Diverse gene targets for Trypanosoma detection in equines ‒ a comparative analysis

Zoonotic parasitic diseases, particularly those caused by Trypanosoma spp., pose a significant global health challenge, resulting in substantial economic losses. The infection caused by this parasite in equines is complex, affecting weight gain and the working capacity of animal. The present study was designed to estimate the occurrence of Trypanosoma in equines from the Rahim Yar Khan District, Southern Punjab, Pakistan, by amplifying the variable surface glycoprotein gene (RoTat 1.2 VSG gene) and the nuclear gene (18S rRNA) of the parasite. To ascertain the prevalence of Trypanosoma, a total of 384 blood samples, determined by calculation of the sample size formula, were collected from equines, i.e., horses, mules and donkeys. Microscopic detection of the parasite was performed using the microhematocrit method. For molecular detection and characterization of Trypanosoma evansi, DNA was extracted from blood samples using commercially available kits, and PCR was performed. Data on the associated risk factors were collected through a pre-designed questionnaire. The overall prevalence of Trypanosoma was 7.12 and 21.44% using microscopic and molecular techniques, respectively. Regarding the associated risk factors, the infection was found to be significantly more prevalent in females compared to male animals, in younger animals with low body weight compared to older animals with high body weight, and in donkeys compared to horses and mules. Among other factors, the prevalence of Trypanosoma was significantly higher in pregnant females, non-dewormed, symptomatic, and emaciated animals. The phylogenetic grouping and sequence analysis revealed that the sequences of both genes appeared in the T. evansi clade, reported in various hosts in different countries. In conclusion, the nuclear gene, that is, the 18S rRNA gene was found to be more suitable to detect Trypanosoma compared to the VSG gene, that is, the RoTat 1.2 VSG gene.