Catalase test is used to identify microorganisms that produce the enzyme called catalase. It is used to differentiate catalase-producing organisms (e.g. Staphylococcus species) from non-catalase producing bacteria (e.g. Streptococcus species). Catalase enzyme breakdown hydrogen peroxide (H2O2) to water and oxygen (as shown in Equation 1); and this leads to the release or production of bubbles – which indicates a positive test result (Figure 1). When a culture plate containing cultures of S. aureus is flooded with H2O2, bubbles are released, and this shows that the test organism in the culture plate is S. aureus (Figure 2). The presence of catalase enzyme in the test isolate is detected using hydrogen peroxide. If the bacteria possess catalase (i.e. are catalase-positive), when a small amount of bacterial isolate is added to hydrogen peroxide, bubbles of oxygen are observed.
2H2O2 —————— 2H2O + O2
Equation 1. Chemical reaction between hydrogen peroxide and catalase enzyme
Most aerobic bacteria produce catalase enzyme in varying amounts except the lactic acid bacteria which do not produce the enzyme in detectable amounts. Obligate anaerobes are usually catalase negative. Hydrogen peroxide is a highly toxic product of certain cellular processes such as the reduction of flavoproteins, thus catalase test is used to determine whether or not a particular microorganism produces catalase (the enzyme that catalyzes the breakdown of H2O2). This test can be performed by two methods: the slide method and the test tube method, but only the slide method shall be expanded here.
PROCEDURE FOR SLIDE METHOD OF CATALASE TEST
- Perform this test with pure culture from culture media plate (preferably blood agar).
- Place a loopful or speck of the test organism on a clean glass slide.
- Emulsify the culture with a loopful of freshly prepared H2O2.
- Lookout for the release or presence of bubbles which indicates a catalase positive result. The bubbles results from the production of oxygen gas released from theaction of catalase enzyme (produced by the test organism) on the H2O2 (Figure 2).
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