Expression of 6-Cys Gene Superfamily Defines Babesia bovis Sexual Stage Development within Rhipicephalus microplus

Babesia bovis , an intra-erythrocytic tick-borne apicomplexan protozoan, is one of the causative agents of bovine babesiosis. Its life cycle includes sexual reproduction within cattle fever ticks, Rhipicephalus spp. Six B . bovis 6-Cys gene superfamily members were previously identified ( A , B , C , D , E , F ) where their orthologues in Plasmodium parasite have been shown to encode for proteins required for the development of sexual stages. The current study identified four additional 6-Cys genes ( G , H , I , J ) in the B . bovis genome. These four genes are described in the context of the complete ten 6-Cys gene superfamily. The proteins expressed by this gene family are predicted to be secreted or surface membrane directed. Genetic analysis comparing the 6-Cys superfamily among five distinct B . bovis strains shows limited sequence variation. Additionally, A , B , E , H , I and J genes were transcribed in B . bovis infected tick midgut while genes A , B and E were also transcribed in the subsequent B . bovis kinete stage. Transcription of gene C was found exclusively in the kinete. In contrast, transcription of genes D , F and G in either B . bovis infected midguts or kinetes was not detected. None of the 6-Cys transcripts were detected in B . bovis blood stages. Subsequent protein analysis of 6-Cys A and B is concordant with their transcript profile. The collective data indicate as in Plasmodium parasite, certain B . bovis 6-Cys family members are uniquely expressed during sexual stages and therefore, they are likely required for parasite reproduction. Within B . bovis specifically, proteins encoded by 6-Cys genes A and B are markers for sexual stages and candidate antigens for developing novel vaccines able to interfere with the development of B . bovis within the tick vector.