Laboratory Diagnosis

Question 1

Techniques

Answer 1

• Microscopy
• Isolation and culture
• Detection of antibodies/ antigen
• Identification of a microbial gene or product

Question 2

Describe microscopy

Answer 2

-Most common method for the detection of microorganisms directly in clinical specimens
-Cheap
-Operator dependent
-Types:
=Light
=Phase contrast
=Fluorescent
=Dark-field
=Electron (virus)

Question 3

Microscopy stains

Answer 3

• Gram (+ve and -ve)
• Ziehl-Neelsen (mycobacteria)
• Giemsa (blood films)
• Potassium hydroxide (fungi)
• Lugol (parasite)
• India ink (fungi)

Question 4

Types of solid and liquid media in bacterial culture

Answer 4

-Non selective
-Selective
=Addition of antimicrobials or specific substrates that are designed to support the growth of the desired organism and suppress others
-Broth (fluid)- inoculate suspension in nutrients (blood culture)

Question 5

Advantages and disadvantages of bacterial culture

Answer 5

-A:
=Cheap
=Able to isolate the organism for susceptibility testing
=Able to isolate DNA

-D:
=Labour intensive
=Requires significant staff expertise
=Results can be affected by prior administration of antimicrobials
=Success depends on extrinsic circumstances such as incubation conditions for the organisms in question
=Results can be delayed

Question 6

Non culture techniques

Answer 6

-Antigen detection:
=Latex agglutination
=ELISA
=Toxin detection

-Nucleic Acid Amplification Techniques

Question 7

Describe Latex agglutination

Answer 7

-Antibodies coated onto Latex particles that react with the presence of the organism or its product
=E Coli 0157
=Pneumococcal antigen in urine
=Cryptococcus neoformans in CSF
=Histoplasma capsulatum
=Legionella antigen in urine

Question 8

Describe ELISA

Answer 8

-Immunoassays used to measure antigen concentration
=CMV
=HIV
=VZV
=HBsAg
=Dengue virus
=RSV
=Legionella pneumophila
=S. pneumoniae
=Hepatitis C

Question 9

Advantages and disadvantages of antigen/ antibody detection methods

Answer 9

• More rapid diagnosis than culture
• Risk of cross reaction with other antigens (false positive, IgM)
• Sensitivity and specificity are limiting factors

Question 10

Toxin detection methods

Answer 10

-Detection of exotoxins:
=Use similar methodology to ELISA (clostridium difficile cytotoxin)
=Bioassays: clostridium botulinum by injection of patient’s serum into mice

Question 11

Nucleic Acid-based tests

Answer 11

-Non-amplified
=Have loss relevance due to technical complexities and poor sensitivity
=PNA-FISH (peptic nucleic acid probes- fluorescent in situ hydribidation)

-Nucleic Acid Amplification Techniques (NAAT)
=PCR
=NASBA (nucleic acid sequence based amplification)

-Whole genome sequencing

Question 12

What is PCR used for?

Answer 12

-Commonest way to detect viruses
-Respiratory specimens
-Genital/ ulcer/ skin
-CSF
-Blood
-Stool
=Influenza virus
=RSV
=Mycoplasma
=Parainfluenza virus
=Adenovirus
=Enterovirus
=Pneumocystitis jiroveci
=Norovirus
=Mycobacterium tuberculosis

Question 13

Uses of whole genome sequencing

Answer 13

• COVID

- Mycobacterium tuberculosis

Question 14

Mandatory data set for ordering investigations

Answer 14

• CHI number
• Surname and Forename
• Date of birth
• Requesting location
• Contact details for an urgent result to be communicated to
• Sex