Evaluation of immunogenicity and protective efficacy of combination heat-killed immunogens from three entero-invasive bacteria in rabbit model

Diarrhea is a very common health problem in both developing and developed countries. Among the major entero-invasive bacteria, Shigella, Salmonella and Campylobacter cause serious problems in different geographic regions. Recently we have shown immunogenicity and protective efficacy of heat killed multi-serotype Shigella immunogen in different animal models. In our present study, we have advanced our research by preparing a combination heat-killed immunogen of three different entero-invasive bacteria, Shigella, Salmonella and Campylobacter. After three doses on 0th, 14th and 28th day of oral immunization with tri-valent heat-killed (TVHK) immunogen in rabbit model, the immunogenicity was determined by differential count of white blood cells and immunoglobulin assay at various time points. During oral immunization differential count of lymphocytes increased where as polymorphonuclear leucocytes (PMN) count decreased. Serum IgG and IgA showed significant elevation during oral immunization and remained at a detectable value upto 120 days. Protection study was performed in both, in vitro and in vivo conditions, using bacteriocidal assay and rabbit ligated ileal loop model, respectively, which conferred protection against homologous bacteria. Moreover, immunoblot assay against whole cell lysate and lipopolysaccharide exhibited significant amount of antigen-specific immunoglobulins raised against three different bacteria which proved that proteins along with lipopolysaccharides played a pivotal role in immunogenicity and protective efficacy. This trivalent heat-killed immunogen could be a low-cost, simple, oral, non-living vaccine candidate for future use against invasive diarrhea.