Analysis of Trypanosoma equiperdum Recombinant Proteins for the Serological Diagnosis of Dourine

Simple Summary: Dourine is a contagious disease prevalent in equids that can lead to serious health issues in affected animals, including reproductive problems and nervous system disorders. Stamping out policies implemented during the 20th century has been crucial in reducing the prevalence of dourine. To maintain control and prevent the international spread of dourine, stringent measures are necessary when it comes to the movement of equids across borders. The challenges associated with diagnosing this disease, including its coexistence with closely related trypanosomes, highlight the critical need for accurate and specific diagnostic tools. In this study, three proteins, namely peptidyl-prolyl cis-trans isomerase (A0A1G4I8N3), GrpE protein homolog (A0A1G4I464), and transport protein particle (TRAPP) component, putative (A0A1G4I740), were expressed as recombinant proteins. In a previous study based on bioinformatics analyses, these proteins were unique to T. equiperdum and did not exhibit matches or cross-reactivities with other Trypanosoma species protein; moreover, these proteins were found to be immunogenic because they exhibit the presence of potential B-cell epitopes. The three proteins were screened for reactivity to a small panel of horse sera using indirect enzyme-linked immunosorbent assay and an immunoblotting test. Protein A0A1G4I8N3, giving the best results, will be tested with a larger amount of sera to more comprehensively evaluate its potential use in the diagnosis of dourine. The significance of Trypanosoma equiperdum as the causative agent of dourine cannot be understated, especially given its high mortality rate among equids. International movement of equids should be subject to thorough health checks and screenings to ensure that animals are not infected with Trypanosoma equiperdum. This involves the implementation of quarantine protocols, testing procedures, and the issuance of health certificates to certify the health status of the animals. Three proteins, the peptidyl-prolyl cis-trans isomerase (A0A1G4I8N3), the GrpE protein homolog (A0A1G4I464) and the transport protein particle (TRAPP) component, putative (A0A1G4I740) (UniProt accession numbers SCU68469.1, SCU66661.1 and SCU67727.1), were identified as unique to T. equiperdum by bioinformatics analysis. The proteins were expressed as recombinant proteins and tested using an indirect ELISA and immunoblotting test with a panel of horse positive and negative sera for dourine. The diagnostic sensitivity, specificity and accuracy of the i-ELISAs were 86.7%, 53.8% and 59.0% for A0A1G4I8N3; 53.3%, 58.7% and 57.9% for A0A1G4I464; and 73.3%, 65.0% and 66.3% for A0A1G4I740, respectively, while the diagnostic sensitivity, specificity and accuracy of immunoblotting were 86.7%, 92.5% and 91.6% for A0A1G4I8N3; 46.7%, 81.3% and 75.8% for A0A1G4I464; and 80.0%, 63.8% and 66.3% for A0A1G4I740. Among the three proteins evaluated in the present work, A0A1G4I8N3 provided the best results when tested by immunoblotting; diagnostic application of this protein should be further investigated using a greater number of positive and negative sera.