Serological based monitoring of a cohort of patients with chronic Chagas disease treated with benznidazole in a highly endemic area of northern Argentina.

This study aimed to evaluate well-documented diagnostic antigens, named B13, 1F8 and JL7 recombinant proteins, as potential markers of seroconversion in treated chagasic patients. Prospective study, involving 203 patients treated with benznidazole, was conducted from endemic areas of northern Argentina. Follow-up was possible in 107 out of them and blood samples were taken for serology and PCR assays before and 2, 3, 6, 12, 24 and 36 months after treatment initiation.

Cloning and expression of transgenes using linear vectors in Trypanosoma cruzi.

The identification of new targets for vaccine and drug development for the treatment of Chagas' disease is dependent on deepening our understanding of the parasite genome. Vectors for genetic manipulation in Trypanosoma cruzi basically include those that remain as circular episomes and those that integrate into the parasite's genome. Artificial chromosomes are alternative vectors to overcome problematic transgene expression often occurring with conventional vectors in this parasite.

Proteins associated with SF3a60 in T. brucei.

Trypanosoma brucei relies on Spliced leader trans splicing to generate functional messenger RNAs. Trans splicing joins the specialized SL exon from the SL RNA to pre-mRNAs and is mediated by the trans-spliceosome, which is made up of small nuclear ribonucleoprotein particles and non-snRNP factors. Although the trans spliceosome is essential for trypanosomatid gene expression, not all spliceosomal protein factors are known and of these, only a few are completely characterized.

Evaluation of in-house ELISA using Trypanosoma cruzi lysate and recombinant antigens for diagnosis of Chagas disease and discrimination of its clinical forms.

The aim of this work was to investigate the potential usefulness of Trypanosoma cruzi lysate, recombinant protein JL7, and peptides P013, R13, JL18, JL19, and P0β as serological markers for human Chagas disease. We analyzed 228 sera from Brazilian Chagas disease patients classified into four clinical groups and 108 from non-chagasic patients. We defined the diagnostic sensitivity, specificity, and Kappa index measured by enzyme-linked immunosorbent assay (ELISA).

Selective blockade of trypanosomatid protein synthesis by a recombinant antibody anti-Trypanosoma cruzi P2β protein.

The ribosomal P proteins are located on the stalk of the ribosomal large subunit and play a critical role during the elongation step of protein synthesis. The single chain recombinant antibody C5 (scFv C5) directed against the C-terminal region of the Trypanosoma cruzi P2β protein (TcP2β) recognizes the conserved C-terminal end of all T. cruzi ribosomal P proteins.

Crystal structure of the complex mAb 17.2 and the C-terminal region of Trypanosoma cruzi P2β protein: implications in cross-reactivity.

Patients with Chronic Chagas' Heart Disease possess high levels of antibodies against the carboxyl-terminal end of the ribosomal P2ß protein of Trypanosoma cruzi (TcP2ß). These antibodies, as well as the murine monoclonal antibody (mAb) 17.2, recognize the last 13 amino acids of TcP2ß (called the R13 epitope: EEEDDDMGFGLFD) and are able to cross-react with, and stimulate, the ß1 adrenergic receptor (ß1-AR).

Human recombinant antibodies against Trypanosoma cruzi ribosomal P2β protein.

Patients with chronic Chagas' Heart Disease (cChHD) develop an antibody response that is suspected to be involved in the cardiac pathogenesis. The response against Trypanosoma cruzi ribosomal P proteins is of particular interest, as these antibodies can cross-react with host cardiac receptors causing electrophysiological alterations. To better understand the humoral anti-P response we constructed a single-chain variable fragment library derived from a cChHD patient.

Interaction map of the Trypanosoma cruzi ribosomal P protein complex (stalk) and the elongation factor 2.

The large subunit of the eukaryotic ribosome possesses a long and protruding stalk formed by the ribosomal P proteins. This structure is involved in the translation step of protein synthesis through interaction with the elongation factor 2 (EF-2). The Trypanosoma cruzi stalk complex is composed of four proteins of about 11 kDa, TcP1α, TcP1β, TcP2α, TcP2β and a fifth TcP0 of about 34 kDa.

Antibodies against the Trypanosoma cruzi ribosomal P proteins induce apoptosis in HL-1 cardiac cells.

High levels of antibodies (Abs) against the C-terminal end of the Trypanosoma cruzi ribosomal P2β protein, defined by the R13 peptide, are detected in sera from patients with chronic Chagas heart disease (cChHD). These Abs can cross-react with the β1-adrenergic receptor (β1-AR), inducing a functional response in cardiomyocytes. In this study, we report that a monoclonal Ab against the R13 peptide, called mAb 17.

Trypanosoma cruzi I genotypes in different geographical regions and transmission cycles based on a microsatellite motif of the intergenic spacer of spliced-leader genes.

The intergenic region of spliced-leader (SL-IR) genes from 105 Trypanosoma cruzi I (Tc I) infected biological samples, culture isolates and stocks from 11 endemic countries, from Argentina to the USA were characterised, allowing identification of 76 genotypes with 54 polymorphic sites from 123 aligned sequences. On the basis of the microsatellite motif proposed by Herrera et al. (2007) to define four haplotypes in Colombia, we could classify these genotypes into four distinct Tc I SL-IR groups, three corresponding to the former haplotypes Ia (11 genotypes), Ib (11 genotypes) and Id (35 genotypes); and one novel group, Ie (19 genotypes).

Molecular identification of Trypanosoma cruzi discrete typing units in end-stage chronic Chagas heart disease and reactivation after heart transplantation.

Background: One hundred years after the discovery of Chagas disease, it remains a major neglected tropical disease. Chronic Chagas heart disease (cChHD) is the most severe manifestation. Heart transplantation is the proper treatment for end-stage heart failure, although reactivation of disease may result after receipt of immunosuppressive therapy.

The heavy chain variable segment gene repertoire in chronic Chagas' heart disease.

Patients chronically infected with Trypanosoma cruzi develop chronic Chagas' heart disease (cChHD). Their Ab response is suspected to be involved in the cardiac pathogenesis. Reactivity of serum Abs from these patients has been extensively studied but little is known about the diversity of the in vivo IgG repertoire.

Molecular diagnosis and treatment monitoring of congenital transmission of Trypanosoma cruzi to twins of a triplet delivery.

Congenital transmission of Trypanosoma cruzi was diagnosed in 2 triplets born to a triamniotic bichorionic delivery. Only the sisters sharing the placenta became infected, as diagnosed by microhematocrit and/or polymerase chain reaction of 3 parasite targets. The neonates' parasitologic response to benznidazole was monitored.

Identification of core components of the exon junction complex in trypanosomes.

In animal cells, the exon junction complex (EJC) is deposited onto mRNAs during the second step of splicing, 20-24 nt upstream of the exon-exon junction. The EJC core contains four proteins: Mago, Y14, eIF4AIII and Btz. In trypanosomes, cis-splicing is very rare but all mRNAs are subject to 5'trans-splicing of a 39-nt RNA sequence.

Accurate real-time PCR strategy for monitoring bloodstream parasitic loads in chagas disease patients.

Background: This report describes a real-time PCR (Q-PCR) strategy to quantify Trypanosoma cruzi (T. cruzi) DNA in peripheral blood samples from Chagas disease patients targeted to conserved motifs within the repetitive satellite sequence.

Methodology/principal Findings: The Q-PCR has a detection limit of 0.

The FIP-1 like polyadenylation factor in trypanosomes and the structural basis for its interaction with CPSF30.

In trypanosomes transcription is polycistronic and individual mRNAs are generated by a trans-splicing/polyadenylation coupled reaction. We identified a divergent trypanosome FIP1-like, a factor required for mRNA 3' end formation from yeasts to human. Here we showed that it is a nuclear protein with a speckled distribution essential for trypanosome viability.

Functional characterization and protein-protein interactions of trypanosome splicing factors U2AF35, U2AF65 and SF1.

Early in the assembly of eukaryotes the branch-point binding protein (BBP, also called SF1) recognizes the branch point sequence, whereas the heterodimer U2AF, consisting of a 65 and a 35 kDa subunit, contacts the polypyrimidine tract and the AG splice site, respectively. Herein, we identified, cloned and expressed the Trypanosoma cruzi and Trypanosoma brucei U2AF35, U2AF65 and SF1. Trypanosomatid U2AF65 strongly diverged from yeast and human homologues.

Proteomic analysis of the Trypanosoma cruzi ribosomal proteins.

Trypanosoma cruzi is a parasite responsible for Chagas disease. The identification of new targets for chemotherapy is a major challenge for the control of this disease. Several lines of evidences suggest that the translational system in trypanosomatids show important differences compared to other eukaryotes.

Cross-talk between anti-beta1-adrenoceptor antibodies in dilated cardiomyopathy and Chagas' heart disease.

Anti-beta(1)-adrenoceptor autoantibodies, first described in sera of patients with Chagas' disease, are now well documented in patients with idiopathic dilated cardiomyopathy. The following review summarizes the knowledge we have about the structural basis of receptor-antibody interactions, about their mechanisms of action and about their pathogenicity. While the origin of anti-receptor antibodies with agonist-like activity in Chagas' disease might be ascribed to recognition by anti-parasite antibodies of an epitope, localized on the second extracellular loop of the beta(1)-adrenoceptor, the origin of such antibodies in idiopathic dilated cardiomyopathy remains unknown.

Molecular epidemiology of domestic and sylvatic Trypanosoma cruzi infection in rural northwestern Argentina.

Genetic diversity of Trypanosoma cruzi populations and parasite transmission dynamics have been well documented throughout the Americas, but few studies have been conducted in the Gran Chaco ecoregion, one of the most highly endemic areas for Chagas disease, caused by T. cruzi. In this study, we assessed the distribution of T.

The C-terminal end of P proteins mediates ribosome inactivation by trichosanthin but does not affect the pokeweed antiviral protein activity.

Ribosome inactivating proteins (RIPs) inhibit protein synthesis depurinating a conserved residue in the sarcin/ricin loop of ribosomes. Some RIPs are only active against eukaryotic ribosomes, but other RIPs inactivate with similar efficiency prokaryotic and eukaryotic ribosomes, suggesting that different RIPs would interact with different proteins. The SRL in Trypanosoma cruzi ribosomes is located on a 178b RNA molecule named 28Sdelta.

Molecular identification of Trypanosoma cruzi I tropism for central nervous system in Chagas reactivation due to AIDS.

Trypanosoma cruzi lineages, microsatellite allelic polymorphism, and mithocondrial gene haplotypes were directly typified from peripheral blood and cerebrospinal fluid specimens of a Bolivian patient with Chagas disease with accompanying AIDS and central nervous system severe involvement. Of note, the patient's blood was infected by a mixture of T. cruzi I and T.

Mapping of the protein-binding interface between splicing factors SF3b155 and p14 of Trypanosoma cruzi.

SF3b155 and p14 are essential components of spliceosome core that recognize the branch point adenosine, a critical step in splicing in eukaryotes. Trypanosomes are unusual since every transcribed gene is processed by trans-splicing instead of cis-splicing. Thus, the trans-spliceosome emerges as an interesting anti-parasitic drug target since this process is not present in mammalian hosts.

Direct molecular profiling of minicircle signatures and lineages of Trypanosoma cruzi bloodstream populations causing congenital Chagas disease.

Congenital transmission of Trypanosoma cruzi may occur in some or all the gestations from a T. cruzi-infected mother. Variable rates of congenital transmission have been reported in different geographical areas where different parasitic strains predominate, suggesting that parasitic genotypes might play a role in the risk of congenital transmission.

Gene expression patterns in Euglena gracilis: insights into the cellular response to environmental stress.

To better understand Euglena gracilis gene expression under different stress conditions (Chromium, Streptomycin or darkness), we undertook a survey of the E. gracilis transcriptome by cDNA sequencing and microarray analysis. First, we constructed a non-normalized cDNA library from the E.