Outbreak Sequencing Flashcards
What has reduced AIDS mortality in recent years?
What was utilised to track HIV outbreak?
Increased availability of anti-retroviral drugs
Sanger sequencing
HIV has a small genome. What are its 3 main/large genes and what do they encode?
What are they produced as and what do they get turned into?
gag (Group antigen) gene encodes the structural proteins of the capsid and matrix
pol (polymerase) gene encodes enzymes including the viral reverse transcriptase, integrase and protease
env (envelope) gene encodes the surface proteins which are embedded in the viral membrane
Encoded polyproteins which are cleaved into individual components
HIV is a retrovirus. How does it integrate its genome into the host?
Why is there a high error rate and what does this result in? (hint - proof reading and evolution)
What happens for viral particles to be packaged and then bud off?
2 copies of a ssRNA genome, but during the lifecycle this is reverse transcribed to produce dsDNA, which then integrates into the host genome
Reverse transcriptase lacks a proof-reading function, which means that it is highly error prone, resulting in HIV evolving rapidly
Transcription of the integrated provirus produces unspliced viral RNA, which goes on to be packaged in the viral particle, and spliced RNA which is translated to produce the 15 viral proteins
Viral envelope derived from the host cell membrane
What was the first suggestion of a possible zoonotic origin of AIDS?
Outbreak of a similar condition amongst a colony of macaque monkeys, associated with an increase in opportunistic infections
What is SIVmac?
Wild macaques did not reveal any cases of SIVmac. WHat does this suggest?
Virus isolated from infected macaques that was distinct from HIV but related
Suggested that it had been acquired in captivity
HIV-2 is closely related to SIVmac but not as widespread as HIV-1. How were origins of HIV-2 and SIVmac resolved?
Direct transmission links?
Similar virus, SIVsm, found in sooty mangabey
Direct transmission links between sooty mangabey pets and owners
What virus was discovered to be closely related to HIV-1?
What did this not demonstrate though?
SIVcpz
Didn’t demonstrate that SIVcpz was the precursor of HIV-1 as chimpanzees could have obtained virus from humans
What were the 3 pieces of evidence that suggested chimpanzees were a plausible reservoir from which humans acquired HIV-1 (Main (M))
SIVcpz sequence from chimpanzee more closely related to HIV-1 than captive chimpanzee strain
Observed a divergent virus in subspecies of chimpanzee suggesting long term co-evolution of SIVcpz and chimpanzees
2 distantly related strains co-infected the same animal so infection rates must be sufficiently high
What did the German 2011 outbreak of E. coli cause and what was believed to be the cause?
What did this cause (hint - market)
Haemolytic uraemic syndrome
Cucumbers based on questionnaires
Caused collapse in the market as vegetable imports were banned; Economic impact
What helped for quick response during E. coli outbreak? (hint - sequencing and draft)
Ion Torrent PGM “benchtop”; Much faster sequencing than Illumina
Allowed for high-throughput next-generation sequencing during outbreak
Raw sequence data of the strain was published allowing for draft genome assembly
What was unusual about the E. coli outbreak strain considering it caused haemolytic uraemic syndrome like an EHEC?
It was an enteroaggregative E. coli (EAEC) and caused a stacked brick configuration in the gut
What did sequencing of the outbreak E. coli strain and comparison with other strains uncover? (hint - islands)
Similar backbone but outbreak strains had several genomic islands for antibiotic resistance and shiga toxin
Sequencing uncovered the causative agent
What are 2 key 3rd generation sequencing techniques?
Why are they good?
Oxford Nanopore MinION
PacBio Sequel
Both produce very long reads with similar run cost
Why is MinION good for field sequencing?
Very small
When has MinION been applied?
How quick was it in determining what?
Salmonella enteritidis outbreak in Birmingham
Determined whether samples from the hospital were part of the outbreak or unrelated within 2 hours
