GBS Case

New Guidelines for the Prevention
of Early-Onset Group B
Streptococcal Disease:
Questions & Answers
Roberta B. Carey, Ph.D.
CDC
rcarey@cdc.gov
Gerri Hall, Ph.D.
CCF
hallg@ccf.org
1
2010
Recommendations
MMWR, Vol. 59
(RR-10)
2
Objective: To review revisions
that affect laboratory practice
1.
2.
3.
4.
5.
6.
Molecular testing
Chromogenic media
GBS Identification
Antimicrobial susceptibility tests
Detection of GBS in urine
New algorithm for work-up of
prenatal GBS screening swabs
3
Q1: Why did we need new
recommendations for GBS?
4
GBS Disease
• GBS emerged as important pathogen in 1970s
• Over 7500 cases of GBS sepsis and meningitis
in newborns annually in 1990
– Early-onset disease < 7 days after birth
– Late-onset disease >7 days after birth
• GBS disease remains the leading infectious
cause of morbidity and mortality among
newborns in the U.S.
5
Initial Steps at Prevention
• Early 1980‟s clinical trials showing
intrapartum antibiotic prophylaxis (IAP)
prevented early-onset disease
(Boyer and Gotoff, NEJM 1986)
• Recommendation for IAP issued in 1996
by ACOG and CDC, in 1997 by AAP
– Risk-based approach
– Culture-screening approach
6
1996
GBS Guidelines
7
Later studies showed
screening 50% more
effective
2002
guidelines
recommended AST
for women at high
risk for penicillin
allergies; report any
GBS in urine
pregnant women.
Isolation and
identification
methods same for
1996 and 2002
8
2.5
Group B Strep
Association 1st ACOG & AAP
statements
formed
2
CDC draft
guidelines published
Consensus
guidelines
1.7
1.5
CDC revised
guidelines
1
.32
.6
0.5
Year
2007
2006
2005
2004
2003
2002
2001
2000
1999
1998
1997
1996
1995
1994
1993
1992
1991
1990
0
1989
Cases per 1000 live births
Incidence of Early-Onset Invasive GBS
Disease, Selected Active Bacterial Core
Surveillance Areas, 1989-2007
9
9
Trends in Early-Onset GBS Disease
MMWR 58:109 (2009)
10
GBS by the Numbers
from the Latest Data
• 48% to 85% = woman getting screened for
GBS increased from „98-99 to ‟03-04
• 27% to 32 % = infants exposed to antibiotics
increased
• 85% of GBS+ women get IAP who should
• 63% of women who deliver preterm get IAP
• 74% of GBS cases occurs in term births
• 61% of term infants with GBS born to women
who tested GBS negative
Van Dyke et al. NEJM 360:2626 (2009)
11
New Guidelines to Overcome
False Negative GBS Results from
Antenatal Cultures
•
•
•
•
•
•
•
Improper collection sites
Transport time
Improper culture methods
Antibiotic exposure
Overgrowth with other organisms
Failure to recognize non-hemolytic GBS
Subjectivity in interpreting results
12
Q2: What molecular tests
are available for GBS and
when can they be used?
13
Rapid Commercially Available
and FDA-Cleared Methods for
Molecular Detection of GBS
– Cepheid Smart GBS (Cepheid, Sunnyvale, CA)
• Smart Cycler
• Directly from sample (~1.5 hr) or from LIM broth
(24 hr + 1.5 hr) overnight culture
– Cepheid Xpert GBS (Cepheid)
• GeneXpert format
• < 30 minutes directly from clinical sample
– BD GeneOhm™ StrepB (BD Diag, San Diego, CA)
• Smart Cycler format
– Approval for direct from clinical sample (~ 1- 1.5 hr)
– Validation in lab for use from overnight LIM Broth
(24 hr + 1-1.5 hr)
14
Cepheid Smart GBS
Smart Cycler
• Directly from sample (~1.5 hr) or
• From LIM broth (18- 24 hr + 1.5 hr)
overnight culture
15
Cepheid Smart GBS
Direct
Sensitivity
(%)
Specificity
(%)
Predictive
Value
Positive
(%)
Predictive
Value
Negative
(%)
81.6
96.3
87.8
94.4
98.7
90.5
77
99.5
(Package
Insert)
PCR from
LIM broth
vs. culture
(Jordan et al
JCM 2010; 48:
3193-7)
16
BD GeneOhm™ StrepB
A. SMART CYCLER FORMAT
– Approval for: directly from clinical sample (~ 1-1/5 hr)
– Validation in lab for use from overnight LIM Broth (24 hr + 1-1.5 hr)
B. BD MAX: automated format
- Direct or Lim-enhanced
17
BD GeneOhm™ StrepB
Direct
Sensitivity
(%)
Specificity
(%)
PPV
(%)
NPV
(%)
94
96
84
99
87
95
88
95
100
100
95.3
99.1
(Package insert)
Direct
(Atkins, et al. OB&GYN 2006;
108:488-91; 233 samples)
LIM-enhanced
PCR
(Maloney, J et al. ASM 2004
poster)
LIM-enhanced
PCR*
(Scicchitano and Bourbeau
JCM 2009; 47: 3021-3; 498
samples)
*Sensitivity of Culture = 87%; LIM-enhanced = 92%; Both = 95.3%
18
BD MAX GBS
LIM-enhanced PCR vs. Culture
Site
Prevalence
(%)
Sensitivity
(%)
Specificity
(%)
PPV
(%)
NPV
(%)
1
20
97.4
96.6
86.4
98.4
2
25.1
92
95.9
89.5
97.9
3
23.6
96.2
97.6
88.7
98
Overall
23
95
96.7
88.3
98
Riedlinger J et al. JCM 2010; 48: 4239-41
19
GeneXpert GBS
GeneXpert GBS
• GeneXpert format
• < 30 minutes directly from clinical sample
• Designed to be run in the clinical laboratory
or near-patient (labor and delivery)
20
GeneXpert vs. Culture
Intrapartum PCR vs.
Culture 1
Sensitivity
(%)
Specificity
(%)
PPV
(%)
NPV
(%)
98.5
99.6
97.8
99.7
58.3
92.1
Antenatal Culture 1
(predicting intrapartum)
Intrapartum and
Antenatal vs. Culture 2
91.1
96
88
97
Intrapartum PCR
(colonization rate of
44% by culture) 3
95.8
64.5
68
95
Antenatal culture 3 vs.
Intrapartum Culture
83.3
80.6
1. El Helali et al. CID 2009; 49: 417-23;863 women
2. Edwards RK et al. OB & GYN 2008; 111:1335-41; 784 women
3. Gavino and Wang. Am J Ob & GYN 2007; 197:388; 55 women
21
GBS Status at Delivery
GBS +
GBS -
IAP
No IAP
* If available rapid nucleic acid
amplification testing (NAAT)
may be performed on patients
with unknown GBS status who
present at triage or
labor/delivery with no risk
factors.
Unknown
Risk Factors
No Risk Factors
NAAT *
IAP *
GBS+
IAP
Algorithm for Intrapartum NAAT
GBS -
No IAP
22
GBS PNA FISH®
(AdvanDx, Inc., Wolburn MA)
• LIM broth culture
– 1.5 hr
– Fluorescent
microscope
needed
– FDA-cleared
23
PNA FISH vs. BD vs. Culture
206 samples
Sensitivity
(%)
Specificity
(%)
PPV
(%)
NPV
(%)
BD GBS PCR
(LIMenhanced)
PNA FISH GBS
(LIMenhanced)
100
100
100
100
98.4
100
99.3
100
96.9
100
98.6
100
LIM Culture
Wilson, DA et al. 2010. JCM 48: 1947-8
24
Roche Light Cycler GBS ASR
vs. Culture
200 vaginalrectal swabs
Positive
(%)
(Goodrich JS DMID
2007; 59:17-22)
Culture
Sensitivity
(%)
Based on
Culture
Specificity
(%)
Based on
Culture
26.5
BD-StrepB
(GeneOHM)
30
92.5
92.5
Light Cycler
Strep B
(Roche ASR)
29.5
100
95.9
25
CCF Procedure for GBS
• 2 BD COPAN Swabs collected from each patient:
vaginal-rectal specimen
• Placed in LIM 18-24 hr incubation at 37 C
• Refrigerated after 24 hr if test not performed
• Aliquot of LIM is removed, DNA extracted and
assayed by NAAT in Smart Cycler using BD
GeneOhm GBS
• Performed 5-6 days per week; resulted as
completed
• LIM broths that are positive are kept for 5 d
• If GBS positive and patient is allergic, LIM is
cultured & susceptibility performed
• If invalid, freeze & thaw and repeat NAAT
– Proceed with culture if invalid on repeat
26
Q3: What new tools do we
have for GBS culture?
27
Chromogenic Media
• Majority are a version of Granada
medium
– A selective and differential medium for GBS
– Uses a polyenic red pigment, Granadaene,
to differentiate GBS
• Studies show majority agars and broths
equal to or better than SBA/CNA and Lim
broth for GBS recovery
– Added advantage of detection within 24hrs
– Positives do not require confirmation by
latex agglutination
28
Commercially Available Chromogenic Broths
Media
Agar/Broth
Color
detection
NonHemo
GBS
Incubation
Sensitivity
Granada
Biphasic broth
Broth
Orange /red
No
18-24hrs
(aerobic)
Good
NEL-GBS
Agar/broth
Orange
No
16-24hrs
(aerobic)
Poor
StrepB Carrot
Broth
Broth
Orange / red
No
18-24hrs
(aerobic)
Good
Granada broth
Granada Biphasic broth
StrepB Carrot Broth
29
Commercially Available Chromogenic Agars
Media
Agar/Broth
Color
detection
NonHemo
GBS
Incubation
Sensitivity
ChromID
StreptoB
Agar
Pale pink-red
Yes
18-24hrs
(aerobic)
Good
Granada Agar
Agar
Orange/red
colonies
No
24hrs
(anaerobic)
Good
NEL-GBS
Agar/broth
Orange
No
16-24hrs
(aerobic)
Poor
chromIDStreptoB
Granada agar
30
Non-Hemolytic GBS
• Approx. 4% of invasive GBS isolates
are non-hemolytic
• Chromogenic broths do not detect
non-hemolytic GBS
• Most chromogenic agars do not detect
non-hemolytic GBS
• Need to subculture all negative
chromogenic media if
they don‟t detect
non-hemolytic isolates
31
Q4: What tests are best to
identify GBS from culture?
32
Recovery & Identification
from Enrichment Broths
• If pigmented in selective broth (carrot broth),
report as GBS
• If positive (pigmented) on Granada agar or
Chromogenic Agar, report as GBS
• Culture from LIM / TransVag Broth / nonpigmented carrot broth
– Sub onto BAP or chromogenic agar
– Sub onto GBS Detect Agar
(Hardy Diagnostics)
• Picks up non-hemolytic GBS
from blood agar
or carrot broth
33
Identification of GBS
• Typical colonies on BAP
– Catalase positive gram positive cocci
in chains
• Hippurate: Positive
• CAMP (Christie-Atkins-Munch-Peterson):
Positive
• Latex agglutination confirmation per the
lab‟s “usual method” for identification of
GBS
– AccuProbe (Gen-Probe, San Diego, CA)
– Nucleic Acid Amplification (NAAT)
– Other Molecular tests such as FISH
34
Q5: Can I use automated
susceptibility panels for
GBS?
35
Antimicrobial Susceptibility
Testing
• CLSI M100-S21 Performance
Standards recommend using:
– Disk diffusion
– Broth microdilution
• FDA-cleared/approved commercial system
may also be used
– Testing for inducible clindamycin
resistance
• D-zone or other validated test
36
GBS from Prenatal Cultures
Percent Resistant
160 Loyola Isolates ‟02-03
290 ABCs Isolates ‟06-08
100
90
80
70
60
50
40
30
20
10
0
94
89
47
26 32
17
0 0
PEN
0 0
3RD CLINDA ERY
GEN
CEPH
0 0
TETRA VANC
'02-03
'06-08
37
Antimicrobial Susceptibility
Etest and Disk Diffusion
Zone of
inhibition of
growth for
clindamycin is
≥19 mm
(susceptible)
Erythromycin MIC
= 0.19µg/ml
Etest
Disk
Diffusion
Zone of
inhibition of
growth for
erythromycin
is ≥21 mm
(susceptible)
38
Photo courtesy of Dr. Lesley McGee, CDC
D-zone Test Result for GBS
Disks 12 mm apart
Blunting of the inhibition zone indicating
inducible clindamycin resistance
39
Photo courtesy of Dr. Lesley McGee, CDC
Increasing MICs to Beta-lactams
What Does It Mean Clinically?
• Isolates in Japan and U.S. have elevated MICs
to beta lactams due to mutation in pbp2x gene
– Pen MIC 0.015 wild type
– Pen MIC <0.12 CLSI breakpoint
– Pen 0.25-1.0 (Japan); 0.12 (U.S.)
• Single base change at residue 557 glutamine
>>>glutamate (Q557E)
– 1st step mutation similar to S. pneumoniae
• Accumulate additional mutations lead to
full resistance
Dahesh et al. AAC 52: 2915, 2008
40
Q6: Why do labs no longer
need to report low numbers
of GBS in urine of pregnant
women?
41
Urines as Surrogate Screening
Culture for GBS
• GBS can cause symptomatic and asymptomatic UTI
• GBS bacteriuria is a marker for heavy genital tract
colonization and these women should receive IAP
• Included in the 2002 and new 2010 CDC guidelines, is
a comment about culture of urine in the asymptomatic
pregnant woman as a “surrogate” for detection of
GBS in vaginal/rectal cultures
– The 2002 guidelines recommended detecting and
reporting any quantity of GBS in urine
– The new 2010 guidelines recommend reporting
GBS when > 104 CFU/ml and/or using guidelines
you would use for UTI
• You could still report any quantity if you choose
42
Urines as Surrogate Screening
Culture for GBS
• Report > 104 GBS in pure culture or in
the presence of a second organism
(E. coli + GBS)
• Report should include a comment that
this represents vaginal colonization
unless patient is symptomatic of a UTI
• Treated same as if isolated from the
vaginal / rectal culture
– Results provided to OB / midwife and
available at time of delivery
43
Urines as Surrogate Screening
Culture for GBS
• Vaginal / rectal culture not
needed if urine is positive for
GBS
• Vaginal / rectal culture should be
performed if urine is negative for
GBS
44
Figure 6. Algorithm for recommended prenatal group B streptococcal testing
Vaginal rectal swab†
Enrichment broth (can use non- pigmented or pigmented broth)
Incubate 18-24hrs at 35-37C
Non-pigmented broth
Pigmented broth
Further testing (can subculture or use rapid tests)
Subculture to appropriate media
Incubate 18-24hrs at 35-37C
No indicator
color growth
GBS Indicator
color observed
DNA Probe, latex agglutination or
nucleic acid amplification test (NAAT)
Identify GBS by recommended
method*
GBS -
Re-incubate overnight
GBS+
GBS +
GBS -
GBS +
Report as GBS +
Report as GBS -
Report as GBS +
GBSReport as GBS -
Antimicrobial susceptibility testing if penicillin-allergic and at high risk of anaphylaxis*
45
Summary of Key Changes
• Vaginal / Rectal Culture
–
–
–
–
Expand swab options
Option to use chromogenic broths and agars
Option to use antigen detection, probe, NAAT from broth
Direct plating can accompany broth inoculation (not replace)
• Molecular ID tests
– Not recommended for routine 35-37 wk antepartum screening
– Recommended direct testing for women with no risk factors
and no prenatal screening
– Recommended for use on broth enriched cultures to ID GBS
• Urine Culture Screening
– Report GBS at >104 cfu / ml in pregnant women
• AST
– Test for inducible clindamycin resistance when woman is
at high risk for anaphylaxis due to pen allergy
46
http://www.cdc.gov/groupbstrep
47
2010 Guidelines
Collection and Transport
• Type of swab acceptable for antenatal screening
vaginal/rectal swabs only; cervical swabs or
perianal / perirectal not acceptable
• Recommend use of non-nutritive transport media –
e.g., Amies or Stuart „s (with or without charcoal)
• GBS viability declines over 1-4 days at high
temperatures, refrigerate if delay before processing
• Specimen requisitions for GBS testing should
identify the patient at high risk for anaphylaxis as
penicillin allergic and antibiotic susceptibility testing
for clindamycin and erythromycin should be ordered
48