VIROLOGY

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VIROLOGY
Margaret.Hunt_at_uscmed.sc.edu 733-3293 Building 2,
B-4 (office), B16(lab)
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OUTLINE

• introduction to viruses
• structure and classification
• basic virology
• clinical virology

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VIROLOGY

• different structure
• different method of replication
• implications for
• diagnosis
• treatment
• prevention

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CONTROL METHODS

• INVOLVE KNOWLEDGE OF
• RESERVOIRS
• MODE OF TRANSMISSION
• METHODS TO INACTIVATE VIRUS OF INTEREST
• VACCINES
• ANTI-VIRAL DRUGS
• DEVELOPMENT OF DRUG RESISTANCE

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EMERGING VIRAL DISEASES

• Some North American examples
• HIV/AIDS
• Hantavirus pulmonary syndrome (HPS)
• West Nile encephalitis (WNV)
• Severe acute respiratory syndrome (SARS)
• Monkey pox
• Human metapneumovirus
• 2009 H1N1 pandemic influenza

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Consequences of viral infections

• 50 of all absenteeism
• Children
• 7 or more viral infections per year that
  involve a visit to a physician

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Consequences of viral infections

• Suffering, followed by recovery
• Persistent disease
• Fatal disease
• Congenital disease
• Contributory factor in cancer
• Contributory factor in other diseases

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SOME ARE ASYMPTOMATIC!
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VIRUSES CAN BE USEFUL

• VACCINE DEVELOPMENT
• GENE THERAPY
• TOOLS TO INVESTIGATE HOST CELLS

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WHAT ARE VIRUSES?
A PIECE OF BAD NEWS WRAPPED UP IN A PROTEIN
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WHAT ARE VIRUSES?

• NUCLEIC ACID GENOME
• DNA OR RNA
• PROTEIN COAT
• PROTECTION, ENTRY
• LIPID ENVELOPE IN SOME VIRUSES
• SMALL
• (20-400nm)
• OBLIGATE INTRACELLULAR PARASITES

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Virus particle virion
White, DO and Fenner, FJ. Medical Virology, 4th
Ed. 1994
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Koneman et al. Color Atlas and Textbook of
Microbiology 5th Ed. 1997
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The arenavirus family appears to accidentally
package ribosomes, but these appear to play no
role in protein synthesis.
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CONSEQUENCES

• HEAVILY PARASITIC ON HOST CELL
• NO BROAD RANGE ANTIBIOTICS
• NEED TO LOOK FOR WEAK LINK

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HOST RANGE

• MAY BE WIDE OR NARROW
• MAY BE INSECT/ANIMAL, INSECT/PLANT
• DO NOT CROSS EUCARYOTE / PROCARYOTE BOUNDARY

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FACTORS AFFECTING HOST RANGE - CELL SURFACE
RECEPTORS
CYTOPLASM
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FACTORS AFFECTING HOST RANGE

• CELL SURFACE RECEPTORS

• AVAILABILITY OF REPLICATION MACHINERY
• ABILITY TO GET OUT OF CELL AND SPREAD
• HOST ANTI-VIRAL RESPONSE

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VIRAL STRUCTURE SOME TERMINOLOGY

• virus particle virion
• protein which coats the genome capsid
• capsid usually symmetrical
• capsid genome nucleocapsid
• may have an envelope

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ICOSAHEDRAL SYMMETRY

• 20 faces
• 12 vertices

http//www.tulane.edu/dmsander/WWW/Video/Video.ht
ml
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ICOSAHEDRAL SYMMETRY
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ICOSAHEDRAL SYMMETRY
NUCLEIC ACID IS INSIDE
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ICOSAHEDRAL SYMMETRY
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ICOSAHEDRAL SYMMETRY
PENTON (pentamer)
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ICOSAHEDRAL SYMMETRY
PENTON
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Adenovirus
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Adenovirus
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herpesvirus capsid (membrane removed)
http//www.bocklabs.wisc.edu/Herpesvirus.html
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polypeptides
capsomers
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ICOSAHEDRAL SYMMETRY
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rhinovirus
Jean-Yves Sgro http//www.bocklabs.wisc.edu/multim
edia.html
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SYMMETRY OF NUCLEOCAPSID

• ICOSAHEDRAL
• HELICAL

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TOBACCO MOSAIC VIRUS
nucleic acid
protein
adapted from Klug and Caspar Adv. Virus Res.
7225
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Helical symmetry

• Length controlled by nucleic acid
• Helix may be stiff or flexible

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COMPLEX SYMMETRY
surface view
cross section
POXVIRUS FAMILY
White, DO and Fenner, FJ. Medical Virology, 4th
Ed. 1994
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ENVELOPE

• OBTAINED BY BUDDING THROUGH A CELLULAR MEMBRANE
  (except poxviruses)
• POSSIBILITY OF EXITING CELL WITHOUT KILLING IT
• CONTAINS AT LEAST ONE VIRALLY CODED PROTEIN

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CYTOPLASM
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ENVELOPE

• OBTAINED BY BUDDING THROUGH A CELLULAR MEMBRANE
  (except poxviruses)
• POSSIBILITY OF EXITING CELL WITHOUT KILLING IT
• CONTAINS AT LEAST ONE VIRALLY CODED PROTEIN
• ATTACHMENT PROTEIN
• LOSS OF ENVELOPE RESULTS IN LOSS OF INFECTIVITY

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ENVELOPE
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5 BASIC TYPES OF VIRAL STRUCTURE
nucleocapsid
icosahedral nucleocapsid
lipid bilayer
ENVELOPED ICOSAHEDRAL
ICOSAHEDRAL
helical nucleocapsid
COMPLEX
nucleocapsid
lipid bilayer
glycoprotein spikes peplomers
HELICAL
ENVELOPED HELICAL
Adapted from Schaechter et al., Mechanisms of
Microbial Disease
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UNCONVENTIONAL AGENTS

• VIROIDS
• RNA only
• Small genome
• Do not code for protein
• So far, only known viroids are in plants

• hepatitis delta virus (agent)
• - some viroid, some virus features

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UNCONVENTIONAL AGENTS

• PRIONS
• protein only?
• do not contain any nucleic acid?

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LIVING OR DEAD?
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CLASSIFICATION

• BASIC STRUCTURE AND MOLECULAR BIOLOGY
• particularly important as diagnostic and
  therapeutic abilities expand

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CLASSIFICATION

• NUCLEIC ACID
• CAPSID
• PRESENCE OF ENVELOPE
• REPLICATION STRATEGY

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CLASSIFICATION NUCLEIC ACID

• RNA or DNA
• segmented or non-segmented
• linear or circular
• single-stranded or double-stranded
• if single-stranded RNA
• is genome mRNA () sense or complementary to mRNA
  (-) sense

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CLASSIFICATION CAPSID

• symmetry
• icosahedral, helical, complex
• number of capsomers if icosahedral
• enveloped or non-enveloped

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CLASSIFICATION

• REPLICATION STRATEGY

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DNA VIRUSES
DOUBLE STRANDED
SINGLE STRANDED
COMPLEX
NON-ENVELOPED
ENVELOPED
PARVOVIRIDAE
POXVIRIDAE
ENVELOPED
NON-ENVELOPED
HERPESVIRIDAE
HEPADNAVIRIDAE
CIRCULAR
LINEAR
All families shown are icosahedral except for
poxviruses
PAPILLOMAVIRIDAE POLYOMAVIRIDAE (formerly grouped
together as the PAPOVAVIRIDAE)
ADENOVIRIDAE
Modified from Volk et al., Essentials of Medical
Microbiology, 4th Ed. 1991
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RNA VIRUSES
SINGLE STRANDED
SINGLE STRANDED
DOUBLE STRANDED
positive sense
negative sense
ENVELOPED
NONENVELOPED
ENVELOPED
NONENVELOPED
HELICAL
ICOSAHEDRAL
HELICAL
ICOSAHEDRAL
ICOSAHEDRAL
ORTHOMYXOVIRIDAE
FLAVIVIRIDAE
CORONAVIRIDAE
PICORNAVIRIDAE
REOVIRIDAE
PARAMYXOVIRIDAE
TOGAVIRIDAE
CALICIVIRIDAE ASTROVIRIDAE
RHABDOVIRIDAE
RETROVIRIDAE
FILOVIRIDAE
BUNYAVIRIDAE
ARENAVIRIDAE
Modified from Volk et al., Essentials of Medical
Microbiology, 4th Ed. 1991
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BASIC STEPS IN VIRAL LIFE CYCLE

• ADSORPTION
• PENETRATION
• UNCOATING AND ECLIPSE
• SYNTHESIS OF VIRAL NUCLEIC ACID AND PROTEIN
• ASSEMBLY (maturation)
• RELEASE

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ADSORPTION
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ADSORPTION

• TEMPERATURE INDEPENDENT
• REQUIRES VIRAL ATTACHMENT PROTEIN
• CELLULAR RECEPTORS

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PENETRATION - ENVELOPED VIRUSES

• FUSION WITH PLASMA MEMBRANE
• ENTRY VIA ENDOSOMES

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PENETRATION
herpesviruses, paramyxoviruses, HIV
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PENETRATION - ENVELOPED VIRUSES

• FUSION WITH PLASMA MEMBRANE
• ENTRY VIA ENDOSOMES, FUSION WITH ACIDIC ENDOSOME
  MEMBRANE

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PENETRATION - ENVELOPED VIRUSES
from Schaechter et al, Mechanisms of Microbial
Disease, 3rd ed, 1998
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VIRUS UPTAKE VIA ENDOSOMES

• CALLED
• VIROPEXIS / ENDOCYTOSIS / PINOCYTOSIS

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PENETRATIONNON-ENVELOPED VIRUSES
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PENETRATIONNON-ENVELOPED VIRUSES
entry directly across plasma membrane
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UNCOATING

• NEED TO MAKE GENOME AVAILABLE
• ONCE UNCOATING OCCURS, ENTER ECLIPSE PHASE
• ECLIPSE PHASE LASTS UNTIL FIRST NEW VIRUS
  PARTICLE FORMED

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SYNTHESIS OF VIRAL NUCLEIC ACID AND PROTEIN

• MANY STRATEGIES
• NUCLEIC ACID MAY BE MADE IN NUCLEUS OR CYTOPLASM
• PROTEIN SYNTHESIS IS ALWAYS IN THE CYTOPLASM

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ASSEMBLY AND MATURATION

• NUCLEUS
• CYTOPLASM
• AT MEMBRANE

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smallpox virus cytoplasmic assembly and maturation
F. A. Murphy, School of Veterinary Medicine,
University of California, Davis.
http//www.vetnet.ucdavis.edu/fam_graphics/downloa
d.html
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RELEASE

• LYSIS
• BUDDING THROUGH PLASMA MEMBRANE
• NOT EVERY RELEASED VIRION IS INFECTIOUS

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HIV budding and maturation
Hsiung, GD et al., Diagnostic Virology 1994 p204
(D. Medina)
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HIV mature form
Briggs JA et al. Structure. (2006) 1415-20
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DEFINITIONS - VIRAL PROTEINS

• STRUCTURAL PROTEINS
• ALL PROTEINS IN A MATURE VIRION
• NON-STRUCTURAL PROTEINS
• VIRALLY CODED PROTEINS WHICH ARE NOT PACKAGED IN
  THE VIRION

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EFFECTS ON HOST

• MAY INHIBIT HOST DNA, RNA OR PROTEIN SYNTHESIS
• DETAILS AND MECHANISM VARY

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CYTOPATHIC EFFECT

• ANY DETECTABLE CHANGES IN THE HOST CELL
• MORPHOLOGICAL CHANGES

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uninfected
HIV infected
HIV infected (at higher magnifcation)
Hockley et al. J Gen Virol 692455-2469
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CYTOPATHIC EFFECT

• ANY DETECTABLE CHANGES IN THE HOST CELL
• MORPHOLOGICAL CHANGES
• DEATH
• APOPTOSIS
• INDEFINITE GROWTH

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tissue culture cells
epithelial
epithelioid
fibroblastic
slides from CDC
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epithelial cells - adenovirus
uninfected
early infection
late infection
slides from CDC
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epithelial cells - respiratory syncytial virus
uninfected
respiratory syncytial virus
slides from CDC
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fibroblastic cells - herpes simplex virus
uninfected
early infection
late infection
slides from CDC
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fibroblastic cells - poliovirus
uninfected
early infection
late infection
slides from CDC
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PLAQUE ASSAY
PLAQUE ASSAY
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PLAQUE ASSAY
PLAQUE ASSAY
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Diluted 10 fold
Diluted 100 fold
Diluted 1000 fold
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PLAQUE FORMING UNIT

• P.F.U.
• pfu

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SOME POINTS TO REMEMBER

• INFECTIVITY
• NOT EVERY RELEASED PARTICLE IS INFECTIOUS

• ASSAYS
• detect every particle (e.g. electron microscope)
• detect infectious particles only (e.g. plaque
  assay)

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GLOSSARY