Validation of an indirect ELISA using recombinant proteins as antigen to identify animals exposed toBabesia bigemina

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Abstract
The objective of this study was to instrument a serological assay for the epidemiological diagnosis of bovine babesiosis in Mexico, using the Babesia bigemina recombinant protein RAP‐1 (rRAP‐1α) as antigen. rRAP‐1α, r12d3 and rGP45 were the three recombinant antigens initially tested. Based on the highest titres obtained in the indirect ELISA (iELISA) with the positive control serum, using similar antigen concentrations, rRAP‐1α was selected for further use. The diagnostic sensitivity and specificity rates estimated for the iELISA with rRAP‐1α as antigen were 89.9% and 86.5%, respectively, while for the Indirect Fluorescent Antibody Test (IFAT), the gold standard assay, the sensitivity was 86.66% and the specificity was 95%. The ĸ agreement value determined was 0.52, indicating a moderate agreement between the iELISA and IFAT assays. The instrumented iELISA with rRAP‐1α as antigen shows an excellent specificity rate and an acceptable sensitivity that allows for the detection of antibodies to B. bigemina in cattle naturally exposed to the vector tick Rhipicephalus microplus. By using the iELISA‐rRAP‐1α, along with an iELISA with recombinant Merozoite Surface Antigen (rMSA‐1) for antibody determination against Babesia bovis in the serum samples collected from cattle at ‘La Posta’ experimental station in Mexico, a seroprevalence of 20.3% was estimated for B. bigemina and 19.4% for B. bovis, while 36.89% of samples were positive for both Babesia species. The iELISA test promises to be a safe and low‐cost type of diagnosis available to cattle producers in Mexico and would facilitate the definition of herd immunity status to implement measures of control adapted for the prevention of bovine babesiosis outbreaks.

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