Three bacterial strains named RH1,RH2 and RH3 were isolated from the cultured sea cucumber (Apostichopus japonicus) with skin ulceration. The strain RH2 was proved to be a pathogen by soaking with suspension of bacteria, coelome injection and muscle injection, the LD50 does was 5.68×106CFU/per sea cucumber, and it was proved that soaking with suspension of bacteria and coelome injection could not infect the sea cucumber successfully. By methods of morphological observation, physiological and biochemical characteristic analysis, RH2 was identified as Vibrio alginolyticus. 16S rRNA gene and 60 KD heat shock protein (HSP60) gene sequence detection further proved that strain RH2 belonged to V. alginolyticus. Phylogenetic tree of Vibrio based on 16S rRNA gene and HSP60 gene both indicated that strain RH2 showed the highest level of similarity to V. alginolyticus, and it’s bootstrap was 50% and 99%, respectively. The optimum salinity and temperature for strain RH2 was 25~35 and 14~22℃, the optimum pH was 7.2~8.0. The results of sensitivity to 18 kinds of antibacterial agents showed that strain RH2 was sensitive to norfloxacin, sulfamethoxazolum and spectinomycine. Key words： Apostichopus japonicus；skin ulceration；Vibrio alginolyticus；16S rRNA gene；HSP60 gene；biological characteristic