We examined Babesia bovis sporozoites for the expression of two molecules, merozoite surface antigen 1 (MSA-1) and rhoptry-associated protein 1 (RAP-1), that are postulated to be involved in the invasion of host erythrocytes. Both MSA-1 and RAP-1 were transcribed and expressed in infectious sporozoites. Importantly, monospecific MSA-1 and RAP-1 antisera each inhibited sporozoite invasion of erythrocytes in vitro. This is the first identification of antigens expressed in Babesia sp. sporozoites and establishes that, at least in part, sporozoites and merozoites share common targets of antibody mediated inhibition of erythrocyte invasion. (7,14,15,24). We hypothesized that surface molecules may be shared by both babesial sporozoites and merozoites and represent a common target for antibody-mediated inhibition of erythrocyte invasion. In this study, we tested this hypothesis by using Boophilus microplus transmission of B. bovis sporozoites.
Parasites in the genusBabesial merozoites, like other apicomplexan parasites, use a combination of cell surface and apical complex molecules to bind and invade host cells (2,4,29). In B. bovis, merozoite surface antigen 1 (MSA-1) and rhoptry-associated protein 1 (RAP-1) are postulated to be involved in the merozoite invasion of erythrocytes. MSA-1 is a 42-kDa glycoprotein that belongs to the family of variable merozoite surface antigens (VMSA) and is uniformly distributed on the surface of B. bovis merozoites (8, 9, 16). MSA-1 is encoded by a single gene, and antibodies against either native or recombinant MSA-1 neutralize merozoite infection in vitro, suggesting a role in the early steps of erythrocyte invasion (10,11,30). RAP-1 of B. bovis is a 60-kDa protein localized to the apical surface and within the rhoptries of merozoites (8,26,33). RAP-1 is encoded by two identical, tandemly arranged rap-1a genes and is highly conserved among diverse isolates (1, 31-33). Studies using its orthologue in B. bigemina, a 58-kDa RAP-1, show that monoclonal antibodies (MAbs) against RAP-1 are able to inhibit multiplication in vitro (5, 6). Consistent with a role in invasion, calves immunized with native RAP-1 or a recombinant fusion protein develop a significantly reduced mean peak parasitemia upon merozoite challenge (20,34).To obtain sporozoites for the examination of msa-1 and rap-1 expression, adult B. microplus ticks were allowed to feed on a splenectomized calf by using skin patches (12). A Babesiafree colony of B. microplus ticks (La Minita strain) was used in all the experiments. Adult female ticks start engorgement approximately 21 days after being placed on the calf (22). Calves were inoculated with 5 ml of the Mexico strain of B. bovis at 13 days postattachment so that parasitemia, determined by microscopic examination of Giemsa-stained blood smears, was maximal during the final stages of female tick engorgement. Engorged ticks were washed and placed in individual vials during ovoposition (13). To obtain a high percentage of infected ticks, only those females replete during the pe...