Brainscape's Knowledge GenomeTM

Browse over 1 million classes created by top students, professors, publishers, and experts.


See full index

Virology > Lecture 3 - Viral Agents 2 > Flashcards

Lecture 3 - Viral Agents 2 Flashcards

1
Q 
Why obtain lab identification of a virus?
1)
2)
3)
4)
A

1) Treatment options
2) Public health measures
3) Surveillance
4) Research

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
2
Q 
Methods of viral identification
1)
2)
3)
4)
5)
A

1) Direct visualisation (EM)
2) Viral cultivation (gold standard)
3) Viral protein detection
4) Serology
5) Viral nucleic acid detection

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
3
Q

How stable are viruses?

A

Typically very susceptible to environmental, chemical inactivation.
Particularly enveloped viruses

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
4
Q 
Ways to preserve viral infectivity for cultivation
1)
2)
3)
4)
5)
A

1) 4 degrees for a day or so
2) -70 for long-term storage
3) -196 (liquid N2) for permanent storage
4) Freeze drying
5) Buffered transfer medium (pH buffered)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
5
Q

Freeze drying preservation

A

Particularly useful for naked viruses.
Dehydration of a frozen suspension in a vacuum.
Used for some live-viral vaccines.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
6
Q

How are PC2 or PC3 viral samples handled?

A

Class II biohazard cabinets

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
7
Q

Why should freeze-thaw be avoided?

A

Forms crystals which shear viruses.

Particularly enveloped viruses

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
8
Q

What is buffering used for?

A

Maintain pH, to maintain a metastable viral state

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
9
Q

Types of samples that are EM’d

A

Messy, crudely prepared samples.

EG: ID’ing rotavirus from faecal samples

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
10
Q

Ways to cultivate virus
1)
2)
3)

A

1) Suckling mice
2) Embryonised chicken egg
3) Mammalian cell line

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
11
Q

Why were suckling mice used?

A

Had no significant immune system

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
12
Q 
Places in an embryonised chicken egg that can be inocculated with virus
1)
2)
3)
4)
A

1) Chorioallantoic membrane
2) Amnotic
3) Yolk sac
4) Allantoic

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
13
Q

Where is influenza best cultivated in a chicken egg?

A

Allantoic innoculation

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
14
Q

Problem with egg innoculation

A

Not all human viruses can replicate in avian cells

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
15
Q

How do viral inocculations of chicken eggs present?

A

As ‘pocks’, which are collections of chicken leukocytes on membranes.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
16
Q

Three mammalian cell culture types used to culture virus

A

1) Primary cells
2) Diploid cell culture
3) Transformed cells

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
17
Q

Primary cell lines

A

Tissue culture, often from animals.

Can only divide up to ten times

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
18
Q

Diploid cell lines

A

Up to 100 generations in culture.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
19
Q

Which cell line are most vaccines cultured in?

A

Diploid cell lines

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
20
Q

Transformed cell lines

A

Aneuploid (abnormal chromosomes)
Often have an activated oncogene
Can divide indefinitely in culture
Not used for human vaccines

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
21
Q

Why aren’t transformed cell lines used for human vaccines?

A

Could transfer an oncogene to human

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
22
Q

Example of primary human cell line

A

Foreskin fibroblast line

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
23
Q

Examples of continuous cell lines

A

HeLa, Vero

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
24
Q

How are viral cell cultures prepared?

A

Cells are inoculated, incubated at 37 degrees for a few days.
Inspected for cytopathic effects

25
Q 
Cytopathic effects 
1)
2)
3)
4)
5)
A

1) Nuclear inclusions
2) Cytoplasmic inclusions
3) Lysis
4) Multinucleate syncitia
5) Transformed cells

26
Q

Cytopathic effects from polio

A

Rounding, nuclear pyknosis

27
Q

Inclusion bodies

A

Represent areas of viral protein production.
DNA viruses cause nuclear inclusions.
RNA viruses cause cytoplasmic inclusions.

28
Q

Viruses that can cause intranuclear inclusion bodies

A

Herpesvirus, adenovirus

29
Q

Cytopathic effects of measlesvirus

A

Synsitia, cell inclusions

30
Q

Viruses that cause multinucleate syncitia

A

HIV, measlesvirus

31
Q

How are multinucleate syncitia formed?

A

Measles fusion protein expressed on cell membrane, which causes cells to stick together

32
Q

How is a plaque assay performed?

A

Monolayer of mammalian cells, inoculated with virus.
Sandwich monolayer between two agar gels to immobilise virus.
Observe areas of cell lysis

33
Q

How can cell transformation occur?

A

Some viruses encode oncogenes (EG: retroviruses)

34
Q

Direct detection of viral antigen

A

Fluorescently tagged anti-antigen antibody binds immobilised viral antigen

35
Q

Indirect detection of viral antigen

A 
Antibody against viral antigen immobilised.  Binds viral antigen.
Another antibody (fluorescently tagged) against viral antigen is washed over sample.
36
Q

Haemadsorption

A

Some viral antigens cause red blood cells to agglutinate

37
Q

Name for agglutination of red blood cells

A

Haemadsorption

38
Q

Haemagglutination assay

A

Often used for influenza.
Measures ability of viral particles to agglutinate red blood cells.
Serial dilution of viral sample with constant concentration of red blood cells.
The maximum dilution that results in a shield (rather than a dot) is the haemagglutination titre

39
Q

Haemagglutination inhibition assay

A

Constant virus, red blood cell concentration.

Serial dilutions of antibody. See when it inhibits haemagglutination (dot)

40
Q

What is haemagglutination inhibition used for?

A

Serotyping

41
Q

How do viral titres vary between assays?

A 
In descending order of viral particles:
Electron microscopy
Quantal assay in eggs
Plaque assay in mammalian cells
Haemagglutination asssay
42
Q

Electron microscopy influenza viral titres per mL

A

10^10 virions

43
Q

Quantal assay in eggs influenza viral titres per mL

A

10^9 egg ID50

44
Q

Plaque assay in mammalian cells influenza viral titres per mL

A

10^8 pfu

45
Q

Haemagglutination assay influenza viral titres per mL

A

10^3 HA units

46
Q

Antigen capture assay

A

Known antibody affixed to plate. Binds to antigen of interest. Sample washed over antibodies. Second antibody - with a fluorescent tag - which binds to antigen is washed over plate.

47
Q

Anti-viral antibody assay

A

Viral antigen affixed to plate. Patient serum washed over plate. If patient has antibodies against antigen, these will bind. Fluorescently-tagged anti human Fc antibody washed over.

48
Q

Antigen capture assay vs anti-viral antibody assay

A

Antigen capture has high sensitivity

Anti-viral antibody assay has high specificity

49
Q

Example of antigen capture assay

A

HIV p24 assay

50
Q

Example of anti-viral antibody assay

A

HIV western blot

51
Q 
Western blotting 
1)
2)
3)
4)
A

1) Run antigen samples on a gel
2) Blot gel onto nitrocellulose gel
3) Immunostaining of blot with a tagged antibody
4) Autoradiography of blot

52
Q

HIV western blot asssay
1)
2)
3)

A

1) Detergents used to solubilise cells with potential HIV infection, vs wild-type HIV proteins
2) Western blot performed with anti-HIV antibodies
3) High specificity.

53
Q

Detection of viral antigen vs detection of host antibody assays

A

Detection of viral proteins has a high sensitivity, therefore false positives
Detection of host antibodies has a high specificity, therefore false negatives

54
Q

Problems with infectivity or serology assays

A

Generally slow. Too slow for clinical use

55
Q 
Viral nucleic acid detection assays
1)
2)
3)
4)
5)
6)
A

1) Southern blot for viral DNA
2) Northern blot for viral RNA
3) PCR for DNA
4) Reverse transcriptase PCR for RNA
5) Viral nucleic acid sequencing
6) DNA microarray

56
Q

PCR assay pros and cons

A

1) Highly sensitive, specific
2) Fast
3) Cheap
4) Prone to contamination
5) Low amount of sample needed

57
Q

Difference in PCR technique for DNA vs RNA virus

A

For DNA virus, normal PCR

For RNA virus, need a viral reverse transcriptase enzyme

58
Q

Stages of PCR

A

1) Denaturation (DNA splits)
2) Annealing (primers bind)
3) Extension (DNA polymerase)

59
Q

How was it initially known that SARS was novel

A

No other human coronavirus could grow in Vero cells

Virology (29 decks)

Brainscape helps you reach your goals faster, through stronger study habits.
© 2024 Bold Learning Solutions. Terms and Conditions