Isolation, identification, and drug resistance analysis of a strain of enterotoxigenic Escherichia coli derived from piglet and preparation of its specific yolk antibody

doi:10.14088/j.cnki.issn0439-8114.2023.04.023

Abstract

Abstract: To identify the pathogen of diarrhea of piglets in a large-scale pig farm and prepare the specific yolk antibody, bacterial isolation and culture, biochemical test, 16SrRNA PCR identification, phylogenetic tree analysis, drug sensitivity test and other methods were used to study the disease materials. Antigen was prepared by whole cell inactivation and ultrasonic fragmentation. It was emulsified with different adjuvants（ISA 71VG, ISA201VG and 15AVG） to prepare immunized laying hens, the high immune yolk antibody was extracted with polyethylene glycol, the mass and concentration of the Antibody were identified by SDS-PAGE electrophoresis and Bradford method, and the antibody titer was determined by two-way AGAR diffusion test. Results showed that the shape, size and biochemical characteristics of the isolates were consistent with escherichia coli. The bands of E. coli-estb and elt were 113 and 272 bp, respectively. The elt and estb genes of the isolates were in the same branch as those of the American epidemic strains, such as UMNK88. They were in a different branch from the Danish strain CFSAN018748 and the Swiss strain 14OD0056. The isolates were sensitive to cefotaxime, norfloxacin and cotrimoxazole, mediated to ceftriaxone and polymyxin B, and resistant to doxycycline, furazolidone and many aminoglycosides and quinolones. The heavy chain （about 70 ku） and light chain （about 25 ku） of yolk antibody were obvious in each group, and the purity was high. The yolk antibody protein content of the vaccine produced by the combination of bacteria solution treated with 0.2% formaldehyde and ISA 15AVG at 56 d after the first immunization was the highest （1.462 mg/g yolk）, followed by the yolk antibody protein content of the vaccine produced by the combination of bacteria solution treated with ultrasonic disruption and ISA 71VG at 56 days after the first immunization（1.459 mg/g yolk）. The titers of serum antibody and egg yolk antibody were as high as 1∶16 and 1∶4, respectively, when the ultrasonic broken antigen was combined with ISA 71VG.

Key words: swine pathogenic Escherichia coli, separate culture, evolutionary analysis, drug resistance analysis, egg yolk antibody

CLC Number:

S852.4⁺3

YIN Jia-yin, YU Jiao, TANG Qing-hai, TENG Wei, YANG Kun, CAO Xin, XU Qing, LIU Bo, LI Ze. Isolation, identification, and drug resistance analysis of a strain of enterotoxigenic Escherichia coli derived from piglet and preparation of its specific yolk antibody[J]. HUBEI AGRICULTURAL SCIENCES, 2023, 62(4): 127-134.

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