Past Papers 3 Flashcards
HAV vaccine
What type of vaccine is it?
Inactivated vaccine
HAV outbreak
Who needs PEP?
<14 days since exposure
<14 days post exposure
- liver disease/ immunosuppressed - Vaccine + HNIG*
- <12 months old - Vaccine carers**
- 12 months - 59 years - Vaccine
- > 60 - Vaccine + HNIG
*ideally check HAV IgG before administering
** vaccine can be given >2 months of age, but is unlicensed
HAV outbreak
Who needs PEP?
> 14 days since exposure
> 14 days post exposure
- liver disease/ immunosuppressed - Vaccine + HNIG if <28 days since exposure
1 household contact
- food handler - if high risk of acquiring infection, eg from small child at home with poor hygiene, and has direct contact with food items, then transfer to duties which do not involve food preparation until 30 days post exposure. If low risk of acquisition, just give hand hygiene advice only. Do not give vaccine if >14 days after exposure
- non-food hander - hand hygiene advice only
> 1 household contact
- Vaccine everyone >12 months old. Only give to try and reduce further outbreak
Baltimore classification of viruses
What is in each class?
I - dsDNA
II - ssDNA
III - dsRNA
IV - ssRNA +
V - ssRNA -
VI - ssRNA-RT
VII - dsDNA - RT
What is a plaque reduction neutralisation test? PRNT
The PRNT is a serological test which utilizes the ability of a specific antibody to neutralize a virus, in turn, preventing the virus from causing the formation of plaques in a cell monolayer.
Typically, the assay involves mixing a constant amount of virus with dilutions of the serum specimen being tested, followed by plating of the mixture onto a monolayer of host cells
The concentration of plaque forming units can be determined by the number of plaques formed after a few days.
A vital dye (eg, neutral red) is then added for visualization of the plaques and the number of plaques in an individual plate is divided by the original number of virions to calculate the percentage neutralization.
Depending on the virus, the plaque forming units are measured by microscopic observations, fluorescent antibodies, or specific dyes that react with the infected cell.
Interpretation is typically based on 50% neutralization, which is the last dilution of serum capable of inhibiting 50% of the total plaques (virions). Termed PRNT50 value
Currently, the PRNT test is considered the “gold standard” for detecting and measuring antibodies that can neutralize the viruses that cause many diseases. It has a higher sensitivity and is more specific than many other serological tests for the diagnosis of some viruses.
Very useful for vaccine studies, whereby you want to see what antibody titres are required to help prevent an infection/ reduce risk of severe disease
Time intensive
What are problems with a plaque reduction assay?
Time intensive - so not very helpful for diagnosing an acute infection. Better to do an EIA and measure antibody titres
if use incorrect cell line may not see virus infect cells correctly
What are good uses for plaque reduction assays?
Very useful for vaccine studies, whereby you want to see what antibody titres are required to help prevent an infection/ reduce risk of severe disease
Very useful for diagnosing Flaviviruses such as Dengue. Flaviviruses have lots of cross-reactivity in EIA tests
Solid organ transplant
What infections to screen for in donor?
HIV Ag/Ab
HBsAg/ Anti-HBc
Anti-HCV
HTLV 1/2
CMV IgG
EBV
Syphilis serology
Toxoplasma serology
If travel history -
West Nile Virus
Strongyloides
Coccidioides/ histoplasma
Trypanosomma Cruzi
What is Sanger sequencing, and what is it used for?
determines sequence of nucleotides bases in a piece of DNA
99.99% accurate
What is Next Generation Sequencing?
What are benefits over Sanger method
NGS
- higher throughput- massively parallel testing
- higher depth reads - increased sensitivity
- higher discovery power for mutants/ new viruses
What are the steps of Sanger sequencing?
The Sanger sequencing method consists of 6 steps:
(1) The double-stranded DNA (dsDNA) is denatured into two single-stranded DNA (ssDNA).
(2) A primer that corresponds to one end of the sequence is attached.
(3) Four polymerase solutions with four types of dNTPs (AGCT) but only one type of ddNTP (AGCT) are added. ddNTP are dideoxynucleotide triphsophates, which have oxygen atom removed from ribonucleotide, and cannot form a link to the next nucleotide thereby terminating chain elongation
(4) The DNA synthesis reaction initiates and the chain extends until a termination nucleotide is randomly incorporated.
(5) The resulting DNA fragments are denatured into ssDNA.
(6) The denatured fragments are separated by their size by gel electrophoresis and the sequence is determined.
What are advantages of Sanger V NGS?
Sanger
Advantages
Fast and cost-effective for low target number
Established workflow
Simple data analysis
Longer reads (500-700 bps)
Disadvantages
Low sensitivity (~15–20% detection limit)
Low discovery power
Low scalability due to increasing sample input requirements
What are advantages of NGS V Sanger?
NGS
Advantages
Higher sequencing depth for increased sensitivity (down to 1%)
Higher discovery power
Higher mutation resolution
More data generated with the same DNA quantity
Higher throughput
Disadvantages
Less cost-effective for low target number (1–20 targets)
Time-consuming for low target number (1–20 targets)
What are available covid vaccines?
What is the target?
Spike protein target
mRNA vaccine
Pfizer-BioNTech
Moderna
Vector vaccine - adenovirus
AstraZeneca
Janssen Johnson/ Johnson
protein subunit - spike protein
Novavax
What are reasons for vaccine hesitancy?
poor knowledge
religious beliefs
medical distrust
poor attitude of healthcare workers - e.g not offering or prompting
poor acces to services - e.g it is a big hassle to get the vaccine
