Abstract
Enterotoxigenic Escherichia coli (ETEC) is an important type of pathogenic bacteria that causes diarrhea in humans and young livestock. The pathogen has a high morbidity and mortality rate, resulting in significant economic losses in the pig industry. To effectively prevent piglet diarrhea, we developed a new tetravalent genetically engineered vaccine that specifically targets ETEC. To eliminate the natural toxin activity of ST(1) enterotoxin and enhance the preventive effect of the vaccine, the mutated ST (1), K88ac, K99, and LT (B) genes were amplified by PCR and site-specific mutation techniques. The recombinant strain BL21(DE3)(pXKK3SL) was constructed and achieved high expression. Animal experiments showed that the inactivated vaccine had eliminated the natural toxin activity of ST(1). The immune protection test demonstrated that the inclusion body and inactivated vaccine exhibited a positive immune effect. The protection rates of the inclusion body group and inactivated vaccine group were 96 and 98%, respectively, when challenged with 1 minimum lethal dose, indicating that the constructed K88ac-K99-3ST(1)-LT(B) vaccine achieved a strong immune effect. Additionally, the minimum immune doses for mice and pregnant sows were determined to be 0.2 and 2 mL, respectively. This study suggests that the novel K88ac-K99-3ST(1)-LT(B) vaccine has a wide immune spectrum and can prevent diarrhea caused by ETEC through enterotoxin and fimbrial pathways. The aforementioned research demonstrates that the K88ac-K99-3ST(1)-LT(B) vaccine offers a new genetically engineered vaccine that shows potential for preventing diarrhea in newborn piglets.