Virology

Question 1

what antibodies do we usually test for

Answer 1

usually IgG, but sometimes IgM. IgM takes longer to develop

Question 2

what is a direct ELISA

Answer 2

antigen on plate and patient antibody binds. an enzyme allows visualization

Question 3

what is indirect ELISA

Answer 3

antigen on plate. patient antibody binds then a secondary antibody conjugate is added for visualization

Question 4

what is a competitive ELISA

Answer 4

antigen on plate. patient antibody binds. then an inhibitor antigen binds to decrease color reaction. so, less of a reaction on the plate means more host antibody is present. opposite from other ELISAs

Question 5

describe titers

Answer 5

using ELISA testing, sample is diluted in a serial manner until no reaction occurs. the last well with a reaction is your titer

Question 6

what is lateral flow assay

Answer 6

SNAP tests. may test for antigen or antibody

Question 7

what is agar gel immunodiffusion? examples of its use?

Answer 7

antigen in middle. samples and controls in a circle around it. positive samples will have a white line near it. very reliable. gold standard
ex: coggins

Question 8

what is a hemagglutination inhibition test

Answer 8

some viruses bind RBCs and prevent agglutination. serum is added and if antibodies are present, they will stop the virus from binding and allow the blood to clot. gold standard

Question 9

what is a plate agglutination test

Answer 9

mix serum with stained antigen and form clumps if there is antibody. larger clumps with more antibodies

Question 10

what viral protein is usually tested for

Answer 10

spike proteins

Question 11

what is a capture/sandwich ELISA

Answer 11

antibody fixed to plate. antigen present in serum binds the antibody. add another label antibody for visualization.
useful for BVD where patient wouldn’t have antibody

Question 12

what is fluorescent antibody testing? what is it used for?

Answer 12

direct method. fluorescent labeled antibodies bind target antigens in fresh tissue. look under fluorescent microscope.
ex: rabies

Question 13

what is immunohistochemistry? what is it used for?

Answer 13

like IFA, but use fixed tissue. expose tissue to antibody then visualize under a normal scope. see the effect of virus in tissues, can see viral inclusions in the cell (usually pathopneumonic)
ex: FIP, marek’s dz

Question 14

what are the pros and cons of ELISAs and lateral flow assays

Answer 14

pro - quick, less expensive, readily available, some done in clinic, can get titers with ELISA
cons - less specific/risk of false positives/negatives (esp with lateral flow), not as likely to be accepted for regulatory purposes, cross reactivity possible

Question 15

what are the pros and cons of AGID (agar gel immunodiffusion) and HIs (hemoagglutination inhibition)

Answer 15

pros - specific, gold standard for some tests, often accepted for import/export
cons - harder to find, longer incubation, labor intensive, often more expensive

Question 16

what are the pros and cons of fluorescent antibody tests and immunohistochemistry

Answer 16

pros - both usually quite specific, visualize pathogen location, FA is fast, IHC is an option for fixed tissue
cons - rarely an antemortem test, can be expensive, may be species specific, limits assays

Question 17

differentiate antigen vs antibody tests

Answer 17

antigen - detect current infection, detects in immunocompetent animals
antibody - detect previous/chronic infection, detects vaccination that elicits IgG

Question 18

what is the benefit of real-time PCR

Answer 18

it quantifies viral load

Question 19

what does a higher Ct value mean in PCR

Answer 19

more cycles for the sample to break through the threshold, so less virus present

Question 20

what is a multiplex panel

Answer 20

PCR panel for multiple viruses that cause similar signs

Question 21

what is whole genome sequencing

Answer 21

done on a pure viral sample. usually used for epidemiology and research

Question 22

what is metagenomic sequencing

Answer 22

start with a mixed sample and sequence everything in it. computer helps analyze the data

Question 23

what are pros and cons for PCR

Answer 23

pros - fast, sensitive, reliable
cons - no distinction btwn live and inactive, may miss if virus mutates, contamination issues, need to know what you’re testing for

Question 24

what are the pros and cons for sequencing

Answer 24

pro - large amount of data, ID unknown pathogens, track mutations
cons - expensive, lots of analysis, best on pure samples

Question 25

describe viral isolation

Answer 25

grow virus from diagnostic sample to identify, gold standard, only test to prove viable virus is present

Question 26

what media are used for viral culture

Answer 26

embryos and special cell lines. cell lines should be similar to tissue the virus infects

Question 27

how do you know virus is growing on your culture

Answer 27

usually thru cytopathic effects or electron microscopy can also use fluorescent antibody test, hemagglutination, PCR, sequencing

Question 28

what is cytopathic effects

Answer 28

damage done by viruses on cells

Question 29

why might you use virus isolation

Answer 29

with low viral load, unknown pathogens, and for species without serological or molecular tests

Question 30

describe electron microscopy

Answer 30

stained with heavy metal salts to visualize surface structures, can search blindly, can detect primary and secondary pathogens

Question 31

T/F: if you don't see inclusion bodies on a group of cells, there are no viruses present that can cause inclusion bodies

Answer 31

F - usually only present at certain stages of disease

Question 32

pros and cons of virus isolation

Answer 32

pros - detect living viruses, detect and identify unknown viruses, sensitive, amplify and collect live virus cons - difficult, long wait time, expensive, sensitive to contamination

Question 33

pros and cons of electron microscopy

Answer 33

pros - visualize pathogen, work even if mutated, false positives rare, see all actors involved cons - expensive, less common, longer turnaround time, may pick up an innocent bystander

Question 34

pros and cons to histopath

Answer 34

pros - false positives unlikely, inclusions often distinct and pathognomonic, virus inactivated so no risk of spread cons - absence of inclusions ≠ negative, longer turnaround time

Question 35

what kind of swabs should not be used for viral testing

Answer 35

wood - antiviral, interferes with PCR calcium alginate - interferes with viral isolation and PCR testing

Question 36

what kind of swabs pick up more virus

Answer 36

flocked - has bristles

Question 37

with a case of avian influenza where you are trying to decide if it is safe to bring in new birds, which test should you use to be sure your flock is virus free? a. lateral flow assay b. real time PCR c. virus isolation

Answer 37

c is best. tells if any viable virus present a- quick, but risk of false positives b- sensitive, but detects inactivated virus as well as live. would be ok, but would end up waiting longer than necessary to bring in new birds

Question 38

what test is accepted from the regulatory bodies for avian influenza

Answer 38

virus isolation

Question 39

how much tissue is needed for viral testing

Answer 39

not a lot. a few grams

Question 40

what test should be used with a herd of cattle with respiratory illness

Answer 40

PCR resp panel. histology helpful but may not be diagnostic

Question 41

you have a cat with clinical signs consistent for FIP. a fecal sample is PCR positive for coronavirus. does this cat have FIP?

Answer 41

not necessarily. normal coronavirus and FIP are both shed in the feces

Question 42

is PCR, fluorescent antibody test, or lateral flow assay helpful in testing for FIP

Answer 42

no, do not differentiate typical coronavirus from FIP

Question 43

does a FCoV negative ELISA mean this cat does not have FIP

Answer 43

no. in late stage FIP, antibodies drop and may not be detected. serum testing not helpful for FIP

Question 44

an abdominal fluid sample is PCR positive for coronavirus. does this cat have FIP

Answer 44

not definitive, but is suggestive. good enough to act on this. histopathology and immunochemistry on biopsy tissues are more definitive than PCR

Question 45

how should whole fish be sent to the lab

Answer 45

overnight on ice packs or dry ice. not formalin b/c inactivates virus. not with ground shipping or just an insulated envelope. will rot