Research Article: Trypanosoma cruzi Detection in Colombian Patients with a Diagnosis of Esophageal Achalasia

Date Published: March 5, 2018

Publisher: The American Society of Tropical Medicine and Hygiene

Author(s): Santiago Panesso-Gómez, Paula Pavia, Iván Enrique Rodríguez-Mantilla, Paola Lasso, Luis A. Orozco, Adriana Cuellar, Concepción J. Puerta, Belén Mendoza de Molano, John M. González.


Achalasia is a motility disorder of the esophagus that might be secondary to a chronic Trypanosoma cruzi infection. Several studies have investigated esophageal achalasia in patients with Chagas disease (CD) in Latin America, but no related studies have been performed in Colombia. The goals of the present study were to determine the presence of anti-T. cruzi antibodies in patients with esophageal achalasia who visited a referral hospital in Bogotá, Colombia, and to detect the presence of the parasite and its discrete typing units (DTUs). This cross-sectional study was conducted in adult patients (18–65 years old) who were previously diagnosed with esophageal achalasia and from whom blood was drawn to assess antibodies against T. cruzi using four different serological tests. Trypanosoma cruzi DNA was detected by conventional polymerase chain reaction (cPCR) and quantitative polymerase chain reaction (qPCR). In total, 38 patients, with an average age of 46.6 years (standard deviation of ±16.2) and comprising 16 men and 22 women, were enrolled. Five (13.15%) patients were found to be positive for anti-T. cruzi antibodies by indirect immunofluorescence assay (IFA), and two patients who were negative according to IFA were reactive by both enzyme-linked immunosorbent assay and immunoblot (5.3%). Parasite DNA was detected in two of these seven patients by cPCR and in one of these by qPCR. The parasite DTU obtained was TcI. In summary, this study identified T. cruzi in Colombian patients with esophageal achalasia, indicating that digestive compromise could also be present in patients with chronic CD.

Partial Text

Chagas disease (CD), which is caused by the intracellular protozoan Trypanosoma cruzi, is a major endemic parasitic infection in Latin America. In Colombia, it is estimated that 437,960 individuals are infected and that nearly 11% of the population is at risk of contracting the disease. However, a recent study indicated that only 1.2% of this population has screening coverage.1 Although the vector transmission of the parasite is confined to the Latin American continent and certain regions of North America, human migration has spread congenital and transfusion infections to Europe and Asia. Symptomatic chronic infection occurs in approximately 30% of infected patients, in whom the cardiac and digestive systems are the most frequently affected.2 The pathogenesis of tissue damage during chronic infection is unclear; however, parasite persistence, cellular tropism, the parasite genotype, and a dysfunctional host immune response have been suggested to participate in the mechanisms that induce damage.3–6 During a chronic symptomatic infection, approximately 90% of patients develop cardiomyopathy, 10% develop enteropathy, and a very low percentage of patients exhibit mixed compromise.2 The currently identified parasite genotypes, or discrete typing units (DTUs), include seven variants, termed TcI to TcVI and TcBat. Although certain studies have attempted to associate a parasite’s DTU with tissue tropism and clinical presentation, these associations are not completely understood.5

In total, 81 patients with a diagnosis of esophageal achalasia were registered in the EMRs through 2015 and 38 agreed to participate in the study. In all, 22 women and 16 men without cardiac compromise and with an average age of 46.6 (SD of ±16.2) years were enrolled. The majority of the patients lived in Bogotá (79%), the capital of Colombia, where T. cruzi is not transmitted. Nonetheless, more than half of them were born outside Bogotá (58%), as shown in Table 1. Among the risk factors for CD, 57.9% of the patients had no knowledge of the vector. In addition, most patients had resided in brick houses during their childhood (84.2%), and 100% still resided in this type of housing. Only one individual analyzed had a history of CD in the family (2.6%), and 10 had a history of blood transfusion (26.3%) (Table 2). The most prevalent symptom was difficulty swallowing or dysphagia (97.4%); the frequencies of the symptoms in this cohort of patients are shown in Supplemental Figure 1. The observed signs and symptoms show distributions similar to those described for patients with esophageal achalasia due to other causes.7

Studies of patients with chronic CD have focused on cardiomyopathy, which is the most common complication observed during chronic infection because of its high mortality and socioeconomic impact.33 However, several studies from Brazil9,10 and from immigrant populations in Spain and Italy11–13 have examined the gastrointestinal complications of chronic T. cruzi infection. Most of these studies were conducted in patients who had been previously diagnosed with CD and were later examined for symptoms or signs of digestive compromise. The purpose of the present study was to determine the presence of anti-T. cruzi antibodies and parasites in individuals with established esophageal disease. Bogotá, the capital of Colombia, is a city at a high altitude (8,660 ft above sea level) where T. cruzi is not transmitted, but it is surrounded by municipalities and states located below 6,580 ft where CD is endemic.34,35 A high percentage of the population migrates from these places (mainly from rural areas) or travels in search of medical attention.36,37 Considering that the majority of individuals analyzed are located in Bogotá, they likely were not continuously exposed to the parasite.34 Risk factors for T. cruzi transmission were uncommon in our cohort of 38 patients, although they were more frequent in seroreactive patients. Indeed, five of seven seropositive individuals analyzed were born in states with high T. cruzi transmission (Boyacá, Cundinamarca, and North Santander), where the primary circulating DTUs are TcI (80.7%) and TcII (7.2%).35 Although the prevalence of digestive system compromise in CD varies from 7% to 15%,9–12 tests for antibodies against T. cruzi are not normally part of the routine exams performed in Colombia on patients with esophageal motility alterations. Interestingly, one study in Mexico showed that among 28 asymptomatic individuals with serological evidence of T. cruzi infection, 54% had some form of esophageal motor dysfunction, and one presented with achalasia.38 Here, an IFA with trypomastigotes of a Colombian TcI isolate was first used, and reactive samples were further assessed using a commercial ELISA kit (NovaTec Immunodiagnostica GmbH). This ELISA showed good performance in Colombian chronic chagasic patients with OD450 values greater than 2.00.39 Two achalasia patients for whom negative results were obtained by IFA were reactive by ELISA. The ELISA IgG detection kit (TcF antigen) used in this study has been used to detect antibodies in samples derived from patients in Argentina, Brazil, Chile, and Ecuador and shows good performance and reproducibility.23,40–44 Our results are inconsistent with two additional in-house IFA and ELISA assessments from a referral center; despite measuring IgG anti-T. cruzi, all samples were negative. In these assays, they used epimastigotes as antigens, and the type of antigen appears to be important in the detection of anti-T. cruzi antibodies, particularly in the digestive form of CDs. In one study that used IFA, an antigen from amastigotes detected 100% of Chagas digestive cases compared with 87% of cases with trypomastigotes and only 37% of cases with epimastigotes.45 The genetic diversity of T. cruzi infection might also influence the sensitivity of the techniques used.46Trypanosoma cruzi DTUs are geographically distributed and could be associated with the discordance among the serological tests used to detect T. cruzi-specific antibodies.4,47 Indeed, at least two reactive tests with different technical principles are needed for diagnosis.48,49 Repetitive peptide sequences from T. cruzi used in the TcF antigen were originally described in TcII strains [RA strain, clone 2 (PEP-2) and clone 13 (TcD)],44 and TcD was also observed in the Tulahuen TcVI strain.50 The parasite DTUs from which the TcE and TcLo1.2 sequences were obtained has not been defined.24 The third serological assay used (ChLIA) has been tested on samples from patients in Latin American countries, mainly from Argentina, Brazil, Bolivia, and Mexico.25,26 Here, none of the samples that yielded reactive results by the IFA and ELISA were reactive in the ChLIA. The additional peptides in the ChLIA, FP3 (TCR27 or FCaBP), FP6 (TCR39 or FRA), and FP10 (SAPA or MAP), were originally identified in a TcI Sylvio X-10/4 clone.51 The TcF antigen alone is also included in the immunoblot assay, and two patients with reactive bands were also classified as reactive by the ELISA. All serological assays share TcF as a common antigen; however, the additional peptides in the ChLIA could alter the ability of the antibody to recognize B-cell epitopes. A summary of the data produced with the serological tests used here is shown in Supplemental Table 1.




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