08/08/2021
BIOCHEMICAL TESTS:
1. COAGULASE TEST:
A coagulase test is a biochemical test that is used to differentiate Staphylococcus aureus from other Staphylococci species like S. epidermidis and S. saprophyticus on the basis of the ability to produce the coagulase enzyme.
PRINCIPLE:
Coagulase is an enzymatic protein that is a thermostable thrombin-like substance, which converts fibrinogen into fibrin resulting in clotting or clumping. In S. aureus, two different forms of coagulase are found; free coagulase and bound coagulase.
REAGENTS AND SUPPLIES:
- Frozen plasma (preferably rabbit plasma) with EDTA. Human plasma is commonly not used for the test, as it is less sensitive and potentially infectious with human pathogenic viruses.
- 5% CaCl2 (optional)
- Loops or sterile sticks
- Glass or plastic tubes
- Glass slides
PROCEDURE:
Coagulase can be detected by two different methods; tube test and slide test.
1. SLIDE TEST:
- About 10 µl of deionized water or physiological saline is added to a slide.
- Several colonies from a fresh culture are collected with an inoculating loop and are emulsified into the water to obtain a smooth milk-colored suspension.
- A drop of a rabbit or human plasma is added to the slide, and the clumping is observed immediately, not to exceed 10 seconds.
2. TUBE TEST:
- The plasma is diluted with physiological saline. (Add 0.2 ml plasma in 1.8 ml saline)
- 5 ml of the diluted plasma is then added to a test tube. About 5 drops of the test organism culture are added to the test tube.
- The test tube is mixed and incubated at 37°C for an hour.
- The tube is finally observed for clot formation. If no clotting is observed, the tube should be examined at 30 minutes intervals of up to 6 hours.
RESULTS:
SLIDE TEST:
- A positive test is the demonstration of the agglutination of the bacterial cells after the plasma is added.
- negative test is demonstrated by the lack of agglutination.
TUBE TEST:
A positive test meets one of the following criteria:
1. Complete clot formation or any degree of clot formation before 24 hours.
2. No clot formation after the addition of 1 or 2 drops of 5% CaCl2 to a tube without a clot at 24 hours.
A negative test meets one of the following criteria:
1. A lack of clot formation at 24 h at 25°C.
2. No clot after 24 hours at 35°C, but after the addition of 1 or 2 drops of 5% CaCl2 to the tube, a clot forms.
Coagulase Positive organisms: Staphylococcus aureus and other animal host bacteria like S. pseudintermedius, S. intermedius, S. schleiferi, S. delphini, S. hyicus, S. lutrae, S. hyicus
Coagulase Negative organisms: Staphylococcus epidermidis, S. saprophyticus, S. warneri, S. hominis, S. caprae, etc.
