MB practical 2 - viral detection

Question 1

what are the 2 main approaches to detecting a virus?

Answer 1

• direct - detection of the virus itself
• indirect - detection of an immune response to the virus using serology

Question 2

why is it more difficult to test for viruses than bacteria?

Answer 2

• not visible under a light microscope
• can only reproduce in cells and not on an agar plate like bacteria can

Question 3

EM and CPE are usually not used in the lab to identify viruses, what are these methods and how are they used to identify bacteria?

Answer 3

electron microscopy:
- nearest equivalent to a gram stain
- used to visualise the morphology of the virus in black and white

cytopathic effect:
- nearest equivalent to seeing colonies on an agar plate
- viruses are cultured in cell culture over 2-4 weeks and the effects they have on host cells is observed

Question 4

why are EM and CPE not used in real life labs as a method of virus detection?

Answer 4

• take a long time and effort
• cannot put through all clinical specimens

Question 5

as EM and CPE are not viable methods of viral detection for clinical lab use, what methods are used instead?

Answer 5

• PCR - polymerase chain reaction
• NAAT - nucleic acid amplification

Question 6

how does a PCR test work?

Answer 6

• using enzymes to unzip chains of rna or dna
• then makes complimentary copies of the originsl dna / rna by using nucleoside bases to double the original number os bases
• the nucleoside substrate bases are each tagged with a fluorescent molecule (only fluoresces when bound in a chain of other bases)
• the creation of new dna / rna will therefore be detected and quantified by measuring increases in flourescence

Question 7

what factor MUST happen for the PCR to work?

Answer 7

• the original primer sequence in the pcr is able to recognise and bind the viral nucleic acid and is complimentary

Question 8

outline the pros and cons of a PCR test.

Answer 8

pro:
- very sensitive
- very quick

cons:
- because it is so sensitive it is vulnerbale to laboratory contamination
- can only detect nucleic material complimentary to the primer sequence to initiate it so you need to know the sequence of the viral nuclein material
- its not possible to detect a virus unless you detect it in the first place and use the relevant primers

Question 9

name some NAATs other than PCR which can be used to detect viral nucleic material.

Answer 9

• ligase chain reaction
• nucleic acid sequence based amplification
• transcription mediated amplification
• branched chain dna
• nucleic acid hybridization

Question 10

define serology.

Answer 10

• study or detection of antibody responses in the serum
• especially IgG and IgM class of antibody

Question 11

name the different types of serology tests.

Answer 11

• ELISA - enzyme linked immunosorbent assay
• CFT - complement fixation test\HA/HAI - haemagglutination / haemagglutination inhibition
• RIA - rdioimmunoassay
• IF - immunoflourescence

Question 12

outline the principle of how a serology test works and when they are used to test for a recent viral infection.

Answer 12

• test for the presence of IgG and IgM antibodies
• IgM presents initually up to 12 weeks of an actue infection
• IgG appears later and remains for life or many years after the infection

Question 13

name the herpes virus 1-8.

Answer 13

• HSV-1
• HSV II
• VZV (zoster)
• EBV (epstein Barr)
• CMV
• HHV6
• HHV7
• HHV8

Question 14

name the important resp viruses.

Answer 14

• influenza a
• influenza b
• parainfluenza 1-4
• respitory syncitial virus
• corona viruses including sars
• rhinovirus
• measels
• mumps
• rubella
• metapneumovirus
• paravirus
• adenovirus

Question 15

name the viruses associated with diahhroa outbreaks.

Answer 15

• rotaviru
• norovirus