Morphological and Selective Biochemical characterization of some specimen

28. CATALASE TEST
• Principle: Some organisms contain enzyme catalase, which
liberates oxygen from hydrogen peroxide. Test is performed on
bacterial growth. It differentiates Staphylococcus from
Streptococcus.
• Procedure: A small inoculum of test organism is added to a slide or
tube having 3% solution of hydrogen peroxide with a sterile wooden
or glass rod.
• Interpretation: Gas bubbles indicate positive result.
Staphylococcus spp is catalase positive and Streptococcus spp is
catalase negative.
• Control: Use positive and negative controls along with the test
organisms.

30. COAGULASE TEST
Principle: Coagulase causes plasma to clot by converting
fibrinogen to fibrin. It is done to differentiate S.aureus from
other staphylococci.
Two types of coagulase are produced by most strains of
Staphylococcus aureus.
1. Free coagulase: It converts fibrinogen to fibrin by
activating a coagulase reacting factor present in plasma,
detected by tube method.
2. Bound coagulase (clumping factor): It converts
fibrinogen directly to fibrin and is detected by slide test.

31. 1. Procedure (Slide Test): Emulsify a colony in a drop
of isotonic saline on each end of slide to make
suspension. Add a drop of plasma to one and mix
gently. Look for clumping within 5-10 seconds.
Other drop serves as negative control to rule out
autoagglutination.
• Interpretation: Report "Coagulase positive or
negative". In case of negative slide test, tube test must
be done before declaring the organism as coagulase
negative.

32. OXIDASE TEST
• Principle: The test detects oxidase-producing bacteria and helps in
identification of Vibrio, Neisseria and Pseudomonas species.
• Reagents: Freshly prepared 10 g/L solution of tetramethyl-p-
phenylenediaminedihydrochloride.
• Procedure: Place a piece of filter paper in a clear petri dish and add
2-3 drops of oxidase reagent. Using a sterile glass rod or wooden
stick, remove a colony of the test organism from culture plate and
streak onto the filter paper. Look for the development of blue purple
colour within 5-10 seconds.
• Interpretation: Report as oxidase positive if blue purple colour is
produced. Test should be controlled by using Pseudomonas
aeruginosa as positive control and E.coli as negative control.

34. INDOLE TEST
• Principle: This test demonstrates the ability of organisms to
breakdown the amino acid Tryptophan to indole, detected by Ehrlich
or Kovac reagent and helps differentiate enterobacteriaceoe
especially Escherichia coli which is positive and Klebsiella
pneumoniae which is indole negative.
• Reagents: Kovac reagent
• Procedure: Add 0.5 ml Kovac reagent to 3 ml 1-day-old peptone
water culture containing tryptophan. A red colour indicates positive
test.
• Control: Escherichia coli is positive and Enterobacter sp is
negative.

36. Triple Sugar Iron and Urea Agar