Basic Virology

Question 1

What is a virus?

Answer 1

• Obligate IC parasites that replicated by self assembly of individual components rather than by binary fission
• Cannot make energy or proteins independent of host cell
• Contain genome of limited size that is either DNA or RNA, not both

Question 2

What are the physical and biochemical characteristics used to classify viruses?

Answer 2

• Size
• Morphology (shape and presence/absence of envelope)
• Type of genome
• Mechanism of replication

Question 3

What are capsids? Name their 3 forms.

Answer 3

• Rigid structures that withstand environmental conditions
• Result of self-assembly of virally-encoded capsomeres
• Helical, icosahedral or spherical, complex: shape dictated by capsomeres that self-assemble, not by shape of genome
• Genome + capsid = nucleocapsid = virion for “naked” viruses

Question 4

What are the characteristics of enveloped viruses?

Answer 4

• Lipid envelope derived from cellular membranes
• Virally-encoded glycoproteins inserted into membrane and serve as virus attachment/membrane fusion proteins
• Nucleocapsid + membrane = virion for enveloped viruses
• B/c envelope is mostly lipid, enveloped viruses are less stable than naked viruses -> 1) more susceptible to drying, 2) sensitive to detergents and alcohols, 3) can’t survive in GI tract
• Spread in large droplets, secretions, organ transplants, and blood transfusions

Question 5

What are the 8 major steps in viral replication?

Answer 5

1. Attachment
2. Penetration: endocytosis, membrane fusion
3. Uncoating
4. Early transcription and synthesis of nonstructural proteins (A: RNA viruses via virally-encoded RNA-dependent RNA polymerase; B: DNA viruses via host RNA polymerase, except poxvirus)
5. Genome replication (A: RNA viruses cytoplasmic; B: DNA viruses nuclear, except poxvirus)
6. Late transcription and synthesis of structural proteins
7. Assembly of virus particles (A: RNA virus cytoplasmic; B: DNA viruses nuclear, except poxvirus and hepadnaviruses)
8. Release of viral particles: cell lysis, budding (enveloped viruses)

Question 6

What are the 6 elements of viral cytopathogenesis?

Answer 6

1. Inhibition of cellular protein synthesis
2. Inhibition and degradation of cellular DNA
3. Alteration of cell membrane structure
4. Disruption of cytoskeleton
5. Formation of inclusion bodies
6. Toxicity of virion components

Question 7

Describe +RNA virus genome replication.

Answer 7

+RNA genome serves as mRNA and is immediately translated by cellular ribosomes:

1. Translated as polyprotein that must be cleaved into individual proteins (including RNA-dependent RNA polymerase)
2. RNA-dependent RNA polymerase transcribes -RNA strands from the +RNA genome
3. -RNA strands used as templates to make many copies of +RNA genome
4. +RNA copies used as mRNA to make structural proteins, and are encapsidated to make nucleocapsids

Question 8

Describe -RNA virus genome replication.

Answer 8

• RNA genome cannot be used as mRNA; must be used as template to transcribe +RNA (mRNA) strand
  1. To transcribe -RNA genome, incoming virus particle carries RNA-dependent RNA polymerase
  2. Resulting +RNA strand translated into proteins (incl. more RNA-dependent RNA polymerase), and used as template for add’l -RNA genomes
  3. newly produced -RNA genomes encapsidated to produce nucleocapsids

Question 9

Describe retrovirus genome replication.

Answer 9

Retrovirus particles carry RNA-dependent DNA polymerase (reverse transcriptase):

1. +RNA genome reverse transcribed into dsDNA, and integrated into host genome
2. Retrovirus proteins and +RNA genome produced by host enzymes

Question 10

Describe DNA virus genome replication.

Answer 10

DNA virus (except poxvirus) genomes are transcribed by host DNA-dependent RNA polymerases:

1. Many viruses have host shut-off mechanism that degrades host mRNAs
2. Many viruses use specific transcription factors that redirect host polymerases to viral genes and away from cellular genes
3. Viral genome replication dependent on virally-encoded DNA-dependent DNA polymerases for larger viruses, but smaller viruses use host DNA polymerase
4. Newly produced DNA genomes are encapsidated to produce nucleocapsids

Question 11

What is MOI?

Answer 11

Multiplicity of Infection: ratio of the number of infectious particles to number of target cells to be infected

1. MOI = 1 will only infect about 60% of cells in monolayer
2. MOI b/t 5 and 10 needed to ensure all cells are infected

Question 12

What is a plaque assay?

Answer 12

Measure the number of infectious virions in a given volume of lysate:

1. Generally measured as plaque-forming units (pfu) per mL of lysate = titer
2. Biological assay of infectivity

Question 13

What is a lysate?

Answer 13

The suspension of virions in culture medium that results from unrestricted growth of virus on cell monolayer. Not all virus particles produced in lysate are infectious:

1. Physical assays exist to measure # of virus particles
2. Particle-to-pfu ratio measures # of physical particles compared to # of infectious virions

Question 14

What is a plaque?

Answer 14

Hole in confluent monolayer of cells resulting from viruses lysing infected cells

Question 15

What are the 2 periods of the single-cycle growth curve?

Answer 15

1. Eclipse period: post-penetration phase phase until virus can be detected IC (corresponds to uncoating, early transcription, and genome replication steps -> ends at virus assembly)
2. Latent period: post-penetration phase until virus can be detected EC, including the eclipse period (corresponds to uncoating, early transcription, genome replication, virus assembly, and release)

Question 16

Why do viral mutations occur at a relatively high frequency (2)?

Answer 16

1. In part, due to the large # of genome copies produced in every infected cell
2. Also due to polymerase errors, especially for RNA viruses

Question 17

What is complementation?

Answer 17

An exchange of proteins:

1. If genome with lethal mutation in gene X arises along with wild-type genome in same cell, wild-type copy of gene X can provide function for mutant genome
2. Mutant genome packaged in capsid and able to infect second cell upon release
3. Mutant not able to replicate in second cell, b/c genome still contains lethal mutation in gene X

Question 18

What are the three processes virus genomes can undergo in a single cell?

Answer 18

1. Complementation
2. Recombination
3. Reassortment

Question 19

What is recombination?

Answer 19

Exchange of genetic material on same segment of genome:

1. Genome with lethal mutation in gene X arises along with wild-type genome in same cell, then mutation removed by recombination with wild-type genome
2. Virus not able to replicate in second cell because it now contains a wild-type genome
3. Recombination frequent with DNA viruses
4. True recombination doesn’t occur with RNA viruses, but they can exchange genetic material in similar manner

Question 20

What is reassortment?

Answer 20

Exchange of genetic material on different segments of genome:

1. If two segmented viruses infect same cell, they can exchange some of their segments at time of virus assembly
2. Novel strain of virus composed of segments from both parents can be released from infected cell
3. Important mechanism in devo of new influenza strains

Question 21

What is syncytia formation?

Answer 21

Enveloped viruses that fuse infected cell with uninfected cells to directly spread to surrounding cells

Question 22

What is the most common route of viral infection?

Answer 22

Inhalation. Can also enter through breaks in skin or mucosa.

Question 23

How do initial replication and localized spread of viruses occur?

Answer 23

• Initial replication: in cells that express viral receptors and contain appropriate cellular factors for replication
• Localized spread via:
  1. Release of virus from infected cell and subsequent infection of surrounding cells
  2. Syncytia formation (enveloped viruses)

Question 24

What is viremia?

Answer 24

Presence of virions in blood

Question 25

How does secondary spread happen?

Answer 25

- May spread from original site of infection by gaining access to bloodstream or lymphatic system - Can gain access to CNS by circumventing BBB, through cerebrospinal fluid, or by direct uptake in peripheral NN

Question 26

What is the incubation period?

Answer 26

Postinfection period prior to onset of symptoms: 1. 1-2 days is a short incubation period and is characteristic of viruses that do not require secondary spread for symptomology 2. 12-14 days is usually minimum for viruses requiring secondary spread 3. Some viruses have extended incubation periods of months, or years (HIV) 4. W/most diseases, patients can be infectious during incubation period, unknowingly spreading virus

Question 27

What is the acute phase of infection?

Answer 27

The symptomatic phase: 1. Many viral infections are mild or asymptomatic 2. Most viral infections completely resolve following acute phase (virus completely eliminated from body)

Question 28

What are the 3 forms of persistent infection?

Answer 28

1. Chronic (productive): virus produced at low levels, but may not continue to cause disease symptoms (i.e., Hep B) 2. Latent: virus genome remains in cells indefinitely, but virus particles not produced, except during reactivation (i.e., herpes) 3. Transforming: intact or partial virus genome integrates into cellular DNA or is otherwise maintained in cell and immortalizes cell, altering its growth properties (i.e., oncogenic viruses) A: RNA viruses - retroviruses, Hep C B: DNA viruses - Hep B, papilloma virus, polyoma virus, adenovirus type 2, EBV, human herpesvirus-8, pox

Question 29

What is the first defense to viral infection?

Answer 29

- Nonspecific immune response, including NK cells and interferon - Three forms of IFN (alpha, beta, and gamma), but all secreted cytokines that regulate immune and inflammatory responses

Question 30

How are alpha and beta IFN induced? What are the most common PAMPs?

Answer 30

- PRRs like TLRs and cytoplasmic PRRs recognize viral PAMPs | - Main PAMP is dsRNA; others include unmethylated DNA and 5' modified ssRNA

Question 31

How do secreted IFNs prevent viral replication?

Answer 31

Bind to surrounding, uninfected cells and induce pathways that prevent viral replication: 1. Protein kinase (PKR) pathway: inactivates translation initiation factor eIF-2, inhibiting viral protein translation 2. 2-5A system: activates RNAse L, which cleaves viral RNA, destroying RNA genomes or inhibiting viral transcription 3. Mx pathway proteins: GTPases that inhibit RNA polymerase activity

Question 32

What is the major cellular response to primary viral infections?

Answer 32

CD8+ cytotoxic T cells: infected cells targeted for lysis via antigen presentation (antigen-specific responses arise several days post-infection)

Question 33

How are antibodies involved in inhibition of viral infection?

Answer 33

Can neutralize virus binding or facilitate lysis of enveloped viruses with complement

Question 34

Are children defective in Ab production more susceptible to viral infection?

Answer 34

No- indicating the importance of cellular immunity compared to humoral immunity in viral infections

Question 35

List four examples of viral immune defense mechanisms.

Answer 35

1. Antigen variation to escape humoral immune response 2. Inhibition of antigen presentation to escape cellular response 3. Cytokine homologs that down-regulate or block cellular response 4. Latent infection in neurons where is no MHC I

Question 36

List 5 examples of viral immunopathogenesis.

Answer 36

1. Flu-like symptoms caused by IFN and lymphokines 2. Inflammation caused by T cells, macrophages, and PMNs 3. Immune complex disease caused by Ab and complement 4. Hemorrhagic disease caused by T cells, Ab, and complement 5. Immunosuppression

Question 37

When are vaccines not practical?

Answer 37

1. If large number of viral strains (serotypes) that cause disease (e.g., over 100 strains of common cold) 2. If virus undergoes much antigenic variation due to high mutation rates in dominant antigenic structure

Question 38

What are the advantages (2) and disadvantages (3) of attenuated, live vaccine strains?

Answer 38

Advantages: 1. Cheap to produce 2. Long-lasting, strong response (IgG, IgA, T cell) Disadvantages: 1. Can be labile in transport 2. Cannot be given to immunocompromised pts 3. Can revert to virulence in rare cases NOTE: cause subclinical infections

Question 39

What are the advantages (3) and disadvantages (2) of killed virus vaccines?

Answer 39

``` Advantages: 1. Very stable 2. Rare side effects 3. Cannot revert to virulence Disadvantages: 1. More expensive to repair 2. Shorter term immunity mostly limited to IgG ``` NOTE: cannot cause illness

Question 40

What are subunit (recombinant DNA) vaccines? List advantages (2) and disadvantages (1).

Answer 40

Composed of single viral proteins are expressed in yeast using recombinant DNA technology (Hep B and HPV). Advantages: 1) cannot cause disease, 2) not derived from blood (another Hep B vaccine is) Disadvantage: requires multiple injections

Question 41

List some frequently used FDA-approved vaccines.

Answer 41

Polio, Hep B and A, flu, varicella-zoster (chickenpox), and measles, mumps, rubella

Question 42

What are immune globulin antivirals? When are they used?

Answer 42

Passive immunization: used in both pre- and post-exposure prophylaxis