Protease and virulence of the extracellular products produced by Vibrio carchariae after growth on various media

Protease and virulence of the extracellular products (ECP) of Vibrio carcha rine strain EmI82KL, a causative agent of gastroenteritis in Epinephelus co ioides, cultured on different media were studied. The bacteria grown on pep tone agar, nutrient agar or brain heart infusion agar produced higher prote ase activities than that grown on tryptic soy agar (TSA) in terms of protei n content. The addition of ethylenediamine di (o-hydroxyphenylacetic acid) or horse serum in TSA enhanced the ECP protease production while the additi on of grouper serum apparently reduced the enzyme activity indicating the p resence of protease inhibitor(s) in the fish serum. Furthermore, the use of grouper meat or peptone as a single nutrient source remarkably enhanced th e production of ECP protease. Adding proteinaceous materials from animal so urces (horse serum, grouper meat or peptone) on agar manifested higher ECP protease activity than that from plant source (TSA), indicating the intesti ne of carnivorous groupers might favour the existence, survival or infectio n of the bacterium. The protease was a metal-chelator-sensitive serine prot ease since it was inhibited by 3,4-dichloroisocoumarin and phenylmethanesul fonyl fluoride while about 80% of its activity inhibited by chelating agent s (ethylene-diaminetetraacetic acid and ethylene glycol-bis(beta-amino-ethy lether) N,N,N',N'-tetraacetic acid). The ECP obtained from each medium was not lethal to the groupers suggesting that the bacterium is low virulent. A s grouper is carnivorous, therefore, the role of the protease played in the fish intestine probably is competing for nutrients and/or associated with the cause of edema leading to gastroenteritis.