Introduction to Plant Virology • History • Definitions • Classification

Introduction to Plant Virology
• History
• Definitions
• Classification
• Structure
Additional program (student‘s presentation):
Mimivirus (Raoult et al., 2004, SCIENCE 306, p. 1344 ff,
Xiao et al., J. Mol. Biol. (2005) 353, 493–496)
K. Richert-Pöggeler, WS 05/06
Roger Hull: Matthews‘ Plant Virology
(2nd edition, 2004)
http://www.apsnet.org/
http://www.virology.net
First plant virus description 752 AD
Eupatorium yellow-vein (gemini) virus (EpYVV)
Saunders et al., 2003
Dr Robert G. Milne, CNR,
Instituto di Fitovirologica Applicata,
Torino, Italy
Tulip breaking (Poty) virus
Ambrosio Bosschaert,
Dutch painter, 1573-1621
Brunt and Walsh, 2005
1892: Dmitri Iwanowski, Russian scientist, works with tobacco plants with
Tobacco Mosaic Disease
-discovers filtration does not remove infectious agent for TM disease
-could not visualize agent with microscope, nor grow on microbial media
-concluded he had found infectious agent smaller than bacterium
“…infection is not caused by microbes but by
a contagium vivum fluidum”
“…reproduces itself in the diseased plants”
“…other diseases of unknown cause may be
ascribed to a contagium fluidum”
VIRUS (lat.): venom, slime
Martinus W. Beijerinck
(1851-1931)
Über ein contagium vivum fluidum als Ursache der
Fleckenkrankheit der Tabaksblätter. Verhandel.
Acad. Wetensch. Amsterdam 65:3-21 (1898)
John Shaw, 2002
Koch’s Postulates
1843-1910, Nobel Prize 1905
1. The causal agent must be associated in every
case with the disease as it occurs naturally.
2. The causal agent must be isolated in pure culture
3. When the host is inoculated the characteristic
symptoms of the disease must develop.
4. The causal agent must be reisolated.
Visualization of viruses made possible by electron microscope
Magnification up to 106 fold !!!!
nm=10-9 meter
The original electron microscope
as
developed in 1938 in McLennan
Laboratories of the University of
Toronto is now on permanent
exhibition at the Ontario Science
Centre, Toronto, Ontario
Isolation of a crystalline protein possessing the properties of
tobacco mosaic virus. Science 81:644-645, 1935
Wendell
Stanley,
1904-1971
By courtesy of the Molecular Biology &
Virus Laboratory, University of
California, Berkeley
Stanley achieved the first crystallization of a virus (1935), the basis for his
Nobel Prize of 1946 (Chemistry). He later remarked on the unique position
of viruses at the junction of life and non-life:
"The fact that, with respect to size, the viruses overlapped with the
organisms of the biologist at one extreme and with the molecules of the
chemist at the other extreme only served to heighten the mystery regarding
the nature of viruses. Then too, it became obvious that a sharp line dividing
living from non-living things could not be drawn and this fact served to add
fuel for discussion of the age-old question
of 'What is life?'"
Molecular Biology and Biotechnology
• 1969-Restriction endonuclease cloned (Arber & Smith)
• 1970-Reverse transcriptase (Temin & Baltimore)
• 1973-Recombinant plasmid (Cohen & Boyer)
• 1977-DNA sequencing (Gilbert & Sanger)
1977: bacteriophage (ssDNA), 1980: CaMV (dsDNA), 1982: TMV (ssRNA)
• 1984-Polymerase chain reaction (PCR) (Mullis)
• 1995-Entire genome sequenced (Haemophilus influenzae)
Distinguishing viruses from other organisms
What fails to distinguish viruses from cellular organisms?
1. Size
The Mimivirus has a size larger than the smallest
bacteria and, with about 900 genes, a genetic
complexity greater than that of the most reduced
bacteria
http://news.bbc.co.uk/1/hi/health/2895165.stm
Matthews, R. E. F. (1981). Plant Virology. Academic Press.
Closteroviridae: 1.9x104
Nanoviridae: 1x103
Mimiviridae (amoeba):1x106
Distinguishing viruses from other organisms
What fails to distinguish viruses from cellular organisms?
2. Obligate intracellular parasite
Fails to distinguish viruses from many bacteria, Mycoplasma,
Rickettsiae and Chlamydiae
Mycoplasma: 150-300 nm diamenter, bilayer membrane, ribosomes, DNA
no cell wall. Replication by binary fission.
Rickettsiae: nonmotile bacteria (typhus fever), CW, plasmamembrane, ribosomes,
DNA, binary fission, ATP production.
Chlamydiae: psittacosis, elementary-, reticulate bodies (bilayer membrane,
binary fission)
3. Stable, inert phase in life cycle
Many bacterial spores are more stable than some virus particles
General characteristics of viruses
A. Acellular, don’t synthesize a cell membrane (+/- envelope= stolen host cell membrane)
B. Genome = RNA or DNA
C. Protein coat = capsid
D. No ribosomes. Lack ability to synthesize organic molecules
E. No metabolism. Can’t generate own energy therefore are “metabolic parasites”
F. Obligate intracellular parasites-can only replicate inside another host cell
G. Host cell specificity: all cellular organisms may be attacked
1. Viral adhesins must bind specific host cell surface receptors
2. Appropriate host enzymes for viral replication
3. Ability of replicated viruses to be released from host cell
H. Viruses do not grow, nor divide.
Viruses direct synthesis of viral nucleic acid and viral proteins by host cell.
Viruses are “assembled”.
Distinguishing “virion” from “virus”
Virion
the particle that is the extracellular phase of the infection
cycle, typically composed of the genomic nucleic acid and
coat protein but may have a lipid membrane and other
components. Intact non-replicating virus particles, no signs
of life
Virus
virion plus intracellular aspects, including replication
intermediates
Alive?
viruses reproduce; property of life
occur as populations
have variation that is inherited
Why viruses are non-living
Lack a complete protein synthesis system
Lack a complete energy generation system
Virus Disease Symptoms
Local lesions
Yellowing
Stunting
Barley yellow dwarf virus
Beet mild yellowing virus
Ringspot
Necrosis
PV-Y
Mosaic
Abutilon mosaic virus
Tomato spotted wilt virus
Developmental abnormalities
Tobacco mosaic virus
Alfalfa mosaic virus
Zucchini yellow mosaic virus
Microsymptoms
Chloroplast Degeneration
(tymoviruses)
Enlarged Nuclei
(rhabdoviruses)
Disorganized Mitochondria
(aggregation (potyvirus), modification (tombusvirus)
Inclusion Bodies
(caulimoviruses, potyviruses)
Cytoplasmic inclusion bodies
Pinwheel Inclusions (Potyvirus)
Originate and develop in association with
the plasma membrane
CI protein of potyviruses
RNA replication
Inclusion bodies (caulimovirus)
Lesemann and Casper, 1973, Phytopathology 63
Protein enoded by gene 6 of CaMV
Host range
Symptom expression
Translation (transactivator)
DNA replication
Symptoms are not sufficient to classify virus:
• mixed infections
• distinct strains that cause different symptoms in same host
• same symptoms, but different viruses:
Tobacco mosaic virus (ssRNA)
Cauliflower mosaic virus (dsDNA)
Abutilon mosaic virus (ssDNA)
mixed infected petunia (Richert, 1992): PVCV, CMV, PV-Y, TMV
Virus classification
Nucleic acid
Morphology
Genome organisation
Transmission vector
http://www.ncbi.nlm.nih.gov/genomes/VIRUSES/viruses.html
Entrez Genomes currently contains 2139 Reference Sequences
for 1486 viral genomes and 36 Reference Sequences for viroids.
Deltavirus
Retro-transcribing viruses
Satellites
dsDNA viruses, no
RNA stage
dsRNA viruses
ssDNA viruses
ssRNA negativestrand viruses
ssRNA positive-strand viruses, no
DNA stage
unclassified
bacteriophages
unclassified viruses
Comments and suggestions to: [genomes@ncbi.nlm.nih.gov]
Revised: October 18, 2005
Comparitive abundance of different viruses
+ssRNA
600
dsDNA
-ssRNA
dsRNA
300
ssDNA rtDNA
H. Scholthof
Relative abundance of different plant viruses
+ssRNA
dsDNA
-ssRNA
dsRNA
ssDNA
rtDNA
H. Scholthof
Classification of plant viruses
Genome (DNA or RNA)
dsDNA-RT
Caulimoviridae (pararetroviruses, vertebrates)
dsRNA
Partitiviridae (fungi)
Reoviridae (invertebrates,
vertebrates, fungi)
ssDNA
Geminiviridae
Nanoviridae
ssRNA(-)
Rhabdoviridae (invertebrates, vertebrates)
Bunyaviridae (invertebrates, vertebrates)
Tenuivirus, Varicosavirus
ssRNA-RT
Pseudoviridae (invertebrates, fungi)
Metaviridae (invertebrates, fungi)
ssRNA(+)
Bromoviridae*, Comoviridae, Sesquiviridae, Tombusviridae, Luteoviridae, Tymoviridae
Flexivirdae, Potyviridae#, Closteroviridae
Ourmiavirus
Tobamovirus, Tobravirus, Hordeivirus, Benyvirus, Pomovirus, Furovirus
Pecluvirus
* Alphavirus, # Picornaviridae
Flexuous rod
Virus Particle
Structure
Bacilliform
Geminate
Spherical
Rigid rod
Morphology
COMPOSITION OF TMV VIRIONS
TMV virions are rod shaped, 300nm long and about 18nm in diameter. The
virions have helical symmetry and a hollow, cylindrical core.
Component
Number of
molecules in
virion
Molecular weight
RNA [6395
residues]*
1
2.3 x 106
Capsid protein
about 2140
17,500
* The virion RNA has a 5' cap structure at the 5' nucleotide residue
Hull, p. 132
The regular icosahedron: Symmetry
12 vertices, 20 identical triangular faces
5 fold rotational symmetry
center of face - 3 fold symmetry axis
midpoint of each edge - 2 fold symmetry axis
Variations on a Theme
Objective: Make particles larger and more spherical
Strategy: Divide the original 20 faces in smaller faces,
which each again can be filled with subunits
Triangulation: T=Px(f)2
Many viruses: P=3, f=1, T=3; (pentamers and hexamers of subunits)
Page 136
Icosahedral Symmetry
• In higher order icosahedra, the symmetry of the particle is
defined by the triangulation number of the icosahedron.
• The triangulation number,
number T, is defined by:T = f 2 x P
where f is the number of subdivisions of each side of the
triangular face, f 2 is the number of subtriangles on each
face & P = h2 + hk + k2, where h & k are any distinct, nonnegative integers.
Fooling around with P
Fig. 5.17, page 137
Each original face is divided up in 6x1/2 new faces: P=3
Each new sub-triangle again can handle 3 protein units
T=Pxf2: 3x1=3===>60x3=180 protein subunits
Triangulation Numbers
Molecular Virology, 3rd edition, Academic Press
icosahedral Cowpea mosaic (como) virus virion
Three copies
of L coat
protein
Five copies of
S coat protein
(12 vertices) x
(5 S protein per vertex)
= 60 copies of S protein
(20 faces) x
(3 L protein per face)
= 60 copies of L protein
Capsid is composed of equal molar amounts of two coat proteins, L and S
Virus Structure
(Reconstruction based on X-Ray crystal structures)
Tomato Bushy Stunt Virus
Cowpea Chlorotic Mottle Virus
D. M. Rochon, Canada
Genome organisation
The Mimivirus has a size larger than the smallest
bacteria and, with about 900 genes, a genetic
complexity greater than that of the most reduced
bacteria
http://news.bbc.co.uk/1/hi/health/2895165.stm