Characterization of a novel live attenuated Edwardsiella piscicida vaccine based on the overexpressed type III secretion system and systematic deletion of the associated effectors

Edwardsiella piscicida causes edwardsiellosis in a variety of fish species and leads to tremendous economic losses in the global aquaculture industries. Thus, effective and safe prevention and control of this bacterium are urgently needed to combat the related infections. Live attenuated vaccines (LAVs) effectively prevent infectious diseases. However, most of the existing E. piscicida LAVs are based on the deletion of genes encoding the translocon components of the type III secretion system (T3SS), the core virulence system, which is the most prominent protective bacterial antigen with the strongest immunogenicity. In this study, we systematically deleted all of the 9 established T3SS effectors in E. piscicida (aka 9 Delta) and the rpoS gene encoding the alternative sigma factor, the esrB repressor (10 Delta), then we overexpressed esrB and T3SS in E. piscicida to obtain the recombinant strain 10 Delta/esrB(OE). The modified strains 10 Delta and 10 Delta/esrB(OE) exhibited severe attenuation and in vivo colonization defects. Additionally, vaccination by intraperitoneal injection with 10 Delta and 10 Delta/esrB(OE) could significantly upregulate the expression of the antigen recognition related gene (TLR5) and the adaptive immune response-related gene (MHC II) in the spleen/kidney of turbot fish, and it also enhanced the hosts' serum bactericidal capacity. Finally, vaccination with 10 Delta/esrB(OE) led to increased immune protection against the challenge of wild type E. piscicida EIB202 in turbot fish. Collectively, these findings demonstrated that 10 Delta/esrB(OE) was a novel LAV strain and therefore a potential novel strategy for the construction of LAVs against bacterial pathogens.