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Extraintestinal pathogenic Escherichia coli (ExPEC) cause a wide variety of infections that are responsible for significant morbidity, mortality and costs to our healthcare system. An efficacious vaccine against ExPEC would be desirable. Previously, we demonstrated that nasal immunization with a genetically engineered strain in which capsule and O-antigen are no longer expressed (CP923) was immunogenic, generated antibodies that bound a subset of heterologous ExPEC strains, and enhanced neutrophil-mediated bactericidal activity against the homologous and a heterologous strain in vitro. In the work reported here we tested the hypothesis that nasal immunization with CP923 conferred protection in a mouse intravenous sepsis model. Nasal immunization with the wild-type strain CP9 conferred protection against challenge with itself and this protection was enhanced when IL-12 was used as an adjuvant. However, when CP923 was used the immunogen, protection was not observed against challenge with CP9. Next, we hypothesized that the observed lack of protection may be due to capsule and the O-antigen moiety of lipopolysaccharide (LPS) impeding antibody binding to non-capsule and O-antigen epitopes. This hypothesis was substantiated by in vitro binding assays, which demonstrated that binding of polyclonal anti-CP923 antisera was decreased when capsule and/or O-antigen were present. Lastly, neutrophil-mediated bactericidal activity against CP923, opsonisized with anti-CP923 antisera, was significantly increased compared to CP9. Taken together, these results demonstrate that the capsule and O-antigen form a biologically significant barrier against antibodies directed against non-capsular and O-antigen epitopes. This defense against the acquired immune response will need to be overcome for the development of a successful vaccine against ExPEC.

Infections due to extraintestinal pathogenic E. coli (ExPEC) result in significant morbidity, mortality and increased healthcare costs. An efficacious vaccine against ExPEC would be desirable. In this report, we explore the use of killed-whole E. coli as a vaccine immunogen. Given the diversity of capsule and O-antigens in ExPEC, we have hypothesized that alternative targets are viable vaccine candidates. We have also hypothesized that immunization with a genetically engineered strain that is deficient in the capsule and O-antigen will generate a greater immune response against antigens other than the capsular and O-antigen epitopes than a wild-type strain. Lastly, we hypothesize that mucosal immunization with killed E. coli has the potential to generate a significant immune response. In this study, we demonstrated that nasal immunization with a formalin-killed ExPEC derivative deficient in capsule and O-antigen results in a significantly greater overall humoral response compared to its wild-type derivative (which demonstrates that capsule and/or the O-antigen impede the development of an optimal humoral immune response) and a significantly greater immune response against non-capsular and O-antigen epitopes. These antibodies also bound to a subset of heterologous ExPEC strains and enhanced neutrophil-mediated bactericidal activity against the homologous and a heterologous strain. Taken together, these studies support the concept that formalin-killed genetically engineered ExPEC derivatives are whole cell vaccine candidates to prevent infections due to ExPEC.