Development of an Indirect ELISA Based on a Recombinant Chimeric Protein for the Detection of Antibodies against Bovine Babesiosis

The current method for Babesia spp. serodiagnosis based on a crude merozoite antigen is a complex and time-consuming procedure. An indirect enzyme-linked immunosorbent assay (iELISA) based on a recombinant multi-antigen of Babesia bovis (rMABbO) was developed for detection of antibodies in bovines suspected of infection with this parasite. The multi-antigen comprises gene fragments of three previously characterized B. bovis antigens: MSA-2c, RAP-1 and the Heat Shock protein 20 that are well-conserved among geographically distant strains. The cutoff value for the new rMABbo-iELISA was determined using 75 known—positive and 300 known—negative bovine sera previously tested for antibodies to B. bovis by the gold-standard ELISA which uses a merozoite lysate. A cutoff value of ≥35% was determined in these samples by receiver operator characteristic (ROC) curve analysis, showing a sensitivity of 95.9% and a specificity of 94.3%. The rMABbo-iELISA was further tested in a blind trial using an additional set of 263 field bovine sera from enzootic and tick-free regions of Argentina. Results showed a good agreement with the gold standard test with a Cohen’s kappa value of 0.76. Finally, the prevalence of bovine babesiosis in different tick enzootic regions of Argentina was analyzed where seropositivity values among 68–80% were obtained. A certain level of cross reaction was observed when samples from B. bigemina infected cattle were analyzed with the new test, which can be attributed to shared epitopes between 2 of the 3 antigens. This new rMABbo-iELISA could be considered a simpler alternative to detect anti Babesia spp. antibodies and appears to be well suited to perform epidemiological surveys at the herd level in regions where ticks are present

Instituto de Biotecnología

Fil: Jaramillo Ortiz, Jose Manuel. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina

Fil: Montenegro, Valeria Noely. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina.

Fil: De la Fourniere, Sofía Ana María. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina

Fil: Sarmiento, Nestor Fabian. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Mercedes; Argentina

Fil: Farber, Marisa Diana. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina

Fil: Wilkowsky, Silvina Elizabeth. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina