Vibrio Chromogenic Agar

The Vibrio genus consists of microorganisms whose natural habitat is marine and fluvial ecosystems. They are frequently isolated from marine water, especially in warmer months and when the water temperature is higher than 17 °C. Vibrio species are mainly responsible for causing cholera and food poisoning in humans.

The medium contains yeast extract and peptones which are the source of nitrogen, vitamins (particularly the B-group essential for bacterial growth), minerals and amino acids. Special bile inhibits Gram-positive organisms. Sucrose, glucose and lactose are the fermentable carbohydrates, which provide carbon and energy. Sodium chloride supplies essential electrolytes for transport and osmotic balance. Sodium citrate, sodium thiosulfate and sodium cholate are the selective agents, inhibiting the Gram-positive bacteria. Chromogenic substrate is added to detect Vibrio species by means of a color change in the colonies.

This medium is designed for the development and differentiation of 3 types of Vibrio depending on the enzyme activity of each strain. Beta-glucosidase activity will appear as blue-green colonies, as in the case of V. parahaemolyticus. The activity of beta-galactosidase enzyme will show red or pink colonies in the case of V. cholerae. And finally, the yellowish-white colonies will be V. alginolyticus, which has beta-galactosidase, but it is not expressed due to the high concentration of sugars. The alkaline pH of the medium enhances the recovery of V. cholerae.

ISO 21872: Microbiology of the food chain - Horizontal method for the detection of potentially enteropathogenic Vibrio spp. - Detection of Vibrio parahaemolyticus, Vibrio cholerae and Vibrio vulnificus (ISO 21872-1:2017), recommends an alternative selective medium to TCBS for the detection of enteropathogenic Vibrio species.

Preparation

Suspend 90 grams of the medium in one liter of distilled water. Mix well and dissolve by heating with frequent agitation. Boil for one minute until complete dissolution. Avoid overheating. DO NOT AUTOCLAVE. Dispense into appropriate containers.

According to ISO 21872

Detection of potentially enteropathogenic Vibrio parahaemolyticus, Vibrio cholerae and Vibrio vulnificus according to ISO 21872:

1. Take test portions (25 g or 25 mL) and homogenize in 225 mL of enrichment medium ASPW. In the case of large quantities of test portion, the ASPW should be warmed to 37 ± 1 °C / 41.5 ± 1 °C before inoculation.
2. Incubate the initial suspension at 41.5 ± 1 °C / 37 ± 1 °C for 6 ± 1 hours.
3. Transfer 1 mL from the surface into a tube with 10 mL of ASPW.
4. Incubate the ASPW at 41.5 ± 1 °C / 37 ± 1 °C for 18 ± 1 hours.
5. From the culture obtained in the ASPW, inoculate 1 µL in TCBS Agar. Incubate a second selective isolation medium (Vibrio Chromogenic Agar).
6. Incubate at 37 ± 1 °C for 24 ± 3 hours.
7. Confirmation.

• Isabel Mª García Bermejo. Diagnóstico de las infecciones humanas causadas por especies halófilas del género Vibrio. Servicio de Microbiología, Hospital de Getafe.
• Koneman. Diagnostico Microbiológico/Microbiological diagnosis: Texto Y Atlas En Color. Stephen Allen Ed. Médica Panamericana, 30/06/2008.
• Romero Cabello, Raúl. Rodríguez, E; Gamboa, M; Hernández, F; García, J. 2005. Microbiología y Parasitología Humana. Bases etiológicas de las enfermedades infecciosas y parasitarias. 3ª Edición. Médica Panamericana.
• H.Y. Kudo et al. Improved Method for Detection of Vibrio parahaemolyticus in Seafood. ASM. Vol 67, No. 12, pg 5819-5823 (2001).
• Technical Specification ISO/TS 21872-1:2007(E) Microbiology of food and animal feeding stuffs - Horizontal method for the detection of potentially enteropathogenic Vibrio spp. Part 1: Detection of Vibrio parahaemolyticus and Vibrio cholerae.
• Technical Specification ISO/TS 21872-2:2007(E) Microbiology of food and animal feeding stuffs - Horizontal method for the detection of potentially enteropathogenic Vibrio spp. Part 2: Detection of species other than Vibrio parahaemolyticus and Vibrio cholerae.