Fernanda Machado FonsecaI; Dulciene Maria de Magalhães QueirozII; Andreia Maria Camargos RochaII; Aluízio Prata†; Eduardo CremaIII; Virmondes Rodrigues JuniorI; Luiz Eduardo RamirezI; Adriana Gonçalves de OliveiraI
IDepartamento de Ciências Biológicas, Universidade Federal do Triângulo Mineiro, Uberaba, MG IILaboratório de Pesquisa em Bacteriologia, Faculdade de Medicina, Universidade Federal de Minas Gerais, Belo Horizonte, MG IIIDepartamento de Clínica Médica, Universidade Federal do Triângulo Mineiro, Uberaba, MG
INTRODUCTION: In this study, we evaluated the seroprevalence of Helicobacter pylori infection among chagasic and non-chagasic subjects as well as among the subgroups of chagasic patients with the indeterminate, cardiac, digestive, and cardiodigestive clinical forms.
METHODS: The evaluated subjects were from the Triângulo Mineiro region, Minas Gerais, Brazil. Chagasic patients showed positive reactions to the conventional serological tests used and were classified according to the clinical form of their disease. Immunoglobulin G antibodies specific to H. pylori were measured using a commercial enzyme-linked immunosorbent assay kit.
RESULTS: The overall H. pylori prevalence was 77.1% (239/310) in chagasic and 69.1% (168/243) in non-chagasic patients. This difference was statistically significant even after adjustment for age and sex (odds ratio = 1.57; 95% confidence interval, 1.02-2.42; p = 0.04) in multivariate analysis. The prevalence of infection increased with age in the non-chagasic group (p = 0.007, χ2 for trend), but not in the chagasic group (p = 0.15, χ2 for trend). H. pylori infection was not associated with digestive or other clinical forms of Chagas disease (p = 0.27).
CONCLUSIONS: Our findings demonstrate that chagasic patients have a higher prevalence of H. pylori compared to non-chagasic subjects; a similar prevalence was found among the diverse clinical forms of the disease. The factors contributing to the frequent co-infection with H. pylori and Trypanosoma cruzi as well as its effects on the clinical outcome deserve further study.
Keywords: Helicobacter pylori.Chagas disease. Chronic Chagas disease clinical forms. Co-infection.
INTRODUÇÃO: No presente estudo, foi comparada a soroprevalência da infecção por Helicobacter pylori entre os indivíduos chagásicos e não-chagásicos, bem como entre subgrupos de chagásicos com as formas clínicas indeterminada, cardíaca, digestiva e cardiodigestiva.
MÉTODOS: Os indivíduos avaliados eram provenientes da região do Triângulo Mineiro, Minas Gerais, Brasil. Foram realizados testes sorológicos convencionais para diagnóstico da infecção pelo T. cruzi e os chagásicos foram classificados de acordo com a forma clínica. O diagnóstico de infecção por H. pylori foi estabelecido pela detecção de anticorpos IgG específicos utilizando-se um kit comercial de ELISA.
RESULTADOS: A prevalência da infecção por H. pylorifoi 77,1% (239/310) no grupo de pacientes chagásicos e 69,1% (168/243) no grupo de não-chagásicos. Esta diferença foi estatisticamente significativa mesmo após ajuste para idade e sexo (OR = 1,57; 95% CI, 1,02-2,42; p = 0,04) na análise multivariada. A prevalência da infecção aumentou de acordo com a idade no grupo não-chagásicos (p = 0,007, χ2 for trend) mas este aumento não foi observado no grupo dos chagásicos (p = 0,15, χ2 for trend). Não houve associação da infecção por H. pylori com a forma digestiva ou com qualquer outra forma clínica da doença de Chagas (p = 0,27).
CONCLUSÕES: Foi demonstrado que pacientes chagásicos apresentam maior prevalência da infecção por H. pylori quando comparados com não-chagásicos, independente da forma clínica da doença. Os fatores que contribuem para a frequente co-infecção Helicobacter pylori e Trypanosoma cruzi, bem como seus efeitos na evolução clínica das doenças associadas devem ser melhor estudados.
Palavras-chaves: Helicobacter pylori. Doença de Chagas. Formas clínicas da doença de Chagas crônica. Co-infecção.
Chagas disease, a tropical disease strongly associated with poor socioeconomic conditions, is endemic in rural Latin America. However, cases of Chagas disease have been reported throughout the world in the past few decades due to the increase in migrations towards non-endemic countries1. An estimated 10 million people are infected, mostly in Latin America, with the causative agent Trypanosoma cruzi, which is mainly transmitted by blood-sucking triatomine bugs2.
The clinical course of Chagas disease is highly variable. In a substantial proportion of the individuals who survive the acute phase, the disease enters the chronic phase that generally starts with a long period of clinical latency called the indeterminate form, which lasts for 10-30 years or throughout life3. Approximately one-third ofthese infected patients will develop serious lesionsintheheartand/ordigestivetract,characterizing the other clinical forms of chronic Chagas disease (i.e., cardiac, digestive, or cardiodigestive).
Likewise, the gram-negative bacterium Helicobacter pylori persists in most individuals, after being acquired mainly in childhood, as a chronic infection throughout life without complications, although a small percentage of infected individuals will develop severe gastroduodenal disease. Currently, H. pylori infection is recognized as the main cause of chronic gastritis and is directly linked to the development of peptic ulcer disease as well as gastric carcinoma and MALT lymphoma4. Interactions between bacterial virulence, host-related and environmental factors influence the clinical outcome in infected individuals, whereas concurrent infection with other pathogens may also potentially exacerbate or reduce disease severity due to the modulation of the gastric inflammatory process5-8 .
Before the discovery of H. pylori, the chronic gastritis commonly seen in chagasic patients, most often in those with the digestive form, had been formerly attributed to biliary duodenal-gastric reflux as well as to hypomotility and hypochlorhydria, resulting in injury to the enteric nervous system caused by T. cruzi infection9-12. Later, some studies demonstrated a high prevalence of H. pylori infection in patients with Chagas disease13-16. As this small number of studies on T. cruziand H. pylori co-infection had no control groups and/or included a low number of patients, we evaluated the prevalence of H. pylori infection among chagasic and non-chagasic subjects living in a region previously considered endemic for Chagas disease. We also analyzed whether H. pyloriinfection is associated with the clinical forms of chronic Chagas disease.
We evaluated outpatients with Chagas disease and subjects without the disease from the same geographical region living in a rural or urban area of Uberaba and other nearby cities. These cities are located in the west of the State of Minas Gerais, Brazil, in a region called Triângulo Mineiro, which was previously considered an endemic area for Chagas disease. Most of the study population was originally recruited to evaluate the prevalence of Chagas disease in the region. Therefore, the majority of the non-chagasic subjects were relatives of chagasic patients or were living near their residence. The other non-chagasic subjects were patients with dyspeptic complaints who underwent diagnostic endoscopy at the Hospital de Clinicas of the Universidade Federal do Triângulo Mineiro (UFTM). Peripheral blood (5mL) was collected from the subjects into vacuum tubes for serological tests in the period from 1995-2010. The sample was centrifuged and the serum was divided into aliquots and stored in sterile containers at -20ºC before testing.
The presence of immunoglobulin (Ig)G antibodies specific to H. pylori was measured by using a commercial enzyme-linked immunosorbent assay (ELISA) kit (Pyloriset EIA-GIII; Orion Diagnostica, Espoo, Finland). The antigen employed in this kit is a cell surface protein of H. pyloristrain NCTC 11637. The assay was performed according to the manufacturer’s instructions. Patients with positive serology were considered to be infected with H. pylori because spontaneous elimination of this bacterium without a specific eradication therapy is a rare event. The concentrations of IgG antibody in serum samples were determined by interpolation from a standard curve constructed by plotting absorbance values obtained for each of the 4 calibrator sera, which are provided by the manufacturer, against the corresponding anti-H. pyloriconcentrations in U/mL.
The diagnosis of Chagas disease was performed by means of ELISA (Chagatest ELISA-WIENER, Argentina), hemagglutination (Chagatest HAI-WIENER, Argentina), and immunofluorescence (Imuno-ConChagas-WAMA, Brazil)tests. Allchagasicpatients showed positive reactions to at least 2 serological tests, while non-chagasic subjects showed negative reactions to all 3 tests. Chagasic patients were classified according to the clinical form of their disease on the basis of the results of a conventional electrocardiogram and contrast X-ray of the esophagus and colon. Clinical, epidemiological, and demographic data were collected by reviewing the patients’ medical records. Only patientsfor who complete data were availablewere includedinthis study.
Data were analyzed using the SPSS statistical software package (version 17.0; SPSS Inc., Chicago, IL, USA). The association between H. pylori infection and Chagas disease was evaluated using a logistic regression model. Variables such as gender and mean age were included in this analysis. The odds ratio (OR) as well as its 95% confidence interval (CI) were determined. The association between H.pylori infection and the clinical form of chronic Chagas disease was tested by the two-tailed χ2 test with Yates correction or Fisher’s exact test. To evaluate the prevalence of H. pyloriwith aging, the χ2 test for trend was performed using Epi Info statistical software (version 3.5.1; Centers for Disease Control and Prevention, Atlanta, GA, USA). The significance level was set at p < 0.05.
This study was carried out in accordance with the Declaration of Helsinki and was approved by the Ethics Committee of UFTM, Uberaba, State of Minas Gerais, Brazil.
A total of 553 subjects were included in the present study. Of these subjects, 310 (56.1%) were patients with chronic Chagas disease (142 men and 168 women; mean age 57.7 ± 12.6 years, range 20-90) and 243 (43.9%) were non-chagasic subjects (100 men and 143 women; mean age 42.4 ± 15.9 years, range 18-86). Among the 310 patients with Chagas disease, 106 (34.2%) had the indeterminate clinical form of disease, 102 (32.9 %) had the cardiac form, 72 (23.2%) had the digestive form, and 30 (9.7%) had the cardiodigestive form.
The overall prevalence of H. pylori infection was significantly higher in chagasic (77.1%; 239/310) than in non-chagasic (69.1%; 168/243) subjects, even after adjustment for age and sex (OR = 1.57; 95% CI, 1.02-2.42; p = 0.04) in multivariate analysis (Table 1).
Table 2 shows that the prevalence of H. pylori infection increased with age (p = 0.007, χ2 for trend) from 61.9% (18-29-years age group) to 81.6% (50-59-years age group) in the non-chagasic group, but not in the chagasic group (p = 0.15, χ2 for trend). The rate of H. pyloriinfection decreased after 60 years of age among the non-chagasic subjects.
Figure 1 shows the prevalence of H. pylori infection according to the clinical forms of chronic Chagas disease. The rates of H.pylori positivity were 79.2% (84/106) in the group of patients with the cardiac form of the disease, 77.5% (79/102) in those with the indeterminate form, 76.7% (23/30) in those with the cardiodigestive form, and 73.6% (53/72) in those with the digestive form. There was no significant association between H. pylori infection and the clinical forms of Chagas disease (p = 0.27).
There was no significant difference (p = 0.34) in the concentrations of anti-H. pylori IgG for the chagasic (mean, 260.1U/mL) and nonchagasic (mean, 274.1U/mL) subjects. The mean concentration of anti- H.pylori IgG was also similar (p = 0.54) among the subgroups of patients with Chagas disease: digestive (278.1U/mL), indeterminate (mean, 267.3U/mL), cardiac (247.5U/mL), and cardiodigestive (239.5U/mL).
Approximately 50% of the world’s population is estimated to be infected with H. pylori, but individuals living in countries with low socioeconomic conditions have high prevalence rates of H. pylori acquired at an early age4,17. In developing countries, individuals are also more susceptible to acquiring chronic concurrent infections with other pathogens; however, the co-infection prevalence is still largely unknown as are the effects of each other in the natural history of the infections.
After the discovery of H. pylori in 1982, Barbosa et al.13 were the first to investigate the presence of bacteria in the gastric mucosa of patients with Chagas disease, followed by Oliveira et al., who evaluated patients with the digestive and indeterminate clinical forms of the disease, respectively. In those studies, the prevalence of H.pylori infection was very high (~95%). It is worth noting that both were uncontrolled studies with a small number of patients. Later, Nascimento et al. found a significantly higher prevalence of H. pylori in chagasic patients aged 21-50 years compared with non-chagasic subjects living in the same urban or rural area. More recently, Pinazo et al. evaluated a short series of high-risk Latin American immigrants that lived in Barcelona, confirming the frequent coexistence of H.pylori and T. cruzi infections.
In the present study, in which a larger number of patients with chronic Chagas disease were evaluated, a significantly higher prevalence of H. pylori infection was found in the chagasic group compared to the non-chagasic group, even after adjustment for confounding factors such as age and gender in multivariate analysis. Considering that H. pylori, even with its multiple mechanisms for acid adaptation, is susceptible to acid and can only survive for a few minutes at the low pH of the stomach lumen8,18, we might speculate that the hypochlorhydria described in the digestive form of Chagas disease could contribute to the higher prevalence of H. pylori among these patients. However, no significant difference between patients with the digestive and non-digestive forms was observed. Although low gastric acid secretion has been demonstrated only in patients with the digestive form, to date, it is known that patients presenting with one of the clinical forms of chronic Chagas disease can also present with lesions in other systems due to the criteria currently used. For instance, severe impairment of the autonomic control of the heart may occur in patients apparently having only digestive tract involvement19. In addition, a variable number of chronic chagasic patients with the indeterminate form show structural or functional abnormalities of the heart and digestive tract when more accurate diagnostic tests are employed20-22. Taken together, these findings could explain why we found a higher prevalence of H. pylori infection in chagasic patients, but without differences among the different forms of the disease.
Conversely, as it is known that chronic H. pylori infection exerts profound and diverse effects on gastric acid secretion, one should consider the possibility that some gastric physiological alterations such as hypergastrinemia and hypochlorhydria that were originally attributed to Chagas disease9,23,24 may actually be exacerbated by concurrent H. pylori infection. The effects of H. pylori infection on the secretion of gastric acid depend on the degree and localization of the gastritis and are related to the disease outcome25. Briefly, H. pylori infection confined to the antrum results in antralpredominant gastritis with the hypersecretion of acid and is associated with duodenal ulceration, whereas body infection may result in atrophic gastritis and abnormally low gastric acid secretion.
In the present study, the prevalence of H. pylori infection increased with age among non-chagasic subjects aged <60 years of age, but not among chagasic patients, who showed relatively constant rates of infection regardless of age. Multiple epidemiological studies have shown that the prevalence of H. pylori increases with age, which is best explained by a cohort phenomenon with diminished acquisition during childhood as socioeconomic development has occurred, because the risk of acquiring the infection is considered to be low in adulthood4. Our data might indicate that patients with Chagas disease have a high risk for acquiring an H. pylori infection in childhood as well as in adult life. Moreover, as reported in other studies, we observed a decrease in the prevalence of infection among the oldest individuals, which has been explained by the fall in the specific serologic response among older individuals due to the decrease in their general immunity17,26-28.
Considering that the transmission of T. cruzi and H. pylori is associated with poor socioeconomic conditions, the high prevalence of co-infection with these pathogens found in our study (~77%) is not surprising. In Brazil, the overall prevalence of H. pylori is high when compared to that of developed countries, although the rates found vary according to the geographical region studied. In the south of Brazil, the rate of H. pylori prevalence in adults is ~63%, while in the poorest regions such as the northeast and mid-west it is ~90%27-29. Regarding the State of Minas Gerais, where the population evaluated in the present study lives, rates of H. pylori infection ranging from 62% to 86% have been reported among adults17,30. The study population of chagasic and non-chagasic subjects may be considered quite homogeneous since the majority of individuals were adults living in the same geographic region and, presumably, have similar socioeconomic conditions and risk factors for acquiring an H. pylori infection.
In conclusion, our findings demonstrate that chagasic patients have a high prevalence of H. pyloricompared to non-chagasic subjects in the study population. Notably, H. pylori infections were not associated with the digestive or other clinical forms of Chagas disease. Therefore, the factors contributing to this frequent co-infection with H.pylori and T. cruzi as well as the effects on the clinical outcomes deserve further study that will certainly contribute to a better understanding of the gastric physiological alterations induced by both of these important agents of chronic diseases.
We thank Dr. Valdo José D. Silva for providing us with the clinical data of the chagasic patients with the cardiac form.
CONFLICT OF INTEREST
The authors declare that there is no conflict of interest.
This work was supported by grants from Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG) and the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Brazil.
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Dra. Adriana Gonçalves de Oliveira
DCB/UFTM. Praça Manoel Terra 330
38015-050 Uberaba, MG, Brasil
Phone: 55 34 3318-5480; Fax: 55 34 3318-5462
Received in 18/06/2011
Accepted in 30/09/2011
† In memorian.