Vaccination is the most effective means of preventing infectious diseases; however, few vaccines are
effective against Streptococcus iniae (S. iniae) in grouper. This work presents an efficacious and safe
vaccine against S. iniae infections in the grouper Epinephelus coioides. The vaccine candidate was the
S. iniae GSI-310 strain. The vaccination was administered by intraperitoneal injection, and consisted
of formalin-inactivated antigens combined with an AS-F or ISA763A adjuvant. Peripheral blood samples
were collected for RT-qPCR and phagocytosis and agglutination assays. Our results indicated that
immunoglobulin M (igm) was maximally expressed in the two vaccinated groups at 3 months postsecondary
vaccination (PSV). A significant upregulation of mRNA expression for interleukin-1 (il-1ˇ)
and tumor necrosis factor- (tnf-˛) was also observed in fish treated with antigens combined with
ISA763A, which peaked at 3 months PSV. In fish treated with antigens combined with AS-F, il-1ˇ and tnf-˛
expression peaked at 14 days post-primary vaccination (PPV). Phagocytic activity and index increased
significantly in the two vaccinated groups. Furthermore, fish in the two vaccinated groups exhibited
significantly elevated agglutination titers compared to fish in the control group, in which almost no agglutination
reaction was detected. In the efficacy test, the vaccinated and control groupers were treated with
S. iniae at 1, 3, and 6 months PSV. The relative percentage survival (RPS) values of antigens with AS-F and
antigens with ISA763A were both 100% at 1 and 3 months PSV; at 6 months PSV, the RPS values for these
groups were 100% and 97.7%, respectively. Furthermore, the level of protection observed in the field
trial closely resembled that achieved on a laboratory scale. Therefore, the proposed vaccine mixed with
AS-F or ISA763A improved immune responses and provided safe and long-lasting protection in farmed
groupers.