Urease test: Principle, Procedure, interpretation and urease positive organsims

Medium used: Any urea medium, agar or broth

Indicator used: Phenol red
Color change:  

Original: orange yellow color
Final color (in positive test): Bright pink 

Test principle

Many organisms especially those that infect the urinary tract, have a urease enzyme which is able to split urea in the presence of water to release ammonia and carbon dioxide. The ammonia combines with carbon dioxide and water to form ammonium carbonate which turns the medium alkaline, turning the indicator phenol red from its original orange yellow color to bright pink.

Oxidase test: Principle Procedure and oxidase positive organisms

Test requirements for Oxidase test: Moisten filter paper with the substrate (1% tetramethyl-p-phenylenediamine dihydrochloride), or commercially prepared paper disk, wooden wire or platinum wire

Expected results of Oxidase test
Positive: Development of dark purple color (indophenols) within 10 seconds
Negative: Absence of color

Diagnostic Tests: Biochemical Tests: Coagulase Test

Coagulase test is used to differentiate Staphylococcus aureus (positive) from Coagulase Negative Staphylococcus (CONS). Coagulase is an enzyme produced by S. aureus that converts (soluble) fibrinogen in plasma to (insoluble) fibrin. Staphylococcus aureus produces two forms of coagulase, bound and free.

1. Slide coagulase test is done to detect bound coagulase or clumping factor.
2. Tube coagulase test is done to detect free coagulase.

Slide Coagulase Test Procedure

1. Take a clean grease free and dry glass slide
2. Place a drop of plasma
3. Place a drop of distilled water or saline next to the drop of plasma as a negative control
4. Emulsify a portion of the isolated colony being tested in each drop, and make a smooth  milky suspension (not clumpy).
5. Rock the slide gently for 5 to 10 seconds and look for clumping (coagulation).

Diagnostic Test for Bacterial Identification: Indole Test

The indole test is used to determine the ability of an organism to spilt amino acid tryptophan to form the compound indole. Tryptophan is hydrolysed by tryptophanase to produce three possible end products – one of which is indole.

Indole production is detected by Kovac's or Ehrlich's  reagent which contains 4 (p)-dimethylamino benzaldehyde, this reacts with indole to produce a red coloured compound.
Indole test is a commonly used biochemical test (eg in IMVIC test, SIM test etc). Indole test helps to differentiate Enterobacteriaceae and other genera.

Two methods are in used;

1. a spot indole test, which detects rapid indole producing organisms and
2. a conventional tube method requiring  overnight incubation, which identifies weak indole producing organisms.