| Avian pathogenic Escherichia coli (APEC) is a frequent cause of extraintestinal infections, collectively called colibacillosis in poultry. Each year, the poultry industry absorbs important financial losses due to the high morbidity and mortality rates caused by APEC.Four avian pathogenic Escherichia strains were identified as intestinal origin through the methods of morphology and biochemistry. The inactivated vaccine was prepared by using isolated strains from the local. The immunity of the chicks inoculated with the vaccine was greatly improved and the strong protection was obtained.In order to amplify pilA gene and OmpC gene of avian pathogenic Escherichia(AEPC) strain, two pairs of primers were designed according to the GenBank sequences, and a 549bp pilA gene and a 1104bp OmpC gene were obtained by PCR separately. Sequence analysis indicated that the homology of the nucleotide sequence of AEPC strain to those other reference strains was 98.18% of the pilA gene and 97.28% of the OmpC gene.Two expression plasmids pETpilA and pETOmpC were constructed by inserting pilA gene and OmpC gene into the prokaryotic expression vector pET-28a. The two plasmids were transformated into E.coli BL21 separately and two recombinant strains BL21(DE3)(pETpilA) and BL21(DE3) (pETOmpC) were obtained. The type I fimbraie and the outer membrane protein were highly expressed when the recombinant strain were induced by IPTG and the express condition were optimazed separately.Two specific protein were detected by SDS-PAGE and immunogenicity of the expressed protein were confirmed by ELISA and western blot, which seems to be immunogenic crucially. The expressed fimbraie and OmpC were transformed into vaccines. The protective immune response was proved after the mice was immunized with the two vaccines. The results showed that the recombinant strain BL21(pETpilA) and BL21(pETOmpC) could be as candidate vaccines to provide protective immune response against AEPC infection. |
