Diagnostic virology Flashcards
Example of a flat and a raised rash
Flat= macular rash Raised= vesicular (=blister)
What kind of rash would be found in measles and herpes
Measles= macular
also macular: rubella, parvovirus, EBV, CMV
Herpes= vesicular, but a discrete distribution across just a couple of dermatomes (shingles is a infection of just a couple of dermatomal nerves)
also vesicular: VZV/entero
What can be detected for viruses
Infectious virus (virus isolation & EM)
Protein components (antigens) of the virus (p24 antigen in HIV, surface antigen in HBV, etc)
Genetic components of the virus (DNA or RNA) - quantitative or qualitative tests are available
The host response (e.g. antibody or cell responses)
What does old vs current virus detection involved
OLD: cell culture and electron microscopy
CURRENT:
Antibody detected for the antigen in the blood (serology enzyme immunoassay= EIA)
Antigen can be detected by immunoflorescence (IF) or enzyme immunoassay (EIA)
PCR
Differentiate sensitivity and specificity
Sensitivity: the test’s ability to correctly identify positive samples
Specificity: the test’s ability to correctly identify negative samples
As well as detecting antibodies/viruses/DNA/protein components,
what else in virology testing is useful and why
- Quantity of antibody found (antibody is much higher in vaccinated people than people with infection, so helps to distinguish)
- Serotyping (=classification based on cell surface antigens) i.e. with HIV
- Quantification of genomes= VIRAL LOAD (essential for diagnosis and monitoring of HIV, HBV, HCV, CMV, EBV in immunocompromised)
- Genome sequencing —genotyping as different virus types have different prognosis
- antivrial resistance testing (to find out what treatment will work)
Wat could following samples test
Throat swab, Nasopharyngeal aspirate (NPA), bronchoalveolar lavage (BAL), ET secretions –
Stools
Urine
CSF
Blood (clotted)
Blood (EDTA)
Saliva
Throat swab, Nasopharyngeal aspirate (NPA), bronchoalveolar lavage (BAL), ET secretions – for detection of respiratory viruses by (IF or) PCR
Stools - for rotavirus, adenovirus & norovirus antigen detection (EIA) or PCR
Urine – for BK virus & adenovirus PCR
CSF - for herpes viruses and enteroviruses PCR
Blood (clotted) - for serology (antibody detection)
Blood (EDTA) - for PCR / viral load testing
Saliva – for serology &/or PCR (eg measles)
SUMMARY: we do PCR on all apart from clotted blood, which is used for antibody detection against the virus, not to identify the virus
Which antibody is shown at which time
IgM iMmediate (acute infection) IgG after infection (chronic infection for life)
What is serology and outline HBV serology
Serology is looking at human response to infection, looking at antigens etc.
Hep B… surface antigen present in acute or chronic HBV. Surface antibody present in recovery/immunity for HBV (incl. vaccinated).
Core antibody persists throughout life…. if the core antobody is IgM it indicated HIV within last 6 months
https://epi.publichealth.nc.gov/cd/lhds/manuals/hepB/docs/hbv_serology.pdf
Which serology for HIV
4th generation (test for AB and anitgen)… so looks for Ab + p24 (which is in the protein capsid)
confirm testing for to exlude non-specific reactivity (false positive)
Typing- HIV 1 or 2
Repeat and measure HIV viral load
Which type of sample can be used for viral load testing
Blood (EDTA) i.e. anticoagulated blood
When is IgM and IgG observed
IgM–> around 3 months
IgG –> lifelong
What is antibody avitdity testing
Over time, the antibodies produced against an antigen increases affinity….
you can test the strenght of antibody binidng 4 weeks apart… if the affinity increases this is consistent with an old infection
When is virus isolation in cell culture used
Rarely
phenotypic antiviral susceptibility testing (HSV)
i.e. to test which antivirals work on a virus
Electron microscope rarely used, but which samples if it was
stool and vesicle fluids
Enveloped virus with 20 faces
Herpes
Bullet shaped viruses
Rabies
Filamentous virus… long and windey
Ebola
How is IF still used
Still occasionally used for the direct detection of viral antigens in clinical samples (DIF) …. inexpensive but subjective
Cause of respiratory distress due to bronchiolitis
RSV
How can respiratory viruses be identified
respiratory virus PCR
How does multiplex PCR work
You put in primers for lots of different genes for many viruses…. if the gene is present then that DNA will be replicated lots of times and will be detected
What is PCR
Method for amplifying specific RNA (RT-PCR) or DNA sequences
What is required for RNA identification
reverse transcription (hence RT-PCR)
3 stages PCR
Denature, primer annealing, chain elongation
What can be used in CNS disease for virus testing
PCR:
CSF (for HSV, VZV, enterovirus)
Stools and throat swap for enterovirus
Blood for serology and/or PCR (west nile, japanese encephalitis virus and other arbovirus)
Which virus are you testing for meningitis and encephalitis
HSV, VZV and enterovirus
What can you do for D&V for virus testing
Stool sample…. PCR or EIA for norovirus, rotavirus,adenovirus, sapovirus, astrovirus
How many cycles in PCR
around 30
How was PCR interpreted previously vs now
In past: with gel
Now: real time PCR melt curve analtysis
Why is sequencing helpful
Genotyping virus
Antiviral resistance testing
Phyogenetic analysis… to see how related viruses are and see if the same virus in different people!
What is CMV
CMV, cytomegalovirus, is similar to the herpes virus that causes cold sores and chickenpox.
Once you have the virus, it stays in your body for the rest of your life
Can get flu like symptoms first time you catch it, after that only dnagerous if past onto new born
Viral cause of fever and rash
HIV (important), EBV, CMV
