A multinested PCR for detection of the equine piroplasmids Babesia caballi and Theileria equi.

Abstract

Two haemoparasites, Theileria equi and Babesia caballi, cause equine piroplasmosis (EP), one of the most prevalent tick-borne diseases in horses. The main aim of the present study was to develop and evaluate a multinested PCR (mn-PCR) for simultaneous detection of the equine piroplasmids T. equi and B. caballi, by amplification of five genetic markers (18S rRNA, β-tubulin, cytB, EMA-1 and RAP-1). This novel assay detected a high prevalence of equine piroplasmids in 235 horse blood samples collected in Castilla-León and Extremadura, Spain. The overall prevalence of infection with equine piroplasmids by mn-PCR was 72.8% (171/235), with 66.0% (155/235) of the animals positive for T. equi and 29.4% (69/235) positive for B. caballi. The seroprevalence obtained by cELISA for the same set of samples was lower than the infection prevalence recorded by mn-PCR, for either of the two equine piroplasmids (62.6%) as well as for T. equi alone (61.7%) or B. caballi alone (3.8%). There was high agreement among the mn-PCR and cELISA assays for diagnosis of EP caused by T. equi (κ = 0.83) but not for B. caballi (κ = 0.06). A phylogenetic analysis based on the RAP-1 gene of B. caballi showed that the strains from Spain clustered with those from Israel.