Properties of Viruses

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Tom German 637 Russell Lab 262-2956 tlg_at_entomology
.wisc.edu Virology/Virus vector relationships
Schuman and Darcy-Essential Plant
Pathology Chapter 5-What are the Causes of Plant
Diseases- Viruses
Appendix 3-Rapids Assays for Pathogens
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What are viruses and viroids ? What are they not
? Why do we care ? Can we control them ?
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Tulip Flower Break Virus
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Tobacco Mosaic Virus (TMV) has played a central
role in the history of virology
The germ theory gained acceptance in the 19th
century with a recognition of bacteria as
infectious agents of disease.
- -Louis Pasteur -Robert Lister -Robert Koch
(1876, 1882)
Infectious filterable agents -Foot and
mouth disease (1898) - Loeffler and Frosch
-Tobacco mosaic disease (1886-1898) -
Mayer - Ivanowski - Beijerinck Stanley 1935
Bawdin Pirie 50s
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Tobacco mosaic virus symptoms on tobacco
Tobacco Mosaic Virus
From Mathews Plant Virology, Fourth Edition, 2001
http//www.apsnet.org/education/k-12plantpathways/
TeachersGuide/Activities/TMV/purpose.htm
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Fig. 10. TMV infection of plants with or without
the dominant N gene. (Left) Systemic mosaic
symptoms of TMV infection on nn tobacco plants.
(Right) Localized necrotic lesions typical of the
hypersensitive response on a leaf from a
TMV-infected tobacco plant carrying the N gene.
Photographs courtesy of Dr. Milt Zaitlin, Cornell
University.
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Tospovirus Symptoms
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Maize fine streak virus causes fine chlorotic
streaks and dwarfing on maize.
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Tobacco Rattle Virus
Corky Ringspot Disease
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CMV-Fny CMV-M
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Potato Virus Y complex
PVYntn
PVYo
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Cucumber mosaic virus (CMV)
Rugosity
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PVX PVY PVY ONLY
PVX ONLY
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Symptoms of Viroid Diseases
CCCVd
PSTVd
CSVd
CEVd
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Viroids are common plant pathogens which are a
serious economic problem. 25 different viroid
sequences have been determined and numerous
variants identified
Pospiviroidae are a large group, only 2 members
of the Avsunviroidae are currently known.
 
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The RNA genomes of viroids are 246-375
nucleotides in length and share many
similarities They are all single stranded
covalent circles There is extensive
intramolecular base pairing A DNA-directed RNA
polymerase makes both plus and minus strands
Replication does not depend on the presence of
a helper virus No proteins are encoded
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Circular and linear forms of the Potato Spindle
Tuber Viroid observed using a nucleic acid
spreading technique followed by high angle
shadowing with platinum metal.Magnified
350,000X.
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Host Range
Host range the Viral Perspective

• Wide host range One virus infects more than one
  plant species  
• Narrow host range Some viruses are host specific 

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Host TRange
The Plant Perspective

• Each plant species is usually susceptible to
  several different kinds of viruses
• A plant can be infected with more than one kind
  of virus at the same time
• mixed infections
• synergistic effect

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What are viruses and viroids ? What are they not
? Why do we care ? Can we control them ?
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VIRUSEGeneS!! What they are ??
General Characteristics of Viruses

• Nucleoprotein
• nucleic acid surrounded by a protein coat
• package of nucleic acid wrapped in a protein coat
  
• acellular, lacks a metabolism of its own
• Obligate parasites
• can only multiply in living cell
• has the ability to cause disease
• Submicroscopic
• too small to be seen individually with a light
  microscope
• electron microscope (EM)

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Viruses The Ultimate Parasites

• No physiologic activity
• No energy production
• Do not divide
• Do not produce any kind of specialized
  reproductive structures
• Multiply by inducing host cells to make more
  virus
• Cause disease by utilizing cellular substances
  during multiplication
• Do not consume cells or kill them with toxins

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Viruses The Ultimate Parasites

• Smaller than the smallest prokaryotic cell
• Variable size range
• small--25 nm
• larger--300 nm

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Fig. 3. Classification of plant viruses. From
Van Regenmortel, M.H.V., Fauquet, C.M., Bishop,
D. et al. (1999). Virus Taxonomy Seventh Report
of the International Committee on Taxonomy of
Viruses.
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Plant Viruses Nucleic Acids
ss RNA viruses () RNA Tobacco mosaic virus
(-) RNA Tomato spotted wilt virus ds RNA
viruses Rice fiji disease virus ds DNA
viruses Cauliflower mosaic virus ss DNA
viruses Tomato yellow leaf curl virus
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What it is a VIRION?

• The intact, infectious viral particle
• Monopartite one type of virus particles
• Multicomponent (split genomes)
• bipartite two types of virus particles
• tripartite three types of virus particles
• All must be in plant for disease development
• Different length rods
• Different density isometric particles

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Luteoviruses / Potyviruses
Geminiviruses
Tobacco mosaic virus
Tospoviruses
Rhabdoviruses
The Big Picture Book of Viruses
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Fig. 20. Structure and genome organization of
sonchus yellow net virus (SYNV), a
nucleorhabdovirus. From Jackson et al. (1999)
Encyc. of Virology (3rd Edition), pp. 1531-1541
with permission.
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Icosahedral structure of viruses in the
Bromoviridae
For these types of icosahedral viruses (T3),
virions consist of 180 coat protein subunits
Speir et al., Structure 363-78 (1995)
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Speir et al., Structure 363-78 (1995)
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Coat Protein

• Provides protective sheathing for NA
• Determines vector transmissibility
• Determines symptoms
• Assembly
• Inclusion bodies

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Coat protein Composition?

• Protein shell composed of repeating subunits,
  constant for a given virus
• TMV protein subunit 158 amino acids
• Protein subunits arranged in a helix
• Diameter 18 nm
• Central hole diameter 4 nm
• NA packed tightly between the helices
• NA in polyhedral viruses folded in unknown matter
  inside the shell

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Diagnosis and Identification
Symptoms

• Virus morphology
• Viral nucleic acid components
• Viral Protein characteristics
• ELISA
• Protein physical properties
• Inclusion Bodies in infected cells

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Cucumber Mosaic Virus
Three genomic RNAs
Single Coat Protein
Isometric Virions
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Enzyme Linked Immunosorbent Assay ELISA
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The Process
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The Process
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Light Microscopic Identification
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Viral Life Cycle

• Enter host cell
• Produce viral nucleic acid and proteins
• Avoid the host defense mechanism
• Move cell-to-cell
• Systemic movement
• Move to the next plant

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animal cell
plasmodesmata
mechanical or vector inoculation
cell wall
plant cell
Modified from Levy et al. (1994)
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Biological vectors
Mechanical Transmission
seed pollen epiphytes nematodes fungi mites insect
s
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Insect transmitters of plant viruses
Clemson University
nationalgardening.com
Aphids
Whiteflies
UNL Dept. Entomology
J. Cho
Beetles
Leafhoppers / Planthoppers
Thrips
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Gene Expression Production of viral gene
products (proteins) Replication Synthesis of
new copies of viral genomic RNA
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DNA
RNA
Protein
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Eukaryotic mRNA
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3
AUG
UAA
ORF

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Generic RNA Virus Genome
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3
Replication Movement Coat

• Helicase
• NTPase
• Polymerase

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Gene Expression Stratagies of RNA Viruses
Divided genomes Subgenomic mRNA synthesis Read
through of termination codons Translational
frame shift Internal initiation of
translation Proteolytic processing of a
polyprotein precursor Differential translation
efficiency
Viruses use one or a combination of these
methods
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BMV Gene Expression
Replication
Transcription
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TMV Gene Expression
126/183 30K CP
VS ()
STOP
SgP
VC (-)
3 5
XXXXX XXXXX

5 3
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Gene Expression Production of viral gene
products Replication Synthesis of new copies
of viral genomic RNA
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Viral RNA Replication
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The replication cycle of (TMV)
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Infection and Replication

• Enter cells
• Wounds (mechanically or vectors)
• Deposition into ovule by pollen grain
• RNA freed from the coat protein
• messenger RNA ( strand RNA)
• Cell machinery translates RNA polymerase
• Form complementary RNA (- strand RNA)
• dsRNA (replicative form)
• (-) strand work as template for new ssRNA
• Translation of other genes
• viral proteins (coat, movement, etc.)
• Assembly RNA protein subunits form virion

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Forces acting on virus populations

• 1. Genetics
• Mutation
• Recombination
• Reassortment
• 2. Selection

selection an interaction between the viral
genome and the environment that
determines fitness.
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Virus Blueprint
ATAACGCTAGCTAGCTATATTTAGACCCGTATGCATGCTCG ATAGCAGA
AGTCGCATCGATTAGACTCAGAGCATCGATCGA ATCGTGCGTGCGTAGG
GCTCCGTCCGTACGATCGTCCGACC CCGATCGATCATAAAATTGGATCA
CGTATCGCATATCGGCA ACGGATCGATTATAGCGACTAGCAGATCGTAG
CATCCGCAC GTATCTACGATGCTACTACGTACGTATGCATGTACGTGCG
C GGGATTCAGCATATTATACGCCGTTTCGATCTACGTATCTAC TACTC
CGCATCGATCTACGGCCGCCGGGTTAAAAGTCGCTC ATGTCGTACTGAT
CGAGATGCTACTACGTACTACTGACTGGT TTCCACACGTAGCCAGCATG
CAGCTGACGACGTATGCATGC CCCTTCTTTACGATCGACGATCGTACGT
ACGACGTACTCGAA GCATGCTAGCTAGTCTACGTAGTACGAGTCTGACT
GACGTC CCATTACGTACGTGACTGTACGTAGCTGCATGCAGACGCTG
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Virus Blueprint
ATAACGCTAGCTAGCTATATTTAGACCCGTATGCATGCTCG ATAGCAGA
AGTCGCATCGATTAGACTCAGAGCATCGATCGA ATCGTGCGTGCGTAGG
GCTCCGTCCGTACGATCGTCCGACC CCGATCGATCATAAAATTGGATCA
CGTATCGCATATCGGCA ACGGATCGATTATAGCGCCTAGCAGATCGTAG
CATCCGCAC GTATCTACGATGCTACTACGTACGTATGCATGTACGTGCG
C GGGATTCAGCATATTATACGCCGTTTCGATCTACGTATCTAC TACTC
CGCATCGATCTACGGCCGCCGGGTTAAAAGTCGCTC ATGTCGTACTGAT
CGAGATGCTACTACGTACTACTGACTGGT TTCCACACGTAGCCAGCATG
CAGCTGACGACGTATGCATGC CCCTTCTTTACGATCGACGATCGTACGT
ACGACGTACTCGAA GCATGCTAGCTAGTCTACGTAGTACGAGTCTGACT
GACGTC CCATTACGTACGTGACTGTACGTAGCTGCATGCAGACGCTG
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REASSORTMENT OF GENOME SEGMENTS OCCURS BETWEEN
GENOTYPES WITHIN A TOSPOVIRUS SPECIES
Coinoculation

• Reassortants may have altered properties, e.g.
  responses to plant resistance, thrips
  transmission properties.
• Reassortment is one way in which tospovirus
  populations may respond to selection pressure
  (e.g. from the plant or insect host).

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RNA viruses undergo recombination
RNA recombination is the formation of chimeric
molecules from segments previously separated on
the same molecule or present on different
parental molecules (Nagy and Simon, 1997).
A
B
Parental RNAs
a
b
RNA recombination
Recombinant RNA
a
B
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Template-switching model for viral RNA
recombination
A
B
Parental RNAs
a
b
Polymerase
a
Recombinant RNA
B
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Tobacco Rattle Virus
Transmitted by nematodes Course sandy
soils Depths of 80-100 cm Virus survives in
dormant nematodes 2-4 yrs Both virus and vector
have large host range Symptoms vary with
cultivar Symptom expression more pronounced at
lt20C (68F) Nematode feeding site influences
symptom expression Poor transmission through
plant Poor transmission from seed to daughter
tubers Virus rapidly diluted in absence of vector