Diagnostic application of recombinant equine merozoite surface antigen-1 in elisa for detection of Theileria equi specific antibodies.

Abstract

Theileria equi merozoite surface antigens have been an important candidate for development of diagnostics. We developed ELISA based on EMA-1 recombinant antigen, so as to widen our diagnostic confidence in detection of antibodies against T. equi in sero-surveillance studies. The 547 bp EMA-1 gene fragment encoding high hydrophilic antigenic region was expressed with glutathione-S-transferase tag in prokaryotic system and purified protein (43 kDa) was used for development of ELISA (EMA-1t/ELISA). The EMA-1t/ELISA clearly differentiated T. equi-infected from Babesia caballi-infected horse sera or normal horse sera. The results of the study were validated with previously developed (EMA-2)ELISA on serum samples of known T. equi infection status and a very high correlation (0.93) was recorded between the relative percent positivity (RPP) values. Further diagnostic sensitivity of EMA-1t/ELISA was 0.92 while specificity was 1.0, indicating its suitability for sero-epidemiological studies. This assay was applied on serum samples (n = 240) collected from field horses in northern part of India. High sero-prevalence of T. equi antibodies were diagnosed in serum samples collected from Haryana state (74%) and Uttarakhand state (36.31%). Results of this study suggested that the 43 kDa EMA-1 expressed protein could be a reliable immunodiagnostic antigen in ELISA for T. equi sero-prevalence studies.