The thermostable direct hemolysin (TDH) and TDH-related hemolysin (TRH) are the main virulence factors of Vibrio parahaemolyticus. We isolated V. parahaemolyticus from seawater, fish, and oysters obtained from the Pueblo Viejo Lagoon in Veracruz, determined the serogroups, phenotypically and genotypically characterized TDH and TRH, and investigated the presence of the toxR gene. A total of 46 V. parahaemolyticus strains were isolated, and all of them amplified the 368-bp toxR gene fragment. The trh gene was not identified in any of the strains; 4 of the 46 strains were Kanagawa phenomenon (KP) positive and amplified the 251-bp tdh gene fragment. The most frequent serogroup was serogroup O3. This is the first report of the presence of KP-positive tdh-positive environmental V. parahaemolyticus strains in Mexico.Vibrio parahaemolyticus is a halophilic gram-negative bacterium that is widely distributed in coastal waters worldwide and is associated with gastroenteritis, wound infections, and septicemia (5). V. parahaemolyticus infections are frequently reported in coastal areas, apparently because of the high consumption of sea products and direct contact with estuarine waters (19).Epidemiological studies have revealed an association between the Kanagawa phenomenon (KP) and gastroenteritis (23,25). KP is a type of beta-hemolysis induced by the thermostable direct hemolysin (TDH) in Wagatsuma agar. Most (90%) of the strains isolated from clinical cases show this type of hemolysis, while only 1 to 2% of the strains of environmental origin are KP positive (20).Several cases of gastroenteritis caused by hemolytic but KPnegative TDH-negative V. parahaemolyticus strains were reported in the 1980s (11), which led to identification of a new hemolysin known as TDH-related hemolysin (TRH) (12,13,30). TDH and TRH are encoded by the tdh and trh genes, respectively; these two genes both have 567-bp open reading frames, and they show 68.6% sequence similarity. These hemolysins are considered the main virulence factors of this microorganism (20).In the present study we determined the prevalence of V. parahaemolyticus in seawater, oyster, and fish samples collected from the Pueblo Viejo Lagoon in Veracruz, an important estuary on the coastline of the Gulf of Mexico. The strains isolated were serotyped and screened for hemolytic activity and for the presence of the toxR, tdh, and trh genes. MATERIALS AND METHODSA total of 266 seawater, oyster, and fish samples were collected from 12 different sites in the Pueblo Viejo Lagoon (Fig. 1) The fish and oysters were transported in individually labeled and sealed plastic bags to avoid contamination. Seawater samples were collected in labeled plastic jars. The samples were placed in sealed containers with dry ice and transported frozen to the laboratory for analysis. The time between sample collection and analysis was approximately 24 h.V. parahaemolyticus strain isolation and identification. V. parahaemolyticus was isolated and identified as described in the Bacteriological Analytical Manua...