Virus Quantification + Virus Replication Flashcards
Define virus quantification
Counts the # of viruses in a specific volume to determine virus concentration
What is a virus titer measuring?
lowest conc. of virus that still infects cells
or infectious units per ml of sample
What is a biological quantification test dependent on?
the virus particle initiating successful replication cycle
What is a physical quantification test dependent on?
- does NOT depend on any biological activity of the virus particle
What are examples of biological quantification tests?
- Plaque assay
- Pock assay
- Various end point titration methods
What are examples of physical quantification tests?
- Electron microscopy
- Hemagglutination
- Immuno assays (ELISA)
- qPCR
- Flow cytometry
What is the most direct method to determine the concentration of virus particles in a sample?
Transmission Electron Microscopy (TEM)
** PHYSICAL TEST
Give examples of a physical test that directly counts viral particles in solution
- TEM
- Virus counter 2100
What physical tests are an assessment based on antigen concentration?
- Hemagglutination assay
- Single radial Immunodiffusion (SRID)
What is a plaque?
circular zone of necrotic cells surrounded by viable cells in a monolayer
What physical tests are an assessment based on gene expression?
qPCR
Which biological test measures the number of virus particles capable of forming plaques per unit volume?
Monolayer plaque assay
If a solution has a PFU of 500 pfu/mL, what does this mean?
every mL of solution contains enough virus to form 500 plaques
What is a pock?
Necrotic area on chorioallantoic membrane (CAM) of embronated egg
Pock assay is a biological or physical test?
Biological
Which biological test is a quantitative determination of oncogenic viruses?
Transformation assay
What are the units used in a transformation assay?
Focus-forming units / mL
What is the endpoint of a quantal assay?
Virus dilution that affects 50% of test subjects
What is TCID50?
tissue culture infectious dose which will infect 50% of the cell monolayers
Which of the following is not a physical assay for quantification of viruses?
A. ELISA
B. Electron microscopy
C. Pock assay
D. qPCR
C. Pock assay - biological
What is a permissive cell?
A cell that allows a virus to replicate
cell machinary that supports replication
What is a non permissive cell?
A cell that lacks necessary factors for viral replication
Ex: lack of appropriate receptors
What is MOI?
Multiplicity of Infection
Refers to # of virions added per cell during infection
What is the latent period?
After uncoating and until just before 1st appearance of EXTRACELLULAR new virus particle
**no extracellular virions are detected at this stage
What is eclipse period?
After uncoating and until just before 1st appearance of INTRACELLULAR new virus particle
What is burst size?
of infectious virions released per avg cell
What are the steps of virus replication?
- Attachment
- Penetration
- Uncoating
- Synthesis of viral components
- Assembly + maturation
- Release in large numbers
True or False: Each virus has its own specific receptor(s) on specific host cells
TRUE - lock and key!
What virus uses more that 1 host cell receptor for attachment?
HIV
What is a co-receptor?
An additional surface molecule on host cells that is required for entry of viruses
How do non-enveloped/naked viruses penetrate and the host cell?
- Receptor mediated endocytosis (most common)
- Pore mediated penetration
How do enveloped viruses penetrate and uncoat in the host cell?
** depends on type of fusion protein
- Surface membrane fusion -pH independent fusion protein
- Receptor mediated endocytosis - pH dependent fusion protein
What are other uncommon methods of entry for viruses?
Ab mediated attachment and penetration
** seen in FIP virus
What viruses inject their genome into the host cytoplasm through creation of a pore in the host membrane?
Non enveloped viruses
**pore mediated penetration
How does FIP virus infect the host cell?
Antibody mediated attachment and penetration
-enters host macrophage by attaching spike proteins to CD13 receptor on host cell
At which phase in viral replication can virions no longer be detected?
Viral uncoating
What are the functions of the parent virus?
- Multiple copies for new viruses
- Viral proteins for capsid and successful replication
What does reverse transcriptase do?
Converts viral RNA to cDNA during viral replication
How does processing of primary RNA transcript (pre-RNA) occur?
- Capping- adding 7-methylguanosine to 5’ end
- Adding poly-A tail to 3’ end
- Splicing
What is capping?
Addition of 7-methylguanosine to the 5’ end of RNA
What is RNA splicing?
process that REMOVES INTRONS and JOINS EXONS
_____ are the portion of a gene that codes for AA
exons
________ are the portion of the gene that does not code for AA
Introns
What is constitutive splicing?
all introns are spliced out, all exons are spliced in
What is alternative splicing?
All introns spliced out, only selected exons are spliced in
Where does assembly and maturation of the virus occur?
- Nucleus
- Cytoplasm
- Plasma/cell membrane (most enveloped viruses)
What types of viral mRNA are there?
- Monocistronic - mRNA that encodes 1 polypeptide
- Polycistronic - mRNA that encodes many polypeptides
Where does assembly and maturation take place in most enveloped viruses?
Plasma membrane / cell membrane
How do naked viruses release progeny of new virions?
- Lysis of host cell
How do enveloped viruses release progeny of new virions?
Budding + exocytosis
________ viruses can not exit the host cell by budding
Naked viruses
bc no envelope
___________ integrates viral DNA into the host genome
Integrase
How do viruses spread from cell to cell?
- Extracellular spread
- Intercellular spread
- Nuclear spread of virus genome
________ results in rapid virus dissemination, evasion of immune system, and persistent infections
Intercellular spread of viral genome
