Bivalent vaccination with Edwardsiella tarda and Aeromonas salmonicida protected freshwater fish after challenge and consists a strategic approach against pathogens

Abstract

The delivery of antigens into host cells via the type III secretion system (T3SS) represents a highly effective strategy employed by bacteria to elicit host immune responses and protect against bacterial infections. In this study, we engineered recombinant strains by heterologously biosynthesizing the T3SS from Photorhabdus luminescens TT01 in the avirulent Aeromonas salmonicida DBFF01 strain using tilapia and carp as model. The GAPDH from Edwardsiella tarda was N-terminally fused with the translocation signal of the type III effector LopT and co-expressed with its chaperone, SlcT, in T3SS-expressing A. salmonicida DBFF01, thereby constructing a bivalent vaccine targeting both E. tarda and A. salmonicida. The detection of GAPDH delivery in eukaryotic cells indicates that the heterologous biosynthetic T3SS in A. salmonicida DBFF01 possesses functional activity and can effectively transport GAPDH to carp epithelial cells (EPCs). In vivo trials revealed that tilapia immunized with the bivalent vaccine achieved 100 % relative percentage survival rate when challenged with lethal E. tarda EIB202. Furthermore, the recombinant strains effectively colonized fish tissues without inducing pathology. The vaccination significantly enhanced innate immune enzyme activities and elevated serum IgM levels as well as proinflammatory cytokines (IL-6, TNF-α, IFN-α), and antigen-presenting cell (APC) activation. These findings establish T3SS as a versatile and potent tool for vaccine delivery in aquaculture, combining high efficacy, safety, and robust immune activation to combat bacterial pathogens.