Veterinary Clinical Biochemistry and Laboratory Diagnosis-II

OXIDASE TEST

Principle

• Cytochrome oxidase is an enzyme found in some bacteria that transfers electrons to oxygen, the final electron acceptor in some electron transport chains.
• Presence of cytochrome oxidase can be detected through the use of dyes such as P-phenylenediamine dihydrocholide, that substitute for oxgen as artificial electron acceptors.
• In reduced state the dye is colorless; however in the presencec of cytochrome oxidase and atmospheric oxygen, the dye is oxidised ,forming indophenol blue. 
• This test is most helpful in screening colonies suspected of being one of the Enterobacteriaceae (all negatives) and other genera such as Aeromonas,Pseudomonas,Neisseria ,Campylobacter and Pasteurella(positive)

Methodology

• Inoculate a nutrient agar plate with the test organism.
• Incubate at 37⁰C for 24 hours.
• Transfer the colony with a platinum loop or glass rod to a wet filter paper containing oxidase reagent (tetra methyl para phenylene diamine hydrochloride) or commercial oxidase disc.

Result

• Bacterial colonies having cytochome oxidase activity develop a deep violet colour with 10 seconds indicates a positive reaction. Stainless steel or nichrome inoculating loop or wires should not be used for this test because surface oxidation products formed when flame sterilizing may result in false positive reaction.

 

Oxidase test

Control

• Positive    :  Pseudomonas aeruginosa
• Negative  :  Enterobacteriaceae