Pathogenesis and Diagnosis of Viral Infection Flashcards

Question

Koch's Postulates

Answer 1

**_Published in 1980, 4 criteria to establish causative relationship between microbe and specific disease:_** 1. The microbe must be found in organisms afflicted by the disease, but not in healthy organisms 2. The organism must be isolated from the disease host and grown in culture 3. The cultured microbe should cause disease when introduced into a healthy organism 4. The same organism must be re-isolated from the experimentally infected host

Answer 2

* Some microbes with pathogenic potential can be found in asymptomatic hosts (carriers): - Vibrio cholera, many viruses (not discovered in Koch’s day), Poliovirus causes paralytic disease in ~1% ofthose infected, FeLV causes leukaemia in ~1% persistent infections * Not all pathogens can be grown in culture or no suitable animal model exists * Not all potential hosts exposed to infection will develop disease

Answer 3

* Present in **most** cases of an infectious disease * Found preferentially in those organs known to be diseased (not those lacking pathology)

Answer 4

**Blood sample** **-**Clotted sampled for serum -EDTA anticoagulant for cells (haematology) **Tissue Biopsy** - Lymph Node - Lesion **Sampling**- If unsure contact lab prior to sampling to find out: Amount of material required, suitable swab material, transport medium (with/without antibiotics), refrigeration

Answer 5

**Detection of pathogen at site of infection:** * Observation of **viral pathogenesis (histopathology)** * Direct visualisation of **virus particles (morphology)** * Presence of **virus genome (nucleic acid)** * Presence of **virus antigen (protein)** **Evidence of host immune response to infection**: - **Antibodies in serum (serology)** - **(Cell-mediated immune response)**

Answer 6

**Grow virus in the laboratory, observe viral pathogenesis** **_Important Concepts:_** * Viruses need cells to replicate * Virus has to attach to cell to infect * Viral surface protein(s) bind to receptor on host cell surface * Species specificity and tissue specificity * Cytopathic effects on cultured cells **E.g. cytotoxicity, inclusion bodies & syncytia** **Advantages**: * Virus is amplified and therefore easier to detect **Disadvantages:** * Need to have a suspect in mind * Takes several days (depending on virus) * Requires live virus * Not all viruses can be cultured * Further identification required

Answer 7

**Negative Staining** * Uses heavy metal stains that don’t bind to sample but stain the background * Stain is non-permeable for electrons & appears dark on image * Virus= **permeable & appears light in colour** * Can Identify virus based on its **morphology**: Type of **capsid symmetry & whether it is naked/ enveloped** **Advantages**: * Can be used for viruses that can’t be cultured - Allows identification of new agents **Disadvantages:** * Requires specialised equipment & experienced personnel * Low sensitivity may require concentration of sample * Often requires purification of sample

Answer 8

* Polymerase chain reaction (PCR) * Based on ability of **Taq polymerase to amplify DNA** * Use short **oligonucleotide primers**, specific for virus sequence * Generation of amplicon of predicted size indicates **presence of virus nucleic acid in sample**. * DNA vs RNA viruses - detection of RNA viruses - PCR template = **DNA or cDNA** - RNA might need to be converted--\> cDNA via **Reverse Transcription-PCR** **Advantages:** * Very sensitive, can use very small amounts of starting material * Does not require live virus * Specific as primers are designed for target DNA * **Real time PCR**- allows quantification, uses fluorescent technology- quicker as no gel stage **Disadvantages:** * Very sensitive, danger of contamination (can--\> **false positives)**

Answer 9

- **Monoclonal** antibodies: **specific for one antigenic epitope** - **Polyclonal** antibodies: mixture of antibodies, **detect several epitopes**, e.g. anti-serum from immunised animal

Answer 10

**(biological fluids)- aka a Sandwich ELISA** * Plates coated with antibody specific to virus * Virus in sample binds to antibody * Second virus-specific, **enzyme-labelled** antibody is added * Enzyme acts on substrate to generate a **colour change**

Answer 11

-(tissue sections/ cells, smears) * Virus antigen in cells is detected by specific antibody * **Antibody carries a label**: - Either fluorescent (**immunofluorescence assay)** - Or enzyme which reacts with colour substrate **(immunoperoxidase** assay) **Example**: BVDV can be detected in epithelium of ear notch biopsy

Answer 12

(biological fluids) * Rapid Immunomigration used- **FeLV testing** * Fast, easily transportable kit

Answer 13

**Advantages:** - fast - Does **not** require live virus - Specific **Disadvantages:** -No amplification included, relatively **↓ sensitivity**

Answer 14

Diagnostic methods based on identifying host immune response * Antibodies are evidence of previous infection * Usually antibodies are detectable from **1-2 weeks post exposure- lag phase**

Answer 15

1. **When animal shows clinical signs of disease (acute sample)** 2. **2-3 weeks later when animal has recovered and produced antibodies (convalescent sample)** * _But useful for:_ - Diagnosis of persistent infections e.g. FIV - Checking disease-free status of herd - Screening for exposure to exotic diseases - Assessing vaccination status * **Rising antibody titre shows that recent infection has taken place**

Answer 16

* Some tests are able to detect IgM which indicates an **early immune response to infection** and therefore requires only one sample (at time of clinical signs) * IgM is formed primarily in infection

Answer 17

* Serum sample is mixed with virus * Antibodies in patient’s sample bind to virus * Mixture is added to cells * If antibodies present: virus can’t infect cells as bound by antibodies, cells remain healthy * If no antibodies present: virus infects cells, cytopathic effect is detected

Answer 18

* Based on ability of some viruses to **agglutinate red blood cells** ***(particularly influenza)*** * Serum sample is mixed with virus * If antibodies are present in the serum they bind to virus * Red blood cells are added to mixture * If **antibodies were present in serum**: _virus is bound_ and red blood cells **don’t agglutinate** * **If no antibodies** were present virus is free and can **agglutinate red blood cells**

Answer 19

* Plate is coated with antigen (whole virus/ purified virus protein) * Antibody in serum sample binds to antigen * Second antibody which recognises IgG is added * Second antibody carries an enzyme label * Enzyme converts colour substrate and colour change is detected

Answer 20

**Advantages**: - Cheap - Specific - Relatively Sensitive **Disadvantages** - Majority of Tests only allow **retrospective diagnosis** - Negative test does not mean no exposure **[NB LAG PHASE]** - Antibody Titre

Answer 21

* To assess quantity of antibodies in a patient’s sample * Serial dilution of serum sample * All dilutions are tested for antibodies * Last positive dilution gives the antibody titre

Pathogenesis and Diagnosis of Viral Infection Flashcards

(46 cards)