Antibodies to an epitope from the Cha human autoantigen are markers of Chagas' disease

Abstract

Chagas' disease is a prevalent disease in South America that is thought to have an autoimmune etiology. We previously identified human Cha as a new autoantigen recognized by chagasic sera. Those sera recognized an epitope spanning amino acids 120 to 129 of Cha, named R3. In the present study we have used the synthetic R3 peptide for the detection of serum immunoglobulin G antibodies from patients at different stages of Chagas' disease, including a therapeutically treated group. The immunoreactivity with R3 by enzyme-linked immunosorbent assay (ELISA) showed 92.4% sensitivity and 100% specificity for Chagas' disease sera. This sensitivity and specificity were higher than for any other autoantigen described to date. No anti-R3 antibodies were detected in sera from Leishmania-infected or idiopathic dilated cardiomyopathy patients or healthy controls from the same areas. Moreover, anti-R3 antibody reactivity detected by ELISA correlated with conventional serological tests as indirect immunofluorescence and ELISA assays with Trypanosoma cruzi extracts and other diagnostic tests as indirect hemagglutination. The levels of anti-R3 antibodies increased with progression and symptomatology of Chagas' disease. More interestingly, a statistically significant fall in anti-R3 antibody titer was observed in patients treated with antiparasitic drugs. Those results suggest that the presence of anti-R3 antibodies is a highly specific marker of Chagas' disease and that R3 ELISA could be helpful in the diagnosis and monitoring of this disease.

FIG. 1

Reactivity of sera from chronic chagasic patients against the R3 peptide. The reactivity of 79 sera with positive serology for Chagas' was tested in ELISA with the R3 peptide. Ten sera from individuals infected with Leishmania infantum, 6 sera from patients suffering (IDC, and samples from 10 healthy individuals living in an area were Chagas' disease is endemic (EHS) were used as controls. Reactivity is expressed as OD at 450 nm. The horizontal dashed line represents the cutoff value.

FIG. 2

Correlation of anti-R3 IgG antibodies with symptomatology and treatment. (A) Correlation of IgG antibody levels against R3 and S1 peptides with Chagas' disease progression. Sera were grouped as control (EHS), asymptomatic, and symptomatic and plotted against the mean OD₄₉₀ ± standard deviation (SD) values obtained with the R3 (solid circles) and S1 (solid triangles) peptide ELISA. The number of individual sera tested in each group is indicated. Sera were assayed at 1:100 dilution. (B) Correlation of IgG antibody levels against R3 and S1 peptides with the treatment of asymptomatic chagasic patients. The results obtained with sera from untreated and treated chagasic patients were plotted against the mean OD₄₉₀ ± SD values obtained with the R3 (solid circles) and S1 (solid triangles) peptide ELISA. The number of individuals in each group is indicated.

FIG. 3

Correlation of IgG antibodies against R3 and S1 peptides by ELISA with other serological tests (IIF, IHA, and parasite ELISA). Individual sera were grouped based on the results (titer inversa) of each serological test and plotted against the mean of OD₄₉₀ ± SD values obtained with the R3 (solid circles) and S1 (solid triangles) peptide ELISA. The number of serum samples in each group is indicated. (A) Correlation with IIF. A value of 0 is negative and indicates titers <1:32. (B) Correlation with IHA. A value of 0 is negative (titer < 1:16). (C) Correlation with a commercial ELISA using crude T. cruzi extract at 1:100 serum dilution.