Three strains of pathogenic bacteria, namely EcGS020802 , EcGS021001 and EcGS020801 were isolated from diseased orange spotted grouper ( Epinephelus coioides) . By physiological and biochemical analyzing , they belonged to Vibrio. In order to further classify them , 16S rRNA and heat shock protein ( HSP60) genes partial sequences were determined. Phylogenetic analysis based on 16S rRNA gene sequences indicated that three strains of pathogeny showed high levels of similarity to Vibrio harveyi , V. alginolyticus and V. parahaemolyticus. The identities among them , above 98. 6 % , were very high. Sequences analysis of HSP60 genes showed that they were very similar to V. harveyi (AF230934) , V. alginolyticus (AF230931) and V. parahaemolyticus (AF230951) , and their nucleotide identities were 95. 7 % , 99. 8 % and 99. 8 % , respectively. Moreover , the similarities with other Vibrio species were all less than 90. 6 %. The nucleotide similarities of HSP60 genes among three isolates were less than 91. 0 %. Phylogenetic tree of Vibrio based on HSP60 genes showed that strains EcGS020802 ,EcGS021001 and EcGS020801 clustered together with V. harveyi , V. alginolyticus and V. parahaemolyticus , respectively. Based on the results obtained, the strains EcGS020802 , EcGS021001 and EcGS020801 were identified as V. harveyi , V. alginolyticus and V. parahaemolyticus , respectively. Results showed that HSP60 gene was fitter to classify marine Vibrios than 16S rRNA gene.