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 produced against isolated rhoptries is directed predominantly against RAP-1 and reacts with both the merozoite and the membrane of parasitized erythrocytes. 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 antibodies (mAbs) were generated from mice immunized with rhoptries isolated from the JG-29 clone of the Mexico strain. The anti-RAP-1 mAbs bound only merozoites in a punctate immunofluorescence pattern. A second group of four mAbs, none of which were reactive with RAP-1, bound the parasitized erythrocyte. Two of these latter mAbs, 64/44.17.3 and 64/05.7.2, reacted only with parasitized erythrocytes that had been permeabilized. MAb 64/44.17.3 bound a 54-kDa merozoite polypeptide while 64/05.7.2 bound a > or = 225-kDa merozoite polypeptide. MAbs 64/32.8.5 and 64/38.5.3 recognized epitopes on 17.5- and 76-kDa polypeptides exposed on the external surface of intact parasitized erythrocytes. The results indicate that the identified RAP-1 epitopes are not associated with the erythrocyte cytoskeleton or membrane and that anti-RAP-1 immunity is most likely generated against the free merozoite. All new mAbs reacted with every B. bigemina strain tested (Mexico, Puerto Rico, St. Croix, Texcoco, Jaboticabal). The conservation of RAP-1 epitopes among these strains supports the continued testing of RAP-1 as a vaccine component. In addition, the identification of epitopes expressed on the surface of erythrocytes infected with all five strains provides new candidate immunogens.
Department of Veterinary Microbiology and Pathology, Washington State University, Pullman 99164-7040, USA.
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Vidotto
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McElwain
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Machado
L E
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Perryman
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Suarez
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Palmer
D000818:Animals; D000911:Antibodies, Monoclonal; D000953:Antigens, Protozoan / Q000276:immunology*; D001402:B-Lymphocytes / Q000276:immunology*; D001403:Babesia / Q000276:immunology*; D016793:Babesia bovis / Q000276:immunology; D015153:Blotting, Western; D002417:Cattle; D000939:Epitopes; D004912:Erythrocytes / Q000469:parasitology*; D005455:Fluorescent Antibody Technique; D011233:Precipitin Tests; D015800:Protozoan Proteins / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000953:Antigens, Protozoan; D000939:Epitopes; D015800:Protozoan Proteins; C097551:rhoptry-associated protein-1, Babesia