IgG Isotypes Targeting a Recombinant Chimeric Protein of Trypanosoma cruzi in Different Clinical Presentations of Chronic Chagas Disease.

Chagas disease (CD) is caused by the protozoan Trypanosoma cruzi, which leads to a spectrum of clinical presentations that range from asymptomatic to severe cardiac involvement. The host immune response plays a pivotal role in disease progression. Ig isotypes may contribute to disease pathogenesis. Investigating these components can provide insights into the immunopathogenic mechanisms underlying CD. This cross-sectional study aims to establish a correlation between the Ig profile of individuals infected with T. cruzi with the clinical forms of chronic CD. Serum samples were collected from partner institutions in different states of Brazil. Individuals diagnosed with chronic CD were categorized based on the clinical form of the disease. The indirect ELISA method using the recombinant chimeric Molecular Biology Institute of Paraná membrane protein 8.4 as the antigen was used to determine the Ig profile, including total IgG, IgG1, IgG2, IgG3, and IgG4. Ninety-seven serum samples from patients classified as negative (NEG, n = 38), indeterminate (IND, n = 24), mild cardiac (MC, n = 20), and severe cardiac (SC, n = 15) forms were analyzed. IgG1 exhibited greater levels compared with the other isotypes, showing a significant difference between the MC and IND groups. IgG3 levels were greater in individuals from the MC group compared with the SC group. IgG1 and IgG3 isotypes can serve as biomarkers to evaluate the progression of CD because they exhibit variations across clinical groups. Additional longitudinal studies are necessary to explore the relationship between antibody kinetics and the development of tissue damage.

A multicenter comparative study of the performance of four rapid immunochromatographic tests for the detection of anti-Trypanosoma cruzi antibodies in Brazil.

Diagnosis of Trypanosoma cruzi (T. cruzi) infection in the chronic phase of Chagas disease (CD) is performed by serologic testing. Conventional tests are currently used with very good results but require time, laboratory infrastructure, and expertise. Rapid diagnostic tests (RDTs) are an alternative as the results are immediate and do not require specialized knowledge, making them suitable for epidemiologic studies and promising as a screening tool. Nevertheless, few studies conducted comparative evaluations of RDTs to validate the results and assess their performance. In this study, we analyzed four trades of rapid tests (OnSite Chagas Ab Combo Rapid Test-United States, SD Bioline Chagas AB-United States, WL Check Chagas-Argentina, and TR Chagas Bio-Manguinhos-Brazil) using a panel of 190 samples, including sera from 111 infected individuals, most of whom had low T. cruzi antibody levels. An additional 59 samples from uninfected individuals and 20 sera from individuals with other diseases, mainly visceral leishmaniasis, were included. All tests were performed by three independent laboratories in a blinded manner. Results showed differences in sensitivity from 92.8 to 100%, specificity from 78.5 to 92.4%, and accuracy from 90.5 to 95.3% among the four assays. The results presented here show that all four RDTs have high overall diagnostic ability. However, WL Check Chagas and TR Chagas Bio-Manguinhos were considered most suitable for use in screening studies due to their high sensitivity combined with good performance. Although these two RDTs have high sensitivity, a positive result should be confirmed with other tests to confirm or rule out reactivity/positivity, especially considering possible cross-reactivity with individuals with leishmaniasis or toxoplasmosis.

Western blot using Trypanosoma cruzi chimeric recombinant proteins for the serodiagnosis of chronic Chagas disease: A proof-of-concept study.

BACKGROUND: Chagas disease (CD) is caused by Trypanosoma cruzi. The chronic phase of CD is characterized by the presence of IgG anti-T. cruzi antibodies; and diagnosis is performed by serological methods. Because there is no reliable test that can be used as a reference test, WHO recommends the parallel use of two different tests for CD serodiagnosis. If results are inconclusive, samples should be subjected to a confirmatory test, e.g., Western blot (WB) or PCR. PCR offers low sensitivity in the chronic phase, whereas few confirmatory tests based on the WB method are commercially available worldwide. Therefore, new diagnostic tools should be evaluated to fill the gap in CD confirmatory tests. In recent years, four chimeric recombinant antigens (IBMP-8.1, IBMP-8.2, IBMP-8.3 and IBMP-8.4) have been evaluated in phase I, II and III studies using ELISA, liquid microarray and immunochromatography with 95-100% accuracy. Given the high diagnostic performance of these antigens, the present study investigated the ability of these molecules to diagnose chronic CD using a WB testing platform. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we analyzed the diagnostic potential of four chimeric antigens using 40 T. cruzi-positive, 24-negative, and three additional positive samples for visceral leishmaniasis (i.e., potentially cross-reactive) using WB as the diagnostic platform. Checkerboard titration with different dilutions of antigens, conjugated antigens, and serum samples was performed to standardize all assays. All IBMP antigens achieved 100% sensitivity, specificity, and accuracy, with the exception of IBMP-8.3, which had 100% specificity despite lack of significance, but lower sensitivity (95%) and accuracy (96.9%). No cross-reactivity was observed in samples positive for leishmaniasis. CONCLUSIONS/SIGNIFICANCE: The present phase I (proof-of-concept) study demonstrated the high diagnostic potential of these four IBMP antigens to discriminate between T. cruzi-positive and -negative samples, making them candidates for phase II and confirmatory testing with WB.

Constipation Is Related to Small Bowel Disturbance Rather Than Colonic Enlargement in Acquired Chagasic Megacolon.

BACKGROUND: Constipation is the main symptom of acquired chagasic megacolon. However, a number of patients with Chagas disease without colon involvement also have the same complain. This study evaluated the role of small bowel in constipated patients with Chagas disease with and without megacolon. METHODS: Orocecal transit time (OCTT) and oral glucose tolerance test (OGTT) in constipated non-chagasic and chagasic patients with and without megacolon were performed. One hundred fifteen patients were included in this study and were divided into two groups based on the presence or absence of constipation, which is defined as at least 7 days without bowel movements for more than 1 year. These two groups were further divided into three subgroups based on the serology test results for Trypanosoma cruzi and the presence and absence of megacolon on barium enema. All patients were subjected to OCTT and OGTT. RESULTS: Among 70 constipated patients, 64.3% had OCTT longer than 120 min, higher than the non-constipated patients (31.1%, P < 0.000). The proportion of patients within the three subgroups in the non-constipated group was not different from each other (P = 0.345). Among the constipated subgroup, 94.44% of the chagasic megacolon subgroup had OCTT longer than 120 min, higher than the other two subgroups (P = 0.005). Chagas patients with constipation, without or without megacolon, showed higher blood glucose levels at 30, 60, and 90 min after oral ingestion of 70 g glucose than normal subjects with or without constipation. CONCLUSIONS: Constipated, either non-chagasic or chagasic, patients have a prolonged OCTT. This result suggests that slow small bowel transit may be a significant factor for constipation.

Proportional mortality ratio due to chagas disease is five times higher for the state of Goias than the rest of Brazil

Mortality data due to Chagas disease for the endemic State of Goias, Brazil, was retrieved from the National System of Information on Mortality, between 2006 and 2011. A total of 29,041 deaths were attributed to Chagas disease in the country, of which 4,293 (14.8%) occurred in the State of Goias. The proportion of deaths attributable to Chagas disease was 0,4% for the country overall and 2.4% for State of Goias. Seventy-two percent of the records were from individuals over 60 years of age, and heart disease was the main cause of death in 80.3%. Chagas disease is a major cause of death in Goias and, proportionally, 5.3 times higher than for the rest of the country.

Detecção de doenças transmissíveis em gestantes no Estado de Goiás: o teste da mamãe / Detection of transmissible diseases in pregnant women in the State of Goias, Brazil. Maternal screening

Neste trabalho é descrita a triagem de doenças infecciosas em gestantes do estado de Goiás para detecção de agravos que podem ser transmitidos durante a gravidez e causar sequelas na criança. A triagem é realizada por meio de programa da Secretaria Estadual da Saúde em parceria com a Associação de Pais e Amigos dos Excepcionais (APAE) e as Secretarias Municipais de Saúde. A experiência da APAE na detecção de algumas doenças congênitas por teste simples em papel filtro foi aproveitada e seu uso expandido para a detecção de doenças infecciosas/transmissíveis. De setembro de 2003 até junho de 2009 foram examinadas amostras de 348.037 gestantes. Implantada progressivamente a partir de dois municípios, a triagem para doenças infecciosas está disponível em 245 dos 246 municípios do estado de Goiás. Os agravos triados foram: sífilis, HIV/Aids, toxoplasmose, rubéola, hepatites B e C, infecção pelo Trypanosoma cruzi, HTLV e citomegalovirose. A triagem foi realizada em papel filtro por testes imunoenzimáticos (ELISA) para cada marcador e os resultados positivos foram confirmados por coleta de sangue venoso, cujo soro foi encaminhado a diferentes centros de referência. Foram identificadas 11.061 gestantes com resultados positivos. A confirmação após os testes com soro foi obtida em 10.496 (94,9%) amostras com as seguintes prevalências: sífilis: 4.028 (1,2%); toxoplasmose: 2.320 (0,7%), anticorpos anti-T. cruzi: 1.768 (0,5%); hepatite B: 956 (0,3%); HIV: 469 (0,1%), hepatite C: 334 (0,1%), HTLV: 312 (0,1%), rubéola: 181(0,05%) e fase aguda de citomegalovirose: 128 (0,04%). Os resultados foram encaminhados ao pré-natalista e ao núcleo de vigilância epidemiológica municipal.

Applicability of an optimized non-conventional flow cytometry method to detect anti-Trypanosoma cruzi immunoglobulin G for the serological diagnosis and cure assessment following chemotherapeutic treatment of Chagas disease.

One of the challenges on immunodiagnostic of Chagas disease in endemic areas has been the search for more practical and safe antigenic preparation that provides tests with higher sensitivity and specificity, with low cross-reactivity. A new approach using fixed Trypanosoma cruzi epimastigotes to detect IgG reactivity was investigated previously. In order to continue this investigation, this study aimed at optimizing the flow cytometry-based method to the diagnosis of Chagas disease patients after specific chemotherapy. To achieve our goal, serum samples from 93 subjects - 52 adults chronically infected by T. cruzi, and 41 uninfected controls were tested by flow cytometry. Secondly, serum samples from patients Treated Cured and Treated Uncured from Chagas disease were also tested to evaluate the potential of the method on assessing cure. After establishing the ideal serum dilution and cut off, 121 serum samples from patients with other endemic infections were tested to check cross-reactivity. The results showed that parasite staining with Evan's blue dye eliminated debris, allowing trustworthy analysis of anti-fixed epimastigote IgG reactivity. The applicability of the method to diagnose Chagas disease was confirmed by the high sensitivity (98.1%) and specificity (100%) found. This method also contributed for post-therapeutic assessment of cure, identifying 94.1% of Treated Uncured and 83.3% of Treated Cured patients. Cross-reactivity was observed in a very low number (6.7%). On the whole, these data highly recommend the use of anti-fixed T. cruzi epimastigote IgG reactivity by flow cytometry to the diagnosis and cure monitoring of Chagas disease in endemic areas.

Chagas disease: increased parasitemia during pregnancy detected by hemoculture.

One hundred fifty-two Trypanosoma cruzi seropositive women were submitted to a single hemoculture; 101 were pregnant, and 51 were not pregnant. Seven tubes from each individual were harvested with liver infusion tryptose (LIT) medium and observed monthly until the fifth month. Hemocultures were positive in 50% (76 of 152) of the women. Results showed that the positivity was 29.4% (15 of 51) among non-pregnant women and 60.4% (61 of 101) in pregnant women (P < 0.05). In relation to gestational age, there were significant differences in positivity, with a higher proportion of women with positive hemocultures (20 of 25) before 21 weeks and lower after 30 weeks (10 of 21; P = 0.02). We conclude that pregnancy enhances the parasitemia in Chagas disease, with a higher effect early in pregnancy.

Specific treatment for Trypanosoma cruzi: lack of efficacy of allopurinol in the human chronic phase of Chagas disease.

Thirty-five individuals from endemic areas of Central Brazil (age range, 18-64 years; 19 women) in the chronic phase of Chagas disease, with positive serology and presence of circulating parasites detected by one or more recent positive xenodiagnosis, were selected for this study. Allopurinol (900 mg/d) or placebo was administered in a double-blind clinical trial for 60 days. After codes were broken, 23 had been allocated to the intervention group and 12 to the placebo group. Side effects were observed in 11 patients in the intervention group and in 1 in the placebo group. Seventeen patients in the intervention group and 10 in the placebo group completed the trial. Follow-up was performed by monthly xenodiagnosis and serologic tests every 3 months during the first year and at the end of the trial. Xenodiagnosis remained positive in all 17 of the treated group and in all 10 of the placebo group. Serologic tests were persistently positive in both groups after treatment. We concluded that, at the doses used, allopurinol was not effective to clear, in our region, Trypanosoma cruzi from peripheral blood of infected individuals.

Detection of parasitemia profiles by blood culture after treatment of human chronic Trypanosoma cruzi infection.

The change in parasitemia profile, measured by sequential blood cultures of 27 benznidazole (Bz) treated patients compared with 13 untreated patients, on the chronic phase of chagas disease, is described. All patients were adults (age limits: 23-88) with positive serology (three tests); 23 of them were females. All patients were submitted to six blood cultures, three before and three after Bz treatment. The parasitemia was classified as nondetected (with three negative blood cultures), medium (one positive culture in three), and high (two or three positive cultures). From the eight patients with nondetected parasitemia before Bz, seven still had the same profile and only one switched to medium; from eight with medium parasitemia, seven shifted to nondetected, and one to high parasitemia. From the 11 patients with high parasitemia before Bz, ten converted to nondetected and only one was positive after Bz. Nineteen of the 27 patients changed the parasitemia profile (70.4%), and the rate of therapeutic failure was 11.1% (3/27) during the first 24 months of follow-up after Bz. The shift to nondetected parasitemia profile was from 8/27 to 24/27 patients after Bz treatment for the first 2 years. Only 46.2% (6/13) of the nontreated individuals changed the parasitemia profile. We conclude that there is a strong trypanocide effect of Bz (88.8%) and a rate of therapeutic failure of 11.1% during the first 2 years after trypanocidal treatment.

Gastric and transverse colonic volvulus in patient with chagasic megagastria and megacolon

In edemic regions for Chagas disease as in central part of Brazil digestive manifestations are common and colonic volvulus is a possible complication. Sigmoid volvulus is the most frequent type. Aim - report an uncommon case of volvulus is a possible complication. Sigmoid volvulus is the most frequent type / O volvo colônico é frequente nas regiões endêmicas da doença de Chagas em que o megacólon é manifestação comum. O sigmóide é o segmento mais acometido e a literatura é escassa sobre o padrão do volvo no megacólon chagásico...

Importância do método de obtençäo das dejeçöes dos triatomíneos na avaliaçäo da suscetibilidade triatomínica para Trypanosoma cruzi / Importance of the method for obtaining feces from triatominae in the assessment of triatomine susceptibility to Trypanosoma cruzi

Foi estudada a suscetibilidade de Dipetalogaster maximus, Rhodnius neglectus, R. prolixus, R. robustus, Triatoma infestans e T. rubrovaria ao Trypanosoma cruzi, através do xenodiagnóstico, em dois pacientes na fase aguda da doença de Chagas. Usaram-se como parâmetros o número de triatomíneos infectados e o número de tripanosomas excretados. A leitura do senodiagnóstico foi realizada pelos métodos da compressäo abdominal e o das dejeçöes espontâneas. Este foi mais eficiente que aquele, em relaçäo ao número de T. cruzi por lâmina pelo teste de Wilcoxon. Em um dos pacientes, tendo como parámetro o número de triatomíneos infectados, a suscetibilidade das espécies obedeceu a seguinte ordem: D. maximus e R. neglectus (100 por cento), R. robustus (95 por cento), R. prolixus e T. rubrovaria (90 por cento) e T. infestans (85 por cento). Nesse mesmo paciente, a suscetibilidade das espécies de triatomíneos, avaliada pelo número de T. cruzi excretados, usando-se compressäo abdominal, encontra-se na seguinte ordem: R. neglectus, R. prolixus, D. maximus e R. robustus, T. infestans e T. rubrovaria. Pelo método das dejeçöes, a ordem foi: D. maximus e r. prolixus, T. rubrovaria, R. robustus, T. infestans e R. negelctus. No outro paciente, esse parâmetro, em ambas as técnicas de leitura, mostrou a mesma ordem de suscetibilidade (R.neglectus, T. rubrovaria e T. infestans), porém, o número de tripanosomas excretados foi significativamente maior pelo método das dejeçöes espontâneas