Viruses: Their Structure and Replication

What are Viruses

• Small, Filterable, infectious agents– Cannot be seen by light microscope – Electron Microscope

• Obligatory Intracellular Parasites– Not enough ATP by itself

Characteristics Of Viruses• Genetic Material: DNA or RNA – never both!

– Single stranded (ss) or double stranded (ds)– Linear or circular

• Capsid – Protein subunits (shell)– Protect the genetic material– May be involved in cell entry

• Envelope (required for entry if present)– Lipid, protein and carbohydrate– Protein: viral origin– Lipid, carbohydrate: host origin – Involved in cell entry (located outside the capsid)

• Subunit Replication only– Assembled after parts are made (quick log phase)

• NEVER contain enzymes for protein synthesis or ATP metabolism!!– Always appropriate the host cell machinery

Size of Viruses

Virus Classification• Based on Host Range (determined by specific receptors)

– Bacterial Viruses (Bacteriophages)– Animal Viruses – Plant Viruses– Others – (amoeba, insects)

• Based on genome structure– DNA or RNA (never both)

• ssDNA viruses• dsDNA viruses• ssRNA viruses

– Plus Strand (+RNA viruses)» Same

– Negative strand (- RNA viruses) » Complementary

– Retroviruses» Converted into complementary DNA, then into the cell for replication

• dsRNA viruses– Linear or circular– Size

• 2000 to 200,000 nucleotides

Virus Classification (cont.d)• Based on Shape

– Polyhedral viruses (icosahedral -20 faces, 12 vertices)– Helical Viruses– Complex viruses

• Based on Envelope– Naked viruses– Enveloped viruses

• Based on Disease caused– Respiratory viruses– Gastrointestinal viruses– Sexually transmitted viruses

Viral Structure: Capsid

Capsid = protein coat that encloses and protects the nucleic acid of a virus•Accounts for most of the viral mass•Composed of single or multiple proteins

• Each subunit = capsomeres

Viral Structure: Envelope

Sometimes, Capsid covered with envelope•spikes = carbohydrate-protein complexes (glycoproteins) that project from the envelope

• Can be used to attach to host cell• Influenza virus causes hemagglutination – clumping

of red blood cells by use of spikes

Non-enveloped viruses/ Naked Viruses = viruses whose capsids are not covered by an envelope

An Enveloped Virus

Virus mutations and immunity

When a virus infects a host cell:•Host immune system produces antibodies•Antibodies = proteins that inactivate the virus by reacting with virus surface proteins

Stops further infection

Why you can get some viruses more than once:• Genes that code for viral surface proteins are

susceptible to mutation• Antibodies can’t react with the altered surface

proteins infection• Ex) influenza; frequent mutations in its spikes

Non-neutralizing antibodies do not inhibit functionNeutralizing inhibits function

Viral Structure:General Morphology

Capsid Structure determines shape:

Helical Viruses = nucleic acid is inside a hollow cylindrical capsid with a helical structure

• Rabies, Ebola viruses, Tobacco Mosaic Virus

Polyhedral viruses = many sided; icosahedron is common with 20 equilateral triangles as sides and 12 vertices

• Poliovirus, Adenovirus, herpes, others??

Viral Structure:General Morphology

Enveloped Viruses = can be helical or polyhedral, but the capsid is surrounded by an envelope

• Helical: influenza virus• Polyhedral (icosahedral): Herpes simplex virus

Complex viruses = Complex structures; additional structures attached to capsids, combos of helical and polyhedral, may have several coats around nucleic acid

• Bacteriophage, poxviruses

Virus Structure

Virus TaxonomyInternational Committee on Taxonomy of Viruses (ICTV) groups viruses based on:

• Nucleic acid type• Mode of replication• Morphology

Viral species = defined as a group of viruses sharing the same genetic information and host range

• Viral species are given common names• Ex) human herpesvirus

• The suffix –virus is used for genus names• Ex) Simplexvirus

• Family of viruses is given the suffix –viridae• Ex) herpesviridae

Some Viral Families & Genera affecting humans

Some Viral Families & Genera affecting humans:Continued

Isolation & cultivation of virusesOur understanding of viruses comes mostly from bacteriophages, as they are easily grown on bacterial cultures

• Liquid suspensions or solid media

Plaque method for detecting & counting viruses:1)bacteriophages are mixed with host bacteria and melted agar, poured onto petri plate with existing layer of solid growth medium2)Top layer solidifies ~ one cell thick3)After several rounds of infection, multiplication and lysis, bacteria surrounding the virus are destroyed plaque

Bacteriophage lambda on a lawn of E. coli

• Each plaque is from a single virus (theoretically)• Number of plaques can be used to calculate

plaque forming units (PFU) in initial suspension

Bacterial Viruses

• Bacteriophages• DNA and RNA viruses

– ds and ss– Linear and circular

• Important tool – Alternative to antibiotic therapy

• Basis to study viral replication in host cells– Lytic cycle (lyses cell)– Lysogenic cycle

http://www.youtube.com/watch?v=gU8XeqI7yts

It is very easy to grow viruses in bacteria, so it another reason it’s an important tool

Never marketed phage therapy because it doesn’t work too well

Lytic Cycle of a T-Even Bacteriophage

1

2

3

Figure 13.11

HAS TO BE A DNA VIRUSOnly infects cells with specific receptors

“syringes in”

4

Figure 13.11

Lytic Cycle of a T-Even BacteriophageIntegrates genome into host genome

Steps for Replication

• Adsorption• Penetration• Biosynthesis

– Uncoating– Replication

• Assembly• Release

Memorize

Lysogenic cycle: bacteriophage lambda

Prophage: DNA incorporated and now it will make bacteriophages

DNA randomly pops off in lysogenic cycle then goes through lytic

Lysogeny: importance• Lysogenic cells are immune to infection by the same

phage (but not to other phages)• Phage conversion = tendency of host cell to exhibit new

properties when carrying lysogenic phage• Ex) Cornyebacterium diptheriae produces toxin only

when carrying lysogenic phage diptheria• Same is true for shiga toxin by pathogenic E. coli

• Specialized transduction = since bacterial DNA is incorporated with phage DNA, adjacent genes on host DNA may remain attached when phage DNA is excised for initiation of the lytic cycle

• Introduce foreign genes into a new cell’s genome

Specialized transduction – horizontal gene transfer

Multiplication of Animal Viruses

• Entry– Adsorption (not misspelled)

• Sticks to the surface– Entry/ Penetration– Uncoating (if capsid goes in)

• Replication/ Synthesis• Assembly/Maturation• Release

– Affect on host cell

DNA matures in the nucleus, RNA matures in the cytoplasm

Multiplication of Animal Viruses: Attachment

• Attachment– Virus attachment sites

• Spikes or capsid proteins

– Receptor Sites• Proteins, glycoproteins on host cell membrane

Multiplication of Animal Viruses: Entry

• Endocytosis– Pinocytosis (unseen)

• Plasma membrane folds inward into vesicles

– Receptor mediated Endocytosis (creates own vesicles)

• Influenza virus

• Fusion (membranes are the same)– Viral envelope fuses with cell membrane

Multiplication of Animal Viruses: Entry

Above: Togavirus entering a cell through pinocytosis

Multiplication of Animal Viruses: Entry

Above: Herpesvirus entering a cell through fusion

Multiplication of Animal Viruses: Uncoating

• Separation of nucleic acid from protein capsid– Capsid digested by host enzymes– Viral proteins synthesized to uncoat

• poxviruses

Replication/Biosynthesis/Maturation/Release: DNA viruses

• Gene expression in most viruses – Stage specific– Temporal Cascade

• DNA viruses– DNA shuttled to host nucleus – EARLY Gene transcription

• Uses host RNA polymerase• mRNA shuttled to protein translation sites in cytoplasm• Viral Enzymes (proteins) shuttled back to nucleus

– Viral DNA replication• Viral DNA polymerase replicates DNA

– LATE gene expression• Uses host RNA polymerase• mRNA shuttled to protein translation sites in cytoplasm• Viral structural proteins Proteins shuttled back to nucleus

– VIRIONS assembled, shuttled back to cytoplasm– Trafficked via ER, Golgi, to Cell membrane for release

“Too much detail, I just need you to remember that some genes are expressed before replication, some are expressed after. It just depends on when it needs that expression.”

It’s acidity is what causes the endosome to fuse with the envelopeand release the capsid

budding

biosynthesis of DNA viruses example

Back into nucleus

RNA viruses• Multiplication is same as that of DNA viruses except

mechanisms of how mRNA is generated

• Four nucleic acid types of RNA viruses• Single (+) strand of RNA

• Ex) picornaviridae, togaviridae• Single (–) strand of RNA

• Ex) rhaboviridae• Double stranded RNA

• Ex) reoviridae• “Reverse transcriptase” RNA (retroviruses)

• Ex) retroviridae

+ transcription - translation +

- strand makes message, so - stays -

- Strand will make one part (like capsid)+ will make other part (like the RNA)

Prophage – bacteriophage DNA incorporated???Provirus – any other virus DNA incorporated???

RNA polymerase is more likely to make errors than DNA polymerase

Avian Influenza

Clinical Focus, p. 371

Antigenic drift – mutations in how it looksAntigenic shift – so many mutations, very virulent, no antibodies to fight it

Biosynthesis of DNA vs RNA viruses

Multiplication of Animal Viruses: Release

Non-enveloped viruses = released from host through ruptures in plasma membrane host cell death

Enveloped viruses = the envelope develops around the capsid by budding: virus takes portion of plasma membrane as it pushes through it to extracellular space

• Doesn’t immediately kill host cell

Budding: formation of the envelope

Viruses and Cancer

Oncogenic viruses (cancer generating) = viruses capable of inducing tumors in animals (aka oncoviruses)

• ~10% of cancers induced by viruses

Oncogenes = parts of the genome that cause cancer when mutated; expressed at high levels in tumor cells

Transformation = viral genetic material integrates into host DNA and replicates with it (like bacterial lysogeny)

• Used by all oncogenic viruses

Oncolytic – viruses that grow in and lyse cancer cells

DNA oncogenic virusesHerpesviridae •Epstein Barr virus causes infectious mononucleosis

- remains latent in some throat and blood cells throughout life various lymphomas (Hodgkin’s, Burkitt’s)

Papovaviridae •all uterine (cervical) cancers are caused by human papillomavirusHepadnaviridae (Hep B) & Flaviviridae (Hep C)•hepatitis B and C can cause liver cancer

RNA oncogenic viruses• ONLY the retroviridae family of RNA viruses cause

cancer

Human T-cell leukemia viruses 1 & 2 = cause adult T-cell (white blood cell) leukemia and lymphoma

Mechanism of tumor generation:Viral reverse transcriptase generates the double

stranded viral DNA (provirus) that integrates into the host chromosome• Changes in genetic material always put the cell at

risk for tumor formation

Know oncogenic viruses and diseases with them

Latent viral infections

Viruses may infect host cells but cause disease only after a long period of time = latent infections

All human herpesviruses can remain in host cells for a person’s lifespan, until reactivation:

• Immune suppression (ex: AIDS)• Fever, sunburn (cold sores from herpes simplex)

•Reactivation may never occur no symptoms

Chronic can be latent or persistant

Persistent viral infections

Persistent (chronic) viral infections occur gradually over a long period of time•Infectious virus builds up over time, rather than appearing suddenly (like latent infections)•Typically fatal

Example:Subacute sclerosing panencephalitis (SSPE) = a progressive, debilitating, and deadly brain disorder

• Caused by immune resistant measles• No cure; may be managed with medication

Remember this one

Figure 13.21

Latent and Persistent Viral Infections

Peaks, looks like it will go away, then spikes (can be lethal)

PrionsPrions = proteinaceous infectious particles

• No nucleic acid, just purely protein• Cause infections diseases - neurological

•Bovine spongiform encephalophathy (mad cow)•Creutzfeldt-Jakob disease (CJD)•Gerstmann-Straussler-Sheinker syndrome

Run in families, indicating genetic componentnot purely genetic:

Eating infected meat transmits mad cow CJD transmitted via transplanted nerve tissue

Only killed by formaldehyde, very resistantKNOW ALL PRIONS

Prion Diseases

• Crutzfedt-Jakob• Kuru• Bovine Spongiform

encephalopathy (mad cow)

• vCJDhttp://www.beatricebiologist.com/2010/08/watch-out-for-prions.html

Plant viruses and viroidsPlant viruses = similar in morphology and nucleic acid types to animal viruses

Common crop viruses:-Bean mosaic virus-Wound tumor virus

corn and sugarcane-Potato yellow dwarf virus

Must penetrate cell wall by:-Wounds-Parasites

Ex) aphids that eat sap

Result = color change, deformed/stunted growth, wilting

Only destructive

Plant viruses and viroids

Infected plant spreads virus via pollen and seeds

viroids = short pieces of RNA with no protein coat• Known to cause some plant diseases• Pathogens of plants only

• Potato spindle tuber viroid

Prions are only proteinViroids are only RNA

Some major plant viruses

DNA virus families

Adenoviridae = cause acute respiratory disease (common cold)

Poxviridae = cause skin lesion diseases•Pox = pus-filled lesions

• Smallpox• Cowpox

Herpesviridae = named after herpetic (spreading) appearance of cold sores

• Genus simplexvirus (cold sores)• Genus varicellovirus (chickenpox)• Genus lymphocryptovirus (mononucleosis)

Figure 13.5b

Poxviridae• Double-stranded DNA, enveloped

viruses– Orthopoxvirus (vaccinia

and smallpox viruses)– Molluscipoxvirus– Smallpox– Molluscum

contagiosum– Cowpox

Mastadenovirus (adenoviridae)

Herpes simplex virus(herpesviridae)

DNA virus families

DNA virus families

Papovaviridae = named for the papillomas (warts) polyomas (tumors) and vacuolation (development of cytoplasmic vacuoles)

• Genus papillomavirus causes warts- HPV: cervical cancer and cauliflower-like growths

in cervix- Vaccine: Gardasil

• Polyomavirus diseases primarily affect the immunocompromised tumors

DNA virus families

Hepadnaviridae = named for their role in causing hepatitis and containing DNA

• Only one genus causes hepatitis B• The other hepatitis viruses (A,C,D,E,F,G) are RNA

viruses

Hepatitis = inflammation of liver• Hep B is similar to Hep C (an RNA virus) • Both transmitted through blood

- Associated with intravenous drug use• Cirrhosis, liver failure, liver cancer• Vaccine for Hep B, no vaccine for Hep C!

RNA virusesPicornaviridae = small (-pico) and contain RNA

• Single stranded RNA viruses

Important genera:Rhinovirus = responsible for >50% of common colds

Enterovirus = fecal oral transmission

poliovirus, coxsackie virus (aseptic meningitis)

Hepatovirus = only species in the genus causes Hep A•Fecal-oral transmission

• Contaminated food or water• Primarily affects less developed countries

•Replication: mucosa intestine liver•Symptoms: fever, nausea, diarrhea, jaundice•Prevention: vaccine

Togaviridae are enveloped (toga = covering)•Like picornaviruses, have a single strand of RNA

Important genera:Rubivirus = only member is rubella virus•Part of MMR vaccination series•Rubella = (latin: little red) aka german measles

- Itchy red rash- Swollen glands, fever

•Transmission: respiratory droplets•Treatment: none, usually subsides in days

- Less severe than measles (rubeola virus)

RNA viruses

RNA virusesParamyxoviridae = enveloped viruses with spikes

• single stranded RNA viruses

Important genera:Rubulavirus = contains the species Mumps virus •Transmitted by respiratory droplets•Was common before MMR vaccine (1960s)

•Symptoms: fever, headache, muscle aches, tiredness- swelling of parotid (salivary) glands!- Orchitis = swelling of testicles (~30% of males)

RNA virusesRhabdoviridae = bullet-shaped viruses with a single strand of RNA

• ~150 viruses of vertebrates, invertebrates and plants

Lyssavirus = genus that contains the species rabies virus

• Transmission: animal bite• Salivary glands highly concentrated with virus

• Spreads from muscle cells into CNS• Fatal if not treated prior to severe symptoms

RNA virusesOrthomyxoviridae = enveloped helical viruses with a single strand of RNA

Influenza virus = three genera (A,B,C) that cause influenza, a contagious respiratory illness

Symptoms: cough, sore throat, aches, fatigue and serious complications:

- Pneumonia- Bronchitis- Worsening of chronic health problems

TEM of H1N1 Influenza

Avian Influenza

Clinical Focus, p. 371

RNA virusesReoviridae = respiratory, enteric, orphan•Affect gastrointestinal system, respiratory tract•Double-stranded RNA viruses

Rotavirus = genus in family reoviridae•Most common cause of severe diarrhea among infants and children

Fecal-oral transmission•2009: included into U.S. recommended vaccination program by W.H.O.

Stylized SEM: rotavirus

RNA virusesRetroviridae = reverse transcriptase viruses•Reverse transcriptase = uses viral RNA as template to produce double-stranded DNA

• integrated into host chromosome provirus- protected from host immune system & antivirals- Replicates with host DNA- Can be expressed to produce new virions and infect

adjacent cells

Human Immunodeficiency virus (HIV)•Infects immune cells, progresses to AIDS•No cure: hard to target latent infected cells

Retroviruses