Unraveling cryptic epizootiology of equid trypanosomosis in Punjab state of India by parasitological and sero-molecular techniques

To unravel equid trypanosomosis caused by Trypanosoma evansi in Punjab state of India, a cross sectional study was designed by utilizing parasitological and sero-molecular tools with objective to assess the prevalence of T. evansi in association with various risk factors in all agroclimatic zones of Punjab state of India. Parasitological Romanowksy stained thin blood smears (RSTBS) to detect patent infection, molecular techniques polymerase chain reaction I (PCR I; TBR 1/2 primers; targeting minichromosomal satellite DNA of T. evansi), polymerase chain reaction II (PCR II; TR 3/4 primers; targeting variable surface glycoprotein region DNA of T. evansi) & LAMP (Loop mediated isothermal amplification) assay to detect latent infection and serological assays card agglutination test (CATT/T. evansi) & ELISA (Enzyme linked immunosorbent assay) to detect exposure status of trypanosomosis were utilized in the present study. A total 429 equid blood and serum samples from all the five agroclimatic zones of Punjab state tested by these techniques showed a prevalence of 1.39% (CL: 0–15.28) by RSTBS, 6.52% (10.94–45.09) by both TBR 1/2 PCR and LAMP assay, 5.82% (11.57–38.42) by TR 3/4 PCR, 15.15% (36.57–135.42) with CATT/T. evansi and 22.84% (17.77–840.22) with ELISA. Interpretation of various risk factors revealed that the donkey/mules population (RR = 5.46, 95% [CI] = 0.15–15.56) was found to be at higher risk of T. evansi infection predominantly at ‘unorganized’ farms (RR = 4.06, 95% [CI] = 0.12–4.51). Animal used for commercial purposes (RR = 3.25, 95% [CI] = 0.06–7.42), rearing of equids with other domestic animals (RR = 2.36, 95% [CI] = 0.10–17.11) and farms without application of fly repellant/insecticides/net (RR = 3.68, 95% [CI] = 0.08-5.94) made them more prone to the disease. This comprehensive report utilizing the classical, serological and molecular diagnostic tools for epidemiology of T. evansi establishes the endemic stability of this infection in all agro climatic zones of Punjab with LAMP assay to be a promisingly sensitive and specific technique for the diagnosis of T. evansi under isothermal conditions in field situations.