Detection of antibodies to Toxoplasma gondii in domesticated ruminants by recombinant truncated SAG2 enzyme-linked immunosorbent assay

Abstract

An antibody detection recombinant enzyme-linked immunosorbent assay (ELISA) specific for Toxoplasma gondii was laboratory standardized using recombinant truncated surface antigen 2 (SAG2) protein of T. gondii. A 483-bp sequence coding for truncated tachyzoite stage-specific SAG2 protein was amplified and ligated in pPROExHT-b expression vector to transform Escherichia coli DH5α cells. A high-level expression of the histidine-tagged fusion protein was obtained after 8 h of incubation. The recombinant protein was affinity purified using Ni-NTA agarose column and characterized by SDS-PAGE and Western blot analysis. Subsequently, the diagnostic potential of the recombinant protein was assessed with 168 field sera samples from sheep, goats and cattle. Among the small ruminants, 50% (n = 60) sheep sera samples and 41.26% (n = 63) goat samples were detected positive for T. gondii-specific antibodies. As far as seroprevalence of toxoplasmosis in cattle is concerned, 64.44% (n = 45) of sera samples assayed were found to be positive. When compared to indirect fluorescent antibody test (IFAT), the sensitivity of the recombinant truncated SAG2 antigen-based ELISA (rec-SAG2-ELISA) ranged from 81.25 to 87.10% while the specificity was 85.71 to 91.43% with substantial agreement between the tests.