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Chagas disease (CD) has become a worldwide problem due to globalization. In Europe, most cases are imported and are diagnosed in the chronic phase by two serological tests, as recommended by the World Health Organization. Chemiluminescent microparticle immunoassays (CMIAs) are an emerging alternative to the diagnostic standard. We aimed to validate the CMIA Alinity s Chagas® as a primary diagnostic test for chronic CD following its replacement of Architect Chagas®, with an amended signal-to-cut-off (S/CO) ratio of ≥6. Laboratory results and clinical data were collected retrospectively from 774 sera from an at-risk population tested for CD in Barcelona during 2020–2022. Negative results required no further testing, and those with a S/CO ratio ≥ 0.8 were confirmed by a second serological assay, according to the common practice. Four per cent of the samples (31/774) were determined to be seropositive by Alinity s, 93.5% of which (29/31) had an S/CO ratio ≥ 6. Almost all the samples could be directly classified by the corrected S/CO. Alinity s Chagas® was validated as a single test for chronic CD diagnosis by raising the S/CO to ≥6. Its implementation could provide faster results and help reduce CD underdiagnosis in non-endemic countries. [ABSTRACT FROM AUTHOR]

The objective of this study was to compare, by qPCR, the circulating blood parasite load of Trypanosoma cruzi in the buffy coat, and in whole blood mixed with boiled and unboiled guanidine hydrochloride-EDTA buffer, of individuals with chronic ChD. The concentration and purity of DNA were evaluated in a Nanodrop Denovix DS-11FX Series Spectrophotometer (DeNovix Inc., Wilmington, NC, USA). The parasite load was determined with the Taqman® qPCR system using a Stratagene Mx3000P thermocycler (Agilent Technologies, Santa Clara, CA, USA) with Cruzi 1 and Cruzi 2 satellite primers. Student's t-test with Bonferroni correction, Chi-squared (χ2) tests and Spearman's correlation coefficient were applied. The concentration and purity of DNA were higher in the buffy coat. Parasite DNA was detected and quantifiable in the three types of samples in seven patients, without statistically significant differences in the parasite load obtained. Higher correlations were found between the total DNA concentrations and the parasite loads obtained in the samples of the buffy coat. [ABSTRACT FROM AUTHOR]

Chagas disease (CD) has become a worldwide problem due to globalization. In Europe, most cases are imported and are diagnosed in the chronic phase by two serological tests, as recommended by the World Health Organization. Chemiluminescent microparticle immunoassays (CMIAs) are an emerging alternative to the diagnostic standard. We aimed to validate the CMIA Alinity s Chagas® as a primary diagnostic test for chronic CD following its replacement of Architect Chagas®, with an amended signal-to-cut-off (S/CO) ratio of ≥6. Laboratory results and clinical data were collected retrospectively from 774 sera from an at-risk population tested for CD in Barcelona during 2020–2022. Negative results required no further testing, and those with a S/CO ratio ≥ 0.8 were confirmed by a second serological assay, according to the common practice. Four per cent of the samples (31/774) were determined to be seropositive by Alinity s, 93.5% of which (29/31) had an S/CO ratio ≥ 6. Almost all the samples could be directly classified by the corrected S/CO. Alinity s Chagas® was validated as a single test for chronic CD diagnosis by raising the S/CO to ≥6. Its implementation could provide faster results and help reduce CD underdiagnosis in non-endemic countries.

Chagas disease is caused by the Trypanosoma cruzi parasite and is transmitted by infected triatomine bugs. This infection affects approximately 8 million people in the Americas, and due to globalisation and displacement, it is becoming increasingly common to find infected patients worldwide. Diagnosis of the disease in its acute form is relatively simple, as the parasite can be detected in peripheral blood smears, and symptoms are visible. However, in its chronic condition, the parasite is almost undetectable, and indirect tests are necessary to determine the presence of antibodies in infected patients. It is important to note that a single test is not enough to confirm the disease in this phase, as a second serological test should confirm the diagnosis. If the results are contradictory, a third test should be performed to confirm or discard the disease. Unfortunately, laboratories may not have access to all necessary tests in many rural areas where the disease is more frequent. Rapid tests to diagnose this disease present problems, such as significant variations in sensitivity and specificity in different countries. Therefore, searching for new biomarkers that allow for optimal correlation is essential. In this work, we have searched scientific literature from the last 10 years for mentions of novel biomarkers for diagnosis, treatment follow‐up, and prediction of cardiac complications in Chagas disease in its chronic phase. [ABSTRACT FROM AUTHOR]

Chagas disease (CD), caused by the parasite Trypanosoma cruzi, is a neglected tropical disease with life-threatening implications. In this study, we conducted a seroepidemiological survey to determine the prevalence and clinical profiles of CD in 217 individuals from an impoverished rural community in Southern Bahia, Brazil. The overall prevalence of CD in the studied community was 0.92%, detected through latent class analysis (LCA). Two individuals tested positive for anti-T. cruzi IgG, both being male farmers. One case was a 22-year-old man born in Camamu, with no evidence of congenital transmission, suggesting other routes of transmission such as vector-borne transmission due to migratory activities. The other case was a 69-year-old man born in São Felipe, who had lived in an adobe/brick house and had a pacemaker due to cardiac involvement caused by CD. The prevalence in this community was lower than expected, given the socioeconomic conditions and environmental factors that contribute to T. cruzi transmission. This could be attributed to the implementation of preventive measures and vector control programs by the Brazilian Government. However, continuous monitoring and surveillance are essential to sustain control efforts and detect any potential re-emergence of the disease. While the overall prevalence was low, the detection of positive cases underscores the need for continued surveillance and control measures in vulnerable populations, such as rural communities. Active surveillance, early diagnosis, and timely treatment are crucial in preventing disease progression and complications, thereby enhancing the effectiveness of screening and treatment programs. [ABSTRACT FROM AUTHOR]

Antigen‐specific T cells are central to the adaptive immune response against T. cruzi infection and underpin the efficacy of on‐going vaccine strategies. In this context, the present study focuses on T‐cell assays that define the parasite‐specificity on the basis of upregulation of TCR stimulation‐induced surface markers. For this purpose, we tested different dual marker combinations (OX40, CD25, CD40L, CD137, CD69, PD‐L1, CD11a, CD49d, HLA‐DR, CD38) to reliably identify activated CD4+ and CD8+ T‐cell populations from PBMCs of chronic Chagas disease (CCD) patients after 12 or 24 h of stimulation with T. cruzi lysate. Results demonstrated that activation‐induced markers (AIM) assays combining the expression of OX40, CD25, CD40L, CD137, CD69 and/or PD‐L1 surface markers are efficient at detecting T. cruzi‐specific CD4+ T cells in CCD patients, in comparison to non‐infected donors, after both stimulation times. For CD8+ T cells, only PD‐L1/OX40 after 24 h of antigen exposure resulted to be useful to track a parasite‐specific response. We also demonstrated that the agnostic activation is mediated by different T. cruzi strains, such as Dm28c, CL Brener or Sylvio. Additionally, we successfully used this approach to identify the phenotype of activated T lymphocytes based on the expression of CD45RA and CCR7. Overall, our results show that different combinations of AIM markers represent an effective and simple tool for the detection of T. cruzi‐specific CD4+ and CD8+ T cells. [ABSTRACT FROM AUTHOR]

IntroductionIn Brazil, an estimated 1.1 million people are infected with Trypanosoma cruzi, the causative agent of Chagas disease (CD). Despite the high number of cases, the estimated prevalence of infection per 100 inhabitants is low (0.03). However, the actual number of chronically infected individuals is still unknown. Therefore, we sought to determine the prevalence of chronic CD in at-risk individuals in Caraíbas (Bahia, Brazil) through active case finding.MethodsA total of 572 individuals living in rural or urban areas of Caraíbas were eligible for the study. A serum sample was collected from 226 individuals, and the diagnosis performed according to international guidelines.ResultsThe overall prevalence of anti-T. cruzi IgG was 4.42%. The median age of anti-T. cruzi IgG-positive individuals was 54.5 years, and the female-to-male ratio was 1.5:1. The prevalence of anti-T. cruzi IgG was similar in rural (4.29%) and urban areas (4.65%).DiscussionCompared with national estimates, we concluded that Caraíbas had a high prevalence for chronic CD and a high risk for persistent transmission. Through our study, it was possible to monitor individuals who were unaware of their clinical condition, thus improving their quality of life.

The objective of this study was to compare, by qPCR, the circulating blood parasite load of Trypanosoma cruzi in the buffy coat, and in whole blood mixed with boiled and unboiled guanidine hydrochloride-EDTA buffer, of individuals with chronic ChD. The concentration and purity of DNA were evaluated in a Nanodrop Denovix DS-11FX Series Spectrophotometer (DeNovix Inc., Wilmington, NC, USA). The parasite load was determined with the Taqman® qPCR system using a Stratagene Mx3000P thermocycler (Agilent Technologies, Santa Clara, CA, USA) with Cruzi 1 and Cruzi 2 satellite primers. Student’s t-test with Bonferroni correction, Chi-squared (χ2) tests and Spearman’s correlation coefficient were applied. The concentration and purity of DNA were higher in the buffy coat. Parasite DNA was detected and quantifiable in the three types of samples in seven patients, without statistically significant differences in the parasite load obtained. Higher correlations were found between the total DNA concentrations and the parasite loads obtained in the samples of the buffy coat.

ABSTRACT: Objectives: This study aimed to describe the electrocardiographic and echocardiographic status of chronic Chagas disease (cChD) patients treated with nifurtimox. Methods: An observational study was performed in 146 cChD patients followed over a mean of 7.9 years. Results: Of the 146 patients, 41 (28.1%) with normal electrocardiogram (ECG) at baseline maintained this condition, 34 (23.3%) with altered ECG at baseline normalised the alterations, and 46 (31.5%) with ECG abnormalities at baseline maintained this condition [23 (15.8%) with small alterations]. Finally, 25 cases (17.1%) in indeterminate phase altered the ECG. Differences before and after follow-up (P < 0.001) were found. The percentage of beneficial treatment was different than expected by chance (Z = 4.8; P < 0.001) and the annual percentage of cases that developed ECG alterations was lower than that of a historical cohort of untreated patients (P < 0.001). An echocardiogram was performed in 68 patients with baseline ECG alterations. The ejection fraction (EF) was normal in 57 (83.8%) and abnormal in 11 (16.2%). In 38 patients with ECG abnormalities that did not progress after treatment, EF and segmental motility (SM) were normal in 31 (81.6%) and 26 (68.4%), respectively. In 17 patients with ECG abnormalities, EF and SM were normal in 15 (88.2%) and 14 (82.4%) cases, respectively. Conclusion: Less progression to cardiomyopathy compared with a historical untreated cohort as well as the EF/SM results in patients with abnormal ECG that did not progress and in indeterminate cChD that altered the ECG suggests a beneficial effect of nifurtimox.

Chagas disease (CD), caused by the parasite Trypanosoma cruzi, is a neglected tropical disease with life-threatening implications. In this study, we conducted a seroepidemiological survey to determine the prevalence and clinical profiles of CD in 217 individuals from an impoverished rural community in Southern Bahia, Brazil. The overall prevalence of CD in the studied community was 0.92%, detected through latent class analysis (LCA). Two individuals tested positive for anti-T. cruzi IgG, both being male farmers. One case was a 22-year-old man born in Camamu, with no evidence of congenital transmission, suggesting other routes of transmission such as vector-borne transmission due to migratory activities. The other case was a 69-year-old man born in São Felipe, who had lived in an adobe/brick house and had a pacemaker due to cardiac involvement caused by CD. The prevalence in this community was lower than expected, given the socioeconomic conditions and environmental factors that contribute to T. cruzi transmission. This could be attributed to the implementation of preventive measures and vector control programs by the Brazilian Government. However, continuous monitoring and surveillance are essential to sustain control efforts and detect any potential re-emergence of the disease. While the overall prevalence was low, the detection of positive cases underscores the need for continued surveillance and control measures in vulnerable populations, such as rural communities. Active surveillance, early diagnosis, and timely treatment are crucial in preventing disease progression and complications, thereby enhancing the effectiveness of screening and treatment programs.

Extracellular vesicles (EVs) are lipid bilayer envelopes that encase several types of molecules. Their contents mostly reflect their cell origin and possible targets at other locations in the organism and can be modified in pathological conditions to interfere with intercellular communication, thus promoting disease establishment and development. These characteristics, in addition to their presence in virtually all body fluids, make such vesicles ideal for biomarker discovery in human diseases. Here, we describe the effect of different anticoagulants and the combination of two purification methods for isolation and characterization of circulating EVs from blood of chronic Chagas disease (CCD) patients. We illustrated this procedure by studying a population of patients with Chagas disease at the indeterminate chronic stage, in which the Trypanosoma cruzi is very scarce in circulation. EVs were harvested from blood collected without or with different anticoagulants. Protein and nanoparticle tracking analysis was used to measure EVs size and concentration. The EVs were purified by ultracentrifugation, followed by size‐exclusion chromatography and characterized by chemiluminescent enzyme‐linked immunosorbent assay and dot blot using antibodies that recognized parasite‐derived EVs, such as hyperimmune sera, polyclonal and monoclonal antibodies against trans‐sialidase and mucins. In parallel, antibodies against classical human EV markers CD9, CD63, CD81, and CD82, were also analyzed. The results showed that anticoagulants did not interfere with the analyzed parameters and circulating EVs from CCD patients contain T. cruzi antigens and classical human exosomal markers. Overall, our protocol is adequate for the isolation of the total circulating EVs and can serve as an important basis for further studies on biomarker discovery in Chagas' disease. [ABSTRACT FROM AUTHOR]

Chronic Chagas disease cardiomyopathy (CCC) is the most important clinical manifestation of infection with Trypanosma cruzi (T. cruzi) due to its frequency and effects on morbidity and mortality. Peripheral blood mononuclear cells (PBMC) infiltrate the tissue and differentiate into inflammatory macrophages. Advances in pathophysiology show that myeloid cell subpopulations contribute to cardiac homeostasis, emerging as possible therapeutic targets. We previously demonstrated that fenofibrate, PPARα agonist, controls inflammation, prevents fibrosis and improves cardiac function in a murine infection model. In this work we investigated the spontaneous release of inflammatory cytokines and chemokines, changes in the frequencies of monocyte subsets, and fenofibrate effects on PBMC of seropositive patients with different clinical stages of Chagas disease. The results show that PBMC from Chagas disease patients display higher levels of IL-12, TGF-β, IL-6, MCP1, and CCR2 than cells from uninfected individuals (HI), irrespectively of the clinical stage, asymptomatic (Asy) or with Chagas heart disease (CHD). Fenofibrate reduces the levels of pro-inflammatory mediators and CCR2 in both Asy and CHD patients. We found that CHD patients display a significantly higher percentage of classical monocytes in comparison with Asy patients and HI. Besides, Asy patients have a significantly higher percentage of non-classical monocytes than CHD patients or HI. However, no difference in the intermediate monocyte subpopulation was found between groups. Moreover, monocytes from Asy or CHD patients exhibit different responses upon stimulation in vitro with T. cruzi lysates and fenofibrate treatment. Stimulation with T. cruzi significantly increases the percentage of classical monocytes in the Asy group whereas the percentage of intermediate monocytes decreases. Besides, there are no changes in their frequencies in CHD or HI. Notably, stimulation with T. cruzi did not modify the frequency of the non-classical monocytes subpopulation in any of the groups studied. Moreover, fenofibrate treatment of T. cruzi -stimulated cells, increased the frequency of the non-classical subpopulation in Asy patients. Interestingly, fenofibrate restores CCR2 levels but does not modify HLA-DR expression in any groups. In conclusion, our results emphasize a potential role for fenofibrate as a modulator of monocyte subpopulations towards an anti-inflammatory and healing profile in different stages of chronic Chagas disease. [ABSTRACT FROM AUTHOR]

Chronic Chagas disease cardiomyopathy (CCC) is the most important clinical manifestation of infection with Trypanosma cruzi (T. cruzi) due to its frequency and effects on morbidity and mortality. Peripheral blood mononuclear cells (PBMC) infiltrate the tissue and differentiate into inflammatory macrophages. Advances in pathophysiology show that myeloid cell subpopulations contribute to cardiac homeostasis, emerging as possible therapeutic targets. We previously demonstrated that fenofibrate, PPARα agonist, controls inflammation, prevents fibrosis and improves cardiac function in a murine infection model. In this work we investigated the spontaneous release of inflammatory cytokines and chemokines, changes in the frequencies of monocyte subsets, and fenofibrate effects on PBMC of seropositive patients with different clinical stages of Chagas disease. The results show that PBMC from Chagas disease patients display higher levels of IL-12, TGF-β, IL-6, MCP1, and CCR2 than cells from uninfected individuals (HI), irrespectively of the clinical stage, asymptomatic (Asy) or with Chagas heart disease (CHD). Fenofibrate reduces the levels of pro-inflammatory mediators and CCR2 in both Asy and CHD patients. We found that CHD patients display a significantly higher percentage of classical monocytes in comparison with Asy patients and HI. Besides, Asy patients have a significantly higher percentage of non-classical monocytes than CHD patients or HI. However, no difference in the intermediate monocyte subpopulation was found between groups. Moreover, monocytes from Asy or CHD patients exhibit different responses upon stimulation in vitro with T. cruzi lysates and fenofibrate treatment. Stimulation with T. cruzi significantly increases the percentage of classical monocytes in the Asy group whereas the percentage of intermediate monocytes decreases. Besides, there are no changes in their frequencies in CHD or HI. Notably, stimulation with T. cruzi did not modify the frequency of the non-classical monocytes subpopulation in any of the groups studied. Moreover, fenofibrate treatment of T. cruzi-stimulated cells, increased the frequency of the non-classical subpopulation in Asy patients. Interestingly, fenofibrate restores CCR2 levels but does not modify HLA-DR expression in any groups. In conclusion, our results emphasize a potential role for fenofibrate as a modulator of monocyte subpopulations towards an anti-inflammatory and healing profile in different stages of chronic Chagas disease.

Chagas cardiomyopathy affects a considerable number of patients infected with the protozoan Trypanosoma cruzi (T. cruzi) and remains one of the most neglected tropical diseases despite being a significant contributor to morbidity and mortality in both endemic regions of Latin America and non-endemic countries like the United States. Since its discovery almost a century ago, knowledge gaps still exist in the mechanisms involved in the pathogenesis of Chagas cardiomyopathy, and numerous challenges exist in its diagnosis and treatment. This article reviews the main pathogenetic mechanisms involved in the progression of Chagas cardiomyopathy, which has been proposed as a result of years of research. It also emphasizes the challenges involved in the diagnosis of the asymptomatic indeterminate phase and has focused on several diagnostic techniques, including echocardiography, electrocardiogram (ECG), magnetic resonance imaging (MRI), and nuclear imaging in diagnosing symptomatic Chagas cardiomyopathy. In this article, we have also provided a brief overview of the current treatment of Chagas cardiomyopathy, which is not etiology-specific but instead derived from the knowledge acquired from the treatment of other cardiomyopathies., Competing Interests: Conflicts of interest: In compliance with the ICMJE uniform disclosure form, all authors declare the following: Payment/services info: All authors have declared that no financial support was received from any organization for the submitted work. Financial relationships: All authors have declared that they have no financial relationships at present or within the previous three years with any organizations that might have an interest in the submitted work. Other relationships: All authors have declared that there are no other relationships or activities that could appear to have influenced the submitted work., (Copyright © 2024, Veluswami et al.)

Infection by the Trypanosoma cruzi parasite causes Chagas disease and triggers multiple immune mechanisms in the host to combat the pathogen. Chagas disease has a variable clinical presentation and progression, producing in the chronic phase a fragile balance between the host immune response and parasite replication that keeps patients in a clinically silent asymptomatic stage for years. Since the parasite is intracellular and replicates within cells, the cell-mediated response of the host adaptive immunity plays a critical role. This function is mainly orchestrated by T lymphocytes, which recognize parasite antigens and promote specific functions to control the infection. However, little is known about the immunological markers associated with this asymptomatic stage of the disease. In this large-scale analysis, the differential expression of 106 immune system-related genes has been analyzed using high-throughput qPCR in T. cruzi antigen-stimulated PBMC from chronic Chagas disease patients with indeterminate form (IND) and healthy donors (HD) from endemic and non-endemic areas of Chagas disease. This analysis revealed that there were no differences in the expression level of most genes under study between healthy donors from endemic and non-endemic areas determined by PCA and differential gene expression analysis. Instead, PCA revealed the existence of different expression profiles between IND patients and HD (p < 0.0001), dependent on the 32 genes included in PC1. Differential gene expression analysis also revealed 23 upregulated genes (expression fold change > 2) and 11 downregulated genes (expression fold change < 0.5) in IND patients versus HD. Enrichment analysis showed that several upregulated genes in IND patients participate in relevant immunological pathways such as antigen-dependent B cell activation, stress induction of HSP regulation, NO2-dependent IL12 pathway in NK cells, and cytokine-inflammatory response. The antigen-specific differential gene expression profile detected in these patients and the relevant immunological pathways that seem to be activated could represent potential biomarkers of the asymptomatic form of Chagas disease, helpful to diagnosis and infection control.

The clinical evolution of patients with chronic Chagas disease (CCD) is mainly associated with an excessive inflammation and a defective immunomodulatory profile caused by the interaction between T. cruzi and the host. Regulatory B (Breg) cells exert immune suppression mostly through IL-10 production (B10 cells), but also through IL-10-independent mechanisms. Previously, we demonstrated that CCD patients with cardiomyopathy show changes in the ex vivo Breg cell phenotypic distribution although maintain IL-10 production capacity. Here, we sought to identify potential alterations on Breg cells upon in vitro stimulation. Isolated B cells from CCD patients with or without cardiomyopathy and non-infected (NI) donors were stimulated with T. cruzi lysate or CpG + CD40L, and characterized by flow cytometry based on the expression of CD24, CD27, CD38, and the regulatory molecules IL-10 and PD-L1. IL-10 and IL-17 secretion in the supernatant of B cells was evaluated by ELISA. Data showed that T. cruzi stimulation diminished the expression of CD24 and CD38 on CD27− B cells while reducing the percentage of CD24high inside CD27+ B cells. Furthermore, T. cruzi induced a regulatory B cell phenotype by increasing B10 cells and IL-10 secretion in all the groups. The innate-like B10 cells expansion observed in patients with cardiomyopathy would be associated with CD27− B10 cell subsets, while no predominant phenotype was found in the other groups. Patients with cardiomyopathy also displayed higher IL-17 secretion levels in T. cruzi–activated B cells. CpG + CD40L stimulation revealed that B cells from CCD patients and NI donors had the same ability to differentiate into B10 cells and secrete IL-10 in vitro. Additionally, CCD patients showed an increased frequency of CD24−CD27− B cells and a reduction in the percentage of CD24highCD27+ Breg cells, which appeared to be inversely correlated with the presence of T. cruzi DNA in blood. Finally, CCD patients exhibited a higher frequency of PD-L1+ B cells in T. cruzi–stimulated samples, suggesting that IL-10-independent mechanisms could also be tangled in the control of inflammation. Altogether, our results provide evidence about the potential role of Breg cells in the immune response developed against T. cruzi and its contribution to chronic Chagas cardiomyopathy.

Infection by the Trypanosoma cruzi parasite causes Chagas disease and triggers multiple immune mechanisms in the host to combat the pathogen. Chagas disease has a variable clinical presentation and progression, producing in the chronic phase a fragile balance between the host immune response and parasite replication that keeps patients in a clinically silent asymptomatic stage for years. Since the parasite is intracellular and replicates within cells, the cell-mediated response of the host adaptive immunity plays a critical role. This function is mainly orchestrated by T lymphocytes, which recognize parasite antigens and promote specific functions to control the infection. However, little is known about the immunological markers associated with this asymptomatic stage of the disease. In this large-scale analysis, the differential expression of 106 immune system-related genes has been analyzed using high-throughput qPCR in T. cruzi antigen-stimulated PBMC from chronic Chagas disease patients with indeterminate form (IND) and healthy donors (HD) from endemic and non-endemic areas of Chagas disease. This analysis revealed that there were no differences in the expression level of most genes under study between healthy donors from endemic and non-endemic areas determined by PCA and differential gene expression analysis. Instead, PCA revealed the existence of different expression profiles between IND patients and HD (p < 0.0001), dependent on the 32 genes included in PC1. Differential gene expression analysis also revealed 23 upregulated genes (expression fold change > 2) and 11 downregulated genes (expression fold change < 0.5) in IND patients versus HD. Enrichment analysis showed that several upregulated genes in IND patients participate in relevant immunological pathways such as antigen-dependent B cell activation, stress induction of HSP regulation, NO2-dependent IL12 pathway in NK cells, and cytokine-inflammatory response. The antigen-specific differential gene expression profile detected in these patients and the relevant immunological pathways that seem to be activated could represent potential biomarkers of the asymptomatic form of Chagas disease, helpful to diagnosis and infection control. [ABSTRACT FROM AUTHOR]

The clinical evolution of patients with chronic Chagas disease (CCD) is mainly associated with an excessive inflammation and a defective immunomodulatory profile caused by the interaction between T. cruzi and the host. Regulatory B (Breg) cells exert immune suppression mostly through IL-10 production (B10 cells), but also through IL-10-independent mechanisms. Previously, we demonstrated that CCD patients with cardiomyopathy show changes in the ex vivo Breg cell phenotypic distribution although maintain IL-10 production capacity. Here, we sought to identify potential alterations on Breg cells upon in vitro stimulation. Isolated B cells from CCD patients with or without cardiomyopathy and non-infected (NI) donors were stimulated with T. cruzi lysate or CpG + CD40L, and characterized by flow cytometry based on the expression of CD24, CD27, CD38, and the regulatory molecules IL-10 and PD-L1. IL-10 and IL-17 secretion in the supernatant of B cells was evaluated by ELISA. Data showed that T. cruzi stimulation diminished the expression of CD24 and CD38 on CD27− B cells while reducing the percentage of CD24high inside CD27+ B cells. Furthermore, T. cruzi induced a regulatory B cell phenotype by increasing B10 cells and IL-10 secretion in all the groups. The innate-like B10 cells expansion observed in patients with cardiomyopathy would be associated with CD27− B10 cell subsets, while no predominant phenotype was found in the other groups. Patients with cardiomyopathy also displayed higher IL-17 secretion levels in T. cruzi –activated B cells. CpG + CD40L stimulation revealed that B cells from CCD patients and NI donors had the same ability to differentiate into B10 cells and secrete IL-10 in vitro. Additionally, CCD patients showed an increased frequency of CD24−CD27− B cells and a reduction in the percentage of CD24highCD27+ Breg cells, which appeared to be inversely correlated with the presence of T. cruzi DNA in blood. Finally, CCD patients exhibited a higher frequency of PD-L1+ B cells in T. cruzi –stimulated samples, suggesting that IL-10-independent mechanisms could also be tangled in the control of inflammation. Altogether, our results provide evidence about the potential role of Breg cells in the immune response developed against T. cruzi and its contribution to chronic Chagas cardiomyopathy. [ABSTRACT FROM AUTHOR]

Background The diversity of individuals at risk for Trypanosoma cruzi infection in the United States poses challenges for diagnosis. We evaluated the diagnostic accuracy of Food and Drug Administration (FDA)-cleared tests in the Washington Metropolitan area (WMA). Methods In total, 1514 individuals were evaluated (1078 from Mexico, Central and northern South America [TcI-predominant areas], and 436 from southern South America [TcII/V/VI-predominant areas]). Optical density (OD) values from the Hemagen EIA and Chagatest v.3 Wiener, and categorical results of the IgG-TESA-blot (Western blot with trypomastigote excretory-secretory antigen), and the Chagas detect plus (CDP), as well as information of area of origin were used to determine T. cruzi serostatus using latent class analysis. Results We detected 2 latent class (LC) of seropositives with low (LC1) and high (LC2) antibody levels. A significantly lower number of seropositives were detected by the Wiener, IgG-TESA-blot, and CDP in LC1 (60.6%, P <.001, 93.1%, P =.014, and 84.9%, P =.002, respectively) as compared to LC2 (100%, 100%, and 98.2%, respectively). LC1 was the main type of seropositives in TcI-predominant areas, representing 65.0% of all seropositives as opposed to 22.8% in TcII/V/VI-predominant areas. The highest sensitivity was observed for the Hemagen (100%, 95% confidence interval [CI]: 96.2–100.0), but this test has a low specificity (90.4%, 95% CI: 88.7–91.9). The best balance between positive (90.9%, 95% CI: 83.5–95.1), and negative (99.9%, 95% CI: 99.4–99.9) predictive values was obtained with the Wiener. Conclusions Deficiencies in current FDA-cleared assays were observed. Low antibody levels are the main type of seropositives in individuals from TcI-predominant areas, the most frequent immigrant group in the United States. [ABSTRACT FROM AUTHOR]

We compared the accuracy of the Stat-Pak and Chagas Detect Plus with a latent class analysis. Sensitivity values of 89.7% and 91.9% and specificities of 97.1% and 80.3%, respectively, were seen in the serodiagnosis of Chagas disease in Hispanic immigrants, revealing the limitations of these tests in diverse populations. [ABSTRACT FROM AUTHOR]

Introduction: Chagas disease (ChD) is one of the main parasitic diseases in Latin‐America. Its heart involvement is the most important cause of death. The aim of this study is to evaluate if Doppler Tissue Imaging (DTI) may have a predictive value for later events in subjects with chronic ChD. Methods: we analyses DTI variables of 543 patients with chronic ChD for the evaluation of predicting factors of events. Major adverse cardiovascular events (MACE) were considered as stroke, heart failure resistant to treatment, sustained ventricular tachycardia, implantable cardioverter‐defibrillator, sudden death, and cardiovascular death. The following findings were also included in total evens: heart failure, bradycardia, ventricular arrhythmia, new conduction system abnormalities, and new echocardiographic abnormalities. Multivariate analysis with logistic regression was used in order to assess the Doppler and DTI parameters predicting events. Variables with a P‐value ≤.5 in the univariate analysis were included in the multivariate analysis. Results: In patients with chronic ChD, the analysis of DTI parameters showed that S′ wave and E′ wave of the lateral wall of the left ventricle were significant predictors of MACE (OR: 0.83; 95% CI: 0.71–0.96; P‐value:.015 and OR: 0.80; 95% CI: 0.66–0.98; P‐value:.031, respectively). Conclusions: This study found that patients with chronic ChD who had events showed significantly lower parameters in the DTI. What is more, this study showed that even lower DTI parameters are significant predictors of events. [ABSTRACT FROM AUTHOR]

In the late phase of Trypanosoma cruzi infection, parasite persistence and an exaggerated immune response accompanied by oxidative stress play a crucial role in the genesis of Chronic Chagasic Cardiomyopathy (CCC). Current treatments (Benznidazole (BNZ) and Nifurtimox) can effect only the elimination of the parasite, but are ineffective for late stage treatment and for preventing heart damage and disease progression. In vivo trypanocidal and cardioprotective activity has been reported for Lippia alba essential oils (EOs), ascribed to their two major terpenes, limonene and caryophyllene oxide. To investigate the role of antioxidant and immunomodulatory mechanisms behind these properties, chronic-T. cruzi-infected rats were treated with oral synergistic mixtures of the aforementioned EOs. For this purpose, the EOs were optimized through limonene-enrichment fractioning and by the addition of exogenous caryophyllene oxide (LIMOX) and used alone or in combined therapy with subtherapeutic doses of BNZ (LIMOXBNZ). Clinical, toxicity, inflammatory, oxidative, and parasitological (qPCR) parameters were assessed in cardiac tissue. These therapies demonstrated meaningful antioxidant and immunomodulatory activity on markers involved in CCC pathogenesis (IFN-γ, TNF-α, IL-4, IL-10, and iNOS), which could explain their significant trypanocidal properties and their noteworthy role in preventing, and even reversing, the progression of cardiac damage in chronic Chagas disease.

Purpose: To describe and give an estimation of the prevalence of urinary disorders in chronic Chagas disease, since most clinical research has been centered on the description of the cardiac and digestive forms. Methods: To explore this topic, a cross-sectional study was conducted in 137 Bolivian adults of both sexes suffering from symptomatic chronic Chagas disease. All patients presenting confirmed chagasic cardiomyopathy, megacolon or both underwent a urologic symptom questionnaire, uroflowmetry, urinary tract ultrasonography and a creatinine assay. When urinary abnormality was detected, a complete urodynamic study was proposed including cystometry, pressure-flow studies and urethral pressure profile. Results: Out of all study patients, 35 (26%) had a Chagas cardiomyopathy, 81 (59%) a megacolon, and 21 (15%) a megacolon associated with cardiomyopathy. In all, 63% presented urinary disorders defined by IPSS > 7 and/or ICIQ SF > 1. Among them, 62% were incontinent, mainly by bladder overactivity, and 45% presented grade 2 or 3 renal insufficiency. Of 49 patients, the urodynamic study identified 34 patients with detrusor overactivity (69%), mostly in those with Chagas megacolon. Median bladder functional capacity, urethral closure pressure and bladder compliance had normal values. Moreover, 36% of these patients presented moderate hypocontractility, without significant post-void residual. Conclusions: This study evidenced lower urinary tract dysfunction in a majority of chronic chagasic patients; those presenting megacolon were more likely to suffer from urinary incontinence. These results strongly suggest including routine urological clinical investigation in chronic Chagas patients, as urinary incontinence due to overactive bladder is frequently observed in this population. [ABSTRACT FROM AUTHOR]

This study aimed to evaluate well-documented diagnostic antigens, named B13, 1F8 and JL7 recombinant proteins, as potential markers of seroconversion in treated chagasic patients. Prospective study, involving 203 patients treated with benznidazole, was conducted from endemic areas of northern Argentina. Follow-up was possible in 107 out of them and blood samples were taken for serology and PCR assays before and 2, 3, 6, 12, 24 and 36 months after treatment initiation. Reactivity against Trypanosoma cruzi lysate and recombinant antigens was measured by ELISA. The rate of decrease of antibody titers showed nonlinear kinetics with an abrupt drop within the first three months after initiation of treatment for all studied antigens, followed by a plateau displaying a low decay until the end of follow-up. At this point, anti-B13, anti-1F8 and anti-JL7 titers were relatively close to the cut-off line, while anti-T. cruzi antibodies still remained positive. At baseline, 60.8% (45/74) of analysed patients tested positive for parasite DNA by PCR and during the follow-up period in 34 out of 45 positive samples (75.5%) could not be detected T. cruzi DNA. Our results suggest that these antigens might be useful as early markers for monitoring antiparasitic treatment in chronic Chagas disease.

Chagas disease (CD), a neglected tropical disease caused by the protozoan Trypanosoma cruzi, affects around six million individuals in Latin America. Currently, CD occurs worldwide, becoming a significant public health concern due to its silent aspect and high morbimortality rate. T. cruzi presents different escape strategies which allow its evasion from the host immune system, enabling its persistence and the establishment of chronic infection which leads to the development of chronic Chagas cardiomyopathy (CCC). The potent immune stimuli generated by T. cruzi persistence may result in tissue damage and inflammatory response. In addition, molecular mimicry between parasites molecules and host proteins may result in cross-reaction with self-molecules and consequently in autoimmune features including autoantibodies and autoreactive cells. Although controversial, there is evidence demonstrating a role for autoimmunity in the clinical progression of CCC. Nevertheless, the exact mechanism underlying the generation of an autoimmune response in human CD progression is unknown. In this review, we summarize the recent findings and hypotheses related to the autoimmune mechanisms involved in the development and progression of CCC.

Summary: Trypanosoma cruzi, the aetiological agent of Chagas disease, has a highly efficient detoxification system to deal with the oxidative burst imposed by its host. One of the antioxidant enzymes involved is the cytosolic tryparedoxin peroxidase (c‐TXNPx), which catalyses the reduction to hydrogen peroxide, small‐chain organic hydroperoxides and peroxynitrite. This enzyme is present in all parasite stages, and its overexpression renders parasites more resistant to the oxidative defences of macrophages, favouring parasite survival. This work addressed the study of the specific humoral and cellular immune response triggered by c‐TXNPx in human natural infection. Thus, sera and peripheral blood mononuclear cells (PBMC) were collected from chronically infected asymptomatic and cardiac patients, and non‐infected individuals. Results showed that levels of IgG antibodies against c‐TXNPx were low in sera from individuals across all groups. B‐cell epitope prediction limited immunogenicity to a few, small regions on the c‐TXNPx sequence. At a cellular level, PBMC from asymptomatic and cardiac patients proliferated and secreted interferon‐γ after c‐TXNPx stimulation, compared with mock control. However, only proliferation was higher in asymptomatic patients compared with cardiac and non‐infected individuals. Furthermore, asymptomatic patients showed an enhanced frequency of CD19+CD69+ cells upon exposure to c‐TXNPx. Overall, our results show that c‐TXNPx fails to induce a strong immune response in natural infection, being measurable only in those patients without any clinical symptoms. The low impact of c‐TXNPx in the human immune response could be strategic for parasite survival, as it keeps this crucial antioxidant enzyme activity safe from the mechanisms of adaptive immune response. c‐TXNPx induces proliferation and interferon‐γ secretion of peripheral blood mononuclear cells from asymptomatic and cardiac patients with chronic Chagas disease, with only proliferation being significantly higher in patients with no clinical symptoms. Furthermore, asymptomatic patients show an increment of CD19+ CD69+ cells upon exposure to c‐TXNPx. At a humoral level, c‐TXNPx fails to induce a strong IgG response in patients with chronic Chagas disease. [ABSTRACT FROM AUTHOR]

Chagas disease (CD), a neglected tropical disease caused by the protozoan Trypanosoma cruzi , affects around six million individuals in Latin America. Currently, CD occurs worldwide, becoming a significant public health concern due to its silent aspect and high morbimortality rate. T. cruzi presents different escape strategies which allow its evasion from the host immune system, enabling its persistence and the establishment of chronic infection which leads to the development of chronic Chagas cardiomyopathy (CCC). The potent immune stimuli generated by T. cruzi persistence may result in tissue damage and inflammatory response. In addition, molecular mimicry between parasites molecules and host proteins may result in cross-reaction with self-molecules and consequently in autoimmune features including autoantibodies and autoreactive cells. Although controversial, there is evidence demonstrating a role for autoimmunity in the clinical progression of CCC. Nevertheless, the exact mechanism underlying the generation of an autoimmune response in human CD progression is unknown. In this review, we summarize the recent findings and hypotheses related to the autoimmune mechanisms involved in the development and progression of CCC. [ABSTRACT FROM AUTHOR]

Introduction: In Brazil, an estimated 1.1 million people are infected with Trypanosoma cruzi , the causative agent of Chagas disease (CD). Despite the high number of cases, the estimated prevalence of infection per 100 inhabitants is low (0.03). However, the actual number of chronically infected individuals is still unknown. Therefore, we sought to determine the prevalence of chronic CD in at-risk individuals in Caraíbas (Bahia, Brazil) through active case finding., Methods: A total of 572 individuals living in rural or urban areas of Caraíbas were eligible for the study. A serum sample was collected from 226 individuals, and the diagnosis performed according to international guidelines., Results: The overall prevalence of anti- T. cruzi IgG was 4.42%. The median age of anti- T. cruzi IgG-positive individuals was 54.5 years, and the female-to-male ratio was 1.5:1. The prevalence of anti- T. cruzi IgG was similar in rural (4.29%) and urban areas (4.65%)., Discussion: Compared with national estimates, we concluded that Caraíbas had a high prevalence for chronic CD and a high risk for persistent transmission. Through our study, it was possible to monitor individuals who were unaware of their clinical condition, thus improving their quality of life., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Pavan, Dias, Cangussú, Dutra, Sampaio and Santos.)

Introduction CD4+CD25+ T lymphocytes have been implicated in the regulation of host inflammatory response against Trypanosoma cruzi, and may be involved in the clinical course of the disease. Methods Peripheral blood mononuclear cells from patients with chronic Chagas disease were cultured in the presence of T. cruzi recombinant antigens and assayed for lymphocytes at distinct time points. Results It was possible to differentiate clinical forms of chronic Chagas disease at days 3 and 5 according to presence of CD4+CD25+ T cells in cell cultures. Conclusions Longer periods of cell culture proved to be potentially valuable for prospective evaluations of CD4+CD25+ T lymphocytes in patients with chronic Chagas disease.

In the indeterminate chronic period of Chagas disease (ChD) the treatment has not been conclusive, because the serological negativization requires many years. This study aims to evaluate the efficacy of nifurtimox (NF) in the treatment of chronic ChD in prolonged follow-up by serological techniques of indirect immunofluorescence assay (IFA) and enzyme-linked immunosorbent assay (ELISA) IgG comparing 2 groups of patients, treated and non treated. Mann-Whitney test was performed for ELISA and IFA, with significant difference between the groups (P<0.05). IgG levels were lower in individuals treated compared with untreated patients, indicating chemotherapeutic efficacy in prolonged follow-up. [ABSTRACT FROM AUTHOR]

We evaluate the elimination of the microscopic stage of conventional xenodiagnosis (XD) to optimize the parasitological diagnosis of Trypanosoma cruzi in chronic Chagas disease. To this purpose we applied under informed consent two XD cages to 150 Chilean chronic chagasic patients. The fecal samples (FS) of the triatomines at 30, 60 and 90 days post feeding were divided into two parts: in one a microscopic search for mobile trypomastigote and/or epimastigote forms was performed. In the other part, DNA extraction-purification for PCR directed to the conserved region of kDNA minicircles of trypanosomes (PCR-XD), without previous microscopic observation was done. An XD was considered positive when at least one mobile T. cruzi parasite in any one of three periods of incubation was observed, whereas PCR-XD was considered positive when the 330 bp band specific for T. cruzi was detected. 25 of 26 cases with positive conventional XD were PCR-XD positive (concordance 96.2%), whereas 85 of 124 cases with negative conventional XD were positive by PCR-XD (68.5%). Human chromosome 12 detected by Real-time PCR used as exogenous internal control of PCR-XD reaction allowed to discounting of PCR inhibition and false negative in 40 cases with negative PCR-XD. Conclusion: PCR-XD performed without previous microscopic observation is a useful tool for detection of viable parasites with higher efficiency then conventional XD.

Currently there are no biological markers to indicate which individuals with chronic indeterminate period of Chagas disease develop heart disease and who will remain all his life in this phase. The aim of this survey was to determine if T rypanosoma cruzi burden is related to the presence of heart disease in patients with chronic Chagas disease. 200 patients who had not been treated, 100 with cardiopathy and 100 without, groups A and B respectively, were submitted to clinical study and electrocardiogram, Echo-Doppler was performed for group A in which all important known causes of cardiopathy were discarded. In both groups xenodiagnosis, conventional PCR and quantitative PCR were undertaken. The 100 cardiopaths had 133 electrocardiographic alterations most of them in grade II of the New York Heart Association classification. 98 cardiopaths were classified in grade I by Echo-Doppler and only 2 cases were in grade III due to low ejection fraction. The difference in average parasitemia in patients of group A and B was not significant and no statistically differences were observed between average parasitemia of cardiopaths grade II versus grade I of NYHA. This results allow to characterize same clinical, electrocardiographical and parasitological features in chagasic cardiopaths of Chile. [ABSTRACT FROM AUTHOR]

Background: Trypanosoma cruzi multiplies and differentiates in the digestive tract of triatomine insects. Xenodiagnosis (XD) is a parasitological tool in which the insect vectors acts as a biological culture medium to amplify and detect T. cruzi infection in mammals. The sensitivity of XD has been overcome by the application of PCR in fecal samples (FS) of XD (PCR-XD). In this study, T. cruzi amplified in Triatoma infestans fed by XD on individuals with chronic Chagas disease (CChD) is quantified by real-time PCR (qPCR-XD). Findings: Under informed consent, 100 individuals were evaluated. In 21 of them XD, PCR-XD and qPCR-XD were positive. For the contrary, 79 were negative XD. In 58 (73.4 %) and 66 cases (83.5 %) of them, PCR-XD (Fisher's exact test P = 0.005) and qPCR-XD (Fisher's exact test: P = 0.037) respectively, were positive. In cases with positive XD, qPCR-XD allowed to establish that in 9/21 cases (42.9 %) the parasite burden fluctuated between 100 and 1,000 par. eq./ml. Otherwise, in 32/79 (40.5 %) cases with negative XD, a parasite burden between 1 and 10 par. eq./ml was determined. All samples showed amplification of exogenous internal control (X12, Ct average: 31.8), so problems in the DNA extraction (excess or loss of genetic material), unspecific amplification and/or inhibition in qPCR-XD reactions were ruled out. Additionally, in all the patients qPCR in blood (qPCR-B) was performed. In the cases with positive XD, the concordance between the positivity of qPCR-XD and qPCR-B was 100 %, nevertheless, the parasite burden in blood was lower and different than XD (Chi-square test: χ² =91.82,df =5, P = 0.0001). In the cases with negative XD the ranges of qPCR-XD and qPCR-B were similar (Chi-square test: χ² =6.71, df =5, P = 0.1520). Conclusions: This study allowed the detection and quantification of T. cruzi by qPCR-XD in FS of Tr. infestans fed on patients with CChD. The highest parasite burden was observed in positive XD cases. qPCR-XD could be used in different studies related with the complex T. cruzi-vector-host interactions. [ABSTRACT FROM AUTHOR]

Trypanosoma cruzi is mainly transmitted by blood-sucking triatomines, but other routes also have epidemiological importance, such as blood transfusion and congenital transmission. Although the possibility of sexual transmission of T . cruzi has been suggested since its discovery, few studies have been published on this subject. We investigated acquisition of T . cruzi by sexual intercourse in an experimental murine model. Male and female mice in the chronic phase of Chagas disease were mated with naive partners. Parasitological, serological and molecular tests demonstrated the parasites in tissues and blood of partners. These results confirm the sexual transmission of T . cruzi in mice. [ABSTRACT FROM AUTHOR]

PCR and FC-ALTA were used to monitor parasite clearance in 54 chronic chagasic patients who had completed therapy with allopurinol (ALLO, n = 31) or itraconazole (ITRA, n = 23) ten years earlier. All patients maintained positive conventional serology. 25 of them showed positive XD (ALLO, n = 11 and ITRA, n = 14) and 29 negative XD (ALLO, n = 20 and ITRA, n = 9). 43 patients were positive by both techniques (ALLO, n = 23 and ITRA, n = 20). Seven of 54 patients were negative by PCR and positive by FC-ALTA and three of 54 were positive by PCR and negative by FC-ALTA. Only one case with both tests negative should be considered cured. Of 29 patients with negative XD, 14 treated ALLO (70 %) and nine with ITRA (77.8 %) showed positive PCR and FC-ALTA. These results do not show differences of efficacy among the drugs, and reinforce the relevance of using sensitive tools such as PCR and FC-ALTA for the follow-up of patients with chronic Chagas disease.

Purpose: Regulatory T cells are involved in the clinical course of chronic Chagas disease, possibly because they exercise a control in the patient's inflammatory response to Trypanosoma cruzi. This study analyzed the levels of CD4 + CD25+ T cells in chronic Chagas disease patients after in vitro stimulation of the peripheral blood mononuclear cells with CRA (Cytoplasmic Repetitive Antigen) or FRA (Flagellar Repetitive Antigen) T. cruzi antigens. Methods: Groups of patients with the cardiac form and indeterminate form; and non-infected individuals, were selected. The CD4 + CD25+ T lymphocyte population, as well as the FoxP3 expression and the IL10 production, were evaluated by flow cytometry after stimulation with CRA or FRA. Result: The IND group presented higher levels of CD4 + CD25+ T cells than the CARD group. However, there was no evidence of a relationship between FoxP3 and IL10 with any of the chronic forms. Conclusions: Our results suggest the possible involvement of CD4 + CD25+ T cells specific to CRA and FRA in controlling the progression of clinical outcomes. Though, further studies are needed to define which mechanisms activate regulatory T cells and lead to pathology control in chronic human Chagas disease. [ABSTRACT FROM AUTHOR]

Antibodies (Ab) recognizing G-protein coupled receptors, such as β1 and β2 adrenergic (anti-β1-AR and anti-β2-AR, respectively) and muscarinic cholinergic receptors (anti-M2-CR) may contribute to cardiac damage, however their role in chronic chagasic cardiomyopathy is still controversial. We describe that Trypanosoma cruzi-infected C3H/He mice show increased P and QRS wave duration, and PR and QTc intervals, while the most significant ECG alterations in C57BL/6 are prolonged P wave and PR interval. Echocardiogram analyses show right ventricle dilation in infected animals of both mouse lineages. Analyses of heart rate variability (HRV) in chronically infected C3H/He mice show no alteration of the evaluated parameters, while C57BL/6 infected mice display significantly lower values of HRV components, suggesting autonomic dysfunction. The time-course analysis of anti-β1-AR, anti-β2-AR and anti-M2-CR Ab titres in C3H/He infected mice indicate that anti-β1-AR Ab are detected only in the chronic phase, while anti-β2-AR and anti-M2-CR are observed in the acute phase, diminish at 60 dpi and increase again in the chronic phase. Chronically infected C57BL/6 mice presented a significant increase in only anti-M2-CR Ab titres. Furthermore, anti-β1-AR, anti-β2-AR and anti-M2-CR, exhibit significantly higher prevalence in chronically T. cruzi-infected C3H/He mice when compared with C57BL/6. These observations suggest that T. cruzi infection leads to host-specific cardiac electric alterations. [ABSTRACT FROM AUTHOR]

Treatment for Chagas disease has limited efficacy in the chronic phase. We evaluated benznidazole (BZ) and itraconazole (ITZ) individually and in association in dogs 16 months after infection with a BZ-resistant Trypanosoma cruzi strain. Four study groups (20 animals) were evaluated and treated for 60 days with BZ, ITZ, or BZ + ITZ, and maintained in parallel to control group infected and not treated (INT). All dogs were evaluated in the first, sixth, 12th, 18th and 24th months of study. Polymerase chain reaction (PCR) was negative in 2 of 3 animals in the BZ + ITZ group, 2 of 5 in the BZ group, and 4 of 5 in the ITZ group. Hemoculture performed in the 24th month was negative in all groups. Enzyme-linked immunoassay remained reactive in all treated animals. Echocardiography differentiated treated animals from control animals. Quantitative PCR analysis of cardiac tissue was negative in the BZ + ITZ and BZ groups, positive in 2 of 5 dogs in the ITZ group and in 2 of 3 dogs in the control group, but negative in colon tissue in all groups. Inflammation was significantly reduced in the right atrium and left ventricle of dogs treated with BZ + ITZ and BZ compared with those receiving ITZ alone. Fibrosis was absent in most dogs treated with BZ + ITZ, mild in those treated with BZ or ITZ alone, and intense in the control group. Parasitological and histopathological evaluations showed that BZ + ITZ treatment improved or stabilized the clinical condition of the dogs. [Display omitted] • Comparison of Benznidazole + Itraconazole and monotherapy in the chronic T. cruzi infection of dogs. • Drugs association showed more reduction of parasitemia, inflammation, fibrosis and mortality. • Echocardiograph differentiated the ITZ group from BZ group, but not all groups from each other. • BZ + ITZ showed less lesions and clinical stability than monotherapy and control group. [ABSTRACT FROM AUTHOR]

Previously, our group showed a prothrombotic state in asymptomatic patients with chronic Chagas disease. The current paper studies the inflammatory status and endothelial function in these patients. Methods: In 40 patients and 40 healthy volunteers, we evaluated prothrombotic state, blood parasitemia (molecular biology: polymerized chain reaction [PCR]-amplification), tissue factor pathway inhibitor antibodies (aTFPI), interleukin 6 (IL-6), and vascular cell adhesion molecule-1 (VCAM-1). Endothelial function was determined by reactive hyperemia (pulse plethysmography). Results: In patients, prothrombin fragment 1 + 2, d-dimer, PAI-1, and fibrinogen were higher. Amplification of 121/122 primers (Trypanosomacruzi) was positive in 45% of the patients. Patients presented higher values of aTFPI- immunoglobulin G (IgG; P < .05), aTFPI-IgM (P < .001), IL-6 (P = .004), and VCAM-1 (P = .00001). In both groups, endothelial function was preserved. Conclusions: We found that asymptomatic patients with chronic Chagas disease presented a prothrombotic/inflammatory status. The fact that endothelial function is still preserved suggests that prothrombosis and inflammation are primarily implicated in the beginning of cardiovascular damage. [ABSTRACT FROM PUBLISHER]

Abstract: PCR and Southern blot hybridization were used to determine the distribution of Trypanosoma cruzi clones in 37 chronic chagasic cardiopathic and non-cardiopathic patients. Parasite DNA amplified from peripheral blood or dejections of Triatoma infestans fed on patient blood was hybridized with probes containing hypervariable minicircle nucleotide sequences capable of detecting three sublineages of T. cruzi. Probes Z-I and Z-IIb detect unique sequences in lineages TcI and TcIIb, respectively. Probe Z-hybrid detects sequences of lineages TcIId and TcIIe. T. cruzi clones of the Z-I sublineage were detected in 62.2% of T. infestans dejections and 5.4% of peripheral blood samples. Clones of Z-IIb and Z-hybrid sublineages had similar distribution in blood and dejection samples. Interestingly, clones of the Z-IIb sublineage were significantly lower in cardiopathic than in non-cardiopathic patients (23.5% versus 75%; P =0.0006). Clones of the Z-hybrid sublineage were found in 29.4% of cardiopathic and 75% of non-cardiopathic patients, respectively (P =0.0051). By contrast, clones of sublineage Z-I were similarly distributed in both groups of patients. The low frequency of Z-IIb and Z-hybrid sublineage clones detected in cardiopathic patients suggests that the immunological mechanisms involved in controlling and eliminating these T. cruzi parasites may be detrimental to the host, leading to the development of chagasic cardiomyopathy. [Copyright &y& Elsevier]

Abstract: In this study, we compare the sensitivity of detecting Trypanosoma cruzi in dejections of Triatoma infestans nymphs that had fed on the blood of chronic chagasic patients, with detection of T. cruzi in peripheral blood, using a polymerase chain reaction assay (PCR-D and PCR-B, respectively). Fifty-seven chronic patients were evaluated who were positive (group I) or negative (group II) by xenodiagnosis (XD). Patients showed 84.8 and 75% positive PCR results in both kinds of samples in groups I and II, respectively. Six cases (10.5%) showed positive PCR-D and negative PCR-B, five of them belonged to group I. In contrast, five cases of group II showed negative PCR-D and positive PCR-B. Overall, the PCR-D assay gave positive results in 52 out of 57 samples (91.2%), while 51 out of 57 (89.5%) were positive by PCR-B. In comparison, only 57.9% were positive by XD (p =0.0001). In conclusion, PCR performed in dejection or blood was more sensitive for the parasite detection than xenodiagnosis. All patients (100%) were detected positive when both, PCR-D and PCR-B, were applied. [Copyright &y& Elsevier]

The presence of Trypanosoma cruzi in chronic chagasic patients with negative xenodiagnosis (XD) after 6 years following completion of therapy with either itraconazole or allopurinol was assessed by polymerase chain reaction (PCR) and hybridization assays. A 330-bp DNA fragment amplified from the hypervariable regions of T. cruzi kinetoplastid minicircles was hybridized with total 32P-labeled kinetoplast DNA as probes. PCR alone enabled the identification of T. cruzi nucleotide sequences in 40% of the patients treated with itraconazole and in 60% of patients treated with allopurinol. PCR used in combination with hybridization detected parasite DNA in 60% and 53% of XD negative individuals treated with itraconazole or allopurinol, respectively. These results show that PCR and hybridization are more sensitive than conventional parasitological techniques in diagnosing patients that have undergone chemotherapy with itraconazole or allopurinol. [Copyright &y& Elsevier]