Stable expression of Shigella dysenteriae serotype 1 O-antigen genes integrated into the chromosome of live Salmonella oral vaccine vector Ty21a

Typhoid fever and shigellosis cause high morbidity and mortality worldwide, yet no anti-Shigella vaccine is currently available. However, to protect against typhoid fever, an approved vaccine, based on the attenuated Salmonella enterica serovar Typhi strain Ty21a is available. We have investigated Ty21a as a live oral vaccine vector for expression of heterologous foreign antigens to protect against other diseases (e.g. shigellosis, anthrax and plague). Shigella lipopolysccharide (LPS) is a potent vaccine antigen for serotype-specific protection against Shigellae. We previously reported the construction of a Ty21a derivative expressing Shigella sonnei O-antigen by insertion of a large (similar to 12.5 kb) operon comprising the S. sonnei O-antigen biosynthetic genes into a targeted site within the Ty21a chromosome using modified lambda red recombineering methods. In the current study, S. dysenteriae 1 O-antigen biosynthetic genes from two separate genetic loci, rfp and rfb, were assembled and inserted into the Ty21a chromosome by lambda red-mediated recombineering to construct strain Ty21a-Sd. To obtain a high level of heterologous LPS expression, the native upstream promoter was replaced with the constitutive lpp promoter, which resulted in Ty21a-Sdl with enhanced heterologous LPS expression. Both Ty21a-Sd and Ty21a-Sdl elicited significant serum antibody responses in mice against both Ty21a and this heterologous Shigella LPS, and conferred protection against virulent S. dysenteriae 1 challenge. This work represents progress toward the goal of a safe and effective vaccine against Shigella.