Comparison of a commercial ELISA, an in-house indirect ELISA, and a dot-ELISA developed for the serodetection of Toxoplasma gondii antibodies in farm animals.

Abstract

Toxoplasma gondii is a widespread intracellular protozoan that can infect humans and animals. We isolated T. gondii strains from sheep, goats, cattle, buffaloes, and camels to develop and evaluate a modified in-house dot-ELISA for the detection of Toxoplasma antibodies in farm animals, and compared the results with a commercial ELISA (IDvet; gold standard). Animal tissue samples (n = 430) were examined microscopically, and infected tissues were bioassayed in mice as a viability test. Egyptian Toxoplasma strains were isolated from sheep, cattle, and camels and identified via PCR using the B1 gene (GenBank OR837022.1, OR837021.1, OR837020.1 from sheep, cattle, and camels, respectively). A T. gondii tachyzoite antigen from a sheep strain had the highest potential for the detection of specific T. gondii antibodies. We characterized this antigen using SDS-PAGE and separated it into 10 polypeptides of 96-12 kDa. Our modified in-house dot-ELISA detected T. gondii seropositivity in 172 of 430 (40%) farm animals with a sensitivity of 96.6% and specificity of 100%. The results of our dot-ELISA were confirmed in comparison with those of our indirect ELISA and the commercial ELISA. In a western blot, a predominant immunogenic reactive antigen band of 65 kDa was detected in T. gondii-positive sera of sheep, cattle, buffaloes, and camels; no cross-reaction occurred with antibodies to other parasitic infections or samples from healthy controls. Our modified in-house dot-ELISA is a rapid and simple test that showed promise for the detection of Toxoplasma antibodies in farm animals.