High-throughput mutational analysis of Trypanosoma cruzi antigenic epitopes reveal consistent conservation of key residues across human Chagas Disease populations.

NOLAN M; ALTCHEH J; AGÜERO F; BRACCO L; TORRICO F; KESPER JR N; RAMSEY J; MARCIPAR I; VILLAR JC

Buenos Aires

Simposio; XIX Simposio Internacional sobre Enfermedades Desatendidas; 2019

Fundación Mundo Sano

Chagas Disease is a major health problem in America for which no vaccine forlarge-scale public health interventions are yet available. Accurate diagnosis isessential for the identification of infection and ecoepidemiological surveillance.Diagnosis is routinely performed using serological methods, which require wellcharacterized antigens. Although available tests give satisfactory results in mostcases, discordant results remain as a possible cause of undetected infectedpatients. We have previously conducted a large-scale screening of T. cruzi linearB-cell epitopes using high-density peptide arrays, leading to the development ofa new proof-of-principle multiepitope diagnostic test that has excellent diagnosticperformance 1 . However, further improvement is possible. Understanding whichresidues in an epitope are important for antibody binding can lead to improvedreagents. To further characterize known antigens, we performed Alanine scansof 649 different proteins (881 antibody-binding peaks/epitopes, represented by2,913 overlapping peptides). This experiment was based on replacing eachamino acid residue in each peptide for an Alanine (or a Glycine if the originalresidue was itself an Alanine), and assessing the impact on reactivity of eachmodified epitope. Using this peptide array design we have assayed 45,492peptide variants against 108 Chagas Disease serum samples (from 6 differentcountries). We developed an algorithm to integrate and analyze the effect ofthese epitope mutations, and to visualize key residues for each antigen andsample. As a result, we identified precise residue positions in epitopes that playa fundamental role in the seroreactivity. In summary, we observe an average of~6 key residues per epitope. As an example Ag2/CA-2, a known antigen,displays the same 7 key residues in all reactive sera. In contrast SAPA, Ag36,and B12 display different degrees of reactivity conservation of their key residues.This variable responses for different antigens, can be used in the design ofimproved antigens for diagnostic applications.1. Mucci, J. et al. Next-generation ELISA diagnostic assay for Chagas Diseasebased on the combination of short peptidic epitopes. PLoS Negl. Trop. Dis. 11,e0005972 (2017).