Escherichia coli (E. coli) is one of the most recognized bacteria in microbiology, existing commonly in the environment and in the intestines of humans and animals. While many strains are harmless components of the normal gut flora, others can cause severe infections, necessitating rapid and accurate laboratory identification. Differentiating E. coli from thousands of other bacterial species often begins with simple biochemical tests. These tests exploit metabolic differences between organisms, providing a quick screening method to narrow down the possible identity of an unknown microbe.
Understanding the Oxidase Test
The oxidase test is a fundamental biochemical assay used to classify bacteria based on their electron transport chain components. This test specifically looks for the presence of the enzyme cytochrome c oxidase. This enzyme is a protein complex that serves as the final component in the aerobic respiration pathway of many organisms, catalyzing the transfer of electrons to oxygen.
To perform the test, a bacterial sample is exposed to a chromogenic reducing agent, such as tetramethyl-p-phenylenediamine dihydrochloride. If cytochrome c oxidase is present, it will oxidize the reagent, causing an immediate and visible color change. A positive result is indicated by the development of a deep purple or blue color within seconds, while a lack of color change signifies a negative result.
The Oxidase Result for E. coli
The definitive answer is that E. coli is consistently oxidase negative. When a sample is subjected to the test reagent, no blue or purple color develops. The absence of a color change indicates that the organism does not possess the specific cytochrome c oxidase enzyme the test is designed to detect.
This negative result is tied to E. coli’s metabolic flexibility as a facultative anaerobe. Unlike strict aerobes, E. coli employs a branched respiratory chain. It utilizes different terminal oxidases that are not reactive with the standard test reagents, allowing it to efficiently use oxygen or switch to anaerobic metabolism when oxygen is scarce.
Using the Oxidase Result for Preliminary Identification
The negative oxidase result for E. coli provides an immediate diagnostic shortcut in the microbiology laboratory. It acts as a rapid screening tool, placing the unknown bacterium into the family Enterobacteriaceae, often referred to as the “enterics.” A negative result immediately suggests the organism belongs to this large and clinically significant group of Gram-negative rods.
This single test is instrumental because it excludes a vast number of other oxidase-positive Gram-negative organisms. For instance, a positive result would immediately point toward genera like Pseudomonas, Neisseria, Vibrio, or Aeromonas, ruling out E. coli and the Enterobacteriaceae family. The negative result focuses the identification process onto a smaller group that includes Salmonella, Shigella, and Klebsiella.
Confirmatory Steps for Precise Identification
While the oxidase test narrows the field to the Enterobacteriaceae, further biochemical analysis is necessary to confirm the specific identity of E. coli. This is because the family contains many members that are also oxidase negative. Precise identification requires a panel of tests that look for specific metabolic traits unique to E. coli.
Key Confirmatory Tests
A characteristic trait used for confirmation is the Indole test, where E. coli yields a positive result. It is also positive for the fermentation of the sugar lactose. Furthermore, E. coli is characterized by a negative result on the Citrate utilization test and a negative Urease test. Combining the oxidase negative result with this specific set of reactions provides a reliable biochemical fingerprint for confirming E. coli.