Diagnostics Final

Question 1

What are our detection goals for viruses?

Answer 1

-Prevent introduction
-Target care (treatment of secondary infections)
-Limit spread (to both animals and humans)

Question 2

How can we detect viruses?

Answer 2

-Host antibodies or antigen
Antigen -> viral proteins, nucleic acid, live virus

Question 3

What are tests we use to detect anibodies?

Answer 3

ELISAS, lateral flow assays, agar gel immunodiffusion, other agglutination assays

Question 4

How do antibody detecting tests work?

Answer 4

-test contains the antigen
-patient antibodies react with test antigen
-reaction is visualized
-some tetss for iGM, others IgG
-lag time from infection

Question 5

Direct Elisa?

Answer 5

The primary antibody conjugate has the antigen and enzyme

Question 6

Indirect ELISA?

Answer 6

Has a secondary antibody conjugate

Question 7

Competitive ELISA?

Answer 7

There is an inhibitor antigen

Question 8

Describe a lateral flow assay?

Answer 8

-labeled antigen
-binds patient antibodies
-easy and cheap
Example :FIV

Question 9

Agar gel immunodiffusion test

Answer 9

-equine infectious anemia (coggins test)
avian influenza

-postive will have a band of precipiate

Question 10

Hemoagglutination inhibiton test

Answer 10

Figure on slide 12

Question 11

Detection of viral proteins
Lateral flow assay

Answer 11

labeled antibodies
-bind antigen
easy and cheap
Example : parvo

Question 12

Viral proteins
ELISA

Answer 12

-capture or sandwhich assay

Question 13

Viral proteins
fluorescent antibody testing

Answer 13

-direct method
-fluroescent labeled antibodies (bind target antigens)
-rabies testing

Question 14

Viral proteins
Immunohistochemistry

Answer 14

-fixed tissue is treated to expose antigen
-sections exposed to antibody
-example: FIP, Mareks disease

Question 15

What are the pros and cons of ELISAs and lateral flow assays?

Answer 15

Pros -> quick, less expensive, readily avaiable, ELISAS have titers
Cons ->less specific, risk of false positives, not as likely to be accepted for regulatory purposes, cross reactivity

Question 16

Pros and cons of AGID and IH

Answer 16

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

Question 17

Pros and cons of FA and IHCs

Answer 17

Pros ->usually quite specific
FAs: fast, visualize pathogen location
IHC: option for fixed tissue
visualize pathogen location

Cons -> rarely an antemortem test, can be expensive, may be species specific, limited assays

Question 18

Why would we pick antigen vs antibody test?

Answer 18

-antigen -> detecting current infection , detects in immunotolerant animals

Antibody ->detecting previous/chronic infection
Detects vaccination that elicits IgG

Question 19

PCR

Answer 19

-real time PCR quantifies viral load
DNA virsues
-reverse transcriptase PCR for RNA viruses

Question 20

Sequencing

Answer 20

Sequencing -> whole genome, metagenomic
-In development/limited use

Question 21

What are some examples of sequencing?

Answer 21

-sequencing mutated FMD
-testing samples for canine respiratory outbreak
-tracking vaccines related ILT outbreak

Question 22

What are the pros and cons for PCR?

Answer 22

Pros -> fast, sensitive, reliable
Cons ->no distinction live vs inactive
may miss if virus mutated
contamination can be an issue
need to know what you are looking for

Question 23

What are pros and cons of sequencing?

Answer 23

Pros -> large amount of data
ID unknown pathogens
track mutations

Cons ->expensive, lots of analyaiss, best on pure sample

Question 24

How do we detect viruses?

Answer 24

viral isolation, electron microspy, histopathology,

Question 25

Describe viral isolation

Answer 25

Goal : grow virus from diagnostic sample then identify usually tissue samples, body swabs as well Gold standard -:only test to prove viable virus is present

Question 26

What cell lines can be used for viral isolation?

Answer 26

embryonic or adult kidney liver others

Question 27

How do we identify that we isolated a virus?

Answer 27

cyotopathic effects electron microsopy PCR sequencing aggluination

Question 28

What are some examples of why we try to isolate viruses?

Answer 28

Detect low viral load: BVD in milk, IBR in semen (grow enough virus to detect) -unknown pathogens

Question 29

What is Electron Microscopy?

Answer 29

transmission electron microscope stained with heavy metal salts visualize surface structures used on diseases samples or purified virus can search blindly can detect primary, secdonary antigens

Question 30

Histopathology?

Answer 30

Visualize viral inclusions inside cells standard H and E stain usually pathopnomonic usually present at a certain stage of infection

Question 31

What are the pros and cons of virus isolation?

Answer 31

Pros -> detecting live virus detect and identify unknown virus Sensitive amplify and collect live virus Cons -> Diffcult, long wait time, expensive, sensitive to contamination

Question 32

What are pros and cons of electron microscopy?

Answer 32

Pros -> visualize pathogen will work even if mutated false positives are rare Cons -> expsensive, less common, longer turnaround time

Question 33

Pros and cons of histopathology?

Answer 33

Pros -> false positives are unlikely inclusions are often distinct and pathopneumonic -Virus is inactivated Cons ->Absences does not equal negative Longer turn around time

Question 34

What swabs are safe to use?

Answer 34

Polyester, dacron, nylon, ryon, Flocked = catch more virus

Question 35

What swabs should we aboid?

Answer 35

wooden handles Calcium alginate ->inactivate virus and interfere with PCR

Question 36

Does a FCoV neagative ELISA mean this cat doesnt have FIP

Answer 36

No, in late stage FIP antibodies drop and may not be detected

Question 37

What are more defintiive options than PCR on biopsy tissues?

Answer 37

Histopathology and immunohistochemistry

Question 38

How do we ship samples for virus isolation?

Answer 38

ship whole fish overnight on icepacks ship overnight on dry ice Formalin inactivates virus