Construction and pathogenic identification of aes-31 gene mutant of Avian Pathogenic Escherichia coli strain E058

Abstract:

Abstract: [Objective] To find the primary function of aes-31 fragment through construction of defined mutation of Avian Pathogenic Escherichia coli strain E058 and animal experiments. [Methods] The fragment of aes-31 was generated by PCR and cloned into pGEM-T-easy vector. A resultant suicide vector containing the aes-31 fragment named pMEG375-aes-31 was constructed and transformed to a receptor strain SM10. Then recombinant strain SM10 was hybridized with E058 strain in solid state. Mutant derivatives of strain E058 were generated by homologous recombination and were named E058(?aes-31). The 50% lethal dose (LD50) of E058 and E058(?aes-31) in commercial day-old chickens experimentally inoculated via intratrachea were 104.3CFU and 103.5CFU, respectively. The same way was used to inoculate with 108 CFU to obtain the pathogenic ability of E058 and E058(?aes-31) in 35-days-old SPF chickens. In the chicken challenge model, the mutant was tested to determine the individual function for virulence and persistence in 2-week-old SPF chicks. [Results] The pathogenicity test for E058 strain and E058(?aes-31) strain showed that the mutant had a higher mortality (75%) to 35-day-old specific pathogen-free (SPF) chicks than that of E058 (62.5%). In the chicken challenge model, there was no obviously CFUs difference in blood and lung in chicks of E058 group and E058(?aes-31) group 6 hours after inoculation. After 24 hours there was obvious CFUs difference in heart, liver, spleen, lung and blood in chicks of E058 group and E058(?aes-31) group. After 48 hours, there was also obvious CFUs difference in heart, liver and spleen in chicks of E058 group and E058(?aes-31) group E058(?aes-31) had a trend of increasing virulence in chicks. [Conclusion] Aes-31 might be associated with negative regulatory gene for E058 virulence and its actual function needed further study.