Diagnosis and control of
dourine is strongly based on serological evidence, but knowledge of the humoral response of horses during
infection is limited. In this study we developed a chemiluminescent immunoblotting (cIB) assay to characterise the Trypanosoma equiperdum
antigen pattern recognised by IgGs from naturally or experimentally
dourine-infected horses and analyse the kinetics of
IgG humoral response following the
infection. One compounding factor is that sera from uninfected animals often cross-react with T. equiperdum
antigens. Development of the cIB assay was based on the hypothesis that serum IgGs from healthy and infected animals recognise different T. equiperdum
antigen patterns. We used sera from 8 naturally infected horses which had recovered from Italian outbreaks and 2 experimentally infected mares. In addition, sera from 10 healthy control animals, eight of which were CFT positive but IFA negative for
dourine, were collected from disease free regions. Sera were compared by the
complement fixation test (CFT), indirect immune fluorescence (IFA) and the cIB assay. cIB analysis revealed that IgGs from infected horses, in contrast to IgGs from healthy horses, specifically recognise a T. equiperdum antigenic profile with low molecular weight bands ranging between 16 and 35 kDa. A time course experiment indicated that IgGs specific for the 16-35 kDa parasite
protein fraction appear 17 days post-
infection. The cIB assay confirmed all ten infected animals as positive and all controls as negative. This study demonstrated that analysis of IgGs by cIB can provide clear confirmation of trypanosome
infection in horses, suggesting that this technique can be applied as a confirmatory serological test for
dourine infection.