SERODIAGNOSIS OF BOVINE TRYPANOSOMOSIS CAUSED BY NON-TSETSETRANSMITTED TRYPANOSOMA (DUTTONELLA) VIVAX PARASITES USING THESOLUBLE FORM OF A TRYPANOZOON VARIANT SURFACE GLYCOPROTEIN ANTIGEN

Abstract

Previous studies have shown that a 64-kDa antigen (p64) that was purified from the VenezuelanTeAp-N/D1 isolate of Trypanosoma (Trypanozoon) equiperdum corresponds to the soluble form of itspredominant variant surface glycoprotein (VSG), and exhibited cross-reactivity with Trypanosoma (Dut-tonella) vivax. The course of experimental acute infections of bovines with T. vivax were followed bymeasuring whole anti-p64 antibodies and specific anti-p64 IgG and IgM antibodies in animal sera by indi-rect enzyme-linked immunosorbent assay (ELISA). The value of p64 to diagnose bovine trypanosomosiswas also examined using 350 sera from healthy and T. vivax-infected cows living in a trypanosomosis-endemic and enzootic stable area, and 48 sera obtained during a trypanosomosis outbreak. Serologicalassays showed that ∼70–80% of the infected sera contained anti-p64 antibodies, based on the compar-ative immunodetection of the T. equiperdum clarified antigenic fraction used as a reference test. In theabsence of a gold standard, Bayesian analysis for multiple testing estimated a sensitivity and specificityof 71.6% and 98.8%, respectively, for the indirect ELISA using p64 as antigen. An apparent prevalence of37.7% for bovine trypanosomosis infection was also estimated with a Bayesian approach when the p64ELISA test was used. Employing blood from acute infected cows, the indirect ELISA response against p64was contrasted with the microhematocrit centrifuge method and analyses by polymerase chain reaction(PCR) using specific primers targeting the inter-specific length variation of the internal transcribed spacer1 region of the 18S ribosomal gene. The efficiency of p64 for the detection of anti-trypanosome antibodiesin acute infected bovines was also corroborated serologically by comparing its response to that of theIndonesian Trypanosoma evansi Rode Trypanozoon antigen type (RoTat) 1.2 VSG, which possesses highspecificity and sensitivity. As expected, PCR was the best method to detect parasites and diagnose bovinetrypanosomosis; however, a substantial level of concordance (Cohen’s = 0.667) was obtained whenserological tests using p64 and RoTat 1.2 VSG were compared. Additionally, an agglutination assay wasdesigned using p64 covalently coupled to carboxylate-modified latex microparticles, which was provenhere to be suitable for a fast qualitative diagnosis of bovine trypanosomosis.