Remodeling Yersinia pseudotuberculosis to generate a highly immunogenic outer membrane vesicle vaccine against pneumonic plague

A recombinant enteric Yersinia pseudotuberculosis PB1(+) strain (Yptb) was designed to synthesize an adjuvant form of lipid A (monophosphoryl lipid A [MPLA]), and tailor an Asd(+) plasmid pSMV13 for high synthesis of the Yersinia pestis LcrV antigen. The recombinant Yptb mutant harboring the pSMV13 dramatically increased the production of outer membrane vesicles (OMVs) enclosing high amounts of LcrV in comparison to its Y. pestis counterpart. Intramuscular (i.m.) immunization with 40 μg of OMVs from YptbS44(pSMV13) (termed OMV(YptbS44)-Bla-V) afforded complete protection to mice against medium (5 ×10(3) colony-forming units [CFU], 50 median lethal dose [LD(50)]) and high (400 LD(50)) doses of pulmonary Y. pestis infection, as well as against subcutaneous (s.c.) infection with 5 ×10(5) CFU (50,000 LD(50)) of Y. pestis. In addition, i.m. immunization with 40 μg of detoxified OMVs from YptbS45(pSMV13) (termed OMV(YptbS45)-Bla-V) afforded 90% protection against pulmonary challenge with 50 LD(50) of Y. pestis and complete protection against s.c. challenge with 50,000 LD(50) of Y. pestis. The protective efficacy was superior to that of vaccination with the F1V subunit vaccine or OMVs from a previous recombinant Y. pestis strain (termed OMV(Yp)-Bla-V). Further, vaccination with OMV(YptbS44)-Bla-V induced robust humoral and cellular immune responses that were correlated with rapid bacterial clearance, unremarkable tissue damage, and low inflammatory cytokine production in the lungs during pulmonary Y. pestis challenge. Our results imply that the recombinant Yptb OMV delivering the Y. pestis protective antigen(s) merits further development as a next-generation plague vaccine candidate.