Ex. 5-4 Catalase test MCB 1000 Lab. Prof. Meléndez

Catalase test The catalase test is used to detect the presence of the enzyme catalase in bacteria. Catalase serves to neutralize the bactericidal effects of hydrogen peroxide. Its concentration in bacteria has been correlated with pathogenicity. This enzymatic test is essential in the scheme of identification for gram-positive organisms and certain gram-negative organisms. It is a primary test used in the differentiation of staphylococci and streptococci.

Catalase test Theory The catalase enzyme serves to neutralize the bactericidal effects of hydrogen peroxide . Catalase expedites the breakdown of hydrogen peroxide (H2O2- (toxic) into water (H2O) and oxygen(O2) 2H2O2 + Catalase → 2H2O + O2(gas bubbles) This reaction is evident by the rapid formation of bubbles

Catalase test The catalase test facilitates the detection of the enzyme catalase in bacteria. It is essential for differentiating catalase-positive Micrococcaceae from catalase-negative Streptococcaceae. The catalase test is also valuable in differentiating aerobic and obligate anaerobic bacteria, as anaerobes are generally known to lack the enzyme . In this context, the catalase test is valuable in differentiating aerotolerant strains of Clostridium, which are catalase negative, from Bacillus, which are catalase positive

Slide catalase test results Slide catalase test results. Hydrogen peroxide was added directly to the culture on a microscope slide. A positive reaction produced by Staphylococcus aureus is indicated by bubbling; a negative reaction produced by Streptococcus pyogenes is indicated by lack of bubbling. (Karen Reiner, Andrews University, Berrien Springs, MI)

Practice questions Read lab. manual pages 243-246 and fill out following chart: Determine if situation on first column will cause a false positive or a false negative. Explain why? Description of situation Effect on test Why? 1. Adding hydrogen peroxide to slide first, then adding cells and mixing cells with hydrogen peroxide using loop. False positive Metal wire of loop can react with hydrogen peroxide causing bubbles due to oxidation of metal, not the enzyme activity. 2. Hydrogen peroxide is expired (old) 3. Cells are heat fixed into slide 4. Cells are very old 5. Contamination of culture with aerobic microorganism 6. Dirty slide was used

References http://www.microbelibrary.org/index.php/l ibrary/laboratory-test/3226-catalase-test- protocol Leboffe, Michael J. , and Burton E. Pierce. Microbiology Laboratory, Theory and application: (brief edition). Colorado: Morton Publishing Company, 2008.