virology

Question 1

diagnostic pathway to virological laboratory tests

Answer 1

history -> physical examination -> “non-specific” laboratory tests -> virological laboratory tests (make list of possible causes then determine which tests to request)

Question 2

what is a common virological symptom in physical examination

Answer 2

rash

Question 3

what 4 things can be detected, and how to detect it

Answer 3

infectious virus (isolation and electron microscopy), protein components (antigens e.g. in CSF, serum), genetic components (DNA or RNA), host response (antibody/cell)

Question 4

5 diagnostic methods

Answer 4

cell culture (very rare - use PCR), electron microscopy (very rare - use PCR), antibody and antigen detection and quantification, genome detection

Question 5

outcome of limitations of laboratory test assays for cell culture and electron microscopy

Answer 5

give rise to false negatives and false positives

Question 6

define sensitivity

Answer 6

test’s ability to correctly identify positive samples

Question 7

define specificity

Answer 7

test’s ability to correctly identify negative samples

Question 8

methods of diagnostics for antibodies and antigens

Answer 8

quantification, serotyping (e.g. HIV)

Question 9

what does quantification of genomes provide, and what is it essential for

Answer 9

viral load, essential for HIV and other blood viruses in immunocompromised

Question 10

2 types of genome sequencing

Answer 10

genotyping, antiviral resistance testing

Question 11

7 samples used for testing

Answer 11

throat (and maybe nose) swab, nasopharyngeal aspirate, bronchoalveolar lavage, endotracheal secretions; stools; urine; CSF; blood (clotted); blood (EDTA); saliva

Question 12

what are throat (and maybe nose) swab, nasopharyngeal aspirate, bronchoalveolar lavage, ET secretions used for and how

Answer 12

detection of respiratory viruses by PCR

Question 13

what are stools used for and how

Answer 13

rotavirus, adenovirus, norovirus antigen detection or PCR

Question 14

what is urine used for and how

Answer 14

BK virus and adenovirus or PCR

Question 15

what is CSF used for and how

Answer 15

herpes viruses and enteroviruses or PCR

Question 16

what is blood (clotted) used for and how

Answer 16

serology (antibody detection)

Question 17

what is blood (EDTA) used for

Answer 17

PCR/viral load testing

Question 18

what is saliva used for

Answer 18

serology and/or PCR (e.g. measles)

Question 19

serology prevalence

Answer 19

most common test

Question 20

serology investigations

Answer 20

HBV surface Ag/Ab, eAg/eAb, core Ab, core IgM

Question 21

serology: IgM vs IgG - when present and duration

Answer 21

both acute phase of disease, but IgM approx 3 months whilst IgG is lifelong

Question 22

antibody avidity testing: when used

Answer 22

IgM is very non-specific (sensitive), so often false positives; use avidity test to determine strength of antibody binding (mix with denaturisation agent to wash away binding; only if non-specific will it wash away and poorly bind)

Question 23

HIV serology technique

Answer 23

4th generation EIA: Ab + p24 Ag detection

Question 24

HIV serology: what must all samples undergo to excluse non-specific reactivity (false positives)

Answer 24

confirmatory testing in second essay

Question 25

HIV serology: what do confirmed positives undergo

Answer 25

typing (HIV 1 vs 2)

Question 26

HIV serology: what is required from all new positives

Answer 26

repeat blood sample and EDTA blood for HIV viral load

Question 27

automated serology equipment

Answer 27

multidisciplinary automated laboratory

Question 28

what is point-of-care testing, and disadvantages

Answer 28

virology test kit (e.g. influenza PCR test kit) at point-of-care to detect virus; not as sensitive or specific

Question 29

when is virus isolation in cell culture used

Answer 29

rarely - only in reference lab

Question 30

disadvantages of virus isolation in cell culture

Answer 30

slow, time consuming, expensive

Question 31

when is virus isolation in cell culture useful

Answer 31

phenotypic antiviral susceptibility testing, but low sensitivity and specificity

Question 32

what is (rarely) used to visualise viruses

Answer 32

electron microscope

Question 33

sample types for electron microscopy

Answer 33

stool, vesicle fluids

Question 34

when is immunofluorescence used

Answer 34

occasionally for direct detection of viral antigens in clinical samples e.g. respiratory virus

Question 35

advantages of immunofluorescence

Answer 35

rapid, inexpensive

Question 36

disadvantages of immunofluorescence

Answer 36

subjective, very dependent on skill of technician and quality of sample

Question 37

what technique is used in testing respiratory tract infections

Answer 37

multiplex PCR

Question 38

what does multiplex PCR allow you to do

Answer 38

look for multiple pathogens, simultaneously, in same sample; amplify viral DNA/RNA to detect if virus using electrophoresis

Question 39

what samples are taken for testing CNS disease e.g. meningitis, encephalitis, and techniques used

Answer 39

CSF for PCR, stools and throat swab for PCR, blood for serology and/or PCR

Question 40

clinical history vs CNS disease

Answer 40

travel, age, immunocompromised; always history and symptoms to be matched up with most relevant diagnostic test

Question 41

what samples are taken if diarrohoea and vomiting

Answer 41

stool (preferred), vomit (lower yield)

Question 42

what techniques are used in investigating diarrohoea and vomiting

Answer 42

PCR or antigen detection assays

Question 43

purpose of PCR

Answer 43

polymerase chain reaction: amplifies specific RNA or DNA sequences

Question 44

PCR method

Answer 44

cycle of: denature (hot) -> primer annealing (cool) -> chain elongation with Taq polymerase using dsDNA (RNA with reverse transcriptase)

Question 45

PCR machines

Answer 45

"real time" PCR, "real time" PCR melt curve analysis (PCR samples melt at different temperatures, so can determine which virus); automated system, viral load system

Question 46

sequencing applications

Answer 46

genotyping, antiviral resistance testing, phylogenetic analysis (determine if transmission between patients)