USE OF A SPECIFIC IMMUNOGENIC REGION ON THE COWDRIA-RUMINANTIUM MAP1 PROTEIN IN A SEROLOGICAL ASSAY

Currently available serological tests for cowdriosis (Cowdria ruminant ium infection) in domestic ruminants are hampered by their low specifi cities because of cross-reactivity with Ehrlichia spp, The use of reco mbinant major antigenic protein (MAP1) of C. ruminantium for serodiagn osis was investigated, Overlapping fragments of the MAP1 protein were expressed in Escherichia coli and were reacted with sera from sheep in fected with either C. ruminantium or Ehrlichia ovina, Two immunogenic regions on the MAP1 protein, designated MAP1-A and MAP1-B, were identi fied. MAP1-A was reactive with C. ruminantium antisera, E. ovina antis era, and three MAP1-specific monoclonal antibodies, whereas MAP1-B rea cted only with C. ruminantium antisera, An indirect enzyme-linked immu nosorbent assay (ELISA) based on MI-B was further developed and valida ted with sera from animals experimentally infected with C. ruminantium or several Ehrlichia spp, Antibodies raised in sheep, tattle, and goa ts against nine isolates of C. ruminantium reacted with MAP1-B. Cross- reactivity with MAP1-B was limited to Ehrlichia canis and Ehrlichia ch affeensis, two rickettsias which do not infect ruminants. Antibodies t o Ehrlichia spp, which do infect ruminants (E. bovis, E. ovina, and E. phagocytophila) did not react with MAP1-B. Antibody titers to C. rumi nantium in sera from experimentally infected cattle, goats, and sheep were detectable for 50 to 200 days postinfection. Further validation o f the recombinant MAP1-B-based ELISA was done with sera obtained from sheep raised in heartwater-free areas in Zimbabwe and from several Car ibbean islands. A total of 159 of 169 samples which were considered to be false positive by immunoblotting or indirect ELISA did not react w ith MAP1-B, In conclusion, recombinant MAP1-B may be a suitable antige n for a sensitive serological test for cowdriosis, with dramatically i mproved specificity.