Next-generation ELISA diagnostic assay for Chagas Disease based on the combination of short peptidic epitopes

Chagas Disease, caused by the protozoan Trypanosoma cruzi, is a major health and economic problem in Latin America for which no vaccine or appropriate drugs for large-scale public health interventions are yet available. Accurate diagnosis is essential for the early identification and follow up of vector-borne cases and to prevent transmission of the disease by way of blood transfusions and organ transplantation. Diagnosis is routinely performed using serological methods, some of which require the production of parasite lysates, parasite antigenic fractions or purified recombinant antigens. Although available serological tests give satisfactory results, the production of reliable reagents remains laborious and expensive. Short peptides spanning linear B-cell epitopes have proven ideal serodiagnostic reagents in a wide range of diseases. Recently, we have conducted a large-scale screening of T. cruzi linear B-cell epitopes using high-density peptide chips, leading to the identification of several hundred novel sequence signatures associated to chronic Chagas Disease. Here, we performed a serological assessment of 27 selected epitopes and of their use in a novel multipeptide-based diagnostic method. A combination of 7 of these peptides were finally evaluated in ELISA format against a panel of 199 sera samples (Chagas-positive and negative, including sera from Leishmaniasis-positive subjects). The multipeptide formulation displayed a high diagnostic performance, with a sensitivity of 96.3% and a specificity of 99.15%. Therefore, the use of synthetic peptides as diagnostic tools are an attractive alternative in Chagas’ disease diagnosis.
Author summary Chagas disease, caused by the parasite Trypanosoma cruzi, is a life-long and debilitating illness of major significance throughout Latin America, and an emergent threat to global public health. Diagnostic tests are key tools to support disease surveillance, and to ultimately help stop transmission of the parasite. However currently available diagnostic methods have several limitations. Identification of novel biomarkers with improved diagnostic characteristics is a main priority. Recently, we conducted a large-scale screening looking for new T. cruzi antigens using short peptides displayed on a solid support at high-density. This led to the identification of several hundred novel antigenic epitopes. In this work we validated the serodiagnostic performance of 27 of these against an extended panel of human serum samples. Based on this analysis, we developed a proof-of-principle multiplex diagnostic kit by combining different validated reactive peptides. Overall, our data support the applicability of high-density peptide microarrays for the rapid identification and mapping epitopes that could be readily translated into novel and useful tools for diagnosis of Chagas disease.