Trypanosoma cruzi: Biological characterization of a isolate from an endemic area and its susceptibility to conventional drugs

We describe some biological and molecular characteristics of a Trypanosoma cruzi isolate derived from a Triatomine captured in Nicaragua. PCR based typification showed that this isolate, named Nicaragua, belonged to the lineage Tc I. Nicaragua infected culture cells were treated with allopurinol, showing different behavior according to the cellular compartment, being cardiomyocyte primary cultures more resistant to this drug. The course of the infection in a mice experimental model and its susceptibility to benznidazole and allopurinol was analyzed. In benznidazole treatment, mice reverted the high lethal effect of parasites during the acute infection, however, a few parasites were detected in the heart of 88% of mice 1 year post-infection. Since T. cruzi is a heterogeneous species population it is important to study and characterize different parasites actually circulating in humans in endemic areas. In this work we show that T. cruzi Nicaragua isolate, is sensitive to early benznidazole treatment.     

Fat tissue regulates the pathogenesis and severity of cardiomyopathy in murine chagas disease

Chronic Chagas cardiomyopathy (CCC) caused by a parasite Trypanosoma cruzi is a life-threatening disease in Latin America, for which there is no effective drug or vaccine. The pathogenesis of CCC is complex and multifactorial. Previously, we demonstrated T. cruzi infected mice lose a significant amount of fat tissue which correlates with progression of CCC. Based on this an investigation was undertaken during both acute and chronic T. cruzi infection utilizing the FAT-ATTAC murine model (that allows modulation of fat mass) to understand the consequences of the loss of adipocytes in the regulation of cardiac parasite load, parasite persistence, inflammation, mitochondrial stress, ER stress, survival, CCC progression and CCC severity. Mice were infected intraperitoneally with 5x10⁴ and 10³ trypomastigotes to generate acute and chronic Chagas models, respectively. Ablation of adipocytes was carried out in uninfected and infected mice by treatment with AP21087 for 10 days starting at 15DPI (acute infection) and at 65DPI (indeterminate infection). During acute infection, cardiac ultrasound imaging, histological, and biochemical analyses demonstrated that fat ablation increased cardiac parasite load, cardiac pathology and right ventricular dilation and decreased survival. During chronic indeterminate infection ablation of fat cells increased cardiac pathology and caused bi-ventricular dilation. These data demonstrate that dysfunctional adipose tissue not only affects cardiac metabolism but also the inflammatory status, morphology and physiology of the myocardium and increases the risk of progression and severity of CCC in murine Chagas disease.     

Trypanosoma cruzi: desferrioxamine decreases mortality and parasitemia in infected mice through a trypanostatic effect

Desferrioxamine (DFO) is a potent iron chelator that is also known to modulate inflammation and act as an efficient antioxidant under normal conditions and under oxidative stress. Many in vitro and in vivo studies have shown the efficacy of DFO in the treatment of viral, bacterial and protozoan infections. DFO is known to reduce the intensity of Trypanosoma cruzi infections in mice even during a course of therapy that is not effective in maintaining anaemia or low iron levels. To further clarify these findings, we investigated the action of DFO on mouse T. cruzi infection outcomes and the direct impact of DFO on parasites.Infected animals treated with DFO (5 mg/animal/day) for 35 days, beginning 14 days prior to infection, presented lower parasitemia and lower cumulative mortality rate. No significant effect was observed on iron metabolism markers, erythrograms, leukograms or lymphocyte subsets.In the rapid method for testing in vivo T. cruzi susceptibility, DFO also induced lower parasitemia.In regard to its direct impact on parasites, DFO slightly inhibited the growth of amastigotes and trypomastigotes in fibroblast culture. Trypan blue staining showed no effects of DFO on parasite viability, and only minor apoptosis in trypomastigotes was observed. Nevertheless, a clear decrease in parasite mobility was detected.In conclusion, the beneficial actions of DFO on mice T. cruzi infection seem to be independent of host iron metabolism and free of significant haematological side effects. Through direct action on the parasite, DFO has more effective trypanostatic than trypanocidal properties.     

Trypanosoma cruzi: 4-aminopyrazolopyrimidine in the treatment of experimental Chagas' disease

An allopurinol metabolite, 4-aminopyrazolopyrimidine, was tested on two different strains of mice (NMRI-IVIC and C57Bl/6J) that had been infected 4 days earlier with the virulent Ya strain of Trypanosoma cruzi. Low doses of 4-aminopyrazolopyrimidine (0.125-0.500 mg/kg body wt/day) for 10 days induced a significant reduction in parasitemia (direct counts and subinoculation experiments) and increased survival time (without any evidence of toxicity) compared with untreated animals. When tested in vitro, 4-aminopyrazolopyrimidine was sixfold more active than allopurinol as a trypanostatic drug. The low therapeutic doses of 4-aminopyrazolopyrimidine suggest that this drug may be useful in the treatment of acute Chagas' disease.     

High Fat Diet Modulates Trypanosoma cruzi Infection Associated Myocarditis

Background

Trypanosoma cruzi, the causative agent of Chagas disease, has high affinity for lipoproteins and adipose tissue. Infection results in myocarditis, fat loss and alterations in lipid homeostasis. This study was aimed at analyzing the effect of high fat diet (HFD) on regulating acute T. cruzi infection-induced myocarditis and to evaluate the effect of HFD on lipid metabolism in adipose tissue and heart during acute T. cruzi infection.

Methodology/Principal Findings

CD1 mice were infected with T. cruzi (Brazil strain) and fed either a regular control diet (RD) or HFD for 35 days following infection. Serum lipid profile, tissue cholesterol levels, blood parasitemia, and tissue parasite load were analyzed to evaluate the effect of diet on infection. MicroPET and MRI analysis were performed to examine the morphological and functional status of the heart during acute infection. qPCR and immunoblot analysis were carried out to analyze the effect of diet on the genes involved in the host lipid metabolism during infection. Oil red O staining of the adipose tissue demonstrated reduced lipolysis in HFD compared to RD fed mice. HFD reduced mortality, parasitemia and cardiac parasite load, but increased parasite load in adipocytes. HFD decreased lipolysis during acute infection. Both qPCR and protein analysis demonstrated alterations in lipid metabolic pathways in adipose tissue and heart in RD fed mice, which were further modulated by HFD. Both microPET and MRI analyses demonstrated changes in infected RD murine hearts which were ameliorated by HFD.

Conclusion/Significance

These studies indicate that Chagasic cardiomyopathy is associated with a cardiac lipidpathy and that both cardiac lipotoxicity and adipose tissue play a role in the pathogenesis of Chagas disease. HFD protected mice from T. cruzi infection-induced myocardial damage most likely due to the effects of HFD on both adipogenesis and T. cruzi infection-induced cardiac lipidopathy.     

Identification of Three Classes of Heteroaromatic Compounds with Activity against Intracellular Trypanosoma cruzi by Chemical Library Screening

The development of new drugs against Chagas disease is a priority since the currently available medicines have toxic effects, partial efficacy and are targeted against the acute phase of disease. At present, there is no drug to treat the chronic stage. In this study, we have optimized a whole cell-based assay for high throughput screening of compounds that inhibit infection of mammalian cells by Trypanosoma cruzi trypomastigotes. A 2000-compound chemical library was screened using a recombinant T. cruzi (Tulahuen strain) expressing β-galactosidase. Three hits were selected for their high activity against T. cruzi and low toxicity to host cells in vitro: PCH1, NT1 and CX1 (IC₅₀: 54, 190 and 23 nM, respectively). Each of these three compounds presents a different mechanism of action on intracellular proliferation of T. cruzi amastigotes. CX1 shows strong trypanocidal activity, an essential characteristic for the development of drugs against the chronic stage of Chagas disease where parasites are found intracellular in a quiescent stage. NT1 has a trypanostatic effect, while PCH1 affects parasite division. The three compounds also show high activity against intracellular T. cruzi from the Y strain and against the related kinetoplastid species Leishmania major and L. amazonensis. Characterization of the anti–T. cruzi activity of molecules chemically related to the three library hits allowed the selection of two compounds with IC₅₀ values of 2 nM (PCH6 and CX2). These values are approximately 100 times lower than those of the medicines used in patients against T. cruzi. These results provide new candidate molecules for the development of treatments against Chagas disease and leishmaniasis.     

Structural Insights into Inhibition of Sterol 14α-Demethylase in the Human Pathogen Trypanosoma cruzi

Trypanosoma cruzi causes Chagas disease (American trypanosomiasis), which threatens the lives of millions of people and remains incurable in its chronic stage. The antifungal drug posaconazole that blocks sterol biosynthesis in the parasite is the only compound entering clinical trials for the chronic form of this infection. Crystal structures of the drug target enzyme, Trypanosoma cruzi sterol 14α-demethylase (CYP51), complexed with posaconazole, another antifungal agent fluconazole and an experimental inhibitor, (R)-4′-chloro-N-(1-(2,4-dichlorophenyl)-2-(1H-imid-azol-1-yl)ethyl)biphenyl-4-carboxamide (VNF), allow prediction of important chemical features that enhance the drug potencies. Combined with comparative analysis of inhibitor binding parameters, influence on the catalytic activity of the trypanosomal enzyme and its human counterpart, and their cellular effects at different stages of the Trypanosoma cruzi life cycle, the structural data provide a molecular background to CYP51 inhibition and azole resistance and enlighten the path for directed design of new, more potent and selective drugs to develop an efficient treatment for Chagas disease.     

Mesenchymal Bone Marrow Cell Therapy in a Mouse Model of Chagas Disease. Where Do the Cells Go?

Background

Chagas disease, resulting from infection with the parasite Trypanosoma cruzi (T. cruzi), is a major cause of cardiomyopathy in Latin America. Drug therapy for acute and chronic disease is limited. Stem cell therapy with bone marrow mesenchymal cells (MSCs) has emerged as a novel therapeutic option for cell death-related heart diseases, but efficacy of MSC has not been tested in Chagas disease.

Methods and Results

We now report the use of cell-tracking strategies with nanoparticle labeled MSC to investigate migration of transplanted MSC in a murine model of Chagas disease, and correlate MSC biodistribution with glucose metabolism and morphology of heart in chagasic mice by small animal positron emission tomography (microPET). Mice were infected intraperitoneally with trypomastigotes of the Brazil strain of T. cruzi and treated by tail vein injection with MSC one month after infection. MSCs were labeled with near infrared fluorescent nanoparticles and tracked by an in vivo imaging system (IVIS). Our IVIS results two days after transplant revealed that a small, but significant, number of cells migrated to chagasic hearts when compared with control animals, whereas the vast majority of labeled MSC migrated to liver, lungs and spleen. Additionally, the microPET technique demonstrated that therapy with MSC reduced right ventricular dilation, a phenotype of the chagasic mouse model.

Conclusions

We conclude that the beneficial effects of MSC therapy in chagasic mice arise from an indirect action of the cells in the heart rather than a direct action due to incorporation of large numbers of transplanted MSC into working myocardium.     

Behavioural alterations are independent of sickness behaviour in chronic experimental Chagas disease

The existence of the nervous form of Chagas disease is a matter of discussion since Carlos Chagas described neurological disorders, learning and behavioural alterations in Trypanosoma cruzi-infected individuals. In most patients, the clinical manifestations of the acute phase, including neurological abnormalities, resolve spontaneously without apparent consequence in the chronic phase of infection. However, chronic Chagas disease patients have behavioural changes such as psychomotor alterations, attention and memory deficits, and depression. In the present study, we tested whether or not behavioural alterations are reproducible in experimental models. We show that C57BL/6 mice chronically infected with the Colombian strain of T. cruzi (150 days post-infection) exhibit behavioural changes as (i) depression in the tail suspension and forced swim tests, (ii) anxiety analysed by elevated plus maze and open field test sand and (iii) motor coordination in the rotarod test. These alterations are neither associated with neuromuscular disorders assessed by the grip strength test nor with sickness behaviour analysed by temperature variation sand weight loss. Therefore, chronically T. cruzi-infected mice replicate behavioural alterations (depression and anxiety) detected in Chagas disease patients opening an opportunity to study the interconnection and the physiopathology of these two biological processes in an infectious scenario.     

Treatment of chronically Trypanosoma cruzi-infected mice with a CCR1/CCR5 antagonist (Met-RANTES) results in amelioration of cardiac tissue damage

The comprehension of the molecular mechanisms leading to Trypanosoma cruzi-elicited heart dysfunction might contribute to design novel therapeutic strategies aiming to ameliorate chronic Chagas disease cardiomyopathy. In C3H/He mice infected with the low virulence T. cruzi Colombian strain, the persistent cardiac inflammation composed mainly of CCR5⁺ T lymphocytes parallels the expression of CC-chemokines in a pro-inflammatory IFN-γ and TNF-α milieu. The chronic myocarditis is accompanied by increased frequency of peripheral CCR5⁺LFA-1⁺ T lymphocytes. The treatment of chronically T. cruzi-infected mice with Met-RANTES, a selective CCR1/CCR5 antagonist, led to a 20–30% decrease in CD4⁺ cell numbers as well as IL-10, IL-13 and TNF-α expression. Further, Met-RANTES administration impaired the re-compartmentalization of the activated CD4⁺CCR5⁺ lymphocytes. Importantly, Met-RANTES treatment resulted in significant reduction in parasite load and fibronectin deposition in the heart tissue. Moreover, Met-RANTES treatment significantly protected T. cruzi-infected mice against connexin 43 loss in heart tissue and CK-MB level enhancement, markers of heart dysfunction. Thus, our results corroborate that therapeutic strategies based on the modulation of CCR1/CCR5-mediated cell migration and/or effector function may contribute to cardiac tissue damage limitation during chronic Chagas disease.     

Adipose Tissue-Derived Mesenchymal Stromal Cells Protect Mice Infected with Trypanosoma cruzi from Cardiac Damage through Modulation of Anti-parasite Immunity

Background

Chagas disease, caused by the protozoan Trypanosoma cruzi (T.cruzi), is a complex disease endemic in Central and South America. It has been gathering interest due to increases in non-vectorial forms of transmission, especially in developed countries. The objective of this work was to investigate if adipose tissue-derived mesenchymal stromal cells (ASC) can alter the course of the disease and attenuate pathology in a mouse model of chagasic cardiomyopathy.

Methodology/Principal Findings

ASC were injected intraperitoneally at 3 days post-infection (dpi). Tracking by bioluminescence showed that cells remained in the abdominal cavity for up to 9 days after injection and most of them migrated to the abdominal or subcutaneous fat, an early parasite reservoir. ASC injection resulted in a significant reduction in blood parasitemia, which was followed by a decrease in cardiac tissue inflammation, parasitism and fibrosis at 30 dpi. At the same time point, analyses of cytokine release in cells isolated from the heart and exposed to T. cruzi antigens indicated an anti-inflammatory response in ASC-treated animals. In parallel, splenocytes exposed to the same antigens produced a pro-inflammatory response, which is important for the control of parasite replication, in placebo and ASC-treated groups. However, splenocytes from the ASC group released higher levels of IL-10. At 60 dpi, magnetic resonance imaging revealed that right ventricular (RV) dilation was prevented in ASC-treated mice.

Conclusions/Significance

In conclusion, the injection of ASC early after T. cruzi infection prevents RV remodeling through the modulation of immune responses. Lymphoid organ response to the parasite promoted the control of parasite burden, while the heart, a target organ of Chagas disease, was protected from damage due to an improved control of inflammation in ASC-treated mice.     

The effect of BCG on the course of experimental Chagas' disease in mice

Experiments were done to determine the effect of BCG treatment on longevity, development of parasitemia, and in vivo distribution of ⁵¹Cr-labelled trypanosomes in C3H(He) female mice infected with a Brazil strain of Trypanosoma cruzi. BCG sensitization of mice was accomplished by a single IV injection of 3·0 mg (wet weight) of BCG. Twenty-one days after BCG injection mice were infected with 5 × 10⁴ blood-form trypomastigotes. Parasitemia determinations were made on alternate days during the experiment while in vivo distribution of exogenously supplied ⁵¹Cr-epimastigotes was made in groups of BCG or PBS stimulated mice on day 15 of the T. cruzi infection.It was found that BCG sensitization had no effect on longevity or parasitemia development in T. cruzi infected C3H(He) female mice. There were, however, some differences in the in vivo distribution of parasites between BCG treated and control mice. BCG stimulated mice accumulated greater numbers of radiolabelled trypanosomes in the kidneys and small intestines while PBS treated mice were found to have greater numbers of labelled parasites in the liver. Although no significant differences were observed in longevity of BCG or PBS treated mice, it was noted that BCG treated animals which were bled for parasitemia determinations lived significantly longer than those which were merely observed for longevity.     

L-arginine supplementation increases cardiac collagenogenesis in mice chronically infected with Berenice-78 Trypanosoma cruzi strain

Chagas disease, caused by Trypanosoma cruzi, is a major neglected tropical disease that occurs mainly as chronic infection and systemic infection. Currently, there is no suitable and effective drug to treat this parasitic disease. Administration of nutrients with immunomodulatory properties, such as arginine and nitric oxide radicals, may be helpful as antiparasitic therapy. In this study, we evaluated the effects of arginine supplementation during the acute phase of infection under the development of chronic Chagas' heart disease in Swiss mice inoculated with the Berenice-78 strain of T. cruzi. The effectiveness of arginine was determined by daily detection of the parasite in the blood and long-term serum levels of nitric oxide and tumor necrosis factor-alpha, in addition to evaluation of heart tissue damage. Arginine could flatten parasitemia and prevent elevation of tumor necrosis factor-alpha in T. cruzi-infected mice. Regarding chronic inflammatory myocardial derangements, similar findings were verified among T. cruzi-infected groups. Arginine promoted collagenogenesis in the heart muscle tissue of T. cruzi-infected arginine-supplemented group. These data show the paradoxical benefits of arginine in improving the outcome of Chagas chronic cardiomyopathy.     

Trypanosoma cruzi infection of human induced pluripotent stem cell-derived cardiomyocytes: an in vitro model for drug screening for Chagas disease

Chagas disease, caused by Trypanosoma cruzi, is an important global public health problem which, despite partial efficacy of benznidazole (Bz) in acute phase, urgently needs an effective treatment. Cardiotoxicity is a major safety concern for conduction of more accurate preclinical drug screening platforms. Human induced pluripotent stem cells derived cardiomyocytes (hiPSC-CM) are a reliable model to study genetic and infectious cardiac alterations and may improve drug development. Herein, we introduce hiPSC-CM as a suitable model to study T. cruzi heart infection and to predict the safety and efficacy of anti-T. cruzi drugs.     

High throughput selection of effective serodiagnostics for Trypanosoma cruzi infection

Background

Diagnosis of Trypanosoma cruzi infection by direct pathogen detection is complicated by the low parasite burden in subjects persistently infected with this agent of human Chagas disease. Determination of infection status by serological analysis has also been faulty, largely due to the lack of well-characterized parasite reagents for the detection of anti-parasite antibodies.

Methods

In this study, we screened more than 400 recombinant proteins of T. cruzi, including randomly selected and those known to be highly expressed in the parasite stages present in mammalian hosts, for the ability to detect anti-parasite antibodies in the sera of subjects with confirmed or suspected T. cruzi infection.

Findings

A set of 16 protein groups were identified and incorporated into a multiplex bead array format which detected 100% of >100 confirmed positive sera and also documented consistent, strong and broad responses in samples undetected or discordant using conventional serologic tests. Each serum had a distinct but highly stable reaction pattern. This diagnostic panel was also useful for monitoring drug treatment efficacy in chronic Chagas disease.

Conclusions

These results substantially extend the variety and quality of diagnostic targets for Chagas disease and offer a useful tool for determining treatment success or failure.     

α-Gal immunization positively impacts Trypanosoma cruzi colonization of heart tissue in a mouse model

Chagas disease, caused by the parasite Trypanosoma cruzi, is considered endemic in more than 20 countries but lacks both an approved vaccine and limited treatment for its chronic stage. Chronic infection is most harmful to human health because of long-term parasitic infection of the heart. Here we show that immunization with a virus-like particle vaccine displaying a high density of the immunogenic α-Gal trisaccharide (Qβ-αGal) induced several beneficial effects concerning acute and chronic T. cruzi infection in α1,3-galactosyltransferase knockout mice. Approximately 60% of these animals were protected from initial infection with high parasite loads. Vaccinated animals also produced high anti-αGal IgG antibody titers, improved IFN-γ and IL-12 cytokine production, and controlled parasitemia in the acute phase at 8 days post-infection (dpi) for the Y strain and 22 dpi for the Colombian strain. In the chronic stage of infection (36 and 190 dpi, respectively), all of the vaccinated group survived, showing significantly decreased heart inflammation and clearance of amastigote nests from the heart tissue.     

High fat diet aggravates cardiomyopathy in murine chronic Chagas disease

Infection with Trypanosoma cruzi, the etiologic agent in Chagas disease, may result in heart disease. Over the last decades, Chagas disease endemic areas in Latin America have seen a dietary transition from the traditional regional diet to a Western style, fat rich diet. Previously, we demonstrated that during acute infection high fat diet (HFD) protects mice from the consequences of infection-induced myocardial damage through effects on adipogenesis in adipose tissue and reduced cardiac lipidopathy. However, the effect of HFD on the subsequent stages of infection – the indeterminate and chronic stages – has not been investigated. To address this gap in knowledge, we studied the effect of HFD during indeterminate and chronic stages of Chagas disease in the mouse model. We report, for the first time, the effect of HFD on myocardial inflammation, vasculopathy, and other types of dysfunction observed during chronic T. cruzi infection. Our results show that HFD perturbs lipid metabolism and induces oxidative stress to exacerbate late chronic Chagas disease cardiac pathology.     

Immune-neuroendocrine and metabolic disorders in human and experimental T. cruzi infection: New clues for understanding Chagas disease pathology

Studies in mice undergoing acute Trypanosoma cruzi infection and patients with Chagas disease, led to identify several immune-neuroendocrine disturbances and metabolic disorders. Here, we review relevant findings concerning such abnormalities and discuss their possible influence on disease physiopathology.     

Synergy Testing of FDA-Approved Drugs Identifies Potent Drug Combinations against Trypanosoma cruzi

An estimated 8 million persons, mainly in Latin America, are infected with Trypanosoma cruzi, the etiologic agent of Chagas disease. Existing antiparasitic drugs for Chagas disease have significant toxicities and suboptimal effectiveness, hence new therapeutic strategies need to be devised to address this neglected tropical disease. Due to the high research and development costs of bringing new chemical entities to the clinic, we and others have investigated the strategy of repurposing existing drugs for Chagas disease. Screens of FDA-approved drugs (described in this paper) have revealed a variety of chemical classes that have growth inhibitory activity against mammalian stage Trypanosoma cruzi parasites. Aside from azole antifungal drugs that have low or sub-nanomolar activity, most of the active compounds revealed in these screens have effective concentrations causing 50% inhibition (EC₅₀''s) in the low micromolar or high nanomolar range. For example, we have identified an antihistamine (clemastine, EC₅₀ of 0.4 µM), a selective serotonin reuptake inhibitor (fluoxetine, EC₅₀ of 4.4 µM), and an antifolate drug (pyrimethamine, EC₅₀ of 3.8 µM) and others. When tested alone in the murine model of Trypanosoma cruzi infection, most compounds had insufficient efficacy to lower parasitemia thus we investigated using combinations of compounds for additive or synergistic activity. Twenty-four active compounds were screened in vitro in all possible combinations. Follow up isobologram studies showed at least 8 drug pairs to have synergistic activity on T. cruzi growth. The combination of the calcium channel blocker, amlodipine, plus the antifungal drug, posaconazole, was found to be more effective at lowering parasitemia in mice than either drug alone, as was the combination of clemastine and posaconazole. Using combinations of FDA-approved drugs is a promising strategy for developing new treatments for Chagas disease.     

Global Metabolomic Profiling of Acute Myocarditis Caused by Trypanosoma cruzi Infection

Chagas disease is caused by Trypanosoma cruzi infection, being cardiomyopathy the more frequent manifestation. New chemotherapeutic drugs are needed but there are no good biomarkers for monitoring treatment efficacy. There is growing evidence linking immune response and metabolism in inflammatory processes and specifically in Chagas disease. Thus, some metabolites are able to enhance and/or inhibit the immune response. Metabolite levels found in the host during an ongoing infection could provide valuable information on the pathogenesis and/or identify deregulated metabolic pathway that can be potential candidates for treatment and being potential specific biomarkers of the disease. To gain more insight into those aspects in Chagas disease, we performed an unprecedented metabolomic analysis in heart and plasma of mice infected with T. cruzi. Many metabolic pathways were profoundly affected by T. cruzi infection, such as glucose uptake, sorbitol pathway, fatty acid and phospholipid synthesis that were increased in heart tissue but decreased in plasma. Tricarboxylic acid cycle was decreased in heart tissue and plasma whereas reactive oxygen species production and uric acid formation were also deeply increased in infected hearts suggesting a stressful condition in the heart. While specific metabolites allantoin, kynurenine and p-cresol sulfate, resulting from nucleotide, tryptophan and phenylalanine/tyrosine metabolism, respectively, were increased in heart tissue and also in plasma. These results provide new valuable information on the pathogenesis of acute Chagas disease, unravel several new metabolic pathways susceptible of clinical management and identify metabolites useful as potential specific biomarkers for monitoring treatment and clinical severity in patients.