The Expression of apxIVA Gene of Actinobacillus pleuropneumoniae Serotype 5 and Establishment of an Indirect ELISA for Distinguishing Infection and Immunization

Actinobacillus pleuropneumoniae is the etiological agent of Porcine pleuropneumonia, which causes severe losses in pig farming. To establish an serological method to detect A. pleuropneumoniae infection, bioinformatics method was utilized to analyzed the sequence of apxIVA gene of A. pleuropneumoniae serotype 5 completed by our lab, a 1152 bp fragment of the Nʹ-terminal of apxIVA gene was amplified and cloned into the prokaryotic expression vector pET-32a(+), a recombinant protein about 62KD was expressed upon isopropy 1-P-D- thiogalactoside (IPTG) induction. The protein could react specifically with antiserum from live A. pleuropneumoniae infection in western-blot. After further purification by Ni-NTA, this protein was used to establish an indirect ELISA to detect A. pleuropneumoniae infection. This method showed high specificity and could react positively with antibodies of live A. pleuropneumoniae infection while negatively with those of inactivated A. pleuropneumoniae immunization. In conclusion, this indirect ELISA could be used to detect A.pleuropneumoniae infection in mice model.