A Recombinant Attenuated Yersinia pseudotuberculosis Vaccine Delivering a Y. pestis YopENt138-LcrV Fusion Elicits Broad Protection against Plague and Yersiniosis in Mice

In this study, a novel recombinant attenuated Yersinia pseudotuberculosis P81+ strain (chi 10069) engineered with Delta yopK Delta yopJ Delta asd triple mutations was used to deliver a Y. pestis fusion protein. YopE amino acid 1 to 138-LcrV (YopE(Nt138)(-)LcrV), to Swiss Webster mice as a protective antigen against infections by yersiniae. chi 10069 bacteria harboring the pYA5199 plasmid constitutively synthesized the YopE(Nt138)-LcrV fusion protein and secreted it via the type 3 secretion system (T3SS) at 37 degrees C under calcium -deprived conditions. The attenuated strain chi 10069(pYA5199) was manifested by the establishment of controlled infection in different tissues without developing conspicuous signs of disease in histopathological analysis of microtome sections. A single -dose oral immunization of chi 10069(pYA5199) induced strong serum antibody titers (log(10) mean value, 4.2), secretory IgA in bronchoalveolar lavage (BAL) fluid from immunized mice, and Yersinia-specific CD4(+) and CD8(+) T cells producing high levels of tumor necrosis factor alpha (TNF-alpha), gamma interferon (IFN-gamma), and interleukin 2 (IL -2), as well as IL -17, in both lungs and spleens of immunized mice, conferring comprehensive Th1- and Th2-mediated immune responses and protection against bubonic and pneumonic plague challenges, with 80% and 90% survival, respectively. Mice immunized with chi 10069(pYA5199) also exhibited complete protection against lethal oral infections by Yersinia enterocolitica WA and Y. pseudotuberculosis PB1+. These findings indicated that chi 10069(pYA5199) as an oral vaccine induces protective immunity to prevent bubonic and pneumonic plague, as well as yersiniosis, in mice and would be a promising oral vaccine candidate for protection against plague and yersiniosis for human and veterinary applications.