BABESIA-BIGEMINA - IDENTIFICATION OF B-CELL EPITOPES ASSOCIATED WITH PARASITIZED ERYTHROCYTES

Rhoptries are involved in host cell invasion and rhoptry polypeptides, including the Babesia bigemina rhoptry-associated protein-1 (RAP-1), are targets for protective immune responses. Polyclonal antisera produ ced against isolated rhoptries is directed predominantly against RAP-1 and reacts with both the merozoite and the membrane of parasitized er ythrocytes. To determine whether these B cell epitopes associated with the parasitized erythrocyte are derived from RAP-1 or, alternatively, from previously undetected merozoite polypeptides, monoclonal antibod ies (mAbs) were generated from mice immunized with rhoptries isolated from the JG-29 clone of the Mexico strain. The anti-RAP-1 mAbs bound o nly merozoites in a punctate immunofluorescence pattern. A second grou p of four mAbs, none of which were reactive with RAP-1, bound the para sitized erythrocyte. Two of these latter mAbs, 64/44.17.3 and 64/05.7. 2, reacted only with parasitized erythrocytes that had been permeabili zed. MAb 64/44.17.3 bound a 54-kDa merozoite polypeptide while 64/05.7 .2 bound a greater than or equal to 225-kDa merozoite polypeptide. MAb s 64/32.8.5 and 64/38.5.3 recognized epitopes on 17.5- and 76-kDa poly peptides exposed on the external surface of intact parasitized erythro cytes. The results indicate that the identified RAP-1 epitopes are not associated with the erythrocyte cytoskeleton or membrane and that ant i-RAP-1 immunity is most likely generated against the free merozoite. All new mAbs reacted with every B. bigemina strain tested (Mexico, Pue rto Rico, St. Croix, Texcoco, Jaboticabal). The conservation of RAP-1 epitopes among these strains supports the continued testing of RAP-1 a s a vaccine component. In addition, the identification of epitopes exp ressed on the surface of erythrocytes infected with all five strains p rovides new candidate immunogens. (C) 1995 Academic Press, inc.