Diagnostic

Question 1

What are the principles for sample collection

Answer 1

1. Specimens obtained aseptically representative of disease
2. A sufficient quantity of material
3. Specimens collected prior to antimicrobial therapy
4. If cultures not immediately initiated must be refrigerated

Question 2

What needs to be on a sample submission form

Answer 2

1. Case history - clinical signs previous treatment ect.
2. Required tests - what does clinician want to know
3. Sample type - which is most appropriate urine blood ect.

Question 3

What does microscope observation tell us and why is it good

Answer 3

Cost effective, rapid, shows presence of bacteria/fungi, number of organisms, can check morphological characteristics and stain - gram positive, provides info on host cellular response.

Question 4

What stain can we use for fungal identification

Answer 4

KOH 10% with Parker blue acts as a clearing agent, when Parker ink is added the fungal spores take up the ink and appear blue-violet

Question 5

How to use microscopic observation in virus

Answer 5

Pox inclusion is characteristic of a virus. Only other way is electron microscope

Question 6

What does bacterial culture aim to achieve

Answer 6

1. Isolation of organism in pure culture inoculating the specimen onto appropriate artificial culture media
2. Identification by microscopic examination, biochemical reactions, state of the art techniques
3. Antimicrobial susceptibility testing after growing in pure culture

Question 7

What are the different types of media

Answer 7

1. Enrichment media - supplemented by blood or other nutrients to encourage growth
2. Differential media - distinguish between different bacteria on basis of their biological characteristics
3. Selective media - favour growth of particular microorganisms and inhibit growth of others

Question 8

When using 5% sheep blood agar what will u see

Answer 8

Non-haemolytic the agar stays red.
Beta-haemolytic the agar turns yellow.
As some bacteria secrete enzymes that lyse the erythrocytes in agar producing zones of haemolysis

Question 9

What happens with gram staining

Answer 9

Stages = fixation, crystal violet, iodine treatment, decolorisation, counter stain with Safranin.
Gram positive stop at crystal violet stage and stay purple
Gram negative go all the way to the end and go pink

Question 10

What does the oxidase test for and show

Answer 10

To determine the presence or absence of specific metabolic enzymes in an isolate. Here is to test for the presence of cytochrome C oxidase, this enzyme reduces TMOD a redox die
Oxidase positive = dark blue/purple
Oxidase negative = colourless

Question 11

What does the catalase test check for and show

Answer 11

Detects for the presence of the catalase enzyme. If present the external hydrogen peroxide is Brocken down to oxygen and water. Seen as bubbles

Question 12

Summary of what can we use to identify bacteria

Answer 12

Oxygen requirements
Culture characteristics
Staining properties
Microscopic morphology
Biochemical reactions
MALDI-TOF skips steps can do many at once

Question 13

Once identified what do we test for

Answer 13

Antimicrobial resistance
Qualitative methods - disc diffusion. Disc impregnated with antimicrobial resistance agents are placed onto the agar leave over night to measure zones of inhibition.
Bacteria can be susceptible and inhibited so can be treated with normal, dose of antibiotic . Or resistant which means the infection could not be treated with normal dose. Or it can be an intermediate which is a buffer zone to avoid misinterpretation

Question 14

How do viruses differ to bacteria

Answer 14

Obligate intracellular organisms (tissue cell cultures, embryonate eggs, experimental animals)
Need specialist lab to grow
Must inhibit bacterial growth
Not all virus grow in culture
Takes a long time very labour intensive

Question 15

What effects might we see if a virus is present in culture

Answer 15

Rounding of cells
Syncytia
Inclusion bodies

Question 16

What is viral haemagglutination

Answer 16

When virus clump red blood cells preventing them from settling at the bottom of a well
Viral glycoproteins cause this - hemagglutinins

Question 17

How can we obtain a sample off a fungal infection

Answer 17

Plucking hairs from lesions
Toothbrush
Skin scrapings
Exudates
Biopsies

Question 18

How is fungi cultured

Answer 18

Sabouraud Dextrose Agar (SDA) low pH (5.5) which inhibits the growth of most bacteria. Incubated at 25 degrees and 35 degrees for 1-3 weeks.
Fungal identification is based off macroscopic and microscopic morphology

Question 19

What is an indirect method of diagnosing ring worm (Microsporum canis)

Answer 19

UV fluorescence. Some fungi produce metabolites that fluoresce a green colour

Question 20

How can we use an Antibody Antigen test

Answer 20

1. Known antigen added to detect specific antibodies in animal e.g. elisa
2. Known antibodies to detect bacterial or viral antigens e.g. immunofluorescent antigen test

Question 21

What is a serological test and what can it detect

Answer 21

1. The test detects for the presence of antibodies
   So see if the animal has ongoing infection, past exposure to a pathogen, maternally derived antibodies or been vaccinated
   Examples = Brucella canis or FIV

Question 22

What is a delayed hypersensitivity reaction

Answer 22

Skin tuberculin test for bovine TB.
The intradermal injection of bovine TB or purified protein derivative. Then follows subsequent swelling at the site of injection 72 hours later
Positive test = an increase in 4mm in skin fold thickness in 72 hours

Question 23

What are the advantages and disadvantages of PCR

Answer 23

Advantages = very sensitive, detects microorganisms that are unculturable or slow growing, faster result
Disadvantages = susceptibility of contamination, failure to differentiate live and dead organisms

Question 24

What is PCR used for

Answer 24

Mycoplasmas
Mycobacteria
Spirochetes
Viruses
Rapid detection of MRSA

Question 25

What is rapid molecular diagnostics

Answer 25

Single step cartridge based molecular testing devise. Targets pathogens implicated in clinical syndromes all at once. But be aware as a large number of clinical specimens may contain unknown pathogens. It can miss some

Question 26

What is point of care testing good

Answer 26

Rapid result so appropriate treatment can be implemented almost immediately.
Example = snap test of FeLV and FIV

Question 27

What are the sources of error in diagnostic test

Answer 27

No test is 100% accurate
2 sources of error = test (false positive/negative) or interpreter of the test

Question 28

What is sensitivity in an analytic term and clinical term

Answer 28

Analytical term = the detection limit (lowest amount a test can detect) defined at 95%
Clinical term = the tests ability to correctly designate an individual with the disease as positive to the test

Question 29

What is specificity in an analytical term and a clinical term

Answer 29

Analytical term = the ability to not react with other substances other than the analyte of interest
Clinical term = the test ability to correctly designate an individual who does not have the disease as negative to the test