Virus Classification, Structure, and Replication Flashcards
how is a virus different from a bacterium
size
obligate intracellular parsite
how is a virus different for a toxin
toxin does not replicate
How did we discover that viruses were small
Dimitri IVanofsky showed that Tobacco Mosaic Virtus was able to pass through a filter while bacteria could not
then afterwards the electron micrograph
how did we discover the growth(non-toxin) of viruses
Martin Beijerinck showed that the titer of Tobacco mosaic Virus increased after infecting a plant
how was the bacteriophage discovered
Frederick W. Twort while trying to grow vaccinia virus (1915)
The first animal virus discovered
Foot and Mouth disease (1898)
The first Human virus discovered
yellow fever virus (1901)
research use of bacteriophage
instrumental in developing the field of virology and biology
are Virus autopoietic
no (obligate intracellular parasites
Theories for virus origin
Cellular origin
Autopoietic origin
the Cellular origin of viruses state that
viruses were once cellular components, but over time they evolved separately
the autopoietic origin of viruses state that
viruses, once autopoietic entities, became dependent on cells for replication
ways to classify viruses
Virus particle strucutre Genome (size, genes, copies) replication features Serology (antibody recognition) Stability
what defines Virus PArticle strucutre
Composition
Shape
Size
envelope or nonenvelop
What makes up the Nucleocapsid
RNA or DNA in a core that is protected by a protein coat (capsid)
tyoes of Nucleocapside strucutres
Helical
Pleomorphic
Icosahedral
Helical Nucleocapsid Structure
Genome coiled around with proteins surrounding it
pleomorphic nucleocapside strucutre
blob
the repeating portein subunites of the nucleocapside
Capsomere
Virus-modified cellular membranes acqueired upon exit from a host
Envelopes
what does exposure to lipid solvents in the lab (Alcohol, ether, acetone, freon, etc) do to enveloped viruses
makes them noninfectious because evelope proteins are important for the infectious cycle
what makes up the envelope of a virus
lipid bilayer
viral proteins and host cell proteins
is the shape of the envelope related to the shape of the nucleocapside
no relation (nucleocapsid can be any shape desired)
smallest and largest virus
18nm:
300nm: pox-virus family
advantage of small virus
taken up easier
move easier
advantage of large virus
can bring more stuff to aid in infection
possible types of Genomes of Viruses
DNA: double stranded, Single stranded
RNA: Double Stranded
Sing Stranded: plus sense, minus sense, or ambisense
plus sense ssRNA
ready for translation
minus sense ssRNA
must be copied before it can be translated
ambisense ssRNA
can be both minus or plus sense ssRNA
Virus Genome structures
Linear
Circular
Segmented
Diploid
Central Dogma
Replication of DNA
DNA transcribed into RNA or RNA reverse transcribed into DNA(viruses only
RNA translated to protein
replication via ssDNA hairpin
ssDNA folds to look like dsDNA right were DNA polymerase binds
Replication via dsDNA rolling circle
makes a copy with a lagging strand also to eventually give a double stranded DNA
replication via +ssRNA
genome enters, and is template for protein
these proteins made can be a polymerase that binds to RNA struutre, to make a negative sense to serve a template to make a bunch of offspring
Relication via -ssRNA
polymerase comes in with the virus to sit on -ssRNA to make the plus sense copies that are eventualy turned into proteins
advantage and disadvantage of large genomes
ad: more proteins
disadvantge: higher chance of detection, longer time, greater chance of error
what do most DNA viruses need that RNA viruses do not?
need transcption machinery
access to the nucleus
what do most RNA viruses need that DNA viruses do not
need an RNA depended RNA polymerase
steps of Virus Replication
Attachment Entry TRanscription Translation Replication Assembly Release
the binding of a virus receptor to a cellular receptor
Attachment
Cellular receptors(parts of the cell that actually have cellular roles) that are targets for viruses to bind
Singaling Molecules
Cell adhesion molecules
Transport molecules
do viral receptors mimnic cell receptor’s normal ligands
No
what do viral receptors look like
spike like projections on particle surface
what may be needed to a viral receptor to bind?
A co-receptor
how can you change receptor recognition
Genetic engineering
types of Genetic engineering to change receptor recognition
Changing to RGD sequeces
pseudotyping particles
do Viruses need to only bind to one receptor on a cell
no, some may need to bind to multiple
what determines Virus Tropism(host range)
attachment is a major determinant
what can virus’s infect
essentially all known forms of life, but have specific host ranges and not shared across more divergent host
what is a major factor in eradication of a virus
Host rnage
who could Smallpox infect
only human
who could Yellow fever infect
Mosquitoes and humans
pathways of how a virus can enter a cell
Receptor mediated endocytosis
Direct penetration of the plasma membrane
how do non-eveloped viruses enter the cell
No well understood
Pores
Membrane disruption
what is non-eveloped virus that enters cell through pores
Picornavirus
what is a non-eveloped cirus that enters cells through membrane disruption
adenovirius, reovirus
how does the enveloped (influenza) virus enter the cell
Via membrane fusion
HA protein allows for arrachment and fusion
this caused a drop in pH and attachment proteins undergo a confirmational change and virus lipid bilayer and cellular bilayer come close togther and mingle, creating a pore and virus gets in
what must a virus do once inside the cell to begin replication process
Uncoating
types of Virus replication
Nuclear replication
Cytoplasmic replication
where uncoating occures in nuclear replication
genome and remaining protein coat are transported to the nuclear membrane to deliver the genome to the nucleus
where uncoating occures in cytoplasmic replication
Relase of the genome into the cytoplasm
transportation of the genome to intracellular site of replication
where do many RNA virus replication in dytoplasmic replication
In membrane associated complexes
do dsRNA viruses release their genomic material from the entering particle
No
DNA and RNA nucleocapsides prefer what kinds of replication
DNA: Nuclear
Cytoplasmic: Cytoplasmic
Pruduction of mRNA templates for protein synthesis
Transcription
what do DNA viruses depend on for TRanscription
Cellular RNA polymerases
the genome of what type of virus can serve as mRNA
+ssRNA
how does +ssRNA create new transcriptis
use a -ssRNA template
what types of virus must bring their own polymerases into the cell for transcription
(-)ssRNA and dsRNA
what must a virus do to cellular transcription
must subvert the cells transcription
The production of proteins
Translation
what do all viruses need from the cell to produce protein
need ribosomes (no exceptions)
where can viral protein production be regulated
at the trascript (mRNA) level of translation level
what type of viral protein are made in high quantities
structural proteins
what type of proteins are only seen in the infected cell
non-structural proteins
Objective of genome Replication
make aditional genome copies
what does the order of viral replication depend on
Virus genome
(+)ssRNA has what as a template for translation
genome
how does (+)ssRNA make new genomes
Polymerase(made from the (+)ssRNA) makes (-)ssRNA copy as template for new genomes
what must be included with (-)ssRNA Virus particles for viral replication
Viral Polymerase
how is the Genome of (-)ssRNA replicated
through full-length (+)ssRNA intermediate
what does dsRNA contain for viral replication
Viral Polymerase
why must dsRNA stay inside the particle
induces innate immune response
how is mRNA made for dsRNA viruses
synthesized in particle and exported to the cytoplasm
how is viral genome replicated in dsRNA
mRNA serves (+) strand in virus genome, (-) strand synth during assembly
where does ssDNA and dsDNA go to do genome replication
must gain access to the nucleus
what dsDNA/ssDNA virus does not need to go to the nucleus to do replication
Poxviruses
why does the Poxviruse not need to go to the nucleus to do genome replication
virion contains necessary RNA polymerase and the genome encodes the DNA polymerase for replication
what does the ssDNA and dsDNA of a virus do once in the nucleus and why
prepares the cell for DNA replication (growth phase, dNTP production, replication machinery) to ensure that the genome ends up copied
Packaging new genomes into functional particles
Assembly
what aids in assembly
localize structural proteins
Genome contains packaging signals
localize structural proteins that aid in assembly
cellular viral “factories”
mechanisms of Assembly
Adenovirus
Reovirus
Retrovirus
when an empty protein coat imports a genome
adenovirus
when RNA is packaged during capside assembly
Reovirus
preassembly of virus on a membrane
Retrovirus
type of infections
Lytic: cell is exploded
Non-lytic: cell does not explode and may be continuously secreted as a result
how Viruses may be released
Lysis
Weak Lysis
Budding
what type of virus is best known for lysis
BActeriophages
when viral molecules rupture cellular membrane
Lysis
weak lysis depends on
membrane breakdown after cell death
what type of virus can do budding
only enveloped
when enveloped virus use cell membrane as the outer coat of the virus particle
Budding
what disadvantage come with targeting virus replication in the nucleus
Disadvantage: have to get across membrane
how does understanding a virus replication process help design antivirals
find the unique parts of the virus, minimizing illness
advantage and disadvnatage of lytic infectinos
advantage: dumps contents of cell with lots of viral products can infect other cells quickly, releases a lot of viruses
disadvantage: allows the immune sytem to respond to broken cells, hard to go undetected
how to measure growth curve of Viruses
One step growth curve
what is a one step growth curve
infect every cell at same time
every cell dies at end of infection
phases of Virus Replication
Eclipse
Exponential growth
Plateau
Eclipse phase
Attachment and uptake
Exponential growth phase
replication and assembly
plateau phase
cell death
uses of Viral Kinetics (one step)
Mutations
Cell entry
Process Design
time for one step growth
start of infection to beginning of plateau
time for one step growth of bacteriophage
30 min
time for one step growth of Vesicular stomatitis virus (VSV)
6 hours
time for Vaccinia one step growth
24 hours
productivity of a virus meaures
amplification of the virus
productivity of VSV
1:1000
productivity of Vaccinia
1:100
how do we initialy discover a virus
Disease in a host
Contaminant in cell culture
how do we confirm a virus
Purification and confirmation of disease (cell Culture is prefered)
where do we confirm a disease
Animals, eggs, or cell cultures
detection and Quantification methods of a virus
Infectivity
physical
Genome
what method is used to detect and quantify viruses that connot be cultivated
Molecular Methods
types of Infection assays
Cytopathic effect
Fluorescent Focus Assay
Plaque Assays
Infectious Dose
what are cytopathic effects
Cell Rounding
Syncytia Formation
Inclusion bodies
when viruses cause cell death that causes cells to round out and separate
Cell Rounding
what is formed when a virus has proteins on its surface that instigate membrane fusion that cause cells to fuse together and have multiple nuclei in one cells
Syncytia formation
how is a Fluorescent Focus Assay used to detect and Quantify Viruses in Infection Assays
Infect cells
Expose Virus Antigen
Stain with labeled antibody
Count areas that fluoresce
How is a Plaque assay used to detect and quantify viruses in infection assays
there is a Cell monolayer
Innoculate with a v. dilute virus
Use a new layer of auger to ensure that virus and cells do not move
Infected cells die leaving a clear area- a plaque
examples of infectious doses
ID50, IU50, TCID50
systems used to measure infectious dose
Tissue Culture, Eggs, Animals
how to measure infectious dose
inoculate with different dilutions of virus
Calculate concentration based on number infected
Types of particle assays
Electron Microscopy
Hemaggultinin assay
what does Electron microscopy show of a virus
a direct image of virus particles
compare to latex bead standard size
how to calibrate electron microscopes
latex bead standard
what type of virus is used in Hemagglutinin assays
Viruses that bind red blood cells (RBCs)
how to do Hemagglutinin assays
mix constant number of RBCs with various virus dilutions
if virus concentration is sufficient, a matrix of RBCs and viris is formed
matrix does not allow RBC to pellet out in a centrifuge
Genome assay types
Polymerase CHain Reaction
Southern (DNA) blots
Northern (RNA) blots
How a polymerase chain reaction works
DNA primer is specific to a virus
Amplify that one gene
sensitivity of Polymerase Chain reactions
Very sensitive
what is done in a southern and Northern Blot
Isolate DNA (Southern blot) or RNA (Northern blot) by electrophoresis-lebeled Use a labeled DNA probe to detect
types of Serological assays
Viral Neutralization
Enzyme link Immunosorbant Assay (ELISA)
Western (Protein) Blot
antibody binding to a virus does what
block infection
how to determine Virus Concentration
by the amount of antibody needed (Virus neutralization
what is an enzyme Immunosorbant ASSAY (ELISA
Antibody recognizes virus
Amplication by enzyme linked to antibody
Western blot
separate proteins by electrophoresis
Probe proteins using an antibody
does the way we quantify a virus give different answers?
yes
particle to PFU ration ranges from
1:1-1:10000
how could someone get a .3 PFU
if humans were a better candidate for infection than a plaque
