Microm 445 Exam 1

Discovery of virus - tobacco mosaic disease

Virus propagate in host cells but not sap

Animal cell culture

breakthrough for virology

Physical characteristics of viruses

Genome either RNA or DNA

Nucleic acid enclosed in protein coat

Viruses are not living organisms but still evolve:

Obligate intracellular parasites

No independent energy metabolism

Require host cell protein synthesis to make progeny

Do not grow or divide - rather are assembled

Emerging infectious disease (EID)

Disease caused by pathogen that has not before been observed within a population or geographic location

Re-emerging infectious disease

caused by an established pathogen that was once controlled by begins appearing at a higher incidence

Components of Naked Virus

Capsid - surround nucleic acid

Nucleic Acid - RNA/DNA genome

Capsomeres

subunits that make up capsid

Protomers

individual protein subunits of a capsomer

Nucleocapsid

capsid and genome together

Components of Envelope Viruses

Capsid - surround nucleic acid

Nucleic Acid - RNA/DNA genome

Envelope - lipid bilayer surrounding capsid

Matrix - proteins that attach capsid to envelope

Surface proteins: proteins inserted in lipid envelope

Baltimore Classification

how viruses produce mRNA from their genome

mRNA is (+)RNA

ssDNA in same polarity - (+)

Complements are (-) strands

Ambisense means both (+) ORFs and (-) ORFs on same ss genome segment

X-ray Crystallography

used for small or large molecules

getting crystal can be difficult

diffraction pattern used to make map in which peptide sequence is fit

high resolution provide atomic detail

CryoEM Reconstruction

1. cryoplunge samples

2. collection electron micrographs

3. digitally select particle images

4. 3D image processing

5. Analyze and model structure

Helical capsids

Repeating identical units

Icosahedral capsids

• Most efficient way to get a symmetrical shell with maximum internal volume from non-symmetrical building blocks

• 20 hexons, 12 pentons

Poxviral structure

Viral culture

• Primary cell culture: from organ tissue

• Cell lines: cells from tumors

• Organoids: more complicated mixture of cells

• Embryonated chicken eggs

• Not all viruses can be cultured!

Cytopathic effect (CPE)

• changes in cell morphology due to viral infection

• detected by light microscopy

• focal area of CPE is a plaque

Multiplicity of infection (MOI)

Number of some viral measure per cell

Direct Contact - Horizontal

Saliva/mucus, blood, genital secretions, mucosal surfaces, skin

Direct contact - vertical

• Pregnancy - virus must cross placenta to infect fetus

• Delivery - virus in birth canal or blood

• Breastfeeding - virus present in milk

Airborne/fomite transmission

• airborne particles remain suspended in air

• fomites: inanimate objects that can spread virus by touch

Fecal/oral transmission

• Portals of exit and entry (rarely direct)

• Water and soil may be vehicles

Vector Transmission

• Vector: living organism that can transmit infectious disease between humans from animals or humans

• Mosquitos/ticks

Zoonoses

any disease or infection that is naturally transmissible from vertebrate animals to humans

R0 (R naught)

• basic reproduction number

• number of people each infected person will infect in a susceptible population

• product of transmissibility, duration, and average rate of contact

• greater than 1: spread throughout population

• less than 1: virus will die out

Viral properties - Enveloped viruses

• Generally more fragile

• Do not persist very long outside of host cell

• inactivated by heat, drying, detergents, low pH

Viral properties - Non-enveloped viruses

• Generally more stable

Portals of Entry

Skin - highly effective barrier (dead cells), require bypass of skin

Mucous membranes: most common route, single layer epithelium and damp environment

Tropism

capacity of a virus to infect specific cells, tissues, or species

Primary Infection

Virus infections initial susceptible cell

• overcome host defenses and environment

• localized spread by cell to cell spread of limited diffusion

• some viruses remain localized to site of entry

Systemic Spread

• amplification at the primary site of infection to body

• Route 1: bloodstream and lymphatic system (either in cells or liquid blood/lymph)

• Route 2: nervous system (brain via blood-brain barrier, CNS via peripheral nerves)

Viremia

virus in the blood

Steps of Viral Replication

1. Attachment

2. Penetration

3. Uncoating of nucleic acid

4. Viral protein expression

5. Replication of viral genome

6. Assembly

7. Release

8. Maturation

Attachment

Virus attachment protein (VAP) binds to receptor on surface of cell

• Envelope viruses have glycoprotein spikes in membrance that act as VAP

• Naked viruses use proteins on the capsid

Cellular receptors

Normal cellular molecules that the virus evolves to bind to

• major determinant of tropism

Co-Receptors

• two-step process of binding and penetration

• Step 1: cell attachment through binding primary receptor

• Step 2: Co-receptor binding triggers cell entry

Penetration

• Two possible sites: plasma membrane or intracellular vesicle

• Virus in vesicle is still outside and must escape from vesicle

• Naked viruses may poke hole in host membrane and release genome into cytoplasm or lytic protein that disrupts host membrane (triggered by low pH in endosome!)

Fusion (penetration)

• Necessary for enveloped viruses

• can occur at cytoplasmic membrane or endocytic membrane

• Requires fusion proteins to join cellular membranes (may be VAP)

Uncoating

• when capsid starts to come apart after interaction with the cell

• Releases the genome

• Overcomes most proximal barrier between genome and replicative niche imposed by the capsid

• both naked and enveloped viruses uncoat

(+)ssRNA viruses

• genome functions as an mRNA

• often encode a single protein that is cut into smaller proteins

• cleavage by a protease is good target for antivirals

(-)ssRNA viruses

• Genome cannot function as an mRNA

• Must encapsidate a viral RNA-dependent RNA polymerase to convert (-) RNA to mRNA

Retroviruses

• Have (+)ssRNA genome that is converted to DNA before transcription and translation

• Enzyme reverse transcriptase

Protein Expression

• Viruses overcome “one gene one protein”

• temporal patterns to gene expression - early vs late

Assembly

May be in nucleus or cytoplasm or other compartments

Maturation

• stage of replication cycle where virus becomes infectious

• often involves structural changes in viral particle (proteases/enzymes)

• maturation can occur inside cell or after release

Release

• Lytic viruses (non-enveloped): infected cell breaks open and releases the virus

• Enveloped viruses: acquire their lipid membrane as the virus buds out of the cell (usually cytoplasmic membrane, can be nuclear or golgi membrane)

Viral Pathogenesis

the process by which viruses produce disease in the host

• advantages and disadvantages, may cause no disease

• mechanisms: directly killing cells, altering cellular functions, stimulating an immune response

Stages of Infection

1. Incubation period - interval between infection and onset of illness

2. Prodromal phase - start of nonspecific symptoms

3. Illness - symptoms of particular disease, may have unique pathognomonic symptoms

4. Decline - symptoms improve due to immune function

5. Convalescence - symptoms resolve

Asymptomatic

an infection that does not cause any symptoms of disease

• dictated by immune response

• still able to transmit virus

Sequelae

a chronic complication of an acute infection

• symptoms that linger after the viral infection is cleared

Virulence

the relative capacity of virus to cause disease

• only applicable to closely related viruses

• mutations can make a virus more or less virulent

• host factors can affect virulence