Isolation, identification and evaluation of inactivated vaccine of enterotoxigenic Escherichia coli (ETEC) in large-scale pig farms in northern Jiangsu

Abstract:

[Objective] Enterotoxigenic Escherichia coli (ETEC) is an important pathogen causing diarrhea of piglets. In this study, the prevalence and biological characteristics of ETEC in large-scale pig farms in northern Jiangsu was investigated, and protective and inactivated vaccines of predominant serotype were developed to provide guidance and suggestions for the prevention and control of ETEC in northern Jiangsu. [Methods] Fresh fecal samples, anal swabs and small intestinal tissue samples of piglets aged 3-30 days were collected from large-scale pig farms in northern Jiangsu province, ETEC was isolated, and serotypes, antibiotic resistance, pathogenicity on mice of these isolates were determined. Finally, the inactivated vaccine of predominant serotype was developed and the immune protection effect of mice was evaluated. [Results] We collected 562 samples from 21 large-scale pig farms. We identified 141 ETEC strains containing ST or LT virulence genes by PCR. O serotypes of 85 isolates were determined by serum agglutination tests. O8, O101 and O128 were dominant serotypes, accounting for 61.2% of the confirmed strains, besides, O9, O3, O20, O148, O149 and the like were identified. The drug resistance of 141 ETEC isolates to 14 common antibiotics was analyzed. The isolates were highly resistant to neomycin, erythromycin, tetracycline, gentamicin, doxycycline, amoxicillin, and trimethoprim/sulfamethoxazole, with resistance rate higher than 80%. Strains were most sensitive to enrofloxacin, with the rate of 50.4% (71/141). 66% (93/141) and 51.8% (73/141) strains were intermediate to polymyxin B and cefotaxime. Multiple drug resistance is serious. 19% (27/141) of strains were resistant to 10 drugs. The virulence of the dominant serotype strains was obtained by mice challenge test. LD₅₀ of one strong strain YC-6 of O8 serotype was measured as 1.4×10⁷ CFU, while the MLD₅₀ was 3×10⁷ CFU. The monovalent inactivated vaccines prepared by O8 virulent strain YC-6 and O101 virulent strain LYG-3 both have a protection rate of 100% for mice. The bivalent inactivated vaccine of YC-6 and LYG-3 were developed as well, the protection rate of which in mice was more than 83%. [Conclusion] Based on the epidemiological investigation of ETEC in northern Jiangsu, dominant serotypes and drug resistance were tested, and bivalent inactivated vaccine with superior immune protection effect against isolated strains were developed, which will provide reference for monitoring and prevention in pig industry.