The use of IgG antibodies in conventional and non-conventional immunodiagnostic tests for early prognosis after treatment of Chagas disease

Abstract

Treatment success of chronically infected Chagas disease patients is laborious and a positive prognosis often is made only after repetitive serological and/or parasitological examinations with continuous negative results. Recently, we have developed a non-conventional flow-cytometric method in order to detect immunoglobulin G antibodies against live trypomastigote forms of Trypanosoma cruzi and showed its usefulness in the prognosis of treatment success. In the present study, we investigated the performance of flow-cytometric anti-live trypomastigote IgG antibodies (FC-ALTA) and flow-cytometric anti-fixed epimastigote IgG antibodies (FC-AFEA), as well as conventional serological methods, for early monitoring of benznidazole treated Chagas disease patients, e.g. 5 years after treatment. The analysis of individual FC-ALTA reactivity along the titration curve before and after treatment, we were able to show, that between 4% and 13% of treated patients under evaluation presented with reduced serological reactivity and segregated from the other patient groups. Similar results were obtained with semi-quantitative, conventional indirect hemagglutination or indirect immunofluorescence. Our data therefore suggest that the combined use of conventional and non-conventional serological methods could provide more suitable cure criteria in early post-therapeutic prognosis of Chagas disease.

Introduction

Human infection with Trypanosoma cruzi is endemic mainly in Latin America, affecting approximately eight million people in South and Central America and a further 100 million people are considered at risk (Weekly, 2007). However, with population movements from endemic to non-endemic countries there are estimates of more than 300,000 chagasic patients in the USA, > 5500 in Canada, > 80,000 in Europe and in the western Pacific region, turning Chagas disease into a public health problem outside the commonly considered countries and regions (Coura and Viñas, 2010).

The treatment of Chagas disease in both acute and recent chronic infections may lead to cure or prevent pathological outcome in the later stages of disease (Ferreira, 1990, Urbina, 1999, Garcia et al., 2005). The indication of the etiological treatment in the chronic phase is still controversial because most treated patients continue to have positive conventional serology, even though their hemocultures become less frequently positive than those of the untreated, chronically infected patients (Galvão et al., 1993). Therefore, one of the major challenges has been the establishment of laboratory tools to assess treatment effectiveness, considering that conventional approaches usually remain positive several years after chemotherapy and therefore fail when used as a cure criterion. During the acute phase of infection, diagnosis is done by detection of the parasites directly in the blood or in artificial hemocultures. During the chronic phase, the direct detection of parasites by xenodiagnosis or hemoculture is difficult and time consuming and the diagnosis is usually done by detecting circulating antibodies with conventional or non-conventional methods (Portela-Lindoso and Shikanai-Yasuda, 2003, Lunardelli et al., 2007). Apart from the usefulness for diagnosis, it has been recently shown that the determination of such antibodies by non-conventional flow cytometry methods is a promising tool to monitor treatment efficacy in Chagas disease patients (Martins-Filho et al., 1995, Martins-Filho et al., 2002, Vitelli-Avelar et al., 2007).

Here, we extended our previous studies and compared non-conventional flow cytometry assays and conventional assays (indirect hemagglutination [IHA], indirect immunofluorescence [IIF], enzyme-linked immunosorbent assay [ELISA], and particle gel agglutination [PaGIA]) for their usefulness for prognosis and treatment efficacy in patients 5 years after chemotherapy against Chagas disease.