Evaluation of two direct plating methods using nonradioactive probes for enumeration, of Vibrio parahaemolyticus in oysters

Oysters (Crassostrea virginica) were collected monthly from May 1998 to Apr il 1999 from Mobile Bay, Ala,, and analyzed to determine Vibrio parahaemoly ticus densities at zero time and after 5, 10, and 24 h of postharvest stora ge at 26 degreesC. After 24 h of storage at 26 degreesC, oysters were trans ferred to a refrigerator at 3 degreesC and then analyzed 14 to 17 days late r. The V. parahaemolyticus numbers were determined by the most-probable-num ber procedure using alkaline phosphatase-labeled DNA probe VPAP, which targ ets the species-specific thermolabile hemolysin gene (tlh), to identify sus pect isolates (MPN-VPAP procedure), Two direct plating methods, one using a VPAP probe (Direct-VPAP) and one using a digoxigenin-labeled probe (Direct -VPDig) to identify suspect colonies, were compared to "the MPN-VPAP proced ure. The results of the Direct-VPAP and Direct-VPDig techniques were highly correlated (r = 0.91), as were the results of the Direct-VPAP and MPN-VPAP procedures (r = 0.91), The correlation between the Direct-VPDig and MPN-VP AP results was 0.85, The two direct plating methods in which nonradioactive DNA probes were used were equivalent to the MPN-VPAP procedure for identif ication of total V. parahaemolyticus, and they were more rapid and less lab or-intensive.