Outline the 10 steps of DNA virus replication
- Attachment or adsorption
- Penetration
- Uncoating of virus & transport of genome to site of replication
- Early Transcription (mRNA)
- Early translation & early viral protein production
- Viral DNA synthesis
- Late transcription of further mRNA
- Late translation & synthesis of structural proteins
- Assembly
- Release
*If one step blocked then the other steps in replication process are also blocked
Describe the one step Growth Curve
- Viral attachment and penetration into cells
- shortly after infection - only low amounts of parental infectious virus identified
- Eclipse phase: virus replicates inside cells (but progeny virus not yet produced (seen as a plateau)
- Mature infectious progeny virions released = exponential phase (released into surrounding medium)
- after few hours cells infected become metabolically disordered and virus production ceases (plateau)
Step 1: Attachment
-neutralisation
- AKA adsorption
- virus attaches via surface protein and a complementary protein (receptor) on the surface of the cell
- occurs after random collision between virus and cell bearing right receptor
- Most of virus attaches in short period of time (mins), more time needed for residual virions to attach (hours)
- Viral attachment protein on surface of protein coat or inserted into envelope
- if antibody present - attaches to viral attachment protein and prevents virus from attaching to appropriate receptor = NEUTRALISATION
- right receptors not present on all cells - means cells exhibit tropism
- cellular receptor site usu. a glycoprotein
Step 2: Penetration
-2 ways;
-Fusion of envelop of virus w/ plasma membrane & direct release of nucleocapsid into cytoplasm [mediated by specific proteins or glyoproteins]
OR
-Engulfment of virus by cell in adsorptive endocytosis before the viral nuclei acid genome is released.
Step 3: Uncoating
Methods for;
- Poxvirus
- Herpesvirus
- Circovirus
-refers to freeing the viral genome
-occur in cytoplasm or in nucleus (where most DNA viruses replicate)
DIFF VIRUSES HAVE DIFF STRATEGIES:
-Poxvirus: replicate in cytoplasm - host factors induce disruption of virus core to release DNA
-Herpesvirus - replicate in nucleus - nucleocapsid migrates to cell nucleus via microtubules to nuclear pore and virus genomes released into nucleus
-Circoviruses replicate in nucleus - gain entry to nucleus during mitosis (Need cell division for replication to occur)
Step 4 & 5: Early transcription & translation of viral proteins
- Where these processes occur
- Early coded proteins
- formation of mRNA from virus DNA & early viral-coded proteins (translation)
- Most transcriptional events occur in nucleus - mRNA transported to cytoplasm where translation of coded proteins occurs using host cell ribosomes.
- proteins transported back to nucles where they are need before more steps can occur
- early coded proteins = enzymes necessary for DNA synthesis
- early transcription stops when viral DNA synthesis begins
Step 6: Viral DNA Synthesis
-e.g. of ones that use own polymerase & those that hijack cell’s polymerase
- Viral DNA synthesised to progeny viral DNA
- Some DNA virus families encode a DNA-dependent DNA polymerase (i.e. Herpesviridae, adenoviridae, poxviridae)
- Others rely on host cell’s DNA-dependent DNA polymerase (i.e. circoviridae, parvoviridae, polymaviridae, papillomaviridae)
Step 7 & 8: Late transcription and Translation
- Late transcriptional events occur AFTER DNA synthesis has occurred
- late mRNAs transported to cytopalsm for translation into structural proteins by host ribosomes
- May be translated as polyproteins & need to be cleaved by proteases
- May be glycoslated w/in trans-Golgi network
Step 9: Assembly of Virions
- Proteins & synthesized viral DNA packaged together = progeny virus
- inner core created first (ass. w/ DNA), capsid proteins last.
- Most DNA viruses assembled in nucleus
- Poxvirus: protein and DNA accumulate in common cytoplasm virus factory (inclusion body)
- Assembly occurs after structural proteins migrate back to nucleus where DNA replication & transcription have occurred
Step 10: Release of Viral Particles
-2 ways
2 ways to occur:
- Autolysis of cell: used by most DNA viruses (virus induced apoptosis may be involved)
- Herpesvirus mature by more complex process during which the viruses acquire an envelope and endocytosise out of cell
Parvoviruses;-
Genome virion enveloped replication target fragility host range
- 5kb, (-) single-stranded, linear DNA
- Icosahedral, 18-26 nm diameter
- No
- Intracellular
- Rapidly dividing cells
- V. resistant in enviro.
- Arthropods and mammals
Parvovirus - DNA in more detail
- Has hairpin (palindromic) structure at 3’ end - used as self-primer to start syn. of plus sense DNA (ss loop back on themselves) [unique to this family]
- dependent on cell division for replication (initiates division after host cell has completed S phase [uses cellular polymerases available in early G phase]
- replicate in nucleus - often producing intranuclear inclusions
Parvovirus: autonomous viruses & defective virus replication
-survival in enviro
Autonomous: need cells to pass through S-phase
Defective: require host machinery PLUS helper virus for replication (called dependoviruses - can’t replicate on own are not considered pathogenic)
-Very stable in enviro - survive pH 3-9 & at high temps
-susceptible to formaldehyde and chloramines and bleach
(though appropriate contact time necessary for inactivation)
-typically results in cell death (necrosis is common feature)
Parvovirus: Pathogenesis (where they infect)
*cells w/ high turnover rates, e.g.
- actively dividing cells in villi of intestine
- lymphoid tissue and bone marrow
- virus affects actively dividing Purkinje & cerebellar cortical cells in brain
Human Parvovirus (B19 Virus)
- major, medically relevant human parvovirus
- spread via respiratory route
- approx. 60% adults seropositive
- replicates in erythroid progenitor cells (found in bone marrow)
- results in rash & arthraligia/arthritis
*called B19 = blood sample no. that was first found to have virus in Australia
4 Human disease associated w/ B19
- Erythema infectiosum (5th disease): intense rash (often starts on cheeks & spreads outwards)
- occurs in children and adults
- little infectious virus in blood at time of disease because it’s due to immune complex formation - Transient aplastic crisis: individuals w/ anemia - acute parvovirus infect. can lead to probs because of cessation of RBC production in bone marrow
- Hydrops fetalis: related to mothers infected in pregnancy - can cause fetal infection (hydrops fetalis) or fetal loss due to severe anaemia
- Persistent B19 Infection: may be persistent for some - ass. w/ failure to make neutralising antibody response (congenital or immunodeficiency)
- no rash or arthritis - virus continues to replicate and destroy RBC progenitors. leads pure red cell aplasia (characterised by anaemia)
Papillomaviruses:-
Genome Virion Enveloped Replication Target Fragility Host Range
- Circular, ds DNA; 7-8kb
- Icosahedral
- no
- intranuclear
- epithelial cells
- resistant
- amniotes (tetrapod vertebrates that lay eggs on land)
Similarities b/w papillomaviruses and polyomaviruses
-Differences (w/ focus on papillomavirus
-both small, non-enveloped icosahedral viruses w/ ds circular DNA
-replicate in nuclei of host
-formerly classified in same family
-recognised as being distinctly different
Papillomavirus: slightly larger, larger genome, genes on one strand (polyomavirus has genes on bothe strands)
Why can’t we use serology to distinguish papillomaviruses
- > 100 genotypes of human PVs exist
- serology [using antibodies] not useful as there is significant antigenic cross-reactivity between genotypes
Papillomavirus Genome
- Long control region contains origin of DNA replication & important regulatory sequences
- has 3 different reading frames w/ overlapping genes
- has mix of early and late genes
Papillomavirus Lifecycle
- most cells remain latently infected
- most PV prefer infecting keratinocytes
- squames containing large no. of viral particles are released at surface
- infectious viral particles transmitted mechanically or direct/indirect physical contact
- anogenital typcially spread during unprotected sexual activities
- genotypes determines its anatomic predilection site & risk it poses (cancer causing)
- Nearly all humans have it - healthy immune system means benign warts should resolve eventually
- lifecycle confined to intracellular locations w/in epithelial cells (helps it hide from immune system)
- infect basal epithelial cells
- v. resistant in enviro
Preventing papillomavirus infection
- Neutralizing antibodies protect host from infection by that type
- gardasil = recombinant vaccine against 4 types
- contains non infectious HVP like particles (empty shells made of capsid proteins & no viral material
- gardasil = recombinant vaccine against 4 types
Treating Warts (Papillomavirus)
- Prophylactic vaccination
- Therapeutic Vaccination
- notoriously difficult to treat
- local recurrence common
- surgical excision
- Cryotherapy
- photodynamic therapy
- radiotherapy
Prophylactic: *all about setting up a provention - neutralizing antibodies (induced by parts of virus - not active)
Therapeutic Vaccination: *if disease is present: aims to cure an established disease
-any vaccine should aim to induce t cell mediated immune response (target early proteins E6 & E7 or papillomavirus DNA (in deeper layers of stratified squamous epithelium))
Cervical Cancer and Papillomavirus
- 3rd commonest concer in women
- vast majority associated w/ presence of high risk HPV types (HPV 16, 18, 31 & 45)
- Pap smear test: cytological evaluation of exfoliated epithelial cells from cervis to detect dysplasia and neoplasia
- is a good screening test (large time frame between development of dysplasia and invasive cervical cancer
