The genus Vibrio is characterized by a large number of species and some of them are human pathogens causing gastrointestinal and wound infections through the ingestion or manipulation of contaminated fishes and shellfish including Vibrio parahaemolyticus and Vibrio alginolyticus. In this study, we reported the phenotypic and molecular characterization of 9 V. parahaemolyticus and 27 V. alginolyticus strains isolated from outbreaks affecting cultured Gilthead sea bream (Sparus aurata L.) and Sea bass (Dicentrarchus labrax) along the Tunisian coast from 2008 to 2009. All isolates were tested for the presence of DNase, caseinase, protease, lipase, amylase, gelatinase, hemolytic activity and antibacterial resistance to different drugs. Randomly amplified polymorphic DNA was used to examine the genetic relatedness among the V. parahaemolyticus and V. alginolyticus strains.