Cloning And Expression Of The Gene Fragments Encoding Surface Protein OmpA And OmpB Of Spotted Fever Group Rickettsiae And Study Of The Antigenicities Of The Recombinant Proteins

The spotted fever group (SPF) rickettsae were the pathogens that cause spotted fever in genus Rickettsia, including R. rickettsii, R. heilongjiangii, and so on.The major immunodominant surface-exposed proteins of SFG rickettsiae, outer membrane proteins A (OmpA) and B (OmpB), are the principal protective antigens. The objects of the study are to clone and express the gene fragments encoding the OmpA and OmpB of SFG rickettsiae and to analyze the antigenicities of the recombinant OmpA and OmpB.The gene fragments encoding partial OmpB and OmpA were amplified by PCR from the genomic DNAs of R. rickettsii and R. heilongjiangii, respectively. The gene fragments were cloned into prokaryotic expression vector pQE30, respectively. E. coli cells were transformed with the recombinant plasmid pQE30/ompB and pQE30/ompA; and the transformants were induced to express the recombinant proteins by IPTG.. A 44kDa (OmpB) recombinant protein expressed in the pQE30/ompB-transformed cells and a 56kDa (OmpA) recombinant protein expressed in the pQE30/ompA-transformed cells were identified in SDS-PAGE analysis.The OmpB recombinant protein was recognized by sera from rabbits immunized with R. rickettsii, but did not by immunosera to other rickettsial agents as demonstrated by immunoblotting assay. R. rickettsii organism, but not other rickettsial agents, was detected by IFA with the immunoserum to the recombinant proteins It indicates that the OmpB recombinant protein is species-specific for R. rickettsii and the immunosera to it may be useful to identify R. rickettsii.To study the immunogenicity of the OmpA recombinant protein of R. heilongjiangii, the purified OmpA recombinant protein were used to immunize BALB/c mice and guinea pigs. The high levels of IgGs to R. heilongjiangii and other rickettsial agents were detected in the sera from the immunized animals by IFA. The high levels of antigen-specific spleen cell proliferative responses were demonstrated in the immunized mice by MTT method.The OmpA-immunized guinea pigs were challenged with R. rickettsii or R. heilongjiangii.