Prioritizing and testing sub-dominant A. marginale antigens for protective capacity

Deirdre R. Ducken

Anaplasma marginale is a tick-borne rickettsial pathogen of cattle that is found worldwide. A. marginale infects erythrocytes leading to anemia and reduced production, and death in some cases. Currently in the USA, there are no USDA-licensed vaccines. Whole outer membrane protein extract or a more well- defined protein complex are protective against infection in 40-70% of animals, and protective against anemia and bacteremia in nearly all animals. However, the specific antigen or antigens that are responsible for protection are unknown. Am202, am368, am854, am936, am1041, and am1096 (oma87) have been identified as components of these protective outer membrane preparations or are thought to mediate cell invasion and thus are primary vaccine candidates. Ideally, a vaccine would induce broad protection against geographically diverse organisms. The goals of these experiments were to 1) determine if any of these candidates were broadly conserved among geographically diverse isolates; 2) determine if these candidates were recognized by antibody from protectively immunized animals; and 3) determine if candidates that were recognized by protectively immunized animals would protect animals from challenge. To address the first goal, each gene was cloned and sequenced from seven geographically diverse strains. Am202, am854, am936, and am1096 were conserved among all isolates with 100% amino acid identity. Am1041 and am368 had some strain variation, with non-synonymous nucleotide polymorphisms in several isolates. To address the second question, we determined the titers of total IgG directed against Am202, Am854, Am936 and Am1096 that were produced in animals immunized with outer membranes, linked protein complexes, or unlinked proteins complexes and shown to be protected against disease from challenge. As the titers to Am854 and Am936 were the highest and these proteins were the most widely recognized by protectively immunized animals, they were used in an immunization and challenge experiment. Outer membrane immunized animals and recombinant protein immunized animals had similar total IgG and IgG2 response to Am854 and Am936. However, animals immunized with Am854 and Am938 had statistically significantly higher bacteremia and more severe anemia than the group that received only adjuvant, while the animals immunized with outer membrane proteins were protected against severe disease from challenge, as expected.

Prioritizing and testing sub-dominant A. marginale antigens for protective capacity

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