The use of peptides for immunodiagnosis of human Chagas disease.

Chagas disease, caused by the protozoa Trypanosoma cruzi, continues to be a serious public health problem in Latin America, worsened by the limitations in its detection. Given the importance of developing new diagnostic methods for this disease, the present review aimed to verify the number of publications dedicated to research on peptides that demonstrate their usefulness in serodiagnosis. To this end, a bibliographic survey was conducted on the PubMed platform using the keyword "peptide" or "epitope" combined with "Chagas disease" or "Trypanosoma cruzi"; "diagno*" or "serodiagnosis" or "immunodiagnosis", without period restriction. An increasing number of publications on studies employing peptides in ELISA and rapid tests assays was verified, which confirms the expansion of research in this field. It is possible to observe that many of the peptides tested so far originate from proteins widely used in the diagnosis of Chagas, and many of them are part of commercial tests developed. In this sense, as expected, promising results were obtained for several peptides when tested in ELISA, as many of them exhibited sensitivity and specificity values above 90%. Furthermore, some peptides have been tested in several studies, confirming their diagnostic potential. Despite the promising results observed, it is possible to emphasize the need for extensive testing of peptides, using different serological panels, in order to confirm their potential. The importance of producing an effective assay capable of detecting the clinical stages of the disease, as well as new immunogenic antigens that enable new serological diagnostic tools for Chagas disease, is evident.

Hypovitaminosis D and reduced cathelicidin are strongly correlated during the multidrug therapy against leprosy.

Mycobacterium leprae infection depends on the competence of the host immune defense to induce effective protection against this intracellular pathogen. The present study investigated the serum levels of vitamin D and the antimicrobial peptide cathelicidin, to determine the statistical correlation between them in leprosy patients before and post-six months of multidrug therapy (MDT), household contacts, and healthy individuals. Previous studies associated these molecules with high risks to develop mycobacterial diseases, such as tuberculosis and leprosy. A total of 34 leprosy patients [paucibacillary (n = 14), multibacillary (n = 20)], and 25 household contacts were recruited. Eighteen healthy adults were selected as a control group. Serum concentrations of vitamin D (25(OH)VD3) and cathelicidin were measured using high-performance liquid chromatography (HPLC), and an enzyme-linked immunosorbent assay (ELISA) kit, respectively. There were no significant differences in serum levels of 25(OH)VD3 between all groups, and the overall prevalence rate of vitamin D deficiency was 67.1%. Cathelicidin levels were significantly lower in both untreated and treated patients when compared to controls and household contacts (p < 0.05). Strong correlations between hypovitaminosis D and reduced cathelicidin in untreated (r = 0.86) and post-six months of MDT (r = 0.79) leprosy patients were observed. These results suggest that vitamin D status and cathelicidin levels are strongly correlated during multidrug therapy for leprosy and nutritional supplementation from the beginning of treatment could strengthen the immune response against leprosy.

Recombinant Leishmania eukaryotic elongation factor-1 beta protein: A potential diagnostic antigen to detect tegumentary and visceral leishmaniasis in dogs and humans.

The laboratorial diagnosis of leishmaniasis is based on parasitological methods, which are invasive, present high cost, require laboratorial infrastructure and/or trained professionals; as well as by immunological methods, which usually present variable sensitivity and/or specificity, such as when they are applied to identify asymptomatic cases and/or mammalian hosts presenting low levels of antileishmanial antibodies. As consequence, new studies aiming to identify more refined antigens to diagnose visceral (VL) and tegumentary (TL) leishmaniasis are urgently necessary. In the present work, the Leishmania eukaryotic elongation factor-1 beta (EF1b) protein, which was identified in L. infantum protein extracts by antibodies in VL patients' sera, was cloned and its recombinant version (rEF1b) was expressed, purified and tested as a diagnostic marker for VL and TL. The post-therapeutic serological follow-up was also evaluated in treated and untreated VL and TL patients, when anti-rEF1b antibody levels were measured before and after treatment. Results showed that rEF1b was highly sensitive and specific to diagnose symptomatic and asymptomatic canine VL, as well as human TL and VL. In addition, low cross-reactivity was observed when sera from healthy subjects or leishmaniasis-related diseases patients were tested. The serological follow-up showed also that rEF1b-specific antibodies declined significantly after treatment, suggesting that this protein could be also evaluated as a prognostic marker for human leishmaniasis.

The prognostic value of health-related quality of life in patients with Chagas heart disease.

PURPOSE: To verify the prognostic value of health-related quality of life (HRQoL) and the differences in HRQoL and clinical variables between groups of Chagas heart disease (CHD) patients with and without cardiovascular adverse events. METHODS: Seventy-five CHD patients were evaluated by echocardiography, maximal exercise testing, and Short-form of Health Survey (SF-36) Questionnaire. Patients were followed during 6 years. In the statistical analysis, uni- and multivariate Cox regression were performed to verify the accuracy of the HRQoL in predicting cardiovascular events. RESULTS: After the follow-up period (41 ± 12 months), 20 patients (27%) had adverse events. Those with poor outcome had lower left ventricular ejection fraction (LVEF) (p = 0.002), higher left ventricular end-diastolic diameter (LVDd) (p = 0.019), and worse scores in general health perceptions (p = 0.047), social role functioning (p = 0.026), and mental component summary (p = 0.043) of SF-36. Patients with lower LVEF (p = 0.003), higher LVDd (p = 0.022), worse HRQoL in the general heath perceptions domain (p = 0.022), and mental component summary (p = 0.031) were associated with worse prognosis. In the multivariate Cox regression, LVEF (HR 0.94, 95% CI from 0.90 to 0.98, p = 0.007) and mental component summary (HR 0.98, 95% CI from 0.94 to 1.00, p = 0.047) remained as independent predictors of adverse events in CHD patients. CONCLUSION: The assessment of HRQoL, especially the mental component, should be taken into account to provide an accurate prognosis in addition to other well-established predictors of poor outcomes in CHD patients.

Immunogenomic screening approach to identify new antigens for the serological diagnosis of chronic Chagas' disease.

Serological tests are preferentially used for the diagnosis of Chagas' disease (CD) during the chronic phase because of the low parasitemia and high anti-Trypanosoma cruzi antibody titers. However, the current methods showed several disadvantages, as contradictory or inconclusive results, mainly related to the characteristics of the antigens used, in general, crude or whole parasites, but also due to antigen production protocol and the experimental conditions used in serological tests. Thus, better-quality serological assays are urgently needed. Here, we performed a wide immunogenomic screen strategy to identify conserved linear B-cell epitopes in the predicted proteome based on genome sequence from T. cruzi strains to will be applied as synthetic peptides in the serodiagnosis of the chronic CD. Three B-cell epitopes derived from mucin-associated surface protein (MASP) family, expressed in both infective parasite stages, trypomastigote and amastigotes, conserved in T. cruzi strains, and highly divergent as compared with Leishmania spp. proteome, were selected for this study. The results demonstrated that synthetic peptide 2 and a mixture of peptides (Mix II: peptides 2 and 3) were able to identify all chronic CD cases, indeterminate or Chagas cardiomyopathy clinical presentation, and simultaneously able to discriminate infections caused by Leishmania parasites, with high accuracy (98.37 and 100.00%, respectively) and agreement (kappa index = 0.967 and 1.000, respectively) with direct methods as compared to current diagnostic pipeline performed by reference laboratories in Brazil. This study represents an interesting strategy for the discovery of new antigens applied to serologic diagnosis of infectious diseases and for the technological development of platforms for large-scale production of diagnostic tests.

Prognostic value of serum brain-derived neurotrophic factor levels in patients with Chagas cardiomyopathy.

BACKGROUND: Serum brain-derived neurotrophic factor (BDNF) levels have been shown to be lower in patients with Chagas cardiomyopathy (ChC) than in patients with non-dilated chagasic cardiomyopathy. However, its prognostic value was not established in patients with ChC. METHODS: Forty-nine patients with ChC (50 ± 7 years, New York Heart Association "NYHA" I-III); were evaluated by echocardiography, exercise testing, and blood analysis. Serum BDNF levels were determined using enzyme-linked immunosorbent assay sandwich. Patients were followed-up, and cardiac death was considered the end-point. The survival analyses were performed using Kaplan-Meier and Cox regression. RESULTS: After 39 ± 14 months of follow-up, 12 patients (25%) died. The concentration of 2.5 ng/mL was the optimal cut-off value to predict survival with significant difference between the groups with low (≤ 2.5 ng/mL) and high (> 2.5 ng/mL) BDNF levels (p = 0.006). Lower serum BDNF levels (hazards ratio (HR) 1.1, 95% confidence interval (CI) 1.1-1.4; p = 0.001), peak oxygen uptake (HR 1.2, 95% CI 1.0-1.3; p = 0.009), and left ventricular ejection fraction (HR 0.8, 95% CI 0.7-0.9; p = 0.001) were the independent predictors of survival. The combination of low serum BDNF levels and reduced left ventricular ejection fraction were highly predictive of death (HR 5.6, 95% CI: 1.2-9.7; p = 0.026). CONCLUSION: In patients with ChC, reduced serum BDNF levels, especially if associated with systolic function, may provide useful prognostic information.

Immunoregulatory mechanisms in Chagas disease: modulation of apoptosis in T-cell mediated immune responses.

BACKGROUND: Chronic Chagas disease presents different clinical manifestations ranging from asymptomatic (namely indeterminate) to severe cardiac and/or digestive. Previous results have shown that the immune response plays an important role, although no all mechanisms are understood. Immunoregulatory mechanisms such as apoptosis are important for the control of Chagas disease, possibly affecting the morbidity in chronic clinical forms. Apoptosis has been suggested to be an important mechanism of cellular response during T. cruzi infection. We aimed to further understand the putative role of apoptosis in Chagas disease and its relation to the clinical forms of the disease. METHODS: Apoptosis of lymphocytes, under antigenic stimuli (soluble T. cruzi antigens - TcAg) where compared to that of non-stimulated cells. Apoptosis was evaluated using the expression of annexin and caspase 3(+) by T cells and the percentage of cells positive evaluated by flow cytometry. In addition activation and T cell markers were used for the identification of TCD4(+) and TCD8(+) subpopulations. The presence of intracellular and plasma cytokines were also evaluated. Analysis of the activation status of the peripheral blood cells showed that patients with Chagas disease presented higher levels of activation determined by the expression of activation markers, after TcAg stimulation. PCR array were used to evaluate the contribution of this mechanism in specific cell populations from patients with different clinical forms of human Chagas disease. RESULTS: Our results showed a reduced proliferative response associated a high expression of T CD4(+)CD62L(-) cells in CARD patients when compared with IND group and NI individuals. We also observed that both groups of patients presented a significant increase of CD4(+) and CD8(+) T cell subsets in undergoing apoptosis after in vitro stimulation with T. cruzi antigens. In CARD patients, both CD4(+) and CD8(+) T cells expressing TNF-α were highly susceptible to undergo apoptosis after in vitro stimulation. Interestingly, the in vitro TcAg stimulation increased considerably the expression of cell death TNF/TNFR superfamily and Caspase family receptors genes in CARD patients. CONCLUSIONS: Taken together, our results suggest that apoptosis may be an important mechanism for the control of morbidity in T. cruzi infection by modulating the expression of apoptosis genes, the cytokine environment and/or killing of effector cells.

Galectin-3 (Gal-3) and the tissue inhibitor of matrix metalloproteinase (TIMP-2) as potential biomarkers for the clinical evolution of chronic Chagas cardiomyopathy.

BACKGROUND: Chronic Chagas cardiomyopathy (CCC) is responsible for the highest morbidity and worst prognosis in Chagas disease patients. However, predicting factors that correlate with disease progression, morbidity, and mortality is challenging. It is necessary to have simple, quantitative, and economical risk biomarkers that add value to conventional methods and assist in the diagnosis and prognosis of patients with CCC or in evolution. OBJECTIVES: We evaluated molecules related to cardiac remodeling and fibrosis, such as MMP-2, MMP-9, TIMP-2, TIMP-1, PICP, CTXI, and Gal-3, and correlated these biomarkers with echocardiographic variables (LVDD, LVEF, and E/e' ratio). METHODS: Blood samples from Chagasic patients without apparent cardiopathy (WAC), CCC patients, and healthy individuals were used to perform plasma molecule dosages using Luminex or ELISA. RESULTS: MMP-2 and TIMP-2 presented higher levels in CCC; in these patients, the inhibitory role of TIMP-2 over MMP-2 was reinforced. The ratio of MMP-2/TIMP-2 in WAC patients showed a bias in favor of the gelatinase pathway. MMP-9 and TIMP-1 showed higher levels in Chagas patients compared to healthy subjects. PICP and CTXI are not associated with cardiac deterioration in Chagas disease. Increased levels of Gal-3 are associated with worse cardiac function in CCC. Receiver operating characteristic (ROC) curve analysis identified Gal-3 and TIMP-2 as putative biomarkers to discriminate WAC from cardiac patients. CONCLUSIONS: Among the molecules evaluated, Gal-3 and TIMP-2 have the potential to be used as biomarkers of cardiac remodeling and progressive myocardial fibrosis in Chagas disease.

Recombinant proteins as promising antigens applied to the immunodiagnosis of Chagas disease: a scoping review.

Chagas disease (CD), caused by the protozoan Trypanosoma cruzi, is an important public health problem, occurring mainly in Latin America. The disease has a major social and economical effect, negatively impacting the life of the infected individuals, and bringing great costs to public health. An early and accurate diagnosis is essential for administration of early treatment. In addition, prognostic tests may aid disease management, decreasing hospitalization costs. However, the serological diagnostic scenario for CD still faces several challenges, making the development of new diagnostic kits a pressing matter. Facing this scenario, several researchers have expanded efforts in developing and testing new antigens, such as recombinant proteins and recombinant multiepitope proteins, with promising results. These recombinant antigens offer several advantages, such as improved sensitivity and specificity, in addition to facilitated scaling. Also, it has been possible to observe a rising number of studies using ELISA and point-of-care platforms, employing these antigens in the past few years. Among them, recombinant proteins were the most applied antigens, demonstrating great capacity to discriminate between positive and negative samples. Although fewer in number, recombinant multiepitope proteins also demonstrated an improved diagnostic performance. Indeed, a great number of studies employing these antigens showed sensitivity and specificity values above 90%, greatly impacting diagnostic accuracy. Nevertheless, despite the good results found, it is still possible to observe some bottlenecks in the development of new antigens, such as the scarcity of tests with sera from the acute phase and the variability of results in different geographic areas. In this sense, aiming to contribute to control and health programs, the continuous search for a more accurate serological diagnosis is essential, both for the acute and chronic phases of the disease.

Chagas cardiomyopathy is associated with a high susceptibility to T. cruzi infection in monocyte-derived macrophages and a predominance of CD4+CD45RO+ T-cells with immunoregulatory patterns.

The pathogenesis of Chronic Chagas Cardiomyopathy (CCC) is still not fully understood, and the persistence of the parasite in tissues seems to be essential for the onset and progression of heart disease, tissue destruction, and chronic inflammation. It is clear that the polarity found between the asymptomatic (IND) and cardiac clinical forms refers mainly to the mechanisms involved in the regulation of the host's immune response. Thus, to elucidate aspects of the susceptibility of host phagocytes to T. cruzi infection, the present study explored novel aspects of innate immune response, integrating data on susceptibility to infection and intracellular replication, using monocyte-derived macrophages from CCC patients, together with memory CD4+ T-cells (CD45RO+). The isolation of PBMC was conducted by means of in vitro infection assay with T. cruzi trypomastigotes and flow cytometry analysis of the intracytoplasmic cytokine production by CD4+T-cells. Our findings indicated that monocytes derived from individuals with CCC are more susceptible to the infection and replication of intracellular amastigotes. Moreover, the stimulation of CD4+ T-cells from CCC patients, together with T. cruzi trypomastigotes, induces a predominance of a regulatory response over a type 1 response, demonstrated by an increase in IL-10 production and a reduction in the IFN-γ and IFN-γ/IL-10. Suppression of the function of monocyte-derived macrophages, from CCC patients, to control trypomastigote infection and intracellular replication sheds light on a potential susceptibility of these cells isolated from peripheral blood, which may reflect the ineffectiveness of parasite control by phagocytes in cardiac tissues, which can subsequently result in serious heart disease.

rMELEISH: A Novel Recombinant Multiepitope-Based Protein Applied to the Serodiagnosis of Both Canine and Human Visceral Leishmaniasis.

BACKGROUND: visceral leishmaniasis (VL) is a critical public health problem in over ninety countries. The control measures adopted in Brazil have been insufficient when it comes to preventing the spread of this overlooked disease. In this context, a precise diagnosis of VL in dogs and humans could help to reduce the number of cases of this disease. Distinct studies for the diagnosis of VL have used single recombinant proteins in serological assays; however, the results have been variable, mainly in relation to the sensitivity of the antigens. In this context, the development of multiepitope-based proteins could be relevant to solving such problem. METHODS: a chimeric protein (rMELEISH) was constructed based on amino acid sequences from kinesin 39 (k39), alpha-tubulin, and heat-shock proteins HSP70 and HSP 83.1, and tested in enzyme-linked immunosorbent (ELISA) for the detection of L. infantum infection using canine (n = 140) and human (n = 145) sera samples. RESULTS: in the trials, rMELEISH was able to discriminate between VL cases and cross-reactive diseases and healthy samples, with sensitivity and specificity values of 100%, as compared to the use of a soluble Leishmania antigenic extract (SLA). CONCLUSIONS: the preliminary data suggest that rMELEISH has the potential to be tested in future studies against a larger serological panel and in field conditions for the diagnosis of canine and human VL.

Proof of Concept of a Novel Multiepitope Recombinant Protein for the Serodiagnosis of Patients with Chagas Disease.

Chagas disease remains a neglected disease that is considered to be a public health problem. The early diagnosis of cases is important to improve the prognosis of infected patients and prevent transmission. Serological tests are the method of choice for diagnosis. However, two serological tests are currently recommended to confirm positive cases. In this sense, more sensitive and specific serological tests need to be developed to overcome these current diagnosis problems. This study aimed to develop a new recombinant multiepitope protein for the diagnosis of Chagas disease, hereafter named rTC. The rTC was constructed based on amino acid sequences from different combinations of Trypanosoma cruzi antigens in the same polypeptide and tested using an enzyme-linked immunosorbent assay (ELISA) to detect different types of Chagas disease. rTC was able to discriminate between indeterminate (IND) and cardiac (CARD) cases and cross-reactive diseases, as well as healthy samples, with 98.28% sensitivity and 96.67% specificity, respectively. These data suggest that rTC has the potential to be tested in future studies against a larger serological panel for the diagnosis of Chagas disease.

B-Cell Epitopes-Based Chimeric Protein from SARS-CoV-2 N and S Proteins Is Recognized by Specific Antibodies in Serum and Urine Samples from Patients.

The impact of the COVID-19 pandemic caused by the SARS-CoV-2 virus underscored the crucial role of laboratorial tests as a strategy to control the disease, mainly to indicate the presence of specific antibodies in human samples from infected patients. Therefore, suitable recombinant antigens are relevant for the development of reliable tests, and so far, single recombinant proteins have been used. In this context, B-cell epitopes-based chimeric proteins can be an alternative to obtain tests with high accuracy through easier and cheaper production. The present study used bioinformatics tools to select specific B-cell epitopes from the spike (S) and the nucleocapsid (N) proteins from the SARS-CoV-2 virus, aiming to produce a novel recombinant chimeric antigen (N4S11-SC2). Eleven S and four N-derived B-cell epitopes were predicted and used to construct the N4S11-SC2 protein, which was analyzed in a recombinant format against serum and urine samples, by means of an in house-ELISA. Specific antibodies were detected in the serum and urine samples of COVID-19 patients, which were previously confirmed by qRT-PCR. Results showed that N4S11-SC2 presented 83.7% sensitivity and 100% specificity when using sera samples, and 91.1% sensitivity and 100% specificity using urine samples. Comparable findings were achieved with paired urine samples when compared to N and S recombinant proteins expressed in prokaryotic systems. However, better results were reached for N4S11-SC2 in comparison to the S recombinant protein when using paired serum samples. Anti-N4S11-SC2 antibodies were not clearly identified in Janssen Ad26.COV2.S COVID-19-vaccinated subjects, using serum or paired urine samples. In conclusion, this study presents a new chimeric recombinant antigen expressed in a prokaryotic system that could be considered as an alternative diagnostic marker for the SARS-CoV-2 infection, with the potential benefits to be used on serum or urine from infected patients.

Galectin-3 and fibrosis intensity in Chronic Chagas Cardiomyopathy: a systematic review.

Chronic Chagas Cardiomyopathy (CCC) is the most prevalent type of myocarditis and the main clinical form of the Chagas disease, which has peculiarities such as focal inflammation, structural derangement, hypertrophy, dilation, and intense reparative fibrosis. Many cellular compounds contribute to CCC development. Galectin-3 is a partaker in inflammation and contributes to myocardial fibrosis formation. Some studies showed the connection between Galectin-3 and fibrosis in Chagas disease but are still inconclusive on the guidance for the early implementation of pharmacological therapy. This systematic review evaluated Galectin-3 as a biomarker for fibrosis intensity in CCC. Two independent reviewers have searched five databases (PubMed, EMBASE, Cochrane Library, Scopus, and Lilacs), using the following search terms: galectin-3, biomarkers, fibrosis, Chagas cardiomyopathy, and Chagas disease. Overall, seven studies met the inclusion criteria and made up this review. There were four trials conducted through animal model experiments and three trials with humans. Experimental data in mice indicate an association between Galectin-3 expression and fibrosis in CCC (75% of studies). Data from human studies showed no direct connection between myocardial fibrosis and Galectin-3 expression (80% of studies). Thus, human findings do not provide significant evidence indicating that Galectin-3 is related to fibrosis formation in Chagas disease. Based on the analyzed studies, it is suggested that Galectin-3 might not be a good fibrosis marker in CCC.

Reduced vitamin D receptor (VDR) and cathelicidin antimicrobial peptide (CAMP) gene expression contribute to the maintenance of inflammatory immune response in leprosy patients.

Leprosy is an infectious disease influenced by genetic, immunological, and environmental factors. Reduced gene expressions may be associated with the immunological response pattern and leprosy susceptibility. We investigated the direct and indirect effects of Vitamin D Receptor (VDR) and Cathelicidin Antimicrobial Peptide (CAMP) gene expressions on the serum levels of vitamin D, Cathelicidin, and cytokines in newly-diagnosed leprosy patients and post-six-months of multidrug therapy (MDT). Thirty-four leprosy patients were assessed, paucibacillary (PB; n = 14) and multibacillary (MB; n = 20) cases, untreated or having received six months of MDT, 18 healthy controls, and 25 household contacts. VDR and CAMP gene expression levels were strongly correlated to some important cytokines in both, untreated leprosy patients (PB, r = 0.9319; MB, r = 0.9569) and patients who had undergone MDT (PB, r = 0.9667; MB, r = 0.9569). We observed that both gene expressions directly influenced IL-2, IFN-γ, and IL-17F serum levels in leprosy patients compared to the household contacts and healthy individuals. VDR and CAMP gene expressions induced a persistent inflammatory response in PB and MB leprosy patients, even after six months of MDT, to fight the Mycobacterium leprae infection. Due to the persistent inflammatory profile, multidrug therapy is suggested to be maintained for more than six months, especially for MB patients. Vitamin D supplementation is recommended from the onset as a transcription factor to improve VDR and CAMP gene expression in leprosy patients.

Potential of recombinant LiHyQ, a novel Leishmania infantum protein, for the diagnosis of canine visceral leishmaniasis and as a diagnostic and prognostic marker for human leishmaniasis and human immunodeficiency virus co-infection: A preliminary study.

Laboratory diagnosis of leishmaniasis shows variable efficacy in detecting infected mammalian hosts and there is a need to identify suitable antigens to improve the accuracy of diagnostic tests. In the present study, a L. infantum hypothetical protein called LiHyQ was evaluated for the diagnosis of tegumentary (TL) and visceral (VL) leishmaniasis using canine and human samples. A collection of dog sera (n=155) were tested and contained samples from asymptomatic (n=20) and symptomatic (n=25) VL animals, from healthy dogs living in endemic (n=25) or non-endemic (n=25) areas of disease, from Leish-Tec® vaccinated dogs (n=20) or from dogs infected with Ehrlichia canis (n=15), Babesia canis (n=10) and Trypanosoma cruzi (n=15). Sensitivity (Se), Specificity (Sp), Positive Predictive Value (PPV) and Negative Predictive Value (NPV) of 100% were observed for rLiHyQ with these samples, whereas the Se, Sp, PPV and NPV values with L. infantum Soluble Leishmania Antigen (SLA) preparation were 60.0%, 99.0%, 96.0% and 86.0%, respectively. A collection of human sera (n=305) were tested and contained samples from TL (n=50) and VL (n=40) patients, from VL/HIV co-infected patients (n=35), from patients infected with HIV alone (n=30), Chagas Disease (n=30), malaria (n=10), tuberculosis (n=10), paracoccidioidomycosis (n=15), leprosy (n=30) or aspergillosis (n=15); and from healthy subjects (n=40). Se, Sp, PPV and NPV values of 100% were observed for rLiHyQ with these samples, whereas the Se, Sp, PPV and NPV values with SLA were 58.0%, 76.0%, 50.0% and 82.0%, respectively. The antibody reactivity against the protein was compared with commercial kits, and the kappa index varied from 0.95 to 1.00 for rLiHyQ, and of 0.55 to 0.82 for the kits. In addition, the serological follow-up of treated patients showed a significant reduction in rLiHyQ-specific IgG antibody levels. All canine and human samples were tested at the same time using the same reagents, in order to reduce experimental variation and interference in data interpretation. In conclusion, our preliminary data suggest a diagnostic and prognostic role for rLiHyQ against leishmaniasis.

Radionuclide esophageal transit scintigraphy in chronic indeterminate and cardiac forms of Chagas disease.

OBJECTIVES: This study was designed to investigate digestive and heart associations, using esophageal transit scintigraphy in three different groups: patients exclusively with the dilated cardiac form of Chagas disease (DCCh), an indeterminate form of Chagas disease, and healthy controls. As a hypothesis, we assumed that autonomic chagasic denervation is a global process. MATERIALS AND METHODS: Twenty healthy controls and 75 outpatients with Chagas disease, divided into indeterminate form (n = 33) and (DCCh, n = 42), underwent esophageal scitigraphy. The esophageal transit time (ETT) and percentage of esophageal emptying (%EE) were analyzed by group and correlated with left ventricular ejection fraction (LVEF). RESULTS: ETT alterations were found in 57% of indeterminate form and 80% of DCCh. The observed values of ETT and %EE in Chagas disease groups were significantly different from the controls (P-value <0.001). The lowest median ETT was observed for the controls (median = 8.0), followed by indeterminate form (median = 16.5) and DCCh (median = 60.0). Regarding %EE, a higher median value was observed for the controls (median = 92.3), followed by indeterminate form (median = 86.7) and DCCh (median = 56.9). In the DCCh group, ETT and %EE were significantly correlated with LVEF values (r = 0.398; P = 0.015) and (r = 0.475; P = 0.003), respectively. CONCLUSION: An association between left ventricular systolic dysfunction and functional esophageal alterations was observed which was interpreted as indirect evidence of concomitance of gastrointestinal and cardiac disorders. We also found that the greater the impairment of the esophagus's autonomic function, the worse the cardiac dysfunction.

Liver damage in schistosomiasis is reduced by adipose tissue-derived stem cell therapy after praziquantel treatment.

BACKGROUND: In view of the potential immunosuppressive and regenerative properties of mesenchymal stem cells (MSC), we investigated whether transplantation of adipose tissue-derived stem cells (ASC) could be used to control the granulomatous reaction in the liver of mice infected with Schistosoma mansoni after Praziquantel (PZQ) treatment. METHODOLOGY/PRINICPAL FINDINGS: C57BL/6 mice infected with S. mansoni were treated with PZQ and transplanted intravenously with ASC from uninfected mice. Liver morpho-physiological and immunological analyses were performed. The combined PZQ/ASC therapy significantly reduced the volume of hepatic granulomas, as well as liver damage as measured by ALT levels. We also observed that ASC accelerated the progression of the granulomatous inflammation to the advanced/curative phase. The faster healing interfered with the expression of CD28 and CTLA-4 molecules in CD4+ T lymphocytes, and the levels of IL-10 and IL-17 cytokines, mainly in the livers of PZQ/ASC-treated mice. CONCLUSIONS: Our results show that ASC therapy after PZQ treatment results in smaller granulomas with little tissue damage, suggesting the potential of ASC for the development of novel therapeutic approaches to minimize hepatic lesions as well as a granulomatous reaction following S. mansoni infection. Further studies using the chronic model of schistosomiasis are required to corroborate the therapeutic use of ASC for schistosomiasis.

Characterization of the presence and distribution of Foxp3(+) cells in chagasic patients with and without megacolon.

Patients with Chagas's disease in the chronic phase regularly present with the chagasic megacolon. This form is characterized by inflammation, neuronal destruction, and organ dilatation. Chagasic patients with megacolon always present with inflammatory process near the enteric plexuses of the colon, as previously demonstrated. The aim of this study is to characterize the presence and distribution of Foxp3(+) cells in the muscle layers and neuronal plexuses area of the colon from chagasic patients with and without megacolon. Our results demonstrated that chagasic patients without megacolon presented with an increased concentration of Foxp3(+) cells in all colon layers compared with chagasic patients with megacolon and noninfected individuals. These cells were situated mainly near the blood vessels and rarely were associated with the inflammatory foci. We believe that the presence of Foxp3(+) cells may help to control the inflammatory process through the management of lymphocyte migration and, consequently, prevent neuronal destruction and chagasic megacolon development.

Reading of the Mitsuda test: comparison between diameter and total area by means of a computerized method.

The Mitsuda test is a skin test based on the individual's immune response through late and highly specific hypersensitivity reaction to the Mycobacterium leprae bacillus. A negative reaction identifies individuals who present a higher risk of becoming ill if exposed to M. leprae and, if they become ill, to develop the virchowian form of disease. The Mitsuda test reading is performed by means of a millimeter ruler. The dermatoscopy is a method that has not been used in the evaluation of cutaneous tests, although its use has increased in several areas. The study aimed to compare the results between the standardized reading and the total area of the Mitsuda test obtained by a computerized method which was structured by the association of digital dermatoscopy, the Dermatology Web system and the Image Tool 3.0 software. Data collection was performed at the Dermatology Outpatient Clinic of the Eduardo de Menezes Hospital, in Belo Horizonte, from November 2015 to August 2016. The sample consisted of 100 leprosy domiciliary contacts. There was an excellent agreement between the Mitsuda test (diameter and area), with a coefficient greater than 80%, and an excellent correlation with the Spearman's correlation coefficient (0.936). The intraclass correlation coefficient indicated a low (0.219) but significant agreement between the two measurements. In conclusion, there is a significant correlation between the standardized reading and the total area of the Mitsuda test. Digital dermoscopy can be an alternative instrument of evaluation, allowing the computerization and recording of the Mitsuda test.