A novel neutralization sensitive and subdominant RAP-1-related antigen (RRA) is expressed byBabesia bovismerozoites

SUMMARYObjective.TheBabesia bovisgenome encodes arap-1related gene denominated RAP-1 related antigen (RRA). In this study, we analysed the pattern of expression, immunogenicity and functional relevance of RRA.Methods.Phylogenetic analysis was performed using the program Phylip. Expression ofrrawas analysed by Northern blots, RT-PCR, immunoprecipitation, Western blots and immunofluorescence. RRA antigenicity was tested by T-cell proliferation and Western blot analysis, and functional relevance was determined in anin vitroneutralization assay.Results.RRA is more closely related to RAP-1b ofBabesia bigeminathan toB. bovisRAP-1, and it is highly conserved among distinct strains. Transcriptional analysis suggests lower numbers ofrratranscripts compared torap-1.Immunoprecipitation of metabolically labelledB. bovisproteins with antibodies against synthetic peptides representing predicted antigenic regions of RRA confirmed the expression of a ∼43 kDa RRA in cultured merozoites. Antibodies present inB. bovishyperimmune sera, but not in field-infected cattle sera, reacted weakly with recombinant RRA, and no significant stimulation was obtained using recombinant RRA as antigen in T-cell proliferation assays, indicating that RRA is a subdominant antigen. Antibodies against RRA synthetic peptides reacted with merozoites using immunofluorescence, and were able to significantly inhibit erythrocyte invasion inin vitroneutralization tests, suggesting functional relevance for parasite survival.Conclusion.B. bovisexpress a novel subdominant RAP-1-like molecule that may contribute to erythrocyte invasion and/or egression by the parasite.