Cloning And Expression Of A Gene Encoding An Apical Membrane Antigen-1 Homologue Of Babesia Gibsoni And Its Study In Serodiagnosis

A cDNA expression library prepared from Babesia gibsoni merozoite mRNA was screened with B. gibsoni-infected dog serum. Twenty-seven repeat positive cDNA clones were obtained. The sequence of these cDNA is not integral, lacking 5'-end sequence. Polymerase chain reaction was performed to amplify 5'-end sequence. The complete nucleotide sequence of the gene was 2,108 bp. Computer analysis suggested that sequence contains an open reading frame of 1,794 bp with a coding capacity of approximately 66 kDa. The deduced amino acid sequence shows a high homology to the apical membrane antigen-1 (AMA-1) of Plasmodium species. This gene was designated B. gibsoni AMA-1 (BgAMA-1). The BgAMA-1 gene was expressed in E. coli BL21 strain, and used as the antigen in an enzyme-linked immunosorbent assay (ELISA). The ELISA was able to differentiate between B. gibsoni-infected dog sera and B. gibsoni-free dog sera or the sera from dogs infected with B. canis canis, B. canis rossi, B. canis vogeli, and T. gondii. Furthermore, the antibody response against the recombinant BgAMA-1 was maintained until the chronic stage of infection in a dog experimentally infected with B. gibsoni. On the other hand, the native BgAMA-1 of B. gibsoni was identified with an anti-recombinant BgAMA-1 mouse serum. These results demonstrate that the recombinant BgAMA-1 might be a useful diagnostic reagent for detection of antibodies to B. gibsoni in dogs.