Evaluation of a double layer agar plate for direct enumeration of Vibrio parahaemolyticus

A double layer agar plate (DLAP) was developed according to the thin agar layer (TAL) method as a 1-step procedure for direct enumeration of injured Vibrio parahaemolyticus cells based on the formation of unique purple colonies by V. parahaemolyticus. The DLAP was prepared by overlaying an equal volume (10 mL) of a nonselective medium (tryptic soy agar supplemented with 1.5% NaCl) onto a selective medium (Bio-Chrome Vibrio medium). The DLAP was capable of detecting V. parahaemolyticus in mixed cultures containing non-Vibrio bacteria. Production of purple colonies by V. parahaemolyticus on DLAP was not affected by the growth of other bacteria, even when V. parahaemolyticus was only a small fraction (5%) of the entire bacterial population. Direct plating on DLAP was found as effective as the most probable number (MPN) method for recovering heat-and cold-injured V. parahaemolyticus cells, which could not be detected by direct plating on Bio-Chrome Vibrio medium or thiosulfate-citrate-bile salts-sucrose agar. The DLAP offers an alternative to the MPN method for detecting injured V. parahaemolyticus cells and can be used as a simple 1-step procedure for quick screening of V. parahaemolyticus in foods.

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