Application of a Novel Epitope- and Structure-Based Vaccinology-Assisted Fimbria-Toxin Multiepitope Fusion Antigen of Enterotoxigenic Escherichia coli for Development of Multivalent Vaccines against Porcine Postweaning Diarrhea

Abstract

Enterotoxigenic Escherichia coli (ETEC) strains producing K88 (F4) or F18 fimbriae and enterotoxins are the predominant cause of pig postweaning diarrhea (PWD). We recently identified neutralizing epitopes of fimbriae K88 and F18, heat-labile toxin (LT), heat-stable toxins type I (STa) and type II (STb), and Shiga toxin 2e (Stx2e). In this study, we explored a novel epitope- and structure-based vaccinology platform, multiepitope fusion antigen (MEFA), for PWD vaccine development. By using an epitope substitution LT toxoid, which lacks enterotoxicity but retains immunogenicity, as the backbone to present neutralizing epitopes of two ETEC fimbriae and four toxins, we generated PWD fimbria-toxin MEFA to mimic epitope native antigenicity. We then examined MEFA protein immunogenicity and evaluated MEFA application in PWD vaccine development. Mice subcutaneously immunized with PWD MEFA protein developed strong IgG responses to K88, F18, LT, and STb and moderate responses to the toxins Stx2e and STa. Importantly, MEFA-induced antibodies inhibited adherence of K88 or F18 fimbrial bacteria to pig intestinal cells and also neutralized LT, STa, STb, and Stx2e toxicity. These results indicated that PWD fimbria-toxin MEFA induced neutralizing antibodies against an unprecedent two fimbriae and four toxins and strongly suggested a potential application of this MEFA protein in developing a broadly protective PWD vaccine.IMPORTANCE ETEC-associated postweaning diarrhea (PWD) causes significant economic losses to swine producers worldwide. Currently, there is no effective prevention against PWD. A vaccine that blocks ETEC fimbriae (K88 and F18) from attaching to host receptors and prevents enterotoxins from stimulating water hypersecretion in pig small intestinal epithelial cells can effectively protect against PWD and significantly improves pig health and well-being. The fimbria-toxin MEFA generated from this study induced neutralizing antibodies against both ETEC fimbriae and all four ETEC toxins, suggesting a great potential of this fimbria-toxin MEFA in PWD vaccine development and further supporting the general application of this novel MEFA vaccinology platform for multivalent vaccine development.

Keywords: ETEC; MEFA; PWD; enterotoxigenic E. coli; fimbria; multiepitope fusion antigen; novel vaccinology; postweaning diarrhea; toxin; vaccine.