic record values. While, Gender, BMI,
obesity, GFR ＜ 60mL/minute, GFR ＜ 90mL/minute, and infusion were
not shown to have a significant difference.Conclusion:We found 7 factors
were shown have a significant difference between predicted and electronic
record value and caused low accuracy of T-formula.
Keynote Speech
PH-09
Keynote Speech
PH-13
Utility of urinary hemosiderin test in the assessment of
treatment for PNH
PH-14
Wataru Kobayashi 1, Masaki Hotta1, Aya Iwata1, Ikuhiro Maeda1, Toru Takano2, Yoh Hidaka2,
Yasutaka Ueda3, Nishimura Junichi3
Keita Kamiyama 1, Tetsuo Matida1, Akihiro Yoshida2, Osamu Araki2, Takao Kimura2, Masami Murakami2
1 Clinical
Laboratory Center, Gunma University Hospital, Japan
2 Department
of Clinical Laboratory Medicine, Gunma University Graduate School of Medicine
1 Department
of Medical Technology, Osaka University Hospital, Japan
2 Laboratory
for Clinical Investigation, Osaka University Hospital
3 Department
of Hematology and Oncology, Osaka University Graduate School of Medicine
Invited Lecture
Special Lecture
Educational Lecture
The sodium glucose co-transporter 2 receptor (SGLT-2) inhibitors are the
newest class of drugs for type 2 diabetes. Inhibition of SGLT-2 results in
a decrease of renal glucose reabsorption and an increase in urinary glucose excretion, with a related reduction of plasma glucose levels. Women
taking the SGLT2 inhibitors have an increased risk of urinary tract and
genital tract infection. In this study we investigated the influence of the
SGLT2 inhibitors on urinalysis. We performed the urinalysis of 30 diabetics treated with the SGLT2 inhibitors. We compared the result of urinalysis
performed by two independent methods; tested by urine dipsticks and
quantitative value measured by automated urine analyzer. The results
of urinary protein, protein/creatinine (P/C) ratio, albumin, albumin/
creatinine (A/C) ratio and glucose were in good agreement for the value
tested by urine dipsticks and quantitative value measured by automated
chemical analysis. The value of specific gravity tested by urine dipsticks
was significantly lower than that of quantitative value measured by the
refraction method. High dose of urinary glucose did attenuate the specific
gravity tested by urine dipsticks. The value of leukocytes tested by urine
dipsticks was significantly lower than that of quantitative value measured
by fluorescent flow cytometry. In the next experiment, we added various
concentrations of glucose to the urine of the patient, then urine analysis
was performed by urine dipsticks and fluorescent flow cytometry. The
results of leukocytes tested by urine dipsticks, but not by fluorescent flow
cytometry, were attenuated by glucose in a concentration dependent manner. In conclusion, the values of specific gravity and leukocytes tested by
urine dipsticks were attenuated by high concentration of urinary glucose
in diabetic patients treated by SGLT2 inhibitors.
Background: Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired
hematopoietic stem cell disorder characterized by complement-mediated
intravascular hemolysis, bone marrow failure, and thrombosis.Eculizumab
is a humanized monoclonal antibody which binds to C5, a