CATALASE TEST

CATALASE TEST

AIM

            To test the Catalase enzyme producing efficiency of Bacteria.

PRINCIPLE   

     Catalase test is used to differentiate the bacteria that produce an enzyme Catalase, such as Staphylococcus sp. from Non-catalase producing bacteria such as Streptococcus sp. It is also used for differentiating Bacillus sp. from Clostridium sp. In 1893, a publication by Gottstein brought attention to bacterial catalase, making it one of the first bacterial enzymes.

           Aerobic and facultative anaerobic organisms produce two toxins during normal metabolism. One is Hydrogen peroxide (H₂O₂) and another one is Superoxide radical (O₂⁻). These bacteria have two enzymes that detoxify the products of normal metabolism. One of these enzymes, Catalase, is capable of converting Hydrogen peroxide to Water and Oxygen. Normally, 5 % H₂O₂ is used for the detection of Catalase in bacteria. The presence of the Catalase enzyme in a bacterial isolate is evident when a small inoculum is introduced into hydrogen peroxide, and the rapid elaboration of oxygen bubbles occurs. The lack of Catalase is evident by a lack of or weak bubble production. The culture should not be more than 24 hours old.

          Catalase positive bacteria include Strict aerobes as well as Facultative anaerobes, although they all have the ability to respire using oxygen as a terminal electron acceptor. Catalase negative bacteria may be Anaerobes, or they may be Facultative anaerobes that only ferment and do not respire using oxygen as a terminal electron acceptor (Streptococcus sp.).

 MATERIALS REQUIRED

1. Test bacteria
2. 3 % Hydrogen peroxide (H₂O₂)
3. Glass slide or Test tube
4. Sterile Wooden stick or Glass rod

PROCEDURE

I) Tube Catalase Test

a)     Pour 1 - 2 ml of 3 % Hydrogen peroxide (H₂O₂) solution into a test tube.

b)  Using a sterile wooden stick or a glass rod, take several colonies of the 18 to 24 hours test organism and immerse in the 5 % Hydrogen peroxide (H₂O₂) solution.

c)     Observe for immediate bubbling.

II) Slide Catalase Test

a)    Use a sterile wooden stick or glass rod to transfer a small amount of colony growth in the surface of a clean, dry glass slide.

b)     Place a drop of 5 % Hydrogen peroxide (H₂O₂) in the glass slide.

c)     Observe for the evolution of oxygen bubbles.

OBSERVATION AND RESULTS

·       Catalase Positive Test – Immediate bubble formation

·       Catalase Negative Test – No bubble formation

 

Figure – 1: Catalase test (Slide test and Tube test) (Source: microbiologyinfo.com)