Molecular characterization and expression of a 47- kDa merozoite surface protein of Babesia gibsoni for serodiagnosis by enzyme-linked immunosorbent assay

Abstract

A novel gene encoding a 47-kDa surface protein was identified by bioinformatic analyses of Expressed Sequence Tags (EST) prepared from mRNA of Babesia gibsoni. The Bgp47 cDNA encodes a 47-kDa polypeptide having an-N terminal signal peptide, a central hydrophilic core and a C-terminal transmembrane domain. This protein shares homology with 50-kDa (BgP50) and 32-kDa (BgP32) proteins of B. gibsoni. The expressed recombinant protein without the signal peptide and the transmembrane domain (BgP47t) was approximately 46-kDa after glutathione S-transferase （GST) cleavage, similar to an approximately 47-kDa mature native protein identified in the parasite merozoite after probing with mouse anti-BgP47t serum. A 32-kDa band was also identified using the same antiserum suggesting that BgP47 and BgP32 might share common B cell epitopes. The BgP47 was localized predominantly on the parasite cell surface. An enzyme-linked immunosorbent assay (ELISA) revealed that the rBgP47t did not cross-react with B. canis subspecies and other genetically related apicomplexans indicating that the antigen is specific. Out of 106 sera from dogs with B. gibsoni-like symptoms, 79.2% were seropositive by rBgP47t-ELISA. Therefore, the BgP47 is prospective serodiagnostic antigen.