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Abstract Background Conventional serological tests, using total soluble proteins or a cocktail of recombinant proteins from T. cruzi as antigens, are highly sensitive for Chagas disease diagnosis. This type of tests, however, does not seem to be reliable tools for short- and medium-term monitoring of the evolution of patients after antiparasitic treatment. The aim of the present study was to search for immunological markers that could be altered in the sera from Chagas disease patients after benznidazole treatment, and therefore have a potential predictive diagnostic value. Methods We analyzed the reactivity of sera from chagasic patients during different clinical phases of the disease against a series of immunodominant antigens, known as KMP11, PFR2, HSP70 and Tgp63. The reactivity of the sera from 46 adult Chronic Chagas disease patients living in a non-endemic country without vector transmission of T. cruzi (15 patients in the indeterminate stage, 16 in the cardiomiopathy stage and 16 in the digestive stage) and 22 control sera from non-infected subjects was analyzed. We also analyzed the response dynamics of sera from those patients who had been treated with benznidazole. Results Regardless of the stage of the sickness, the sera from chagasic patients reacted against KMP11, HSP70, PFR2 and Tgp63 recombinant proteins with statistical significance relative to the reactivity against the same antigens by the sera from healthy donors, patients with autoimmune diseases or patients suffering from tuberculosis, leprosy or malaria. Shortly after benznidazole treatment, a statistically significant decrease in reactivity against KMP11, HSP70 and PFR2 was observed (six or nine month). It was also observed that, following benznidazole treatment, the differential reactivity against these antigens co-relates with the clinical status of the patients. Conclusions The recombinant antigens KMP11, PFR2, Tgp63 and HSP70 are recognized by Chagas disease patients' sera at any clinical stage of the disease. Shortly after benznidazole treatment, a drop in reactivity against three of these antigens is produced in an antigen-specific manner. Most likely, analysis of the reactivity against these recombinant antigens may be useful for monitoring the effectiveness of benznidazole treatment.

Introduction: Treatment of Chagas disease frequently causes distress to patients due to a high incidence of adverse effects. Different preemptive tests have been researched to prevent these effects and to allow focus to be given to certain predisposed patients. Benznidazole is the most prescribed Chagas disease treatment in Spain. In this work, we analyzed the genetic markers HLA-B*35 allele group and HLA-B*35:05 allele specifically, as well as an allergy patch test, as benznidazole's most frequent adverse effects are cutaneous., Methods: HLA-B intermediate-resolution genotyping was performed followed by a high-resolution level analysis. Cutaneous allergies were tested using strips impregnated with a mixture of benznidazole and placed on the upper back of patients before starting treatment., Results: In our sample of more than 400 patients, there was almost no relationship between any kind of side effect and either of the HLA-B alleles studied. The patch testing was quickly discarded as a preemptive test due to its low sensitivity (16.7%)., Conclusion: In conclusion, we were unable to replicate and corroborate genetic markers identified by other groups and there is currently no test that can anticipate the adverse effects of benznidazole, therefore, more investigation should be carried out in this field., (Copyright © 2024 Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. Published by Elsevier España, S.L.U. All rights reserved.)

Introduction: Chagas disease, caused by the Trypanosoma cruzi parasite infection, is a potentially life-threatening neglected tropical disease with a worldwide distribution. During the chronic phase of the disease, there exists a fragile balance between the host immune response and parasite replication that keeps patients in a clinically-silent asymptomatic stage for years or even decades. However, in 40% of patients, the disease progresses to clinical manifestations mainly affecting and compromising the cardiac system. Treatment is recommended in the chronic phase, although there are no early markers of its effectiveness. The aim of this study is to identify differential expression changes in genes involved in the immune response in antigen-restimulated PBMC from chronic patients with Chagas disease due to benznidazole treatment., Methods: Thus, high-throughput real-time qPCR analysis has been performed to simultaneously determine global changes in the expression of 106 genes involved in the immune response in asymptomatic (IND) and early cardiac manifestations (CCC I) Chagas disease patients pre- and post-treatment with benznidazole., Results and Discussion: The results revealed that 7 out of the 106 analyzed genes were differentially expressed (4 up- and 3 downregulated) after treatment in IND patients and 15 out of 106 (3 up- and 12 downregulated) after treatment of early cardiac Chagas disease patients. Particularly in CCC I patients, regulation of the expression level of some of these genes towards a level similar to that of healthy subjects suggests a beneficial effect of treatment and supports recommendation of benznidazole administration to early cardiac Chagas disease patients. The data obtained also demonstrated that both in asymptomatic patients and in early cardiac chronic patients, after treatment with benznidazole there is a negative regulation of the proinflammatory and cytotoxic responses triggered as a consequence of T. cruzi infection and the persistence of the parasite. This downregulation of the immune response likely prevents marked tissue damage and healing in early cardiac patients, suggesting its positive effect in controlling the pathology., 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 © 2024 Gómez, Egui, Palacios, Carrilero, Benítez, Simón, Segovia, Carmelo, López and Thomas.)

Introduction: We report the case of a fatal hemorrhagic varicella primary infection in an immunocompetent man and whole-genome characterization of the virus for the investigation of biomarkers of virulence., Case: A 38-year-old patient born in Nigeria presented to the emergency department with abdominal pain and subsequently developed fatal hemorrhagic disease without skin rash. Extensive laboratory tests including serology and PCR for arenaviruses, bunyaviruses and ebolaviruses were negative. Varicella-zoster virus (VZV) PCR of sera, liver and spleen tissue samples from autopsy revealed the presence of VZV DNA. Primary infection by varicella-zoster virus with hemorrhagic manifestations was diagnosed after virological testing. The VZV genome was sequenced using a mWGS approach. Bioinformatic analysis showed 53 mutations across the genome, 33 of them producing non-synonymous variants affecting up to 14 genes. Some of them, such as ORF11 and ORF 62, encoded for essential functions related to skin or neurotropism. To our knowledge, the mutations reported here have never been described in a VZV causing such a devastating outcome., Discussion: In immunocompetent patients, viral factors should be considered in patients with uncommon symptoms or severe diseases. Some relevant mutations revealed by using whole genome sequencing (WGS) directly from clinical samples may be involved in this case and deserves further investigation., Conclusion: Differential diagnosis of varicella-zoster virus in immunocompetent adults should be considered among patients with suspected VHF, even if the expected vesicular rash is not present at admission and does not arise thereafter. Whole genome sequencing of strains causing uncommon symptoms and/or mortality is needed for epidemiological surveillance and further characterization of putative markers of virulence. Additionally, this report highlights the recommendation for a VZV vaccination policy in non-immunized migrants from developing countries., (© 2023. The Author(s).)

Background: Patients with chronic Chagas disease present marked clinical and immunological heterogeneity. During the disease, multiple immune mechanisms are activated to fight the parasite. The purpose of this study was to investigate the expression patterns of genes involved in relevant immunological processes throughout the disease in patients with chronic Chagas disease., Methodology/principal Findings: High-throughput RT-qPCR with QuantStudio 12K Flex real-time PCR system was used to evaluate the expression of 106 immune-related genes in PBMC from a cohort of cardiac Chagas disease patients (CCC I), asymptomatic patients (IND) and healthy donors (HD) after being stimulated with T. cruzi soluble antigens. Principal component analysis (PCA), cluster analysis and volcano plots were used to identify differentially expressed genes. In addition, gene set enrichment analysis (GSEA) was employed to identify the enriched immunological pathways in which these genes are involved. PCA revealed the existence of a statistically divergent expression profile of the 36 genes correlated with PC1 between CCC I patients and HD (p < 0.0001). Differential gene expression analysis revealed upregulation of 41 genes (expression fold-change > 1.5) and downregulation of 14 genes (expression fold-change < 0.66) (p = 8.4x10-13 to p = 0.007) in CCC I patients versus HD. Furthermore, significant differences in the expression level of specific genes have been identified between CCC I and IND patients (8 up and 1 downregulated). GSEA showed that several upregulated genes in CCC I patients participate in immunological pathways such as antigen-dependent B cell activation, stress induction of HSP regulation, NO2-dependent IL12 pathway in NK cells, cytokines-inflammatory response and IL-10 anti-inflammatory signaling., Conclusions: Cardiac Chagas disease patients show an antigen-specific differential gene expression profile in which several relevant immunological pathways seem to be activated. Assessment of gene expression profiles reveal unique insights into the immune response that occurs along chronic Chagas disease., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Gómez et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: Chagas disease (CD) treatment is commonly associated with a high incidence of adverse effects. It is crucial to study and update these adverse effects to improve the existing knowledge of which drugs to use and to clarify the information presented to patients., Methods: We analyzed the adverse effects of benznidazole in two cohorts of patients: a large retrospective study and a small prospective study., Results: This large retrospective study described the most and least common adverse effects in our area and characterized our Chagas disease population. This prospective study, along with a close follow-up of the treatment, detected more adverse effects and enhanced the patients' perception of the disease and treatment., Conclusions: This information is important for preventing non-medical-related withdrawals and for removing baseless fears. Better knowledge of patients could help us provide better care.

Congenital Chagas disease (CCD) has become a global health problem. Historically, the diagnosis of CCD has been carried out using parasitological methods and traditional serological techniques, however, new serological techniques such as chemiluminescent microparticle immunoassays (CMIA) have been developed in the last few years with many advantages compared with traditional serological tests. A total of 75 children born to 72 Latin American Chagas-infected mothers were consecutively enrolled and studied by CMIA and indirect immunofluorescence (IIF) at 0-2, 6, 9, and 12 months of age. At the end of the follow-up, 74 out of 75 children were considered uninfected and one child was diagnosed with CCD. Our study emphasizes the need to carry out serological follow-up on every newborn from a mother with Chagas disease and shows that CMIA assay is a great diagnostic tool as a single serological test at 9 months of age to rule out CCD or to identify possible transmission.

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., 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 © 2021 Gómez, Thomas, Palacios, Egui, Carrilero, Simón, Valladares, Segovia, Carmelo and López.)

Background: Signs of senescence and the late stages of differentiation associated with the more severe forms of Chagas disease have been described in the Trypanosoma cruzi antigen-specific CD4+ T-cell population. However, the mechanisms involved in these functions are not fully known. To date, little is known about the possible impact of benznidazole treatment on the T. cruzi-specific functional response of CD4+ T cells., Methodology/principal Findings: The functional capacity of CD4+ T cells was analyzed by cytometric assays in chronic Chagas disease patients, with indeterminate form (IND) and cardiac alterations (CCC) (25 and 15, respectively) before and after benznidazole treatment. An increase in the multifunctional capacity (expression of IFN-γ, IL-2, TNF-α, perforin and/or granzyme B) of the antigen-specific CD4+ T cells was observed in indeterminate versus cardiac patients, which was associated with the reduced coexpression of inhibitory receptors (2B4, CD160, CTLA-4, PD-1 and/or TIM-3). The functional profile of these cells shows statistically significant differences between IND and CCC (p<0.001), with a higher proportion of CD4+ T cells coexpressing 2 and 3 molecules in IND (54.4% versus 23.1% and 4.1% versus 2.4%, respectively). A significant decrease in the frequencies of CD4+ T cells that coexpress 2, 3 and 4 inhibitory receptors was observed in IND after 24-48 months of treatment (p<0.05, p<0.01 and p<0.05, respectively), which was associated with an increase in antigen-specific multifunctional activity. The IND group showed, at 9-12 months after treatment, an increase in the CD4+ T cell subset coproducing three molecules, which were mainly granzyme B+, perforin+ and IFN-γ+ (1.4% versus 4.5%)., Conclusions/significance: A CD4+ T cell dysfunctional process was detected in chronic Chagas disease patients, being more exacerbated in those patients with cardiac symptoms. After short-term benznidazole treatment (9-12 months), indeterminate patients showed a significant increase in the frequency of multifunctional antigen-specific CD4+ T cells., Competing Interests: The authors have declared that no competing interests exist.

Background: Chagas disease has become a global health problem, with the pediatric population being especially vulnerable. Our aim was to describe the clinical-epidemiologic aspects of disease in this population, as well as tolerance and adherence to treatment and the subsequent evolution of the disease., Methods: A prospective study involving 949 children 0-14 years of age screened from 2007 to 2018. Diagnosis was performed by polymerase chain reaction and/or microhematocrit in <1-year-old children or serology in those ≥1 year of age. After diagnosis, children were examined for the clinical manifestation of Chagas disease and were treated with benznidazole. Treatment response was monitored by polymerase chain reaction and serology., Results: Forty children were infected (4.2% of the population screened). Twelve children were diagnosed during the acute phase (≤1-year-old), 3 of whom were symptomatic, and 28 (4- to 14-year-olds) were in the chronic phase: 18 in the indeterminate phase and 10 presented cardiac and/or digestive involvement. Regarding treatment, 10 (25.6%) children had side effects (6 mild, 2 moderate and 2 severe reactions), leading to treatment interruption in 3 of them. No side effects were detected in ≤1-year-old children (P < 0.05). Cure was confirmed in 29.4% of the children during follow-up, and the age of the children at treatment (≤1 year) was clearly associated with the effectiveness of treatment (P < 0.05)., Conclusions: Effectiveness and safety of treatment were optimum in ≤1-year-old children. Increased side effects, cardiac and/or digestive disorder incidence and lower treatment effectiveness were detected in older children, highlighting the need for early screening.

In recent decades and because of migration, Chagas disease has become a global public health problem. A significant focus has been placed on pregnant women who can transmit the disease to their offspring. Here, we report four cases of women who did not know that they were pregnant while they were being treated with benznidazole. A diagnosis was established according to serology and Trypanosoma cruzi polymerase chain reaction (PCR)-standardized tests. Treatment was discontinued when pregnancy was confirmed, and a thorough follow-up was carried out. Although each case was different, none of the mothers developed health problems during pregnancy, and their newborns were delivered without any teratogenic effects.

Objectives: Congenital Chagas Disease (CCD) has become a global health problem. Early diagnosis and treatment is essential for the cure of the disease. Our aim was to evaluate techniques and samples used for the diagnosis of CCD in order to improve diagnostic strategies., Methods: A total of 181 children born in Spain from Latin American Chagas-infected mothers were consecutively enrolled and studied by microhematocrit, PCR and serology tests at 0-2, 6 and 9-12 months of age and followed up when it was required. Samples of cord blood and peripheral blood were collected for T. cruzi detection by PCR. Parasite culture was performed in patients with a positive PCR., Results: Of 181 children, 7 children (3.9%) were lost to follow-up. A total of 174 children completed follow-up, 12 were diagnosed with CCD (6.9%) and 162 (93.1%) as uninfected children (negative serology tests at the end of the follow-up). Traditional parasitological diagnosis by microhematocrit had a poor performance (sensitivity was 10%), while PCR in peripheral blood showed high sensitivity (90.9%) and specificity (100%), allowing the early diagnosis of 9 infected children during the first 6-months-old. In the other 3 congenital cases, diagnosis was only possible at 12 months by serological and molecular techniques. However, PCR in cord blood showed low sensitivity (33.3%) and less specificity (96.4%) for the diagnosis., Conclusion: PCR in peripheral blood has proven to be the most adequate strategy for the diagnosis of CCD, allowing an early and reliable diagnosis., (Copyright © 2019 Elsevier B.V. All rights reserved.)

One of the current greatest challenges of Chagas disease is the establishment of biomarkers to assess the efficacy of drugs in a short period of time. In this context, the reactivity of sera from 66 adults with chronic indeterminate Chagas disease (IND) for a set of four Trypanosoma cruzi antigens (KMP11, PFR2, HSP70, and 3973 d ) was analyzed before and after benznidazole treatment. The results showed that the reactivity against these antigens decreased at 9, 24, and 48 months after treatment. Moreover, the 42.4% and 68.75% of IND patients met the established standard criteria of therapeutic efficacy (STEC) at 24 and 48 months posttreatment, respectively. Meeting the STEC implied that there was a continuous decrease in the reactivity of the patient sera against the four antigens after treatment and that there was a substantial decrease in the reactivity for at least two of the antigens. This important decrease in reactivity may be associated with a drastic reduction in the parasite load, but it is not necessarily associated with a parasitological cure. After treatment, a positive PCR result was only obtained in patients who did not meet the STEC. The percentage of granzyme B + /perforin + CD8 + T cells was significantly higher in patients who met the STEC than in those who did not meet the STEC (35.2% versus 2.2%; P  < 0.05). Furthermore, the patients who met the STEC exhibited an increased quality of the multifunctional response of the antigen-specific CD8 + T cells compared with that in the patients who did not meet the STEC., (Copyright © 2019 American Society for Microbiology.)

Congenital T. cruzi infections involve multiple factors in which complex interactions between the parasite and the immune system of pregnant women play important roles. In this study, we used an experimental murine model of chronic infection with T. cruzi to evaluate the changes in the expression of inhibitory receptors and the polyfunctionality of T cells during gestation and their association with congenital transmission rate of T. cruzi infection. The results showed that pregnant naïve mice had a higher percentage of CD4+ and CD8+ T cells that expressed inhibitory receptors than cells from non-pregnant naïve mice. However, in mice chronically infected with T. cruzi, gestation induced a significant decrease in the frequency of T cells that expressed or co-expressed inhibitory receptors, as well as an increase in the frequency of polyfunctional CD4+ and CD8+ T cells. This different behavior may be due to the breakdown in the infected mice of the gestation-induced immune homeostasis, probably to control the parasite load. Remarkably, it was observed that the mothers that transmitted the parasite had a higher frequency of T cells that expressed and co-expressed inhibitory receptors as well as a lower frequency of polyfunctional parasite-specific T cells than those that did not transmit it, even though the parasitemia load was similar in both groups. All together these data suggest that the maternal immune profile of the CD4+ and CD8+ T cells could be a determining factor in the congenital transmission of T. cruzi.

Background: We evaluated the effectiveness of treating women of childbearing age with benznidazole to prevent congenital Chagas disease (CCD), as well as the usefulness of polymerase chain reaction (PCR) as a tool to predict the risk of transmission., Methods: Prospective study involving 144 T. cruzi seropositive pregnant women. The parasitological status was studied by PCR in 159 pregnancies, 38 of which involved a cohort of previously treated mothers. One hundred sixty children were examined by PCR and serologically studied at 0-6, 9 and 12 months and annually after treatment., Results: PCR was seen to be useful for predicting the risk of congenital transmission: 18.8% of mothers with a positive PCR result transmitted the infection (16 infected children out of 85 pregnancies). No infected infants were detected among 74 pregnancies when PCR was negative. Of the treated mothers, 92.1% had negative PCR results, compared with 32.2% of untreated mothers. No infected infants were detected from previously treated mothers, compared with 13.2% among untreated mothers (P = .019; χ2). All infants treated before the first year of life were cured., Conclusions: Treating infected women of childbearing age prevents congenital Chagas disease. Polymerase chain reaction screening of T. cruzi-infected pregnant women is a useful tool for predicting the risk of congenital transmission., (© The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.)

Chagas disease is a chronic infection caused by Trypanosoma cruzi, an intracellular protozoan parasite. Chronic chagasic patients (CCPs) have dysfunctional CD8 + T cells that are characterized by impaired cytokine production, high coexpression of inhibitory receptors, and advanced cellular differentiation. Most patients diagnosed in the chronic phase of Chagas disease already exhibit heart involvement, and there is no vaccination that protects against the disease. Antiparasitic treatment is controversial as to its indication for this stage of the disease. There is a lack of biological markers to evaluate the effectiveness of antiparasitic treatment, and little is known about the effect of the treatment on CD8 + T cells. Thus, the aim of the current study was to analyze the early effects of antiparasitic treatment on CD8 + T cells from CCPs with asymptomatic clinical forms of disease. To evaluate the CD8 + T cell subsets, expression of inhibitory receptors, and functionality of T cells in CCPs, PBMCs were isolated. The results showed that treatment of CCPs with the asymptomatic form of the disease induces an increase in the frequency of CD8 + central memory T cells and terminal effector T cells, a decrease in the coexpression of inhibitory receptors, an improved Ag-specific CD8 + T cell response exhibited by the individual production of IFN-γ or IL-2, and a multifunctional CD8 + T cell profile of up to four functions (IFN-γ + IL-2 + Perforin + Granzyme B + ). These findings suggest that, in CCPs, antiparasitic treatment improved the quality of Ag-specific CD8 + T cell responses associated with a decrease in inhibitory receptor coexpression, which could serve as biomarkers for monitoring the effectiveness of antiparasitic treatment., (Copyright © 2017 by The American Association of Immunologists, Inc.)

Objectives: The objective was to characterize a Trypanosoma cruzi repetitive amino acid sequence that can be used as a marker of therapeutic drug efficacy in patients with chronic Chagas' disease., Methods: Reactivities to the 3973 peptide were measured in 85 patients with Chagas' disease (41 in the asymptomatic stage and 44 in the cardiomyopathy stage) before and 9 and 24 months after benznidazole administration. Additionally, the levels of IL-6 and C-reactive protein were measured in serum samples from patients with cardiomyopathy., Results: In 85% of the asymptomatic patients and 73% of the symptomatic chronic patients, modifications of the reactivity to the 3973 peptide were observed at 9 and 24 months post-benznidazole treatment. Significant variations in reactivities to the total antigens of T. cruzi were not observed at these times. Significant decreases in the reactivity to the 3973 peptide were observed after treatment in 20 of 41 (49%) asymptomatic patients and 15 of 44 (34%) cardiac chagasic patients (P < 0.001). In these patients, the decreases in reactivity at 24 months post-treatment were at least 40% lower than those detected before treatment. No correlations were found of the detected modifications in reactivity to the 3973 peptide after treatment with the levels of C-reactive protein or IL-6., Conclusions: Decreases in reactivity to the 3973 peptide may be relevant in the post-treatment follow-up of chronic chagasic patients., (© The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

It has been reported that the immune response mediated by T CD8+ lymphocytes plays a critical role in the control of Trypanosoma cruzi infection and that the clinical symptoms of Chagas disease appear to be related to the competence of the CD8+ T immune response against the parasite. Herewith, in silico prediction and binding assays on TAP-deficient T2 cells were used to identify potential HLA-A*02:01 ligands in the T. cruzi TcCA-2 protein. The TcCA-2-specific CD8+ T cells were functionality evaluated by Granzyme B and cytokine production in peripheral blood mononuclear cells (PBMC) from Chagas disease patients stimulated with the identified HLA-A*02:01 peptides. The specific cells were phenotypically characterized by flow cytometry using several surface markers and HLA-A*02:01 APC-labeled dextramer loaded with the peptides. In the T. cruzi TcCA-2 protein four T CD8+ epitopes were identified which are processed and presented during Chagas disease. Interestingly, a differential cellular phenotypic profile could be correlated with the severity of the disease. The TcCA-2-specific T CD8+ cells from patients with cardiac symptoms are mainly effector memory cells (TEM and TEMRA) while, those present in the asymptomatic phase are predominantly naive cells (TNAIVE). Moreover, in patients with cardiac symptoms the percentage of cells with senescence features is significantly higher than in patients at the asymptomatic phase of the disease. We consider that the identification of these new class I-restricted epitopes are helpful for designing biomarkers of sickness pathology as well as the development of immunotherapies against T. cruzi infection.

The innate immune response from diamniotic and dichorionic twin brothers congenitally infected with Trypanosoma. cruzi (strain DTU-V) who displayed different clinical symptomatology was studied. While Brother I manifested severe cardiac and digestive disorders, the Brother II showed slight splenomegaly. The secretion level of IL-1β, TNF-α, IL-12, IL-10, IFN-α and IL-6 cytokines produced after stimulation of peripheral blood cells with TLR-2, TLR-4 and TLR-9 ligands was determined pre- and post-benznidazole treatment. Cells from 10 uninfected infants born to mothers seropositive for Chagas disease were included as control. The obtained data show that the cells of Brother I secreted lower levels of the pro-inflammatory cytokines IL-1β and TNF-α (upon TLR-2 and TLR-4 stimulation) relative to those secreted by cells from Brother II and uninfected controls. The cells from Brother II secreted high levels of the IL-1β cytokine following TLR-2 stimulation relative to uninfected controls. The cells from both brothers secreted a higher level of IL-6, following TLR-4 stimulation, than that secreted by uninfected infant cells. After treatments, the cytokine secretion levels were similar in both children and comparable to those of uninfected donors. Treatment success in Brother I and treatment interruption in Brother II was detected by the use of serological biomarkers (KMP11, HSP70, PFR2, Tgp63) as well as follow-up done by PCR. Therefore, the Brother II required a second treatment. The data presented suggest that benznidazol treatment allows the innate immune system to reach a fully functional status similar to that of uninfected subjects., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Introduction: The increasing frequency of malaria infection in our area is due to the rise in international travel and immigration from endemic malaria areas. The aim of this study is to describe the chemoprophylaxis taken and treatment given as well as the clinical, epidemiological and microbiological characteristics for those patients admitted to our hospital with malaria., Methods: A retrospective study of patients with malaria admitted to the Hospital Virgen de la Arrixaca, between January 1998 and December 2010, was carried out., Results: During this period, fifty one cases of malaria were diagnosed. 78.3% of them were immigrants of whom 65% resided in Spain and had travelled to their country of origin for a short stay. Seventy four per cent acquired the infection in central and west Africa, and Plasmodium falciparum was responsible for the majority of the cases (88%). Only four patients had taken antimalarial chemoprophylaxis but none correctly. The most frequently treatment used was a combination of quinine and doxycicline (64.7%). Inappropriate anti-malarial treatment occurred in 9 patients (17.6%). At least one indicator of severe malaria was established in 23.5% of the cases; however, the clinical outcome was successful in every case and no patient died., Conclusions: Imported malaria is observed mostly among immigrants who travel to their countries of origin for a short stay and do not take anti-malarial prophylaxis, increasing the risk of acquiring malaria. Inappropriate malarial treatment is relatively frequent in the case management of imported malaria.

Background: In this longitudinal cohort study we evaluated the congenital transmission of Chagas disease (CD) in a nonendemic area. The aim of this work was to analyze the predictive value of a Trypanosoma cruzi-positive polymerase chain reaction (PCR) result in pregnant women for the diagnosis of vertical transmission and to evaluate the use of PCR as a tool for early detection of infection., Methods: The offspring of 59 seropositive pregnant mothers were followed up. The parasitological status of mothers was studied by PCR in a total of 64 pregnancies; 10 of these women had received treatment before pregnancy. Sixty-five infants (including a pair of twins) were monitored at 0, 6, 9, and 12 months of age by PCR and serology. In cases of congenital transmission, hemoculture and parasite lineage typing were performed., Results: Nine infants had acquired CD congenitally. This represents a transmission rate of 13.8% among seropositive mothers (9 infected newborns of 65 total live births). All infants were infected with T. cruzi discrete typing unit V strain. A statistically significant correlation was found between T. cruzi vertical transmission and a positive PCR result during pregnancy (31%; 9 infected newborns in 29 live births). No infected infants were detected among 10 mothers who were treated before they became pregnant, compared with 16.4% (9 of 55 live births) among untreated mothers., Conclusions: PCR is a useful tool for the detection of congenital CD, and the treatment of infected women of childbearing age seems to be useful for preventing vertical transmission.

Trypanosoma cruzi infection, or Chagas disease, was discovered more than 100 years ago by Carlos Chagas. Although the infection kills more than 15,000 people each year, it is still classified as a neglected tropical disease. Today, this disease affects eight million people in 21 Latin American countries and, due to immigration, is also present in non-endemic countries. In recent years, the size of the immigrant population with chronic imported forms of Chagas disease has increased in Spain. In addition, several cases of congenital transmission have been reported. Some patients have severe infection and require specialized treatment such as pacemaker implantation or even heart transplantation, representing a considerable clinical, social and economic burden, particularly in areas with a large immigrant population. Since the 1960s, the only drugs available for the etiological treatment of this infection have been benznidazole and nifurtimox. Although new, more effective and better tolerated compounds are urgently needed, treatment with these trypanocidal drugs is recommended in both the acute and chronic stages of Chagas disease. New strategies for diagnosis and infection control in chronically infected patients have recently been reported, allowing the effectiveness of treatments to be assessed., (Copyright © 2013 Elsevier España, S.L. All rights reserved.)

The protozoan parasite Trypanosoma cruzi is the etiological agent of Chagas disease. To date, no vaccine is available for protection against T. cruzi infection. The CD8(+) T cells immune response against specific antigens has shown to efficiently control the spread of the parasite in murine experimental infection. However, data concerning CD8(+) response in Chagas patients are still restricted to a few epitopes. We have studied the existence of immunodominant CD8(+) T cell epitopes in the paraflagellar rod proteins 2 and 3 (PFR2 and PFR3) from T. cruzi in a mouse model, and analyzed their recognition by cytotoxic T lymphocytes from Chagas disease patients. Immunization of C57BL/6-A2/K(b) transgenic mice with plasmids coding for the fusion proteins PFR2-HSP70 and PFR3-HSP70 induced a specific CTL response against two PFRs epitopes (PFR2(449-457) and PFR3(481-489)), and showed specific lysis percentages of 24 and 12, respectively. Moreover, the PFR2(19-28), PFR2(156-163), PFR2(449-457), PFR3(428-436), PFR3(475-482) and PFR3(481-489) peptides were observed to have a high binding affinity to the HLA-A*02:01 molecule. Remarkably, these HLA-A*02:01-binding peptides are successfully processed and presented during natural infection by T. cruzi in the context of MHC class I as evidenced by using peptide-pulsed K562-A2 cells as antigen presenting cells. The T cells from Chagas disease chronic patients specific for PFR2/PFR3 selected CD8(+) epitopes showed a pro-inflammatory cytokine secretion profile (IFN-γ, TNF-α and IL-6). A positive Granzime B secretion was observed in three out of 16 patients in response to PFR2(156-163) and PFR2(449-457) peptides, two out of 11 patients in response to PFR2(19-28) peptide and one out of 14 and 11 patients in response to PFR3(428-436) and PFR3(481-489) peptides, respectively. The PFRs-specific cytotoxic activity in purified PBMC was only detected in patients in the indeterminate phase of the disease., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Nowadays, the techniques available for chronic Chagas' disease diagnosis are very sensitive; however, they do not allow discrimination of the patient's clinical stages of the disease. The present paper describes that three out of the five different repeats contained in the Trypanosoma cruzi TcCA-2 membrane protein (3972-FGQAAAGDKPPP, 6303-FGQAAAGDKPAP, and 3973-FGQAAAGDKPSL) are recognized with high sensitivity (>90%) by sera from chronic Chagas' disease patients and that they are not recognized by sera from patients in the acute phase of the disease. A total of 133 serum samples from chagasic patients and 50 serum samples from healthy donors were tested. In addition, sera from 15 patients with different autoimmune diseases, 43 serum samples from patients suffering an infectious disease other than Chagas' disease, and 38 serum samples from patients with nonchagasic cardiac disorders were also included in this study. The residue 3973 peptide shows a specificity of >98%, as it is not recognized by individuals with autoimmune and inflammatory processes or by patients with a nonchagasic cardiomyopathy. Remarkably, the levels of antibody against the 3973 epitope detected by the sera from Chagas' disease patients in the symptomatic chronic phase, involving cardiac or digestive alterations, are higher than those detected by the sera from Chagas' disease patients in the indeterminate phase of the disease. It is suggested that the diagnostic technique described could also be used to indicate the degree of pathology. The amino acids F, Q, and DKP located in the peptide at positions 1, 3, and 8 to 10, respectively, are essential to conform to the immunodominant antigenic epitope.

As a result of population migration, Chagas disease is no longer limited to the North and South American continents. In HIV-infected patients, chronic infection by Trypanosoma cruzi behaves as an opportunistic infection in severely immunosuppressed patients and is responsible for high morbidity and mortality. Unlike other opportunistic infections, information on the natural history, diagnosis, treatment, and prevention of Chagas disease is scarce. Spain has the highest number of cases of Chagas disease outside the North and South American continents, and coinfection with HIV is increasingly prevalent. In this article, the Spanish Society for Tropical Medicine and International Health (Sociedad Española de Medicina Tropical y Salud Internacional) reviews the current situation of coinfection with HIV and T. cruzi infection and provides guidelines on the diagnosis, treatment, and prevention in areas where Chagas disease is not endemic. It also identifies areas of uncertainty where additional research is necessary.

CD8(+) cytotoxic T lymphocyte (CTL) response is critical for controlling the infection of the protozoan parasite Trypanosoma cruzi, the causative agent of Chagas disease. Since only a few CD8 antigens have been described in Chagas disease patients, the identification of new class I-restricted epitopes is urgently needed for the development of immunotherapies against T. cruzi infection. In this study, bioinformatic methods were used to predict HLA-A∗02:01-binders, and 30 peptides were selected, synthesized and tested for HLA-A∗02:01 binding. Among them, sixteen peptides with medium-to-high affinity were assayed for their recognition by CTL from HSP70-immunized or T. cruzi-infected transgenic B6-A2/K(b) mice. Our results show that four immunodominant epitopes (HSP70(210-8), HSP70(255-63), HSP70(316-24) and HSP70(345-53)) are contained in the T. cruzi HSP70 antigen. Indeed two of them (HSP70(210-8) and HSP70(316-24)) were also recognized by CTL of HLA-A∗02:01(+) Chagas disease patients, indicating that these peptides are processed and displayed as MHC class I epitopes during the natural history of T. cruzi infection. The HLA-A∗02:01 restriction was evidenced using peptide-pulsed K562-A2 cells as antigen-presenting cells. Both cytotoxic and cytokine-secreting activities were detected in response to the former two peptides and, moreover, 10/12 patients (83%) recognized at least one of these two HSP70-derived CD8(+) epitopes., (Copyright © 2011 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.)