Describe catalase test and its reaction time.
- Detects catalase enzyme
- - converts H2O2 to H2O and O2
- - uncommon in anaerobes
Rapid detection (within minutes)
Describe the coagulase test and its reaction time.
Two main types: agglutination and tube
- Agglutination: rapid, done by a kit to detect bound coagulase
- - microparticles with bound fibrinogen and IgG
- - agglutinate in presence of coagulase
: detects free coagulase; overnight reaction
Define Free and Bound Coagulase
Free: reacts with coagulase-reactin factor to form staphylothrombin, which catalyzes the breakdown of fibrinogen to fibrin
Bound: attached to bacterial cell wall and enzymatically converts fibrinogen to fibrin
How is fermentation used to differentiate microbes?
Fermentation converts sugars in the medium to alcohol/acids, which lowers the pH.
Different mediums have different indicators that change color based on these pH changes, assisting in identification.
These reactions occur overnight.
Describe the oxidase test.
The microbes are dabbed directly on a wet piece of paper that will react in the presence of cytochrome oxidase.
A positive reaction will occur within 3 minutes, yielding a purplish-maroon spot.
Describe how the urease test works.
Microbes with urease enzyme break down amides in the medium to ammonia, lowering the pH.
A pH indicator in the medium changes color in response; test happens overnight.
What is thioglycollate, and how is it used?
Thioglycollate is added to a semisolid medium to remove all O2 from it.
A tube containing the medium is innoculated, and based on the pattern of growth, we can determine if the microbe is a faculative or obligate aerobe, oxygen tolerant, or an obligate anaerobe.
What staining is this?
Thick peptidoglycan cell wall retains crystal violet staining after alcohol wash
What staining is this?
- Thin peptidoglycan cell wall with outer membrane
- Alcohol wash removes outer membrane and crystal violet, counterstain with pink saffranin
What staining is this? (Look at small rods)
- Acid-Fast Staining
- Some bacteria have unusual cell walls (contain mycolic acid, giving waxy surface) and resist Gram staining
Used especially for microbes like Mycobacterium tuberculosis, leprae,
; also Nocardia
Stain using hot carbolfuschin (pink color), and counterstain with methylene blue (gets everything else)
What genera stain poorly?
Obligate Intracellulars (Chlamydia, Chlamydiophila, Rickettsia, Ehrlichia, Anaplasma, and Coxiella)
Spirochetes (Treponema and Borrelia)
Mycoplasma (do NOT have cell wall!)