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Diagnostic Virology


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Why do diagnostic virology?
-Therapeutic intervention
-Pregnant females, babies
-Rabies, life-threatening ds
Why is it important to test pregnant females for viruses?
If HSV pos, then would do a c-section.
Why important to test babies?
CMV and rubella
What are 3 Diagnostic methods in virology?
1. Direct exam on infctd tissue
2. Indirect exam (isolatd virus)
3. Serology
What are 4 ways to directly examine tissue for virus?
1. Antigen detection via ELISA
2. E-microscopy for morphology
3. Light microscopy for histology and inclusion bodies
4. Virus genome detection via probes and PCR.
What are 3 ways to INDIRECTLY examine for virus?
1. Cell culture
2. Eggs
3. Animals
What is cell culture used for?
1. CPE observation
2. Hemadsorption
3. IFA
What are Eggs used for?
Pocks on CAM, hemagglutination, and inclusion bodies.
What are animals used for?
inoculating with the virus to observe for death or disease.
When using viral serology, what are you looking to see?
-Detection of titers rising >4x between acute and convalescent serums
-IgM indicates acute phase
What are 4 classical serological techniques?
1. Complement fixation
2. Hemagglutination inhibition
3. Immunoflourescence
4. Neutralization
What are 3 Newer techniques?
1. RIA
2. EIA
3. Western Blot
What 3 types of cell cultures are used for isolating viruses?
1. Primary cells (Rhesus)
2. Semi-contins cells - diploid
3. Continuous cells - heteroploid
Which type of cell culture is best? Why?
Primary; supports the widest range of viruses.
Which type is easy to handle?
What two changes may viruses produce in cell cultures?
1. CPE - cytopathic effect
2. Hemadsorption
what is hemadsorption?
When cells in culture acquire the ability to stick to mammalian RBCs. May be characteristic of certain virus.
What viruses are NOT often isolated via cell culture?
1. Measles
2. Mumps
3. Rubella
4. Coxsackie A
5. Poliovirus
What are 4 drawbacks of cellculture?
-Takes a long time
-Poor sensitivity dependent on specimen condition.
-Suscept. to bact/fungal/toxins
-Some viruses won't even grow
Which 4 viruses won't grow in cell culture?
1. Hep B
2. Norovirus
3. Parvovirus
4. Papillomavirus
What technique has sped up the processof cell culture?
Rapid culture - spinning inoculated culture allows faster adsorption and growth.
What is the good thing about testing specimens directly for viral antigen?
Speeds things up immensely!
What are the bad things about it?
-Very reduced sensitivity
-Specimens have to be v. good
-Procedures are tedious, timely, and expensive.
Is Electron microscopy good for diagnostic virology? Why?
No; requires high concentration of virus in the specimen.
What viruses ARE observed better with EM?
adenovirus, rotavirus, norwalk, HSV, smallpox, papillomavirus..
what 3 specimens can viruses be isolated from for EM?
1. Feces
2. Vesicle fluid
3. Skin scrapings
How can electron microscopy be enhanced?
By making it immune EM
What are 2 variants of immune EM?
1. Classical
2. Solid phase
what is classical immune EM?
Treat sample w/ spcf anti-sera to make viral particles clump.
What is solid-phase immune EM?
Antisera added to solid phase so virus particles stick to that.
What are the 4 criteria for diagnosing primary infection with serology?
1. 4x or more incr in Ab titer between acute and convalesc. stages.
2. IgM present
3. Seroconversion
4. Single high titer IgG
What are the 2 diagnostic criteria for Reinfection?
1. IgM absent or only sl. incr.
2. 1x or more inrese in IgG titer btwn acute/convalesc.
What's a common serological method?
Complement fixation in microtiter plates.
Wht is the test screen for HIV?
What is the confirm test for it?
Western Blot
Which viruses are NOT tested for with serology? Why?
Respiratory & Diarrheal.
The antibodies are produced long after symptoms/acute phase.
Name 3 causes of false pos results in serology:
1. Antigenic cross-reactivity
2. Patients with Mono/SLE
3. Passive vaccination via blood product transfusion.
When is CSF used for Ab testing?
To detect HSV or VZV.
What are molecular methods based on?
Detection of viral genome.
What are some molecular methods?
PCR and bDNA

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