Virus Replication

Question 1

What is the one step growth curve?

Answer 1

growth curve used to study viral replication by observing the time of a complete process of infection by a given virus in a permissive host cell until the release of new viral progeny
- one cycle of growth is observed

Question 2

What 7 steps are taken to prepare viruses for one step growth curve analysis?

Answer 2

1. inoculate cell culture with virus
2. incubate virus-infected cells in CO2 incubators
3. examine the viral infected cells daily under inverted microscope
4. harvest cell lysates at various time intervals
5. serially dilute the collected cell lysates
6. do plaque assay and calculate the number of plaques at each time point
7. analyze data and create viral one step growth curve

Question 3

What are the 2 main phases of the viral one step growth curve?

Answer 3

1. ECLIPSE PHASE: time from entry of the virus into the host cell until the assemble of new progeny viruses (1-20 hrs)
2. EXPONENTIAL GROWTH PHASE: number of produced viruses increases exponentially until reaching a plateau where no more viral particles are produced

Question 4

How does the bacterial growth curve compare to the viral one step growth curve?

Answer 4

BACTERIAL - lag phase, logarithmic phase, stationary phase, death

VIRAL - attachment, eclipse, assembly/maturation, release

Question 5

Despite the high variability among viruses, they replicate in a standard of 7 consecutive steps. What are they?

Answer 5

1. attachment/absorption
2. penetration
3. uncoating of envelope/capsid
4. synthesis of viral proteins (translation)
5. synthesis of viral nucleic acids
6. assembly and maturation
7. release of progeny virus

Question 6

How do viruses attach to the target cell?

Answer 6

ligands on the viral surface, called virus attachment proteins, bind to the receptors (proteins, glycoproteins, carbohydrates, lipids) on the plasma membrane of the target cells

Question 7

What is tropism? How is it affected by the presence of viral receptors? What are the 2 types of tropism in viruses?

Answer 7

the affinity of a virus to attach to specific target cells over other ones - the way the virus responds to external stimuli in order to attach to infect cells

viral receptors can be the primary determinant of tropism

1. TISSUE TROPISM: virus is more likely to bind to cells of specific tissues more than others (measles and skin cells; mumps and salivary gland cells)
2. SPECIES TROPISM: virus is more likely to bind to cells of a certain species more than other species (BHV-1 and cattle)

Question 8

How do Influenza viruses attach to target cells? HIV?

Answer 8

hemagglutinins on the viral cell bind to sialic acid on the target cells (sialic acid is rich in respiratory cells!)
GP-120 on the viral cell binds to CD4+ receptors on the target cells (on immune cells!)

Question 9

What 5 factors affect the efficacy of virus attachment to target cells?

Answer 9

1. density of receptors on the host cells
2. density of ligands on the surface on the virus
3. ratio of virus/host cells
4. temperature and pH
5. presence of specific ions, like calcium

Question 10

What tissues do neurotropic, pneumotropic, viscerotropic, dermotropic, and pantropic viruses target? What are some examples of each?

Answer 10

NEUROTROPIC: nervous tissue, brain and spinal cord; Rabies, AEV
PNEUMOTROPIC: respiratory tissue, lung, trachea, bronchi; influenza, rhinovirus, SARS-CoV-2
VISCEROTROPIC: GIT tissues; Rotavirus, NDV, Enterovirus
DERMOTROPIC: skin; Poxvirus, LSDV, HPV, cattle papillomavirus
PANTROPIC: many tissues and organs; SARS-CoV-2, NDV, YFV

Question 11

How do Picornaviruses inject their genomic RNA into their host?

Answer 11

create a pore into either at the plasma or endosomal membrane and release their RNA

Question 12

What are 2 strategies for keeping viruses from attaching to host cell receptors?

Answer 12

1. removal of cellular receptors on host cell (neuraminidase can remove sialic acid)
2. application of monoclonal antibodies to attach to cellular receptors and block viruses

Question 13

What are the 4 overall steps of endocytic viral entry/penetration via vesicle formation?

Answer 13

1. binding of the virus to the host cell receptors
2. invagination of the membrane
3. formulation of an endocytic vesicle
4. release of the virion into the cytoplasm
   (L)

Question 14

How do enveloped viruses undergo receptor-mediated fusion?

Answer 14

1. attach to host cell
2. viral and cellular envelopes fuse
3. capsid contianing viral nucleic acids is released inside the host cell
4. viral envelope forms a patch on the plasma membrane of the host cell
   (R)

Question 15

How do naked viruses tend to enter host cells? What are the 3 steps?

Answer 15

receptor-mediated endocytosis

1. viral ligand-cell surface interaction causes a clathrin-coated pit formation/invagination at the cell surface
2. the clathrin-coated pit (CCP) buds off to form vesicles
3. CCP sheds and the virus containing vesicle fuses with lysosomes

Question 16

What is the environment like inside the endocytic vesicle holding the virus during viral penetration? Why?

Answer 16

low pH and the presence of proteases

dissociated the capsid proteins, allowing for viral nucleic acid release

Question 17

What is the objective of viral uncoating?

Answer 17

make the viral genetic material accessible to the host’s cellular transcription and translation machineries to initiate the downstream viral replication steps

Question 18

How do the 2 types of viruses uncoat themselves?

Answer 18

1. ENVELOPED VIRUSES undergo COMPLETE UNCOATING where the virus fuses with the plasma membrane of the host cell, the nucleocapsid is released directly into the cell cytoplasm, and transcription starts immediately
2. NAKED VIRUSES remove viral capsid proteins and the viral genome performs all function without being released from the viral core

Question 19

Mechanism of viral fusion with host cells:

Answer 19

“drill” enzymes create a pore in the host plasma membrane and negative pressure pushes viral genome into host cells

Question 20

Upon virus entry into the cell, what 2 things happen?

Answer 20

1. production of viral structural proteins and enzymes
2. production of the viral genome

Question 21

In what 2 ways can the viral genome be classified in respect to continuity?

Answer 21

1. monopartite: linear genomes
2. multipartite: segmented genomes

Question 22

What is the point of the genome replication step during viral replication?

Answer 22

creates for genomic DNA or RNA for the progeny virus

Question 23

Where do DNA viruses multiply within the host cell? An exception?

Answer 23

within the nucleus and use the host cell’s DNA-dependent DNA polymerase

Poxvirus replicated in the cytoplasm and has its own DNA polymerase

Question 24

Where do RNA viruses replicate within host cells? What are 2 exceptions?

Answer 24

within the cytoplasm

1. Retroviruses
2. Influenza viruses

Question 25

How are RNA viruses able to replicate in the host's cytoplasm? How is this done with negative sense and positive sense strands? What is unique about Retroviruses?

Answer 25

RNA viruses have their own RNA dependent RNA polymerase NEGATIVE SENSE strands are prepackaged with the genome in a capsid and replicated into a complementary strand before translation POSITIVE SENSE strands are directly translated the Retrovirus has its own reverse transcriptase

Question 26

How do DNA viruses undergo transcription?

Answer 26

utilize the cellular DNA-dependent DNA polymerase - dsDNA can be directly transcribed into mRNA using cellular DNA polymerase II - ssDNA must first have cellular DNA polymerase I create a complementary strand to make it double-stranded, and then transcription into mRNA can occur using cellular DNA polymerase II once mRNA is made, translation can occur

Question 27

How do positive sense single-stranded RNA viruses undergo translation?

Answer 27

the viral genome itself acts as RNA, so it is directly translated into proteins using an RNA-dependent RNA polymerase

Question 28

How do negative sense single-stranded RNA viruses undergo translation?

Answer 28

virus must utlilize its own RNA-dependent polymerase, a transcriptase, to creat a complementary (positive) strand of RNA first, and then it can be translated by the same enzyme into viral protein

Question 29

How do double-stranded RNA viruses undergo translation?

Answer 29

its segments of RNA are translated by their own RNA-dependent RNA polymerase that is able to convert each segment into mRNA that can be translated by the RNA-dependent RNA polymerase

Question 30

What are ambisense viruses? How do they undergo transcription and translation?

Answer 30

viruses containing a genome that has RNA that is both positive and negative sense these viruses use their own RNA-depdendent DNA polymerase (reverse transcriptase) to turn the (+)ssRNA segments into (-)ssDNA, from there, the DNA is able to be transcripted into mRNA using cellular RNA polymerase II and translated into viral proteins

Question 31

Summary of virus replication strategies:

Answer 31

Question 32

Where does assembly of the viral nucleocapsid occur? How does this happen?

Answer 32

at the site of nucleic acid replication, intracytoplasmically or intranuclearly - capsomeres are produced from one or several proteins and bound together - nucleic acid genome is packaged along with other essential virion components - capsid then encloses the nucleic acids to form the nucleocapsid

Question 33

How are viral envelopes acquired? Where is the nucleocapsid located?

Answer 33

viral glycoproteins migrate from the ribosomes via ER toward the cell membrane and grouped into cylindrical spikes, allowing the virus to bud off underneath the plasma/nuclear membrane of host cell

Question 34

What are the 2 ways that viruses are able to release from infected cells based on the type of virus?

Answer 34

1. naked viruses tend to cause the host cell to lyse (aka cytopathic viruses); caused by accumulation of the viral particles in the nucleus and cytoplasm, triggering of apoptotic pathways, or cell necrosis due to the viral release of lysosomal enzymes 2. enveloped viruses tend to bud off the host cell by moving into the extracellular fluid while covered with the plasma membrane of the host cell ---> undergoes second replication cycle in a new cell

Question 35

Which of the following is not true regarding viruses? a. may be DNA, RNA, single, or double-stranded b. all are obligative intracellular parasites c. all have a protein capsid and envelop d. all are true

Answer 35

C

Question 36

Viruses are different from bacteria because... a. viruses do not replicate by binary fission b. viruses are obligate intracellular parasites c. viruses are sensitive to antibiotics d. A and B

Answer 36

D