Coliform bacteria: Biochemical tests – presumptive identification and confirmation

What are coliform bacteria

Coliform bacteria are a group of bacteria that are commonly used as indicators of fecal contamination in water, food, and other environmental samples. They are found in the intestines of humans and animals, and their presence in water or food can indicate the potential presence of harmful pathogens.

Coliform bacteria are gram-negative, rod-shaped bacteria that belong to the family Enterobacteriaceae. The most well-known coliform species is Escherichia coli (E. coli), which is a normal inhabitant of the intestines of warm-blooded animals, including humans.

The presence of coliform bacteria in water or food samples suggests that there may have been fecal contamination, indicating a possible risk of disease-causing organisms (pathogens) being present. While coliform bacteria themselves may not necessarily cause illness, their presence serves as an indicator of potential contamination and the need for further testing and monitoring.

These bacteria are commonly tested in various industries, including food and beverage production, water treatment, and environmental health. Testing for coliform bacteria helps ensure the safety and quality of water and food supplies, as well as the overall sanitary conditions in various settings.

It’s important to note that while these bacteria are useful as indicators, additional testing is required to identify specific pathogens if their presence is suspected.

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Procedure for presumptive identification and confirmation of coliform bacteria

Based on the provided excerpt, here are the steps involved in confirming if the bacteria growing on the selective media (violet red bile agar) are truly coliforms:

  1. Growing bacteria on selective media (VRBA): The first step is to inoculate the bacteria on a selective media such as violet red bile agar (VRBA). This media is specifically designed for the identification of coliform bacteria.
  2. Select representative colonies: After incubation, 10 representative colonies are selected from the VRBA plate. These colonies are chosen to ensure a representative sample for further testing.
  3. Transferring colonies to brilliant green lactose broth: The selected colonies are transferred to sterile tubes containing brilliant green lactose broth. This medium allows the growth and detection of coliform bacteria.
  4. Incubation at 35°C for 24 hours: The tubes containing the transferred colonies are incubated at a temperature of 35°C for a period of 24 hours. This provides optimal conditions for the growth and metabolic activity of coliform bacteria.
  5. Observing for gas production and pellicle formation: After incubation, the tubes are examined for the presence of gas production. Gas-positive tubes are identified, indicating the production of gas as a metabolic byproduct of coliform bacteria. Additionally, the presence of a pellicle (a floating layer) in these tubes is observed, which can be indicative of coliform growth.
  6. Gram staining: Tubes that show gas production and pellicle formation are selected for further testing. Gram staining is performed on these tubes to eliminate the possibility of Gram-positive, lactose-fermenting bacilli. Gram staining allows differentiation between Gram-positive and Gram-negative bacteria based on the staining properties of their cell walls.

By following these steps, the confirmation process ensures that the bacteria growing on the VRBA media are truly coliforms, specifically eliminating the presence of Gram-positive, lactose-fermenting bacilli. The steps are simplified in the flow diagram below:

Identification and confirmation of coliform bacteria

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