Progress 07/15/11 to 03/14/12
Outputs
OUTPUTS: Enterotoxigenic Escherictia coli (ETEC) is a major pathogen of young weaned piglets, yet there currently are no licensed non-living vaccines for the disease, and available living vaccines do not express heat labile enterotoxin (LT) antigen. That toxin is a virulence determinant in the majority of postweaning strains of ETEC. This may limit the efficacy and/or breadth of protection of available live vaccines. We recently developed a model non-living vaccine based on purified K88 fimbriae and LT and demonstrated a high level of efficacy under controlled challenge conditions. This model vaccine elucidates essential vaccine components and a delivery method. However, the product is too cumbersome and expensive to produce for practical commercial application. The purpose of the current project was to simplify the experimental subunit vaccine to produce an inexpensive product with high efficacy for weaned pigs. Several methods of fimbriae and LT extraction were tested with varying extraction successes. The most promising method involved use of 8M urea to strip fimbriae and enterotoxins from the surface of E. coli, with urea removal by step-wise dialysis. It was tested for ability to stimulate antibody production and provide efficacy against virulent ETEC challenge. Both safety and efficacy tests were conducted in young pigs of genetically characterized background (K88 receptor positive). Vaccine was prepared to achieve 10 micrograms LT/dosage and significantly more K88 antigen, and was delivered in two dosages by the intranasal route at 10 and 17 days of piglet age. Piglets were challenged with virulent ETEC (O157:NM; K88/LT/STb) at 24-27 days of age. Parameters measured included clinical signs of disease, change in body weight and serum antibody titers measured by ELISA. PARTICIPANTS: Mojun Zhao, MD, MS, Principal Investigator--also completeing a PhD at the time David H. Francis, Consultant, Professor, South Dakota State University and President of Brookings Biomedical, Inc Denise Owens, Masters Student--assisted with ELISA tests for antibody titers and in the preparation of vaccine antigens TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
While non-toxic when delivered by the intranasal route, immune responses in vaccinated pigs were disappointingly limited to that against LT. While piglets in the study all bore phenotypic characteristics of inherent susceptibility to K88 ETEC, controls were largely refractory to clinical disease. This suggested presence of a here-to-fore undescribed K88+ ETEC resistance phenotype in which K88 receptors are present, but animals are refractory to disease. However, vaccinees trended to more weight gain, while controls did not. Inability of pigs to respond immunologically to K88 antigen was possibly due to a urea-caused denaturation of the antigens. Alternatively poor immunological response to K88 could have been due to presence of a factor or condition in the intestines of these disease-resistant pigs that prevented K88 adhesion to its receptor, a condition previously shown important to antibody production. Alternative methods of antigen production will be explored and different sources of pigs will be identified and assessed for disease susceptibility, in an effort to produce and efficacious, yet inexpensive vaccine for ETEC.
Publications
- No publications reported this period
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