L3 Laboratory Diagnosis Flashcards
Role of laboratory diagnosis
Carry out tests in support of clinical diagnosis of infected disease etc
Function
- Investigative advice
- Interpreting raw data
- Reporting of results
- Provision of treatment advice
What are the types of specimen used?
Urine/ swab/ tissue/ blood/ sputum/ faeces
Methods of microbiological diagnosis
Direct detection - look at sample down microscope
Culture - grow in lab and investigate it
Immunological test - not able to grow or see under microscope but able to see effects on body
Microscopy
e.g. CSF examination in meningitis
Via gram method = detect presence of gram -ve diplococci in diagnosis (meningococcal meningitis)
Pros: Quick and provide info on how to treat
Detect antigen
e.g. C.difficile (using glutamate dehydrogenase
Antigen-antibody complex is added to solution coupled to a dye (dye changes colour is correct complex is present)
- Lateral flow assay: cheap and rapid but not accurate
- 96-well format: manual or semi-automated
- Well based format: automated
Detect nucleic acids
- Can be specific (directed against 1 target)
- Or amplify ‘unknown’ sequences using universal primers targeting 16s rRNA (matching to database)
What are the uses and limitations of direct detection?
- Establish presence of microorganism at a particular site
- Allow use of appropriate empirical antimicrobial therapy
- Provides epidemiological info
- No info on susceptibility or typing
- Fast diagnostic method
Culture requirements
Need culture medium (provides nutrients required for growth) and incubation (physical requirements for growth)
Culture medium
- Non-selective: Growth of all microorganisms
- Selective: Inhibits growth of specific microorganisms (inhibitory components to certain microorganisms)
What are indicator mediums used for?
Indicates certain properties of bacteria e.g. sugar fermentation
MacConkey (indicator medium)
Contains sufficient water and nutrients to support bacterial growth
Dye turns red in presence of acid
Ferment lactose = red (acid) produced)
Can’t ferment lactose = yellow
CLEO (indicator medium)
Lactose fermenter appears yellow
Chromogenic agar
Often able to indicate more than 1 bacterial species
e.g. differentiate between E.Coli, Kiebsella spp. and Proteus spp. in urine
Gram stain (Bacterial identification test)
Distinguish between bacterial cell wall
Ability to retain crystal violet
Gram +ve: purple
Gram -ve: pink (counterstain)
Haemolysis (bacterial identification test)
Breakdown of blood cells
a = opaque
b = completely clear
Oxidase test (bacterial identification test)
See whether have pseudomonas
oxidise to a purple colour
Biochemical tests
- API strips contain lots of biochemical reactions all in 1 strip
- Produces profile to aid identification
- assign score to each positive test in a group of 3 tests
- Unique score from 0 to 7 is obtained
- Compare with database
MALDI-TOF
- Mass spectrometer produces spectrum of bacterial proteins
- Compared to database of known spectra
- Cheap and fast
- Only as good as database
Uses and limitations of bacterial identification tests
- Establishes presence of microorganism at particular site
- Allows use of empirical and targeted antimicrobial therapy
- Provide epidemiological and typing of information
- Usually slower than direct detection
Susceptibility testing
Requires viable microorganisms (usually bacteria or fungi)
Methods
- measure zone of inhibition
- commercial e-test
- breakpoint plates
Uses and limitations of susceptibility testing
- inform decisions on targeted antimicrobial therapy
- initial treatment with ‘empirical’ therapy
- subsequent treatment is ‘targeted’
requires isolation of microorganism and antimicrobial susceptibility testing
Immunological tests
IgM detection
Seroconversion = Change from -ve to +ve result from one test to subsequent test
Fourfold rise in titre = rise in concentration of antibody from one test to subsequent test
Non-antibody immunological test
IFN-Y release assays in TB
Uses and limitations
- Confirms exposure to specific microorganisms
- Provides epidemiological info
- Restricted to patients with detectable antibody response
- Is retrospective (too late to inform antimicrobial therapy decisions)
