, a gram-negative bacterium has been described to be among the most common and economically important aquatic pathogen of fish and shellfish. Vaccine immunization is an effective approach to prevent infection. And attenuated vaccine could stimulate systemic immune response in host administrated. Now there has been little attenuated vaccine of . The type III secretion system (T3SS) is used by bacterial pathogens to inject effector proteins into the cytoplasm of their host cells. And it is an important pathogenic factor of . The T3SS consists of a structure called needle complex, which is made of a multi-ring base that spans the bacterial and a needle-like extension that protrudes several nanometers from the bacterial surface. is located at the “needle” site of T3SS. Its role is to escort the molecular chaperone and effector proteins into host cells, and further induced host cell death. In this paper, combining with chloramphenicol (Cm) and sucrose screening, an in-frame deletion mutant of using overlap PCR and homologous recombination technology was constructed. And the LD₅₀ changes of ZJ03ΔvscO mutant strains compared with the strain ZJ03 were examed. Grouper (), body length (17±3) cm, body weight (50±5) g were injected with ZJ03ΔvscO and ZJ03, respectively. The ΔvscOmutant showed about 150 times decrease in virulence in grouper. The ZJ03ΔvscOwas used to immunize grouper by way of injection and immersion. The result indicated that the specific antibody titers were markedly higher than that in the control group (<0.05).The relative percentage survival (RPS) of injection group was 84%, while the RPS of immersion group was 68%. The results showed that the ZJ03ΔvscO of has a high immunogenicity, and can be a live attenuated vaccine. In addition, . The purpose of this study is to provide technical support for fish diseases caused by