A bivalent vaccine derived from attenuated Salmonella expressing O-antigen polysaccharide provides protection against avian pathogenic Escherichia coli O1 and O2 infection

Avian pathogenic Escherichia coli (APEC), a leading cause of avian airsacculitis and colibacillosis, is responsible for significant economic loss in the poultry industry. APEC serogroups O1, O2, and O78 are predominantly associated with disease. Lipopolysaccharide (LPS) O-antigen has been shown to be a potent antigen for inducing specific protective immune responses. Therefore, we sought to develop a multivalent polysaccharide vaccine to prevent most APEC infections. We previously reported the stable expression of plasmid pSS27 encoding the APEC O1 O-antigen gene cluster (10.8 kb) in attenuated Salmonella enterica serovar Typhimurium S740 provided excellent protection against APEC O1 challenge. In this study, the plasmid pSS28 harboring the APEC O2 O-antigen polysaccharide gene cluster (15.5 kb) was constructed. Biosynthesis of pSS28-encoded APEC O2 O-antigen in Salmonella vaccine strain S740 was validated by Western blot. The recombinant Salmonella vaccine strain S740 (pSS28) elicited homologous protection against virulent wild-type APEC O2 challenge in a chicken model. Furthermore, through equal-volume mixing the two monovalent vaccine strains S740 (pSS27) and S740 (pSS28), a bivalent vaccine candidate against both APEC O1 and O2 was developed. Immunization of chickens with the bivalent vaccine elicited production of serum IgG and mucosal sIgA antibodies against the LPS of both APEC O1 and O2. Moreover, antibodies induced by the bivalent vaccine promoted opsonization, provoked complement-mediated bactericidal activity, and elicited protection against lethal challenge with both virulent APEC O1 and O2 strains. These results demonstrate that the bivalent vaccine comprised of S740 (pSS27) and S740 (pSS28) is a promising vaccine candidate against APEC O1 and O2 infection.