Main Cardiac Histopathologic Alterations in the Acute Phase of Trypanosoma cruzi Infection in a Murine Model.

Symptoms in the acute phase of Chagas disease are usually mild and nonspecific. However, after several years, severe complications like dilated heart failure and even death may arise in the chronic phase. Due to the lack of specific symptoms in the acute phase, the aim of this work was to describe and analyze the cardiac histopathology during this phase in a CD1 mouse model by assessing parasitism, fibrotic damage, and the presence and composition of a cellular infiltrate, to determine its involvement in the pathogenesis of lesions in the cardiac tissue. Our results indicate that the acute phase lasts about 62 days post-infection (dpi). A significant increase in parasitemia was observed since 15 dpi, reaching a maximum at 33 dpi (4.1 × 106). The presence of amastigote nests was observed at 15-62 dpi, with a maximum count of 27 nests at 35 dpi. An infiltrate consisting primarily of macrophages and neutrophils was found in the cardiac tissue within the first 30 days, but the abundance of lymphocytes showed an 8 ≥ fold increase at 40-62 dpi. Unifocal interstitial fibrosis was identified after 9 dpi, which subsequently showed a 16 ≥ fold increase at 40-60 dpi, along with a 50% mortality rate in the model under study. The increased area of fibrotic lesions revealed progression in the extent of fibrosis, mainly at 50-62 dpi. The presence of perivasculitis and thrombus circulation disorders was seen in the last days (62 dpi); finally, cases of myocytolysis were observed at 50 and 62 dpi. These histopathological alterations, combined with collagen deposition, seem to lead to the development of interstitial fibrosis and damage to the cardiac tissue during the acute phase of infection. This study provides a more complete understanding of the patterns of histopathological abnormalities involved in the acute phase, which could help the development of new therapies to aid the preclinical tests of drugs for their application in Chagas disease.

Immunopathological Mechanisms Underlying Cardiac Damage in Chagas Disease.

In Chagas disease, the mechanisms involved in cardiac damage are an active field of study. The factors underlying the evolution of lesions following infection by Trypanosoma cruzi and, in some cases, the persistence of its antigens and the host response, with the ensuing development of clinically observable cardiac damage, are analyzed in this review.

Response to Infection by Trypanosoma cruzi in a Murine Model.

Cardiopathy is a common, irreversible manifestation of the chronic phase of Chagas disease; however, there is controversy as to how the causes for progression from the acute to the chronic phase are defined. In this work, the presence of the parasite is correlated with the occurrence of cell infiltration and fibrosis in cardiac tissues, as well as IgG detection and disease progression in a murine model. Fifty CD1 mice were infected intraperitoneally with Trypanosoma cruzi, while 30 control were administered with saline solution. Parasitemia levels were determined, and IgG titers were quantified by ELISA. At different times, randomly selected mice were euthanized, and the heart was recovered. Cardiac tissue slides were stained with HE and Masson trichrome stain. A significant increase in parasitemia levels was observed after 15 days post-infection (dpi), with a maximum of 4.1 × 106 parasites on 33 dpi, ending on 43 dpi; amastigote nests were observed on 15-62 dpi. Histological analysis revealed lymphocytic infiltration and fibrotic lesions from 8 dpi until the end of the study, on 100 dpi. The presence of plasma cells in the myocardium observed on 40-60 dpi, accompanied by seropositivity to ELISA on 40-100 dpi, was regarded as the hallmark of the transition phase. Meanwhile, the chronic phase, characterized by the absence of amastigotes, presence of cell infiltration, fibrotic lesions, and seropositivity, started on 62 dpi. A strong correlation between parasitemia and the presence of amastigote nests was found (r 2 = 0.930), while correlation between the presence of fibrosis and of amastigote nests was weak (r 2 = 0.306), and that between fibrosis and lymphocyte infiltration on 100 dpi was strong (r 2 = 0.899). The murine model is suitable to study Chagas disease, since it can reproduce the chronic and acute phases of the human disease. The acute phase was determined to occur on 1-60 dpi, while the chronic phase starts on 62 dpi, and fibrotic damage is a consequence of the continuous inflammatory infiltration; on the other hand, fibrosis was determined to start on the acute phase, being more apparent in the chronic phase, when Chagas disease-related cardiopathy is induced.

Coinfection by Trypanosoma cruzi and a fungal pathogen increases survival of Chagasic bugs: advice against a fungal control strategy.

Triatomine bugs carry the parasitic protozoa Trypanosoma cruzi, the causal agent of Chagas disease. It is known that both the parasite and entomopathogenic fungi can decrease bug survival, but the combined effect of both pathogens is not known, which is relevant for biological control purposes. Herein, the survival of the triatomine Meccus pallidipennis (Stal, 1872) was compared when it was coinfected with the fungus Metarhizium anisopliae (Metschnikoff) and T. cruzi, and when both pathogens acted separately. The immune response of the insect was also studied, using phenoloxidase activity in the bug gut and hemolymph, to understand our survival results. Contrary to expectations, triatomine survival was higher in multiple than in single challenges, even though the immune response was lower in cases of multiple infection. We postulate that T. cruzi exerts a protective effect and/or that the insect reduced the resources allocated to defend itself against both pathogens. Based on the present results, the use of M. anisopliae as a control agent should be re-considered.

Identification of O-Glcnacylated Proteins in Trypanosoma cruzi.

Originally an anthropozoonosis in the Americas, Chagas disease has spread from its previous borders through migration. It is caused by the protozoan Trypanosoma cruzi. Differences in disease severity have been attributed to a natural pleomorphism in T. cruzi. Several post-translational modifications (PTMs) have been studied in T. cruzi, but to date no work has focused on O-GlcNAcylation, a highly conserved monosaccharide-PTM of serine and threonine residues mainly found in nucleus, cytoplasm, and mitochondrion proteins. O-GlcNAcylation is thought to regulate protein function analogously to protein phosphorylation; indeed, crosstalk between both PTMs allows the cell to regulate its functions in response to nutrient levels and stress. Herein, we demonstrate O-GlcNAcylation in T. cruzi epimastigotes by three methods: by using specific antibodies against the modification in lysates and whole parasites, by click chemistry labeling, and by proteomics. In total, 1,271 putative O-GlcNAcylated proteins and six modification sequences were identified by mass spectrometry (data available via ProteomeXchange, ID PXD010285). Most of these proteins have structural and metabolic functions that are essential for parasite survival and evolution. Furthermore, O-GlcNAcylation pattern variations were observed by antibody detection under glucose deprivation and heat stress conditions, supporting their possible role in the adaptive response. Given the numerous biological processes in which O-GlcNAcylated proteins participate, its identification in T. cruzi proteins opens a new research field in the biology of Trypanosomatids, improve our understanding of infection processes and may allow us to identify new therapeutic targets.

Th-17 cytokines are associated with severity of Trypanosoma cruzi chronic infection in pediatric patients from endemic areas of Mexico.

In Chagas disease the clinical, acute and chronic manifestations are the result of the interaction between the parasite and the host factors. The balance between inflammatory and anti-inflammatory immune responses is essential for the increase or resolution of the manifestations in individuals infected with T. cruzi. To identify if children with chronic Chagas disease and heart injury is related with non-regulated Th1, Th2 and Th17 responses. We included 31 children with T. cruzi confirmed chronic infection from endemic areas of Mexico. Subsequently, they were separated according to their ECHO and ECG results into three groups according to the severity of cardiac involvement. Circulating Th1, Th2 and Th17 cytokine profiles were performed by Luminex assays and the results were analyzed by bivariate and multivariable analysis. Patients were classified in asymptomatic chronic (group 1, N=12); individuals with IRBBB in ECG and incipient lesions in ECHO (Group 2, N=8) and Patients with severe chronic symptomatic disease (Group 3, N=11). The analysis of immune mediators revealed that patients with severe cardiac manifestations had significant higher levels (p <0.05) of Th17 related cytokines including IL-17 and IL-6 as well as IFN-γ and IL-2. Also patients with severe cardiomyopathy exhibit increased levels of IL-13 (p <0.05) after multivariate analysis. High levels of Th17 related cytokines including IL-17, IFN-γ, IL-6 and IL-2 and pro-fibrotic factors such as IL-13 could be associated to the severity of cardiac involvement in children with chronic T. cruzi infection. These cytokines could be useful as indicators for the early identification of cardiac damage associated to the T. cruzi infection.

Chagas Disease in Mexico: Report of 14 Cases of Chagasic Cardiomyopathy in Children.

Chagas disease is a parasitic infection mainly found in Latin America; it is transmitted by a triatomine, also known as assassin bug or kissing bug. In humans, the parasite causes mostly cardiac disorders. Two-thirds of the Mexican territory are regarded as risk areas for vector transmission of Trypanosoma cruzi, the causal agent. The parasite can be found as a blood-borne trypomastigote or as an intracellular amastigote. The progression and severity of lesions could be due to frequent reinfections or to infection by highly virulent strains. A total of 3,327 individuals younger than 18 years old, living in risk areas for this disease in the rural setting of the States of Queretaro, San Luis Potosi, and Veracruz, underwent a seroepidemiological study. Among them, 37 subjects were seropositive for T. cruzi, and were studied to look for signs of cardiac pathology, which has only been reported in adults. A clinical record was prepared for all included individuals, and electrocardiography (ECG) and echocardiography (ECHO) studies were performed; 25 cases showed lesions compatible with the onset of Chagas cardiomyopathy. The other 12 patients showed either normal ECG and ECHO data or showed abnormal parameters that were not regarded as significant. Lesions found in the onset of Chagas cardiomyopathy in children are herein reported, along with 14 cases of cardiac pathology compatible with Chagas disease. Our results indicate that patients younger than 18 years can show a cardiac pathology similar to that observed in adults.

Enfermedad de Chagas en México / Chagas disease in Mexico

Resumen México es un país endémico para la enfermedad de Chagas, donde dos terceras partes del territorio pueden ser consideradas en riesgo de transmisión vectorial, es decir que 1'100,000 individuos podrían estar infectados con Trypanosoma cruzi y 29'500,000 en riesgo de contraer la infección. En la morbimortalidad del padecimiento son importantes las características de la vivienda, condiciones biológicas, ambientales y factores socioculturales. El tamizaje en bancos de sangre, a la fecha, es de observancia obligatoria con una cobertura mayor al 92%. El diagnóstico no se establece frecuentemente debido al desconocimiento de la enfermedad por parte del personal de salud y de la población. La fase aguda generalmente pasa desapercibida y en la crónica, la patología se presenta principalmente en el corazón, con evolución lenta. La patogénesis de la miocardiopatía crónica es muy compleja y se presentan lesiones con mayor frecuencia en el sistema nervioso autónomo y miocardio, lo que genera trastornos en la conductibilidad y contractilidad del órgano. Se describen los principales mecanismos patogénicos implicados en el desarrollo de la enfermedad.

Abstract Mexico is a country endemic for Chagas disease in which two thirds of the territory can be considered at risk of vector-borne infection. This means that 1.1 million people could be infected with Trypanosoma cruzi and 29.5 million at risk of infection. Dwelling characteristics of poverty in these rural areas linked with biological conditions, lifestyle, environmental and sociocultural factors are important in the morbidity and mortality of the disease. Nowadays, the screening for the parasite is mandatory and at least 92% of blood banks are covered. The inadequate knowledge of the disease by the health personnel and the population limits the possibility of the diagnosis. The acute phase of the disease courses undetected. The main affected organ in Chagas disease is the heart, with a slow evolution; the pathogenesis of chronic cardiomyopathy is complex and lesions occur mainly in the autonomic nervous system and myocardium leading to disturbances in the conductivity and contractility of the organ. The main pathogenic mechanisms involved in the development of the disease are described.

[Identification of immunodominant components of an isolate of Trypanosoma cruzi by immunoblot and its standardization for diagnostic purposes]. / Identificación de componentes inmunodominantes de un aislado de Trypanosoma cruzi por inmunoblot y su estandarización con fines diagnósticos.

INTRODUCTION: Conventional serology was used for the detection of Trypanosoma cruzi infection, with diverse sensitivity and specificity results. Due to the number of samples with doubtful results, it is necessary to develop additional confirmation tests such as the immunoblot. OBJECTIVE: The aim of this study was identify major immunogenic proteins of T. cruzi isolate and establish criteria for immunoblot positivity with diagnostic purposes. METHODS: Immunoblot initial standardization was performed, determining optimal concentrations of antigen, serum, and second antibody. Thirty-five positive and thirty negative sera were assayed to evaluate different criteria of positivity and determine which provides greater sensitivity and specificity. RESULTS: Immunoblot of T. cruzi positive sera shared a rich pattern of components with molecular weights between 10-250 kDa. Twelve components had a recognition rate higher than 50%, of which the polypeptides of 27, 32, 34, and 38 kDa were close to 100%. Of the positivity criteria evaluated, the recognition of the components of 27 and 32 kDa provided sensitivity and specificity of 100%. DISCUSSION: The Immunoblot is suitable for confirmation of infection by T. cruzi, so it is strongly recommended for confirmation and discrimination of discordant cases.

Chagas disease as a cause of symptomatic chronic myocardopathy in Mexican children.

We report the first case series of children in Mexico living with symptomatic Chagas disease causing chronic myocardopathy. The findings suggest that children with Chagas disease may develop symptomatic chronic myocardopathy earlier than previously recognized. Our findings emphasize the importance of longitudinal cardiologic follow-up of all children identified with acute Chagas disease.In a cohort of 826 children from the state of Queretaro in Mexico, 11 were identified with positive serology (ELISA and IFI) for Chagas and were tested for electrocardiogram alterations and symptoms and signs. Four children had ECG alterations with 3 of these reporting signs and symptoms associated with the chronic phase of Chagas disease (27%; 95% CI: 6%-61%). The most common chronic symptom was chest pain, with one child also reporting dyspnea and tachycardia.