Diagnostic microbiology Flashcards
What are some examples of culture independent tests?
molecular testing (incl. toxin detection, NA amplification)
Rapid tests and immunoassays (biochemical identification, ELISA)
Microscopy
What are some examples of culture dependent tests?
Biochemical tests and culture
What is sensitivity?
no false positives, everyone with disease gets diagnosed
What is specificity?
there are no false positives, no one without the disease gets diagnosed
Biochemical tests
Usually based on differential features of closely related organisms or those from similar sites
e.g. g- or +, catalase, oxidase, urease, grows on certain agar
Speeding up biochemical tests
rolled into kits for specific organisms
- API test strips: series of cupules containing various freeze dried reagents and color indicators designed for biochemical tests
- automated culture systems
What is commonly diagnosed with staining and why?
Tuberculosis ~ 36.8% sensitivity, used because it is cheap and easier in low resource settings
Microscopic detection
- can examine smears directly with stains
- or can examine the binding of fluorescent antibodies > often used to view difficult to grow pathogens
Molecular detection methods
PCR ~ used for difficult to grow organisms, or when speed of diagnosis is essential
DNA sequencing ~ used to confirm other tests
- dependent on the sample - if there is too much DNA it can be bad
GeneXpert for TB diagnosis
automates DNA extraction and amplification form sputum samples
MALDI-TOF MS
sort of culture dependent
Rapid, mass spectrometry based method
○ Based on identification of ‘protein fingerprint’
○ Builds up profile for different organisms then compare to database
Works best with blood or urine for direct sample analysis -> because they are less complex than faeces for example
Otherwise culture is required
Cells in sample are lysed then overlaid with a matrix which helps ionize proteins
- Laser is fired at cells, which ionizes proteins
Proteins are detected in mass spectrometer
○ Small proteins ‘fly’ faster than larger proteins, will hit the plate quicker AKA time of flight
Immunodiagnostic
- Serology (antibodies) - looking for the antibody ‘footprint’ left by infection e.g. syphilis, ELISA for Vi (typhoid)
- Measure cytokine released from blood T cells when stimulated with peptides, e.g. TB, where TB peptides incubated with blood – measure interferon (QuantiFERON)
automated culture system
BACTEC - Inoculate bottles with predefined media in them, machine checks bottles for growth of bacteria
VITEK 2 - card with well that are filled with media and antibiotics
Full lab automation - WASP system
fully automates plating, biochemical tests
