Micro - Viruses

Viruses are Nonliving Pathogens

• recent studies indicated about 40,000 viruses in mammals

  • 270 infect humans

  • ~320,000 mammalian viruses remain uncharacterized

• ~70% that infect humans are harbored in other animals

• viruses are extremely small, acellular & obligate intracell pathos

Viruses Exhibit diverse structural & genomic features

• can infect every branch in tree of life

• animal viruses infect animals & humans

• virion = single, infectious virus particles that have an exterior protective protein capsid & contain gene material

Viral Capsids

• protein shell that packages genome

• accounts for bulk of virion mass

• made of capsomere subunits

• most animal viruses have helical (hollow tube) or icosahedral capsids (3-dimensional polygons)

• deviated from 2 structures are complex capsids

• held together by protein-protein interactions based upon hydrophobic binding

Bacteriophages Capsid

• complex capsid structure

• usually have icosahedral symmetry

• often associated with additional complex structures that enable them to inject genome into target cells

Viral Envelopes

• lipid based envelope around capsid

  • arise from budding off host cell (take portion of cell mem with them)

• Naked viruses lack envelope

  • arise from lysing (bursting) host cell

• Animal viruses = envelope or naked

• bacteriophages lyse host, always naked

Viral Spikes (peplomers)

• many viruses have spikes that produce from viral capsid or envelope

• glycoprotein extensions help virus attach & gain entry to host

• only bind to specific factors on given host

• influzena frequently mutate & experience small changes in spike proteins

  • flu A spikes: Hemagglutinin, Neurominidase: Make up subtype name of flu variants

Viral Genomes

• most viruses have fewer than 300

• viral genes encode:

  • capsomere proteins

  • enzymes needed fro viral repli.

  • structural factors

• viral genomes can be either:

  • rna or dna

  • single or double

  • single or segmented

  • circular or linear

• viruses need to hijak a host cell to be able to make viral proteins so more virions can be made

Viral Genomes - dsDNA, ssDNA, ssRNA+, ssRNA-, dsRNA

• double stranded DNA virus (dsDNA)

  • viral dna is transcribed using host rna polymerases

  • mRNA is then translated → protein

• single strand DNA (ssDNA)

  • convert to double stranded form before transcription

• single strand positive rna (ssRNA+)

  • rna genome is complementary to mRNA

  • transcribed into mrna by rna-dependent rna polymerases (RdRPs)

• single stranded retroviruses

  • RNA genome made into → dna by reverse transcripts

  • dna is inserted into host dna

  • dna is transcribed into mrna

• double stranded rna genome (dsRNA)

  • transcribed to make mrna

  • requires ran-dependent rna polymerase

Viral genomes change over time

• virus exhibit a faster rate of genome change than do living because

  • quick replication time

  • large quantity of virions are produced

  • rna genomes mutate more than dna

    • dna poly lack proofreading capabilities

    • rna poly lack proofreading

• genetic changes that limit infectivity lead to attenuated strains (knocked down in virulence & used in vaccines)

• beneficial mutations may allow virus to:

  • escape host immune system detection

  • broaden host range

  • expand tropism (type of cell or tissues the virus infects)

  • increase infectivity

• reassortment may occur when 2 different viral strains co-infect single host cell → leads to new viral strains

Antigenic drift & shift

• drift: influenza RNA genome mutates frequently causing minor changes to HA & NA spikes (slow changes) **airprone rna & rna poly = need new vaccine**

• shift: occasionally flu undergo a major genetic reassortment (quick & deadly), lead to increased infectivity or expanded host range, people have no residual immune protection from infections or vaccines set stage for pandemic **different zoonotic mix & get a new virus**

Diverse Features are used to classify & name viruses

• international committee on taxonomy of viruses

  • develop criteria for naming & refine naming conventions for viruses SARS-CoV-2

• viruses are group by follow properties

  • type of NA present

  • capsid symmetry

  • presence or absence of envelope

  • genome architecture

medically Important DNA virus families

• papillomaviridae = huamn papilloma viruses (warts, some strains cause cervical cancer ~ naked icosahedral dsDNA circular

• herpesviridae = herpes simplex virus (oral & genital herpes) & varicella-zoster virus (chickenpox) ~ enveloped icosahedral dsDNA linear

• poxivirdae = smallpox virus ~ complex envelopes dsDNA

medically Important RNA virus families

• flaviviridae = hep c, west nile & dengue ~ icosahedral enveloped ssRNA nonseg no reverse transcriptase

• retroviridae = hiv & leukemia ~ icosahedral enveloped ssRNA nonseg w/ reverse transcriptae

• coronavirus = SARS, SARS-CoV2, common cold ~ helical, enveloped ssRNA+ nonseg

• Orthomyxoviridae = influenza ~ helical enveloped ssRNA- segmented

Host Range & Tropism

• host range = collection of species that a virus can infect

• some viruses infect more than 1 species while other only infect 1 species

• can infect both animals & humans = zoonotic

• measles = only humans

• tropism = tissues or cell specificity

  • due to viral surface factors

  • some infect wide range of cells/tissues = broad *ebola ~ 1st into macrophage → liver → epithelial → bleeding kills host

  • others infect only 1 type of host cell or tissue = narrow *HIV ~ require 2 receptors, very specific, only get into 2 cell types, asymp, CD4T cells & macrophages

Virus Sizes

• exhibit wide range of sizes

• rhinoviruses & polioviruses have a diameter as small as 30 nm

• ebola & pandoraviruses = 1,000 nm

• pithovirus is one of largest viruses = 1,500 nm *E. coli = largest @ 2,000nm

Viruses are names using standardized rules

• unlike organisms, viruses are not assigned to domains or kingdom

• phylum level is highest taxon for viruses

  • order

  • family

  • genus

  • species

• international committee on taxonomy of viruses

  • develop criteria for naming

  • refine naming conventions for viruses

Viruses hijack host cell machinery to multiply

• once inside, a virus commandeers the host cells energy, enzymes, organelles & molecular building block to build new virions

• host cell proteins required for HIV infection as an example

generalized animal virus replication

1. attachment (non envel viruses attach to host cell mem through capsid proteins)

2. penetration (enveloped = fusion/endocytosis, non = endocytosis)

3. uncoating (capsid is digested by enzymes in endocytic vesicles or cyto or nucleus)

4. replication (genome is rep & viral proteins are made)

5. assembly (new virions assembled)

6. release (enveloped virions are release by budding & non = lysis)

Some animal viruses have unique replication mechanisms that cause persistent infection

• acute = infect host cell & new virions are made immediately

  • viral rep is quick

• persistent = viruses have replication strategies that allow them to avoid immune system clearance (chronic or latent)

  • chronic = continuous at low levels

    • characterized by: continuous release of virions over time, slow progression & EBV ~90%, HIV ~ 1%

  • latent = initial infection, its DNA stays in ganglia & can reactivate *silenced

Endogenous Retroviruses

• hiv integrates into host cell genome

• ancient retro inserted bits of DNA into primate genome millions ago

• 8% of human genetic code consists of endogenous retro

• researchers propose that insertion of DNA by ancient retro helped genes like p53 become master gene reg that can switch on & off

Chronic Persistent Infections

• HIV

• Provirus = integration of viral genome into host cell

Latent Persistent Infections

• distinguished by flare up with intermittent periods of dormancy

• during flare up virions are shed & person experience symptoms

• flare ups can be triggered to come out of latency by stress

• members of herpesviridiae family are notorious for causing latent infection

  • HSV-1 = cold sores

  • HSV-2 = genital herpes

  • HHV-3 = chickenpox & shingles

Persistent Infections that can lead to cancer

• oncogenic viruses cause ~ 10-15% of cancers

• cause cancer by stimulating uncontrolled host cell division and/or decreasing host cell responsiveness to death signals

• Human papilloma viruses (HPVs) & Human T-lymphotropic viruses (HTLV)

Viruses can be propagated in lab

• in order to develop vaccines & drugs to combat viruses, researchers must be able to propagate viruses in lab setting

• requires host cell

Plaque Assay

1. initial virus stock is sequentially diluted

2. portion of diluted samples are added to cell culture plate

3. clear zones (plaque) form where host cells are killed by viruses

4. following incubation plaques are counted & initial viral titer is calculated & presented as PFU/ml

• human cells on a dish in layer, put agarose on it, add virus that infects specific cell; virus won’t float around & is stuck in augarose & is confined & can only go to neighbors

• gives # of infectious viral particles

growing/propogating animal viruses

• animal viruses are more difficult to cultivate than bacteriophages

• most animal viruses are grown using tissue culture tech

• live animal hosts may be required to support growth of some viruses

• embryonate eggs are also useful for propagating certain viruses

Diagnostic tests determine presence of certain viruses

• accurate diagnostics are necessary to ensure safe, virus free transplant tissues, pharmaceutical products & clinical samples

• clinical useful detection = specific, sensitive & short turn around times

• specificity = test only tests virus of interest *no false pos

• sensitivity = detects very low levels of target *no false neg

Agglutination Tests

• latex

  • viral antigens linked to tiny latex beads

  • mixed with sample

  • patient antibodies bind to viral antigen

  • beads agglutinate

• ELISA

  • some virus detection methods involve searching for viral proteins

  • utilizes purified antibodies to bond to viral antigens

  • adapted to detect antigen or antibodies

  • target adheres to surface

  • change of color indicate binding

Limitations of ELISA & agglutination assays

• sample tested must be liquid

• antigens must be fairly well characterized

• viruses can undergo an antigenic shift making it no longer detect it

• takes time to build up detectable antibodies (seroconversion window)

• its helpful to use a combo of detection methods

Detecting viral genetic material

• detecting viral NA is a growing trend in diagnostic

• NA detection techniques are more sensitive & sometimes more rapid

• to perform test:

  • clinal sample is collected

  • dna & rna are extracted

  • very specific seg of viral nucleic acid are detected by fluorescent-labeled proteins & PCR ~ denature @ 95-96 C, anneal @ 68 C, elongate @ 72 C

RT-qPCR

• viral genome is first made into complimentary DNA by HIV enzyme reverse transcriptase

• DNA put through polymerase chain rxn & amplification is read quantitatively in real time

• Results:

  • reported as cycle threshold value (# of cycles for signal to reach threshold where it can be seen over background signal)

Antiviral drugs treat infections, but don’t cure

• any step in replication = potential drug target

• most cases antiviral drugs only limit infections rather than cure

• specific difficulties that come into play when designing antiviral drugs:

  • **viruses = obligate intracell pathos**

  • antivirals = selective toxic

  • viruses have fewer chem distinct targets than living pathos

• few effective antiviral agents

• drug usually target viral enzymes

• prevent serious viral disease through vaccine

• vaccines train immune to recognize viruses & are effective means to limit infection

Drugs that block viral attachment, penetration & uncoat

• few antiviral drugs prevent viral entry into cells by blocking attach or penetration

• postexposure prophylaxis

  • lab prepared mix of injectable antibodies prevents viruses from binding & entering host

  • used shortly after suspected exposure & treats rabies & HIV

• nucleoside reverse trans. inhib

  • target reverse trans. enzymes

  • azidothymidine

• Oseltamivir & Zanamivir (tamiflu & relenza)

  • prevent influ A & B virions from budding off host surface

Nucleoside analogs

• drug that blocks replication

• at least 1 dozen drugs in class

• activated into compounds that mimic normal nucleotides

• chemical dead end for NA replication

• Acyclovir

  • inhibit DNA rep

  • effective against HIV-1, HHV-2 & varicella zoster virus

• ribavirin

  • target rna poly

  • effective against respiratory syncytial virus & hep c virus

interferons

• naturally occurring sub released by cells in reponse to viral infections

• single the presence of a virus

• neighboring uninfected cells make defensive changes that limit viral entry & replication

• can be produced in lab & administered to help limit progression of certain viral infections

Prions

• infections proteins; no gene materials

• dont replicate

• cause transmissible spongiform encephalopathies (TSE)

types of spongiform encephalopathies

• gerstmann-straussler-schienker Syndrome = fatal familial insomnia, inheritied

• creuzfeldt-jakob disease = acquired

Some neurodegenerative disease exhibit prion like features

• associated with misfolded proteins in brain

• alzheimers, parkinsons, amyotriphic lateral sclerosis