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Principles of Molecular Virology > Lecture 1 > Flashcards

Lecture 1 Flashcards

1
Q

Definition of a virus (3 parts)

A

An ultramicroscopic (20nm-400nm) metabolically inert, infections agent,

that replicates only within cells of living hosts, mainly bacteria plants and animals,

composed of an RNA or DNA core, a protein coat and in more complex types, a surrounding envelope.

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2
Q

Three ways of escaping our body’s defences

A

The city virus: changing quickly to escape becoming recognisable to the immune system, playing a fitness cost game.

The buy what I can afford virus: been around long enough to have a safe way to escape from our immune system.

The infect whatever moves virus: not particularly good at escaping defences, just highly infectious so move onto next host before response kicks in

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3
Q

What are Koch’s postulates?

A

Four criteria that must be adhered to to identify the causative agent of a particular disease.

  1. The organism must always be present, in every case of the disease.
  2. The organism must be isolated from a host containing the disease and grown in a pure culture.
  3. Samples of the organism taken from pure culture must cause the same disease with inoculated into a healthy, susceptible animal in the laboratory.
  4. The organism must be isolated from the inoculated animal and must be identified as the same original organism first isolated from the originally diseased host.
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4
Q

What do they do? Basic functions.

A

Introduced into the population. Spread. Escape bodily defences. Cause disease. Major cause of mortality or morbidity worldwide. Attract publicity.

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5
Q

How are viruses classified?

A

Based on what their genetic material is. The Baltimore classification.

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6
Q

Class I?

A

Double-stranded DNA virus

Herpesviruses: herpes simplex, chickenpox, EBV,
Papillomaviruses: warts (cerival cancer)
Poxiviruses: small pox

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7
Q

Class II?

A

Single-stranded DNA viruses

Anelloviridae, Circoviridae, and Parvoviridae (which infect vertebrates), the Geminiviridae and Nanoviridae (which infect plants), and the Microviridae (which infect prokaryotes).

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8
Q

Class III?

A

Double-stranded RNA viruses

Reoviridae (reovirus, rotavirus) and Birnaviridae.

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9
Q

Class IV?

A

Single-stranded RNA viruses - Positive-sense

Picornaviruses: polio, rhinovirus (cold), hepatitis A, hepatitis C
Flaviviridae: Dengue, West Nile Virus

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10
Q

Class V

A

Single-stranded RNA viruses - Negative-sense

Arenaviridae, Orthomyxoviridae (influenza), Paramyxoviridae, Bunyaviridae, Filoviridae (Ebola) and Rhabdoviridae (rabies)

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11
Q

Class VI

A

Positive-sense single stranded RNA viruses that replicate through a DNA intermediate

HIV and HTLV

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12
Q

Class VII

A

Double-stranded DNA viruses that replicate through a single stranded RNA intermediate

Hepatitis B

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13
Q

What is the extracellular phase of the virus called?

A

virion

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14
Q

What is the range of the viral genome in bp?

A

5000-230000

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15
Q

What is the capsid?

A

Highly repetitive protein ‘casing’ that surrounds the nucleic acid

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16
Q

Complex visions can also be surrounded by?

A

A membrane, making them an enveloped virus.

Without the membrane, naked virus.

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17
Q

How is attachment to host cell mediated?

A

By proteins or spikes on surface of virus, which bind to host surface molecules, often glycoproteins, known as viral receptors.

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18
Q

How does enveloped virus enter host cell?

A

Membrane fusion followed by endocytosis.

Naked virus just endocytosis.

19
Q

How does Class I replicate?

A

Viral genome must be transported to nucleus.

  1. Viral DNA transported to host nucleus.
  2. Host RNA polymerase converts viral DNA to viral mRNA in nucleus, then transported to cytoplasm.
  3. Viral RNAs translated to new viral proteins, transported back to nucleus.
  4. Viral DNA replicate by host DNA polymerase and viral factors.
  5. Reassembly of viral genome and proteins, followed by release.
20
Q

How does Class IV replicate?

A

Viral genome can stay in cytoplasm.

  1. Viral RNA translated to proteins by cytoplasmic ribosomes.
  2. The viral RNA genome is replicated by viral synthetase.
  3. Viral proteins form the capsid and encapsulate the replicated RNA genomes forming new visions.
  4. The new visions bud from the infected cell.
21
Q

How does Class V replicate?

A
  1. Taken up by endocytosis.
  2. RNA negative strand converts to mRNA using viral RNA polymerase.
  3. mRNA translated in cytoplasm to produce viral proteins (HA, NA, NP, RNApoly, M2 protein).
22
Q

How does Class VI (retrovirus) replicate?

A
  1. Entrance via binding and fusion of viral envelope proteins to specific host cell receptors.
  2. Virion uncaring.
  3. Viral reverse transcriptase reverse transcribes one RNA strand into ssDNA and then into dsDNA, enters nucleus.
  4. The viral DNA integrates into host DNA, through function of viral integrate, provirus is formed.
  5. Transcription of viral DNA to viral mRNA and progeny RNA.
  6. Synthesis of HIV poly proteins.
  7. Processing of HIV-polyproteins by the viral protease and generation of mature functional proteins.
  8. Encapsidation of viral ssRNA.
  9. Budding of visions.
  10. Release of virions.
23
Q

How does Class VII replicate?

A
  1. HBV binds to host cell membrane.
  2. Fuses with membrane, core enters cytoplasm.
  3. Double stranded genome is completed by enzymes contained within the core.
  4. Entry of DNA into nucleus and transcription of DNA to genomic and mRNA.
  5. RNA enters cytoplasm and viral proteins translated.
  6. Core assembles around + sense RNA, and + sense RNA copied to - sense DNA.
    • Sense DNA partly copied to + sense.
  8. Core associated with HbS antigen in cell membrane.
  9. Virus buds through host cell membrane and virus with partly double strand DNA leaves the cell.
24
Q

What are consequences of viral infections?

A

Cell transformed to tumour cell.

Death of cell and release of virus in lytic cycle.

Slow release of virus without cell death, persistent infection (for a lifetime).

Latent infection if virus infects cell but does not initially ‘act’ late onset lytic infection.

25
Q

How do we detect viral infections?

A
  • Plaque forming units
  • Enzyme linked immunoassay (EIA)
  • Immunofluorescence (IF)
  • Serology
  • Electron microscopy (EM)
  • PCR
26
Q

How does the Plaque forming unit assay detect viral infections?

A

Measures infectious visions. Diluting virus stock until individual plaques are countable on cell plate. Calculate original virus number in stock.

27
Q

Advantage to Plaque forming unit assay?

A

Detects infectious units, not just virions present.

28
Q

Disadvantage to Plque forming unit assay?

A

Slow (2days-3weeks), labour intensive, requires skill/experience.

29
Q

How does IF detect viral infections?

A

Flurochrome-tagged antisera used to detect viral antigens in cells from specimen.

30
Q

Advantage to IF?

A

Quick (1-3h), semi-quantitative. Feedback on specimen quality.

31
Q

Disadvantage to IF?

A

Requires specific antisera, ruthlessly assessed. Requires skill and experience. Requires fluorescence microscope.

32
Q

How does EIA detect viral infections?

A

Similar to IF, but using an enzyme as a tag and usually done in micro titre plate. Measures antigen levels from samples.

33
Q

Advantage to EIA?

A

Quick (2-3h), can be read by machine, suitable for large numbers. No special skills required. Commercial kits available.

34
Q

Disadvantage to EIA?

A

Requires specific antisera, ruthlessly assessed. No feedback on specimen quality, problems over false positives and borderline results. Uncertain sensitivity.

35
Q

How does EM detect viral infections?

A

Use high-resolution to visualise virus particles directly in specimen.

36
Q

Advantages to EM?

A

Very quick (15min-3hr), specific, can detect mixed infection. Most conclusive.

37
Q

Disadvantages to EM?

A

Expensive machinery. Requires trained microscopist.

38
Q

How does Serology detect viral infections?

A

Detection of antibody response to virus.

39
Q

Advantages to Serology?

A

Automated and read by machine. Presence of IgM indicates recent stimulus.

40
Q

Disadvantage to Serology?

A

Test can be quick, but patients response to virus can take up to 10 days to develop. All results require interpretation.

41
Q

How does molecular amplification methods such as PCR detect viral infection?

A

Amplification of viral nucleic acid to assessable levels. Measures the genomic copies of the virus.

42
Q

Advantage to PCR

A

Highly sensitive, can be same day. Can detect mixed infections and can be automated for larger numbers.

43
Q

Disadvantage to PCR?

A

May be over-sensitive. No indication of specimen quality. Expensive. Requires good technique to avoid cross-contamination.

Principles of Molecular Virology (4 decks)

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