Influenza and hepatitis virus Flashcards
a) Influenza structural features (8)
b) Classification of influenza
c) Overview of influenza A viruses
a) i) Around ~100nm ii) Lipid membrane iii) Haemagglutinin, HA - used to attach to host cell sialic acid iv) Neuraminidase, NA - enzyme to degrade sialic acid v) M2 ion channel - disassembly vi) Matrix vii) Nucleoprotein viii) Polymerase (PB1, PB2, PA) ix) RNA genome
b) Family Orthomyxoviridae
There are four types: A, B, C, and D (type A causes most diseases in humans)
Strains are named according to the type, country of isolation, isolate number, and year
Within a specific type (eg A), influenza viruses are grouped into subtypes based on the type of their HA and NA proteins (eg H1N1)
c) There are 18 HA and 11 NA subtypes (so many different combinations possible)
Birds are the natural hosts of influenza virus and migratory birds can spread influenza virus long distances (very problematic in fowl production)
Influenza virus entry
(see image)
1) HA binds to host cell sialic acid
2) Trigegrs endocytosis into an endosome
3) Endosome is acidified causing a conformational change in shape of HA so that it can function as a fusion device
4) Vesicular and viral membrane fuse
5) Viral nucleic acid is released into the cytosol, then into nucleus where replication takes place
a) Overview of haemagglutinin (HA)
b) The need for a change in pH
c) Entry of influenza and drugs that effect
a) i) A trimer ii) each monomer consists of HA1 and HA2 polypeptides, cleaved from the precursor HA0. HA1 is the globular head (with sialic acid binding site) and HA2 is the stalk iii) HA binds to sialic acid. Antibodies can be used that bind to HA1 and prevent the interaction with sialic acid, preventing infection and neutralising the virus iv) HA2 contains a fusogenic peptide buried in the timer. Acidification causes a conformational change, exposing the fusion peptide and it can insert into the host vesicular membrane to cause fusion
b) At a neutral pH, HA1 globular head covers the fusogenic peptide, but under the fusion pH (acidified), the globular head moves out of the way and the HA2 stalk extends, so the fusion peptide is more at the tip. This allows the host vesicular and viral membrane to fuse.
c) Entry and fusion requires a low pH, and fusion is mediated by HA. Acidification of the viral core occurs by protons being imported by the ion channel M2
Amantadine and rimantadine are drugs used to inhibit the M2 protein, so prevents protonation of the core, and virion uncoating, hence replication is not possible. However, drug resistance arises easily.
Replication of influenza virus and the action of drugs on this
The ribonucleoprotein complexes for each of the 8 RNA segments are transported into the nucleus, the 5’ and 3’ ends hybridise to make dsRNA (panhandle) and the rest is a loop. The panhandle is where the viral RNA dependent RNA polymerase (RdRp) will bind. The -ve sense RNA is transcribed to make viral mRNA, which can be translated into viral proteins.
However, the viral mRNA is not an exact copy of the genome as it differs at the 5’ and 3’ end (so can’t be used for replication). At the 5’ end there is a cap and a few nucleotides derived from cellular mRNA (derived by cap-snatching from host mRNAs, hence why influenza needs the host DNA dependent RNA polymerase II - to provide a supply of caps that the influenza viral RdRp uses to synthesise viral mRNA). At the 3’ end the virus mRNAs terminate before the end of the viral genome template, and have a polyA tail.
To replicate the genome, complete copies of the genome viral RNAs are needed. These +ve ssRNAs are also produced by the viral RdRp and are then copied back into more -ve ssRNAs (cRNA). The new -ve ssRNA may either be packaged into new virions, or act as templates for more mRNA to be made
Favipiravir inhibits transcription of cRNA and hence replication of the viral genome
Pimodivir prevents translation of viral protein
Release of influenza virus and drugs that affects this
Released by budding through the plasma membrane. The new virions acquire NA and HA. NA is essential for release: i) NA removes sialic acid from the cell surface, which means the virions wont be bound via HA to the cell surface. NA also removes sialic acid from virions, prevents virion clumping (otherwise HA would bind to sialic acid on HA or NA). Therefore NA is the target of drugs tamiflu (oseltamivir), and relenza and rapivab peramivir. However, these drugs must be administered at a precise time during the viral replication cycle: if given too early - the drugs may be removed before they are needed to have their effect; if given too late - the virus would have already been able to replicate and will have been released already
Antigenic variation
a) Describe how influenza shows antigenic drift
b) Describe how influenza shows antigenic shift
a) i) Due to amino acid mutations in the HA. These accumulate with time - the H3 HA in man now has asurface very different to earlier H3 HA ii) Due to continual change. New variants that escape exisiting antibodies are selected for in the presence of antibodies. This change is less dramatic than antigenic shift
b) (see image). This occurs when the same cell is infected by different strains of influenza virus at the same time. eg human virus with avian HA, there is no existing immunity, hence pandemics can occur (if the virus’ structure is mainly human elements, but just has an avian HA, this can be deadly)
a) H5N1 influenza
b) How might pathogenic avian influenza adapt to man - overview
c) Avian influenza binding to human cells
a) Highly pathogenic strain. H5N1 influenza viruses are endemic in avian species and is spread globally by migratory birds. It is transmitted to other birds, eg poultry industry where there is a high density of hosts, hence high virus titres. This has been transmitted to man, where it has very high mortality (~60%). There is no human to human transmission, but the virus may adapt - serious threat
b) i) better binding to human cells ii) better replication in human cells iii) better escape from human innate immunity iv) better transmission between humans. Adaptation may be multi-factoral
c) HA binds to host cell sialic acid with either α2’ - 3’ or α2’ - 6’ linkage to galactose. HA from avian viruses binds to α2’ - 3’ better than α2’ - 6’. However, HA from human viruses binds to α2’ - 6’. Just a single aa change (L226Q) in H3 is needed to alter this specificity, if that happened, avian influenza could be a big threat.
How might pathogenic avian influenza adapt to man
a) Replication efficiency in human cells
b) Resistance to host innate immunity
a) Influenced by the RNA polymerase. Avian flu PB2 has E627 and replicates well in avian cells, but poorly in human. Human flu PB2 has K627 and replicates well in avian and human cells. Experimental change (E627K) enables better replication of avian influenza in human cells. Avian strains that have adapted to man have K627
b) The non-structural protein 1 (NS1) decreases production of interferon. Avian influenza NS1 protein may do this better in avian cells than human. Changes in avian NS1 may make it a more potent inhibitor of human interferon production
Overview of hepatitis viruses (3)
1) Hepatitis A virus (HAV) - a picornavirus transmitted by the faecal-oral route
2) Hepatitis B virus (HBV) - a hepadnavirus that causes acute or chronic infection (depending on age), the latter predisposing to liver cancer
3) Hepatitis C virus (HCV) - a flavivirus that causes acute or chronic infection, the latter predisposing to liver cancer
Hepatitis A virus
a) Overview
b) Transmission
c) Disease caused
d) Prevention
a) A picornavirus, genus Hepatovirus. +ve sense ssRNA genome, 7.5kb. Icosahedral capsid, 27-32 nm, non-enveloped, stable at pH 1. Causes infectious hepatitis (distinct from serum hepatitis, HBV)
b) Transmitted via faecal-oral route. Ingestion of infected material → infection of epithial cells → local replication → viraemia → Infection of hepatocytes and liver macrophages (Kupffer cells) →virus secreted from liver bile → released in faeces for ~2 months after the infection
c) Acute hepatitis - jaundice. In 90% children, infection is asymptomatic, but in adults 80% develop acute hepatitis (worse if older). Fever, fatigue, appetite loss, diarrhoea, jaundice, elevated levels of liver enzymes (alanine amino transferase, ALT) in serum.. Symptoms may last for 2-6 weeks. Full recovery in most cases
d) Clean water, sanitation. Good standards of hygiene. Vaccination - inactivated vaccine and live attenuated vaccine.
Hepatitis B virus
a) Overview
b) How first discovered
c) HBV genome
a) A hepadnavirus. Icosahedral capsid, lipid envelope, surface glycoprotein (HBsAg). There are teo types of particles present. Virion (Dane particle) 42nm. 22nm lipoprotein particles abundant in serum of chronically infected patients. Virus doesn’t grow in cultured cells. 300m people infected chronically, and chronic infection leads to hepatocellular carcinoma (HCC)
b) Blumberg studies blood antigen polymorphisms in different polulations. Noted abundant antigen common in australian aboriginals (australia Ag). This Ag was bound by antibody from patients who has recovered from serum hepatitis. Ag also found in patients with i) chronic hepatitis and ii) acute hepatitis following blood transfusion. Australia Ag represents HBV surface antigen HBsAg. HBV was a cause of serum hepatitis. Led to development of diagnostic tests to screen blood and products for HBV, aswell as a HBV vaccine. Hence a prevention of chronic hepatitis and liver cancer
c) Partially dsDNA, 3.2kb. Extensive overlapping open reading frames. Encodes reverse transcriptase. HBsAg, with 2 N-terminal extensions (preS1 & preS2). Core (capsid) (see image)
Hepatitis B virus
a) Transmission
b) What is meant by its action as a reversivirus
c) Pathogenesis
d) Prevention
a) Perinatal - infected mother transmits to infant at, or close to, birth. Infected mother or siblings during the first years of life. Previously, a large amount of transmission through use of infected blood products to blood transfusion recipients and haemophiliacs. Contaminated needles (drug abusers)
b) Like a retrovirus, HBV replicates via reverse transcription. However, HBV packages DNA rather than RNA in retroviruses. Also, HBV doesn’t integrate the genome into host chromosomes during replication, hence doesn’t establish latency. HBV cannot be grown in culture
c) Acute infection - jaundice, seroconversion, clearance of HBsAg from blood, clearance of virus, good prognisis. Chronic infection - jaundice, virus not cleared, high levels of circulating HBsAg in blood, persistent liver disease, inflammation, development of HCC. Age is the major factor influencing acute versus chronic infection.
d) Screening of blood products. Vaccination i) from serum (the 22nm particles) ii) from recombinant Ag expressed from yeast. Mass vacination is having positive impact in preventing HBV infection and related liver cancer. Therapies needed to treat those infected chronically
Hepatitis C virus
a) Overview
b) Discovery
a) A flavivirus. Icosahedral capsid, lipid envelope, 50nm. Two surface glycoproteins E1 and E2 used for binding and inducing entry into host
b) After screening blood products for HAV and HBV, incidence of post-transfusion hepatitis was reduced, but not eliminated. Originally called post-transfusion non-A, non-B hepatitis (PT-NANBH), suggested another infectious agent responsible. Found that agent was removed by a 30nm filter, but not 80nm, and was destroyed by ether, so likely a virus with a lipid envelope. Was transmissable to chimpanzees, and virus genome was found in infected chimps, a flavivirus
Hepatitis C virus
a) Genome
b) Pathogenesis
c) Prevention and treatment
d) Brief comparison to HBV
a) +ve ssRNA, 9.5kb. Translated into a polyprotein using an IRES, then cleaved by proteases. Genome has huge diversity (6 major clades and up to 30% nucleotide diversity) due to major antigenic diversity and variation - a major challenge for vaccine development
b) Acute or chronic hepatitis. Unlike HBV, chronic infection is much more likely (70% infections). Predisposes to liver cirrhosis within 30 years. HCC.
c) Screen blood products. Treatment with anti-viral drugs: very effective drugs that block replication and cure patient - targets non-structural proteins like the proteases NS2 and NS3, phosphoprotein NS5A and RNA polymerase NS5B. All used in combination to essentially eliminate infection
d) HBV - effective vaccine, no effective drugs. HCV - no vaccine, very effective drugs
