Exam 1 Flashcards
What are Koch’s postulates?
- The pathogen must be present in every case of the disease
- The pathogen must be isolated from the host with the disease and grown in pure culture
- The specific disease must be reproduced when a pure culture of the pathogen is inoculated into a healthy susceptible host
- The pathogen must be recoverable from the experimentally infected host
Who created the smallpox vaccine? When?
Edward Jenner
1700s
What is the tobacco mosaic virus?
A virus that spoils the tobacco plant
What did Adolf Mayer and Martinus Beijerinck do?
They studied the tabacco plant and discovered it was not a bacteria nor a fungus
What experiment did Mayer and Beijerinck do on the tobacco plant?
Mayer and Beijerinck
1. took the leaves and ground them up
2. extracted the juice
3. smeared it on a leaf of a healthy plant
4. healthy plant got sick
This follows Koch’s postulates without realizing it
How did Mayer of Beijerinck do to determine it wasn’t a fungus or bacteria that was harming the tobacco plant?
Filtration Take a ceramic with small holes and run the plant leaf juice through it and then tested to see if it could still make the plant sick. Removed any fungi = still contagious Removed any bacteria = still contagious Therefore it must be something else
What is the difference between a pathogen and a toxin? (2)
A toxin isn’t living and a virus is. This effects
- Transmissibility (a virus is easily transferred from one organism to another)
- Maintenance of Potency (a virus will maintain potency no matter how many times it is transferred, but a toxin will decrease in toxicity.)
It took the invention of what instrument to visual viruses?
Electron Microscope
Why can we learn about our own cells using viruses?
Since we are the host for viruses they must conform to our bodies and our machinery
What is life?
Life is the condition that distinguishes active animals and plants from inorganic matter, including the capacity for growth, functional activity, and continual change preceding death -OED
What are some reasons (3) that make the claim “viruses are living things” a little grey
- Some viruses can be crystallized and kept in this form for a very long time
- Viruses are only active after they enter a living cell
- They lack all the metabolic processes needed to generate energy and the machinery to make proteins
Did viruses evolve out of ‘rogue’ pieces of DNA? Or did they originate independent of their host cells and only later acquire infectivity? What kind of evidence exists for this?
- Virus First Hypothesis
- Reduction Hypothesis
- Escape Hypothesis
https: //www.ncbi.nlm.nih.gov/pmc/articles/PMC3575434/
What are the 5 main structures of a generic virus?
- Envelope
- Tegument
- Capsid
- Glycoprotein ‘spikes’
- Genome (DNA or RNA)
What are viruses (definition)?
Subcellular, infectious agents that are obligate intracellular parasites
What is a virion?
A mature, extracellular virus
What is a genome?
An organism’s complete set of genetic instructions
What is the nucleocapsid?
A protein coat containing the genome
What is the envelope?
A lipid envelope that surrounds the nucleocapsid
Where are glycoproteins located?
They are located in the envelope and are transmembrane
What is the most likely shape of a virus? Why?
An icosahedron
It is the closest viruses can get to a sphere and a sphere is the most efficient volume to surface ratio
How many sides does an icosahedron have?
20 sides
What are capsomeres?
Repeating protein subunits that make up the capsid
What is the drawback of an icosahedron?
The genome is limited in size
What is the function of a virus particle? (ie why not have a naked genome) (4)
- Protects the genome from being degraded
- Aids in host recognition, binding, and entering
- Fidelity; viral genome recognition and collection
- Self-assembly
What are some of the uniform structures viruses will take? (2) Why do they take these structures?
- Helical Nucleocapsid (limitless genome)
- Icosahedron (best volume to SA ratio)
The capsid is usually regular due to the small number of repeating proteins
What is the capsid?
The protein coat of the virus which is made up of capsomeres
What are the capsomere encoded by?
They are encoded for by the viral genome.
Constitute some of the structural genes (late-stage)
Are viral genomes limited or unlimited in their coding capacity? What is the implication of this?
They are limited in their coding capacity
Virons may be formed using a limited number of different proteins
Why is it that capsids can self-assemble? (3)
- Small amount of genetic information
- Assembly and disassembly are low energy processes
- It is self-correcting
Talk me through the process of self-assembly
- Virus infect the cell
- Virus begins replication of the genome
- In the late stage genes = formation of capsomeres
- Caposmeres have an RNA binding domain that recognizes the encapsidation sequence of the viral RNA and binds to it
- The capsomeres then “click” into place with one another due to affinities and nucleation events and create the capsid
- Some = proteolysis of the capsid proteins into their mature form triggers assembly
How does Palanoma Virus Vaccine work?
Deliver an empty capsid
The body recognizes the capsid from the glycoproteins and develops antibodies
But there is no viral genome to actually make you sick
What do packaging signals do?
Direct incorporation of viral genomes into virions
What are core proteins?
Accompany the viral genome inside the capsid
Possible examples: RDRP, Reverse Transcriptase
What are scaffolding proteins?
They help in virion assembly but are not incorporated into the mature virion
How is the tobacco mosaic virus organized?
Tobacco Mosaic Virus is organized as a Helical nucleocapsid
How many subunits does an icosahedron have? How many types of symmetry?
20 faces
60 subunits
2-fold (at vertex), 3-fold (at triangle), and 5-fold symmetry (at axis)
Viruses can vary in size, what are the 4 common sizes?
- 20 nm (parovirus)
- 80 nm (adenovirus)
- 200 nm (phycodavirus)
- 500 nm (Mimivirus)
Can viruses be infected by other viruses?
Yes, if they are large enough they can be infected by another smaller virus
What is the minimum number of subunits required to make an icosahedron shape?
60
Viruses can differ in two ways when it comes to capsids, what are they?
- Size of capsomeres
2. Variety of capsomeres
How many vertices, faces, and edges does an icosahedron have?
12 vertices
60 faces
30 edges
What is the dumbbell shape? What is an example of a virus that has this shape?
Ovoid particle with a dumbbell-shaped (constricts in the middle) nucleocapsid
Pox Virus
What is the bullet shape? What is an example of a virus that has this shape?
Elongated end and rounded end
Coiled helix
Ex. Rhabdovirus
What is the envelope of enveloped viruses made of? What do they contain?
Lipid bilayer membranes
They contain viral glycoprotiens
How is budding driven in enveloped viruses?
Budding is driven by interactions between viral protiens
How is fusion driven in enveloped viruses?
Fusion is driven by the fact that the lipids on the viral envelope and the lipids of the host membrane are both hydrophobic
What property of enveloped viruses are we exploiting when we wash our hands?
When we wash our hands we use detergent (soap) which is are long chains of hydrophobic carbons. The hydrophobic carbons fuse with the viral envelope and ruin its membrane
What is intercalation?
The process of fusing two membranes or inserting something into the membrane
What property of enveloped viruses can also be exploited?
Enveloped viruses cannot be dried out because the membrane will fall apart therefore they survive better in more humid environments
What are fomites? How long can enveloped fomites last? How long can non-enveloped fomites last?
Fomites are virus particles left on nonliving things
Enveloped viruses can only last hours
Nonevneloped viruses can last years
What is an example of a non-enveloped virus?
Adenovirus = common cold
How are non-enveloped viruses often shed?
Non -enveloped viruses are often shed in feces
Which is more fragile, an enveloped or non-enveloped virus?
An enveloped virus
Define tropism
The affinity of a virus for a particular cell type
It is determined by the type of receptor on the cell
What can determine an enveloped virus’s tropism?
The glycoproteins embedded in the envelope
What is hemagglutinattion? What is responsible for the hemagglutination properties of enveloped viruses?
Hemagglutination is an assay to determine the antibodies of the virus
It is determined by the glycoproteins embedded in the envelope
What mediates the fusion of the viral membrane with the cell membrane for enveloped viruses?
Glycoproteins embedded in the envelope
What are often the primary targets of the immune system of enveloped viruses?
Glycoproteins embedded in the envelope
Can glycoproteins possess enzymatic activity? If so, provide an example
Yes
Neuraminidase of the orthomyxoviruses
What is an example of an enveloped virus?
Herpes simplex virus Type I
What is the shape of the adenovirus?
Three copies of the capsomere protein (hexon protein) come together to form the hexon
12 vertices with projections of long fibers that end in spherical extensions used for viral attachment to host cells
What about bacteriophages make them unconventional in shape? What are their three additional appendages?
Icosahedral head with a helical tail
- Baseplates
- Collars
- Tail Fibers
What is the tropism of bacteriophages?
Bacteria
What are baseplates?
On bacteriophages
Cell adhesion molecule (unproven to exist?)
What are collars?
On bacteriophages
Attaches tail fibers
Helps with viral assembly
What are tail fibers?
On bacteriophages
Improves efficiency of infection by facilitating viral entry
What are the 3 criteria viral classification are based on?
- Molecular architecture
- Genetic relatedness
- Host organism
How are viruses grouped?
Viruses are grouped into species, genera, and families
Viruses may be divided into three broad classes. What are they?
- DNA Viruses
- RNA Viruses
- Retroviruses
What are the two subdivisions within DNA viruses?
ssDNA
dsDNA
How does replication occur in DNA viruses? What type of polymerase does it use?
Replication occurs by direct DNA to DNA copying
They are the same as us, so they can use the same cellular machinery
Uses DDDP (already made in our cells)
What is an example of a ssDNA virus?
Banana Bunchy Top Virus
What is an example of a dsDNA virus?
Human adenovirus
What are the two subdivisions within RNA viruses?
Plus strand RNA
Minus strand RNA
What is an example of a plus-strand RNA virus?
Poliovirus
What is an example of a minus-strand RNA virus?
Measles virus
What are arenaviruses?
Viruses that have both forms of RNA (plus and minus-strand)
All viruses with negative-strand RNA genomes have what kind of nucleocapsids? Is the genome fragmented or nonfragmented?
Helical nucleocapsids
Some have fragmented genomes
How does replication occur for plus-strand RNA? What polymerase does it use?
Plus strand RNA can be read normally
Its genome encodes for RDRP
So it goes + strand to complementary - strand and then to an exact copy of the + strand
What does RDRP stand for? What does it do?
RNA Dependent RNA Polymerase
An enzyme that catalyzes the synthesis of RNA from an RNA template
What is DDRP? What does it do?
DNA Dependent RNA Polymerase
Catalyzes the synthesis of transcription of RNA from a DNA template
What is DDDP? What does it do?
DNA Dependent DNA Polymerase
Catalyzes the synthesis of DNA from DNA template
How does replication occur for minus-strand RNA? What polymerase does it use?
Must first convert from minus-strand to plus-strand before translating (?) genome
Uses RDRP but must bring it already made in order to translate the minus strand to the plus strand!
What do retroviruses must have in the capsid?
Reverse transcriptase
How does replication occur for retroviruses?
Converted RNA to DNA using reverse transcriptase, then use DNA to create proteins
Are viral genomes haploid?
Most viruses are haploid with the Exception of retroviruses which have diploid genomes
What is the advantage of having a segmented genome?
It allows for genetic recombination
What is an example of a retrograde virus?
Hepatitis B Virus
What does it mean to have a segmented genome?
Analogous to chromosomes
Does each subunit of segmented genomes have to be present in each capsid?
Yes for animal viruses
No for plant viruses (though the complete genome is needed)
What are the seven classes based on the Baltimore Classification?
dsDNA ssDNA dsRNA \+ssRNA -ssRNA RNA reverse transcribing DNA reverse transcribing
The evolutionary origin of viruses
RNA world
Viroids and RNA viruses may have originated in the RNA world
Transition to DNA based world
Small and medium DNA viruses could have arisen as independently replicating genetic elements
Large DNA viruses could have evolved from cellular forms that became obligatory intracellular parasites
Define MOI
Multiplicity of Infection
Number of virus particles infecting each cell
It is a ratio (# viral particles/#cells)
Define Pfu
Plaque forming units = clear spots on the plate because the infected cells have died
Tells us the number of infectious virus particles
Define Pfu/cell
Number of infectious virus particles infecting each cell
Eclipse Period
The time between infection and the appearance of a mature virus within the cell
You cant see the virus until late-stage genes are expressed and viral particles start to appear
Rise Period
The period of grown after the eclipse period
If there are 1,000,000 viral particles and you put them on 100,000 cells. What is the MOI?
10
What are the steps in a plaque-forming assay?
- Infect cells in multiple tissue culture dishes at the same MOI
- Lyse the cells at each hour
- Take the cell lysate and growth medium. Plate increasing amount of host cells
What happens during the eclipse period?
The viruses attached to the cell, then the virus falls apart - releasing its genome. The viral genome is not always infections (it is not fully functional??) and therefore you can’t see it
What is an LD50 assay? What does it count? What does it not count?
Answers the question: how much of this virus is needed to kill 50% of X?
It does not count the number of infectious particles
What is a hemagglutination assay? What does it count? What does it not count?
Looks at the amount of RBC clumping
It does not count the number of infectious particles
Which step of the virus life cycle is best to inhibit?
?
What is an example of a drug that inhibits viral attachment?
Tamiflu
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7149618/
What is an example of a drug that inhibits viral penetration?
Enfuvirtide
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7149618/
What is an example of a drug that inhibits viral uncoating?
Amantadine
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7149618/
What is an example of a drug that inhibits viral gene expression?
Azidothymidine
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7149618/
What is an example of a drug that inhibits viral assembly?
Verdinexor
https://www.frontiersin.org/articles/10.3389/fmicb.2015.00517/full
What are the 6 stages of the viral life cycle?
- Attachment
- Penetration
- Uncoating
- Gene expression
4a. Early genes
4b. Late genes - Assembly
- Egress
Why is learning the attachment site so important?
It can tell you about the pathology, which cells will be affected, treatment options, and viral therapies
Talk me through attachment
- Virion binds to accessory receptor (low affinity)
- This slows viral movement and allow the virus to try to find the high-affinity receptor
- Virion binds to the high-affinity receptor (binding occurs between one of the viral surface proteins and a receptor)
- This arrests viral movement
Does each virus have a specific receptor?
Yes
Can different viruses share the same receptor?
Yes
Can viruses of the same family have different receptors?
Yes
Can viruses of the same family have different receptors?
Yes
Viruses often use members of the _________ as high affinity receptors
Ig Superfamily
Is a tropism narrow or broad?
It can be either
What are some examples of viral receptors? Also say if that would result in a narrow or broad tropsim (3)
- GLUT-1 ; broad (on all cells)
- CD21 ; narrow (only B cells)
- CD4 ; narrow (only T cells)
What is the receptor for SARS-CoV-2?
ACE2
If viruses can use different species as a host, does that mean their receptor is different in each host?
No, it will be the same receptor no matter the species
Define Arbovirus
Virus that infect insects
Talk me through 3 ways to discover what the viral receptor is
1.
What are 3 ways to discover what the viral receptor is
- Use an enzyme to cleave the receptor
- Use antibodies
- Knockin gene for receptor in a species that doesn’t normally have it
- Knockout gene for the receptor
What are 3 ways to discover what the viral receptor is
- Use an enzyme to cleave the receptor
- Use antibodies
- Knockin gene for the receptor in a species that doesn’t normally have it
- Knockout gene for the receptor
Talk me through an experimental setup for using an enzyme to cleave a potential viral receptor
Control group:
- Plate cells
- Treat with viral particles
- Count pfu
- Calculate MOI
Test group:
- Plate cells
- Treat with the enzyme to cleave potential viral receptor
- Treat with viral particles
- Count pfu
- Calculate MOI
If the MOI is significantly smaller in the test group then it is likely that the viral receptor was correctly identified. In the test group, the viral receptor would have been cleaved by the enzyme thus inhibiting viral attachment and therefore viral infection resulting in fewer pfu.
Talk me through an experimental setup for using antibodies to identify a potential viral receptor
Control group:
- Plate cells
- Treat with viral particles
- Count pfu
- Calculate MOI
Test Group:
- Plate cells
- Treat with antibody against potential viral receptor
- Treat with viral particles
- Count pfu
- Calculate MOI
If MOI is significantly smaller in the test group than the control group, it is likely the correct viral receptor has been identified. The antibodies against the viral receptor would bind to the viral receptor creating a physical barrier. The virus would not be able to bind to the receptor due to the antibody blocking it, this would block viral attachment and thus block infection, decreasing the MOI.
Talk me through an experimental setup for using knock-in genes to identify a potential viral receptor.
Control group:
- Raise the mice
- Infect with the virus
- Monitor for symptoms
Trest Group:
- Knock-in gene for the potential viral receptor
- Raise the mice
- Infect with the virus
- Monitor for symptoms
This experiment hinges on the fact that mice or the organism of choice does not normally code for the potential viral receptor. The control group should experience no symptoms since the viral receptor will not exist and therefore infection cannot take place. The test group will experience symptoms since their cells will express the viral receptor.
Variant of Concern vs Variant of Interest
Variant of Concern: Variants currently in circulation that have increased rate of spread, increased severity, decreased vaccine efficiency, or decreased treatment efficiency
Variant of Interest: Variants that have genetic changes in parts of the genome responsible for virus transmission, disease severity, immune system evasion, and treatment effectiveness
What is penetration?
The introduction of the viral genome into the cytoplasm of the host cell
The genome could be in the form of the nucleocapsid or
Talk me through penetration of an enveloped virus
Fusion!
- (Fusion domain on a viral surface protein)
- Glycoprotein protein-ligand bind to viral receptor
- Fusion of viral and cellular envelopes
- Nucleocapsid released inside of the cell
- Viral envelop forms patch on plasma membrane
Talk me through receptor mediated endocytosis
- Attachment
- Formation of clathrin-coated pit
- Uncoating of vessel
- Fusion with endosome
- Acidification
- Disaggregation of viral nucleocapsid
- Release of the viral genome
What are the two ways an enveloped virus can penetrate the cell?
- fusion
2. receptor-mediated endocytosis
How do non-enveleoped viruses penetrate the host cell?
Unclear
Pass through via interactions with viral proteins and the receptor
Talk me through uncoating - what is it? What is the mechanism?
When the capsid falls apart
Mechanism is unclear
- Ribosome extracts the RNA –> capsid disintegration
- Specific factors int he host cell –> uncoating
What makes the penetration of bacteriophages different than other viruses? What is the mechanism of penetration?
Bacteriophages have to penetrate the rigid cell wall of bacteria
The tail of the bacteriophage attaches to the surface and drills a hole into it. This is also mediated by enzymes that eat into the cell wall
The DNA is delivered into the bacteria
Talk to me about gene expression
The synthetic phase:
Replication of genome, synthesis of viral proteins, formation of viral particles
Where does viral genome replication occur? (2)
- Cytoplasm of host cell
2. Nucleus of host cell
Viral protein is synthesized using ___ ribosomes and ___ tRNA in the cytoplasm
Host
What are the two phases of gene expression?
- Early Genes
2. Late Genes
Is gene expression tightly regulated?
Yes
What are some characteristics of early stage genes? (3)
- Non-structured
- Regulatory in nature
- Required for Viral Replication (ex Transcription Factors)
What are some characteristics of late stage genes? (2)
- Structural
2. Required for viral morphogenesis = capsid formation
How is viral gene expression regulated?
It is temporarily regulated
How do you end up with such high production rates of late stage genes?
The late mRNA is primarily transcribed from progeny genomes
There are multiple copies of progeny genomes –> greater amount of proteins
Talk me through the replication cycle of dsDNA viruses
- Virsus enters the cell
- Uncoats and releases genome
- DNA enters the nucleus
- Replication
- Assembly
- Egress
Talk me through the replication cycle of ssDNA viruses
- Virus enters the cell
- Uncoats and releases genome
- ss is converted to ds in the cytosol by viral proteins (DDDP)
- DNA enters the nucleus
- Replication
- Assembly
- Egress
Talk me through the replication cycle of plus-strand RNA viruses
- Virus enters the cell
- Uncoats and releases genome
- Translation to get RDRP
- Transcription to create the negative complementary strand
- Replication to create progeny positive strands
- Assembly
- Egress
Talk me through the replication cycle of negative-strand RNA viruses
- Virus enters the cell
- Uncoats and releases genome
- Create complimentary plus-strand RNA from RDRP
- Replication
- Assembly
- Egress
Talk me through the replication cycle of retroviruses?
- Virus enters the cell
- Complementary strand undergoes reverse transcription by RDDP
- Uncoats and releases genome
- Integrates into host DNA
- Replication
- Assembly
- Egress
T/F: Cell needs to be going through the cell cycle for the replication of DNA viruses
True
Since the replication cycle begins by translating the RNA genome to produce the enzymes for RNA synthesis, then ____
The naked RNA is infectious
What is an ambisense RNA virus?
One that contains both plus and minus-strand RNA
How do you know if a ambisense RNA virus will be infectious?
It depends on whether or not the positive side encodes for all necessary proteins such as RDRP
What is the stage of the life cycle of the virus when it become infectious?
Maturation
One or more of the capsid or envelop proteins may undergo what during maturation?
Proteolytic cleavage
Do all viruses egress the same way?
No
What are 3 common ways viruses egress?
- Enveloped viruses can bud out
- Infected cells can disintegrate
- Viruses can pass from cell to cell by syncytia formation
What is a drug that inhibits viral egress?
Tamiflu
