Vibrio Millichrome^™ plus Agar

Vibrio Millichrome^™ plus Agar is a medium used to isolate and detect Vibrio parahaemolyticus, Vibrio vulnificus, and Vibrio cholerae. Peptones and yeast extract provide nitrogenous nutrients for growth and other essential growth factors. Salts provide osmotic balance, stimulate growth, and offer essential ions. High salt concentration, along with the alkaline pH, contribute to the selectivity of Vibrio species. The chromogenic mix contains chromogenic substrates for color differentiation based on the ability tocleave the substrate by characteristic enzymes. Agar is added as the solidifying agent. The medium presents clear color differentiation, with V. parahaemolyticus growing as mauve colonies, V. cholerae and V. vulnificus giving blue colonies, and V. alginolyticus remaining colorless. The recovery rate of Vibrio Millichrome^™ plus Agar is higher than with TCBS agar, and fewer false negatives are obtained. The final identification of the Vibrio species must be confirmed by biochemical tests or mass spectrophotometry.

Vibrio Millichrome^™ plus Agar is a medium used for the isolation and detection of Vibrio parahaemolyticus, Vibrio vulnificus, and Vibrio cholerae in the environmental and food samples.

Step 1 (Preparation of 1L media)

• Disperse slowly 74.7 g of powder base in 1 L of purified water.
• Stir until agar is well thickened.
• Heat and bring to boil (100 °C) while swirling or stirring regularly.

DO NOT HEAT TO MORE THAN 100 °C. DO NOT AUTOCLAVE AT 121 °C.
Warning 1: If using an autoclave, do so without pressure.
Advice 1: For the 100 °C heating step, mixture may also be brought to a boil in a microwave oven: after initial boiling, remove from oven, stir gently, then return to oven for short repeated bursts of heating until complete fusion of the agar grains has taken place (large bubbles replacing foam).
Step 2 (Pouring)

• Cool in a water bath at 45-50 °C, swirling or stirring gently.

Advice 2: In case of samples with a high presence of Aeromonas, 50 mg of cefsulodin can be added to the mix once cooled down at 45-50 °C (50 mg/L).

• Pour medium into sterile Petri dishes.
• Let it dry and gel

V. parahaemolyticus grows as mauve colonies while V. cholerae and V. vulnificus give blue colonies. V. alginolyticus remains colorless which helps to avoid any interference with the detection of other species. The recovery rate of Vibrio Millichrome™ plus Agar is higher than with TCBS agar, even if using an enrichment broth. Also, fewer false negative results are obtained than with TCBS agar.

MilliChrome is a trademark of Merck KGaA, Darmstadt, Germany