Protective efficacy of the optimized GluB-MTAP fusion subunit vaccine against Nocardia seriolae infection in largemouth bass (Micropterus salmoides).

Nocardia seriolae is the predominant causative agent of piscine nocardiosis, exhibiting broad piscine host specificity along with prolonged latency periods and high mortality rates, thereby posing a severe threat to global fish health. However, reliably effective control measures remain limited. Subunit vaccines, derived from virulence factors or immunodominant proteins of the pathogen, are widely acknowledged as one of the most secure and potent strategies for mitigating disease outbreaks. Here we focused on five T7SS gene cluster proteins (MTAP, HP05, HP23, HP24, and VST) and the dominant N. seriolae secretory protein (GluB) as subunit vaccine candidates through whole-genome sequencing and HPLC-MS analysis, identifying GluB and MTAP as superior antigens through efficacy-based screening assessing immunogenicity, serum antibody titers, and protective efficacy. To further enhance the protective efficacy, we engineered a novel fusion antigen composed of GluB and MTAP. In Micropterus salmoides, immunization with the GluB-MTAP fusion antigen elicited robust defensive responses at 28 days post-immunization, characterized by enhanced antioxidant capacity and amplified inflammatory and humoral immunity. The fusion vaccine conferred significant protection against N. seriolae challenge with a relative percentage survival (RPS21d) of 70 %, achieving 72.3 % absolute survival compared to 8 % in controls, which represented absolute survival improvements of 6.3 % over GluB-monovalent (66 %) and 18.3 % over MTAP-monovalent immunization (54 %). Collectively, the GluB-MTAP subunit vaccine demonstrates substantial efficacy against nocardiosis in M. salmoides, providing a mechanistic platform for investigating Nocardia-host interplay. This study further establishes a rational framework for anti-Nocardia seriolae vaccine development and advances sustainable aquaculture disease control strategies.