DEVELOPMENT AND EVALUATION OF A RECOMBINANT ANTIGEN, MONOCLONAL ANTIBODY-BASED COMPETITIVE ELISA FOR HEARTWATER SERODIAGNOSIS

Cowdria ruminantium is the etiologic agent of heartwater, a tick-trans mitted foreign animal disease with considerable potential for entrance into the USA. A competitive enzyme-linked immunosorbent assay (cELISA ) was developed to detect serologic responses to C. ruminantium infect ion. The cELISA utilized a recombinant form of the C. ruminantium majo r antigenic protein (MAP-1) as the antigen and an anti-MAP-1 monoclona l antibody as the competing indicator reagent. Experimental antisera t o C. ruminantium and a wide variety of related ehrlichial organisms we re used to evaluate cELISA reactivity. Only sera against C. ruminantiu m, Ehrlichia canis, E. chaffeensis, and a recently discovered cervine ehrlichia-like organism reacted positively in the cELISA. Specificity of the cELISA was greater than or equal to 99.5% in a survey of 1,774 southeastern US and Puerto Rican slaughter cattle sera but was only 85 % in a group of 79 hunter-killed white-tailed deer (Odocoileus virgini anus) from the southeastern USA. Reference true-positive and cELISA fa lse-positive sera were further analyzed by end point titrations using the cELISA and by indirect fluorescent antibody (IFA) tests for reacti vity with C. ruminantium, E. canis, and E. chaffeensis antigens. True heartwater-positive sera were significantly more reactive using the cE LISA and C. ruminantium IFA procedures (P < 0.05), whereas false-posit ive sera were significantly more reactive with the antigens used in th e E. chaffeensis IFA procedure (P < 0.05). A group of sera from 210 fi eld-origin ruminants residing on known or potentially heartwater-endem ic Caribbean islands revealed a substantial (12.4%) prevalence of cELI SA-positive specimens. The cELISA is a relatively specific serodiagnos tic test for heartwater in cattle and could be used to monitor for pos sible introduction of the disease into the USA. The cELISA may also be an excellent tool for monitoring the success of an ongoing Caribbean Amblyomma tick eradication program designed to eliminate the biologica l vector responsible for the perpetuation and spread of this dangerous foreign animal disease.