Immunization of cattle with a purified Anaplasma marginale major surface protein, AmF36, induced protection against homologous challenge with the Florida isolate. Similarly, immunized cattle were protected from challenge with the antigenically and structurally distinct Washington-O isolate of A. marginale. The degree of protection in AmF36-immunized cattle varied from complete prevention of rickettsemia to significant delay in the onset of rickettsemia compared with control immunized cattle. A single AmF36 vaccinate was not protected against homologous challenge despite development of a strong antibody response. Immunoprecipitation of A. marginale proteins with a monoclonal antibody to AmF36 identified minor molecular size heterogeneity in this protein from different isolates, including the Florida and Washington-O isolates. The apparent molecular size of this surface protein in the Florida isolate was 36 kilodaltons, whereas the analogous proteins in Washington-O and four other isolates of A. marginale from the United States had molecular masses of 33 to 34 kilodaltons. Significantly, the surface-exposed peptides of these proteins appear to be conserved among the different isolates. These results demonstrate the potential of AmF36 as a subunit immunogen for bovine anaplasmosis and indicate a structural basis for its cross-protective ability.

A major surface protein complex from the Florida isolate of Anaplasma marginale has been previously shown to induce protection in immunized cattle and has been proposed as the basis of a subunit vaccine against anaplasmosis. This complex in the Florida isolate is composed of two noncovalently associated polypeptides with molecular masses of 105 and 100 kilodaltons (kDa). The analogous protein complex from four geographically different isolates of A. marginale was immunoprecipitated and compared with the protein complex of the Florida isolate. The polypeptides of the complex varied in apparent molecular mass among the isolates. By using antibodies recognizing epitopes on each polypeptide of the Florida isolate, the antigenic identity of the polypeptides in the analogous complexes was determined. The polypeptides recognized by the neutralizing monoclonal antibody 22B1, which recognizes a 105-kDa polypeptide in the Florida isolate, ranged from 70 to 100 kDa in the other isolates. Those polypeptides recognized by rabbit antiserum R911, which recognizes a 100-kDa polypeptide in the Florida isolate, ranged from 97 to 100 kDa. The surface-exposed peptides in the complexes were compared by limited enzymatic digestion to assess structural homology among isolates. Despite the marked variations in molecular weight, there were conserved peptides between the 22B,-reactive polypeptides and between the R911-reactive peptides. Determination of the role of the conserved peptides in inducing immunity will be critical in the application of these polypeptides as the basis of a subunit vaccine for bovine anaplasmosis.

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Defining conserved, protective epitopes is essential to the design of an effective vaccine against bovine anaplasmosis. MSP4, one of six initial body proteins recognized by a neutralizing serum, is conserved among Anaplasma mainale isolates at both the protein and the DNA levels. Sera from cattle immunized with an outer membrane fraction ofA. malginale and protected from a virulent challenge bind MSP4. The gene for MSP4 has been cloned, and the recombinant protein has been expressed, isolated, and demonstrated to share epitopes with the native protein expressed on initial bodies. MSP4 may have a greater potential to protect cattle from a challenge by heterologous isolates than other A. marginake surface proteins, which vary widely in size and structure.