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Sample records for bloodstream stage trypanosoma

  1. In vitro cultivation of Trypanosoma acomys: production of insect stages and bloodstream forms.

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    Maraghi, S; Wallbanks, K R; Molyneux, D H; Abdel-Hafez, S K

    1995-01-01

    When Trypanosoma acomys bloodstream forms were cultivated at 37 degrees C in Schneider's Drosophila medium supplemented with 20% (v/v) heat-inactivated foetal calf serum (FCS), with Microtus agrestis embryonic fibroblasts in RPMI 1640 medium supplemented with 20% FCS or in Baltz's medium supplemented with 10% FCS, the parasites transformed and largely remained as epimastigotes. Epimastigotes were also usually the commonest stage observed when the parasites were co-cultivated with a mosquito cell line at 27 degrees C. However, if these cultures were initiated with the supernatant suspensions from fibroblast cultures that had been cryopreserved, trypomastigotes, including bloodstream-like forms, were the predominant stage for the first 4 days of culture. It is suggested that the glycerol supplement or the temperature changes stimulated this unusual morphogenesis. At 27 degrees C, T. acomys was incapable of multiplying and died when cultured in fresh Schneider's Drosophila medium supplemented with 20% FCS, but co-cultivation with the mosquito cell lines or cultivation in cell-free supernatants from 1-week-old mosquito cell cultures was successful at this temperature; most of the parasites multiplied as epimastigotes.

  2. Diverse effects on mitochondrial and nuclear functions elicited by drugs and genetic knockdowns in bloodstream stage Trypanosoma brucei.

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    Christal Worthen

    Full Text Available BACKGROUND: The options for treating the fatal disease human African trypanosomiasis are limited to a few drugs that are toxic or facing increasing resistance. New drugs that kill the causative agents, subspecies of Trypanosoma brucei, are therefore urgently needed. Little is known about the cellular mechanisms that lead to death of the pathogenic bloodstream stage. METHODOLOGY/PRINCIPAL FINDINGS: We therefore conducted the first side by side comparison of the cellular effects of multiple death inducers that target different systems in bloodstream form parasites, including six drugs (pentamidine, prostaglandin D(2, quercetin, etoposide, camptothecin, and a tetrahydroquinoline and six RNAi knockdowns that target distinct cellular functions. All compounds tested were static at low concentrations and killed at high concentrations. Dead parasites were rapidly quantified by forward and side scatter during flow cytometry, as confirmed by ethidium homodimer and esterase staining, making these assays convenient for quantitating parasite death. The various treatments yielded different combinations of defects in mitochondrial potential, reactive oxygen species, cell cycle, and genome segregation. No evidence was seen for phosphatidylserine exposure, a marker of apoptosis. Reduction in ATP levels lagged behind decreases in live cell number. Even when the impact on growth was similar at 24 hours, drug-treated cells showed dramatic differences in their ability to further proliferate, demonstrating differences in the reversibility of effects induced by the diverse compounds. CONCLUSIONS/SIGNIFICANCE: Parasites showed different phenotypes depending on the treatment, but none of them were clear predictors of whether apparently live cells could go on to proliferate after drugs were removed. We therefore suggest that clonal proliferation assays may be a useful step in selecting anti-trypanosomal compounds for further development. Elucidating the genetic or

  3. Genome-wide expression profiling of in vivo-derived bloodstream parasite stages and dynamic analysis of mRNA alterations during synchronous differentiation in Trypanosoma brucei

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    Ghazal Peter

    2009-09-01

    Full Text Available Abstract Background Trypanosomes undergo extensive developmental changes during their complex life cycle. Crucial among these is the transition between slender and stumpy bloodstream forms and, thereafter, the differentiation from stumpy to tsetse-midgut procyclic forms. These developmental events are highly regulated, temporally reproducible and accompanied by expression changes mediated almost exclusively at the post-transcriptional level. Results In this study we have examined, by whole-genome microarray analysis, the mRNA abundance of genes in slender and stumpy forms of T.brucei AnTat1.1 cells, and also during their synchronous differentiation to procyclic forms. In total, five biological replicates representing the differentiation of matched parasite populations derived from five individual mouse infections were assayed, with RNAs being derived at key biological time points during the time course of their synchronous differentiation to procyclic forms. Importantly, the biological context of these mRNA profiles was established by assaying the coincident cellular events in each population (surface antigen exchange, morphological restructuring, cell cycle re-entry, thereby linking the observed gene expression changes to the well-established framework of trypanosome differentiation. Conclusion Using stringent statistical analysis and validation of the derived profiles against experimentally-predicted gene expression and phenotypic changes, we have established the profile of regulated gene expression during these important life-cycle transitions. The highly synchronous nature of differentiation between stumpy and procyclic forms also means that these studies of mRNA profiles are directly relevant to the changes in mRNA abundance within individual cells during this well-characterised developmental transition.

  4. KREX2 is not essential for either procyclic or bloodstream form Trypanosoma brucei.

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    Jason Carnes

    Full Text Available BACKGROUND: Most mitochondrial mRNAs in Trypanosoma brucei require RNA editing for maturation and translation. The edited RNAs primarily encode proteins of the oxidative phosphorylation system. These parasites undergo extensive changes in energy metabolism between the insect and bloodstream stages which are mirrored by alterations in RNA editing. Two U-specific exonucleases, KREX1 and KREX2, are both present in protein complexes (editosomes that catalyze RNA editing but the relative roles of each protein are not known. METHODOLOGY/PRINCIPAL FINDINGS: The requirement for KREX2 for RNA editing in vivo was assessed in both procyclic (insect and bloodstream form parasites by methods that use homologous recombination for gene elimination. These studies resulted in null mutant cells in which both alleles were eliminated. The viability of these cells demonstrates that KREX2 is not essential in either life cycle stage, despite certain defects in RNA editing in vivo. Furthermore, editosomes isolated from KREX2 null cells require KREX1 for in vitro U-specific exonuclease activity. CONCLUSIONS: KREX2 is a U-specific exonuclease that is dispensable for RNA editing in vivo in T. brucei BFs and PFs. This result suggests that the U deletion activity, which is required for RNA editing, is primarily mediated in vivo by KREX1 which is normally found associated with only one type of editosome. The retention of the KREX2 gene implies a non-essential role or a role that is essential in other life cycle stages or conditions.

  5. Ribose 5-phosphate isomerase B knockdown compromises Trypanosoma brucei bloodstream form infectivity.

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    Loureiro, Inês; Faria, Joana; Clayton, Christine; Macedo-Ribeiro, Sandra; Santarém, Nuno; Roy, Nilanjan; Cordeiro-da-Siva, Anabela; Tavares, Joana

    2015-01-01

    Ribose 5-phosphate isomerase is an enzyme involved in the non-oxidative branch of the pentose phosphate pathway, and catalyzes the inter-conversion of D-ribose 5-phosphate and D-ribulose 5-phosphate. Trypanosomatids, including the agent of African sleeping sickness namely Trypanosoma brucei, have a type B ribose-5-phosphate isomerase. This enzyme is absent from humans, which have a structurally unrelated ribose 5-phosphate isomerase type A, and therefore has been proposed as an attractive drug target waiting further characterization. In this study, Trypanosoma brucei ribose 5-phosphate isomerase B showed in vitro isomerase activity. RNAi against this enzyme reduced parasites' in vitro growth, and more importantly, bloodstream forms infectivity. Mice infected with induced RNAi clones exhibited lower parasitaemia and a prolonged survival compared to control mice. Phenotypic reversion was achieved by complementing induced RNAi clones with an ectopic copy of Trypanosoma cruzi gene. Our results present the first functional characterization of Trypanosoma brucei ribose 5-phosphate isomerase B, and show the relevance of an enzyme belonging to the non-oxidative branch of the pentose phosphate pathway in the context of Trypanosoma brucei infection.

  6. JVG9, a benzimidazole derivative, alters the surface and cytoskeleton of Trypanosoma cruzi bloodstream trypomastigotes

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    Díaz-Chiguer, Dylan L; Hernández-Luis, Francisco; Nogueda-Torres, Benjamín; Castillo, Rafael; Reynoso-Ducoing, Olivia; Hernández-Campos, Alicia; Ambrosio, Javier R

    2014-01-01

    Trypanosoma cruzi has a particular cytoskeleton that consists of a subpellicular network of microtubules and actin microfilaments. Therefore, it is an excellent target for the development of new anti-parasitic drugs. Benzimidazole 2-carbamates, a class of well-known broad-spectrum anthelmintics, have been shown to inhibit the in vitro growth of many protozoa. Therefore, to find efficient anti-trypanosomal (trypanocidal) drugs, our group has designed and synthesised several benzimidazole derivatives. One, named JVG9 (5-chloro-1H-benzimidazole-2-thiol), has been found to be effective against T. cruzi bloodstream trypomastigotes under both in vitro and in vivo conditions. Here, we present the in vitro effects observed by laser scanning confocal and scanning electron microscopy on T. cruzi trypomastigotes. Changes in the surface and the distribution of the cytoskeletal proteins are consistent with the hypothesis that the trypanocidal activity of JVG9 involves the cytoskeleton as a target. PMID:25317703

  7. Bloodstream form pre-adaptation to the tsetse fly in Trypanosoma brucei.

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    Rico, Eva; Rojas, Federico; Mony, Binny M; Szoor, Balazs; Macgregor, Paula; Matthews, Keith R

    2013-01-01

    African trypanosomes are sustained in the bloodstream of their mammalian hosts by their extreme capacity for antigenic variation. However, for life cycle progression, trypanosomes also must generate transmission stages called stumpy forms that are pre-adapted to survive when taken up during the bloodmeal of the disease vector, tsetse flies. These stumpy forms are rather different to the proliferative slender forms that maintain the bloodstream parasitaemia. Firstly, they are non proliferative and morphologically distinct, secondly, they show particular sensitivity to environmental cues that signal entry to the tsetse fly and, thirdly, they are relatively robust such that they survive the changes in temperature, pH and proteolytic environment encountered within the tsetse midgut. These characteristics require regulated changes in gene expression to pre-adapt the parasite and the use of environmental sensing mechanisms, both of which allow the rapid initiation of differentiation to tsetse midgut procyclic forms upon transmission. Interestingly, the generation of stumpy forms is also regulated and periodic in the mammalian blood, this being governed by a density-sensing mechanism whereby a parasite-derived signal drives cell cycle arrest and cellular development both to optimize transmission and to prevent uncontrolled parasite multiplication overwhelming the host. In this review we detail recent developments in our understanding of the molecular mechanisms that underpin the production of stumpy forms in the mammalian bloodstream and their signal perception pathways both in the mammalian bloodstream and upon entry into the tsetse fly. These discoveries are discussed in the context of conserved eukaryotic signaling and differentiation mechanisms. Further, their potential to act as targets for therapeutic strategies that disrupt parasite development either in the mammalian bloodstream or upon their transmission to tsetse flies is also discussed.

  8. Differential Gel Electrophoresis (DIGE) Evaluation of Naphthoimidazoles Mode of Action: A Study in Trypanosoma cruzi Bloodstream Trypomastigotes

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    Brunoro, Giselle Villa Flor; Faça, Vitor Marcel; Caminha, Marcelle Almeida; Ferreira, André Teixeira da Silva; Trugilho, Monique; de Moura, Kelly Cristina Gallan; Perales, Jonas; Valente, Richard Hemmi; Menna-Barreto, Rubem Figueiredo Sadok

    2016-01-01

    Background The obligate intracellular protozoan Trypanosoma cruzi is the causative agent of Chagas disease, a neglected illness affecting millions of people in Latin America that recently entered non-endemic countries through immigration, as a consequence of globalization. The chemotherapy for this disease is based mainly on benznidazole and nifurtimox, which are very efficient nitroderivatives against the acute stage but present limited efficacy during the chronic phase. Our group has been studying the trypanocidal effects of naturally occurring quinones and their derivatives, and naphthoimidazoles derived from β-lapachone N1, N2 and N3 were the most active. To assess the molecular mechanisms of action of these compounds, we applied proteomic techniques to analyze treated bloodstream trypomastigotes, which are the clinically relevant stage of the parasite. Methodology/Principal Findings The approach consisted of quantification by 2D-DIGE followed by MALDI-TOF/TOF protein identification. A total of 61 differentially abundant protein spots were detected when comparing the control with each N1, N2 or N3 treatment, for 34 identified spots. Among the differentially abundant proteins were activated protein kinase C receptor, tubulin isoforms, asparagine synthetase, arginine kinase, elongation factor 2, enolase, guanine deaminase, heat shock proteins, hypothetical proteins, paraflagellar rod components, RAB GDP dissociation inhibitor, succinyl-CoA ligase, ATP synthase subunit B and methionine sulfoxide reductase. Conclusion/Significance Our results point to different modes of action for N1, N2 and N3, which indicate a great variety of metabolic pathways involved and allow for novel perspectives on the development of trypanocidal agents. PMID:27551855

  9. Probing the metabolic network in bloodstream-form Trypanosoma brucei using untargeted metabolomics with stable isotope labelled glucose.

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    Darren J Creek

    2015-03-01

    Full Text Available Metabolomics coupled with heavy-atom isotope-labelled glucose has been used to probe the metabolic pathways active in cultured bloodstream form trypomastigotes of Trypanosoma brucei, a parasite responsible for human African trypanosomiasis. Glucose enters many branches of metabolism beyond glycolysis, which has been widely held to be the sole route of glucose metabolism. Whilst pyruvate is the major end-product of glucose catabolism, its transamination product, alanine, is also produced in significant quantities. The oxidative branch of the pentose phosphate pathway is operative, although the non-oxidative branch is not. Ribose 5-phosphate generated through this pathway distributes widely into nucleotide synthesis and other branches of metabolism. Acetate, derived from glucose, is found associated with a range of acetylated amino acids and, to a lesser extent, fatty acids; while labelled glycerol is found in many glycerophospholipids. Glucose also enters inositol and several sugar nucleotides that serve as precursors to macromolecule biosynthesis. Although a Krebs cycle is not operative, malate, fumarate and succinate, primarily labelled in three carbons, were present, indicating an origin from phosphoenolpyruvate via oxaloacetate. Interestingly, the enzyme responsible for conversion of phosphoenolpyruvate to oxaloacetate, phosphoenolpyruvate carboxykinase, was shown to be essential to the bloodstream form trypanosomes, as demonstrated by the lethal phenotype induced by RNAi-mediated downregulation of its expression. In addition, glucose derivatives enter pyrimidine biosynthesis via oxaloacetate as a precursor to aspartate and orotate.

  10. Quantitative Proteomic and Phosphoproteomic Analysis of Trypanosoma cruzi Amastigogenesis

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    Queiroz, Rayner M L; Charneau, Sebastien; Mandacaru, Samuel C;

    2014-01-01

    Chagas disease is a tropical neglected disease endemic in Latin America and it is caused by the protozoan Trypanosoma cruzi. The parasite has four major life stages: epimastigote, metacyclic trypomastigote, bloodstream trypomastigote and amastigote. The differentiation from infective trypomastigo......Chagas disease is a tropical neglected disease endemic in Latin America and it is caused by the protozoan Trypanosoma cruzi. The parasite has four major life stages: epimastigote, metacyclic trypomastigote, bloodstream trypomastigote and amastigote. The differentiation from infective...

  11. Isolation and characterization of kinetoplast DNA from the bloodstream form of Trypanosoma brucei.

    NARCIS (Netherlands)

    A.H. Fairlamb; P.O. Weislogel; J.H.J. Hoeijmakers (Jan); P. Borst (Piet)

    1978-01-01

    textabstractWe have used restriction endonucleases PstI, EcoRI, HapII, HhaI, and S1 nuclease to demonstrate the presence of a large complex component, the maxi-circle, in addition to the major mini-circle component in kinetoplast DNA (kDNA) networks of Trypanosoma brucei (East African Trypanosomiasi

  12. Alkaloids Induce Programmed Cell Death in Bloodstream Forms of Trypanosomes (Trypanosoma b. brucei

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    Michael Wink

    2008-10-01

    Full Text Available The potential induction of a programmed cell death (PCD in Trypanosoma b. brucei by 55 alkaloids of the quinoline, quinolizidine, isoquinoline, indole, terpene, tropane, steroid, and piperidine type was studied by measuring DNA fragmentation and changes in mitochondrial membrane potential. For comparison, the induction of apoptosis by the same alkaloids in human leukemia cells (Jurkat APO-S was tested. Several alkaloids of the isoquinoline, quinoline, indole and steroidal type (berberine, chelerythrine, emetine, sanguinarine, quinine, ajmalicine, ergotamine, harmine, vinblastine, vincristine, colchicine, chaconine, demissidine and veratridine induced programmed cell death, whereas quinolizidine, tropane, terpene and piperidine alkaloids were mostly inactive. Effective PCD induction (EC50 below 10 µM was caused in T. brucei by chelerythrine, emetine, sanguinarine, and chaconine. The active alkaloids can be characterized by their general property to inhibit protein biosynthesis, to intercalate DNA, to disturb membrane fluidity or to inhibit microtubule formation.

  13. Mathematical modelling of polyamine metabolism in bloodstream-form Trypanosoma brucei: an application to drug target identification.

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    Xu Gu

    Full Text Available We present the first computational kinetic model of polyamine metabolism in bloodstream-form Trypanosoma brucei, the causative agent of human African trypanosomiasis. We systematically extracted the polyamine pathway from the complete metabolic network while still maintaining the predictive capability of the pathway. The kinetic model is constructed on the basis of information gleaned from the experimental biology literature and defined as a set of ordinary differential equations. We applied Michaelis-Menten kinetics featuring regulatory factors to describe enzymatic activities that are well defined. Uncharacterised enzyme kinetics were approximated and justified with available physiological properties of the system. Optimisation-based dynamic simulations were performed to train the model with experimental data and inconsistent predictions prompted an iterative procedure of model refinement. Good agreement between simulation results and measured data reported in various experimental conditions shows that the model has good applicability in spite of there being gaps in the required data. With this kinetic model, the relative importance of the individual pathway enzymes was assessed. We observed that, at low-to-moderate levels of inhibition, enzymes catalysing reactions of de novo AdoMet (MAT and ornithine production (OrnPt have more efficient inhibitory effect on total trypanothione content in comparison to other enzymes in the pathway. In our model, prozyme and TSHSyn (the production catalyst of total trypanothione were also found to exhibit potent control on total trypanothione content but only when they were strongly inhibited. Different chemotherapeutic strategies against T. brucei were investigated using this model and interruption of polyamine synthesis via joint inhibition of MAT or OrnPt together with other polyamine enzymes was identified as an optimal therapeutic strategy.

  14. Involvement of lysosomes in the uptake of macromolecular material by bloodstream forms of Trypanosoma brucei.

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    Opperdoes, F R; Van Roy, J

    1982-09-01

    To investigate whether the lysosomes of Trypanosoma brucei are capable of uptake of macromolecules after internalization by the cell, we used Triton WR-1339, a non-digestible macromolecular compound, which is known to cause a marked decrease in the density of hepatic lysosomes due to massive intralysosomal storage. Intraperitoneal administration of 0.4 g/kg Triton WR-1339 to rats infected with T. brucei led to the development of a large vacuole in the trypanosomes between nucleus and kinetoplast within 22 h. Higher doses (2 g/kg) led to the disappearance of the trypanosomes from the blood and resulted in permanent cures (greater than 100 days). Lysosomes isolated from the trypanosomes of animals treated with a sub-curative dose showed a decrease in equilibrium density of 0.03 g/cm3 in sucrose gradients. These lysosomes were partly damaged as evidenced by a reduction in latency and an increase in the non-sedimentable part of lysosomal enzymes. We conclude that acid proteinase and alpha-mannosidase-containing organelles of T. brucei take up exogenous macromolecules and must therefore be considered as true lysosomes and that Triton WR-1339 acts in T. brucei as a true lysosomotropic drug. Its trypanocidal action probably results from an interference with lysosomal function.

  15. Evaluation of some organic compounds on bloodstream forms of Trypanosoma cruzi

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    João S. Silva

    1992-09-01

    Full Text Available Accidental transmission of Chagas' disease to man by blood transfusion is a serious problem in Latin-America. This paper describes the testing of several synthetic, semi-synthetic, and natural compounds for their activity against blood trypomastigotes in vitro at 4-C. The compounds embody several types of chemical structures: benzoquinone, naphthoquinone, anthracenequinone, phenanthrenequinone, imidazole, piperazine, quinoline, xanthene, and simple benzenic and naphthalenic derivates. Some of them are for the first time tested against Trypanosoma cruzi. The toxic effect these compounds on this parasite was done by two quite distinct sets of experiments. In one set, the compounds were added to infected blood as ethanolic solution. In this situation the most active one was a furan-1, 2-naphthoquinone, in the same range as gentian violet, a new fact to be considered in the assessment of structure-activity relationships in this class of compounds. In other set, we tentatively evaluated the biological activity of water insoluble compounds by adding them in a pure form without solvent into infected blood. In this way some appear to be very active and it was postulated that the effectiveness of such compounds must result from interactions between them and specific blood components.

  16. Paraflagellar rod proteins administered with alum and IL-12 or recombinant adenovirus expressing IL-12 generates antigen-specific responses and protective immunity in mice against Trypanosoma cruzi.

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    Wrightsman, R A; Manning, J E

    2000-01-31

    Successful vaccination of mice against an otherwise lethal challenge with the Peru strain of Trypanosoma cruzi necessitates the induction of a strong cell mediated immune response. Previously, immunization of mice with the paraflagellar rod proteins from Trypanosoma cruzi90% reduction in parasitemia in immunized mice challenged with the bloodstream stage of Trypanosoma cruzi.

  17. Biosynthesis and uptake of thiamine (vitamin B1) in bloodstream form Trypanosoma brucei brucei and interference of the vitamin with melarsen oxide activity.

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    Stoffel, Sabine A; Rodenko, Boris; Schweingruber, Anne-Marie; Mäser, Pascal; de Koning, Harry P; Schweingruber, M Ernst

    2006-02-01

    Bloodstream forms of Trypanosoma brucei brucei were cultivated in the presence and absence of thiamine (vitamin B1) and pyridoxine (vitamin B6). The vitamins do not change growth behaviour, indicating that Trypanosoma brucei is prototrophic for the two vitamins even though in silico no bona-fide thiamine-biosynthetic genes could be identified in the T. brucei genome. Intracellularly, thiamine is mainly present in its diphosphate form. We were unable to detect significant uptake of [3H]thiamine and structural thiamine analogues such as pyrithiamine, oxithiamine and amprolium were not toxic for the bloodstream forms of T. brucei, indicating that the organism does not have an efficient uptake system for thiamine and its analogues. We have previously shown that, in the fission yeast Saccharomyces pombe, the toxicity of melarsen oxide, the pharmacologically active derivative of the frontline sleeping sickness drug melarsoprol, is abolished by thiamine and the drug is taken up by a thiamine-regulated membrane protein which is responsible for the utilization of thiamine. We show here that thiamine also has weak effects on melarsen oxide-induced growth inhibition and lysis in T. brucei. These effects were consistent with a low affinity of thiamine for the P2 adenosine transporter that is responsible for uptake of melaminophenyl arsenicals in African trypanosomes.

  18. A simple method to purify biologically and antigenically preserved bloodstream trypomastigotes of Trypanosoma cruzi using Deae-cellulose columns

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    Maria Auxiliadora de Sousa

    1983-09-01

    Full Text Available A method to purify trypanosomastigotes of some strains of Trypanosoma cruzi (Y, CL, FL, F, "Berenice", "Colombiana" and "São Felipe" from mouse blood by using DEAE-cellulose columns was standardized. This procedure is a modification of the Lanham & Godfrey methods and differs in some aspects from others described to purify T. cruzi bloodstream trypomastigotes, mainly by avoidance of prior purifications of parasites. By this method, the broad trypomastigotes were mainly isolated, accounting for higher recoveries obtained with strains having higher percentages of these forms: processing of infected blood from irradiated mice could be advantageous by increasing the recovery of parasites (percentage and/or total number and elution of more slender trypomastigotes. Trypomastigotes purified by this method presented normal morphology and motility, remained infective to triatomine bugs and mice, showing in the latter prepatent periods and courses parasitemia similar to those of control parasites, and also reproducing the polymorphism pattern of each strain. Their virulence and pathogenicity also remained considerably preserved, the latter property being evaluated by LD 50 tests, mortality rates and mean survival time of inoculated mice. Moreover, these parasites presented positive, clear and peripheral immunofluorescence reaction at titres similar to those of control organisms, thus suggesting important preservation of their surface antigens.Usando colunas de DEAE-cellulose foi padronizado um método para purificação de tripomastigotas de várias cepas de Trypanosoma cruzi (Y, CL, FL, F, "Berenice", "Columbiana" e "São Felipe" a partir do sangue de camundongos. Este método é uma modificação daqueles descritos por Lanham & Godfrey e difere em vários aspectos de outros descritos para purificar as formas sanguíneas deste parasita, particularmente na dispensa de pré-purificações. Por ele foram isolados principalmente os tripomastigotas largos

  19. Comprehensive glycoprofiling of the epimastigote and trypomastigote stages of Trypanosoma cruzi

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    Alves, Maria Julia Manso; Kawahara, Rebeca; Viner, Rosa

    2017-01-01

    Trypanosoma cruzi, the protozoan that causes Chagas disease, has a complex life cycle involving insect and mammalian hosts and distinct developmental stages. During T. cruzi developmental stages, glycoproteins play important role in the host-parasite interaction, such as cellular recognition, hos...

  20. Transcriptomic analysis reveals metabolic switches and surface remodeling as key processes for stage transition in Trypanosoma cruzi

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    Greif, Gonzalo; Rodriguez, Matias; Alvarez-Valin, Fernando

    2017-01-01

    American trypanosomiasis is a chronic and endemic disease which affects millions of people. Trypanosoma cruzi, its causative agent, has a life cycle that involves complex morphological and functional transitions, as well as a variety of environmental conditions. This requires a tight regulation of gene expression, which is achieved mainly by post-transcriptional regulation. In this work we conducted an RNAseq analysis of the three major life cycle stages of T. cruzi: amastigotes, epimastigotes and trypomastigotes. This analysis allowed us to delineate specific transcriptomic profiling for each stage, and also to identify those biological processes of major relevance in each state. Stage specific expression profiling evidenced the plasticity of T. cruzi to adapt quickly to different conditions, with particular focus on membrane remodeling and metabolic shifts along the life cycle. Epimastigotes, which replicate in the gut of insect vectors, showed higher expression of genes related to energy metabolism, mainly Krebs cycle, respiratory chain and oxidative phosphorylation related genes, and anabolism related genes associated to nucleotide and steroid biosynthesis; also, a general down-regulation of surface glycoprotein coding genes was seen at this stage. Trypomastigotes, living extracellularly in the bloodstream of mammals, express a plethora of surface proteins and signaling genes involved in invasion and evasion of immune response. Amastigotes mostly express membrane transporters and genes involved in regulation of cell cycle, and also express a specific subset of surface glycoprotein coding genes. In addition, these results allowed us to improve the annotation of the Dm28c genome, identifying new ORFs and set the stage for construction of networks of co-expression, which can give clues about coded proteins of unknown functions. PMID:28286708

  1. Transcriptomic analysis reveals metabolic switches and surface remodeling as key processes for stage transition in Trypanosoma cruzi

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    Luisa Berná

    2017-03-01

    Full Text Available American trypanosomiasis is a chronic and endemic disease which affects millions of people. Trypanosoma cruzi, its causative agent, has a life cycle that involves complex morphological and functional transitions, as well as a variety of environmental conditions. This requires a tight regulation of gene expression, which is achieved mainly by post-transcriptional regulation. In this work we conducted an RNAseq analysis of the three major life cycle stages of T. cruzi: amastigotes, epimastigotes and trypomastigotes. This analysis allowed us to delineate specific transcriptomic profiling for each stage, and also to identify those biological processes of major relevance in each state. Stage specific expression profiling evidenced the plasticity of T. cruzi to adapt quickly to different conditions, with particular focus on membrane remodeling and metabolic shifts along the life cycle. Epimastigotes, which replicate in the gut of insect vectors, showed higher expression of genes related to energy metabolism, mainly Krebs cycle, respiratory chain and oxidative phosphorylation related genes, and anabolism related genes associated to nucleotide and steroid biosynthesis; also, a general down-regulation of surface glycoprotein coding genes was seen at this stage. Trypomastigotes, living extracellularly in the bloodstream of mammals, express a plethora of surface proteins and signaling genes involved in invasion and evasion of immune response. Amastigotes mostly express membrane transporters and genes involved in regulation of cell cycle, and also express a specific subset of surface glycoprotein coding genes. In addition, these results allowed us to improve the annotation of the Dm28c genome, identifying new ORFs and set the stage for construction of networks of co-expression, which can give clues about coded proteins of unknown functions.

  2. Cell surface proteome analysis of human-hosted Trypanosoma cruzi life stages

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    Queiroz, Rayner M L; Charneau, Sébastien; Bastos, Izabela M D;

    2014-01-01

    Chagas' disease is a neglected infectious illness, caused by the protozoan Trypanosoma cruzi. It remains a challenging health issue in Latin America, where it is endemic, and so far there is no immunoprophylatic vaccine or satisfactory chemotherapic treatment for its chronic stage. The present work...

  3. A luciferase based viability assay for ATP detection in 384-well format for high throughput whole cell screening of Trypanosoma brucei brucei bloodstream form strain 427

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    Avery Vicky M

    2009-11-01

    Full Text Available Abstract Background Human African Trypanosomiasis (HAT is caused by two trypanosome species, Trypanosoma brucei rhodesiense and Trypanosoma brucei gambiense. Current drugs available for the treatment of HAT have significant issues related to toxicity, administration regimes with limited effectiveness across species and disease stages, thus there is a considerable need to find alternative drugs. A well recognised approach to identify new drug candidates is high throughput screening (HTS of large compound library collections. Results We describe here the development of a luciferase based viability assay in 384-well plate format suitable for HTS of T.b.brucei. The parameters that were explored to determine the final HTS assay conditions are described in detail and include DMSO tolerability, Z', diluents and cell inoculum density. Reference compound activities were determined for diminazene, staurosporine and pentamidine and compared to previously published IC50 data obtained. The assay has a comparable sensitivity to reference drugs and is more cost effective than the 96-well format currently reported for T.b.brucei. Conclusion Due to the reproducibility and sensitivity of this assay it is recommended for potential HTS application. As it is commercially available this assay can also be utilised in many laboratories for both large and small scale screening.

  4. Proteomic analysis of Trypanosoma cruzi developmental stages using isotope-coded affinity tag reagents.

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    Paba, Jaime; Ricart, Carlos A O; Fontes, Wagner; Santana, Jaime M; Teixeira, Antonio R L; Marchese, Jason; Williamson, Brian; Hunt, Tony; Karger, Barry L; Sousa, Marcelo V

    2004-01-01

    Comparative proteome analysis of developmental stages of the human pathogen Trypanosoma cruzi was carried out by isotope-coded affinity tag technology (ICAT) associated with liquid cromatography-mass spectrometry peptide sequencing (LC-MS/MS). Protein extracts of the protozoan trypomastigote and amastigote stages were labeled with heavy (D8) and light (D0) ICAT reagents and subjected to cation exchange and avidin affinity chromatographies followed by LC-MS/MS analysis. High confidence sequence information and expression levels for 41 T. cruzi polypeptides, including metabolic enzymes, paraflagellar rod components, tubulins, and heat-shock proteins were reported. Twenty-nine proteins displayed similar levels of expression in both forms of the parasite, nine proteins presented higher levels in trypomastigotes, whereas three were more expressed in amastigotes.

  5. Immunospecific immunoglobulins and IL-10 as markers for Trypanosoma brucei rhodesiense late stage disease in experimentally infected vervet monkeys

    DEFF Research Database (Denmark)

    Ngotho, Maina; Kagira, J.M.; Jensen, Henrik Michael Elvang

    2009-01-01

    OBJECTIVE: To determine the usefulness of IL-10 and immunoglobulin M (IgM) as biomarkers for staging HAT in vervet monkeys, a useful pathogenesis model for humans. METHODS: Vervet monkeys were infected with Trypanosoma brucei rhodesiense and subsequently given sub-curative and curative treatment 28...

  6. Nuclear Compartmentalization Contributes to Stage-Specific Gene Expression Control in Trypanosoma cruzi

    Science.gov (United States)

    Pastro, Lucía; Smircich, Pablo; Di Paolo, Andrés; Becco, Lorena; Duhagon, María A.; Sotelo-Silveira, José; Garat, Beatriz

    2017-01-01

    In the protozoan parasite Trypanosoma cruzi, as in other trypanosomatids, transcription of protein coding genes occurs in a constitutive fashion, producing large polycistronic transcription units. These units are composed of non-functionally related genes which are pervasively processed to yield each mRNA. Therefore, post-transcriptional processes are crucial to regulate gene expression. Considering that nuclear compartmentalization could contribute to gene expression regulation, we comparatively studied the nuclear, cytoplasmic and whole cell transcriptomes of the non-infective epimastigote stage of T. cruzi, using RNA-Seq. We found that the cytoplasmic transcriptome tightly correlates with the whole cell transcriptome and both equally correlate with the proteome. Nonetheless, 1,200 transcripts showed differential abundance between the nuclear and cytoplasmic fractions. For the genes with transcript content augmented in the nucleus, significant structural and compositional differences were found. The analysis of the reported epimastigote translatome and proteome, revealed scarce ribosome footprints and encoded proteins for them. Ontology analyses unveiled that many of these genes are distinctive of other parasite life-cycle stages. Finally, the relocalization of transcript abundance in the metacyclic trypomastigote infective stage was confirmed for specific genes. While gene expression is strongly dependent on transcript steady-state level, we here highlight the importance of the distribution of transcripts abundance between compartments in T. cruzi. Particularly, we show that nuclear compartmentation is playing an active role in the developmental stage determination preventing off-stage expression. PMID:28243589

  7. Nitroheterocyclic drugs cure experimental Trypanosoma cruzi infections more effectively in the chronic stage than in the acute stage

    Science.gov (United States)

    Francisco, Amanda Fortes; Jayawardhana, Shiromani; Lewis, Michael D.; White, Karen L.; Shackleford, David M.; Chen, Gong; Saunders, Jessica; Osuna-Cabello, Maria; Read, Kevin D.; Charman, Susan A.; Chatelain, Eric; Kelly, John M.

    2016-01-01

    The insect-transmitted protozoan parasite Trypanosoma cruzi is the causative agent of Chagas disease, and infects 5–8 million people in Latin America. Chagas disease is characterised by an acute phase, which is partially resolved by the immune system, but then develops as a chronic life-long infection. There is a consensus that the front-line drugs benznidazole and nifurtimox are more effective against the acute stage in both clinical and experimental settings. However, confirmative studies have been restricted by difficulties in demonstrating sterile parasitological cure. Here, we describe a systematic study of nitroheterocyclic drug efficacy using highly sensitive bioluminescence imaging of murine infections. Unexpectedly, we find both drugs are more effective at curing chronic infections, judged by treatment duration and therapeutic dose. This was not associated with factors that differentially influence plasma drug concentrations in the two disease stages. We also observed that fexinidazole and fexinidazole sulfone are more effective than benznidazole and nifurtimox as curative treatments, particularly for acute stage infections, most likely as a result of the higher and more prolonged exposure of the sulfone derivative. If these findings are translatable to human patients, they will have important implications for treatment strategies. PMID:27748443

  8. Bloodstream infection in patients with end-stage renal disease in a teaching hospital in central-western Brazil

    Directory of Open Access Journals (Sweden)

    Tamara Trelha Gauna

    2013-08-01

    Full Text Available Introduction Vascular access in patients undergoing hemodialysis is considered a critical determinant of bloodstream infection (BSI and is associated with high morbidity and mortality. The purpose of this study was to investigate the occurrence of BSI in patients with end-stage renal disease using central venous catheters for hemodialysis. Methods A cohort study was conducted in a public teaching hospital in central-western Brazil from April 2010 to December 2011. For every patient, we noted the presence of hyperemia/exudation upon catheter insertion, as well as fever, shivering, and chills during hemodialysis. Results Fifty-nine patients were evaluated. Thirty-five (59.3% patients started dialysis due to urgency, 37 (62.7% had BSI, and 12 (20% died. Hyperemia at the catheter insertion site (64.9% was a significant clinical manifestation in patients with BSI. Statistical analysis revealed 1.7 times more cases of BSI in patients with hypoalbuminemia compared with patients with normal albumin levels. The principal infective agents identified in blood cultures and catheter-tip cultures were Staphylococcus species (24 cases, non-fermentative Gram-negative bacilli (7 cases of Stenotrophomonas maltophilia and 5 cases of Chryseobacterium indologenes, and Candida species (6. Among the Staphylococci identified, 77.7% were methicillin-resistant, coagulase-negative Staphylococci. Of the bacteria isolated, the most resistant were Chryseobacterium indologenes and Acinetobacter baumannii. Conclusions Blood culture was demonstrated to be an important diagnostic test and identified over 50% of positive BSI cases. The high frequency of BSI and the isolation of multiresistant bacteria were disturbing findings. Staphylococcus aureus was the most frequently isolated microorganism, although Gram-negative bacteria predominated overall. These results highlight the importance of infection prevention and control measures in dialysis units.

  9. [Early stages of development of Trypanosoma rotatorium (Mayer, 1843) from peripheral blood and internal organs of Anurans Bufo bufo (Linnaeus) and Rana sp. (Anura)].

    Science.gov (United States)

    Malysheva, M N

    2014-01-01

    The data on the fauna of trypanosomes of Anura of the Leningrad Province are given. The initial development stages of Trypanosoma rotatorium in peripheral blood and internal organs of the frog are described for the first time.

  10. Variation in antigenic determinants specific to the infective stage of Trypanosoma cruzi.

    Science.gov (United States)

    Wrightsman, R A; Leon, W; Manning, J E

    1986-08-01

    Monoclonal antibodies reactive with the surface antigens of the Peru strain of Trypanosoma cruzi were analyzed by Western blots and immunofluorescence assays to determine their reactivity with three life cycle stages and five strain isolates of T. cruzi. One monoclonal antibody, 7.6, recognized a 68-kilodalton (kDa) polypeptide in Western blots of Peru strain trypomastigotes, epimastigotes, and amastigotes. A 68-kDa polypeptide was also detected by monoclonal antibody 7.6 in trypomastigotes of the CL and Y strains and in the clonal isolates Esmeraldo clone 3 and Silvio X10 clone 1. Positive immunofluorescence results were obtained for all life cycle stages of the five strains that were reacted with monoclonal antibody 7.6, thus indicating that the antigen recognized by monoclonal antibody 7.6 is universally present in all T. cruzi strains tested. In contrast, monoclonal antibody 4.2 reacted with a polypeptide doublet of 90 and 105 kDa in Western blots of Peru strain trypomastigotes, but it did not detect these antigens in epimastigotes or amastigotes. The same polypeptide doublet of 90 and 105 kDa was also detected in Western blots of Y strain trypomastigotes; however, no bands were detected in blots of strain CL or isolate Silvio X10 clone 1 trypomastigotes. In blots of Esmeraldo clone 3 trypomastigotes, a single band of 130 kDa was detected by monoclonal antibody 4.2. In immunofluorescence assays of monoclonal antibody 4.2, positive reactions were obtained only with trypomastigotes of Peru, Y, and Esmeraldo clone 3 strains. Thus, monoclonal antibody 4.2 recognizes a trypomastigote-specific antigen which is not universally present on all strains of T. cruzi.

  11. Identification of monoclonal antibodies against the trypomastigote stage of Trypanosoma cruzi by use of iminobiotinylated surface polypeptides.

    Science.gov (United States)

    Beard, C A; Wrightsman, R A; Manning, J E

    1985-08-01

    The surface polypeptides of epimastigotes and tissue culture-derived trypomastigotes of Trypanosoma cruzi have been isolated free of most cytosolic components by use of the 2-iminobiotin-avidin interaction. Polypeptides of the trypomastigote stage obtained by this technique are recognized by serum antibodies from Chagasic patients and T. cruzi-infected mice. These polypeptides have been used as the detecting antigen for the identification of hybridoma cells producing monoclonal antibodies against the surface proteins of the trypomastigote stage of T. cruzi. These experiments document a practical approach for obtaining T. cruzi surface proteins in sufficient quantity and purity for use in immunological studies.

  12. Trypanosoma cruzi alkaline 2-DE: Optimization and application to comparative proteome analysis of flagellate life stages

    Directory of Open Access Journals (Sweden)

    Santana Jaime M

    2008-09-01

    Full Text Available Abstract Background Trypanosoma cruzi, a flagellate protozoan, is the etiological agent of Chagas disease, a chronic illness that causes irreversible damage to heart and digestive tract in humans. Previous 2-DE analyses of T. cruzi proteome have not focused on basic proteins, possibly because of inherent difficulties for optimizing 2-DE in the alkaline pH range. However, T. cruzi wide pH range 2-DE gels have shown few visible spots in the alkaline region, indicating that the parasite either did not have an appreciable amount of alkaline proteins or that these proteins were underrepresented in the 2-DE gels. Results Different IEF conditions using 6–11 pH gradient strips were tested for separation of T. cruzi alkaline proteins. The optimized methodology described here was performed using anodic "paper bridge" sample loading supplemented by increased concentration of DTT and Triton X-100 on Multiphor II (GE Healthcare equipment and an electrode pad embedded in DTT- containing solution near the cathode in order to avoid depletion of reducing agent during IEF. Landmark proteins were identified by peptide mass fingerprinting allowing the production of an epimastigote 2-DE map. Most identified proteins corresponded to metabolic enzymes, especially those related to amino acid metabolism. The optimized 2-DE protocol was applied in combination with the "two-in-one gel" method to verify the relative expression of the identified proteins between samples from epimastigote and trypomastigote life stages. Conclusion High resolution 2-DE gels of T. cruzi life forms were achieved using the optimized methodology and a partial epimastigote alkaline 2-DE map was built. Among 700 protein spots detected, 422 were alkaline with a pI above 7.0. The "two-in-one gel" method simplified the comparative analysis between T. cruzi life stages since it minimized variations in spot migration and silver-stained spot volumes. The comparative data were in agreement with

  13. Validation of N-myristoyltransferase as Potential Chemotherapeutic Target in Mammal-Dwelling Stages of Trypanosoma cruzi.

    Directory of Open Access Journals (Sweden)

    Linda J Herrera

    2016-04-01

    Full Text Available Trypanosoma cruzi causes Chagas disease, an endemic and debilitating illness in Latin America. Lately, owing to extensive population movements, this neglected tropical disease has become a global health concern. The two clinically available drugs for the chemotherapy of Chagas disease have rather high toxicity and limited efficacy in the chronic phase of the disease, and may induce parasite resistance. The development of new anti-T. cruzi agents is therefore imperative. The enzyme N-myristoyltransferase (NMT has recently been biochemically characterized, shown to be essential in Leishmania major, Trypanosoma brucei, and T. cruzi¸ and proposed as promising chemotherapeutic target in these trypanosomatids.Here, using high-content imaging we assayed eight known trypanosomatid NMT inhibitors, against mammal-dwelling intracellular amastigote and trypomastigote stages and demonstrated that three of them (compounds 1, 5, and 8 have potent anti-proliferative effect at submicromolar concentrations against T. cruzi, with very low toxicity against human epithelial cells. Moreover, metabolic labeling using myristic acid, azide showed a considerable decrease in the myristoylation of proteins in parasites treated with NMT inhibitors, providing evidence of the on-target activity of the inhibitors.Taken together, our data point out to the potential use of NMT inhibitors as anti-T. cruzi chemotherapy.

  14. On opportunist infections by Trypanosoma lewisi in humans and its differential diagnosis from T. cruzi and T. rangeli.

    Science.gov (United States)

    de Sousa, Maria Auxiliadora

    2014-12-01

    Trypanosoma lewisi is a cosmopolitan species originally found in Rattus spp., being nonpathogenic, host-restricted, and transmitted by rat fleas. This species has been recorded as an opportunist blood parasite of human beings mainly in Asia, with a case in Africa. In Brazil, this species was recently recorded in captive monkeys. As T. lewisi can share vertebrate hosts both with Trypanosoma rangeli and Trypanosoma cruzi, some markers for the differential diagnosis of these species were examined and discussed herein. The identification of T. lewisi was based on morphological features of bloodstream stages at the initial phase of infection in mammals, isoenzyme electrophoresis at the MDH locus, and PCR products of kinetoplast DNA (kDNA) minicircles using the primers TC121/TC122.

  15. Stage and strain specific expression of the tandemly repeated 90 kDa surface antigen gene family in Trypanosoma cruzi.

    Science.gov (United States)

    Beard, C A; Wrightsman, R A; Manning, J E

    1988-04-01

    A recombinant cDNA library constructed in the expression vector lambda gtll using mRNA from the trypomastigote stage of Trypanosoma cruzi was screened with two monoclonal antibodies that have been shown to react with a 105 kDa and a 90 kDa surface antigen in trypomastigotes of the Peru and Y strains of T. cruzi. One recombinant lambda phage, designated Tcc-20, was reactive to both monoclonals. The beta-galactosidase/T. cruzi hybrid protein encoded in Tcc-20 is recognized by the monoclonal antibodies and by serum antibodies from mice infected with strains of T. cruzi which contain the 90 kDa antigen. Antibodies immunoselected from serum of mice infected with the Peru strain by adsorption to Tcc-20 fusion protein react specifically with a 90 kDa polypeptide in trypomastigote but not epimastigote lysates of T. cruzi. The mRNA complementary to the DNA insert in Tcc-20 is present only in those stages and strains of T. cruzi which express the 90 kDa surface antigen. These characteristics are strong evidence that the T. cruzi DNA fragment cloned into Tcc-20 encodes a portion of the 90 kDa surface antigen. The gene(s) which encodes this polypeptide is shown to be present in approximately 20 copies per haploid genome and most, and possibly all, of the copies are found in a tandemly linked multigene family.

  16. Trypanosoma evansi contains two auxiliary enzymes of glycolytic metabolism: Phosphoenolpyruvate carboxykinase and pyruvate phosphate dikinase.

    Science.gov (United States)

    Rivero, Luz Amira; Concepción, Juan Luis; Quintero-Troconis, Ender; Quiñones, Wilfredo; Michels, Paul A M; Acosta, Héctor

    2016-06-01

    Trypanosoma evansi is a monomorphic protist that can infect horses and other animal species of economic importance for man. Like the bloodstream form of the closely related species Trypanosoma brucei, T. evansi depends exclusively on glycolysis for its free-energy generation. In T. evansi as in other kinetoplastid organisms, the enzymes of the major part of the glycolytic pathway are present within organelles called glycosomes, which are authentic but specialized peroxisomes. Since T. evansi does not undergo stage-dependent differentiations, it occurs only as bloodstream forms, it has been assumed that the metabolic pattern of this parasite is identical to that of the bloodstream form of T. brucei. However, we report here the presence of two additional enzymes, phosphoenolpyruvate carboxykinase and PPi-dependent pyruvate phosphate dikinase in T. evansi glycosomes. Their colocalization with glycolytic enzymes within the glycosomes of this parasite has not been reported before. Both enzymes can make use of PEP for contributing to the production of ATP within the organelles. The activity of these enzymes in T. evansi glycosomes drastically changes the model assumed for the oxidation of glucose by this parasite.

  17. Deletion of a novel protein kinase with PX and FYVE-related domains increases the rate of differentiation of Trypanosoma brucei.

    Science.gov (United States)

    Vassella, E; Krämer, R; Turner, C M; Wankell, M; Modes, C; van den Bogaard, M; Boshart, M

    2001-07-01

    Growth control of African trypanosomes in the mammalian host is coupled to differentiation of a non-dividing life cycle stage, the stumpy bloodstream form. We show that a protein kinase with novel domain architecture is important for growth regulation. Zinc finger kinase (ZFK) has a kinase domain related to RAC and S6 kinases flanked by a FYVE-related zinc finger and a phox (PX) homology domain. To investigate the function of the kinase during cyclical development, a stable transformation procedure for bloodstream forms of differentiation-competent (pleomorphic) Trypanosoma brucei strains was established. Deletion of both allelic copies of ZFK by homologous recombination resulted in reduced growth of bloodstream-form parasites in culture, which was correlated with an increased rate of differentiation to the non-dividing stumpy form. Growth and differentiation rates were returned to wild-type level by ectopic ZFK expression. The phenotype is stage-specific, as growth of procyclic (insect form) trypanosomes was unaffected, and Deltazfk/Deltazfk clones were able to undergo full cyclical development in the tsetse fly vector. Deletion of ZFK in a differentiation-defective (monomorphic) strain of T. brucei did not change its growth rate in the bloodstream stage. This suggests a function of ZFK associated with the trypanosomes' decision between either cell cycle progression, as slender bloodstream form, or differentiation to the non-dividing stumpy form.

  18. Expression and subcellular localization of kinetoplast-associated proteins in the different developmental stages of Trypanosoma cruzi

    Directory of Open Access Journals (Sweden)

    Cavalcanti Danielle

    2009-06-01

    Full Text Available Abstract Background The kinetoplast DNA (kDNA of trypanosomatids consists of an unusual arrangement of circular molecules catenated into a single network. The diameter of the isolated kDNA network is similar to that of the entire cell. However, within the kinetoplast matrix, the kDNA is highly condensed. Studies in Crithidia fasciculata showed that kinetoplast-associated proteins (KAPs are capable of condensing the kDNA network. However, little is known about the KAPs of Trypanosoma cruzi, a parasitic protozoon that shows distinct patterns of kDNA condensation during their complex morphogenetic development. In epimastigotes and amastigotes (replicating forms the kDNA fibers are tightly packed into a disk-shaped kinetoplast, whereas trypomastigotes (non-replicating present a more relaxed kDNA organization contained within a rounded structure. It is still unclear how the compact kinetoplast disk of epimastigotes is converted into a globular structure in the infective trypomastigotes. Results In this work, we have analyzed KAP coding genes in trypanosomatid genomes and cloned and expressed two kinetoplast-associated proteins in T. cruzi: TcKAP4 and TcKAP6. Such small basic proteins are expressed in all developmental stages of the parasite, although present a differential distribution within the kinetoplasts of epimastigote, amastigote and trypomastigote forms. Conclusion Several features of TcKAPs, such as their small size, basic nature and similarity with KAPs of C. fasciculata, are consistent with a role in DNA charge neutralization and condensation. Additionally, the differential distribution of KAPs in the kinetoplasts of distinct developmental stages of the parasite, indicate that the kDNA rearrangement that takes place during the T. cruzi differentiation process is accompanied by TcKAPs redistribution.

  19. In or out? On the tightness of glycosomal compartmentalization of metabolites and enzymes in Trypanosoma brucei

    NARCIS (Netherlands)

    Haanstra, Jurgen R.; Bakker, Barbara M.; Michels, Paul A. M.

    2014-01-01

    Trypanosomatids sequester large parts of glucose metabolism inside specialised peroxisomes, called glycosomes. Many studies have shown that correct glycosomal compartmentalization of glycolytic enzymes is essential for bloodstream-form Trypanosoma brucel. The recent finding of pore-forming activitie

  20. Pure paraflagellar rod protein protects mice against Trypanosoma cruzi infection.

    OpenAIRE

    1995-01-01

    The paraflagellar rod proteins (PAR) purified from Trypanosoma cruzi epimastigotes were shown to protect mice against an otherwise lethal challenge inoculum of 10(3) bloodstream-form trypomastigotes. The injection route used for immunization was shown to have a marked impact on the development of protective immunity. Mice receiving subcutaneous (s.c.) injections of PAR proteins had reduced bloodstream parasitemias and showed 100% survival following challenge. In contrast, mice immunized via t...

  1. Sesquiterpene lactones and the diterpene 5-epi-icetexone affect the intracellular and extracellular stages of Trypanosoma cruzi

    NARCIS (Netherlands)

    Lozano, E.; Barrera, P.; Salinas, R.; Vega, I.; Nieto, M.; Tonn, C.; Kemmerling, U.; Mortara, R.A.; Sosa, M.A.

    2012-01-01

    Chagas disease is a major health problem in Latin America and is caused by the parasitic protozoan Trypanosoma cruzi. Although many drugs have been used to alleviate the disease, these have been ineffective in the chronic phase and have also presented numerous side effects on patients. In this study

  2. Kinetoplast adaptations in American strains from Trypanosoma vivax

    Energy Technology Data Exchange (ETDEWEB)

    Greif, Gonzalo [Unidad de Biología Molecular, Institut Pasteur de Montevideo (Uruguay); Rodriguez, Matías [Sección Biomatemática, Facultad de Ciencias, Universidad de la Republica (Uruguay); Reyna-Bello, Armando [Departamento de Ciencias de la Vida, Carrera en Ingeniería en Biotecnología, Universidad de las Fuerzas Armadas (Ecuador); Centro de Estudios Biomédicos y Veterinarios, Universidad Nacional Experimental Simón Rodríguez-IDECYT, Caracas (Venezuela, Bolivarian Republic of); Robello, Carlos [Unidad de Biología Molecular, Institut Pasteur de Montevideo (Uruguay); Departamento de Bioquímica, Facultad de Medicina, Universidad de la República Uruguay (Uruguay); Alvarez-Valin, Fernando, E-mail: falvarez@fcien.edu.uy [Sección Biomatemática, Facultad de Ciencias, Universidad de la Republica (Uruguay)

    2015-03-15

    Highlights: • American T. vivax strains exhibit a drastic process of mitochondrial genome degradation. • T. vivax mitochondrial genes have among the fastest evolutionary rates in eukaryotes. • High rates of kDNA evolution are associated with relaxation of selective constrains. • Relaxed selective pressures are the result of mechanical transmission. • The evolutionary strategy of T. vivax differs from that of T. brucei-species complex. - Abstract: The mitochondrion role changes during the digenetic life cycle of African trypanosomes. Owing to the low abundance of glucose in the insect vector (tsetse flies) the parasites are dependent upon a fully functional mitochondrion, capable of performing oxidative phosphorylation. Nevertheless, inside the mammalian host (bloodstream forms), which is rich in nutrients, parasite proliferation relies on glycolysis, and the mitochondrion is partially redundant. In this work we perform a comparative study of the mitochondrial genome (kinetoplast) in different strains of Trypanosoma vivax. The comparison was conducted between a West African strain that goes through a complete life cycle and two American strains that are mechanically transmitted (by different vectors) and remain as bloodstream forms only. It was found that while the African strain has a complete and apparently fully functional kinetoplast, the American T. vivax strains have undergone a drastic process of mitochondrial genome degradation, in spite of the recent introduction of these parasites in America. Many of their genes exhibit different types of mutations that are disruptive of function such as major deletions, frameshift causing indels and missense mutations. Moreover, all but three genes (A6-ATPase, RPS12 and MURF2) are not edited in the American strains, whereas editing takes place normally in all (editable) genes from the African strain. Two of these genes, A6-ATPase and RPS12, are known to play an essential function during bloodstream stage

  3. Trypanosoma cruzi: a stage-specific calpain-like protein is induced after various kinds of stress

    Directory of Open Access Journals (Sweden)

    Viviane Giese

    2008-09-01

    Full Text Available Calpains are calcium-dependent cysteine proteinases found in all living organisms and are involved in diverse cellular processes. Calpain-like proteins have been reported after in silico analysis of the Tritryps genome and are believed to play important roles in cell functions of trypanosomatids. We describe the characterization of a member of this family, which is differentially expressed during the life-cycle of Trypanosoma cruzi.

  4. Trypanosoma cruzi: a stage-specific calpain-like protein is induced after various kinds of stress.

    Science.gov (United States)

    Giese, Viviane; Dallagiovanna, Bruno; Marchini, Fabricio K; Pavoni, Daniela P; Krieger, Marco A; Goldenberg, Samuel

    2008-09-01

    Calpains are calcium-dependent cysteine proteinases found in all living organisms and are involved in diverse cellular processes. Calpain-like proteins have been reported after in silico analysis of the Tritryps genome and are believed to play important roles in cell functions of trypanosomatids. We describe the characterization of a member of this family, which is differentially expressed during the life-cycle of Trypanosoma cruzi.

  5. A Novel Trypanosoma cruzi Protein Associated to the Flagellar Pocket of Replicative Stages and Involved in Parasite Growth.

    Science.gov (United States)

    Durante, Ignacio M; Cámara, María de Los Milagros; Buscaglia, Carlos A

    2015-01-01

    The flagellar pocket constitutes an active and strategic site in the body of trypanosomatids (i.e. parasitic protozoa that cause important human and/or livestock diseases), which participates in several important processes such as cell polarity, morphogenesis and replication. Most importantly, the flagellar pocket is the unique site of surface protein export and nutrient uptake in trypanosomatids, and thus constitutes a key portal for the interaction with the host. In this work, we identified and characterized a novel Trypanosoma cruzi protein, termed TCLP 1, that accumulates at the flagellar pocket area of parasite replicative forms, as revealed by biochemical, immuno-cytochemistry and electron microscopy techniques. Different in silico analyses revealed that TCLP 1 is the founding member of a family of chimeric molecules restricted to trypanosomatids bearing, in addition to eukaryotic ubiquitin-like and protein-protein interacting domains, a motif displaying significant structural homology to bacterial multi-cargo chaperones involved in the secretion of virulence factors. Using the fidelity of an homologous expression system we confirmed TCLP 1 sub-cellular distribution and showed that TCLP 1-over-expressing parasites display impaired survival and accelerated progression to late stationary phase under starvation conditions. The reduced endocytic capacity of TCLP 1-over-expressors likely underlies (at least in part) this growth phenotype. TCLP 1 is involved in the uptake of extracellular macromolecules required for nutrition and hence in T. cruzi growth. Due to the bacterial origin, sub-cellular distribution and putative function(s), we propose TCLP 1 and related orthologs in trypanosomatids as appealing therapeutic targets for intervention against these health-threatening parasites.

  6. A Novel Trypanosoma cruzi Protein Associated to the Flagellar Pocket of Replicative Stages and Involved in Parasite Growth.

    Directory of Open Access Journals (Sweden)

    Ignacio M Durante

    Full Text Available The flagellar pocket constitutes an active and strategic site in the body of trypanosomatids (i.e. parasitic protozoa that cause important human and/or livestock diseases, which participates in several important processes such as cell polarity, morphogenesis and replication. Most importantly, the flagellar pocket is the unique site of surface protein export and nutrient uptake in trypanosomatids, and thus constitutes a key portal for the interaction with the host. In this work, we identified and characterized a novel Trypanosoma cruzi protein, termed TCLP 1, that accumulates at the flagellar pocket area of parasite replicative forms, as revealed by biochemical, immuno-cytochemistry and electron microscopy techniques. Different in silico analyses revealed that TCLP 1 is the founding member of a family of chimeric molecules restricted to trypanosomatids bearing, in addition to eukaryotic ubiquitin-like and protein-protein interacting domains, a motif displaying significant structural homology to bacterial multi-cargo chaperones involved in the secretion of virulence factors. Using the fidelity of an homologous expression system we confirmed TCLP 1 sub-cellular distribution and showed that TCLP 1-over-expressing parasites display impaired survival and accelerated progression to late stationary phase under starvation conditions. The reduced endocytic capacity of TCLP 1-over-expressors likely underlies (at least in part this growth phenotype. TCLP 1 is involved in the uptake of extracellular macromolecules required for nutrition and hence in T. cruzi growth. Due to the bacterial origin, sub-cellular distribution and putative function(s, we propose TCLP 1 and related orthologs in trypanosomatids as appealing therapeutic targets for intervention against these health-threatening parasites.

  7. Cytosolic peroxidases protect the lysosome of bloodstream African trypanosomes from iron-mediated membrane damage.

    Directory of Open Access Journals (Sweden)

    Corinna Hiller

    2014-04-01

    Full Text Available African trypanosomes express three virtually identical non-selenium glutathione peroxidase (Px-type enzymes which preferably detoxify lipid-derived hydroperoxides. As shown previously, bloodstream Trypanosoma brucei lacking the mitochondrial Px III display only a weak and transient proliferation defect whereas parasites that lack the cytosolic Px I and Px II undergo extremely fast lipid peroxidation and cell lysis. The phenotype can completely be rescued by supplementing the medium with the α-tocopherol derivative Trolox. The mechanism underlying the rapid cell death remained however elusive. Here we show that the lysosome is the origin of the cellular injury. Feeding the px I-II knockout parasites with Alexa Fluor-conjugated dextran or LysoTracker in the presence of Trolox yielded a discrete lysosomal staining. Yet upon withdrawal of the antioxidant, the signal became progressively spread over the whole cell body and was completely lost, respectively. T. brucei acquire iron by endocytosis of host transferrin. Supplementing the medium with iron or transferrin induced, whereas the iron chelator deferoxamine and apo-transferrin attenuated lysis of the px I-II knockout cells. Immunofluorescence microscopy with MitoTracker and antibodies against the lysosomal marker protein p67 revealed that disintegration of the lysosome precedes mitochondrial damage. In vivo experiments confirmed the negligible role of the mitochondrial peroxidase: Mice infected with px III knockout cells displayed only a slightly delayed disease development compared to wild-type parasites. Our data demonstrate that in bloodstream African trypanosomes, the lysosome, not the mitochondrion, is the primary site of oxidative damage and cytosolic trypanothione/tryparedoxin-dependent peroxidases protect the lysosome from iron-induced membrane peroxidation. This process appears to be closely linked to the high endocytic rate and distinct iron acquisition mechanisms of the infective

  8. In Vivo Antiprotozoal Activity of the Chloroform Extract from Carica papaya Seeds against Amastigote Stage of Trypanosoma cruzi during Indeterminate and Chronic Phase of Infection

    Directory of Open Access Journals (Sweden)

    Matilde Jimenez-Coello

    2014-01-01

    Full Text Available In order to evaluate the antiprotozoal activity of the chloroform extract of Carica papaya seeds during the subacute and chronic phase of infection of Trypanosoma cruzi, doses of 50 and 75 mg/kg were evaluated during the subacute phase, including a mixture of their main components (oleic, palmitic, and stearic acids. Subsequently, doses of 50 and 75 mg/kg in mice during the chronic phase of infection (100 dpi were also evaluated. It was found that chloroform extract was able to reduce the amastigote nests numbers during the subacute phase in 55.5 and 69.7% (P > 0.05 as well as in 56.45% in animals treated with the mixture of fatty acids. Moreover, the experimental groups treated with 50 and 75 mg/kg during the chronic phase of the infection showed a significant reduction of 46.8 and 53.13% respectively (P < 0.05. It is recommended to carry out more studies to determine if higher doses of chloroformic extract or its administration in combination with other antichagasic drugs allows a better response over the intracellular stage of T. cruzi in infected animal models and determine if the chloroform extract of C. papaya could be considered as an alternative for treatment during the indeterminate and chronic phase of the infection.

  9. In vitro investigation of Brazilian Cerrado plant extract activity against Plasmodium falciparum, Trypanosoma cruzi and T. brucei gambiense.

    Science.gov (United States)

    Charneau, Sébastien; de Mesquita, Mariana Laundry; Bastos, Izabela Marques Dourado; Santana, Jaime Martins; de Paula, José Elias; Grellier, Philippe; Espindola, Laila Salmen

    2016-06-01

    The threatened Brazilian Cerrado biome is an important biodiversity hotspot but still few explored that constitutes a potential reservoir of molecules to treat infectious diseases. We selected eight Cerrado plant species for screening against the erythrocytic stages of Plasmodium falciparum, human intracellular stages of Trypanosoma cruzi and bloodstream forms of T. brucei gambiense, and for their cytotoxicity upon the rat L6-myoblast cell line. Bioassays were performed with 37 hexane, ethyl acetate and ethanol extracts prepared from different plant organs. Activities against parasites were observed for 24 extracts: 9 with anti-P. falciparum, 4 with anti-T. cruzi and 11 with anti-T. brucei gambiense activities. High anti-protozoal activity (IC50 values Cerrado conservation and sustainable development.

  10. An Overview of Trypanosoma brucei Infections: An Intense Host–Parasite Interaction

    Science.gov (United States)

    Ponte-Sucre, Alicia

    2016-01-01

    Trypanosoma brucei rhodesiense and T. brucei gambiense, the causative agents of Human African Trypanosomiasis, are transmitted by tsetse flies. Within the vector, the parasite undergoes through transformations that prepares it to infect the human host. Sequentially these developmental stages are the replicative procyclic (in which the parasite surface is covered by procyclins) and trypo-epimastigote forms, as well as the non-replicative, infective, metacyclic form that develops in the vector salivary glands. As a pre-adaptation to their life in humans, metacyclic parasites begin to express and be densely covered by the Variant Surface Glycoprotein (VSG). Once the metacyclic form invades the human host the parasite develops into the bloodstream form. Herein the VSG triggers a humoral immune response. To avoid this humoral response, and essential for survival while in the bloodstream, the parasite changes its cover periodically and sheds into the surroundings the expressed VSG, thus evading the consequences of the immune system activation. Additionally, tools comparable to quorum sensing are used by the parasite for the successful parasite transmission from human to insect. On the other hand, the human host promotes clearance of the parasite triggering innate and adaptive immune responses and stimulating cytokine and chemokine secretion. All in all, the host–parasite interaction is extremely active and leads to responses that need multiple control sites to develop appropriately. PMID:28082973

  11. Identification of immunodominant epitopes in Trypanosoma cruzi trypomastigote surface antigen-1 protein that mask protective epitopes.

    Science.gov (United States)

    Wrightsman, R A; Dawson, B D; Fouts, D L; Manning, J E

    1994-10-01

    The gene that encodes trypomastigote surface Ag-1 (TSA-1), a major surface Ag of the bloodstream trypomastigote stage of Trypanosoma cruzi, was expressed in a baculovirus expression system. To determine the epitope(s) in TSA-1 that was recognized during T. cruzi infection and after immunization with TSA-1, subregions of the TSA-1 gene were expressed in a bacterial expression system. As seen by Western blotting, both mice and rabbits immunized with recombinant TSA-1 protein, as well as T. cruzi-infected mice, developed strong immune responses to the carboxyl-proximal region of TSA-1, but show no reaction to the amino-proximal portion of TSA-1. When mice were immunized with either recombinant TSA-1 protein or the carboxyl-proximal region of TSA-1, they did not survive challenge with 10(3) bloodstream trypomastigotes. However, 70% of the mice immunized with the amino-proximal portion of TSA-1 survived challenge with 10(3) bloodstream trypomastigotes. Thus, the immune responses elicited by recombinant TSA-1 or the carboxyl-proximal portion of TSA-1 are nonprotective during T. cruzi infection. In contrast, vaccination with the amino proximal region of TSA-1 elicits a protective immune response. These results suggest that responses to immunodominant epitope(s) within the carboxyl-proximal portion of TSA-1 mask epitopes within the amino-proximal portion that are capable of stimulating host-protective immune responses. It is suggested that immunodominant regions in surface molecules such as TSA-1 may provide a mechanism for the parasite to evade the host immune response by directing the response away from epitopes that have the potential to elicit a reaction that is damaging to the parasite.

  12. A core MRB1 complex component is indispensable for RNA editing in insect and human infective stages of Trypanosoma brucei.

    Directory of Open Access Journals (Sweden)

    Michelle L Ammerman

    Full Text Available Uridine insertion/deletion RNA editing is a unique and vital process in kinetoplastids, required for creation of translatable open reading frames in most mitochondrially-encoded RNAs. Emerging as a key player in this process is the mitochondrial RNA binding 1 (MRB1 complex. MRB1 comprises an RNA-independent core complex of at least six proteins, including the GAP1/2 guide RNA (gRNA binding proteins. The core interacts in an RNA-enhanced or -dependent manner with imprecisely defined TbRGG2 subcomplexes, Armadillo protein MRB10130, and additional factors that comprise the dynamic MRB1 complex. Towards understanding MRB1 complex function in RNA editing, we present here functional characterization of the pentein domain-containing MRB1 core protein, MRB11870. Inducible RNAi studies demonstrate that MRB11870 is essential for proliferation of both insect vector and human infective stage T. brucei. MRB11870 ablation causes a massive defect in RNA editing, affecting both pan-edited and minimally edited mRNAs, but does not substantially affect mitochondrial RNA stability or processing of precursor transcripts. The editing defect in MRB1-depleted cells occurs at the initiation stage of editing, as pre-edited mRNAs accumulate. However, the gRNAs that direct editing remain abundant in the knockdown cells. To examine the contribution of MRB11870 to MRB1 macromolecular interactions, we tagged core complexes and analyzed their composition and associated proteins in the presence and absence of MRB11870. These studies demonstrated that MRB11870 is essential for association of GAP1/2 with the core, as well as for interaction of the core with other proteins and subcomplexes. Together, these data support a model in which the MRB1 core mediates functional interaction of gRNAs with the editing machinery, having GAP1/2 as its gRNA binding constituents. MRB11870 is a critical component of the core, essential for its structure and function.

  13. Trypanosoma cruzi trans-sialidase in complex with a neutralizing antibody: structure/function studies towards the rational design of inhibitors.

    Directory of Open Access Journals (Sweden)

    Alejandro Buschiazzo

    2012-01-01

    Full Text Available Trans-sialidase (TS, a virulence factor from Trypanosoma cruzi, is an enzyme playing key roles in the biology of this protozoan parasite. Absent from the mammalian host, it constitutes a potential target for the development of novel chemotherapeutic drugs, an urgent need to combat Chagas' disease. TS is involved in host cell invasion and parasite survival in the bloodstream. However, TS is also actively shed by the parasite to the bloodstream, inducing systemic effects readily detected during the acute phase of the disease, in particular, hematological alterations and triggering of immune cells apoptosis, until specific neutralizing antibodies are elicited. These antibodies constitute the only known submicromolar inhibitor of TS's catalytic activity. We now report the identification and detailed characterization of a neutralizing mouse monoclonal antibody (mAb 13G9, recognizing T. cruzi TS with high specificity and subnanomolar affinity. This mAb displays undetectable association with the T. cruzi superfamily of TS-like proteins or yet with the TS-related enzymes from Trypanosoma brucei or Trypanosoma rangeli. In immunofluorescence assays, mAb 13G9 labeled 100% of the parasites from the infective trypomastigote stage. This mAb also reduces parasite invasion of cultured cells and strongly inhibits parasite surface sialylation. The crystal structure of the mAb 13G9 antigen-binding fragment in complex with the globular region of T. cruzi TS was determined, revealing detailed molecular insights of the inhibition mechanism. Not occluding the enzyme's catalytic site, the antibody performs a subtle action by inhibiting the movement of an assisting tyrosine (Y₁₁₉, whose mobility is known to play a key role in the trans-glycosidase mechanism. As an example of enzymatic inhibition involving non-catalytic residues that occupy sites distal from the substrate-binding pocket, this first near atomic characterization of a high affinity inhibitory molecule

  14. Lead optimization of a pyrazole sulfonamide series of Trypanosoma brucei N-myristoyltransferase inhibitors: identification and evaluation of CNS penetrant compounds as potential treatments for stage 2 human African trypanosomiasis.

    Science.gov (United States)

    Brand, Stephen; Norcross, Neil R; Thompson, Stephen; Harrison, Justin R; Smith, Victoria C; Robinson, David A; Torrie, Leah S; McElroy, Stuart P; Hallyburton, Irene; Norval, Suzanne; Scullion, Paul; Stojanovski, Laste; Simeons, Frederick R C; van Aalten, Daan; Frearson, Julie A; Brenk, Ruth; Fairlamb, Alan H; Ferguson, Michael A J; Wyatt, Paul G; Gilbert, Ian H; Read, Kevin D

    2014-12-11

    Trypanosoma brucei N-myristoyltransferase (TbNMT) is an attractive therapeutic target for the treatment of human African trypanosomiasis (HAT). From previous studies, we identified pyrazole sulfonamide, DDD85646 (1), a potent inhibitor of TbNMT. Although this compound represents an excellent lead, poor central nervous system (CNS) exposure restricts its use to the hemolymphatic form (stage 1) of the disease. With a clear clinical need for new drug treatments for HAT that address both the hemolymphatic and CNS stages of the disease, a chemistry campaign was initiated to address the shortfalls of this series. This paper describes modifications to the pyrazole sulfonamides which markedly improved blood-brain barrier permeability, achieved by reducing polar surface area and capping the sulfonamide. Moreover, replacing the core aromatic with a flexible linker significantly improved selectivity. This led to the discovery of DDD100097 (40) which demonstrated partial efficacy in a stage 2 (CNS) mouse model of HAT.

  15. Zinc finger nuclease technology: A stable tool for high efficiency transformation in bloodstream form T. brucei.

    Science.gov (United States)

    Schumann, Gabriela; Kangussu-Marcolino, Monica M; Doiron, Nicholas; Käser, Sandro; de Assis Burle-Caldas, Gabriela; DaRocha, Wanderson D; Teixeira, Santuza M; Roditi, Isabel

    2017-02-20

    In Trypanosoma brucei, the generation of knockout mutants is relatively easy compared to other organisms as transfection methods are well established. These methods have their limitations, however, when it comes to the generation of genome-wide libraries that require a minimum of several hundred thousand transformants. Double-strand breaks with the meganuclease ISce-I dramatically increase transformation efficiency, but are not widely in use as cell lines need to be generated de novo before each transfection. Here we show that zinc finger nucleases are a robust and stable tool that can enhance transformation in bloodstream forms by more than an order of magnitude.

  16. Stearoyl-CoA desaturase is an essential enzyme for the parasitic protist Trypanosoma brucei

    Energy Technology Data Exchange (ETDEWEB)

    Alloatti, Andres [Instituto de Biologia Molecular y Celular de Rosario (IBR), CONICET, Facultad de Ciencias Bioquimicas y Farmaceuticas, Universidad Nacional de Rosario, Santa Fe (Argentina); Gupta, Shreedhara; Gualdron-Lopez, Melisa; Nguewa, Paul A. [Research Unit for Tropical Diseases, de Duve Institute and Laboratory of Biochemistry, Universite Catholique de Louvain, Brussels (Belgium); Altabe, Silvia G. [Instituto de Biologia Molecular y Celular de Rosario (IBR), CONICET, Facultad de Ciencias Bioquimicas y Farmaceuticas, Universidad Nacional de Rosario, Santa Fe (Argentina); Deumer, Gladys; Wallemacq, Pierre [Department of Clinical Chemistry, Cliniques Universitaires Saint-Luc, LTAP, Universite Catholique de Louvain, Brussels (Belgium); Michels, Paul A.M. [Research Unit for Tropical Diseases, de Duve Institute and Laboratory of Biochemistry, Universite Catholique de Louvain, Brussels (Belgium); Uttaro, Antonio D., E-mail: toniuttaro@yahoo.com.ar [Instituto de Biologia Molecular y Celular de Rosario (IBR), CONICET, Facultad de Ciencias Bioquimicas y Farmaceuticas, Universidad Nacional de Rosario, Santa Fe (Argentina)

    2011-08-26

    Highlights: {yields} Inhibiting {Delta}9 desaturase drastically changes T. brucei's fatty-acid composition. {yields} Isoxyl specifically inhibits the {Delta}9 desaturase causing a growth arrest. {yields} RNA interference of desaturase expression causes a similar effect. {yields} Feeding T. brucei-infected mice with Isoxyl decreases the parasitemia. {yields} 70% of Isoxyl-treated mice survived the trypanosome infection. -- Abstract: Trypanosoma brucei, the etiologic agent of sleeping sickness, is exposed to important changes in nutrients and temperature during its life cycle. To adapt to these changes, the fluidity of its membranes plays a crucial role. This fluidity, mediated by the fatty-acid composition, is regulated by enzymes named desaturases. We have previously shown that the oleoyl desaturase is essential for Trypanosoma cruzi and T. brucei. In this work, we present experimental support for the relevance of stearoyl-CoA desaturase (SCD) for T. brucei's survival, in both its insect or procyclic-form (PCF) and bloodstream-form (BSF) stages. We evaluated this essentiality in two different ways: by generating a SCD knocked-down parasite line using RNA interference, and by chemical inhibition of the enzyme with two compounds, Isoxyl and a thiastearate with the sulfur atom at position 10 (10-TS). The effective concentration for 50% growth inhibition (EC{sub 50}) of PCF was 1.0 {+-} 0.2 {mu}M for Isoxyl and 5 {+-} 2 {mu}M for 10-TS, whereas BSF appeared more susceptible with EC{sub 50} values 0.10 {+-} 0.03 {mu}M (Isoxyl) and 1.0 {+-} 0.6 {mu}M (10-TS). RNA interference showed to be deleterious for both stages of the parasite. In addition, T. brucei-infected mice were fed with Isoxyl, causing a reduction of the parasitemia and an increase of the rodents' survival.

  17. Cloning of a major surface-antigen gene of Trypanosoma cruzi and identification of a nonapeptide repeat.

    Science.gov (United States)

    Peterson, D S; Wrightsman, R A; Manning, J E

    The parasitic protozoan Trypanosoma cruzi can establish infection in humans and other vertebrate hosts through direct penetration of host cells by trypomastigotes transmitted by the insect vector. Although the molecular processes involved in trypomastigote interiorization of vertebrate cells are unknown, several studies suggest that surface glycoproteins are involved. It is likely that the proteins involved are specific to the trypomastigote stage of the parasite, since only trypomastigotes found in both the insect vector and the vertebrate host bloodstream are capable of invading vertebrate cells. In contrast, the epimastigote stage, found exclusively in the vector, and the amastigote stage, an intracellular stage in the vertebrate host, cannot penetrate the cell directly. We have therefore concentrated our efforts on trypomastigote surface proteins and, along with others, have identified two trypomastigote-specific surface glycoproteins of relative molecular mass (Mr) 90,000 (90K) and 85,000 (85K). Antibody neutralization experiments indicate that the 85K glycoprotein is necessary for efficient interiorization of trypomastigotes in mammalian cells. Here we describe the molecular cloning of a genomic DNA fragment that encodes antigenic determinants present in the 85K trypomastigote surface antigen. The polypeptide fragment encoded by the cloned DNA is recognized by serum from a T. cruzi-infected host and is inferred by DNA sequence analysis to contain a nonapeptide unit that is tandemly repeated five times. Also, the messenger complementary to the cloned DNA fragment is present only in the trypomastigote stage of the parasite.

  18. Pure paraflagellar rod protein protects mice against Trypanosoma cruzi infection.

    Science.gov (United States)

    Wrightsman, R A; Miller, M J; Saborio, J L; Manning, J E

    1995-01-01

    The paraflagellar rod proteins (PAR) purified from Trypanosoma cruzi epimastigotes were shown to protect mice against an otherwise lethal challenge inoculum of 10(3) bloodstream-form trypomastigotes. The injection route used for immunization was shown to have a marked impact on the development of protective immunity. Mice receiving subcutaneous (s.c.) injections of PAR proteins had reduced bloodstream parasitemias and showed 100% survival following challenge. In contrast, mice immunized via the intraperitoneal (i.p.) route developed parasitemia levels equivalent to those of unimmunized controls and did not survive infection. Western blotting (immunoblotting) demonstrated that sera from both i.p. and s.c. immunized mice reacted specifically with PAR proteins; however, the antibody titer of the i.p. immunized mice was approximately 64-fold greater than that of the s.c. immunized mice, suggesting that the protective response in the s.c. immunized mice is cell mediated rather than humoral.

  19. Procyclic Trypanosoma brucei do not use Krebs cycle activity for energy generation.

    Science.gov (United States)

    van Weelden, Susanne W H; Fast, Beate; Vogt, Achim; van der Meer, Pieter; Saas, Joachim; van Hellemond, Jaap J; Tielens, Aloysius G M; Boshart, Michael

    2003-04-11

    The importance of a functional Krebs cycle for energy generation in the procyclic stage of Trypanosoma brucei was investigated under physiological conditions during logarithmic phase growth of a pleomorphic parasite strain. Wild type procyclic cells and mutants with targeted deletion of the gene coding for aconitase were derived by synchronous in vitro differentiation from wild type and mutant (Delta aco::NEO/Delta aco::HYG) bloodstream stage parasites, respectively, where aconitase is not expressed and is dispensable. No differences in intracellular levels of glycolytic and Krebs cycle intermediates were found in procyclic wild type and mutant cells, except for citrate that accumulated up to 90-fold in the mutants, confirming the absence of aconitase activity. Surprisingly, deletion of aconitase did not change differentiation nor the growth rate or the intracellular ATP/ADP ratio in those cells. Metabolic studies using radioactively labeled substrates and NMR analysis demonstrated that glucose and proline were not degraded via the Krebs cycle to CO(2). Instead, glucose was degraded to acetate, succinate, and alanine, whereas proline was degraded to succinate. Importantly, there was absolutely no difference in the metabolic products released by wild type and aconitase knockout parasites, and both were for survival strictly dependent on respiration via the mitochondrial electron transport chain. Hence, although the Krebs cycle enzymes are present, procyclic T. brucei do not use Krebs cycle activity for energy generation, but the mitochondrial respiratory chain is essential for survival and growth. We therefore propose a revised model of the energy metabolism of procyclic T. brucei.

  20. A transcription-independent epigenetic mechanism is associated with antigenic switching in Trypanosoma brucei

    OpenAIRE

    Aresta-Branco, Francisco; Pimenta, Silvia; Figueiredo, Luisa M.

    2015-01-01

    Antigenic variation in Trypanosoma brucei relies on periodic switching of variant surface glycoproteins (VSGs), which are transcribed monoallelically by RNA polymerase I from one of about 15 bloodstream expression sites (BES). Chromatin of the actively transcribed BES is depleted of nucleosomes, but it is unclear if this open conformation is a mere consequence of a high rate of transcription, or whether it is maintained by a transcription-independent mechanism. Using an inducible BES-silencin...

  1. Purification of extracellular and intracellular amastigotes of Trypanosoma cruzi from mammalian host-infected cells

    OpenAIRE

    sprotocols

    2015-01-01

    Authors: Alexandre Marques, Ernesto Nakayasu & Igor Almeida ### Abstract The protozoan parasite Trypanosoma cruzi is the etiologic agent of Chagas disease, which affects millions of people in Latin America. T. cruzi has a complex life cycle characterized by several developmental forms present in vertebrate and invertebrate hosts. In vertebrate mammalian hosts T. cruzi is found as intracellular amastigotes and bloodstream trypomastigotes. On the other hand, in the intestine of the ...

  2. Cloning and characterization of a gene for the stage-specific 82-kDa surface antigen of metacyclic trypomastigotes of Trypanosoma cruzi.

    Science.gov (United States)

    Araya, J E; Cano, M I; Yoshida, N; da Silveira, J F

    1994-05-01

    We have cloned and sequenced a cDNA clone coding for a metacyclic trypomastigote-specific surface glycoprotein with a molecular mass of 82 kDa (MTS-gp82). By immunoblotting and immunoprecipitation, antibodies against the recombinant protein recognized an 82-kDa protein of metacyclic trypomastigotes, without any detectable reaction towards amastigotes, epimastigotes or tissue culture-derived trypomastigotes. The insert of the MTS-gp82 cDNA clone strongly hybridized with a single 2.2-kb metacyclic trypomastigote mRNA, suggesting that the steady-state levels of mRNAs for MTS-gp82 are developmentally regulated. MTS-gp82 is encoded by a multigene family whose members are distributed in several chromosomes. Sequence analysis revealed 40-56% identity at amino acid level between MTS-gp82 and members of Trypanosoma cruzi gp85/sialidase family (TSA-1, Tt34c1, SA85-1.1). MTS-gp82 showed several amino acid motifs that are characteristic of gp85/sialidase family, such as the Asp box (SxDxGxTW), the subterminal (VTVxNVFLYNR) motif and the putative GPI-anchor sequence. On the basis of its structural features, the MTS-gp82 gene could be included in the T. cruzi gp85/sialidase family, but constituting a distinct group which is preferentially expressed in metacyclic trypomastigotes.

  3. Identification and characterization of an unusual class I myosin involved in vesicle traffic in Trypanosoma brucei.

    Directory of Open Access Journals (Sweden)

    Diana Spitznagel

    Full Text Available Myosins are a multimember family of motor proteins with diverse functions in eukaryotic cells. African trypanosomes possess only two candidate myosins and thus represent a useful system for functional analysis of these motors. One of these candidates is an unusual class I myosin (TbMyo1 that is expressed at similar levels but organized differently during the life cycle of Trypanosoma brucei. This myosin localizes to the polarized endocytic pathway in bloodstream forms of the parasite. This organization is actin dependent. Knock down of TbMyo1 results in a significant reduction in endocytic activity, a cessation in cell division and eventually cell death. A striking morphological feature in these cells is an enlargement of the flagellar pocket, which is consistent with an imbalance in traffic to and from the surface. In contrast TbMyo1 is distributed throughout procyclic forms of the tsetse vector and a loss of approximately 90% of the protein has no obvious effects on growth or morphology. These results reveal a life cycle stage specific requirement for this myosin in essential endocytic traffic and represent the first description of the involvement of a motor protein in vesicle traffic in these parasites.

  4. Identification and characterization of an unusual class I myosin involved in vesicle traffic in Trypanosoma brucei.

    Science.gov (United States)

    Spitznagel, Diana; O'Rourke, John F; Leddy, Neal; Hanrahan, Orla; Nolan, Derek P

    2010-01-01

    Myosins are a multimember family of motor proteins with diverse functions in eukaryotic cells. African trypanosomes possess only two candidate myosins and thus represent a useful system for functional analysis of these motors. One of these candidates is an unusual class I myosin (TbMyo1) that is expressed at similar levels but organized differently during the life cycle of Trypanosoma brucei. This myosin localizes to the polarized endocytic pathway in bloodstream forms of the parasite. This organization is actin dependent. Knock down of TbMyo1 results in a significant reduction in endocytic activity, a cessation in cell division and eventually cell death. A striking morphological feature in these cells is an enlargement of the flagellar pocket, which is consistent with an imbalance in traffic to and from the surface. In contrast TbMyo1 is distributed throughout procyclic forms of the tsetse vector and a loss of approximately 90% of the protein has no obvious effects on growth or morphology. These results reveal a life cycle stage specific requirement for this myosin in essential endocytic traffic and represent the first description of the involvement of a motor protein in vesicle traffic in these parasites.

  5. Trypanosoma congolense Infections: Induced Nitric Oxide Inhibits Parasite Growth In Vivo

    Directory of Open Access Journals (Sweden)

    Wenfa Lu

    2011-01-01

    Full Text Available Wild-type (WT C57BL/6 mice infected intraperitoneally with 5×106 Trypanosoma congolense survive for more than 30 days. C57BL/6 mice deficient in inducible nitric oxide synthase (iNOS−/− and infected with 103 or 5×106 parasites do not control the parasitemia and survive for only 14±7 or 6.8±0.1 days, respectively. Bloodstream trypanosomes of iNOS−/− mice infected with 5×106  T. congolense had a significantly higher ratio of organisms in the S+G2+M phases of the cell cycle than trypanosomes in WT mice. We have reported that IgM anti-VSG-mediated phagocytosis of T. congolense by macrophages inhibits nitric oxide (NO synthesis via CR3 (CD11b/CD18. Here, we show that during the first parasitemia, but not at later stages of infection, T. congolense-infected CD11b−/− mice produce more NO and have a significantly lower parasitemia than infected WT mice. We conclude that induced NO contributes to the control of parasitemia by inhibiting the growth of the trypanosomes.

  6. Taxonomy Icon Data: Trypanosoma brucei [Taxonomy Icon

    Lifescience Database Archive (English)

    Full Text Available Trypanosoma brucei Trypanosoma brucei Trypanosoma_brucei_L.png Trypanosoma_brucei_NL.png Trypanosoma_bruce...i_S.png Trypanosoma_brucei_NS.png http://biosciencedbc.jp/taxonomy_icon/icon.cgi?i=Trypanosoma+bruce...i&t=L http://biosciencedbc.jp/taxonomy_icon/icon.cgi?i=Trypanosoma+brucei&t=NL http://bioscie...ncedbc.jp/taxonomy_icon/icon.cgi?i=Trypanosoma+brucei&t=S http://biosciencedbc.jp.../taxonomy_icon/icon.cgi?i=Trypanosoma+brucei&t=NS http://togodb.biosciencedbc.jp/togodb/view/taxonomy_icon_comment_en?species_id=121 ...

  7. Global Gene Expression Profiling through the Complete Life Cycle of Trypanosoma vivax.

    Directory of Open Access Journals (Sweden)

    Andrew P Jackson

    Full Text Available The parasitic flagellate Trypanosoma vivax is a cause of animal trypanosomiasis across Africa and South America. The parasite has a digenetic life cycle, passing between mammalian hosts and insect vectors, and a series of developmental forms adapted to each life cycle stage. Each point in the life cycle presents radically different challenges to parasite metabolism and physiology and distinct host interactions requiring remodeling of the parasite cell surface. Transcriptomic and proteomic studies of the related parasites T. brucei and T. congolense have shown how gene expression is regulated during their development. New methods for in vitro culture of the T. vivax insect stages have allowed us to describe global gene expression throughout the complete T. vivax life cycle for the first time. We combined transcriptomic and proteomic analysis of each life stage using RNA-seq and mass spectrometry respectively, to identify genes with patterns of preferential transcription or expression. While T. vivax conforms to a pattern of highly conserved gene expression found in other African trypanosomes, (e.g. developmental regulation of energy metabolism, restricted expression of a dominant variant antigen, and expression of 'Fam50' proteins in the insect mouthparts, we identified significant differences in gene expression affecting metabolism in the fly and a suite of T. vivax-specific genes with predicted cell-surface expression that are preferentially expressed in the mammal ('Fam29, 30, 42' or the vector ('Fam34, 35, 43'. T. vivax differs significantly from other African trypanosomes in the developmentally-regulated proteins likely to be expressed on its cell surface and thus, in the structure of the host-parasite interface. These unique features may yet explain the species differences in life cycle and could, in the form of bloodstream-stage proteins that do not undergo antigenic variation, provide targets for therapy.

  8. Trypanosoma evansi is alike to Trypanosoma brucei brucei in the subcellular localisation of glycolytic enzymes

    Directory of Open Access Journals (Sweden)

    S Andrea Moreno

    2015-06-01

    Full Text Available Trypanosoma evansi, which causes surra, is descended from Trypanosoma brucei brucei, which causes nagana. Although both parasites are presumed to be metabolically similar, insufficient knowledge of T. evansi precludes a full comparison. Herein, we provide the first report on the subcellular localisation of the glycolytic enzymes in T. evansi, which is a alike to that of the bloodstream form (BSF of T. b. brucei: (i fructose-bisphosphate aldolase, glyceraldehyde-3-phosphate dehydrogenase (GAPDH, hexokinase, phosphofructokinase, glucose-6-phosphate isomerase, phosphoglycerate kinase, triosephosphate isomerase (glycolytic enzymes and glycerol-3-phosphate dehydrogenase (a glycolysis-auxiliary enzyme in glycosomes, (ii enolase, phosphoglycerate mutase, pyruvate kinase (glycolytic enzymes and a GAPDH isoenzyme in the cytosol, (iii malate dehydrogenase in cytosol and (iv glucose-6-phosphate dehydrogenase in both glycosomes and the cytosol. Specific enzymatic activities also suggest that T. evansi is alike to the BSF of T. b. brucei in glycolytic flux, which is much faster than the pentose phosphate pathway flux, and in the involvement of cytosolic GAPDH in the NAD+/NADH balance. These similarities were expected based on the close phylogenetic relationship of both parasites.

  9. Trypanosoma evansi is alike to Trypanosoma brucei brucei in the subcellular localisation of glycolytic enzymes.

    Science.gov (United States)

    Moreno, S Andrea; Nava, Mayerly

    2015-06-01

    Trypanosoma evansi, which causes surra, is descended from Trypanosoma brucei brucei, which causes nagana. Although both parasites are presumed to be metabolically similar, insufficient knowledge of T. evansi precludes a full comparison. Herein, we provide the first report on the subcellular localisation of the glycolytic enzymes in T. evansi, which is a alike to that of the bloodstream form (BSF) of T. b. brucei: (i) fructose-bisphosphate aldolase, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), hexokinase, phosphofructokinase, glucose-6-phosphate isomerase, phosphoglycerate kinase, triosephosphate isomerase (glycolytic enzymes) and glycerol-3-phosphate dehydrogenase (a glycolysis-auxiliary enzyme) in glycosomes, (ii) enolase, phosphoglycerate mutase, pyruvate kinase (glycolytic enzymes) and a GAPDH isoenzyme in the cytosol, (iii) malate dehydrogenase in cytosol and (iv) glucose-6-phosphate dehydrogenase in both glycosomes and the cytosol. Specific enzymatic activities also suggest that T. evansi is alike to the BSF of T. b. brucei in glycolytic flux, which is much faster than the pentose phosphate pathway flux, and in the involvement of cytosolic GAPDH in the NAD+/NADH balance. These similarities were expected based on the close phylogenetic relationship of both parasites.

  10. The phosphoarginine energy-buffering system of trypanosoma brucei involves multiple arginine kinase isoforms with different subcellular locations.

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    Frank Voncken

    Full Text Available Phosphagen energy-buffering systems play an essential role in regulating the cellular energy homeostasis in periods of high-energy demand or energy supply fluctuations. Here we describe the phosphoarginine/arginine kinase system of the kinetoplastid parasite Trypanosoma brucei, consisting of three highly similar arginine kinase isoforms (TbAK1-3. Immunofluorescence microscopy using myc-tagged protein versions revealed that each isoform is located in a specific subcellular compartment: TbAK1 is exclusively found in the flagellum, TbAK2 in the glycosome, and TbAK3 in the cytosol of T. brucei. The flagellar location of TbAK1 is dependent on a 22 amino acid long N-terminal sequence, which is sufficient for targeting a GFP-fusion protein to the trypanosome flagellum. The glycosomal location of TbAK2 is in agreement with the presence of a conserved peroxisomal targeting signal, the C-terminal tripeptide 'SNL'. TbAK3 lacks any apparent targeting sequences and is accordingly located in the cytosol of the parasite. Northern blot analysis indicated that each TbAK isoform is differentially expressed in bloodstream and procyclic forms of T. brucei, while the total cellular arginine kinase activity was 3-fold higher in bloodstream form trypanosomes. These results suggest a substantial change in the temporal and spatial energy requirements during parasite differentiation. Increased arginine kinase activity improved growth of procyclic form T. brucei during oxidative challenges with hydrogen peroxide. Elimination of the total cellular arginine kinase activity by RNA interference significantly decreased growth (>90% of procyclic form T. brucei under standard culture conditions and was lethal for this life cycle stage in the presence of hydrogen peroxide. The putative physiological roles of the different TbAK isoforms in T. brucei are further discussed.

  11. Proline Metabolism is Essential for Trypanosoma brucei brucei Survival in the Tsetse Vector

    Science.gov (United States)

    Mantilla, Brian S.; Dyer, Naomi A.; Biran, Marc; Bringaud, Frédéric; Lehane, Michael J.; Acosta-Serrano, Alvaro

    2017-01-01

    Adaptation to different nutritional environments is essential for life cycle completion by all Trypanosoma brucei sub-species. In the tsetse fly vector, L-proline is among the most abundant amino acids and is mainly used by the fly for lactation and to fuel flight muscle. The procyclic (insect) stage of T. b. brucei uses L-proline as its main carbon source, relying on an efficient catabolic pathway to convert it to glutamate, and then to succinate, acetate and alanine as the main secreted end products. Here we investigated the essentiality of an undisrupted proline catabolic pathway in T. b. brucei by studying mitochondrial Δ1-pyrroline-5-carboxylate dehydrogenase (TbP5CDH), which catalyzes the irreversible conversion of gamma-glutamate semialdehyde (γGS) into L-glutamate and NADH. In addition, we provided evidence for the absence of a functional proline biosynthetic pathway. TbP5CDH expression is developmentally regulated in the insect stages of the parasite, but absent in bloodstream forms grown in vitro. RNAi down-regulation of TbP5CDH severely affected the growth of procyclic trypanosomes in vitro in the absence of glucose, and altered the metabolic flux when proline was the sole carbon source. Furthermore, TbP5CDH knocked-down cells exhibited alterations in the mitochondrial inner membrane potential (ΔΨm), respiratory control ratio and ATP production. Also, changes in the proline-glutamate oxidative capacity slightly affected the surface expression of the major surface glycoprotein EP-procyclin. In the tsetse, TbP5CDH knocked-down cells were impaired and thus unable to colonize the fly’s midgut, probably due to the lack of glucose between bloodmeals. Altogether, our data show that the regulated expression of the proline metabolism pathway in T. b. brucei allows this parasite to adapt to the nutritional environment of the tsetse midgut. PMID:28114403

  12. Beta-interferon inhibits cell infection by Trypanosoma cruzi

    Science.gov (United States)

    Kierszenbaum, F.; Sonnenfeld, G.

    1984-01-01

    Beta interferon has been shown to inhibit the capacity of bloodstream forms of the flagellate Trypanosoma cruzi, the causative agent of Chagas' disease, to associate with and infect mouse peritoneal macrophages and rat heart myoblasts. The inhibitory effect was abrogated in the presence of specific antibodies to the interferon. Pretreatment of the parasites with interferon reduced their infectivity for untreated host cells, whereas pretreament of either type of host cell did not affect the interaction. The effect of interferon on the trypanosomes was reversible; the extent of the inhibitory effect was significantly reduced afer 20 min, and was undetectable after 60 min when macrophages were used as host cells. For the myoblasts, 60 min elapsed before the inhibitory effect began to subside and 120 min elapsed before it became insignificant or undetectable.

  13. Detection of Trypanosoma cruzi by Polymerase Chain Reaction.

    Science.gov (United States)

    Márquez, María Elizabeth; Concepción, Juan Luis; González-Marcano, Eglys; Mondolfi, Alberto Paniz

    2016-01-01

    American Trypanosomiasis (Chagas disease) is an infectious disease caused by the hemoflagellate parasite Trypanosoma cruzi which is transmitted by reduviid bugs. T. cruzi infection occurs in a broad spectrum of reservoir animals throughout North, Central, and South America and usually evolves into an asymptomatic chronic clinical stage of the disease in which diagnosis is often challenging. This chapter describes the application of polymerase chain reaction (PCR) for the detection of Trypanosoma cruzi DNA including protocols for sample preparation, DNA extraction, and target amplification methods.

  14. Inhibition of Trypanosoma cruzi growth in vitro by Solanum alkaloids: a comparison with ketoconazole.

    Science.gov (United States)

    Chataing, B; Concepción, J L; Lobatón, R; Usubillaga, A

    1998-02-01

    The glycoalkaloids alpha-chaconine, alpha-solamargine, alpha-solanine, solasonine, sycophantine, and tomatine, as well as the aglycones demissidine, solanidine, solanocapsine, solasodine, tomatidine, and veratrine were tested as growth inhibitors of Trypanosoma cruzi, strain EP, in LIT medium. Their activity was compared with the antifungal ketoconazole. Glycoalkaloids containing alpha-chacotriose showed trypanolytic activity against the epimastigote form and trypanocidal activity against the bloodstream and metacyclic trypomastigote form of Trypanosoma cruzi in culture medium in micromolar concentrations. Ketoconazole showed a lower activity, at the same concentrations of alpha-chaconine and alpha-solamargine. The observations indicate that the initial target of the compound is at the membrane level with a concomitant change in the parasite morphology. Moreover, internal compartments of the parasites were observed to be affected by the drugs, revealing the dissolution of some organelles as mitocondrias and glycosomes.

  15. Computer algorithms to detect bloodstream infections.

    Science.gov (United States)

    Trick, William E; Zagorski, Brandon M; Tokars, Jerome I; Vernon, Michael O; Welbel, Sharon F; Wisniewski, Mary F; Richards, Chesley; Weinstein, Robert A

    2004-09-01

    We compared manual and computer-assisted bloodstream infection surveillance for adult inpatients at two hospitals. We identified hospital-acquired, primary, central-venous catheter (CVC)-associated bloodstream infections by using five methods: retrospective, manual record review by investigators; prospective, manual review by infection control professionals; positive blood culture plus manual CVC determination; computer algorithms; and computer algorithms and manual CVC determination. We calculated sensitivity, specificity, predictive values, plus the kappa statistic (kappa) between investigator review and other methods, and we correlated infection rates for seven units. The kappa value was 0.37 for infection control review, 0.48 for positive blood culture plus manual CVC determination, 0.49 for computer algorithm, and 0.73 for computer algorithm plus manual CVC determination. Unit-specific infection rates, per 1,000 patient days, were 1.0-12.5 by investigator review and 1.4-10.2 by computer algorithm (correlation r = 0.91, p = 0.004). Automated bloodstream infection surveillance with electronic data is an accurate alternative to surveillance with manually collected data.

  16. Complete in vitro life cycle of Trypanosoma congolense: development of genetic tools.

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    Virginie Coustou

    Full Text Available BACKGROUND: Animal African trypanosomosis, a disease mainly caused by the protozoan parasite Trypanosoma congolense, is a major constraint to livestock productivity and has a significant impact in the developing countries of Africa. RNA interference (RNAi has been used to study gene function and identify drug and vaccine targets in a variety of organisms including trypanosomes. However, trypanosome RNAi studies have mainly been conducted in T. brucei, as a model for human infection, largely ignoring livestock parasites of economical importance such as T. congolense, which displays different pathogenesis profiles. The whole T. congolense life cycle can be completed in vitro, but this attractive model displayed important limitations: (i genetic tools were currently limited to insect forms and production of modified infectious BSF through differentiation was never achieved, (ii in vitro differentiation techniques lasted several months, (iii absence of long-term bloodstream forms (BSF in vitro culture prevented genomic analyses. METHODOLOGY/PRINCIPAL FINDINGS: We optimized culture conditions for each developmental stage and secured the differentiation steps. Specifically, we devised a medium adapted for the strenuous development of stable long-term BSF culture. Using Amaxa nucleofection technology, we greatly improved the transfection rate of the insect form and designed an inducible transgene expression system using the IL3000 reference strain. We tested it by expression of reporter genes and through RNAi. Subsequently, we achieved the complete in vitro life cycle with dramatically shortened time requirements for various wild type and transgenic strains. Finally, we established the use of modified strains for experimental infections and underlined a host adaptation phase requirement. CONCLUSIONS/SIGNIFICANCE: We devised an improved T. congolense model, which offers the opportunity to perform functional genomics analyses throughout the whole life

  17. Anti-Trypanosoma cruzi activity of green tea (Camellia sinensis) catechins.

    Science.gov (United States)

    Paveto, Cristina; Güida, María C; Esteva, Mónica I; Martino, Virginia; Coussio, Jorge; Flawiá, Mirtha M; Torres, Héctor N

    2004-01-01

    The trypanocidal action of green tea catechins against two different developmental stages of Trypanosoma cruzi is reported for the first time. This activity was assayed with the nonproliferative bloodstream trypomastigote and with the intracellular replicative amastigote parasite forms. An ethyl acetate fraction from Camellia sinensis green tea leaves, which contains most of the polyphenolic compounds and the maximal trypanocidal activity, was obtained by fractionation of the aqueous extract with organic solvents. The active compounds present in this extract were further purified by LH-20 column chromatography and were identified by high-performance liquid chromatography analysis with a photo diode array detector and gas chromatography coupled to mass spectroscopy. The following flavan-3-ols derivatives, known as catechins, were identified: catechin, epicatechin, gallocatechin, epigallocatechin, catechin gallate, epicatechin gallate, gallocatechin gallate, and epigallocatechin gallate. The purified compounds lysed more than 50% of the parasites present in the blood of infected BALB/c mice at concentrations as low as 0.12 to 85 pM. The most active compounds were gallocatechin gallate and epigallocatechin gallate, with minimal bactericidal concentrations that inhibited 50% of isolates tested of 0.12 and 0.53 pM, respectively. The number of amastigotes in infected Vero cells decreased by 50% in the presence of each of these compounds at 100 nM. The effects of the catechins on the recombinant T. cruzi arginine kinase, a key enzyme in the energy metabolism of the parasite, were assayed. The activity of this enzyme was inhibited by about 50% by nanomolar concentrations of catechin gallate or gallocatechin gallate, whereas the other members of the group were less effective. On the basis of these results, we suggest that these compounds could be used to sterilize blood and, eventually, as therapeutic agents for Chagas' disease.

  18. The orthologue of Sjogren's syndrome nuclear autoantigen 1 (SSNA1 in Trypanosoma brucei is an immunogenic self-assembling molecule.

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    Helen P Price

    Full Text Available Primary Sjögren's Syndrome (PSS is a highly prevalent autoimmune disease, typically manifesting as lymphocytic infiltration of the exocrine glands leading to chronically impaired lacrimal and salivary secretion. Sjögren's Syndrome nuclear autoantigen 1 (SSNA1 or NA14 is a major specific target for autoantibodies in PSS but the precise function and clinical relevance of this protein are largely unknown. Orthologues of the gene are absent from many of the commonly used model organisms but are present in Chlamyodomonas reinhardtii (in which it has been termed DIP13 and most protozoa. We report the functional characterisation of the orthologue of SSNA1 in the kinetoplastid parasite, Trypanosoma brucei. Both TbDIP13 and human SSNA1 are small coiled-coil proteins which are predicted to be remote homologues of the actin-binding protein tropomyosin. We use comparative proteomic methods to identify potential interacting partners of TbDIP13. We also show evidence that TbDIP13 is able to self-assemble into fibril-like structures both in vitro and in vivo, a property which may contribute to its immunogenicity. Endogenous TbDIP13 partially co-localises with acetylated α-tubulin in the insect procyclic stage of the parasite. However, deletion of the DIP13 gene in cultured bloodstream and procyclic stages of T. brucei has little effect on parasite growth or morphology, indicating either a degree of functional redundancy or a function in an alternative stage of the parasite life cycle.

  19. Biosynthesis of very long chain fatty acids in Trypanosoma cruzi.

    Science.gov (United States)

    Livore, Verónica I; Uttaro, Antonio D

    2015-01-01

    Trypanosoma brucei and Trypanosoma cruzi showed similar fatty acid (FA) compositions, having a high proportion of unsaturated FAs, mainly 18:2Δ9,12 (23-39%) and 18:1Δ9 (11-17%). C22 polyunsaturated FAs are in significant amounts only in T. brucei (12-20%) but represent a mere 2% of total FAs in T. cruzi. Both species have also similar profiles of medium- and long-chain saturated FAs, from 14:0 to 20:0. Interestingly, procyclic and bloodstream forms of T. brucei lack very long chain FAs (VLCFAs), whereas epimastigotes and trypomastigotes of T. cruzi contain 22:0 (0.1-0.2%), 24:0 (1.5-2%), and 26:0 (0.1-0.2%). This is in agreement with the presence of an additional FA elongase gene (TcELO4) in T. cruzi. TcELO4 was expressed in a Saccharomyces cerevisiae mutant lacking the endogenous ScELO3, rescuing the synthesis of saturated and hydroxylated C26 FAs in the yeast. Expression of TcELO4 also rescued the synthetic lethality of a ScELO2, ScELO3 double mutation, and the VLCFA profile of the transformed yeast was similar to that found in T. cruzi. By identifying TcELO4 as the enzyme responsible for the elongation of FA from 16:0 and 18:0 up to 26:0, with 24:0 being the preferred product, this work completed the characterization of FA elongases in Trypanosoma spp.

  20. Profiling the anti-protozoal activity of anti-cancer HDAC inhibitors against Plasmodium and Trypanosoma parasites

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    Jessica A. Engel

    2015-12-01

    Full Text Available Histone deacetylase (HDAC enzymes work together with histone acetyltransferases (HATs to reversibly acetylate both histone and non-histone proteins. As a result, these enzymes are involved in regulating chromatin structure and gene expression as well as other important cellular processes. HDACs are validated drug targets for some types of cancer, with four HDAC inhibitors clinically approved. However, they are also showing promise as novel drug targets for other indications, including malaria and other parasitic diseases. In this study the in vitro activity of four anti-cancer HDAC inhibitors was examined against parasites that cause malaria and trypanosomiasis. Three of these inhibitors, suberoylanilide hydroxamic acid (SAHA; vorinostat®, romidepsin (Istodax® and belinostat (Beleodaq®, are clinically approved for the treatment of T-cell lymphoma, while the fourth, panobinostat, has recently been approved for combination therapy use in certain patients with multiple myeloma. All HDAC inhibitors were found to inhibit the growth of asexual-stage Plasmodium falciparum malaria parasites in the nanomolar range (IC50 10–200 nM, while only romidepsin was active at sub-μM concentrations against bloodstream form Trypanosoma brucei brucei parasites (IC50 35 nM. The compounds were found to have some selectivity for malaria parasites compared with mammalian cells, but were not selective for trypanosome parasites versus mammalian cells. All compounds caused hyperacetylation of histone and non-histone proteins in P. falciparum asexual stage parasites and inhibited deacetylase activity in P. falciparum nuclear extracts in addition to recombinant PfHDAC1 activity. P. falciparum histone hyperacetylation data indicate that HDAC inhibitors may differentially affect the acetylation profiles of histone H3 and H4.

  1. Profiling the anti-protozoal activity of anti-cancer HDAC inhibitors against Plasmodium and Trypanosoma parasites.

    Science.gov (United States)

    Engel, Jessica A; Jones, Amy J; Avery, Vicky M; Sumanadasa, Subathdrage D M; Ng, Susanna S; Fairlie, David P; Adams, Tina S; Andrews, Katherine T

    2015-12-01

    Histone deacetylase (HDAC) enzymes work together with histone acetyltransferases (HATs) to reversibly acetylate both histone and non-histone proteins. As a result, these enzymes are involved in regulating chromatin structure and gene expression as well as other important cellular processes. HDACs are validated drug targets for some types of cancer, with four HDAC inhibitors clinically approved. However, they are also showing promise as novel drug targets for other indications, including malaria and other parasitic diseases. In this study the in vitro activity of four anti-cancer HDAC inhibitors was examined against parasites that cause malaria and trypanosomiasis. Three of these inhibitors, suberoylanilide hydroxamic acid (SAHA; vorinostat(®)), romidepsin (Istodax(®)) and belinostat (Beleodaq(®)), are clinically approved for the treatment of T-cell lymphoma, while the fourth, panobinostat, has recently been approved for combination therapy use in certain patients with multiple myeloma. All HDAC inhibitors were found to inhibit the growth of asexual-stage Plasmodium falciparum malaria parasites in the nanomolar range (IC50 10-200 nM), while only romidepsin was active at sub-μM concentrations against bloodstream form Trypanosoma brucei brucei parasites (IC50 35 nM). The compounds were found to have some selectivity for malaria parasites compared with mammalian cells, but were not selective for trypanosome parasites versus mammalian cells. All compounds caused hyperacetylation of histone and non-histone proteins in P. falciparum asexual stage parasites and inhibited deacetylase activity in P. falciparum nuclear extracts in addition to recombinant PfHDAC1 activity. P. falciparum histone hyperacetylation data indicate that HDAC inhibitors may differentially affect the acetylation profiles of histone H3 and H4.

  2. Bloodstream infections in HIV-infected patients.

    Science.gov (United States)

    Taramasso, Lucia; Tatarelli, Paola; Di Biagio, Antonio

    2016-04-02

    In the combined antiretroviral therapy era, HIV-infected patients remain a vulnerable population for the onset of bloodstream infections (BSI). Worldwide, nontyphoid salmonellae, Streptococcus pneumoniae, Escherichia coli, Staphylococcus aureus and coagulase negative staphylococci are the most important pathogens. Intravenous catheter associated infection, skin-soft tissue infection and endocarditis are associated with Gram-positive bacteremia. Among the Gram-negative, nontyphoidal Salmonella have been previously correlated to sepsis. Other causes of BSI in HIV-infected patients are mycobacteria and fungi. Mycobacteria constitute a major cause of BSI in limited resource countries. Fungal BSI are not frequent and among them Cryptococcus neoformans is the most common life-threatening infection. The degree of immunosuppression remains the key prognostic factor leading to the development of BSI.

  3. Bloodstream Infections with Mycobacterium tuberculosis among HIV patients

    Centers for Disease Control (CDC) Podcasts

    2010-09-23

    This podcast looks at bloodstream infections with Mycobacterium tuberculosis and other pathogens among outpatients infected with HIV in Southeast Asia. CDC health scientist Kimberly McCarthy discusses the study and why bloodstream infections occur in HIV-infected populations.  Created: 9/23/2010 by National Center for Emerging and Zoonotic Infectious Diseases (NCEZID).   Date Released: 9/23/2010.

  4. An Atypical Mitochondrial Carrier That Mediates Drug Action in Trypanosoma brucei.

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    Juan P de Macêdo

    2015-05-01

    Full Text Available Elucidating the mechanism of action of trypanocidal compounds is an important step in the development of more efficient drugs against Trypanosoma brucei. In a screening approach using an RNAi library in T. brucei bloodstream forms, we identified a member of the mitochondrial carrier family, TbMCP14, as a prime candidate mediating the action of a group of anti-parasitic choline analogs. Depletion of TbMCP14 by inducible RNAi in both bloodstream and procyclic forms increased resistance of parasites towards the compounds by 7-fold and 3-fold, respectively, compared to uninduced cells. In addition, down-regulation of TbMCP14 protected bloodstream form mitochondria from a drug-induced decrease in mitochondrial membrane potential. Conversely, over-expression of the carrier in procyclic forms increased parasite susceptibility more than 13-fold. Metabolomic analyses of parasites over-expressing TbMCP14 showed increased levels of the proline metabolite, pyrroline-5-carboxylate, suggesting a possible involvement of TbMCP14 in energy production. The generation of TbMCP14 knock-out parasites showed that the carrier is not essential for survival of T. brucei bloodstream forms, but reduced parasite proliferation under standard culture conditions. In contrast, depletion of TbMCP14 in procyclic forms resulted in growth arrest, followed by parasite death. The time point at which parasite proliferation stopped was dependent on the major energy source, i.e. glucose versus proline, in the culture medium. Together with our findings that proline-dependent ATP production in crude mitochondria from TbMCP14-depleted trypanosomes was reduced compared to control mitochondria, the study demonstrates that TbMCP14 is involved in energy production in T. brucei. Since TbMCP14 belongs to a trypanosomatid-specific clade of mitochondrial carrier family proteins showing very poor similarity to mitochondrial carriers of mammals, it may represent an interesting target for drug

  5. When Prostate Cancer Circulates in the Bloodstream

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    Virginie Vlaeminck-Guillem

    2015-10-01

    Full Text Available Management of patients with prostate cancer is currently based on imperfect clinical, biological, radiological and pathological evaluation. Prostate cancer aggressiveness, including metastatic potential, remains difficult to accurately estimate. In an attempt to better adapt therapeutics to an individual (personalized medicine, reliable evaluation of the intrinsic molecular biology of the tumor is warranted, and particularly for all tumor sites (primary tumors and secondary sites at any time of the disease progression. As a consequence of their natural tendency to grow (passive invasion or as a consequence of an active blood vessel invasion by metastase-initiating cells, tumors shed various materials into the bloodstream. Major efforts have been recently made to develop powerful and accurate methods able to detect, quantify and/or analyze all these circulating tumor materials: circulating tumors cells, disseminating tumor cells, extracellular vesicles (including exosomes, nucleic acids, etc. The aim of this review is to summarize current knowledge about these circulating tumor materials and their applications in translational research.

  6. Functional characterization of methionine sulfoxide reductase A from Trypanosoma spp.

    Science.gov (United States)

    Arias, Diego G; Cabeza, Matías S; Erben, Esteban D; Carranza, Pedro G; Lujan, Hugo D; Téllez Iñón, María T; Iglesias, Alberto A; Guerrero, Sergio A

    2011-01-01

    Methionine is an amino acid susceptible to being oxidized to methionine sulfoxide (MetSO). The reduction of MetSO to methionine is catalyzed by methionine sulfoxide reductase (MSR), an enzyme present in almost all organisms. In trypanosomatids, the study of antioxidant systems has been mainly focused on the involvement of trypanothione, a specific redox component in these organisms. However, no information is available concerning their mechanisms for repairing oxidized proteins, which would be relevant for the survival of these pathogens in the various stages of their life cycle. We report the molecular cloning of three genes encoding a putative A-type MSR in trypanosomatids. The genes were expressed in Escherichia coli, and the corresponding recombinant proteins were purified and functionally characterized. The enzymes were specific for L-Met(S)SO reduction, using Trypanosoma cruzi tryparedoxin I as the reducing substrate. Each enzyme migrated in electrophoresis with a particular profile reflecting the differences they exhibit in superficial charge. The in vivo presence of the enzymes was evidenced by immunological detection in replicative stages of T. cruzi and Trypanosoma brucei. The results support the occurrence of a metabolic pathway in Trypanosoma spp. involved in the critical function of repairing oxidized macromolecules.

  7. Discovery and verification of osteopontin and Beta-2-microglobulin as promising markers for staging human African trypanosomiasis.

    Science.gov (United States)

    Tiberti, Natalia; Hainard, Alexandre; Lejon, Veerle; Robin, Xavier; Ngoyi, Dieudonné Mumba; Turck, Natacha; Matovu, Enock; Enyaru, John; Ndung'u, Joseph Mathu; Scherl, Alexander; Dayon, Loïc; Sanchez, Jean-Charles

    2010-12-01

    Human African trypanosomiasis, or sleeping sickness, is a parasitic disease endemic in sub-Saharan Africa, transmitted to humans through the bite of a tsetse fly. The first or hemolymphatic stage of the disease is associated with presence of parasites in the bloodstream, lymphatic system, and body tissues. If patients are left untreated, parasites cross the blood-brain barrier and invade the cerebrospinal fluid and the brain parenchyma, giving rise to the second or meningoencephalitic stage. Stage determination is a crucial step in guiding the choice of treatment, as drugs used for S2 are potentially dangerous. Current staging methods, based on counting white blood cells and demonstrating trypanosomes in cerebrospinal fluid, lack specificity and/or sensitivity. In the present study, we used several proteomic strategies to discover new markers with potential for staging human African trypanosomiasis. Cerebrospinal fluid (CSF) samples were collected from patients infected with Trypanosoma brucei gambiense in the Democratic Republic of Congo. The stage was determined following the guidelines of the national control program. The proteome of the samples was analyzed by two-dimensional gel electrophoresis (n = 9), and by sixplex tandem mass tag (TMT) isobaric labeling (n = 6) quantitative mass spectrometry. Overall, 73 proteins were overexpressed in patients presenting the second stage of the disease. Two of these, osteopontin and β-2-microglobulin, were confirmed to be potential markers for staging human African trypanosomiasis (HAT) by Western blot and ELISA. The two proteins significantly discriminated between S1 and S2 patients with high sensitivity (68% and 78%, respectively) for 100% specificity, and a combination of both improved the sensitivity to 91%. The levels of osteopontin and β-2-microglobulin in CSF of S2 patients (μg/ml range), as well as the fold increased concentration in S2 compared with S1 (3.8 and 5.5 respectively) make the two markers good

  8. Single molecule analysis of Trypanosoma brucei DNA replication dynamics.

    Science.gov (United States)

    Calderano, Simone Guedes; Drosopoulos, William C; Quaresma, Marina Mônaco; Marques, Catarina A; Kosiyatrakul, Settapong; McCulloch, Richard; Schildkraut, Carl L; Elias, Maria Carolina

    2015-03-11

    Eukaryotic genome duplication relies on origins of replication, distributed over multiple chromosomes, to initiate DNA replication. A recent genome-wide analysis of Trypanosoma brucei, the etiological agent of sleeping sickness, localized its replication origins to the boundaries of multigenic transcription units. To better understand genomic replication in this organism, we examined replication by single molecule analysis of replicated DNA. We determined the average speed of replication forks of procyclic and bloodstream form cells and we found that T. brucei DNA replication rate is similar to rates seen in other eukaryotes. We also analyzed the replication dynamics of a central region of chromosome 1 in procyclic forms. We present evidence for replication terminating within the central part of the chromosome and thus emanating from both sides, suggesting a previously unmapped origin toward the 5' extremity of chromosome 1. Also, termination is not at a fixed location in chromosome 1, but is rather variable. Importantly, we found a replication origin located near an ORC1/CDC6 binding site that is detected after replicative stress induced by hydroxyurea treatment, suggesting it may be a dormant origin activated in response to replicative stress. Collectively, our findings support the existence of more replication origins in T. brucei than previously appreciated.

  9. Telomeric expression sites are highly conserved in Trypanosoma brucei.

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    Christiane Hertz-Fowler

    Full Text Available Subtelomeric regions are often under-represented in genome sequences of eukaryotes. One of the best known examples of the use of telomere proximity for adaptive purposes are the bloodstream expression sites (BESs of the African trypanosome Trypanosoma brucei. To enhance our understanding of BES structure and function in host adaptation and immune evasion, the BES repertoire from the Lister 427 strain of T. brucei were independently tagged and sequenced. BESs are polymorphic in size and structure but reveal a surprisingly conserved architecture in the context of extensive recombination. Very small BESs do exist and many functioning BESs do not contain the full complement of expression site associated genes (ESAGs. The consequences of duplicated or missing ESAGs, including ESAG9, a newly named ESAG12, and additional variant surface glycoprotein genes (VSGs were evaluated by functional assays after BESs were tagged with a drug-resistance gene. Phylogenetic analysis of constituent ESAG families suggests that BESs are sequence mosaics and that extensive recombination has shaped the evolution of the BES repertoire. This work opens important perspectives in understanding the molecular mechanisms of antigenic variation, a widely used strategy for immune evasion in pathogens, and telomere biology.

  10. Synanthropic rodent reservoirs of Trypanosoma (Schizotrypanum cruzi in the valley of Caracas, Venezuela

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    HERRERA Leidi

    1997-01-01

    Full Text Available Direct blood examination and xenodiagnosis of 47 synanthropic rodents (Rattus rattus, R. norvegicus, Mus musculus captured in the valley of Caracas, Venezuela, revealed trypanosomal infections in 12 R. rattus, 10 with T. lewisi and 2 with T. cruzi. Of the latter the course of parasitemia, the pleomorphism of the bloodstream trypomastigotes, tissue tropism in naturally and experimentally infected rats and mice, host mortality, morphology of fecal parasites in Rhodnius prolixus used for xenodiagnosis, and infectivity of the bug feces for NMRI mice, were all characteristic of Trypanosoma (Schizotrypanum cruzi. One rat, with a patent parasitemia, had numerous nests of amastigotes in cardiac muscle and moderate parasitism of the smooth muscle of the duodenum and of skeletal muscle. Mice inoculated with fecal flagellates from the bugs had moderate tissue tropism in the same organs and also in the colon and pancreas. The possible role of R. rattus in the establishment of foci of Chagas? disease in Caracas is discussed

  11. Contribution of Glucose Transport to the Control of the Glycolytic Flux in Trypanosoma brucei

    Science.gov (United States)

    Bakker, Barbara M.; Walsh, Michael C.; Ter Kuile, Benno H.; Mensonides, Femke I. C.; Michels, Paul A. M.; Opperdoes, Fred R.; Westerhoff, Hans V.

    1999-08-01

    The rate of glucose transport across the plasma membrane of the bloodstream form of Trypanosoma brucei was modulated by titration of the hexose transporter with the inhibitor phloretin, and the effect on the glycolytic flux was measured. A rapid glucose uptake assay was developed to measure the transport activity independently of the glycolytic flux. Phloretin proved a competitive inhibitor. When the effect of the intracellular glucose concentration on the inhibition was taken into account, the flux control coefficient of the glucose transporter was between 0.3 and 0.5 at 5 mM glucose. Because the flux control coefficients of all steps in a metabolic pathway sum to 1, this result proves that glucose transport is not the rate-limiting step of trypanosome glycolysis. Under physiological conditions, transport shares the control with other steps. At glucose concentrations much lower than physiological, the glucose carrier assumed all control, in close agreement with model predictions.

  12. Genotypic analysis of Acinetobacter bloodstream infection isolates in a Turkish university hospital.

    NARCIS (Netherlands)

    Alp, E.; Esel, D.; Yildiz, O.; Voss, A.; Melchers, W.J.G.; Doganay, M.

    2006-01-01

    Acinetobacter baumannii is a significant pathogen of bloodstream infections in hospital patients that frequently causes single clone outbreaks. We aimed to evaluate the genetic relatedness and antimicrobial susceptibility of Acinetobacter spp. bloodstream isolates, in order to obtain insight into th

  13. A rare case of human trypanosomiasis caused by Trypanosoma evansi

    Directory of Open Access Journals (Sweden)

    Powar R

    2006-01-01

    Full Text Available Human trypanosoma infections like the ones seen in Africa and South America are unknown in India. The only exception in literature is of two documented cases of a self-limiting febrile illness, being attributed to Trypanosoma lewisi like parasites. We are reporting an unusual case of trypanosomiasis from the rural parts of Chandrapur district in Maharashtra. An adult male farmhand who used to practice veterinary medicine also, presented with history of febrile episodes on and off since five months and drowsiness before admission to this Institute. Though routine blood and other investigations were within normal limits, the peripheral smear showed a large number of trypanosomes which morphologically resembled the species Trypanosoma evansi , the aetiological agent of surra - a form of animal trypanosomiasis. A battery of assays covering the spectrum of parasitology, serology, and molecular biology confirmed the infecting parasite to be T. evansi . Failure to demonstrate the central nervous system (CNS involvement, as evidenced by the absence of parasite in cerebrospinal fluid (CSF advocated the use of suramin - the drug of choice in early stage African trypanosomiasis without any CNS involvement. Suramin achieved cure in our patient. The case is being reported because of its unique nature as the patient was not immunocompromised and showed infestation with a parasite which normally does not affect human beings.

  14. Procyclic Trypanosoma brucei do not use Krebs cycle activity for energy generation

    NARCIS (Netherlands)

    Weelden, van S.W.H.; Fast, B.; Vogt, A.; Meer, van der P.; Saas, J.; Hellemond, van J.J.; Tielens, A.G.M.; Boshart, M.

    2003-01-01

    The importance of a functional Krebs cycle for energy generation in the procyclic stage of Trypanosoma brucei was investigated under physiological conditions during logarithmic phase growth of a pleomorphic parasite strain. Wild type procyclic cells and mutants with targeted deletion of the gene cod

  15. Trypanosoma spp. in Swedish game animals.

    Science.gov (United States)

    Neumüller, Magnus; Nilsson, Kenneth; Påhlson, Carl

    2012-01-01

    Serum and blood samples from 36 game animals, shot during the hunting seasons 2007-2009, were collected and analyzed for the presence of Trypanosoma spp. by three methods: isolation, polymerase chain reaction (PCR), and serology. Only fissiped animals were included, four different ruminants and wild boar. Trypanosomes could be isolated from two of the animals, and eight had detectable parasite DNA. Seven animals had high titers of anti-trypanosoma IgG antibodies. The two isolated strains, one from roe dear and one from European elk, were determined to Trypanosoma theileri by partial DNA sequencing of the 18S ribosomal gene. In the seven boars, no Trypanosoma were detected, but four out of seven strongly positive serological samples came from this group. This is the first study in Scandinavia on the presence of Trypanosoma in game animals. The results indicate that trypanosomiasis is frequently occurring among Swedish game animals.

  16. The flagellar adenylate kinases of Trypanosoma cruzi.

    Science.gov (United States)

    Camara, María de los Milagros; Bouvier, León A; Miranda, Mariana R; Pereira, Claudio A

    2015-01-01

    Adenylate kinases (ADK) are key enzymes involved in cell energy management. Trypanosomatids present the highest number of variants in a single cell in comparison with the rest of the living organisms. In this work, we characterized two flagellar ADKs from Trypanosoma cruzi, called TcADK1 and TcADK4, which are also located in the cell cytosol. Interestingly, TcADK1 presents a stage-specific expression. This variant was detected in epimastigotes cells, and was completely absent in trypomastigotes and amastigotes, while TcADK4 is present in the major life cycle stages of T. cruzi. Both variants are also regulated, in opposite ways, along the parasite growth curve suggesting that their expression depends on the intra- and extracellular conditions. Both, TcADK1 and TcADK4 present N-terminal extension that could be responsible for their subcellular localization. The presence of ADK variants in the flagellum would be critical for the provision of energy in a process of high ATP consumption such as cell motility.

  17. Trypanosoma evansi: identification and characterization of a variant surface glycoprotein lacking cysteine residues in its C-terminal domain.

    Science.gov (United States)

    Jia, Yonggen; Zhao, Xinxin; Zou, Jingru; Suo, Xun

    2011-01-01

    African trypanosomes are flagellated unicellular parasites which proliferate extracellularly in the mammalian host blood-stream and tissue spaces. They evade the hosts' antibody-mediated lyses by sequentially changing their variant surface glycoprotein (VSG). VSG tightly coats the entire parasite body, serving as a physical barrier. In Trypanosoma brucei and the closely related species Trypanosoma evansi, Trypanosoma equiperdum, each VSG polypeptide can be divided into N- and C-terminal domains, based on cysteine distribution and sequence homology. N-terminal domain, the basis of antigenic variation, is hypervariable and contains all the exposed epitopes; C-terminal domain is relatively conserved and a full set of four or eight cysteines were generally observed. We cloned two genes from two distinct variants of T. evansi, utilizing RT-PCR with VSG-specific primers. One contained a VSG type A N-terminal domain followed a C-terminal domain lacking cysteine residues. To confirm that this gene is expressed as a functional VSG, the expression and localization of the corresponding gene product were characterized using Western blotting and immunofluorescent staining of living trypanosomes. Expression analysis showed that this protein was highly expressed, variant-specific, and had a ubiquitous cellular surface localization. All these results indicated that it was expressed as a functional VSG. Our finding showed that cysteine residues in VSG C-terminal domain were not essential; the conserved C-terminal domain generally in T. brucei like VSGs would possibly evolve for regulating the VSG expression.

  18. Flux Analysis of the Trypanosoma brucei Glycolysis Based on a Multiobjective-Criteria Bioinformatic Approach

    Directory of Open Access Journals (Sweden)

    Amine Ghozlane

    2012-01-01

    Full Text Available Trypanosoma brucei is a protozoan parasite of major of interest in discovering new genes for drug targets. This parasite alternates its life cycle between the mammal host(s (bloodstream form and the insect vector (procyclic form, with two divergent glucose metabolism amenable to in vitro culture. While the metabolic network of the bloodstream forms has been well characterized, the flux distribution between the different branches of the glucose metabolic network in the procyclic form has not been addressed so far. We present a computational analysis (called Metaboflux that exploits the metabolic topology of the procyclic form, and allows the incorporation of multipurpose experimental data to increase the biological relevance of the model. The alternatives resulting from the structural complexity of networks are formulated as an optimization problem solved by a metaheuristic where experimental data are modeled in a multiobjective function. Our results show that the current metabolic model is in agreement with experimental data and confirms the observed high metabolic flexibility of glucose metabolism. In addition, Metaboflux offers a rational explanation for the high flexibility in the ratio between final products from glucose metabolism, thsat is, flux redistribution through the malic enzyme steps.

  19. In Vitro Trypanocidal Activity of Antibodies to Bacterially Ex­pressed Trypanosoma brucei Tubulin

    Directory of Open Access Journals (Sweden)

    GW Lubega

    2012-09-01

    Full Text Available Background: There are only four drugs for treating African trypanosomiasis, a devastating disease in sub-Saharan Africa. With slow discovery of better drugs, vaccination is viewed as the best method of control. We previously showed that antibodies to native Trypanosoma brucei brucei tubulin inhibit the growth of trypanosomes in culture. Here, we aimed to determine the effect of antibodies to bacte­rially expressed trypanosome tubulin on T. brucei brucei growth. Methods: T. brucei brucei alpha and beta tubulin genes were individually expressed in Escherichia coli under the tryptophan promoter. Monoclonal tubulin antibodies reacted specifically with the ex­pressed tubulins with no cross-reaction with the opposite tubulin. Rabbits were immunized with 450μg each of the concentrated recombinant tubulin, and production of antibodies assessed by ELISA and Western blotting. The effect of polyclonal antibodies on trypanosome growth was deter­mined by culturing bloodstream T. brucei brucei in up to 25% of antisera. Results: Low antisera dilutions (25% from the immunized rabbits inhibited trypanosome growth. The most cytotoxic antisera were from one rabbit immunized with a mixture of both alpha and beta tubulins. However, the result was not reproduced in other rabbits and there was no apparent effect on growth at higher antisera dilutions. Conclusion: Antibodies to bacterially expressed trypanosome tubulin are not effective at killing cul­tured bloodstream trypanosomes.

  20. Parasite development and host responses during the establishment of Trypanosoma brucei infection transmitted by tsetse fly.

    Science.gov (United States)

    Barry, J D; Emergy, D L

    1984-02-01

    Following inoculation of Trypanosoma brucei into large mammals by the tsetse fly a local skin reaction, the 'chancre', develops due to trypanosome proliferation. We have cannulated the afferent and efferent lymphatics of the draining lymph node in goats and examined the onset of a cellular reaction, the emigration of the parasite from the chancre and the development of both antigenic variation and the specific immune response. The chancre first became detectable by day 3 post-infection, peaked by day 6 and then subsided. Lymphocyte output increased 6- to 8-fold by day 10 and the number of lymphoblasts increased 50-fold in this period. Both then declined. Trypanosomes were detected in lymph 1-2 days before the chancre, peaked by days 5-6, declined during development of the chancre and then peaked again. The bloodstream population appeared by days 4-5 and displayed different kinetics from that in lymph. Recirculation of parasites through the lymphatics ensued. Lymph-borne trypanosome populations were highly pleomorphic. Parasites in lymph expressed firstly a mixture of the Variable Antigen Types (VATs) which are found characteristically in the tsetse fly, this being followed by a mixture of other VATs. The two groups overlapped in appearance. In the bloodstream the same sequence of events occurred although 2 or 3 days later. The specific antibody response, as measured by radioimmunoassay and agglutination, arose within a few days of the first detection of each VAT. Activities appeared first in the lymph and then in plasma.

  1. NLP is a novel transcription regulator involved in VSG expression site control in Trypanosoma brucei.

    Science.gov (United States)

    Narayanan, Mani Shankar; Kushwaha, Manish; Ersfeld, Klaus; Fullbrook, Alexander; Stanne, Tara M; Rudenko, Gloria

    2011-03-01

    Trypanosoma brucei mono-allelically expresses one of approximately 1500 variant surface glycoprotein (VSG) genes while multiplying in the mammalian bloodstream. The active VSG is transcribed by RNA polymerase I in one of approximately 15 telomeric VSG expression sites (ESs). T. brucei is unusual in controlling gene expression predominantly post-transcriptionally, and how ESs are mono-allelically controlled remains a mystery. Here we identify a novel transcription regulator, which resembles a nucleoplasmin-like protein (NLP) with an AT-hook motif. NLP is key for ES control in bloodstream form T. brucei, as NLP knockdown results in 45- to 65-fold derepression of the silent VSG221 ES. NLP is also involved in repression of transcription in the inactive VSG Basic Copy arrays, minichromosomes and procyclin loci. NLP is shown to be enriched on the 177- and 50-bp simple sequence repeats, the non-transcribed regions around rDNA and procyclin, and both active and silent ESs. Blocking NLP synthesis leads to downregulation of the active ES, indicating that NLP plays a role in regulating appropriate levels of transcription of ESs in both their active and silent state. Discovery of the unusual transcription regulator NLP provides new insight into the factors that are critical for ES control.

  2. Cell-penetrating peptide TP10 shows broad-spectrum activity against both Plasmodium falciparum and Trypanosoma brucei brucei.

    Science.gov (United States)

    Arrighi, Romanico B G; Ebikeme, Charles; Jiang, Yang; Ranford-Cartwright, Lisa; Barrett, Michael P; Langel, Ulo; Faye, Ingrid

    2008-09-01

    Malaria and trypanosomiasis are diseases which afflict millions and for which novel therapies are urgently required. We have tested two well-characterized cell-penetrating peptides (CPPs) for antiparasitic activity. One CPP, designated TP10, has broad-spectrum antiparasitic activity against Plasmodium falciparum, both blood and mosquito stages, and against blood-stage Trypanosoma brucei brucei.

  3. The changing epidemiology of Staphylococcus aureus bloodstream infection

    DEFF Research Database (Denmark)

    Laupland, K B; Lyytikäinen, O; Søgaard, M;

    2012-01-01

    Clin Microbiol Infect ABSTRACT: Although the epidemiology of Staphylococcus aureus bloodstream infection (BSI) has been changing, international comparisons are lacking. We sought to determine the incidence of S. aureus BSI and assess trends over time and by region. Population-based surveillance...

  4. Prevention of nosocomial bloodstream infections in preterm infants

    NARCIS (Netherlands)

    K. Helder MScN (Onno)

    2013-01-01

    textabstractProtecting patients from harm is the overarching theme of the studies presented here. More precisely, this thesis places a focus on the prevention of nosocomial or hospitalacquired bloodstream infections in preterm infants, thus saving them from further harm. A nosocomial infection is an

  5. Neonatal bloodstream infections in a pediatric hospital in Vietnam

    DEFF Research Database (Denmark)

    Kruse, Alexandra Yasmin; Thieu Chuong, D.H.; Phuong, C.N.;

    2013-01-01

    Septicemia and bloodstream infections (BSIs) are major causes of neonatal morbidity and mortality in developing countries. We prospectively recorded all positive blood cultures (BSI) among neonates admitted consecutively to a tertiary pediatric hospital in Vietnam during a 12-month period. Among...

  6. Trypanosoma cruzi: strain selection by diferent schedules of mouse passage of an initially mixed infection

    Directory of Open Access Journals (Sweden)

    Maria P. Deane

    1984-12-01

    Full Text Available From an initial double infection in mice, established by simultaneous and equivalent inocula of bloodstream forms of strains Y and F of Trypanosoma cruzi, two lines were derived by subinoculations: one (W passaged every week, the other (M every month. Through biological and biochemical methods only the Y strain was identified at the end of the 10th and 16th passages of line W and only the F strain at the 2nd and 4th passages of line M. The results illustrate strain selection through laboratory manipulation of initially mixed populations of T. cruzi.De uma infecção inicialmente dupla em camundongo, estabelecida por inóculo simultaneo e equivalente de formas sanguíneas das cepas Y e F de Trypanosoma cruzi, duas linhagens foram originadas por subinoculações: uma (W passada casa semana, a outra (M cada mês. Por métodos biológicos e bioquímicos apenas a cepa Y foi identificada ao fim a 10a. e 16a. passagens da linhagem W e apenas a cepa F na 2a. e 4a.passagens de linhagem M. Os resultados demonstram a seleção de cepas através de manipulação em laboratorio de populações inicialmente mistas de T. cruzi.

  7. Regulators of Trypanosoma brucei cell cycle progression and differentiation identified using a kinome-wide RNAi screen.

    Directory of Open Access Journals (Sweden)

    Nathaniel G Jones

    2014-01-01

    Full Text Available The African trypanosome, Trypanosoma brucei, maintains an integral link between cell cycle regulation and differentiation during its intricate life cycle. Whilst extensive changes in phosphorylation have been documented between the mammalian bloodstream form and the insect procyclic form, relatively little is known about the parasite's protein kinases (PKs involved in the control of cellular proliferation and differentiation. To address this, a T. brucei kinome-wide RNAi cell line library was generated, allowing independent inducible knockdown of each of the parasite's 190 predicted protein kinases. Screening of this library using a cell viability assay identified ≥42 PKs that are required for normal bloodstream form proliferation in culture. A secondary screen identified 24 PKs whose RNAi-mediated depletion resulted in a variety of cell cycle defects including in G1/S, kinetoplast replication/segregation, mitosis and cytokinesis, 15 of which are novel cell cycle regulators. A further screen identified for the first time two PKs, named repressor of differentiation kinase (RDK1 and RDK2, depletion of which promoted bloodstream to procyclic form differentiation. RDK1 is a membrane-associated STE11-like PK, whilst RDK2 is a NEK PK that is essential for parasite proliferation. RDK1 acts in conjunction with the PTP1/PIP39 phosphatase cascade to block uncontrolled bloodstream to procyclic form differentiation, whilst RDK2 is a PK whose depletion efficiently induces differentiation in the absence of known triggers. Thus, the RNAi kinome library provides a valuable asset for functional analysis of cell signalling pathways in African trypanosomes as well as drug target identification and validation.

  8. A Protein Complex Map of Trypanosoma brucei.

    Directory of Open Access Journals (Sweden)

    Vahid H Gazestani

    2016-03-01

    Full Text Available The functions of the majority of trypanosomatid-specific proteins are unknown, hindering our understanding of the biology and pathogenesis of Trypanosomatida. While protein-protein interactions are highly informative about protein function, a global map of protein interactions and complexes is still lacking for these important human parasites. Here, benefiting from in-depth biochemical fractionation, we systematically interrogated the co-complex interactions of more than 3354 protein groups in procyclic life stage of Trypanosoma brucei, the protozoan parasite responsible for human African trypanosomiasis. Using a rigorous methodology, our analysis led to identification of 128 high-confidence complexes encompassing 716 protein groups, including 635 protein groups that lacked experimental annotation. These complexes correlate well with known pathways as well as for proteins co-expressed across the T. brucei life cycle, and provide potential functions for a large number of previously uncharacterized proteins. We validated the functions of several novel proteins associated with the RNA-editing machinery, identifying a candidate potentially involved in the mitochondrial post-transcriptional regulation of T. brucei. Our data provide an unprecedented view of the protein complex map of T. brucei, and serve as a reliable resource for further characterization of trypanosomatid proteins. The presented results in this study are available at: www.TrypsNetDB.org.

  9. Differential gene expression during Trypanosoma cruzi metacyclogenesis

    Directory of Open Access Journals (Sweden)

    Marco Aurelio Krieger

    1999-09-01

    Full Text Available The transformation of epimastigotes into metacyclic trypomastigotes involves changes in the pattern of expressed genes, resulting in important morphological and functional differences between these developmental forms of Trypanosoma cruzi. In order to identify and characterize genes involved in triggering the metacyclogenesis process and in conferring to metacyclic trypomastigotes their stage specific biological properties, we have developed a method allowing the isolation of genes specifically expressed when comparing two close related cell populations (representation of differential expression or RDE. The method is based on the PCR amplification of gene sequences selected by hybridizing and subtracting the populations in such a way that after some cycles of hybridization-amplification genes specific to a given population are highly enriched. The use of this method in the analysis of differential gene expression during T. cruzi metacyclogenesis (6 hr and 24 hr of differentiation and metacyclic trypomastigotes resulted in the isolation of several clones from each time point. Northern blot analysis showed that some genes are transiently expressed (6 hr and 24 hr differentiating cells, while others are present in differentiating cells and in metacyclic trypomastigotes. Nucleotide sequencing of six clones characterized so far showed that they do not display any homology to gene sequences available in the GeneBank.

  10. The krebs cycle enzyme α-ketoglutarate decarboxylase is an essential glycosomal protein in bloodstream African trypanosomes.

    Science.gov (United States)

    Sykes, Steven; Szempruch, Anthony; Hajduk, Stephen

    2015-03-01

    α-Ketoglutarate decarboxylase (α-KDE1) is a Krebs cycle enzyme found in the mitochondrion of the procyclic form (PF) of Trypanosoma brucei. The bloodstream form (BF) of T. brucei lacks a functional Krebs cycle and relies exclusively on glycolysis for ATP production. Despite the lack of a functional Krebs cycle, α-KDE1 was expressed in BF T. brucei and RNA interference knockdown of α-KDE1 mRNA resulted in rapid growth arrest and killing. Cell death was preceded by progressive swelling of the flagellar pocket as a consequence of recruitment of both flagellar and plasma membranes into the pocket. BF T. brucei expressing an epitope-tagged copy of α-KDE1 showed localization to glycosomes and not the mitochondrion. We used a cell line transfected with a reporter construct containing the N-terminal sequence of α-KDE1 fused to green fluorescent protein to examine the requirements for glycosome targeting. We found that the N-terminal 18 amino acids of α-KDE1 contain overlapping mitochondrion- and peroxisome-targeting sequences and are sufficient to direct localization to the glycosome in BF T. brucei. These results suggest that α-KDE1 has a novel moonlighting function outside the mitochondrion in BF T. brucei.

  11. Central line-associated bloodstream infections: prevention and management.

    Science.gov (United States)

    Weber, David J; Rutala, William A

    2011-03-01

    Approximately 80,000 central venous line-associated bloodstream infections (CLA-BSI) occur in the United States each year. CLA-BSI is most commonly caused by coagulase-negative staphylococci, Staphylococcus aureus, Candida spp, and aerobic gram-negative bacilli. These organisms commonly gain entrance in into the bloodstream via the catheter-skin interface (insertion site) or via the catheter hub. Use of strict aseptic technique for insertion is the key method for the prevention of CLA-BSI. Various methods can be used to reduce unacceptably high rates of CLA-BSI, including use of an antiseptic- or antibiotic-impregnated catheter, daily chlorhexidine baths/washes, and placement of a chlorhexidine-impregnated sponge over the insertion site.

  12. Trypanosoma evansi isolated from capybara (Hidrochaeris hidrochaeris)

    OpenAIRE

    2001-01-01

    A study was conducted to determine the morphological and biometric characteristics of Trypanosoma isolated from 50 capybaras animals, raised in captivity in the Peruvian Amazon. Trypanosoma was found in 14 blood samples using the microhaematocrit, wide drop, and Giemsa-stain methods and T. evansi was identified through morphological details in all 14 positive samples (the subterminal kinetoplast, the developed undulating membrane, and a long free flagellum were used for the identification of ...

  13. Trypanosoma evansi isolated from capybara (Hidrochaeris hidrochaeris

    Directory of Open Access Journals (Sweden)

    Karina Muñoz

    2001-10-01

    Full Text Available A study was conducted to determine the morphological and biometric characteristics of Trypanosoma isolated from 50 capybaras animals, raised in captivity in the Peruvian Amazon. Trypanosoma was found in 14 blood samples using the microhaematocrit, wide drop, and Giemsa-stain methods and T. evansi was identified through morphological details in all 14 positive samples (the subterminal kinetoplast, the developed undulating membrane, and a long free flagellum were used for the identification of the agent.

  14. Prosthetic valve endocarditis and bloodstream infection due to Mycobacterium chimaera.

    Science.gov (United States)

    Achermann, Yvonne; Rössle, Matthias; Hoffmann, Matthias; Deggim, Vanessa; Kuster, Stefan; Zimmermann, Dieter R; Bloemberg, Guido; Hombach, Michael; Hasse, Barbara

    2013-06-01

    Prosthetic valve endocarditis (PVE) due to fast-growing nontuberculous mycobacteria (NTM) has been reported anecdotally. Reports of PVE with slowly growing NTM, however, are lacking. We present here one case of PVE and one case of bloodstream infection caused by Mycobacterium chimaera. Randomly amplified polymorphic DNA (RAPD)-PCR indicated a relatedness of the two M. chimaera strains. Both patients had heart surgery 2 years apart from each other. A nosocomial link was not detected.

  15. Prosthetic Valve Endocarditis and Bloodstream Infection Due to Mycobacterium chimaera

    OpenAIRE

    Achermann, Yvonne; Rössle, Matthias; Hoffmann, Matthias; Deggim, Vanessa; Kuster, Stefan; Zimmermann, Dieter R.; Bloemberg, Guido; Hombach, Michael; Hasse, Barbara

    2013-01-01

    Prosthetic valve endocarditis (PVE) due to fast-growing nontuberculous mycobacteria (NTM) has been reported anecdotally. Reports of PVE with slowly growing NTM, however, are lacking. We present here one case of PVE and one case of bloodstream infection caused by Mycobacterium chimaera. Randomly amplified polymorphic DNA (RAPD)-PCR indicated a relatedness of the two M. chimaera strains. Both patients had heart surgery 2 years apart from each other. A nosocomial link was not detected.

  16. Endothelial JAM-A promotes reovirus viremia and bloodstream dissemination.

    Science.gov (United States)

    Lai, Caroline M; Boehme, Karl W; Pruijssers, Andrea J; Parekh, Vrajesh V; Van Kaer, Luc; Parkos, Charles A; Dermody, Terence S

    2015-02-01

    Viruses that cause systemic disease often spread through the bloodstream to infect target tissues. Although viremia is an important step in the pathogenesis of many viruses, how viremia is established is not well understood. Reovirus has been used to dissect mechanisms of viral pathogenesis and is being evaluated in clinical trials as an oncolytic agent. After peroral entry into mice, reovirus replicates within the gastrointestinal tract and disseminates systemically via hematogenous or neural routes. Junctional adhesion molecule-A (JAM-A) is a tight junction protein that serves as a receptor for reovirus. JAM-A is required for establishment of viremia and viral spread to sites of secondary replication. JAM-A also is expressed on the surface of circulating hematopoietic cells. To determine contributions of endothelial and hematopoietic JAM-A to reovirus dissemination and pathogenesis, we generated strains of mice with altered JAM-A expression in these cell types and assessed bloodstream spread of reovirus strain type 1 Lang (T1L), which disseminates solely by hematogenous routes. We found that endothelial JAM-A but not hematopoietic JAM-A facilitates reovirus T1L bloodstream entry and egress. Understanding how viruses establish viremia may aid in development of inhibitors of this critical step in viral pathogenesis and foster engineering of improved oncolytic viral vectors.

  17. Inhibitors of the mitochondrial cytochrome b-c1 complex inhibit the cyanide-insensitive respiration of Trypanosoma brucei.

    Science.gov (United States)

    Turrens, J F; Bickar, D; Lehninger, A L

    1986-06-01

    The cyanide-insensitive respiration of bloodstream trypomastigote forms of Trypanosoma brucei (75 +/- 8 nmol O2 min-1(mg protein)-1) is completely inhibited by the mitochondrial ubiquinone-like inhibitors 2-hydroxy-3-undecyl-1,4-naphthoquinone (UHNQ) and 5-n-undecyl-6-hydroxy-4,7-dioxobenzothiazole (UHDBT). The Ki values for UHDBT (30 nM) and UHNQ (2 microM) are much lower than the reported Ki for salicylhydroxamic acid (SHAM) (5 microM), a widely used inhibitor of the cyanide-insensitive oxidase. UHNQ also stimulated the glycerol-3-phosphate-dependent reduction of phenazine methosulfate, demonstrating that the site of UHNQ inhibition is on the terminal oxidase of the cyanide-insensitive respiration of T. brucei. These results suggest that a ubiquinone-like compound may act as an electron carrier between the two enzymatic components of the cyanide-insensitive glycerol-3-phosphate oxidase.

  18. 21 CFR 866.3870 - Trypanosoma spp. serological reagents.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Trypanosoma spp. serological reagents. 866.3870... (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3870 Trypanosoma spp. serological reagents. (a) Identification. Trypanosoma spp. serological reagents are devices...

  19. Tridimensional ultrastructure and glycolipid pattern studies of Trypanosoma dionisii.

    Science.gov (United States)

    Oliveira, Miriam Pires de Castro; Ramos, Thiago Cesar Prata; Pinheiro, Adriana Maria V N; Bertini, Silvio; Takahashi, Helio Kiyoshi; Straus, Anita Hilda; Haapalainen, Edna Freymuller

    2013-12-01

    Trypanosoma (Schizotrypanum) dionisii is a non-pathogenic bat trypanosome closely related to Trypanosoma cruzi, the etiological agent of Chaga's disease. Both kinetoplastids present similar morphological stages and are able to infect mammalian cells in culture. In the present study we examined 3D ultrastructure aspects of the two species by serial sectioning epimastigote and trypomastigote forms, and identified common carbohydrate epitopes expressed in T. dionisii, T. cruzi and Leishmania major. A major difference in 3D morphology was that T. dionisii epimastigote forms present larger multivesicular structures, restricted to the parasite posterior region. These structures could be related to T. cruzi reservosomes and are also rich in cruzipain, the major cysteine-proteinase of T. cruzi. We analyzed the reactivity of two monoclonal antibodies: MEST-1 directed to galactofuranose residues of glycolipids purified from Paracoccidioides brasiliensis, and BST-1 directed to glycolipids purified from T. cruzi epimastigotes. Both antibodies were reactive with T. dionisii epimastigotes by indirect immunofluorescense, but we noted differences in the location and intensity of the epitopes, when compared to T. cruzi. In summary, despite similar features in cellular structure and life cycle of T. dionisii and T. cruzi, we observed a unique morphological characteristic in T. dionisii that deserves to be explored.

  20. The innate resistance of Trypanosoma copemani to human serum.

    Science.gov (United States)

    Austen, J M; Ryan, U; Ditcham, W G F; Friend, J A; Reid, S A

    2015-06-01

    Trypanosoma copemani is known to be infective to a variety of Australian marsupials. Characterisation of this parasite revealed the presence of stercorarian-like life-cycle stages in culture, which are similar to T. rangeli and T. cruzi. The blood incubation infectivity test (BIIT) was adapted and used to determine if T. copemani, like T. cruzi and T. rangeli, has the potential to grow in the presence of human serum. To eliminate any effects of anticoagulants on the complement system and on human high density lipoprotein (HDL), only fresh whole human blood was used. Trypanosoma copemani was observed by microscopy in all human blood cultures from day 5 to day 19 post inoculation (PI). The mechanism for normal human serum (NHS) resistance in T. copemani is not known. The results of this study show that at least one native Australian trypanosome species may have the potential to be infective for humans.

  1. Molecular and Functional Characterization of a Trypanosoma cruzi Nuclear Adenylate Kinase Isoform

    OpenAIRE

    María de los Milagros Cámara; Bouvier, León A.; Canepa, Gaspar E.; Mariana R Miranda; Pereira, Claudio A.

    2013-01-01

    Trypanosoma cruzi, the etiological agent of Chagas’ disease, is an early divergent eukaryote in which control of gene expression relies mainly in post-transcriptional mechanisms. Transcription levels are globally up and down regulated during the transition between proliferating and non-proliferating life-cycle stages. In this work we characterized a nuclear adenylate kinase isoform (TcADKn) that is involved in ribosome biogenesis. Nuclear adenylate kinases have been recently described in a fe...

  2. Bloodstream infection caused by nontoxigenic Corynebacterium diphtheriae in an immunocompromised host in the United States.

    Science.gov (United States)

    Wojewoda, Christina M; Koval, Christine E; Wilson, Deborah A; Chakos, Mary H; Harrington, Susan M

    2012-06-01

    Corynebacterium species are well-known causes of catheter-related bloodstream infections. Toxigenic strains of Corynebacterium diphtheriae cause respiratory diphtheria. We report a bloodstream infection caused by a nontoxigenic strain of C. diphtheriae and discuss the epidemiology, possible sources of the infection, and the implications of rapid species identification of corynebacteria.

  3. An essential signal peptide peptidase identified in an RNAi screen of serine peptidases of Trypanosoma brucei.

    Directory of Open Access Journals (Sweden)

    Catherine X Moss

    Full Text Available The serine peptidases of Trypanosoma brucei have been viewed as potential drug targets. In particular, the S9 prolyl oligopeptidase subfamily is thought to be a good avenue for drug discovery. This is based on the finding that some S9 peptidases are secreted and active in the mammalian bloodstream, and that they are a class of enzyme against which drugs have successfully been developed. We collated a list of all serine peptidases in T. brucei, identifying 20 serine peptidase genes, of which nine are S9 peptidases. We screened all 20 serine peptidases by RNAi to determine which, if any, are essential for bloodstream form T. brucei survival. All S9 serine peptidases were dispensable for parasite survival in vitro, even when pairs of similar genes, coding for oligopeptidase B or prolyl oligopeptidase, were targeted simultaneously. We also found no effect on parasite survival in an animal host when the S9 peptidases oligopeptidase B, prolyl oligopeptidase or dipeptidyl peptidase 8 were targeted. The only serine peptidase to emerge from the RNAi screen as essential was a putative type-I signal peptide peptidase (SPP1. This gene was essential for parasite survival both in vitro and in vivo. The growth defect conferred by RNAi depletion of SPP1 was rescued by expression of a functional peptidase from an RNAi resistant SPP1 gene. However, expression of catalytically inactive SPP1 was unable to rescue cells from the SPP1 depleted phenotype, demonstrating that SPP1 serine peptidase activity is necessary for T. brucei survival.

  4. Comparative analysis of the kinomes of three pathogenic trypanosomatids: Leishmania major, Trypanosoma brucei and Trypanosoma cruzi

    OpenAIRE

    Parsons, Marilyn; Worthey, Elizabeth A.; Ward, Pauline N; Mottram, Jeremy C.

    2005-01-01

    Background The trypanosomatids Leishmania major, Trypanosoma brucei and Trypanosoma cruzi cause some of the most debilitating diseases of humankind: cutaneous leishmaniasis, African sleeping sickness, and Chagas disease. These protozoa possess complex life cycles that involve development in mammalian and insect hosts, and a tightly coordinated cell cycle ensures propagation of the highly polarized cells. However, the ways in which the parasites respond to their environment and coordinate intr...

  5. Trypanosoma epinepheli n. sp. (Kinetoplastida) from a farmed marine fish in China, the brown-marbled grouper (Epinephelus fuscoguttatus).

    Science.gov (United States)

    Su, Youlu; Feng, Juan; Jiang, Jingzhe; Guo, Zhixun; Liu, Guangfeng; Xu, Liwen

    2014-01-01

    An outbreak of trypanosomosis occurred in farmed Epinephelus fuscoguttatus in Xincun Bay, province of Hainan, South China Sea. The infected fish showed loss of appetite, lethargy, emaciation, severe anemia, and splenomegaly. Light and scanning electron microscopic examination of bloodstream trypomastigotes revealed morphological features typical for small-sized marine fish trypanosomes. The trypanosome possesses a short body length (mean 22.3 μm, range 17.6-25.9 μm) and narrow body width (mean1.7 μm, range 1.3-2.0 μm), a central nucleus, a narrow but distinct undulating membrane, and a relatively long free flagellum (mean 10.1 μm, range 7.4-13.3 μm). The kinetoplast is situated at approximately one quarter of body length from posterior extremity. The division process of this trypanosome was observed in the peripheral blood of the host, and occurred by transverse constriction at a point between the kinetoplasts. Comparison of the small subunit rDNA (SSU rDNA) sequences revealed that the trypanosome from E. fuscoguttatus showed 93.4-97.1% identity with the available sequences from Trypanosoma spp. from other piscine hosts. Phylogenetic analysis supported the existence of an aquatic clade, and the present trypanosome grouped with other marine fish trypanosomes, in a subclade together with Trypanosoma senegalense. Based on the differences in morphological characteristics, host species, and molecular data, the trypanosome infecting E. fuscoguttatus is considered to be a new species, for which we propose the name Trypanosoma epinepheli n. sp.

  6. The cell surface of Trypanosoma cruzi

    Directory of Open Access Journals (Sweden)

    Wanderley de Souza

    1984-01-01

    Full Text Available The cell surface of trypanosomatids is formed by the plasma membrane and a layer of sub-pellicular microtubules which are connected to the plasma membrane. The plasma membrane is composed by proteins, lipids and carbohydrates which form the glycocalix. In this paper we will review briefly aspects related to the organization of the cell surface of Trypanosoma cruzi.

  7. The kinetoplast DNA of Trypanosoma equiperdum.

    NARCIS (Netherlands)

    A.C.C. Frasch; S.L. Hajduk; J.H.J. Hoeijmakers (Jan); P. Borst (Piet); F. Brunel; J. Davison

    1980-01-01

    textabstractWe have analyzed the kinetoplast DNA for Trypanosoma equiperdum (American Type Culture Collection 30019) and two dyskinetoplastic strains derived from it. The DNA networks from the kinetoplastic strain are made up of catenated mini-circles and maxi-circles, like the networks from the clo

  8. Structural Insights into Inhibition of Sterol 14[alpha]-Demethylase in the Human Pathogen Trypanosoma cruzi

    Energy Technology Data Exchange (ETDEWEB)

    Lepesheva, Galina I.; Hargrove, Tatiana Y.; Anderson, Spencer; Kleshchenko, Yuliya; Furtak, Vyacheslav; Wawrzak, Zdzislaw; Villalta, Fernando; Waterman, Michael R. (Vanderbilt); (NWU); (Meharry)

    2010-09-02

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

  9. The isolation and identification of Trypanosoma cruzi from raccoons in Maryland

    Science.gov (United States)

    Walton, B.C.; Bauman, P.M.; Diamond, L.S.; Herman, C.M.

    1958-01-01

    Five raccoons trapped at Patuxent Research Refuge, Laurel, Maryland, were found to have trypanosomes in the blood which were morphologically indistinguishable from Trypanosoma cruzi on stained smears. The organism grew well in culture. It developed and reproduced in Triatoma protracta, T. infestans, T. phyllosoma, and Rhodnius prolixus. Experimental infections were produced in raccoons, opossums, mice, rats, and monkeys by inoculation of blood, culture, and triatome forms. Typical leishmaniform bodies were found in tissue sections of cardiac muscle fibers from naturally and experimentally infected animals. Cross agglutinations carried out with Iiving cultural forms and rabbit antisera demonstrated a close antigenic relationship between the raccoon trypanosome and T. cruzi (Brazil strain). On the basis of (1) morphology, (2) presence of leishmaniform tissue stages, (3) development in triatomes, (4) infectivity to a variety of mammals, (5) culture characteristics, and (6) cross reactions in serological tests, this parasite is considered conspecific with Trypanosoma cruzi (Chagas, 1909), the causative agent of American human trypanosomiasis.

  10. Systemic effects of locally injected platelet rich plasma in a rat model: an analysis on muscle and bloodstream.

    Science.gov (United States)

    Borrione, P; Grasso, L; Racca, S; Abbadessa, G; Carriero, V; Fagnani, F; Quaranta, F; Pigozzi, F

    2015-01-01

    Abundant evidence suggests that growth factors, contained in platelets alpha granules, may play a key role in the early stages of the muscle healing process with particular regard to the inflammatory phase. Although the contents of the platelet-rich plasma preparations have been extensively studied, the biological mechanisms involved as well as the systemic effects and the related potential doping implications of this approach are still largely unknown. The aim of the present study was to investigate whether local platelet-rich plasma administration may modify the levels of specific cytokines and growth factors both in treated muscle and bloodstream in rats. An additional aim was to investigate more deeply whether the local platelet-rich plasma administration may exert systemic effects by analyzing contralateral lesioned but untreated muscles. The results showed that platelet-rich plasma treatment induced a modification of certain cytokines and growth factor levels in muscle but not in the bloodstream, suggesting that local platelet-rich plasma treatment influenced directly or, more plausibly, indirectly the synthesis or recruitment of cytokines and growth factors at the site of injury. Moreover, the observed modifications of cytokine and growth factor levels in contralateral injured but not treated muscles, strongly suggested a systemic effect of locally injected platelet-rich plasma.

  11. Concentration of erythrocyte-based magnetic carriers in the bloodstream

    Energy Technology Data Exchange (ETDEWEB)

    Danilov, Y.N.; Il' ina, M.B.; Makharudov, S.Y.; Orekhov, A.N.; Rodchenko, S.A.; Samokhin, G.P.

    1986-04-01

    The writers postulated that magnetic erythrocytes (ME), injected into the bloodstream, may be concentrated in an assigned region of the vascular bed with the aid of the field of a permanent magnet. To test this hypothesis, erythrocytes ''loaded'' with colloidal magnetite were used, and concentrated in experiments in vitro and in vivo. For the experiments in vivo ME were labeled with sodium pertechnetate (Na /SUP 99n/ TcO4) was estimated in ME with a Rackgamma 1270 counter by determining radioactivity in the ME suspension and in the supernatant. For the experiment in vivo, a midline laparotomy was performed on a dog weighing 14 kg under intravenous hexobarbital anesthesia. The distribution of ME was recorded by means of a gamma-camera.

  12. Mortality in enterococcal bloodstream infections increases with inappropriate antimicrobial therapy

    DEFF Research Database (Denmark)

    Suppli, M.; Aabenhus, R.; Harboe, Z.B.;

    2010-01-01

    Enterococcus species are common in nosocomial bloodstream infections and their incidence is rising. Although well recognized in several serious bacterial infections, the influence of appropriate antimicrobial therapy in enterococcal bacteraemia has not been fully settled. The aim of the study...... in Denmark. Patients with growth of non-enterococcus co-pathogens apart from the enterococcal bacteraemia were also included, as were patients with repeated enterococcal bacteraemia. Time to appropriate antimicrobial therapy was counted from the first episode. Appropriate antibiotic therapy was defined...... as any therapy with documented clinical effect, in vitro activity and a minimum treatment length of 6 days. Multivariate regression models were built to determine the independent risk factors for mortality. We included 196 patients with enterococcal bacteraemia. Appropriate antibiotics for at least 6...

  13. Reducing bloodstream infection with a chlorhexidine gel IV dressing.

    Science.gov (United States)

    Jeanes, Annette; Bitmead, James

    The use of vascular access devices (VAD) is common in healthcare provision but there is a significant risk of acquiring an infection. Central venous catheters (CVC) are associated with the highest risk of intravenous catheter-related bloodstream infection (CRBSI). 3M™ Tegaderm™ CHG IV dressing is a semi-permeable transparent adhesive dressing with an integrated gel pad containing chlorhexidine gluconate 2%. This product was reviewed by the National Institute for Health and Care Excellence (NICE) in 2015, recommending that Tegaderm CHG could be used for CVC and arterial line dressings in high-dependency and intensive-care settings. This article discusses issues around CRBSI, interventions to reduce the risk of CRBSI, and the use of Tegaderm CHG dressing.

  14. Antimicrobial resistance predicts death in Tanzanian children with bloodstream infections: a prospective cohort study

    Directory of Open Access Journals (Sweden)

    Msangi Viola

    2007-05-01

    Full Text Available Abstract Background Bloodstream infection is a common cause of hospitalization, morbidity and death in children. The impact of antimicrobial resistance and HIV infection on outcome is not firmly established. Methods We assessed the incidence of bloodstream infection and risk factors for fatal outcome in a prospective cohort study of 1828 consecutive admissions of children aged zero to seven years with signs of systemic infection. Blood was obtained for culture, malaria microscopy, HIV antibody test and, when necessary, HIV PCR. We recorded data on clinical features, underlying diseases, antimicrobial drug use and patients' outcome. Results The incidence of laboratory-confirmed bloodstream infection was 13.9% (255/1828 of admissions, despite two thirds of the study population having received antimicrobial therapy prior to blood culture. The most frequent isolates were klebsiella, salmonellae, Escherichia coli, enterococci and Staphylococcus aureus. Furthermore, 21.6% had malaria and 16.8% HIV infection. One third (34.9% of the children with laboratory-confirmed bloodstream infection died. The mortality rate from Gram-negative bloodstream infection (43.5% was more than double that of malaria (20.2% and Gram-positive bloodstream infection (16.7%. Significant risk factors for death by logistic regression modeling were inappropriate treatment due to antimicrobial resistance, HIV infection, other underlying infectious diseases, malnutrition and bloodstream infection caused by Enterobacteriaceae, other Gram-negatives and candida. Conclusion Bloodstream infection was less common than malaria, but caused more deaths. The frequent use of antimicrobials prior to blood culture may have hampered the detection of organisms susceptible to commonly used antimicrobials, including pneumococci, and thus the study probably underestimates the incidence of bloodstream infection. The finding that antimicrobial resistance, HIV-infection and malnutrition predict fatal

  15. Trypanosoma cruzi extracellular amastigotes and host cell signaling: more pieces to the puzzle

    Directory of Open Access Journals (Sweden)

    Éden Ramalho Ferreira

    2012-11-01

    Full Text Available Among the different infective stages that Trypanosoma cruzi employs to invade cells, extracellular amastigotes have recently gained attention by our group. This is true primarily because these amastigotes are able to infect cultured cells and animals, establishing a sustainable infective cycle. Extracellular amastigotes are thus an excellent means of adaptation and survival for T. cruzi, whose different infective stages each utilize unique mechanisms for attachment and penetration. Here we discuss some features of host cell invasion by extracellular amastigotes and the associated host cell signaling events that occur as part of the process.

  16. Cefotaxime resistance and outcome of Klebsiella spp bloodstream infection.

    Science.gov (United States)

    Ortega, M; Marco, F; Soriano, A; Almela, M; Martínez, J A; López, J; Pitart, C; Mensa, J

    2011-12-01

    We attempt to describe the epidemiology and outcome associated with cefotaxime-resistant (CTX-R) Klebsiella spp bacteraemia. Klebsiella spp bloodstream infection episodes prospectively collected through a blood culture surveillance programme from January 1991 to December 2008 in a single institution were analysed. A total of 910 monomicrobial episodes of Klebsiella spp bacteraemia were identified during the study period. The most important sources were from urinary tract infection, unknown sources, billiary focus and catheter related infection. There were 112 (12%) CTX-R isolates. Out of 112 isolates, 98 were CTX-R by Extended-Spectrum β-Lactamase production. Shock on presentation and mortality were significantly more frequent in CTX-R than in CTX susceptible isolates. Inappropriate empirical therapy was received in 50 (45%) cases in the CTX-R Klebsiella spp group (13 cases of death, 26%). Predictive factors associated with CTX-R Klebsiella spp isolate were: previous β-lactam therapy (OR = 4.16), nosocomial acquired bacteraemia (OR = 1.93), solid organ trasplantation (OR = 2.09) and shock (OR = 1.90). Independent risk factors associated with mortality in Klebsiella spp bacteraemia were: age (OR = 1.03), liver cirrhosis (OR = 2.63), ultimately or rapidly fatal prognosis of underlying disease (OR = 2.44), shock (OR = 8.60), pneumonia (OR = 4.96) or intraabdominal (OR = 3.85) source of bacteraemia and CTX-R isolate (OR = 4.63). Klebsiella spp is an important cause of bloodstream infection. CTX-R isolates have been increasing since 2000. CTX-R is an independent factor associated with mortality in Klebsiella spp bacteraemia.

  17. The Role of Folate Transport in Antifolate Drug Action in Trypanosoma brucei*

    Science.gov (United States)

    Dewar, Simon; Sienkiewicz, Natasha; Ong, Han B.; Wall, Richard J.; Horn, David

    2016-01-01

    The aim of this study was to identify and characterize mechanisms of resistance to antifolate drugs in African trypanosomes. Genome-wide RNAi library screens were undertaken in bloodstream form Trypanosoma brucei exposed to the antifolates methotrexate and raltitrexed. In conjunction with drug susceptibility and folate transport studies, RNAi knockdown was used to validate the functions of the putative folate transporters. The transport kinetics of folate and methotrexate were further characterized in whole cells. RNA interference target sequencing experiments identified a tandem array of genes encoding a folate transporter family, TbFT1–3, as major contributors to antifolate drug uptake. RNAi knockdown of TbFT1–3 substantially reduced folate transport into trypanosomes and reduced the parasite's susceptibly to the classical antifolates methotrexate and raltitrexed. In contrast, knockdown of TbFT1–3 increased susceptibly to the non-classical antifolates pyrimethamine and nolatrexed. Both folate and methotrexate transport were inhibited by classical antifolates but not by non-classical antifolates or biopterin. Thus, TbFT1–3 mediates the uptake of folate and classical antifolates in trypanosomes, and TbFT1–3 loss-of-function is a mechanism of antifolate drug resistance. PMID:27703008

  18. Trypanosoma brucei CYP51: Essentiality and Targeting Therapy in an Experimental Model

    Science.gov (United States)

    Dauchy, Frédéric-Antoine; Bonhivers, Mélanie; Landrein, Nicolas; Dacheux, Denis; Courtois, Pierrette; Lauruol, Florian; Daulouède, Sylvie

    2016-01-01

    Trypanosoma brucei gambiense is the main causative agent of Human African Trypanosomiasis (HAT), also known as sleeping sickness. Because of limited alternatives and treatment toxicities, new therapeutic options are urgently needed for patients with HAT. Sterol 14alpha-demethylase (CYP51) is a potential drug target but its essentiality has not been determined in T. brucei. We used a tetracycline-inducible RNAi system to assess the essentiality of CYP51 in T. brucei bloodstream form (BSF) cells and we evaluated the effect of posaconazole, a well-tolerated triazole drug, within a panel of virulent strains in vitro and in a murine model. Expression of CYP51 in several T. brucei cell lines was demonstrated by western blot and its essentiality was demonstrated by RNA interference (CYP51RNAi) in vitro. Following reduction of TbCYP51 expression by RNAi, cell growth was reduced and eventually stopped compared to WT or non-induced cells, showing the requirement of CYP51 in T. brucei. These phenotypes were rescued by addition of ergosterol. Additionally, CYP51RNAi induction caused morphological defects with multiflagellated cells (ptrypanosomiasis. PMID:27855164

  19. Melarsoprol- and pentamidine-resistant Trypanosoma brucei rhodesiense populations and their cross-resistance.

    Science.gov (United States)

    Bernhard, Sonja C; Nerima, Barbara; Mäser, Pascal; Brun, Reto

    2007-11-01

    Resistance to melarsoprol and pentamidine was induced in bloodstream-form Trypanosoma brucei rhodesiense STIB 900 in vitro, and drug sensitivity was determined for melarsoprol, pentamidine and furamidine. The resistant populations were also inoculated into immunosuppressed mice to verify infectivity and to monitor whether rodent passage selects for clones with altered drug sensitivity. After proliferation in the mouse, trypanosomes were isolated and their IC(50) values to the three drugs were determined. To assess the stability of drug-induced resistance, drug pressure was ceased for 2 months and the drug sensitivity was determined again. Resistance was stable, with a few exceptions that are discussed. Drug IC(50)s indicated cross-resistance among all drugs, but to varying extents: resistance of the melarsoprol-selected and pentamidine-selected trypanosomes to pentamidine was the same, but the pentamidine-selected trypanosome population showed lower resistance to melarsoprol than the melarsoprol-selected trypanosomes. Interestingly, both resistant populations revealed the same intermediate cross-resistance to furamidine. Resistant trypanosome populations were characterised by molecular means, referring to the status of the TbAT1 gene. The melarsoprol-selected population apparently had lost TbAT1, whereas in the pentamidine-selected trypanosome population it was still present.

  20. Role of membrane-bound IgM in Trypanosoma cruzi evasion from immune clearance.

    Science.gov (United States)

    Garcia, I E; Lima, M R; Marinho, C R; Kipnis, T L; Furtado, G C; Alvarez, J M

    1997-04-01

    We have recently described that Trypanosoma cruzi parasites of the reticulotropic Y strain increase their resistance to antibody-induced clearance during their interaction with the vertebrate host immune system. In the present study, we observed that trypomastigotes of the myotropic CL strain isolated from normal host also display an increased resistance to immune clearance when compared to parasites obtained from immunosuppressed donors. Through fluorescence-activated cell sorting analysis, we have observed that the high expression of membrane-bound IgM antibodies on Y and CL trypomastigotes correlates with their enhanced resistance to Ig-induced clearance. Trypomastigotes from normal mice were essentially refractory to the in vitro binding of immunoglobulins, showing that their membrane structures were completely covered by IgM antibodies. These findings suggest that this isotype does not efficiently mediate immune clearance. Moreover, membrane-bound IgM antibodies limited the amount of IgG attached to the parasite and, as a consequence, impaired efficient immune clearance. Through this mechanism, trypomastigotes of T. cruzi could increase their persistence in the bloodstream thus favoring parasite transmission to its hematophagous host vector in the early acute phase of the disease.

  1. Trypanosoma vivax Adhesion to Red Blood Cells in Experimentally Infected Sheep

    Science.gov (United States)

    Boada-Sucre, Alpidio A.; Rossi Spadafora, Marcello Salvatore; Tavares-Marques, Lucinda M.; Finol, Héctor J.; Reyna-Bello, Armando

    2016-01-01

    Trypanosomosis, a globally occurring parasitic disease, poses as a major obstacle to livestock production in tropical and subtropical regions resulting in tangible economic losses. In Latin America including Venezuela, trypanosomosis of ruminants is mainly caused by Trypanosoma vivax. Biologically active substances produced from trypanosomes, as well as host-trypanosome cellular interactions, contribute to the pathogenesis of anemia in an infection. The aim of this study was to examine with a scanning electron microscope the cellular interactions and alterations in ovine red blood cells (RBC) experimentally infected with T. vivax. Ovine infection resulted in changes of RBC shape as well as the formation of surface holes or vesicles. A frequent observation was the adhesion to the ovine RBC by the trypanosome's free flagellum, cell body, or attached flagellum in a process mediated by the filopodia emission from the trypanosome surface. The observed RBC alterations are caused by mechanical and biochemical damage from host-parasite interactions occurring in the bloodstream. The altered erythrocytes are prone to mononuclear phagocytic removal contributing to the hematocrit decrease during infection. PMID:27293960

  2. Advancing Trypanosoma brucei genome annotation through ribosome profiling and spliced leader mapping.

    Science.gov (United States)

    Parsons, Marilyn; Ramasamy, Gowthaman; Vasconcelos, Elton J R; Jensen, Bryan C; Myler, Peter J

    2015-08-01

    Since the initial publication of the trypanosomatid genomes, curation has been ongoing. Here we make use of existing Trypanosoma brucei ribosome profiling data to provide evidence of ribosome occupancy (and likely translation) of mRNAs from 225 currently unannotated coding sequences (CDSs). A small number of these putative genes correspond to extra copies of previously annotated genes, but 85% are novel. The median size of these novels CDSs is small (81 aa), indicating that past annotation work has excelled at detecting large CDSs. Of the unique CDSs confirmed here, over half have candidate orthologues in other trypanosomatid genomes, most of which were not yet annotated as protein-coding genes. Nonetheless, approximately one-third of the new CDSs were found only in T. brucei subspecies. Using ribosome footprints, RNA-Seq and spliced leader mapping data, we updated previous work to definitively revise the start sites for 414 CDSs as compared to the current gene models. The data pointed to several regions of the genome that had sequence errors that altered coding region boundaries. Finally, we consolidated this data with our previous work to propose elimination of 683 putative genes as protein-coding and arrive at a view of the translatome of slender bloodstream and procyclic culture form T. brucei.

  3. Insight into the exoproteome of the tissue-derived trypomastigote form of Trypanosoma cruzi

    Science.gov (United States)

    Queiroz, Rayner; Ricart, Carlos; Machado, Mara; Bastos, Izabela; Santana, Jaime; Sousa, Marcelo; Roepstorff, Peter; Charneau, Sébastien

    2016-11-01

    The protozoan parasite Trypanosoma cruzi causes Chagas disease, one of the major neglected infectious diseases. It has the potential to infect any nucleated mammalian cell. The secreted/excreted protein repertoire released by T. cruzi trypomastigotes is crucial in host-pathogen interactions. In this study, mammalian tissue culture-derived trypomastigotes (Y strain) were used to characterize the exoproteome of the infective bloodstream life form. Proteins released into the serum-free culture medium after 3h of incubation were harvested and digested with trypsin. NanoLC-MS/MS analysis resulted in the identification of 540 proteins, the largest set of released proteins identified to date in Trypanosome spp. Bioinformatic analysis predicted most identified proteins as secreted, predominantly by non-classical pathways, and involved in host-cell infection. Some proteins possess predicted GPI-anchor signals, these being mostly trans-sialidases, mucin associated surface proteins and surface glycoproteins. Moreover, we enriched phosphopeptides and glycopeptides from tryptic digests. The majority of identified glycoproteins are trans-sialidases and surface glycoproteins involved in host-parasite interaction. Conversely, most identified phosphoproteins have no Gene Ontology classification. The existence of various proteins related to similar functions in the exoproteome likely reflects this parasite’s enhanced mechanisms for adhesion, invasion and internalization of different host-cell types, and escape from immune defences.

  4. Fluorinated Sterols Are Suicide Inhibitors of Ergosterol Biosynthesis and Growth in Trypanosoma brucei.

    Science.gov (United States)

    Leaver, David J; Patkar, Presheet; Singha, Ujjal K; Miller, Matthew B; Haubrich, Brad A; Chaudhuri, Minu; Nes, W David

    2015-10-22

    Trypanosoma brucei, the causal agent for sleeping sickness, depends on ergosterol for growth. Here, we describe the effects of a mechanism-based inhibitor, 26-fluorolanosterol (26FL), which converts in vivo to a fluorinated substrate of the sterol C24-methyltransferase essential for sterol methylation and function of ergosterol, and missing from the human host. 26FL showed potent inhibition of ergosterol biosynthesis and growth of procyclic and bloodstream forms while having no effect on cholesterol biosynthesis or growth of human epithelial kidney cells. During exposure of cloned TbSMT to 26-fluorocholesta-5,7,24-trienol, the enzyme is gradually killed as a consequence of the covalent binding of the intermediate C25 cation to the active site (kcat/kinact = 0.26 min(-1)/0.24 min(-1); partition ratio of 1.08), whereas 26FL is non-productively bound. These results demonstrate that poisoning of ergosterol biosynthesis by a 26-fluorinated Δ(24)-sterol is a promising strategy for developing a new treatment for trypanosomiasis.

  5. Synchronous expression of individual metacyclic variant surface glycoprotein genes in Trypanosoma brucei.

    Science.gov (United States)

    Ramey-Butler, Kiantra; Ullu, Elisabetta; Kolev, Nikolay G; Tschudi, Christian

    2015-01-01

    One distinctive feature of the Trypanosoma brucei life cycle is the presence of two discrete populations that are based on differential expression of variant surface glycoproteins (VSGs). Both are adapted to the environmental pressures they face and more importantly, both contribute directly to transmission. Metacyclics in the tsetse fly enable transmission to a new mammalian host, whereas bloodstream trypanosomes must avoid immune destruction to the extent that sufficient numbers are available for transmission, when the insect vector takes a blood meal. At present, there are few investigations on the molecular aspects of parasite biology in the tsetse vector and specifically about the activation of metacyclic VSG gene expression. Here we used an established in vitro differentiation system based on the overexpression of the RNA-binding protein 6 (RBP6), to monitor two metacyclic VSGs (VSG 397 and VSG 653) during development from procyclics to infectious metacyclic forms. We observed that activation of these two mVSGs was simultaneous both at the transcript and protein level, and manifested by the appearance of only one of the mVSGs in individual cells.

  6. Identification of a new EF-hand superfamily member from Trypanosoma brucei

    Science.gov (United States)

    Wong, S.; Kretsinger, R. H.; Campbell, D. A.

    1992-01-01

    We identified several open reading frames between the regions encoding calmodulin and ubiquitin-EP52/1 in the genome of Trypanosoma brucei. One of these, EFH5, encodes a protein 192 amino acids long. The EFH5 transcript is present in poly(A)+ mRNA and is present at similar levels in the mammalian bloodstream form and the insect procyclic form. EFH5 contains four EF-hand homolog domains, two of which are inferred to bind Ca2+ ions. We expressed EFH5 as a fusion protein in Escherichia coli and demonstrated calcium-binding activity of the fusion protein using the 45Ca-overlay technique. The function of EFH5 remains unknown; however, as the fourth EF-hand homolog identified in trypanosomes, it attests to the broad range of functions assumed by calcium functioning as a second messenger. EFH5, which is most closely related to LAV1-2 from Physarum, represents a distinct subfamily among the EF-hand-containing proteins.

  7. Life cycle of Trypanosoma cruzi (y strain in mice

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    Pinto Pedro Luiz Silva

    1999-01-01

    Full Text Available Since 1958, we have studied experimental Chagas' disease (CD by subcutaneous inoculation of 1,000 blood forms of Trypanosoma cruzi (Y strain in Balb/C. mice. Evolution of parasitemia remained constant, beginning on the 5th and 6th day of the disease, increasing progressively, achieving a maximum on about the 30th day. After another month, only a few forms were present, and they disappeared from the circulation after the third month, as determined from direct examination of slides and the use of a Neubauer Counting Chamber. These events coincided with the appearance of amastigote nests in the tissues (especially the cardiac ones, starting the first week, and following the Gauss parasitemia curve, but they were not in parallel until the chronic stage. In 1997, we began to note the following changes: Parasites appeared in the circulation during the first week and disappeared starting on the 7th day, and there was a coincident absence of the amastigote nests in the tissues. A careful study verified that young forms in the evolutionary cycle of T. cruzi (epi + amastigotes began to appear alongside the trypomastigotes in the circulation on the 5th and 7th post-inoculation day. At the same time, rounded, oval, and spindle shapes were seen circulating through the capillaries and sinusoids of the tissues, principally of the hematopoietic organs. Stasis occurs because the diameter of the circulating parasites is greater than the vessels, and this makes them more visible. Examination of the sternal bone marrow revealed young cells with elongated forms and others truncated in the shape of a "C" occupying the internal surface of the blood cells that had empty central portions (erythrocytes?. We hypothesize that there could be a loss of virulence or mutation of the Y strain of Trypanosoma cruzi.

  8. Bacterial bloodstream infections in liver transplantation: etiologic agents and antimicrobial susceptibility profiles.

    Science.gov (United States)

    Sganga, G; Spanu, T; Bianco, G; Fiori, B; Nure, E; Pepe, G; D'inzeo, T; Lirosi, M C; Frongillo, F; Agnes, S

    2012-09-01

    Liver transplantation (OLT) is a lifesaving procedure for the treatment of many end-stage liver diseases, but infection and acute rejection episodes still remain the main causes of morbidity and mortality. Bloodstream infections (BSIs), particularly, are the major cause of mortality among these patients. BSIs in OLT, are from intra-abdominal, biliary, respiratory, urinary, wound and/or central venous catheter sources. A certain percentage are of unknown origin. Using the computerized database of our microbiology laboratory, we analyzed all BSIs in 75 consecutive adult liver transplant patients in a single center between January 2008 and July 2011. BSIs occurred in 21/75 (28%) patients. Thirteen subjects had a single; two, two episodes, and the other six patients each >4 episodes. All episodes occurred in the first 60 days following OLT; the majority (74%), in the first month. Among 44 microorganisms recovered, 52.3% were gram-negative, the most frequent being Pseudomonas aeruginosa and Klebsiella pneumoniae; 47.7% were gram-positive, the most frequent being coagulase-negative staphylococci, particularly Staphylococcus epidermidis. Overall 65.9% of the isolates were resistant to several antibiotics: 40.9% displayed the multiding-resistant and 25% the panding-resistant phenotype. There was a high incidence of gram-negative and most importantly, resistant bacteria, which required appropriate therapy. These data showed that it is imperative to promote strategies to prevention and contain antimicrobial resistance.

  9. ORC1/CDC6 and MCM7 distinct associate with chromatin through Trypanosoma cruzi life cycle.

    Science.gov (United States)

    Calderano, Simone; Godoy, Patricia; Soares, Daiane; Sant'Anna, Osvaldo Augusto; Schenkman, Sergio; Elias, M Carolina

    2014-02-01

    Trypanosoma cruzi alternates between replicative and non-replicative stages. We analyzed the expression of components of the pre-replication machinery TcORC1/CDC6 and TcMCM7 and their interaction with DNA in all T. cruzi stages. TcORC1/CDC6 remains in the nuclear space during all stages of the life cycle and interacts with DNA in the replicative stages; however, it does not bind to DNA in the non-replicative forms. Moreover, TcMCM7 is not present in the non-replicative stages. These data suggest that the lacking of DNA replication during the T. cruzi life cycle may be a consequence of the blocking of TcORC1/CDC6-DNA interaction and of the down regulation of the TcMCM7 expression.

  10. Catheter Removal versus Retention in the Management of Catheter-Associated Enterococcal Bloodstream Infections

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    Jonas Marschall

    2013-01-01

    Full Text Available BACKGROUND: Enterococci are an important cause of central venous catheter (CVC-associated bloodstream infections (CA-BSI. It is unclear whether CVC removal is necessary to successfully manage enterococcal CA-BSI.

  11. 布氏锥虫葡糖基磷脂酰肌醇(GPI)的研究进展%RESEARCH PROSPECTS ON GLYCOSYLPHOSPHATIDYLINOSITOL (GPI) OF TRYPANOSOMA BRUCEI

    Institute of Scientific and Technical Information of China (English)

    原丽红; 林本夫; 王祥生; 李莲瑞

    2004-01-01

    布氏锥虫(Trypanosoma brucei)是一种原虫性寄生虫,通过采采蝇(tsets fly)的传播感染人和其它哺乳动物,导致人的昏睡病和家畜的那卡那病。布氏锥虫有两个截然不同的生活阶段,即血液期(bloodstream stage,寄生于哺乳动物血细胞内)和昆虫期(insect stage,又叫循环期或感染期,寄生于采采蝇的中肠内)。

  12. Trypanosoma rangeli (Tejera, 1920: observations upon pleomorphism

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    S. Urdaneta-Morales

    1992-12-01

    Full Text Available Metatrypomastigotes of Trypanosoma rangeli Tejera, 1920, harvested from LIT medium, were inoculated i.p. or s.c. into 6, 16, and 26g NMRI mice, these representing increasing degrees of immunological maturity. In all cases, similar pleomorphic patterns were observed. Four morphobiometrically differentiable types of trypanosome were encountered in an overlapping temporal sequence. These observations, taken in comparison with those on pleomorphism in this and other species of Trypanosoma by other workers, are consistent with the hypothesis that the pleomorphic types represent the natural development of the parasite, rather than the result of the immune response of the mammal host. Small, slender trypanosomes prevalent at the onset of the parasitemia either reinvade the tissue cells for relatively limited subsequent generations of tissue reproduction, or else differentiate toward the forms that are only capable of colonizing the insect vector.

  13. Transporter protein and drug resistance of Trypanosoma.

    Science.gov (United States)

    Medina, Noraine P; Mingala, Claro N

    2016-01-01

    Trypanosoma infection is one of the most important infections in livestock and humans. One of the main problems of its therapeutic control and treatment is the resurgence of drug resistance. One of the most studied causes of such resistance is the function of its adenosine transporter gene. A trypanosomal gene TbAT1 from Trypanosoma brucei has been cloned in yeast to demonstrate its function in the transport of adenosine and trypanocidal agents. Drug resistant trypanosomes showed a defective TbAT1 variant; furthermore, deletion of the gene and set point mutations in the transporter gene has been demonstrated from isolates from relapse patients. The molecular understanding of the mechanism of action trypanocidal agents and function of transporter gene can lead to control of drug resistance of Trypanosomes.

  14. Bacillus Cereus Catheter Related Bloodstream Infection in a Patient with Acute Lymphoblastic Leukemia

    OpenAIRE

    N Gurler; Oksuz, L; M Muftuoglu; Sargin, FD; Besisik, SK

    2012-01-01

    Bacillus cereus infection is rarely associated with actual infection and for this reason single positive blood culture is usually regarded as contamination . However it may cause a number of infections, such catheter-related bloodstream infections. Significant catheter-related bloodstream infections (CRBSI) caused by Bacillus spp. are mainly due to B. cereus and have been predominantly reported in immunocompromised hosts. Catheter removal is generally advised for management of infection. In t...

  15. Trypanosoma cruzi Infection and Host Lipid Metabolism

    OpenAIRE

    2014-01-01

    Trypanosoma cruzi is the causative agent of Chagas disease. Approximately 8 million people are thought to be affected worldwide. Several players in host lipid metabolism have been implicated in T. cruzi-host interactions in recent research, including macrophages, adipocytes, low density lipoprotein (LDL), low density lipoprotein receptor (LDLR), and high density lipoprotein (HDL). All of these factors are required to maintain host lipid homeostasis and are intricately connected via several me...

  16. Elimination of central line-associated bloodstream infections: application of the evidence.

    Science.gov (United States)

    Posa, Patricia J; Harrison, Denise; Vollman, Kathleen M

    2006-01-01

    Central line-associated bloodstream infections are considered to be an avoidable complication of care delivery. In addition to considerable morbidity and use of resources, central line-associated bloodstream infections carry an attributable morality between 12% and 25%. The estimated cost per infection is approximately 25,000 US dollars. Research over the last decade has focused on a number of care activities that have been shown to reduce the incidence of bloodstream infections related to central line placement in the critically ill patient. A significant reduction or elimination of central line-associated bloodstream infections can occur with implementation of a comprehensive central line-associated bloodstream infection prevention program that includes staff education, hand hygiene, use of maximal sterile barrier precautions, chlorhexidine gluconate skin antisepsis, avoidance of femoral lines, empowerment of staff to stop the procedure if sterile technique is broken, and daily assessment of the continued need for a central line. This article focuses on strategies for implementing a comprehensive central line-associated bloodstream infections prevention program and a tool and process for defect analysis as part of a statewide collaborative in Michigan.

  17. Epidemiology, surveillance, and prevention of bloodstream infections in hemodialysis patients.

    Science.gov (United States)

    Patel, Priti R; Kallen, Alexander J; Arduino, Matthew J

    2010-09-01

    Infections cause significant morbidity and mortality in patients undergoing hemodialysis. Bloodstream infections (BSIs) are particularly problematic, accounting for a substantial number of hospitalizations in these patients. Hospitalizations for BSI and other vascular access infections appear to have increased dramatically in hemodialysis patients since 1993. These infections frequently are related to central venous catheter (CVC) use for dialysis access. Regional initiatives that have shown successful decreases in catheter-related BSIs in hospitalized patients have generated interest in replicating this success in outpatient hemodialysis populations. Several interventions have been effective in preventing BSIs in the hemodialysis setting. Avoiding the use of CVCs in favor of access types with lower associated BSI risk is among the most important. When CVCs are used, adherence to evidence-based catheter insertion and maintenance practices can positively influence BSI rates. In addition, facility-level surveillance to detect BSIs and stimulate examination of vascular access use and care practices is essential to a comprehensive approach to prevention. This article describes the current epidemiology of BSIs in hemodialysis patients and effective prevention strategies to decrease the incidence of these devastating infections.

  18. Bloodstream Infections in a Neonatal Intensive Care Unit

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    Mehmet Sah Ižpek

    2016-09-01

    Full Text Available Aim: To determine the pattern of bloodstream infections (BSIs and antimicrobial susceptibility of pathogens in a neonatal intensive care unit (NICU.Material and Method: Positive hemoculture of neonates diagnosed with nosocomial sepsis from March 2011 to March 2014 in the NICU of Diyarbakir Maternity and Children%u2019s Hospital, in the southeastern region of Anatolia, Turkey, were retrospectively reviewed. Results: A total of 148 pathogens were isolated in 142 neonates. The most common microorganisms isolated were Klebsiella pneumoniae (40.5% and Acinetobacter baumannii (29.7% which was a result of a hospital outbreak. Multi-drug resistant (MDR strains accounted for 20.0% of K. pneumoniae isolates and 93.2% of A. baumannii isolates. The sepsis-attributable mortality rate was higher in cases infected with MDR strains than in cases infected without MDR strains or Candida spp (24% vs. 9.7%, p=0.032. Discussion: In our unit, BSIs were more often caused by Gram negative bacteria. BSIs caused by MDR strains were associated with a higher rate of sepsis-attributable mortality.

  19. Clinical forms of Trypanosoma cruzi infected individuals in the chronic phase of Chagas disease in Puebla, Mexico

    OpenAIRE

    2006-01-01

    In Mexico, despite the relatively high seroprevalence of Trypanosoma cruzi infection in humans in some areas, reported morbidity of Chagas disease is not clear. We determined clinical stage in 71 individuals seropositive to T. cruzi in the state of Puebla, Mexico, an area endemic for Chagas disease with a reported seroprevalence of 7.7%. Diagnosis of Chagas disease was made by two standardized serological tests (ELISA, IHA). Individuals were stratified according to clinical studies. All patie...

  20. Deviating the level of proliferating cell nuclear antigen in Trypanosoma brucei elicits distinct mechanisms for inhibiting proliferation and cell cycle progression.

    Science.gov (United States)

    Valenciano, Ana L; Ramsey, Aaron C; Mackey, Zachary B

    2015-01-01

    The DNA replication machinery is spatially and temporally coordinated in all cells to reproduce a single exact copy of the genome per division, but its regulation in the protozoan parasite Trypanosoma brucei is not well characterized. We characterized the effects of altering the levels of proliferating cell nuclear antigen, a key component of the DNA replication machinery, in bloodstream form T. brucei. This study demonstrated that tight regulation of TbPCNA levels was critical for normal proliferation and DNA replication in the parasite. Depleting TbPCNA mRNA reduced proliferation, severely diminished DNA replication, arrested the synthesis of new DNA and caused the parasites to accumulated in G2/M. Attenuating the parasite by downregulating TbPCNA caused it to become hypersensitive to hydroxyurea. Overexpressing TbPCNA in T. brucei arrested proliferation, inhibited DNA replication and prevented the parasite from exiting G2/M. These results indicate that distinct mechanisms of cell cycle arrest are associated with upregulating or downregulating TbPCNA. The findings of this study validate deregulating intra-parasite levels of TbPCNA as a potential strategy for therapeutically exploiting this target in bloodstream form T. brucei.

  1. In vitro cytocidal effect of novel lytic peptides on Plasmodium falciparum and Trypanosoma cruzi.

    Science.gov (United States)

    Jaynes, J M; Burton, C A; Barr, S B; Jeffers, G W; Julian, G R; White, K L; Enright, F M; Klei, T R; Laine, R A

    1988-10-01

    Plasmodium falciparum and Trypanosoma cruzi were killed by two novel lytic peptides (SB-37 and Shiva-1) in vitro. Human erythrocytes infected with P. falciparum, and Vero cells infected with T. cruzi, were exposed to these peptides. The result, in both cases, was a significant decrease in the level of parasite infection. Furthermore, the peptides had a marked cytocidal effect on trypomastigote stages of T. cruzi in media, whereas host eukaryotic cells were unaffected by the treatments. In view of the worldwide prevalence of these protozoan diseases and the lack of completely suitable treatments, lytic peptides may provide new and unique chemotherapeutic agents for the treatment of these infections.

  2. Drug discovery for Chagas disease should consider Trypanosoma cruzi strain diversity

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    Bianca Zingales

    2014-09-01

    Full Text Available This opinion piece presents an approach to standardisation of an important aspect of Chagas disease drug discovery and development: selecting Trypanosoma cruzi strains for in vitro screening. We discuss the rationale for strain selection representing T. cruzi diversity and provide recommendations on the preferred parasite stage for drug discovery, T. cruzi discrete typing units to include in the panel of strains and the number of strains/clones for primary screens and lead compounds. We also consider experimental approaches for in vitro drug assays. The Figure illustrates the current Chagas disease drug-discovery and development landscape.

  3. The MASP family of Trypanosoma cruzi: changes in gene expression and antigenic profile during the acute phase of experimental infection.

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    Sara Lopes dos Santos

    Full Text Available BACKGROUND: Trypanosoma cruzi is the etiological agent of Chagas disease, a debilitating illness that affects millions of people in the Americas. A major finding of the T. cruzi genome project was the discovery of a novel multigene family composed of approximately 1,300 genes that encode mucin-associated surface proteins (MASPs. The high level of polymorphism of the MASP family associated with its localization at the surface of infective forms of the parasite suggests that MASP participates in host-parasite interactions. We speculate that the large repertoire of MASP sequences may contribute to the ability of T. cruzi to infect several host cell types and/or participate in host immune evasion mechanisms. METHODS: By sequencing seven cDNA libraries, we analyzed the MASP expression profile in trypomastigotes derived from distinct host cells and after sequential passages in acutely infected mice. Additionally, to investigate the MASP antigenic profile, we performed B-cell epitope prediction on MASP proteins and designed a MASP-specific peptide array with 110 putative epitopes, which was screened with sera from acutely infected mice. FINDINGS AND CONCLUSIONS: We observed differential expression of a few MASP genes between trypomastigotes derived from epithelial and myoblast cell lines. The more pronounced MASP expression changes were observed between bloodstream and tissue-culture trypomastigotes and between bloodstream forms from sequential passages in acutely infected mice. Moreover, we demonstrated that different MASP members were expressed during the acute T. cruzi infection and constitute parasite antigens that are recognized by IgG and IgM antibodies. We also found that distinct MASP peptides could trigger different antibody responses and that the antibody level against a given peptide may vary after sequential passages in mice. We speculate that changes in the large repertoire of MASP antigenic peptides during an infection may contribute to

  4. Expanding the tool box for genetic manipulation of Trypanosoma cruzi.

    Science.gov (United States)

    Burle-Caldas, Gabriela de Assis; Grazielle-Silva, Viviane; Laibida, Letícia Adejani; DaRocha, Wanderson Duarte; Teixeira, Santuza Maria Ribeiro

    2015-01-01

    Trypanosoma cruzi is a protozoan parasite that causes Chagas disease, an illness that affects 6-7 million people and for which there is no effective drug therapy or vaccine. The publication of its complete genome sequence allowed a rapid advance in molecular studies including in silico screening of genes involved with pathogenicity as well as molecular targets for the development of new diagnostic methods, drug therapies and prophylactic vaccines. Alongside with in silico genomic analyses, methods to study gene function in this parasite such as gene deletion, overexpression, mutant complementation and reporter gene expression have been largely explored. More recently, the use of genome-wide strategies is producing a shift towards a global perspective on gene function studies, with the examination of the expression and biological roles of gene networks in different stages of the parasite life cycle and under different contexts of host parasite interactions. Here we describe the molecular tools and protocols currently available to perform genetic manipulation of the T. cruzi genome, with emphasis on recently described strategies of gene editing that will facilitate large-scale functional genomic analyses. These new methodologies are long overdue, since more efficient protocols for genetic manipulation in T. cruzi are urgently needed for a better understanding of the biology of this parasite and molecular processes involved with the complex and often harmful, interaction with its human host.

  5. Conservation and divergence within the clathrin interactome of Trypanosoma cruzi.

    Science.gov (United States)

    Kalb, Ligia Cristina; Frederico, Yohana Camila A; Boehm, Cordula; Moreira, Claudia Maria do Nascimento; Soares, Maurilio José; Field, Mark C

    2016-08-09

    Trypanosomatids are parasitic protozoa with a significant burden on human health. African and American trypanosomes are causative agents of Nagana and Chagas disease respectively, and speciated about 300 million years ago. These parasites have highly distinct life cycles, pathologies, transmission strategies and surface proteomes, being dominated by the variant surface glycoprotein (African) or mucins (American) respectively. In African trypanosomes clathrin-mediated trafficking is responsible for endocytosis and post-Golgi transport, with several mechanistic aspects distinct from higher organisms. Using clathrin light chain (TcCLC) and EpsinR (TcEpsinR) as affinity handles, we identified candidate clathrin-associated proteins (CAPs) in Trypanosoma cruzi; the cohort includes orthologs of many proteins known to mediate vesicle trafficking, but significantly not the AP-2 adaptor complex. Several trypanosome-specific proteins common with African trypanosomes, were also identified. Fluorescence microscopy revealed localisations for TcEpsinR, TcCLC and TcCHC at the posterior region of trypomastigote cells, coincident with the flagellar pocket and Golgi apparatus. These data provide the first systematic analysis of clathrin-mediated trafficking in T. cruzi, allowing comparison between protein cohorts and other trypanosomes and also suggest that clathrin trafficking in at least some life stages of T. cruzi may be AP-2-independent.

  6. Production of amastigotes from metacyclic trypomastigotes of Trypanosoma cruzi

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    Víctor T Contreras

    2002-12-01

    Full Text Available Attempts to recreate all the developmental stages of Trypanosoma cruzi in vitro have thus far been met with partial success. It is possible, for instance, to produce trypomastigotes in tissue culture and to obtain metacyclic trypomastigotes in axenic conditions. Even though T. cruzi amastigotes are known to differentiate from trypomastigotes and metacyclic trypomastigotes, it has only been possible to generate amastigotes in vitro from the tissue-culture-derived trypomastigotes. The factors and culture conditions required to trigger the transformation of metacyclic trypomastigotes into amastigotes are as yet undetermined. We show here that pre-incubation of metacyclic trypomastigotes in culture (MEMTAU medium at 37°C for 48 h is sufficient to commit the parasites to the transformation process. After 72 h of incubation in fresh MEMTAU medium, 90% of the metacyclic parasites differentiate into forms that are morphologically indistinguishable from normal amastigotes. SDS-PAGE, Western blot and PAABS analyses indicate that the transformation of axenic metacyclic trypomastigotes to amastigotes is associated with protein, glycoprotein and antigenic modifications. These data suggest that (a T. cruzi amastigotes can be obtained axenically in large amounts from metacyclic trypomastigotes, and (b the amastigotes thus obtained are morphological, biological and antigenically similar to intracellular amastigotes. Consequently, this experimental system may facilitate a direct, in vitro assessment of the mechanisms that enable T. cruzi metacyclic trypomastigotes to transform into amastigotes in the cells of mammalian hosts.

  7. High throughput screening for anti-Trypanosoma cruzi drug discovery.

    Science.gov (United States)

    Alonso-Padilla, Julio; Rodríguez, Ana

    2014-12-01

    The discovery of new therapeutic options against Trypanosoma cruzi, the causative agent of Chagas disease, stands as a fundamental need. Currently, there are only two drugs available to treat this neglected disease, which represents a major public health problem in Latin America. Both available therapies, benznidazole and nifurtimox, have significant toxic side effects and their efficacy against the life-threatening symptomatic chronic stage of the disease is variable. Thus, there is an urgent need for new, improved anti-T. cruzi drugs. With the objective to reliably accelerate the drug discovery process against Chagas disease, several advances have been made in the last few years. Availability of engineered reporter gene expressing parasites triggered the development of phenotypic in vitro assays suitable for high throughput screening (HTS) as well as the establishment of new in vivo protocols that allow faster experimental outcomes. Recently, automated high content microscopy approaches have also been used to identify new parasitic inhibitors. These in vitro and in vivo early drug discovery approaches, which hopefully will contribute to bring better anti-T. cruzi drug entities in the near future, are reviewed here.

  8. High throughput screening for anti-Trypanosoma cruzi drug discovery.

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    Julio Alonso-Padilla

    2014-12-01

    Full Text Available The discovery of new therapeutic options against Trypanosoma cruzi, the causative agent of Chagas disease, stands as a fundamental need. Currently, there are only two drugs available to treat this neglected disease, which represents a major public health problem in Latin America. Both available therapies, benznidazole and nifurtimox, have significant toxic side effects and their efficacy against the life-threatening symptomatic chronic stage of the disease is variable. Thus, there is an urgent need for new, improved anti-T. cruzi drugs. With the objective to reliably accelerate the drug discovery process against Chagas disease, several advances have been made in the last few years. Availability of engineered reporter gene expressing parasites triggered the development of phenotypic in vitro assays suitable for high throughput screening (HTS as well as the establishment of new in vivo protocols that allow faster experimental outcomes. Recently, automated high content microscopy approaches have also been used to identify new parasitic inhibitors. These in vitro and in vivo early drug discovery approaches, which hopefully will contribute to bring better anti-T. cruzi drug entities in the near future, are reviewed here.

  9. Immune Evasion Strategies of Trypanosoma cruzi

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    Ana Flávia Nardy

    2015-01-01

    Full Text Available Microbes have evolved a diverse range of strategies to subvert the host immune system. The protozoan parasite Trypanosoma cruzi, the causative agent of Chagas disease, provides a good example of such adaptations. This parasite targets a broad spectrum of host tissues including both peripheral and central lymphoid tissues. Rapid colonization of the host gives rise to a systemic acute response which the parasite must overcome. The parasite in fact undermines both innate and adaptive immunity. It interferes with the antigen presenting function of dendritic cells via an action on host sialic acid-binding Ig-like lectin receptors. These receptors also induce suppression of CD4+ T cells responses, and we presented evidence that the sialylation of parasite-derived mucins is required for the inhibitory effects on CD4 T cells. In this review we highlight the major mechanisms used by Trypanosoma cruzi to overcome host immunity and discuss the role of parasite colonization of the central thymic lymphoid tissue in chronic disease.

  10. Impact of benznidazole on infection course in mice experimentally infected with Trypanosoma cruzi I, II, and IV.

    Science.gov (United States)

    Gruendling, Ana Paula; Massago, Miyoko; Teston, Ana Paula M; Monteiro, Wuelton M; Kaneshima, Edilson N; Araújo, Silvana M; Gomes, Mônica L; Barbosa, Maria das Graças V; Toledo, Max Jean O

    2015-06-01

    American trypanosomiasis is an emerging zoonosis in the Brazilian Amazon. Studies on benznidazole (BZ) chemotherapy with Trypanosoma cruzi from this region have great relevance, given the different discrete typing units (DTUs) that infect humans in the Amazon and other regions of Brazil. We performed a parasitological, histopathological, and molecular analysis of mice inoculated with strains of T. cruzi I, II, and IV that were BZ-treated during the acute phase of infection. Groups of Swiss mice were inoculated; 13 received oral BZ, whereas the other 13 comprised the untreated controls. Unlike parasitemia, the infectivity and mortality did not vary among the DTUs. Trypanosoma cruzi DNA was detected in all tissues analyzed and the proportion of organs parasitized varied with the parasite DTU. The BZ treatment reduced the most parasitological parameters, tissue parasitism and the inflammatory processes at all infection stages and for all DTUs. However, the number of significant reductions varied according to the DTU and infection phase.

  11. Candida bloodstream infections in intensive care units: analysis of the extended prevalence of infection in intensive care unit study

    NARCIS (Netherlands)

    Kett, D.H.; Azoulay, E.; Echeverria, P.M.; Vincent, J.L.; Pickkers, P.

    2011-01-01

    OBJECTIVES: To provide a global, up-to-date picture of the prevalence, treatment, and outcomes of Candida bloodstream infections in intensive care unit patients and compare Candida with bacterial bloodstream infection. DESIGN: A retrospective analysis of the Extended Prevalence of Infection in the I

  12. Vaccination with Trypanosoma rangeli induces resistance of guinea pigs to virulent Trypanosoma cruzi.

    Science.gov (United States)

    Basso, B; Moretti, E; Fretes, R

    2014-01-15

    Chagas' disease, endemic in Latin America, is spread in natural environments through animal reservoirs, including marsupials, mice and guinea pigs. Farms breeding guinea pigs for food are located in some Latin-American countries with consequent risk of digestive infection. The aim of this work was to study the effect of vaccination with Trypanosoma rangeli in guinea pigs challenged with Trypanosoma cruzi. Animals were vaccinated with fixated epimastigotes of T. rangeli, emulsified with saponin. Controls received only PBS. Before being challenged with T. cruzi, parasitemia, survival rates and histological studies were performed. The vaccinated guinea pigs revealed significantly lower parasitemia than controls (pguinea pigs and dogs. The development of vaccines for use in animals, like domestic dogs and guinea pigs in captivity, opens up new opportunities for preventive tools, and could reduce the risk of infection with T. cruzi in the community.

  13. Bacillus cereus catheter related bloodstream infection in a patient with acute lymphoblastic leukemia.

    Science.gov (United States)

    Gurler, N; Oksuz, L; Muftuoglu, M; Sargin, Fd; Besisik, Sk

    2012-01-01

    Bacillus cereus infection is rarely associated with actual infection and for this reason single positive blood culture is usually regarded as contamination . However it may cause a number of infections, such catheter-related bloodstream infections. Significant catheter-related bloodstream infections (CRBSI) caused by Bacillus spp. are mainly due to B. cereus and have been predominantly reported in immunocompromised hosts. Catheter removal is generally advised for management of infection. In this report, catheter-related bacteremia caused by B. cereus in a patient with acute lymphoblast c leukemia (ALL) in Istanbul Medical Faculty was presented.

  14. Classical clinical signs in rats experimemtally infected with Trypanosoma brucei

    Institute of Scientific and Technical Information of China (English)

    Nwoha Rosemary Ijeoma Ogechi; Omamegbe Joseph Omolathebu

    2015-01-01

    Objective:To investigate clinical signs in Trypanosoma brucei infection in albino rats. Methods:Fourteen rats grouped into 2 with 7 rats in each group were used to determine classical clinical manifestation of Trypanosoma brucei infection in rats. Group A rats were uninfected control and Group B rats were infected with Trypanosoma brucei. Results:Parasitaemia was recorded in Group B by (3.86±0.34) d and the peak of parasitaemia was observed at Day 5 post infection. Classical signs observed included squint eyes, raised whiskers, lethargy, no weight loss, pyrexia, isolation from the other rats, and starry hair coat. Conclusions:These signs could be diagnostic or aid in diagnosis of Trypanosoma brucei infection in rats.

  15. MDL28170, a calpain inhibitor, affects Trypanosoma cruzi metacyclogenesis, ultrastructure and attachment to Rhodnius prolixus midgut.

    Directory of Open Access Journals (Sweden)

    Vítor Ennes-Vidal

    Full Text Available BACKGROUND: Trypanosoma cruzi is the etiological agent of Chagas' disease. During the parasite life cycle, many molecules are involved in the differentiation process and infectivity. Peptidases are relevant for crucial steps of T. cruzi life cycle; as such, it is conceivable that they may participate in the metacyclogenesis and interaction with the invertebrate host. METHODOLOGY/PRINCIPAL FINDINGS: In this paper, we have investigated the effect of the calpain inhibitor MDL28170 on the attachment of T. cruzi epimastigotes to the luminal midgut surface of Rhodnius prolixus, as well as on the metacyclogenesis process and ultrastructure. MDL28170 treatment was capable of significantly reducing the number of bound epimastigotes to the luminal surface midgut of the insect. Once the cross-reactivity of the anti-Dm-calpain was assessed, it was possible to block calpain molecules by the antibody, leading to a significant reduction in the capacity of adhesion to the insect guts by T. cruzi. However, the antibodies were unable to interfere in metacyclogenesis, which was impaired by the calpain inhibitor presenting a significant reduction in the number of metacyclic trypomastigotes. The calpain inhibitor also promoted a direct effect against bloodstream trypomastigotes. Ultrastructural analysis of epimastigotes treated with the calpain inhibitor revealed disorganization in the reservosomes, Golgi and plasma membrane disruption. CONCLUSIONS/SIGNIFICANCE: The presence of calpain and calpain-like molecules in a wide range of organisms suggests that these proteins could be necessary for basic cellular functions. Herein, we demonstrated the effects of MDL28170 in crucial steps of the T. cruzi life cycle, such as attachment to the insect midgut and metacyclogenesis, as well as in parasite viability and morphology. Together with our previous findings, these results help to shed some light on the functions of T. cruzi calpains. Considering the potential roles of

  16. Mitochondrial outer membrane proteome of Trypanosoma brucei reveals novel factors required to maintain mitochondrial morphology.

    Science.gov (United States)

    Niemann, Moritz; Wiese, Sebastian; Mani, Jan; Chanfon, Astrid; Jackson, Christopher; Meisinger, Chris; Warscheid, Bettina; Schneider, André

    2013-02-01

    Trypanosoma brucei is a unicellular parasite that causes devastating diseases in humans and animals. It diverged from most other eukaryotes very early in evolution and, as a consequence, has an unusual mitochondrial biology. Moreover, mitochondrial functions and morphology are highly regulated throughout the life cycle of the parasite. The outer mitochondrial membrane defines the boundary of the organelle. Its properties are therefore key for understanding how the cytosol and mitochondria communicate and how the organelle is integrated into the metabolism of the whole cell. We have purified the mitochondrial outer membrane of T. brucei and characterized its proteome using label-free quantitative mass spectrometry for protein abundance profiling in combination with statistical analysis. Our results show that the trypanosomal outer membrane proteome consists of 82 proteins, two-thirds of which have never been associated with mitochondria before. 40 proteins share homology with proteins of known functions. The function of 42 proteins, 33 of which are specific to trypanosomatids, remains unknown. 11 proteins are essential for the disease-causing bloodstream form of T. brucei and therefore may be exploited as novel drug targets. A comparison with the outer membrane proteome of yeast defines a set of 17 common proteins that are likely present in the mitochondrial outer membrane of all eukaryotes. Known factors involved in the regulation of mitochondrial morphology are virtually absent in T. brucei. Interestingly, RNAi-mediated ablation of three outer membrane proteins of unknown function resulted in a collapse of the network-like mitochondrion of procyclic cells and for the first time identified factors that control mitochondrial shape in T. brucei.

  17. Trypanosoma cruzi: Entry into Mammalian Host Cells and Parasitophorous Vacuole Formation

    Science.gov (United States)

    Barrias, Emile Santos; de Carvalho, Tecia Maria Ulisses; De Souza, Wanderley

    2013-01-01

    Trypanosoma cruzi, the causative agent of Chagas disease, is transmitted to vertebrate hosts by blood-sucking insects. This protozoan is an obligate intracellular parasite. The infective forms of the parasite are the metacyclic trypomastigotes, amastigotes, and bloodstream trypomastigotes. The recognition between the parasite and mammalian host cell, involves numerous molecules present in both cell types, and similar to several intracellular pathogens, T. cruzi is internalized by host cells via multiple endocytic pathways. Morphological studies demonstrated that after the interaction of the infective forms of T. cruzi with phagocytic or non-phagocytic cell types, plasma membrane (PM) protrusions can form, showing similarity with those observed during canonical phagocytosis or macropinocytic events. Additionally, several molecules known to be molecular markers of membrane rafts, macropinocytosis, and phagocytosis have been demonstrated to be present at the invasion site. These events may or may not depend on the host cell lysosomes and cytoskeleton. In addition, after penetration, components of the host endosomal-lysosomal system, such as early endosomes, late endosomes, and lysosomes, participate in the formation of the nascent parasitophorous vacuole (PV). Dynamin, a molecule involved in vesicle formation, has been shown to be involved in the PV release from the host cell PM. This review focuses on the multiple pathways that T. cruzi can use to enter the host cells until complete PV formation. We will describe different endocytic processes, such as phagocytosis, macropinocytosis, and endocytosis using membrane microdomains and clathrin-dependent endocytosis and show results that are consistent with their use by this smart parasite. We will also discuss others mechanisms that have been described, such as active penetration and the process that takes advantage of cell membrane wound repair. PMID:23914186

  18. Trypanosoma cruzi: Entry Into Mammalian Host Cells and Parasitophorous Vacuole Formation

    Directory of Open Access Journals (Sweden)

    Emile Santos Barrias

    2013-08-01

    Full Text Available Trypanosoma cruzi, the causative agent of Chagas disease, is transmitted to vertebrate hosts by blood-sucking insects. This protozoan is an obligate intracellular parasite. The infective forms of the parasite are the metacyclic trypomastigotes, amastigotes and bloodstream trypomastigotes. The recognition between the parasite and mammalian host cell, involves numerous molecules present in both cell types, and similar to several intracellular pathogens, T.cruzi is internalized by host cells via multiple endocytic pathways. Morphological studies demonstrated that after the interaction of the infective forms of T.cruzi with phagocytic or non-phagocytic cell types, plasma membrane protrusions can form, showing similarity with those observed during canonical phagocytosis or macropinocytic events. Additionally, several molecules known to be molecular markers of membrane rafts, macropinocytosis and phagocytosis have been demonstrated to be present at the invasion site. These events may or may not depend on the host cell lysosomes and cytoskeleton. In addition, after penetration, components of the host endosomal-lysosomal system, such as early endosomes, late endosomes and lysosomes, participate in the formation of the nascent parasithophorous vacuole (VP. Dynamin, a molecule involved in vesicle formation, has been shown to be involved in the parasitophorous vacuole release from the host cell plasma membrane. This review focuses on the multiple pathways that T.cruzi can use to enter the host cells until complete VP formation. We will describe different endocytic processes, such as phagocytosis, macropinocytosis, endocytosis using membrane microdomains and clathrin-dependent endocytosis and show results that are consistent with their use by this smart parasite. We will also discuss other mechanisms that have been described, such as active penetration and the process that takes advantage of cell membrane wound repair.

  19. Analysis of the Trypanosoma brucei cell cycle by quantitative DAPI imaging.

    Science.gov (United States)

    Siegel, T Nicolai; Hekstra, Doeke R; Cross, George A M

    2008-08-01

    Trypanosoma brucei has two DNA compartments: the nucleus and the kinetoplast. DNA replication of these two compartments only partially coincides. Woodward and Gull [Woodward R, Gull K. Timing of nuclear and kinetoplast DNA replication and early morphological events in the cell cycle of Trypanosoma brucei. J Cell Sci 1990;95:49-57] comprehensively studied the relative timing of the replication and segregation of nuclear DNA (nDNA) and kinetoplast DNA (kDNA). Others have since assumed the consistency of morphological indicators of cell-cycle stage among strains and conditions. We report the use of quantitative DAPI imaging to determine the cell-cycle stage of individual procyclic cells. Using this approach, we found that kinetoplast elongation occurs mainly during nuclear S phase and not during G2, as previously assumed. We confirmed this finding by sorting cells by DNA content, followed by fluorescence microscopy. In addition, simultaneous quantitative imaging at two wavelengths can be used to determine the abundance of cell-cycle-regulated proteins during the cell cycle. We demonstrate this technique by co-staining for the non-acetylated state of lysine 4 of histone H4 (H4K4), which is enriched during nuclear S phase.

  20. Expression and role of CXCL10 during the encephalitic stage of experimental and clinical African trypanosomiasis

    DEFF Research Database (Denmark)

    Amin, Daniel N; Rottenberg, Martin E; Thomsen, Allan R;

    2009-01-01

    BACKGROUND: Human African trypanosomiasis, caused by Trypanosoma brucei, involves an early hemolymphatic stage followed by a late encephalitic stage. METHODS: We studied the expression of chemokines with use of microarray and enzyme-linked immunosorbent assay in T. brucei brucei-infected mice...

  1. Investigation of the morphological diversity of the potentially zoonotic Trypanosoma copemani in quokkas and Gilbert's potoroos.

    Science.gov (United States)

    Austen, Jill M; Reid, Simon A; Robinson, Derrick R; Friend, James A; Ditcham, William G F; Irwin, Peter J; Ryan, Una

    2015-09-01

    Trypanosomes are blood-borne parasites that can cause severe disease in both humans and animals, yet little is known of the pathogenicity and life-cycles of trypanosomes in native Australian mammals. Trypanosoma copemani is known to be infective to a variety of Australian marsupials and has recently been shown to be potentially zoonotic as it is resistant to normal human serum. In the present study, in vivo and in vitro examination of blood and cultures from Australian marsupials was conducted using light microscopy, immunofluorescence, scanning electron microscopy and fluorescence in situ hybridization. Promastigote, sphaeromastigote and amastigote life-cycle stages were detected in vivo and in vitro. Novel trypanosome-like stages were also detected both in vivo and in vitro representing an oval stage, an extremely thin stage, an adherent stage and a tiny round stage. The tiny round and adherent stages appeared to adhere to erythrocytes causing potential haematological damage with clinical effects similar to haemolytic anaemia. The present study shows for the first time that trypomastigotes are not the only life-cycle stages circulating within the blood stream of trypanosome infected Australian native marsupials and provides insights into possible pathogenic mechanisms of this potentially zoonotic trypanosome species.

  2. The Trypanosoma cruzi nucleic acid binding protein Tc38 presents changes in the intramitochondrial distribution during the cell cycle

    Directory of Open Access Journals (Sweden)

    Nardelli Sheila C

    2009-02-01

    Full Text Available Abstract Background Tc38 of Trypanosoma cruzi has been isolated as a single stranded DNA binding protein with high specificity for the poly [dT-dG] sequence. It is present only in Kinetoplastidae protozoa and its sequence lacks homology to known functional domains. Tc38 orthologues present in Trypanosoma brucei and Leishmania were proposed to participate in quite different cellular processes. To further understand the function of this protein in Trypanosoma cruzi, we examined its in vitro binding to biologically relevant [dT-dG] enriched sequences, its expression and subcellular localization during the cell cycle and through the parasite life stages. Results By using specific antibodies, we found that Tc38 protein from epimastigote extracts participates in complexes with the poly [dT-dG] probe as well as with the universal minicircle sequence (UMS, a related repeated sequence found in maxicircle DNA, and the telomeric repeat. However, we found that Tc38 predominantly localizes into the mitochondrion. Though Tc38 is constitutively expressed through non-replicating and replicating life stages of T. cruzi, its subcellular localization in the unique parasite mitochondrion changes according to the cell cycle stage. In epimastigotes, Tc38 is found only in association with kDNA in G1 phase. From the S to G2 phase the protein localizes in two defined and connected spots flanking the kDNA. These spots disappear in late G2 turning into a diffuse dotted signal which extends beyond the kinetoplast. This later pattern is more evident in mitosis and cytokinesis. Finally, late in cytokinesis Tc38 reacquires its association with the kinetoplast. In non-replicating parasite stages such as trypomastigotes, the protein is found only surrounding the entire kinetoplast structure. Conclusions The dynamics of Tc38 subcellular localization observed during the cell cycle and life stages support a major role for Tc38 related to kDNA replication and maintenance.

  3. Vaccination of dogs with Trypanosoma rangeli induces antibodies against Trypanosoma cruzi in a rural area of Córdoba, Argentina

    OpenAIRE

    2016-01-01

    Dogs play a major role in the domestic cycle of Trypanosoma cruzi, acting as reservoirs. In a previous work we have developed a model of vaccination of dogs in captivity with nonpathogenic Trypanosoma rangeli epimastigotes, resulting in the production of protective antibodies against T. cruzi, with dramatic decrease of parasitaemia upon challenge with 100,000 virulent forms of this parasite. The aim of this work was to evaluate the immunogenicity of this vaccine in dogs living in a rural area...

  4. Trypanosoma cruzi, cancer and the Cold War.

    Science.gov (United States)

    Krementsov, Nikolai

    2009-07-01

    In the summer of 1946, the international community of cancer researchers was inspired by the announcement that two Soviet scientists, Nina Kliueva and Grigorii Roskin, had discovered anticancer properties in culture extracts made from the South American protozoan, Trypanosoma cruzi, and had produced a preparation--named after its discoverers KR--which showed clear therapeutic effects on cancer patients. Research teams from various countries enthusiastically pursued the promising new line of investigation. The story of the rise and fall of interest in the anticancer properties of T. cruzi in different countries suggests that during the second half of the twentieth century, the Cold War competition between the superpowers played an important role in shaping the research agendas of cancer studies.

  5. The haemoculture of Trypanosoma minasense chagas, 1908

    Directory of Open Access Journals (Sweden)

    Mariangela Ziccardi

    1996-08-01

    Full Text Available Trypanosoma minasense was isolated for the first time in blood axenic culture from a naturally infected marmoset, Callithrix penicillata, from Brazil. The parasite grew profusely in an overlay of Roswell Park Memorial Institute medium plus 20% foetal bovine serum, on Novy, McNeal and Nicolle medium (NNN , at 27°C, with a peak around 168 hr. The morphometry of cultural forms of T. minasense, estimates of cell population size and comparative growth in four different media overlays always with NNN, were studied. The infectivity of cultural forms to marmosets (C. penicillata and C. jacchus and transformation of epimastigotes into metacyclic-like forms in axenic culture in the presence of chitin derivates (chitosan were evaluated.

  6. Synergy testing of FDA-approved drugs identifies potent drug combinations against Trypanosoma cruzi.

    Directory of Open Access Journals (Sweden)

    Joseph D Planer

    2014-07-01

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

  7. Using real time process measurements to reduce catheter related bloodstream infections in the intensive care unit

    OpenAIRE

    Wall, R; Ely, E; Elasy, T; Dittus, R; Foss, J.; Wilkerson, K; Speroff, T

    2005-01-01

    

Problem: Measuring a process of care in real time is essential for continuous quality improvement (CQI). Our inability to measure the process of central venous catheter (CVC) care in real time prevented CQI efforts aimed at reducing catheter related bloodstream infections (CR-BSIs) from these devices.

  8. Patients with Central Lines - What You Need to Know to Avoid a Bloodstream Infection PSA (:60)

    Centers for Disease Control (CDC) Podcasts

    2011-03-01

    This 60 second PSA is based on the March, 2011 CDC Vital Signs report which indicates bloodstream infections in patients with central lines are largely preventable when healthcare providers use CDC-recommended infection control steps.  Created: 3/1/2011 by Centers for Disease Control and Prevention (CDC).   Date Released: 3/1/2011.

  9. Routine Surveillance for Bloodstream Infections in a Pediatric Hematopoietic Stem Cell Transplant Cohort: Do Patients Benefit?

    Directory of Open Access Journals (Sweden)

    Heather Rigby

    2007-01-01

    Full Text Available BACKGROUND: Hematopoietic stem cell transplant (HSCT recipients are at a high risk for late bloodstream infection (BSI. Controversy exists regarding the benefit of surveillance blood cultures in this immunosuppressed population. Despite the common use of this practice, the practical value is not well established in non-neutropenic children following HSCT.

  10. Secular Trends in Nosocomial Bloodstream Infections : Antibiotic-Resistant Bacteria Increase the Total Burden of Infection

    NARCIS (Netherlands)

    Ammerlaan, H. S. M.; Harbarth, S.; Buiting, A. G. M.; Crook, D. W.; Fitzpatrick, F.; Hanberger, H.; Herwaldt, L. A.; van Keulen, P. H. J.; Kluytmans, J. A. J. W.; Kola, A.; Kuchenbecker, R. S.; Lingaas, E.; Meessen, N.; Morris-Downes, M. M.; Pottinger, J. M.; Rohner, P.; dos Santos, R. P.; Seifert, H.; Wisplinghoff, H.; Ziesing, S.; Walker, A. S.; Bonten, M. J. M.

    2013-01-01

    Background. It is unknown whether rising incidence rates of nosocomial bloodstream infections (BSIs) caused by antibiotic-resistant bacteria (ARB) replace antibiotic-susceptible bacteria (ASB), leaving the total BSI rate unaffected. Methods. We investigated temporal trends in annual incidence densit

  11. Severe Community-Acquired Bloodstream Infection with Acinetobacter ursingii in Person who Injects Drugs.

    Science.gov (United States)

    Salzer, Helmut J F; Rolling, Thierry; Schmiedel, Stefan; Klupp, Eva-Maria; Lange, Christoph; Seifert, Harald

    2016-01-01

    We report a community-acquired bloodstream infection with Acinteobacter ursingii in an HIV-negative woman who injected drugs. The infection was successfully treated with meropenem. Species identification was performed by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Improved identification of Acinetobacter spp. by using this method will help identify clinical effects of this underdiagnosed pathogen.

  12. Bloodstream Infections in Very Low Birth Weight Infants with Intestinal Failure

    NARCIS (Netherlands)

    Cole, Conrad R.; Hansen, Nellie I.; Higgins, Rosemary D.; Bell, Edward F.; Shankaran, Seetha; Laptook, Abbot R.; Walsh, Michele C.; Hale, Ellen C.; Newman, Nancy S.; Das, Abhik; Stoll, Barbara J.

    2012-01-01

    Objective To examine pathogens and other characteristics associated with late-onset bloodstream infections (BSIs) in infants with intestinal failure (IF) as a consequence of necrotizing enterocolitis (NEC). Study design Infants weighing 401-1500 g at birth who survived for >72 hours and received car

  13. Assessment of the anti-protozoal activity of crude Carica papaya seed extract against Trypanosoma cruzi.

    Science.gov (United States)

    Jiménez-Coello, Matilde; Guzman-Marín, Eugenia; Ortega-Pacheco, Antonio; Perez-Gutiérrez, Salud; Acosta-Viana, Karla Y

    2013-10-11

    In order to determine the in vivo activity against the protozoan Trypanosoma cruzi, two doses (50 and 75 mg/kg) of a chloroform extract of Carica papaya seeds were evaluated compared with a control group of allopurinol. The activity of a mixture of the three main compounds (oleic, palmitic and stearic acids in a proportion of 45.9% of oleic acid, 24.1% of palmitic and 8.52% of stearic acid previously identified in the crude extract of C. papaya was evaluated at doses of 100, 200 and 300 mg/kg. Both doses of the extracts were orally administered for 28 days. A significant reduction (p papaya extract in comparison with the positive control group (allopurinol 8.5 mg/kg). Parasitemia in animals treated with the fatty acids mixture was also significantly reduced (p papaya (from ripe fruit) are able to reduce the number of parasites from both parasite stages, blood trypomastigote and amastigote (intracellular stage).

  14. On the search for potential anti-Trypanosoma cruzi drugs: synthesis and biological evaluation of 2-hydroxy-3-methylamino and 1,2,3-triazolic naphthoquinoidal compounds obtained by click chemistry reactions.

    Science.gov (United States)

    da Silva Júnior, Eufrânio N; de Melo, Isadora M M; Diogo, Emilay B T; Costa, Verenice A; de Souza Filho, José D; Valença, Wagner O; Camara, Celso A; de Oliveira, Ronaldo N; de Araujo, Alexandre S; Emery, Flávio S; dos Santos, Marcelo R; de Simone, Carlos A; Menna-Barreto, Rubem F S; de Castro, Solange L

    2012-06-01

    Five 2-hydroxy-3-substituted-aminomethyl naphthoquinones, nine 1,2,3-triazolic para-naphthoquinones, five nor-β-lapachone-based 1,2,3-triazoles, and several other naphthoquinonoid compounds were synthesized and evaluated against the infective bloodstream form of Trypanosoma cruzi, the etiological agent of Chagas disease, continuing our screening program for new trypanocidal compounds. Among all the substances, 16-18, 23, 25-29 and 30-33 were herein described for the first time and fifteen substances were identified as more potent than the standard drug benznidazole, with IC(50)/24h values in the range of 10.9-101.5 μM. Compounds 14 and 19 with Selectivity Index of 18.9 and 6.1 are important structures for further studies.

  15. Active transcription and ultrastructural changes during Trypanosoma cruzi metacyclogenesis

    Directory of Open Access Journals (Sweden)

    Ludmila R.P. Ferreira

    2008-03-01

    Full Text Available The differentiation of proliferating epimastigote forms of Trypanosoma cruzi , the protozoan parasite that causes Chagas’ disease, into the infective and non-proliferating metacyclic forms can be reproduced in the laboratory by incubating the cells in a chemically-defined medium that mimics the urine of the insect vector. Epimastigotes have a spherical nucleus, a flagellum protruding from the middle of the protozoan cell, and a disk-shaped kinetoplast - an organelle that corresponds to the mitochondrial DNA. Metacyclic trypomastigotes have an elongated shape with the flagellum protruding from the posterior portion of the cell and associated with a spherical kinetoplast. Here we describe the morphological events of this transformation and characterize a novel intermediate stage by three-dimensional reconstruction of electron microscope serial sections. This new intermediate stage is characterized by a kinetoplast compressing an already elongated nucleus, indicating that metacyclogenesis involves active movements of the flagellar structure relative to the cell body. As transcription occurs more intensely in proliferating epimastigotes than in metacyclics, we also examined the presence of RNA polymerase II and measured transcriptional activity during the differentiation process. Both the presence of the enzyme and transcriptional activity remain unchanged during all steps of metacyclogenesis. RNA polymerase II levels and transcriptional activity only decrease after metacyclics are formed. We suggest that transcription is required during the epimastigote-to-metacyclic trypomastigote differentiation process, until the kinetoplast and flagellum reach the posterior position of the parasites in the infective form.A diferenciação de formas epimastigotas (proliferativas do Trypanosoma cruzi, parasita protozoário causador da doença de Chagas, em formas metacíclicas tripomastigotas (infectivas e não proliferativas, pode ser reproduzida em laborat

  16. The Trypanosoma cruzi protease cruzain mediates immune evasion.

    Directory of Open Access Journals (Sweden)

    Patricia S Doyle

    2011-09-01

    Full Text Available Trypanosoma cruzi is the causative agent of Chagas' disease. Novel chemotherapy with the drug K11777 targets the major cysteine protease cruzain and disrupts amastigote intracellular development. Nevertheless, the biological role of the protease in infection and pathogenesis remains unclear as cruzain gene knockout failed due to genetic redundancy. A role for the T. cruzi cysteine protease cruzain in immune evasion was elucidated in a comparative study of parental wild type- and cruzain-deficient parasites. Wild type T. cruzi did not activate host macrophages during early infection (<60 min and no increase in ∼P iκB was detected. The signaling factor NF-κB P65 colocalized with cruzain on the cell surface of intracellular wild type parasites, and was proteolytically cleaved. No significant IL-12 expression occurred in macrophages infected with wild type T. cruzi and treated with LPS and BFA, confirming impairment of macrophage activation pathways. In contrast, cruzain-deficient parasites induced macrophage activation, detectable iκB phosphorylation, and nuclear NF-κB P65 localization. These parasites were unable to develop intracellularly and survive within macrophages. IL 12 expression levels in macrophages infected with cruzain-deficient T. cruzi were comparable to LPS activated controls. Thus cruzain hinders macrophage activation during the early (<60 min stages of infection, by interruption of the NF-κB P65 mediated signaling pathway. These early events allow T. cruzi survival and replication, and may lead to the spread of infection in acute Chagas' disease.

  17. Further characterization of protective Trypanosoma cruzi-specific CD4+ T-cell clones: T helper type 1-like phenotype and reactivity with shed trypomastigote antigens.

    OpenAIRE

    1993-01-01

    We previously reported the isolation from immune mice of a panel of murine clonal T-cell lines which specifically recognize antigens expressed by the trypomastigote stage of the protozoan parasite Trypanosoma cruzi, the causative agent of human Chagas' disease. Our analysis indicated that distinct clones which recognize common as well as strain-specific antigenic determinants were represented. The immunoprotective potential of several of these T-cell clones was demonstrated by adoptive transf...

  18. Proteomics of Trypanosoma evansi infection in rodents.

    Directory of Open Access Journals (Sweden)

    Nainita Roy

    Full Text Available BACKGROUND: Trypanosoma evansi infections, commonly called 'surra', cause significant economic losses to livestock industry. While this infection is mainly restricted to large animals such as camels, donkeys and equines, recent reports indicate their ability to infect humans. There are no World Animal Health Organization (WAHO prescribed diagnostic tests or vaccines available against this disease and the available drugs show significant toxicity. There is an urgent need to develop improved methods of diagnosis and control measures for this disease. Unlike its related human parasites T. brucei and T. cruzi whose genomes have been fully sequenced T. evansi genome sequence remains unavailable and very little efforts are being made to develop improved methods of prevention, diagnosis and treatment. With a view to identify potential diagnostic markers and drug targets we have studied the clinical proteome of T. evansi infection using mass spectrometry (MS. METHODOLOGY/PRINCIPAL FINDINGS: Using shot-gun proteomic approach involving nano-lc Quadrupole Time Of Flight (QTOF mass spectrometry we have identified over 160 proteins expressed by T. evansi in mice infected with camel isolate. Homology driven searches for protein identification from MS/MS data led to most of the matches arising from related Trypanosoma species. Proteins identified belonged to various functional categories including metabolic enzymes; DNA metabolism; transcription; translation as well as cell-cell communication and signal transduction. TCA cycle enzymes were strikingly missing, possibly suggesting their low abundances. The clinical proteome revealed the presence of known and potential drug targets such as oligopeptidases, kinases, cysteine proteases and more. CONCLUSIONS/SIGNIFICANCE: Previous proteomic studies on Trypanosomal infections, including human parasites T. brucei and T. cruzi, have been carried out from lab grown cultures. For T. evansi infection this is indeed the

  19. Phylogenetic analysis of the Trypanosoma genus based on the heat-shock protein 70 gene.

    Science.gov (United States)

    Fraga, Jorge; Fernández-Calienes, Aymé; Montalvo, Ana Margarita; Maes, Ilse; Deborggraeve, Stijn; Büscher, Philippe; Dujardin, Jean-Claude; Van der Auwera, Gert

    2016-09-01

    Trypanosome evolution was so far essentially studied on the basis of phylogenetic analyses of small subunit ribosomal RNA (SSU-rRNA) and glycosomal glyceraldehyde-3-phosphate dehydrogenase (gGAPDH) genes. We used for the first time the 70kDa heat-shock protein gene (hsp70) to investigate the phylogenetic relationships among 11 Trypanosoma species on the basis of 1380 nucleotides from 76 sequences corresponding to 65 strains. We also constructed a phylogeny based on combined datasets of SSU-rDNA, gGAPDH and hsp70 sequences. The obtained clusters can be correlated with the sections and subgenus classifications of mammal-infecting trypanosomes except for Trypanosoma theileri and Trypanosoma rangeli. Our analysis supports the classification of Trypanosoma species into clades rather than in sections and subgenera, some of which being polyphyletic. Nine clades were recognized: Trypanosoma carassi, Trypanosoma congolense, Trypanosoma cruzi, Trypanosoma grayi, Trypanosoma lewisi, T. rangeli, T. theileri, Trypanosoma vivax and Trypanozoon. These results are consistent with existing knowledge of the genus' phylogeny. Within the T. cruzi clade, three groups of T. cruzi discrete typing units could be clearly distinguished, corresponding to TcI, TcIII, and TcII+V+VI, while support for TcIV was lacking. Phylogenetic analyses based on hsp70 demonstrated that this molecular marker can be applied for discriminating most of the Trypanosoma species and clades.

  20. Early oral switch therapy in low-risk Staphylococcus aureus bloodstream infection (SABATO) : Study protocol for a randomized controlled trial

    NARCIS (Netherlands)

    Kaasch, Achim J.; Fätkenheuer, Gerd; Prinz-Langenohl, Reinhild; Paulus, Ursula; Hellmich, Martin; Weiß, Verena; Jung, Norma; Rieg, Siegbert; Kern, Winfried V.; Seifert, Harald; Lewalter, Karl; Lemmen, Sebastian; Stijnis, Cornelis; Van der Meer, Jan; Soriano, Alex; Ruiz, Laura Morata; Arastéh, Keikawus; Stocker, Hartmut; Kluytmans, Jan; Veenemans, Jacobien; Brodt, Hans Reinhard; Stephan, Christoph; Wolf, Timo; Kessel, Johanna; Joost, Insa; Sinha, Bhanu; van Assen, Sander; Wilting, Kasper; Tobias Welte, Welte; Christiane Mölgen, Mölgen; Julia Freise, Freise; Brunkhorst, Frank; Pletz, Mathias; Hagel, Stefan; Becker, Christian; Frieling, Thomas; Kösters, Katrin; Reuter, Stefan; Hsiao, Mikai; Rupp, Jan; Dalhoff, Klaus; Turner, David; Snape, Susan; Crusz, Shanika; Venkatesan, Pradhib; Salzberger, Bernd; Hanses, Frank; Rodriguez-Baño, Jesùs; Méndez, Adoración Valiente; López-Cortés, Luis Eduardo; Cisneros, José Miguel; Navarro-Amuedo, Maria Dolores; Bonten, Marc; Oosterheert, Jan Jelrik; Ekkelenkamp, Miquel

    2015-01-01

    Background: Current guidelines recommend that patients with Staphylococcus aureus bloodstream infection (SAB) are treated with long courses of intravenous antimicrobial therapy. This serves to avoid SAB-related complications such as relapses, local extension and distant metastatic foci. However, in

  1. Evaluation of In Vitro Activity of Essential Oils against Trypanosoma brucei brucei and Trypanosoma evansi

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    Nathan Habila

    2010-01-01

    Full Text Available Essential oils (EOs from Cymbopogon citratus (CC, Eucalyptus citriodora (EC, Eucalyptus camaldulensis (ED, and Citrus sinensis (CS were obtained by hydrodistillation process. The EOs were evaluated in vitro for activity against Trypanosoma brucei brucei (Tbb and Trypanosoma evansi (T. evansi. The EOs were found to possess antitrypanosomal activity in vitro in a dose-dependent pattern in a short period of time. The drop in number of parasite over time was achieved doses of 0.4 g/ml, 0.2 g/mL, and 0.1 g/mL for all the EOs. The concentration of 0.4 g/mL CC was more potent at 3 minutes and 2 minutes for Tbb and T. evansi, respectively. The GC-MS analysis of the EOs revealed presence of Cyclobutane (96.09% in CS, 6-octenal (77.11% in EC, Eucalyptol (75% in ED, and Citral (38.32% in CC among several other organic compounds. The results are discussed in relation to trypanosome chemotherapy.

  2. Trypanosoma cruzi and Trypanosoma rangeli: interplay with hemolymph components of Rhodnius prolixus.

    Science.gov (United States)

    Mello, C B; Garcia, E S; Ratcliffe, N A; Azambuja, P

    1995-05-01

    Studies were carried out on the course of infection of Trypanosoma cruzi (clone Dm28c) and Trypanosoma rangeli (clone San Agustin) and their interactions with hemolymph components of Rhodnius prolixus. These parasites when inoculated into the hemocoel of adult R. prolixus (i) had different courses of infection (T. rangeli had high rates of both multiplication and infection and T. cruzi had no division and disappeared soon from the hemolymph); (ii) induced high but no differential increases in lysozyme levels; (iii) failed to induce any other antibacterial activity; (iv) showed similar patterns of hemolymph agglutination activity for erythrocytes and parasites, although there was evidence of limited, unquantifiable, agglutination of T. cruzi; (v) elicited different hemocyte responses with only the T. rangeli infection resulting in significantly increased hemocyte counts; and (vi) did not induce trypanolytic activity. These experiments, unlike previous studies, also showed (i) an interaction of these trypanosomes with the prophenoloxidase-activating system [phenoloxidase (PO) production was spontaneously activated by both parasites but the number of T. cruzi in the hemolymph was directly correlated with PO levels] and (ii) that the elimination of T. cruzi also corresponded to the formation of nodules in the hemolymph. The significance of these results is discussed in relation to the hypothesis that T. rangeli but not T. cruzi has the ability to escape from and perhaps utilize the vector immune system in order to successfully colonize the R. prolixus hemolymph.

  3. Localization of serum resistance-associated protein in Trypanosoma brucei rhodesiense and transgenic Trypanosoma brucei brucei.

    Science.gov (United States)

    Bart, Jean-Mathieu; Cordon-Obras, Carlos; Vidal, Isabel; Reed, Jennifer; Perez-Pastrana, Esperanza; Cuevas, Laureano; Field, Mark C; Carrington, Mark; Navarro, Miguel

    2015-10-01

    African trypanosomes infect a broad range of mammals, but humans and some higher primates are protected by serum trypanosome lytic factors that contain apolipoprotein L1 (ApoL1). In the human-infective subspecies of Trypanosoma brucei, Trypanosoma brucei rhodesiense, a gene product derived from the variant surface glycoprotein gene family member, serum resistance-associated protein (SRA protein), protects against ApoL1-mediated lysis. Protection against trypanosome lytic factor requires the direct interaction between SRA protein and ApoL1 within the endocytic apparatus of the trypanosome, but some uncertainty remains as to the precise mechanism and location of this interaction. In order to provide more insight into the mechanism of SRA-mediated resistance to trypanosome lytic factor, we assessed the localization of SRA in T. b. rhodesiense EATRO3 using a novel monoclonal antibody raised against SRA together with a set of well-characterized endosomal markers. By three-dimensional deconvolved immunofluorescence single-cell analysis, combined with double-labelling immunoelectron microscopy, we found that ≈ 50% of SRA protein localized to the lysosome, with the remaining population being distributed through the endocytic pathway, but apparently absent from the flagellar pocket membrane. These data suggest that the SRA/trypanolytic factor interaction is intracellular, with the concentration within the endosomes potentially crucial for ensuring a high efficiency.

  4. Immunopathological Aspects of Experimental Trypanosoma cruzi Reinfections

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    Juliana Reis Machado

    2014-01-01

    Full Text Available Chagas disease is caused by Trypanosoma cruzi infection. Besides the host-related factors, such as immune response and genetic background, the parasite, strain, and occurrences of reinfection episodes, may influence disease outcome. Our results demonstrate that both the primary infection and the reinfection with the Colombiana strain are connected with lower survival rate of the mice. After reinfection, parasitaemia is approximately ten times lower than in primary infected animals. Only Colombiana, Colombiana/Colombiana, and Y/Colombiana groups presented amastigote nests in cardiac tissue. Moreover, the mice infected and/or reinfected with the Colombiana strain had more T. cruzi nests, more intense inflammatory infiltrate, and higher in situ expression of TNF-α and IFN-γ than Y strain. Antigen-stimulated spleen cells from infected and/or reinfected animals produced higher levels of TNF-α, IFN-γ, and IL-10. Our results reinforce the idea that Chagas disease outcome is influenced by the strain of the infective parasite, being differentially modulated during reinfection episodes. It highlights the need of control strategies involving parasite strain characterization in endemic areas for Chagas disease.

  5. Inducible suicide vector systems for Trypanosoma cruzi.

    Science.gov (United States)

    Ma, Yanfen; Weiss, Louis M; Huang, Huan

    2015-06-01

    Chagas disease caused by Trypanosoma cruzi is a major neglected tropical parasitic disease. The pathogenesis of this infection remains disputable. There is no suitable vaccine for the prevention. Attenuated live vaccines can provide strong protection against infection; however, there are the concerns about latent infection or reversion to virulence in such attenuated strains. A method to induce T. cruzi death would provide a critical tool for research into the pathophysiological mechanisms and provide a novel design of safe live attenuated vaccines. We established effective inducible systems for T. cruzi employing the degradation domain based on the Escherichia coli dihydrofolate reductase (ecDHFR). The DHFR degradation domain (DDD) can be stabilized by trimethoprim-lactate and can be used to express detrimental or toxic proteins. T. cruzi lines with Alpha-toxin, Cecropin A and GFP under the control of DDD with a hemagglutinin tag (HA) were developed. Interestingly, amastigotes bearing GFP-DDDHA, Alpha-toxin-DDDHA, Cecropin A-DDDHA and DDDHA all resulted in inducible cell death with these fusions, indicating that DDDHA protein is also detrimental to amastigotes. Furthermore, these strains were attenuated in mouse experiments producing no pathological changes and inoculation with these DDDHA strains in mice provided strong protection against lethal wild type infection.

  6. Flagellar Motility of Trypanosoma cruzi Epimastigotes

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    G. Ballesteros-Rodea

    2012-01-01

    Full Text Available The hemoflagellate Trypanosoma cruzi is the causative agent of American trypanosomiasis. Despite the importance of motility in the parasite life cycle, little is known about T. cruzi motility, and there is no quantitative description of its flagellar beating. Using video microscopy and quantitative vectorial analysis of epimastigote trajectories, we find a forward parasite motility defined by tip-to-base symmetrical flagellar beats. This motion is occasionally interrupted by base-to-tip highly asymmetric beats, which represent the ciliary beat of trypanosomatid flagella. The switch between flagellar and ciliary beating facilitates the parasite's reorientation, which produces a large variability of movement and trajectories that results in different distance ranges traveled by the cells. An analysis of the distance, speed, and rotational angle indicates that epimastigote movement is not completely random, and the phenomenon is highly dependent on the parasite behavior and is characterized by directed and tumbling parasite motion as well as their combination, resulting in the alternation of rectilinear and intricate motility paths.

  7. Motility modes of the parasite Trypanosoma brucei

    Science.gov (United States)

    Temel, Fatma Zeynep; Qu, Zijie; McAllaster, Michael; de Graffenried, Christopher; Breuer, Kenneth

    2015-11-01

    The parasitic single-celled protozoan Trypanosoma brucei causes African Sleeping Sickness, which is a fatal disease in humans and animals that threatens more than 60 million people in 36 African countries. Cell motility plays a critical role in the developmental phases and dissemination of the parasite. Unlike many other motile cells such as bacteria Escherichia coli or Caulobacter crescentus, the flagellum of T. brucei is attached along the length of its awl-like body, producing a unique mode of motility that is not fully understood or characterized. Here, we report on the motility of T. brucei, which swims using its single flagellum employing both rotating and undulating propulsion modes. We tracked cells in real-time in three dimensions using fluorescent microscopy. Data obtained from experiments using both short-term tracking within the field of view and long-term tracking using a tracking microscope were analyzed. Motility modes and swimming speed were analyzed as functions of cell size, rotation rate and undulation pattern. Research supported by NSF.

  8. The Uptake of GABA in Trypanosoma cruzi.

    Science.gov (United States)

    Galvez Rojas, Robert L; Ahn, Il-Young; Suárez Mantilla, Brian; Sant'Anna, Celso; Pral, Elizabeth Mieko Furusho; Silber, Ariel Mariano

    2015-01-01

    Gamma aminobutyric acid (GABA) is widely known as a neurotransmitter and signal transduction molecule found in vertebrates, plants, and some protozoan organisms. However, the presence of GABA and its role in trypanosomatids is unknown. Here, we report the presence of intracellular GABA and the biochemical characterization of its uptake in Trypanosoma cruzi, the etiological agent of Chagas' disease. Kinetic parameters indicated that GABA is taken up by a single transport system in pathogenic and nonpathogenic forms. Temperature dependence assays showed a profile similar to glutamate transport, but the effect of extracellular cations Na(+) , K(+) , and H(+) on GABA uptake differed, suggesting a different uptake mechanism. In contrast to reports for other amino acid transporters in T. cruzi, GABA uptake was Na(+) dependent and increased with pH, with a maximum activity at pH 8.5. The sensitivity to oligomycin showed that GABA uptake is dependent on ATP synthesis. These data point to a secondary active Na(+) /GABA symporter energized by Na(+) -exporting ATPase. Finally, we show that GABA occurs in the parasite's cytoplasm under normal culture conditions, indicating that it is regularly taken up from the culture medium or synthesized through an still undescribed metabolic pathway.

  9. ADAPTACIÓN DE LA CEPA MUNANTA DE TRYPANOSOMA CRUZI AL CULTIVO IN VITRO EN CÉLULAS VERO

    OpenAIRE

    Velazco Gamboa, Carolina; Departamento de Microbiología, Facultad de Ciencias, Pontificia Universidad Javeriana, Bogotá; Puentes Corredor, Manuel; Instituto de Inmunología - HSJD, Bogotá; Moreno García, Alberto; Instituto de Inmunología - HSJD, Bogotá; Patarroyo Murillo, Manuel Elkin; Instituto de Inmunología - HSJD, Bogotá; Puerta Bula, Concepción; Departamento de Microbiología, Facultad de Ciencias, Pontificia Universidad Javeriana, Bogotá

    2013-01-01

    The importance of obtaining the different stages of Trypanosoma cruzi in order to recognize the antigens involved in the intracellular invasion and replication processes, makes it necessary to adapt these strains to tissue culture, especially considering the high leve! of biological, biochemical and genetic variation, which is found among the strains and clones ofthe parasite. Within the aforementioned context, in this study, the Colombian strain ofT. cruzi, M un anta, was adapted to tissue c...

  10. Current strategies for the prevention and management of central line-associated bloodstream infections

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    Zhuolin Han

    2010-11-01

    Full Text Available Zhuolin Han, Stephen Y Liang, Jonas MarschallDivision of Infectious Diseases, Washington University School of Medicine in St Louis, St Louis, MO, USAAbstract: Central venous catheters are an invaluable tool for diagnostic and therapeutic purposes in today’s medicine, but their use can be complicated by bloodstream infections (BSIs. While evidence-based preventive measures are disseminated by infection control associations, the optimal management of established central line-associated BSIs has been summarized in infectious diseases guidelines. We prepared an overview of the state-of-the-art of prevention and management of central line-associated BSIs and included topics such as the role of antibiotic-coated catheters, the role of catheter removal in the management, and a review of currently used antibiotic compounds and the duration of treatment.Keywords: central venous catheters, bloodstream infections, guidelines, prevention

  11. Cluster of Candida parapsilosis primary bloodstream infection in a neonatal intensive care unit

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    Silva Carmem Lúcia P. da

    2001-01-01

    Full Text Available Candida parapsilosis is an increasingly important bloodstream pathogen in neonatal intensive care units (NICU. We investigated a cluster of bloodstream infections in a NICU to determine whether nosocomial transmission occurred. During a 3-day period, 3 premature infants hospitalized in the same unit presented with sepsis caused by C. parapsilosis. Electrophoretic karyotype of the organisms was performed by using pulsed field gel electrophoresis in a countour-clamped homogeneous electric field system. The isolate from 1 newborn could not be typed, and the isolates from the remaining 2 infants had identical patterns. All 3 cases are described. We conclude that nosocomial transmission of C. parapsilosis occurred and that neonates under intensive care may represent a risk group for this pathogen.

  12. Megazol and its bioisostere 4H-1,2,4-triazole: comparing the trypanocidal, cytotoxic and genotoxic activities and their in vitro and in silico interactions with the Trypanosoma brucei nitroreductase enzyme.

    Science.gov (United States)

    Carvalho, Alcione Silva de; Salomão, Kelly; Castro, Solange Lisboa de; Conde, Taline Ramos; Zamith, Helena Pereira da Silva; Caffarena, Ernesto Raúl; Hall, Belinda Suzette; Wilkinson, Shane Robert; Boechat, Núbia

    2014-06-01

    Megazol (7) is a 5-nitroimidazole that is highly active against Trypanosoma cruzi and Trypanosoma brucei, as well as drug-resistant forms of trypanosomiasis. Compound 7 is not used clinically due to its mutagenic and genotoxic properties, but has been largely used as a lead compound. Here, we compared the activity of 7 with its 4H-1,2,4-triazole bioisostere (8) in bloodstream forms of T. brucei and T. cruzi and evaluated their activation by T. brucei type I nitroreductase (TbNTR) enzyme. We also analysed the cytotoxic and genotoxic effects of these compounds in whole human blood using Comet and fluorescein diacetate/ethidium bromide assays. Although the only difference between 7 and 8 is the substitution of sulphur (in the thiadiazole in 7) for nitrogen (in the triazole in 8), the results indicated that 8 had poorer antiparasitic activity than 7 and was not genotoxic, whereas 7 presented this effect. The determination of Vmax indicated that although 8 was metabolised more rapidly than 7, it bounds to the TbNTR with better affinity, resulting in equivalent kcat/KM values. Docking assays of 7 and 8 performed within the active site of a homology model of the TbNTR indicating that 8 had greater affinity than 7.

  13. Tsukamurella catheter-related bloodstream infection in a pediatric patient with pulmonary hypertension

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    Kristen A. Wendorf

    2010-03-01

    Full Text Available Catheter-related bloodstream infections (CR-BSI are important complications in patients with long-term indwelling central venous catheters. In this report, we present the case of a 14-year-old male with pulmonary hypertension treated with continuous treprostinil infusion, who presented with a CR-BSI caused by a Tsukamurella species. This case highlights the potential for this unusual organism to cause infection in immunocompetent patients.

  14. Enterococcal bloodstream infection. Design and validation of a mortality prediction rule

    OpenAIRE

    Perez-Garcia, Alejandra; Landecho, Manuel; Beunza Nuin, Juan Jose; Conde-Estévez, D; Horcajada, J.P.; Grau, S.; Gea Sánchez, Alfredo; E. Mauleón; Sorli, L.; Gómez, J.; Terradas, R.; Lucena, J.F. (Juan F.); Alegre Garrido, Félix; A. Huerta; Pozo, José Luis del

    2016-01-01

    To develop a prediction rule to describe the risk of death as a result of enterococcal bloodstream infection. A prediction rule was developed by analysing data collected from 122 patients diagnosed with enterococcal BSI admitted to the Clínica Universidad de Navarra (Pamplona, Spain); and validated by confirming its accuracy with the data of an external population (Hospital del Mar, Barcelona). According to this model, independent significant predictors for the risk of death were being diabet...

  15. Risk of vancomycin-resistant enterococci bloodstream infection among patients colonized with vancomycin-resistant enterococci

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    Ahu Kara

    2015-02-01

    Full Text Available Background:Vancomycin-resistant enterococci colonization has been reported to increase the risk of developing infections, including bloodstream infections.Aim:In this study, we aimed to share our experience with the vancomycin-resistant enterococci bloodstream infections following gastrointestinal vancomycin-resistant enterococci colonization in pediatric population during a period of 18 months.Method:A retrospective cohort of children admitted to a 400-bed tertiary teaching hospital in Izmir, Turkey whose vancomycin-resistant enterococci colonization was newly detected during routine surveillances for gastrointestinal vancomycin-resistant enterococci colonization during the period of January 2009 and December 2012 were included in this study. All vancomycin-resistant enterococci isolates found within 18 months after initial detection were evaluated for evidence of infection.Findings: Two hundred and sixteen patients with vancomycin-resistant enterococci were included in the study. Vancomycin-resistant enterococci colonization was detected in 136 patients (62.3% while they were hospitalized at intensive care units; while the remaining majority (33.0% were hospitalized at hematology-oncology department. Vancomycinresistant enterococci bacteremia was present only in three (1.55% patients. All these patients were immunosuppressed due to human immunodeficiency virus (one patient and intensive chemotherapy (two patients.Conclusion:In conclusion, our study found that 1.55% of vancomycin-resistant enterococcicolonized children had developed vancomycin-resistant enterococci bloodstream infection among the pediatric intensive care unit and hematology/oncology patients; according to our findings, we suggest that immunosupression is the key point for developing vancomycinresistant enterococci bloodstream infections.

  16. Efficacy of an infection control programme in reducing nosocomial bloodstream infections in a Senegalese neonatal unit.

    Science.gov (United States)

    Landre-Peigne, C; Ka, A S; Peigne, V; Bougere, J; Seye, M N; Imbert, P

    2011-10-01

    Neonatal nosocomial infections are public health threats in the developing world, and successful interventions are rarely reported. A before-and-after study was conducted in the neonatal unit of the Hôpital Principal de Dakar, Senegal to assess the efficacy of a multi-faceted hospital infection control programme implemented from March to May 2005. The interventions included clustering of nursing care, a simple algorithm for empirical therapy of suspected early-onset sepsis, minimal invasive care and promotion of early discharge of neonates. Data on nosocomial bloodstream infections, mortality, bacterial resistance and antibiotic use were collected before and after implementation of the infection control programme. One hundred and twenty-five infants were admitted immediately before the programme (Period 1, January-February 2005) and 148 infants were admitted immediately after the programme (Period 2, June-July 2005). The two groups of infants were comparable in terms of reason for admission and birth weight. After implementation of the infection control programme, the overall rate of nosocomial bloodstream infections decreased from 8.8% to 2.0% (P=0.01), and the rate of nosocomial bloodstream infections/patient-day decreased from 10.9 to 2.9/1000 patient-days (P=0.03). Overall mortality rates did not differ significantly. The proportion of neonates who received antimicrobial therapy for suspected early-onset sepsis decreased significantly from 100% to 51% of at-risk infants (Pnosocomial bloodstream infections, and the efficacy of these interventions was long-lasting. Such interventions could be extended to other low-income countries.

  17. Second-Generation central venous catheter in the prevention of bloodstream infection: a systematic review 1

    Science.gov (United States)

    Stocco, Janislei Gislei Dorociaki; Hoers, Hellen; Pott, Franciele Soares; Crozeta, Karla; Barbosa, Dulce Aparecida; Meier, Marineli Joaquim

    2016-01-01

    Abstract Objective: to evaluate the effectiveness and safety in the use of second-generation central venous catheters impregnated in clorhexidine and silver sulfadiazine when compared with other catheters, being them impregnated or not, in order to prevent the bloodstream infection prevention. Method: systematic review with meta-analysis. Databases searched: MEDLINE, EMBASE, CINAHL, LILACS/SciELO, Cochrane CENTRAL; search in Congress Proceedings and records from Clinical Trials. Results: 1.235 studies were identified, 97 were pre-selected and 4 were included. In catheter-related bloodstream infection, there was no statistical significance between second-generation impregnated catheter compared with the non-impregnated ones, absolute relative risk 1,5% confidence interval 95% (3%-1%), relative risk 0,68 (confidence interval 95%, 0,40-1,15) and number needed to treat 66. In the sensitivity analysis, there was less bloodstream infection in impregnated catheters (relative risk 0,50, confidence interval 95%, 0,26-0,96). Lower colonization, absolute relative risk 9,6% (confidence interval 95%, 10% to 4%), relative risk 0,51 (confidence interval 95% from 0,38-0,85) and number needed to treat 5. Conclusion: the use of second-generation catheters was effective in reducing the catheter colonization and infection when a sensitivity analysis is performed. Future clinical trials are suggested to evaluate sepsis rates, mortality and adverse effects. PMID:27508901

  18. Cost-Effectiveness of Surveillance for Bloodstream Infections for Sepsis Management in Low-Resource Settings.

    Science.gov (United States)

    Penno, Erin C; Baird, Sarah J; Crump, John A

    2015-10-01

    Bacterial sepsis is a leading cause of mortality among febrile patients in low- and middle-income countries, but blood culture services are not widely available. Consequently, empiric antimicrobial management of suspected bloodstream infection is based on generic guidelines that are rarely informed by local data on etiology and patterns of antimicrobial resistance. To evaluate the cost-effectiveness of surveillance for bloodstream infections to inform empiric management of suspected sepsis in low-resource areas, we compared costs and outcomes of generic antimicrobial management with management informed by local data on etiology and patterns of antimicrobial resistance. We applied a decision tree model to a hypothetical population of febrile patients presenting at the district hospital level in Africa. We found that the evidence-based regimen saved 534 more lives per 100,000 patients at an additional cost of $25.35 per patient, resulting in an incremental cost-effectiveness ratio of $4,739. This ratio compares favorably to standard cost-effectiveness thresholds, but should ultimately be compared with other policy-relevant alternatives to determine whether routine surveillance for bloodstream infections is a cost-effective strategy in the African context.

  19. Infecção por Trypanosoma cruzi em candidatos a doador de sangue Trypanosoma cruzi infection in blood donors

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    Ana M. Bonametti

    1998-12-01

    Full Text Available INTRODUÇÃO: A transmissão transfusional da tripanossomíase americana tem-se reduzido no Brasil, com a progressiva ampliação do controle de qualidade do sangue. Nesse sentido, realizou-se pesquisa para avaliar a atual soro-prevalência da infecção por Trypanosoma cruzi em candidatos a doador de sangue em Londrina, Paraná (Brasil, e comparar essa taxa com a encontrada em candidatos a doador estudados em 1958 e 1975, na mesma cidade. MÉTODO: Estudo transversal para determinação da soroprevalência. O imuno-diagnóstico de infecção por Trypanosoma cruzi foi realizado através das técnicas imunoenzimática (ELISA e imunofluorescência indireta. RESULTADOS E CONCLUSÃO: A taxa de soroprevalência encontrada foi de 1,3%. Foi detectada tendência de queda temporal da taxa de positividade dos testes sorológicos para o diagnóstico de infecção por Trypanosoma cruzi nos bancos de sangue do município estudado nos anos de 1958, 1975 e 1995.INTRODUCTION: Transmission of American trypanossomiasis by transfusion has been reduced by expansion of control measures of blood quality in Brazil. A research project was, therefore, undertaken to evaluate soropositivity for Trypanosoma cruzi infection on blood donors and to compare this rate with those found in 1958 and 1975 in blood banks. METHOD: A transversal study was carried out on blood donors in Londrina, Paraná, Brazil. ELISA and Immunofluorescence were the serological test techniques used in the diagnosis of Trypanosoma cruzi infection. RESULTS AND CONCLUSION: A serumprevalence rate of 1.3% was found with a tendency for positive serum findings for Trypanosoma cruzi infection on blood donors to decrease over Aime (1958, 1975, and 1995.

  20. Clinical and microbiological characteristics of bloodstream infections in a tertiary hospital in Maceió, Alagoas, Brazil

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    Maria Tereza Freitas Tenório

    2010-04-01

    Full Text Available We observed the clinical and microbiological characteristics of several stages of bloodstream infections (BSI, as well as the mortality attributed to it in a tertiary hospital in the northeast of Brazil (in the city of Maceió, Alagoas. A prospective cohort of 143 patients who had at least one positive blood culture was enrolled in the study. Their clinical evolution was followed up for 30 days from October 2005 to December 2006. The relation among the qualitative variables was verified through Chi-square test. The significance level was 5%. The statistical package adopted was SPSS 15.0 for Windows. Up to the thirtieth day, 30.1% of the patients presented bacteremia and 69.9% developed sepsis.Among these, 20.3% developed severe sepsis and 10.5% septic shock. The mortality attributed to it was 37.8%. In bacteremia, sepsis, severe sepsis, and septic shock conditions, mortality rates were 9.3%, 50%, 65.5%, and 84.6%, respectively. Respiratory (32.2% and urinary (14% sources and the ones related to central venous catheter (14% were prevalent. In the wards 55.12% of the cases developed sepsis, whereas in the intensive care units, the rate was 87.69% (p < 0.05. Chronic renal failure, diabetes melitus, and neuropathy were present in 21.7%, 26.6%, and 29.4% of the cases, respectively. Coagulase-negative Staphylococcus (25.9%, Staphylococcus aureus (21%, and Klebsiella pneumoniae (14% were the most present microorganism in the sample. The high morbidity and mortality rates in this study are attributed to the lack of knowledge on BSI characteristics and on instituted protocols for detection and treatment in early stages.

  1. Reversibility of muscle and heart lesions in chronic Trypanosoma cruzi infected mice after late trypanomicidal treatment

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    M. A. Segura

    1994-06-01

    Full Text Available The effect of trypanomicidal treatment upon established histopathological Trypanosoma cruzi induced lesions was studied in Swiss mice. The animals were inoculated with 50 trypomastigotes and infection was allowed to progress without treatment for 99 days. After this period, the animals were divided in three groups, treated for 30 days with either placebo, benznidazole (200 mg/kg/day or nifurtimox (100 mg/kg/day. These treatments induced 94 and 100 (per cent cure rates respectively as detected by xenodiagnosis and reduction of antibody levels. Autopsies and histopathological studies of heart, urinary bladderand skeletal muscle performed on day 312 after infection showed almost complete healing without residual lesions. As long periods were allowed between infection, treatment and autopsy, the results indicate that tissue lesions depend, up to advances stages, on the continuous presence of the parasite.

  2. Host cell poly(ADP-ribose glycohydrolase is crucial for Trypanosoma cruzi infection cycle.

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    Salomé C Vilchez Larrea

    Full Text Available Trypanosoma cruzi, etiological agent of Chagas' disease, has a complex life cycle which involves the invasion of mammalian host cells, differentiation and intracellular replication. Here we report the first insights into the biological role of a poly(ADP-ribose glycohydrolase in a trypanosomatid (TcPARG. In silico analysis of the TcPARG gene pointed out the conservation of key residues involved in the catalytic process and, by Western blot, we demonstrated that it is expressed in a life stage-dependant manner. Indirect immunofluorescence assays and electron microscopy using an anti-TcPARG antibody showed that this enzyme is localized in the nucleus independently of the presence of DNA damage or cell cycle stage. The addition of poly(ADP-ribose glycohydrolase inhibitors ADP-HPD (adenosine diphosphate (hydroxymethyl pyrrolidinediol or DEA (6,9-diamino-2-ethoxyacridine lactate monohydrate to the culture media, both at a 1 µM concentration, reduced in vitro epimastigote growth by 35% and 37% respectively, when compared to control cultures. We also showed that ADP-HPD 1 µM can lead to an alteration in the progression of the cell cycle in hydroxyurea synchronized cultures of T. cruzi epimastigotes. Outstandingly, here we demonstrate that the lack of poly(ADP-ribose glycohydrolase activity in Vero and A549 host cells, achieved by chemical inhibition or iRNA, produces the reduction of the percentage of infected cells as well as the number of amastigotes per cell and trypomastigotes released, leading to a nearly complete abrogation of the infection process. We conclude that both, T. cruzi and the host, poly(ADP-ribose glycohydrolase activities are important players in the life cycle of Trypanosoma cruzi, emerging as a promising therapeutic target for the treatment of Chagas' disease.

  3. EPIDEMIOLOGÍA MOLECULAR DE TRYPANOSOMA CRUZI

    Directory of Open Access Journals (Sweden)

    Felipe Guhl

    2013-01-01

    Full Text Available La enfermedad de Chagas causada por el parásito Trypanosoma cruzi es una zoonosis compleja, ampliamente distribuida en el continente americano. La infección puede ser adquirida a través de las heces de insectos triatominos, transfusión de sangre, trasplante de órganos, vía oral, por transmisión congénita y por accidentes de laboratorio. El completo entendimiento de la etiología y epidemiología de la enfermedad de Chagas a través de su distribución geográfica es complejo y permanece bajo intensa investigación hasta la actualidad. Los recientes estudios sobre la variabilidad genética del parásito han dado nuevas luces de los diferentes escenarios de los ciclos de transmisión de la enfermedad y su patogénesis en humanos. El propósito principal para la caracterización molecular de T.cruzi y sus múltiples genotipos está dirigido hacia su asociación con la clínica y la patogenesis de la enfermedad, así como al esclarecimiento de los diferentes escenarios de transmisión y los aspectos coevolutivos relacionados con reservorios e insectos vectores. La caracterización molecular de los diferentes aislamientos a partir de humanos, insectos y reservorios, ha permitido identificar la amplia variabilidad genética del parásito, abriendo nuevos caminos hacia la búsqueda de nuevos blancos terapéuticos y pruebas diagnósticas más específicas que contribuyan a mitigar la enfermedad de Chagas.

  4. Nanomolar Inhibitors of Trypanosoma brucei RNA Triphosphatase

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    Paul Smith

    2016-02-01

    Full Text Available Eukaryal taxa differ with respect to the structure and mechanism of the RNA triphosphatase (RTPase component of the mRNA capping apparatus. Protozoa, fungi, and certain DNA viruses have a metal-dependent RTPase that belongs to the triphosphate tunnel metalloenzyme (TTM superfamily. Because the structures, active sites, and chemical mechanisms of the TTM-type RTPases differ from those of mammalian RTPases, the TTM RTPases are potential targets for antiprotozoal, antifungal, and antiviral drug discovery. Here, we employed RNA interference (RNAi knockdown methods to show that Trypanosoma brucei RTPase Cet1 (TbCet1 is necessary for proliferation of procyclic cells in culture. We then conducted a high-throughput biochemical screen for small-molecule inhibitors of the phosphohydrolase activity of TbCet1. We identified several classes of chemicals—including chlorogenic acids, phenolic glycopyranosides, flavonoids, and other phenolics—that inhibit TbCet1 with nanomolar to low-micromolar 50% inhibitory concentrations (IC50s. We confirmed the activity of these compounds, and tested various analogs thereof, by direct manual assays of TbCet1 phosphohydrolase activity. The most potent nanomolar inhibitors included tetracaffeoylquinic acid, 5-galloylgalloylquinic acid, pentagalloylglucose, rosmarinic acid, and miquelianin. TbCet1 inhibitors were less active (or inactive against the orthologous TTM-type RTPases of mimivirus, baculovirus, and budding yeast (Saccharomyces cerevisiae. Our results affirm that a TTM RTPase is subject to potent inhibition by small molecules, with the caveat that parallel screens against TTM RTPases from multiple different pathogens may be required to fully probe the chemical space of TTM inhibition.

  5. Autochthonous Transmission of Trypanosoma Cruzi in Southern California

    Science.gov (United States)

    Hernandez, Salvador; Flores, Carmen A.; Viana, Gracia M.; Sanchez, Daniel R.; Traina, Mahmoud I.

    2016-01-01

    Trypanosoma cruzi usually infects humans via triatomine insects in Latin America. Vector-borne transmission in the United States is exceedingly rare. We describe (1) the first case of probable autochthonous transmission reported in California in more than 30 years and (2) the first ever reported case in the greater Los Angeles area. PMID:28018928

  6. Bestatin Induces Specific Changes in Trypanosoma cruzi Dipeptide Pool

    OpenAIRE

    2015-01-01

    Proteases and peptidases in Trypanosoma cruzi are considered potential targets for antichagasic chemotherapy. We monitored changes in low-mass metabolites in T. cruzi epimastigotes treated with bestatin, a dipeptide metalloaminopeptidase inhibitor. After treatment, multiple dipeptides were shown to be increased, confirming in situ inhibition of the leucine aminopeptidase of T. cruzi (LAPTc) and probably other peptidases.

  7. Trypanosoma brucei mitochondrial respiratome: Composition and organization in procyclic form

    KAUST Repository

    Acestor, Nathalie

    2011-05-24

    The mitochondrial respiratory chain is comprised of four different protein complexes (I-IV), which are responsible for electron transport and generation of proton gradient in the mitochondrial intermembrane space. This proton gradient is then used by F oF 1-ATP synthase (complex V) to produce ATP by oxidative phosphorylation. In this study, the respiratory complexes I, II, and III were affinity purified from Trypanosoma brucei procyclic form cells and their composition was determined by mass spectrometry. The results along with those that we previously reported for complexes IV and V showed that the respiratome of Trypanosoma is divergent because many of its proteins are unique to this group of organisms. The studies also identified two mitochondrial subunit proteins of respiratory complex IV that are encoded by edited RNAs. Proteomics data from analyses of complexes purified using numerous tagged component proteins in each of the five complexes were used to generate the first predicted protein-protein interaction network of the Trypanosoma brucei respiratory chain. These results provide the first comprehensive insight into the unique composition of the respiratory complexes in Trypanosoma brucei, an early diverged eukaryotic pathogen. © 2011 by The American Society for Biochemistry and Molecular Biology, Inc.

  8. A tropical tale: how Naja nigricollis venom beats Trypanosoma brucei

    DEFF Research Database (Denmark)

    Martos Esteban, Andrea; Laustsen, Andreas Hougaard; Carrington, Mark

    Trypanosoma brucei is a parasitic protozoan species capable to infecting insect vectors whose bite further produces African sleeping sickness inhuman beings [1]. During the parasite’s extracellular life in the mammalian host,its outer coat, mainly composed of Variable Surface Glycoproteins (VSGs...

  9. Trypanosoma cruzi: avirulence of the PF strain to Callithrix marmosets

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    Humberto Menezes

    1981-06-01

    Full Text Available Callithrix jacchus geoffroy marmosets (HumBol. 1812 were injected once subcutaneously with 10.000 parasites/g body weight and followed for a period of six months. The PF strain of Trypanosoma cruzi was used. Follow-up was done through blood cultures, xenodiagnosis, serological tests, and ECG. A small number of normaI animais served as control.

  10. National Bloodstream Infection Surveillance in Switzerland 2008-2014: Different Patterns and Trends for University and Community Hospitals.

    Science.gov (United States)

    Buetti, Niccolò; Marschall, Jonas; Atkinson, Andrew; Kronenberg, Andreas

    2016-09-01

    OBJECTIVE To characterize the epidemiology of bloodstream infections in Switzerland, comparing selected pathogens in community and university hospitals. DESIGN Observational, retrospective, multicenter laboratory surveillance study. METHODS Data on bloodstream infections from 2008 through 2014 were obtained from the Swiss infection surveillance system, which is part of the Swiss Centre for Antibiotic Resistance (ANRESIS). We compared pathogen prevalences across 26 acute care hospitals. A subanalysis for community-acquired and hospital-acquired bloodstream infections in community and university hospitals was performed. RESULTS A total of 42,802 bloodstream infection episodes were analyzed. The most common etiologies were Escherichia coli (28.3%), Staphylococcus aureus (12.4%), and polymicrobial bloodstream infections (11.4%). The proportion of E. coli increased from 27.5% in 2008 to 29.6% in 2014 (P = .04). E. coli and S. aureus were more commonly reported in community than university hospitals (34.3% vs 22.7%, P<.001 and 13.9% vs 11.1%, P<.001, respectively). Fifty percent of episodes were community-acquired, with E. coli again being more common in community hospitals (41.0% vs 32.4%, P<.001). The proportion of E. coli in community-acquired bloodstream infections increased in community hospitals only. Community-acquired polymicrobial infections (9.9% vs 5.6%, P<.001) and community-acquired coagulase-negative staphylococci (6.7% vs 3.4%, P<0.001) were more prevalent in university hospitals. CONCLUSIONS The role of E. coli as predominant pathogen in bloodstream infections has become more pronounced. There are distinct patterns in community and university hospitals, potentially influencing empirical antibiotic treatment. Infect Control Hosp Epidemiol 2016;37:1060-1067.

  11. Characterization of Calflagin, a Flagellar Calcium-Binding Protein from Trypanosoma congolense

    Science.gov (United States)

    Eyford, Brett A.; Kaufman, Laura; Salama-Alber, Orly; Loveless, Bianca; Pope, Matthew E.; Burke, Robert D.; Matovu, Enock; Boulanger, Martin J.; Pearson, Terry W.

    2016-01-01

    Background Identification of species-specific trypanosome molecules is important for laboratory- and field-based research into epidemiology and disease diagnosis. Although Trypanosoma congolense is the most important trypanosome pathogen of cattle in Africa, no species-specific molecules found in infective bloodstream forms (BSF) of the parasites have been identified, thus limiting development of diagnostic tests. Methods Immuno-mass spectrometric methods were used to identify a protein that is recognized by a T. congolense-specific monoclonal antibody (mAb) Tc6/42.6.4. The identified molecule was expressed as a recombinant protein in E. coli and was tested in several immunoassays for its ability to interact with the mAb. The three dimensional structure of the protein was modeled and compared to crystal- and NMR-structures of the homologous proteins from T. cruzi and T. brucei respectively, in order to examine structural differences leading to the different immunoreactivity of the T. congolense molecule. Enzyme-linked immunosorbent assays (ELISA) were used to measure antibodies produced by trypanosome-infected African cattle in order to assess the potential for use of T. congolense calflagin in a serodiagnostic assay. Results The antigen recognized by the T. congolense-specific mAb Tc6/42.6.4 was identified as a flagellar calcium-binding protein, calflagin. The recombinant molecule showed immunoreactivity with the T. congolense-specific mAb confirming that it is the cognate antigen. Immunofluorescence experiments revealed that Ca2+ modulated the localization of the calflagin molecule in trypanosomes. Structural modelling and comparison with calflagin homologues from other trypanosomatids revealed four non-conserved regions on the surface of the T. congolense molecule that due to differences in surface chemistry and structural topography may form species-specific epitopes. ELISAs using the recombinant calflagin as antigen to detect antibodies in trypanosome

  12. Protection of mice against Trypanosoma cruzi by immunization with paraflagellar rod proteins requires T cell, but not B cell, function.

    Science.gov (United States)

    Miller, M J; Wrightsman, R A; Stryker, G A; Manning, J E

    1997-06-01

    Previous studies have shown that immunization of mice with the paraflagellar rod proteins (PAR) of Trypanosoma cruzi induces an immune response capable of protecting mice against an otherwise lethal challenge with this parasite. Herein, we define immunologic responses that do or do not play a critical role in PAR-mediated protection. Firstly, PAR-immunized Ab-deficient (muMT) strain mice survived an otherwise lethal T. cruzi challenge, indicating that a B cell response is not required for PAR-induced immunity. However, beta2m -/- mice, which are severely deficient in MHC class I and TCR alphabeta+ CD8+ CD4- T cells, did not survive challenge infection following PAR immunization, indicating that MHC class I/CD8+ T cell function is necessary for protection induced by PAR immunization. Surprisingly, PAR-immunized mice depleted of CD4+ T cells survived a T. cruzi challenge for >84 days postinfection while maintaining a parasitemia that is generally thought to be lethal (i.e., >10(6) trypomastigotes/ml), thus associating CD4+ T cell function with the process of parasite clearance. Consistent with this association, CD4+ T cells from PAR-immunized mice released INF-gamma and stimulated T. cruzi-infected macrophages to release nitric oxide. The importance of IFN-gamma in PAR-induced protective immunity is further indicated by the observation that PAR-immunized INF-gamma knockout mice developed an extremely high parasitemia and did not survive a challenge infection. Thus, while Ab-mediated immune mechanisms are not required for protection induced by PAR immunization, T cell responses are necessary for both elimination of bloodstream parasites and survival.

  13. Functional and structural insights revealed by molecular dynamics simulations of an essential RNA editing ligase in Trypanosoma brucei.

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    Rommie E Amaro

    Full Text Available RNA editing ligase 1 (TbREL1 is required for the survival of both the insect and bloodstream forms of Trypanosoma brucei, the parasite responsible for the devastating tropical disease African sleeping sickness. The type of RNA editing that TbREL1 is involved in is unique to the trypanosomes, and no close human homolog is known to exist. In addition, the high-resolution crystal structure revealed several unique features of the active site, making this enzyme a promising target for structure-based drug design. In this work, two 20 ns atomistic molecular dynamics (MD simulations are employed to investigate the dynamics of TbREL1, both with and without the ATP substrate present. The flexibility of the active site, dynamics of conserved residues and crystallized water molecules, and the interactions between TbREL1 and the ATP substrate are investigated and discussed in the context of TbREL1's function. Differences in local and global motion upon ATP binding suggest that two peripheral loops, unique to the trypanosomes, may be involved in interdomain signaling events. Notably, a significant structural rearrangement of the enzyme's active site occurs during the apo simulations, opening an additional cavity adjacent to the ATP binding site that could be exploited in the development of effective inhibitors directed against this protozoan parasite. Finally, ensemble averaged electrostatics calculations over the MD simulations reveal a novel putative RNA binding site, a discovery that has previously eluded scientists. Ultimately, we use the insights gained through the MD simulations to make several predictions and recommendations, which we anticipate will help direct future experimental studies and structure-based drug discovery efforts against this vital enzyme.

  14. The glycosylphosphatidylinositol-PLC in Trypanosoma brucei forms a linear array on the exterior of the flagellar membrane before and after activation.

    Science.gov (United States)

    Hanrahan, Orla; Webb, Helena; O'Byrne, Robert; Brabazon, Elaine; Treumann, Achim; Sunter, Jack D; Carrington, Mark; Voorheis, H Paul

    2009-06-01

    Bloodstream forms of Trypanosoma brucei contain a glycosylphosphatidylinositol-specific phospholipase C (GPI-PLC) that cleaves the GPI-anchor of the variable surface glycoprotein (VSG). Its location in trypanosomes has been controversial. Here, using confocal microscopy and surface labelling techniques, we show that the GPI-PLC is located exclusively in a linear array on the outside of the flagellar membrane, close to the flagellar attachment zone, but does not co-localize with the flagellar attachment zone protein, FAZ1. Consequently, the GPI-PLC and the VSG occupy the same plasma membrane leaflet, which resolves the topological problem associated with the cleavage reaction if the VSG and the GPI-PLC were on opposite sides of the membrane. The exterior location requires the enzyme to be tightly regulated to prevent VSG release under basal conditions. During stimulated VSG release in intact cells, the GPI-PLC did not change location, suggesting that the release mechanism involves lateral diffusion of the VSG in the plane of the membrane to the fixed position of the GPI-PLC.

  15. Effect of oregano (Origanum vulgare L.) and thyme (Thymus vulgaris L.) essential oils on Trypanosoma cruzi (Protozoa: Kinetoplastida) growth and ultrastructure.

    Science.gov (United States)

    Santoro, Giani F; das Graças Cardoso, Maria; Guimarães, Luiz Gustavo L; Salgado, Ana Paula S P; Menna-Barreto, Rubem F S; Soares, Maurilio J

    2007-03-01

    In the present work, we have investigated the effect of essential oils obtained from Origanum vulgare L. (oregano) and Thymus vulgaris L. (thyme) on growth and ultrastructure of diverse evolutive forms of Trypanosoma cruzi. Culture epimastigotes and bloodstream trypomastigotes were incubated for 24 h with different concentrations of oregano or thyme essential oils and with thymol (the main constituent of thyme), and the inhibitory concentration (IC)(50) was determined by cell counting. Crude extract of oregano essential oil inhibited epimastigote growth (IC(50)/24 h = 175 microg/ml) and also induced trypomastigote lysis (IC(50)/24 h = 115 microg/ml). Thyme essential oil presented IC(50)/24 h values of 77 microg/ml for epimastigotes and 38 mug/ml for trypomastigotes, while treatment with thymol resulted in an IC(50)/24 h of 62 microg/ml for epimastigotes and 53 microg/ml for trypomastigotes. Scanning electron microscopy of treated cells showed few morphological alterations at the plasma membrane. Observation by transmission electron microscopy showed cytoplasmic swelling with occasional morphological alterations in plasma and flagellar membrane. Our data indicate that oregano and thyme essential oils are effective against T. cruzi, with higher activity of thyme, and that thymol may be the main component responsible for the trypanocidal activity.

  16. Dynein Light Chain LC8 Is Required for RNA Polymerase I-Mediated Transcription in Trypanosoma brucei, Facilitating Assembly and Promoter Binding of Class I Transcription Factor A.

    Science.gov (United States)

    Kirkham, Justin K; Park, Sung Hee; Nguyen, Tu N; Lee, Ju Huck; Günzl, Arthur

    2016-01-01

    Dynein light chain LC8 is highly conserved among eukaryotes and has both dynein-dependent and dynein-independent functions. Interestingly, LC8 was identified as a subunit of the class I transcription factor A (CITFA), which is essential for transcription by RNA polymerase I (Pol I) in the parasite Trypanosoma brucei. Given that LC8 has never been identified with a basal transcription factor and that T. brucei relies on RNA Pol I for expressing the variant surface glycoprotein (VSG), the key protein in antigenic variation, we investigated the CITFA-specific role of LC8. Depletion of LC8 from mammalian-infective bloodstream trypanosomes affected cell cycle progression, reduced the abundances of rRNA and VSG mRNA, and resulted in rapid cell death. Sedimentation analysis, coimmunoprecipitation of recombinant proteins, and bioinformatic analysis revealed an LC8 binding site near the N terminus of the subunit CITFA2. Mutation of this site prevented the formation of a CITFA2-LC8 heterotetramer and, in vivo, was lethal, affecting assembly of a functional CITFA complex. Gel shift assays and UV cross-linking experiments identified CITFA2 as a promoter-binding CITFA subunit. Accordingly, silencing of LC8 or CITFA2 resulted in a loss of CITFA from RNA Pol I promoters. Hence, we discovered an LC8 interaction that, unprecedentedly, has a basal function in transcription.

  17. Treatment and Outcomes in Carbapenem-resistant Klebsiella pneumoniae Bloodstream Infections

    Science.gov (United States)

    Neuner, Elizabeth A.; Yeh, Jun-Yen; Hall, Gerri S.; Sekeres, Jennifer; Endimiani, Andrea; Bonomo, Robert A.; Shrestha, Nabin K.; Fraser, Thomas G.; van Duin, David

    2010-01-01

    Carbapenem-resistant K. pneumoniae (CR-Kp) is an emerging multi-drug resistant nosocomial pathogen. This is a retrospective chart review describing the outcomes and treatment of 60 cases of CR-Kp bloodstream infections. All CR-Kp isolated from blood cultures were identified retrospectively from the microbiology laboratory from January 2007 to May 2009. Clinical information was collected from the electronic medical record. Patients with 14 day-hospital mortality were compared to those who survived 14 days. The all-cause in-hospital and 14-day mortality for all 60 CR-Kp bloodstream infections was 58.3% and 41.7%, respectively. In this collection, 98% of tested isolates were susceptible in vitro to tigecycline, compared to 86% to colistimethate, 45% to amikacin and 22% to gentamicin. Nine patients died prior to cultures being finalized, and received no therapy active against CR-Kp. In the remaining 51 patients, those who survived to day14 (n=35) were compared to non-survivor at day 14 (n=16). These patients were characterized by both chronic disease and acute illness. The 90-day readmission rate for hospital survivors was 72%. Time to active therapy was not significantly different between survivors and non-survivors, and hospital mortality was also similar regardless of therapy chosen. Pitt bacteremia score was the only significant factor associated with mortality in Cox regression analysis. In summary, CR-Kp bloodstream infections occur in patients who are chronically and acutely ill. They are associated with high 14-day mortality and poor outcomes regardless of tigecycline or other treatment regimens were selected. PMID:21396529

  18. Epidemic increase in Salmonella bloodstream infection in children, Bwamanda, the Democratic Republic of Congo.

    Science.gov (United States)

    Phoba, M-F; De Boeck, H; Ifeka, B B; Dawili, J; Lunguya, O; Vanhoof, R; Muyembe, J-J; Van Geet, C; Bertrand, S; Jacobs, J

    2014-01-01

    Salmonella enterica is the leading cause of bloodstream infection in children in sub-Saharan Africa, but few data are available from Central-Africa. We documented during the period November 2011 to May 2012 an epidemic increase in invasive Salmonella bloodstream infections in HGR Bwamanda, a referral hospital in Equateur Province, DR Congo. Salmonella spp. represented 90.4 % (103 out of 114) of clinically significant blood culture isolates and comprised Salmonella Typhimurium (54.4 %, 56 out of 103), Salmonella Enteritidis (28.2 %, 29 out of 103) and Salmonella Typhi (17.5 %, 18 out of 103), with Salmonella Enteritidis accounting for most of the increase. Most (82 out of 103, 79.6 %) isolates were obtained from children infected with Salmonella Typhimurium and Salmonella Enteritidis were 14 months (14 days to 64 years) and 19 months (3 months to 8 years) respectively. Clinical presentation was non-specific; the in-hospital case fatality rate was 11.1 %. More than two thirds (69.7 %, 53 out of 76) of children infection. Most (83/85, 97.6 %) non-typhoid Salmonella isolates as well as 6/18 (33.3 %) Salmonella Typhi isolates were multidrug resistant (i.e. resistant to the first-line oral antibiotics amoxicillin, trimethoprim-sulfamethoxazole and chloramphenicol), one (1.0 %) Salmonella Typhimurium had decreased ciprofloxacin susceptibility owing to a point mutation in the gyrA gene (Gly81Cys). Multilocus variable-number tandem-repeat (MLVA) analysis of the Salmonella Enteritidis isolates revealed closely related patterns comprising three major and four minor profiles, with differences limited to one out of five loci. These data show an epidemic increase in clonally related multidrug-resistant Salmonella bloodstream infection in children in DR Congo.

  19. Control method exploration of nosocomial bloodstream infection and its effect evaluation

    Institute of Scientific and Technical Information of China (English)

    CHAI Wen-zhao; WANG Xiao-ting; ZHOU Jiong; LI Xin; LUO Hong-bo; LIU Da-wei

    2012-01-01

    Background Currently,slightly more than 50% of bloodstream infections (BSIs) are hospital acquired.When these infections occur in patients in intensive care units,they are associated with a high mortality rate,additional hospital days and excess hospital costs.Because of multifactor of nosocomial BSIs,measurements of control nosocomial BSIs are wide variety and lead to some confusion in practice.The aim of this study was to explore special way in accordance with self-hospital base on common principle.Methods In one ward of the Intensive Care Unit,Peking Union Medical College Hospital,at first,we divided the all operation about bloodstream way into three sections used as keypoints.By surveying keypoints respectively,some operation faults of blood way were discovered.For decreasing the mobidity of nosocomial BSls,some intervention measurements were executed.The rate of nosocomial BSIs was analyzed by chi-square test.Results According to the statistics from January to June,we received and cured 618 patients in total; among them,there were 13 cases of nosocomial BSI and the average occurrence was 2.3 cases/month.After intervention measurements from July to December 2011,we received and cured 639 patients in total with seven cases of nosocomial BSI,and the average occurrence was 1.2 cases/month (P <0.05).From January to April 2012,no nosocomial BSI occurred in the investigated ward.Conclusion Removing the operation faults of bloodstream way might decrease the nosocomial BSI rapidly and efficiently by utilizing a key point survey.

  20. EFFECT OF INFLIXIMAB ON PARAMETERS OF REMODELING OF ARTERIAL BLOODSTREAM, RANKL AND OSTEOPROTEGERIN LEVELS IN PATIENTS WITH RHEUMATOID ARTHRITIS

    Directory of Open Access Journals (Sweden)

    Larisa Aleksandrovna Knyazeva

    2013-01-01

    Full Text Available Objective. To study the effect of infliximab (INF on serum levels of RANKL and osteoprotegerin (OPG, as well as on structural and functional properties of the vascular wall in patients with rheumatoid arthritis (RA.Material and Methods. A total of 79 RA patients who corresponded to the classification criteria ACR (1987 or ACR/EULAR (2010 and were seronegative for IgM rheumatoid factor (RF were examined. The mean age of patients was 43.6±8.5 years. The serum levels of OPG and RANKL were determined by ELISA (Biomedica, Austria; the common carotid arteries (CCAs were visualized using an Acuson X/10 ultrasonic complex equipped with a 7 MHz linear sensor in the β-mode prior to therapy and after 12-month therapy with INF.Results and Discussion. An increased OPG level was observed mostly in patients with RA duration up to 1 year; an increase in RANKL level was pronounced stronger in patients with PA duration over 2 years. The disturbance of structural and functional properties of the arterial bloodstream was revealed, manifesting itself as an increase in the intimamedia complex thickness, diameter and rigidity index of CCA that were stronger pronounced in patients with late onset RA. A correlation analysis showed the presence of reliable relationship between the RANKL and OPG levels and CCA remodeling parameters. INF therapy showed high clinical effectiveness and correction effect on the RANKL/OPG system. In addition, it was accompanied by a reduction of signs of CCA remodeling, which was stronger pronounced in patients with early RA.Conclusion. The results prove the reasonability of using INF at early stages of RA in order to optimize the therapy and achieve more efficient control of cardiovascular complications.

  1. Bacillus cereus Bloodstream Infection in a Preterm Neonate Complicated by Late Meningitis

    Directory of Open Access Journals (Sweden)

    Toshinobu Horii

    2012-01-01

    Full Text Available Central nervous system infections caused by Bacillus cereus have rarely been reported in infants. In this paper, the case of a 2-month-old low-birth-weight female who developed meningitis 45 days after resolution of a bloodstream infection (BSI is described. The pulsed-field gel electrophoresis results revealed that the patterns of both B. cereus isolates responsible for the acute meningitis and for the prior bacteraemic episode were closely related. Although the source of the infection from within the patient was not clear, it is suggested that the B. cereus BSI developed in the neonate was complicated by acute meningitis.

  2. Taurolidine is effective in the treatment of central venous catheter-related bloodstream infections in cancer patients.

    Science.gov (United States)

    Koldehoff, M; Zakrzewski, J L

    2004-11-01

    Taurolidine is an antimicrobial agent that was originally used in the local treatment of peritonitis and was shown to be effective in the prevention of catheter-related bloodstream infections (CR-BSI). In this pilot study, we used taurolidine solution as an intravenous (i.v.) lock into the totally implantable intravascular devices of 11 consecutive oncological patients with catheter-related bloodstream infections not responding to systemic antimicrobial chemotherapy. All patients recovered completely from the infection. No adverse drug effects were seen. Three patients were successfully retreated for a recurrent infection. Our data suggest a beneficial role of taurolidine i.v. lock for the therapy of catheter-related bloodstream infections in oncological patients. Taurolidine i.v. lock application is feasible and could especially be useful in infections resistant to antibiotic chemotherapy.

  3. Trypanosoma cruzi: single cell live imaging inside infected tissues

    Science.gov (United States)

    Ferreira, Bianca Lima; Orikaza, Cristina Mary; Cordero, Esteban Mauricio

    2016-01-01

    Summary Although imaging the live Trypanosoma cruzi parasite is a routine technique in most laboratories, identification of the parasite in infected tissues and organs has been hindered by their intrinsic opaque nature. We describe a simple method for in vivo observation of live single‐cell Trypanosoma cruzi parasites inside mammalian host tissues. BALB/c or C57BL/6 mice infected with DsRed‐CL or GFP‐G trypomastigotes had their organs removed and sectioned with surgical blades. Ex vivo organ sections were observed under confocal microscopy. For the first time, this procedure enabled imaging of individual amastigotes, intermediate forms and motile trypomastigotes within infected tissues of mammalian hosts. PMID:26639617

  4. Muscle Releases Alpha-Sarcoglycan Positive Extracellular Vesicles Carrying miRNAs in the Bloodstream.

    Directory of Open Access Journals (Sweden)

    Michele Guescini

    Full Text Available In the past few years, skeletal muscle has emerged as an important secretory organ producing soluble factors, called myokines, that exert either autocrine, paracrine or endocrine effects. Moreover, recent studies have shown that muscle releases microRNAs into the bloodstream in response to physical exercise. These microRNAs affect target cells, such as hormones and cytokines. The mechanisms underlying microRNA secretion are poorly characterized at present. Here, we investigated whether muscle tissue releases extracellular vesicles (EVs, which carry microRNAs in the bloodstream under physiological conditions such as physical exercise. Using density gradient separation of plasma from sedentary and physically fit young men we found EVs positive for TSG101 and alpha-sarcoglycan (SGCA, and enriched for miR-206. Cytometric analysis showed that the SGCA+ EVs account for 1-5% of the total and that 60-65% of these EVs were also positive for the exosomal marker CD81. Furthermore, the SGCA-immuno captured sub-population of EVs exhibited higher levels of the miR-206/miR16 ratio compared to total plasma EVs. Finally, a significant positive correlation was found between the aerobic fitness and muscle-specific miRNAs and EV miR-133b and -181a-5p were significantly up-regulated after acute exercise. Thus, our study proposes EVs as a novel means of muscle communication potentially involved in muscle remodeling and homeostasis.

  5.   Bloodstream Bacterial Pathogens and their Antibiotic Resistance Pattern in Dhahira Region, Oman

    Directory of Open Access Journals (Sweden)

    PP Geethanjali

    2011-07-01

    Full Text Available Objectives: To describe the epidemiological, clinical, microbiological characteristics and antimicrobial resistance pattern of Bloodstream infections in Dhahira region, Oman.Methods: Clinical data was collected from all patients with positive blood cultures for two years period. Standard laboratory methods were used for blood culture. Antibiotic sensitivity was tested using Kirby-Bauer disc diffusion method.Results: Of the 360 bacterial pathogens isolated from 348 patients, 57.8�0were gram-positive and 42.2�0were gram-negative. The common isolates were: Streptococcus species 76 (21.1� coagulase-negative Staphylococci 75 (20.8� Escherichia coli 43 (11.9� Staphylococcus aureus 41 (11.4� Overall, mortality was 21.3�0(74/348. Staphylococcus species (Staphylococcus aureus and CoNS were more commonly resistant to Trimethoprim/ Sulphamethoxazole (35.3�20and Penicillin (25.9� Streptococcus species were resistant to Trimethoprim/Sulphamethoxazole (39.1�20and Erythromycin (19.6�Conclusion: Bloodstream infections are important causes of morbidity and mortality in our patients, especially among chronically ill elderly adult males. Prescription of proven resistant antibiotics to suspected bacteremic patients needs attention in Dhahira region.

  6. Coordinated Molecular Cross-Talk between Staphylococcus aureus, Endothelial Cells and Platelets in Bloodstream Infection

    Directory of Open Access Journals (Sweden)

    Carolina D. Garciarena

    2015-12-01

    Full Text Available Staphylococcus aureus is an opportunistic pathogen often carried asymptomatically on the human body. Upon entry to the otherwise sterile environment of the cardiovascular system, S. aureus can lead to serious complications resulting in organ failure and death. The success of S. aureus as a pathogen in the bloodstream is due to its ability to express a wide array of cell wall proteins on its surface that recognise host receptors, extracellular matrix proteins and plasma proteins. Endothelial cells and platelets are important cells in the cardiovascular system and are a major target of bloodstream infection. Endothelial cells form the inner lining of a blood vessel and provide an antithrombotic barrier between the vessel wall and blood. Platelets on the other hand travel throughout the cardiovascular system and respond by aggregating around the site of injury and initiating clot formation. Activation of either of these cells leads to functional dysregulation in the cardiovascular system. In this review, we will illustrate how S. aureus establish intimate interactions with both endothelial cells and platelets leading to cardiovascular dysregulation.

  7. Risk factors for nosocomial bloodstream infection caused by multidrug resistant gram-negative bacilli in pediatrics

    Directory of Open Access Journals (Sweden)

    Mariana V. Arnoni

    2007-04-01

    Full Text Available The aim of this study was to identify the risk factors for nosocomial bloodstream infections by multidrug resistant Gram-negative bacilli. From November 2001 to December 2003, in the Pediatric Department of the Santa Casa de São Paulo, a retrospective case-control study was developed concerning patients who had nosocomial bloodstream infection caused by Gram-negative bacilli. Patients with multidrug resistant infections were designated as case patients, and control patients were those with an infection that did not meet the criteria for multidrug resistance. Previous use of central venous catheter and previous use of vancomycin plus third generation cephalosporins were associated to a higher chance of infections by multidrug resistant Gram-negative bacilli (Odds ratio - 5.8 and 5.2, respectively. Regarding sensitivity of the isolated agents, 47.8% were multidrug resistant, 54.2% were Klebsiella spp. ESBL producers and 36.4% were imipenem resistant Pseudomonas aeruginosa. The lethality rate was 36.9% in the studied cases and this rate was significantly higher in the group of patients with multidrug resistant infections (p=0.013. Risk factor identification as well as the knowledge of the susceptibility of the nosocomial infectious agents gave us the possibility to perform preventive and control strategies to reduce the costs and mortality related to these infections.

  8. Exchange of peripherally inserted central catheters is associated with an increased risk for bloodstream infection.

    Science.gov (United States)

    McCoy, Michael; Bedwell, Susan; Noori, Shahab

    2011-06-01

    It is not uncommon that the peripherally inserted central catheter (PICC) needs to be replaced either due to blockage or migration to a peripheral position. In such circumstances, there are two methods of PICC placement: new-site insertion and exchange by using the old PICC as a guide wire. Our objective was to investigate risk of infection associated with the exchange method. In this retrospective study, data on all PICC insertions in the neonatal intensive care unit in 2004 to 2008 were obtained. In the population who needed removal of existing PICC and insertion of a new one, we compared central line-associated bloodstream infection (CLABSI) within 1 week of insertion between the two insertion methods. Of 1148 PICC insertions reviewed, 164 (103 new-site and 61 exchange insertions) were performed after removal of a blocked/malpositioned PICC and therefore comprised the study population. The rate of CLABSI was higher in the exchange method (9.8% versus 1%, P exchange method (odds ratio 25.2, 95% confidence interval: 2.17 to 292.98; P = 0.01). In infants, insertion of PICCs using the exchange method carries an increased risk of bloodstream infection.

  9. Trypanosoma caninum n. sp. (Protozoa: Kinetoplastida) isolated from intact skin of a domestic dog ( Canis familiaris) captured in Rio de Janeiro, Brazil.

    Science.gov (United States)

    Madeira, M F; Sousa, M A; Barros, J H S; Figueiredo, F B; Fagundes, A; Schubach, A; DE Paula, C C; Faissal, B N S; Fonseca, T S; Thoma, H K; Marzochi, M C A

    2009-04-01

    An unknown Trypanosoma species was isolated from an axenic culture of intact skin from a domestic dog captured in Rio de Janeiro, Brazil, which was co-infected with Leishmania (Viannia) braziliensis. Giemsa-stained smears of cultures grown in different media revealed the presence of epimastigotes, trypomastigotes, spheromastigotes, transitional stages, and dividing forms (epimastigotes or spheromastigotes). The highest frequency of trypomastigotes was observed in RPMI (15.2%) and DMEM (9.2%) media containing 5% FCS, with a mean length of these forms of 43.0 and 36.0 mum, respectively. Molecular analysis by sequential application of PCR assays indicated that this trypanosome differs from Trypanosoma cruzi and T. rangeli when specific primers were applied. On the other hand, a PCR strategy targeted to the D7 domain of 24salpha rDNA, using primers D75/D76, amplified products of about 250 bp in that isolate (stock A-27), different from the amplification products obtained with T. cruzi and T. rangeli. This organism differs from T. cruzi mainly by the size of its trypomastigote forms and kinetoplasts and the absence of infectivity for macrophages and triatomine bugs. It is also morphologically distinct from salivarian trypanosomes reported in Brazil. Isoenzyme analysis at 8 loci demonstrated a very peculiar banding pattern clearly distinct from those of T. rangeli and T. cruzi. We conclude that this isolate is a new Trypanosoma species. The name T. caninum is suggested.

  10. Genetically attenuated Trypanosoma cruzi parasites as a potential vaccination tool

    OpenAIRE

    2012-01-01

    Chagas disease is the clinical manifestation of the infection produced by the parasite Trypanosoma cruzi. Currently there is no vaccine to prevent this disease and the protection attained with vaccines containing non-replicating parasites is limited. Genetically attenuated trypanosomatid parasites can be obtained by deletion of selected genes. Gene deletion takes advantage of the fact that this parasite can undergo homologous recombination between endogenous and foreign DNA sequences artifici...

  11. Surface antigens of metacyclic trypomastigotes of Trypanosoma cruzi.

    OpenAIRE

    1983-01-01

    The surface antigen makeup of metacyclic trypomastigote forms of strain G of Trypanosoma cruzi, which produce a subpatent infection in mice, differed from those of the virulent strains Y and CL. A 100,000-molecular-weight protein, barely detectable on the Y or CL cell surface, appeared as the main surface antigen of the G metacyclic trypomastigotes. In addition, the G metacyclic forms differed from those of the virulent strains in their susceptibility to complement-mediated immunolysis.

  12. Genitourinary changes in hamsters infected and reinfected with Trypanosoma cruzi

    OpenAIRE

    Cabrine-Santos Marlene; Santos Vitorino Modesto dos; Lima Marcus Aurelho; Abreu Marta Elena Araújo de; Lages-Silva Eliane; Ramírez Luís Eduardo

    2003-01-01

    Authors describe genitourinary changes in male hamsters infected and reinfected with Trypanosoma cruzi. Changes in genital organs have been described in human and in experimental chagasic infection. Genital dysfunctions in chronic chagasic patients affect ejaculation, libido and sexual potency, and testis biopsies may show arrested maturation of germ cells, oligozoospermia and azoospermia. Sixty-five male hamsters were inoculated and reinoculated with 2x10³ trypomastigotes of T. cruzi VIC str...

  13. Regulation and spatial organization of PCNA in Trypanosoma brucei

    Energy Technology Data Exchange (ETDEWEB)

    Kaufmann, Doris; Gassen, Alwine [University of Munich (LMU), Department Biology I, Genetics, Grosshaderner Str. 2-4, 82152 Martinsried (Germany); Maiser, Andreas; Leonhardt, Heinrich [University of Munich (LMU), Department Biology II, Grosshaderner Str. 2-4, 82152 Martinsried (Germany); Janzen, Christian J., E-mail: christian.janzen@uni-wuerzburg.de [University of Munich (LMU), Department Biology I, Genetics, Grosshaderner Str. 2-4, 82152 Martinsried (Germany)

    2012-03-23

    Highlights: Black-Right-Pointing-Pointer Characterization of the proliferating cell nuclear antigen in Trypanosoma brucei (TbPCNA). Black-Right-Pointing-Pointer TbPCNA is a suitable marker to detect replication in T. brucei. Black-Right-Pointing-Pointer TbPCNA distribution and regulation is different compared to closely related parasites T. cruzi and Leishmania donovani. -- Abstract: As in most eukaryotic cells, replication is regulated by a conserved group of proteins in the early-diverged parasite Trypanosoma brucei. Only a few components of the replication machinery have been described in this parasite and regulation, sub-nuclear localization and timing of replication are not well understood. We characterized the proliferating cell nuclear antigen in T. brucei (TbPCNA) to establish a spatial and temporal marker for replication. Interestingly, PCNA distribution and regulation is different compared to the closely related parasites Trypanosoma cruzi and Leishmania donovani. TbPCNA foci are clearly detectable during S phase of the cell cycle but in contrast to T. cruzi they are not preferentially located at the nuclear periphery. Furthermore, PCNA seems to be degraded when cells enter G2 phase in T. brucei suggesting different modes of replication regulation or functions of PCNA in these closely related eukaryotes.

  14. Vaccination of dogs with Trypanosoma rangeli induces antibodies against Trypanosoma cruzi in a rural area of Córdoba, Argentina.

    Science.gov (United States)

    Basso, Beatriz; Marini, Vanina; Gauna, Diego; Frias, Maria

    2016-04-01

    Dogs play a major role in the domestic cycle of Trypanosoma cruzi, acting as reservoirs. In a previous work we have developed a model of vaccination of dogs in captivity with nonpathogenic Trypanosoma rangeli epimastigotes, resulting in the production of protective antibodies against T. cruzi, with dramatic decrease of parasitaemia upon challenge with 100,000 virulent forms of this parasite. The aim of this work was to evaluate the immunogenicity of this vaccine in dogs living in a rural area. Domestic dogs, free from T. cruzi infection, received three immunisations with fixed T. rangeli epimastigotes. Dogs were not challenged with T. cruzi, but they were left in their environment. This immunisation induced antibodies against T. cruzi for more than three years in dogs in their natural habitat, while control dogs remained serologically negative.

  15. Incidence of Trypanosoma cruzi Infection in Triatomines Collected at Indio Mountains Research Station

    OpenAIRE

    2015-01-01

    Chagas disease, caused by the parasite Trypanosoma cruzi, is an emerging infectious disease in the United States. In our study, 24 out of 39 triatomines, from the specie Triatoma rubida, were infected with Trypanosoma cruzi. Additionally, only the genotype TcI was characterized among the parasite specimens. Improved knowledge of local epidemiology is needed to prevent transmission of Chagas disease.

  16. Semisolid liver infusion tryptose supplemented with human urine allows growth and isolation of Trypanosoma cruzi and Trypanosoma rangeli clonal lineages

    OpenAIRE

    2016-01-01

    Abstract: INTRODUCTION This work shows that 3% (v/v) human urine (HU) in semisolid Liver Infusion Tryptose (SSL) medium favors the growth of Trypanosoma cruzi and T. rangeli. METHODS Parasites were plated as individual or mixed strains on SSL medium and on SSL medium with 3% human urine (SSL-HU). Isolate DNA was analyzed using polymerase chain reaction (PCR) and pulsed-field gel electrophoresis (PFGE). RESULTS SSL-HU medium improved clone isolation. PCR revealed that T. cruzi strains pr...

  17. Humoral immune response of horses experimentally infected with Trypanosoma evansi/ Resposta imune humoral de eqüinos infectados experimentalmente com Trypanosoma evansi

    Directory of Open Access Journals (Sweden)

    Lúcia Padilha Cury Thomaz de Aquino

    2001-05-01

    Full Text Available Six adult horses were experimentally infected with Trypanosoma evansi (106 parasites. Three other adult horses served as negative control. Serum samples of the experimentally infected horses with T. evansi and non-infected controls horses were obtained before inoculation, and daily thereafter until 14 days post infection (DPI. After that time the serum samples were obtained weekly. Sera of the infected and non-infected control horses was tested by indirect fluorescent antibody test (IFAT and enzyme-linked immunosorbent assay (ELISA for the detection of antibodies against T. evansi. Both ELISA and IFAT detected trypanosomal antibodies shortly after infection and showed progressive increases in antibodies levels during early stages of infection. The responses started on the eighth and eleventh DPI. Maximum IFAT and ELISA values were reached after four weeks of infection and were maintained at this level until the end of the period of study.Seis eqüinos foram inoculados com 106 tripomastigota sangüícolas de Trypanosoma evansi. Três outros animais foram mantidos como testemunhas. Amostras de soro sangüíneo foram obtidas de todos os animais, antes da inoculação, e diariamente até o 14º dia pós inoculação (DPI; após este período uma vez por semana. Pesquisa de anticorpos anti- T. evansi, foram realizadas através da reação de imunofluorescência indireta (RIFI e do ensaio de imunoabsorção enzimática (ELISA. A resposta imune humoral, detectada através da RIFI e do ELISA, iniciou-se, em média, a partir do oitavo DPI, alcançando títulos máximos após quatro semanas de evolução, e os titulos de anticorpos anti- T. evansi mantiveram-se elevadas até o término das observações.

  18. Assessment of the Anti-Protozoal Activity of Crude Carica papaya Seed Extract against Trypanosoma cruzi

    Directory of Open Access Journals (Sweden)

    Karla Y. Acosta-Viana

    2013-10-01

    Full Text Available In order to determine the in vivo activity against the protozoan Trypanosoma cruzi, two doses (50 and 75 mg/kg of a chloroform extract of Carica papaya seeds were evaluated compared with a control group of allopurinol. The activity of a mixture of the three main compounds (oleic, palmitic and stearic acids in a proportion of 45.9% of oleic acid, 24.1% of palmitic and 8.52% of stearic acid previously identified in the crude extract of C. papaya was evaluated at doses of 100, 200 and 300 mg/kg. Both doses of the extracts were orally administered for 28 days. A significant reduction (p < 0.05 in the number of blood trypomastigotes was observed in animals treated with the evaluated doses of the C. papaya extract in comparison with the positive control group (allopurinol 8.5 mg/kg. Parasitemia in animals treated with the fatty acids mixture was also significantly reduced (p < 0.05, compared to negative control animals. These results demonstrate that the fatty acids identified in the seed extracts of C. papaya (from ripe fruit are able to reduce the number of parasites from both parasite stages, blood trypomastigote and amastigote (intracellular stage.

  19. The extraordinary mitochondrion and unusual citric acid cycle in Trypanosoma brucei.

    Science.gov (United States)

    van Hellemond, J J; Opperdoes, F R; Tielens, A G M

    2005-11-01

    African trypanosomes are parasitic protozoa that cause sleeping sickness and nagana. Trypanosomes are not only of scientific interest because of their clinical importance, but also because these protozoa contain several very unusual biological features, such as their specially adapted mitochondrion and the compartmentalization of glycolytic enzymes in glycosomes. The energy metabolism of Trypanosoma brucei differs significantly from that of their hosts and changes drastically during the life cycle. Despite the presence of all citric acid cycle enzymes in procyclic insect-stage T. brucei, citric acid cycle activity is not used for energy generation. Recent investigations on the influence of substrate availability on the type of energy metabolism showed that absence of glycolytic substrates did not induce a shift from a fermentative metabolism to complete oxidation of substrates. Apparently, insect-stage T. brucei use parts of the citric acid cycle for other purposes than for complete degradation of mitochondrial substrates. Parts of the cycle are suggested to be used for (i) transport of acetyl-CoA units from the mitochondrion to the cytosol for the biosynthesis of fatty acids, (ii) degradation of proline and glutamate to succinate, (iii) generation of malate, which can then be used for gluconeogenesis. Therefore the citric acid cycle in trypanosomes does not function as a cycle.

  20. Interactions between Trypanosoma cruzi secreted proteins and host cell signaling pathways

    Directory of Open Access Journals (Sweden)

    Renata Watanabe Costa

    2016-03-01

    Full Text Available Chagas disease is one of the prevalent neglected tropical diseases, affecting at least 6-7 million individuals in Latin America. It is caused by the protozoan parasite Trypanosoma cruzi (T. cruzi, which is transmitted to vertebrate hosts by blood-sucking insects. After infection, the parasite invades and multiplies in the myocardium, leading to acute myocarditis that kills around 5% of untreated individuals. T. cruzi secretes proteins that manipulate multiple host cell signaling pathways to promote host cell invasion. The primary secreted lysosomal peptidase in T. cruzi is cruzipain, which has been shown to modulate the host immune response. Cruzipain hinders macrophage activation during the early stages of infection by interrupting the NF-kB P65 mediated signaling pathway. This allows the parasite to survive and replicate, and may contribute to the spread of infection in acute Chagas disease. Another secreted protein P21, which is expressed in all of the developmental stages of T. cruzi, has been shown to modulate host phagocytosis signaling pathways. The parasite also secretes soluble factors that exert effects on host extracellular matrix, such as proteolytic degradation of collagens. Finally, secreted phospholipase A from T. cruzi contributes to lipid modifications on host cells and concomitantly activates the PKC signaling pathway. Here we present a brief review of the interaction between secreted proteins from T. cruzi and the host cells, emphasizing the manipulation of host signaling pathways during invasion.

  1. Interactions between Trypanosoma cruzi Secreted Proteins and Host Cell Signaling Pathways

    Science.gov (United States)

    Watanabe Costa, Renata; da Silveira, Jose F.; Bahia, Diana

    2016-01-01

    Chagas disease is one of the prevalent neglected tropical diseases, affecting at least 6–7 million individuals in Latin America. It is caused by the protozoan parasite Trypanosoma cruzi, which is transmitted to vertebrate hosts by blood-sucking insects. After infection, the parasite invades and multiplies in the myocardium, leading to acute myocarditis that kills around 5% of untreated individuals. T. cruzi secretes proteins that manipulate multiple host cell signaling pathways to promote host cell invasion. The primary secreted lysosomal peptidase in T. cruzi is cruzipain, which has been shown to modulate the host immune response. Cruzipain hinders macrophage activation during the early stages of infection by interrupting the NF-kB P65 mediated signaling pathway. This allows the parasite to survive and replicate, and may contribute to the spread of infection in acute Chagas disease. Another secreted protein P21, which is expressed in all of the developmental stages of T. cruzi, has been shown to modulate host phagocytosis signaling pathways. The parasite also secretes soluble factors that exert effects on host extracellular matrix, such as proteolytic degradation of collagens. Finally, secreted phospholipase A from T. cruzi contributes to lipid modifications on host cells and concomitantly activates the PKC signaling pathway. Here, we present a brief review of the interaction between secreted proteins from T. cruzi and the host cells, emphasizing the manipulation of host signaling pathways during invasion. PMID:27065960

  2. The PARP promoter of Trypanosoma brucei is developmentally regulated in a chromosomal context

    DEFF Research Database (Denmark)

    Biebinger, S; Rettenmaier, S; Flaspohler, J;

    1996-01-01

    RNA is abundant in procyclic forms and almost undetectable in blood-stream forms. Post-transcriptional mechanisms are mainly responsible for PARP mRNA regulation but results of nuclear run-on experiments suggested that transcription might also be regulated. We measured the activity of genomically-integrated PARP...... not developmentally regulated, but integration at the PARP locus reduced rRNA promoter activity in bloodstream forms. PARP promoter activity was 5-fold down-regulated in bloodstream forms when integrated at either site. Regulation was probably at the level of transcriptional initiation, but elongation through plasmid...

  3. Algorithm for pre-emptive glycopeptide treatment in patients with haematologic malignancies and an Enterococcus faecium bloodstream infection

    NARCIS (Netherlands)

    Zhou, Xuewei; Arends, Jan P; Span, Lambert Fr; Friedrich, Alexander W

    2013-01-01

    INTRODUCTION: Nowadays Enterococcus faecium has become one of the most emerging and challenging nosocomial pathogens. The aim of this study was to determine risk factors in haematology patients who are at risk of an Enterococcus faecium bloodstream infection (BSI) and should be considered for pre-em

  4. The Resistance Phenotype and Molecular Epidemiology of Klebsiella pneumoniae in Bloodstream Infections in Shanghai, China, 2012–2015

    Science.gov (United States)

    Xiao, Shu-zhen; Wang, Su; Wu, Wen-man; Zhao, Sheng-yuan; Gu, Fei-fei; Ni, Yu-xing; Guo, Xiao-kui; Qu, Jie-ming; Han, Li-zhong

    2017-01-01

    Klebsiella pneumoniae (K.pneumoniae) is a common nosocomial pathogen causing bloodstream infections. Antibiotic susceptibility surveillance and molecular characterization will facilitate prevention and management of K. pneumoniae bloodstream infections. K. pneumoniae isolates causing bloodstream infections were consecutively collected between January 2012 and December 2015 in Shanghai. Eighty isolates (20 per year) were randomly selected and enrolled in this study. Drug susceptibility were determined by the disk diffusion method. Polymerase chain reaction (PCR) was employed to detect extended-spectrum β-lactamases (ESBLs), carbapenemases, and seven housekeeping genes of K. pneumoniae. eBURST was used for multi-locus sequence typing (MLST). More than 50% isolates were resistant to cefuroxime, ampicillin-sulbactam, and piperacillin, while carbapenems had lower resistant rates than other antibiotics. Of the 80 isolates, 22 produced ESBLs, and 14 were carbapenemase producers. In the ESBL-producing K. pneumoniae isolates, the most common ESBL genes were blaSHV and blaCTX−M. Thirteen carbapenemase producers harbored blaKPC−2 and one other carried blaNDM−5. ST11 (14/80) was the most frequent sequence type (ST), followed by ST15 (7/80) and ST29 (4/80). Our data revealed high prevalence of antibiotic resistant K. pneumoniae isolates from bloodstream infections but their genetic diversity suggested no clonal dissemination in the region. Also, one K. pneumoniae isolate harbored blaNDM−5 in this study, which was firstly reported in Shanghai. PMID:28280486

  5. Patients with Central Lines — What You Need to Know to Avoid a Bloodstream Infection

    Centers for Disease Control (CDC) Podcasts

    2011-03-01

    This podcast is based on the March, 2011 CDC Vital Signs report which indicates bloodstream infections in patients with central lines are largely preventable when healthcare providers use CDC-recommended infection control steps.  Created: 3/1/2011 by Centers for Disease Control and Prevention (CDC).   Date Released: 3/1/2011.

  6. Bacterial etiology of bloodstream infections and antimicrobial resistance in Dhaka, Bangladesh, 2005–2014

    Directory of Open Access Journals (Sweden)

    Dilruba Ahmed

    2017-01-01

    Full Text Available Abstract Background Bloodstream infections due to bacterial pathogens are a major cause of morbidity and mortality in Bangladesh and other developing countries. In these countries, most patients are treated empirically based on their clinical symptoms. Therefore, up to date etiological data for major pathogens causing bloodstream infections may play a positive role in better healthcare management. The aim of this study was to identify the bacterial pathogens causing major bloodstream infections in Dhaka, Bangladesh and determine their antibiotic susceptibility pattern. Methods From January 2005 to December 2014, a total of 103,679 single bottle blood samples were collected from both hospitalized and domiciliary patients attending Dhaka hospital, icddrb, Bangladesh All the blood samples were processed for culture using a BACT/Alert blood culture machine. Further identification of bacterial pathogens and their antimicrobial susceptibility test were performed using standard microbiological procedures. Results Overall, 13.6% of the cultured blood samples were positive and Gram-negative (72.1% bacteria were predominant throughout the study period. Salmonella Typhi was the most frequently isolated organism (36.9% of samples in this study and a high percentage of those strains were multidrug-resistant (MDR. However, a decreasing trend in the S. Typhi isolation rate was observed and, noticeably, the percentage of MDR S. Typhi isolated declined sharply over the study period. An overall increase in the presence of Gram-positive bacteria was observed, but most significantly we observed the percentage of MDR Gram-positive bacteria to double over the study period. Overall, Gram positive bacteria were more resistant to most of the commonly used antibiotics than Gram-negative bacteria, but the MDR level was high in both groups. Conclusions This study identified the major bacterial pathogens involved with BSI in Dhaka, Bangladesh and also revealed their

  7. Well staged

    Energy Technology Data Exchange (ETDEWEB)

    Budd, Godfrey

    2011-06-15

    Packers Plus Energy Services Inc. has commercially launched QuickFRAC, a multi-stage completition system which can fracture four to five isolated stages in one treatment and set up a record of 23-stage slickwater frac in less than 10 hours. It could take up to 40 days to do 100 fracture treatments with other systems. This technology makes it possible to distribute fluid at each port thanks to the limited entry system. In order to make multiple isolated stages within one treatment zone, each zone includes multiple QuickPORT sleeves with packers on either side. The other technology which made this possible is the repeater port system, it allows them to perform more frac stages. This technology could be useful in the future since the need for stages will be doubling soon with microdarcy shale oil extraction which is more difficult than gas.

  8. Comparative analysis of the kinomes of three pathogenic trypanosomatids: Leishmania major, Trypanosoma brucei and Trypanosoma cruzi

    OpenAIRE

    Ward Pauline N; Worthey Elizabeth A; Parsons Marilyn; Mottram Jeremy C

    2005-01-01

    Abstract Background The trypanosomatids Leishmania major, Trypanosoma brucei and Trypanosoma cruzi cause some of the most debilitating diseases of humankind: cutaneous leishmaniasis, African sleeping sickness, and Chagas disease. These protozoa possess complex life cycles that involve development in mammalian and insect hosts, and a tightly coordinated cell cycle ensures propagation of the highly polarized cells. However, the ways in which the parasites respond to their environment and coordi...

  9. Trading stages

    DEFF Research Database (Denmark)

    Steiner, Uli; Tuljapurkar, Shripad; Coulson, Tim

    2012-01-01

    Interest in stage-and age structured models has recently increased because they can describe quantitative traits such as size that are left out of age-only demography. Available methods for the analysis of effects of vital rates on lifespan in stage-structured models have not been widely applied ...... examples. Much of our approach relies on trading of time and mortality risk in one stage for time and risk in others. Our approach contributes to the new framework of the study of age- and stage-structured biodemography....

  10. Taurolidine lock is superior to heparin lock in the prevention of catheter related bloodstream infections and occlusions.

    Directory of Open Access Journals (Sweden)

    Evelyn D Olthof

    Full Text Available BACKGROUND AND AIMS: Patients on home parenteral nutrition (HPN are at risk for catheter-related complications; mainly infections and occlusions. We have previously shown in HPN patients presenting with catheter sepsis that catheter locking with taurolidine dramatically reduced re-infections when compared with heparin. Our HPN population therefore switched from heparin to taurolidine in 2008. The aim of the present study was to compare long-term effects of this catheter lock strategy on the occurrence of catheter-related bloodstream infections and occlusions in HPN patients. METHODS: Data of catheter-related complications were retrospectively collected from 212 patients who received HPN between January 2000 and November 2011, comprising 545 and 200 catheters during catheter lock therapy with heparin and taurolidine, respectively. We evaluated catheter-related bloodstream infection and occlusion incidence rates using Poisson-normal regression analysis. Incidence rate ratios were calculated by dividing incidence rates of heparin by those of taurolidine, adjusting for underlying disease, use of anticoagulants or immune suppressives, frequency of HPN/fluid administration, composition of infusion fluids, and duration of HPN/fluid use before catheter creation. RESULTS: Bloodstream infection incidence rates were 1.1/year for heparin and 0.2/year for taurolidine locked catheters. Occlusion incidence rates were 0.2/year for heparin and 0.1/year for taurolidine locked catheters. Adjusted incidence ratios of heparin compared to taurolidine were 5.9 (95% confidence interval, 3.9-8.7 for bloodstream infections and 1.9 (95% confidence interval, 1.1-3.1 for occlusions. CONCLUSIONS: Given that no other procedural changes than the catheter lock strategy were implemented during the observation period, these data strongly suggest that taurolidine decreases catheter-related bloodstream infections and occlusions in HPN patients compared with heparin.

  11. APPROACH OF IMPROVING PRECISION IN ULTRASONIC DOPPLER BLOODSTREAM SPEED MEASUREMENT BY CHAOS-BASED FREQUENCY DETECTING

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    It is critical for cerebral vascular disease diagnosis through Doppler to detect the maximum and the minimum of the carotid blood flow speed accurately. A kind of Duffing system under an external periodic power with dump is introduced in the letter, numerical analysis is carried out by four-order Runge-Kutta method. An oscillator array is designed according to the frequency of the ultrasonic wave. When the external signals are inputted, computational algorithm is used to scan the array in turn and analyze the result, and the frequency can be determined. Based on the methods above, detecting the carotid blood flow speed accurately is realized. The Signal-to-Noise Ratio (SNR) of-20.23dB is obtained by the result of experiments. In conclusion, the SNR has been improved and the precision of the measured bloodstream speed has been increased,which can be 0.069% to 0.13%.

  12. Evidence-based measures to prevent central line-associated bloodstream infections: a systematic review

    Directory of Open Access Journals (Sweden)

    Daniele Cristina Perin

    Full Text Available ABSTRACT Objective: to identify evidence-based care to prevent CLABSI among adult patients hospitalized in ICUs. Method: systematic review conducted in the following databases: PubMed, Scopus, Cinahl, Web of Science, Lilacs, Bdenf and Cochrane Studies addressing care and maintenance of central venous catheters, published from January 2011 to July 2014 were searched. The 34 studies identified were organized in an instrument and assessed by using the classification provided by the Joanna Briggs Institute. Results: the studies presented care bundles including elements such as hand hygiene and maximal barrier precautions; multidimensional programs and strategies such as impregnated catheters and bandages and the involvement of facilities in and commitment of staff to preventing infections. Conclusions: care bundles coupled with education and the commitment of both staff and institutions is a strategy that can contribute to decreased rates of central line-associated bloodstream infections among adult patients hospitalized in intensive care units.

  13. Healthcare-associated Gram-negative bloodstream infections: antibiotic resistance and predictors of mortality.

    Science.gov (United States)

    Ergönül, Ö; Aydin, M; Azap, A; Başaran, S; Tekin, S; Kaya, Ş; Gülsün, S; Yörük, G; Kurşun, E; Yeşilkaya, A; Şimşek, F; Yılmaz, E; Bilgin, H; Hatipoğlu, Ç; Cabadak, H; Tezer, Y; Togan, T; Karaoğlan, I; İnan, A; Engin, A; Alışkan, H E; Yavuz, S Ş; Erdinç, Ş; Mulazimoglu, L; Azap, Ö; Can, F; Akalın, H; Timurkaynak, F

    2016-12-01

    This article describes the prevalence of antibiotic resistance and predictors of mortality for healthcare-associated (HA) Gram-negative bloodstream infections (GN-BSI). In total, 831 cases of HA GN-BSI from 17 intensive care units in different centres in Turkey were included; the all-cause mortality rate was 44%. Carbapenem resistance in Klebsiella pneumoniae was 38%, and the colistin resistance rate was 6%. Multi-variate analysis showed that age >70 years [odds ratio (OR) 2, 95% confidence interval (CI) 1.22-3.51], central venous catheter use (OR 2.1, 95% CI 1.09-4.07), ventilator-associated pneumonia (OR 1.9, 95% CI 1.1-3.16), carbapenem resistance (OR 1.8, 95% CI 1.11-2.95) and APACHE II score (OR 1.1, 95% CI 1.07-1.13) were significantly associated with mortality.

  14. Achromobacter Xylosoxidans Bloodstream Infection in Elderly Patient with Hepatocellular Carcinoma: Case Report and Review of Literature.

    Science.gov (United States)

    Raghuraman, Kausalya; Ahmed, Nishat H; Baruah, Frincy K; Grover, Rajesh K

    2015-01-01

    Achromobacter xylosoxidansis a nonfermentative Gram-negative organism, known to cause opportunistic infection in humans. We report a case of septicemia in a 76-year-old male patient with underlying hepatocellular carcinoma due to A. xylosoxidans, which showed a different antimicrobial susceptibility pattern from what is usually reported. From aerobic blood culture of the patient, A. xylosoxidans was isolated which was found to be sensitive to amoxicillin-clavulanic acid, piperacillin-tazobactam, ceftazidime, cefoperazone-sulbactam, meropenem, minocycline, tigecycline, and trimethoprim/sulfamethoxazole. The patient recovered with amoxicillin-clavulanic acid treatment, which was given empirically to the patient. The present case highlights the possible role of amoxicillin-clavulanic acid for treatment of bloodstream infection with A. xylosoxidans.

  15. Achromobacter Xylosoxidans bloodstream infection in elderly patient with Hepatocellular Carcinoma: Case report and review of literature

    Directory of Open Access Journals (Sweden)

    Kausalya Raghuraman

    2015-01-01

    Full Text Available Achromobacter xylosoxidansis a nonfermentative Gram-negative organism, known to cause opportunistic infection in humans. We report a case of septicemia in a 76-year-old male patient with underlying hepatocellular carcinoma due to A. xylosoxidans, which showed a different antimicrobial susceptibility pattern from what is usually reported. From aerobic blood culture of the patient, A. xylosoxidanswas isolated which was found to be sensitive to amoxicillin-clavulanic acid, piperacillin-tazobactam, ceftazidime, cefoperazone-sulbactam, meropenem, minocycline, tigecycline, and trimethoprim/sulfamethoxazole. The patient recovered with amoxicillin-clavulanic acid treatment, which was given empirically to the patient. The present case highlights the possible role of amoxicillin-clavulanic acid for treatment of bloodstream infection with A. xylosoxidans.

  16. Positive deviance as a strategy to prevent and control bloodstream infections in intensive care

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    Francimar Tinoco de Oliveira

    Full Text Available Abstract OBJECTIVE To describe the application of positive deviance as a strategy to prevent and control bloodstream infections. METHOD An intervention study with nursing and medical team members working in an intensive care unit in a university hospital, between June and December 2014. The four steps of the positive defiance methodology were applied: to define, to determine, to discover and to design. RESULTS In 90 days, 188 actions were observed, of these, 36.70% (n=69 were related to catheter dressing. In 81.15% (n=56 of these dressings, the professionals most adhered to the use of flexible sterile cotton-tipped swabs to perform antisepsis at catheter entry sites and fixation dressing. CONCLUSION Positive deviance contributed to the implementation of proposals to improve work processes and team development related to problems identified in central venous catheter care.

  17. Bloodstream-To-Eye Infections Are Facilitated by Outer Blood-Retinal Barrier Dysfunction.

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    Phillip S Coburn

    Full Text Available The blood-retinal barrier (BRB functions to maintain the immune privilege of the eye, which is necessary for normal vision. The outer BRB is formed by tightly-associated retinal pigment epithelial (RPE cells which limit transport within the retinal environment, maintaining retinal function and viability. Retinal microvascular complications and RPE dysfunction resulting from diabetes and diabetic retinopathy cause permeability changes in the BRB that compromise barrier function. Diabetes is the major predisposing condition underlying endogenous bacterial endophthalmitis (EBE, a blinding intraocular infection resulting from bacterial invasion of the eye from the bloodstream. However, significant numbers of EBE cases occur in non-diabetics. In this work, we hypothesized that dysfunction of the outer BRB may be associated with EBE development. To disrupt the RPE component of the outer BRB in vivo, sodium iodate (NaIO3 was administered to C57BL/6J mice. NaIO3-treated and untreated mice were intravenously injected with 108 colony forming units (cfu of Staphylococcus aureus or Klebsiella pneumoniae. At 4 and 6 days postinfection, EBE was observed in NaIO3-treated mice after infection with K. pneumoniae and S. aureus, although the incidence was higher following S. aureus infection. Invasion of the eye was observed in control mice following S. aureus infection, but not in control mice following K. pneumoniae infection. Immunohistochemistry and FITC-dextran conjugate transmigration assays of human RPE barriers after infection with an exoprotein-deficient agr/sar mutant of S. aureus suggested that S. aureus exoproteins may be required for the loss of the tight junction protein, ZO-1, and for permeability of this in vitro barrier. Our results support the clinical findings that for both pathogens, complications which result in BRB permeability increase the likelihood of bacterial transmigration from the bloodstream into the eye. For S. aureus, however, BRB

  18. Port central venous catheters-associated bloodstream infection during outpatient-based chemotherapy.

    Science.gov (United States)

    Mauri, Davide; Roumbkou, Sofia; Michalopoulou, Stella; Tsali, Lamprini; Spiliopoulou, Anastasia; Panou, Charalampos; Valachis, Antonis; Panagopoulos, Angelos; Polyzos, Nikolaos P

    2010-12-01

    Central venous catheters (CVCs) are commonly used for the administration of intravenous chemotherapy in outpatient setting. Nevertheless, outbreaks of catheter-associated bloodstream infections had been reported from oncology centers. We describe a large outbreak of CVCs-associated Klebsiella oxytoca bloodstream infection, occurring in an oncology chemotherapy outpatient unit of northern Greece between October 2006 and May 2007. The outbreak involved approximately 10% of the patients with CVCs who were receiving home-based chemotherapy, and it represents the second larger outbreak of CVCs-associated BSIs due to Klebsiella oxytoca in oncology outpatient centers. We retrospectively analyzed the chain of investigations and prophylactic/diagnostic measures taken to eradicate the infection: (1) patients' chart audit, (2) estimation of the infection among asymptomatic patients, (3) implementation of the level of awareness of medical and paramedical personnel, (4) collection of samples from environment, medications and infusion materials, (5) critical appraisal of chemotherapeutical schemes and (6) cooperation with peripheral institutions. The isolation of Klebsiella oxytoca in a chemotherapy solution (infusional 5-FU in dextrose 5% solution within a 48 h pump) from a peripheral General Hospital and the prompt transmission of the data to the chemotherapy center played a key role for the management of the infection cluster. This is the first report that evidenced the detection of Klebsiella oxytoca within a chemotherapeutical preparation. Data transmission from peripheral hospitals to the central institution resulted in an important feedback that allowed a better estimation of the infection cluster and more tailored actions for the eradication of the infection.

  19. Bloodstream infection among children presenting to a general hospital outpatient clinic in urban Nepal.

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    Rahul Pradhan

    Full Text Available BACKGROUND: There are limited data on the etiology and characteristics of bloodstream infections in children presenting in hospital outpatient settings in South Asia. Previous studies in Nepal have highlighted the importance of murine typhus as a cause of febrile illness in adults and enteric fever as a leading bacterial cause of fever among children admitted to hospital. METHODS: We prospectively studied a total of 1084 febrile children aged between 2 months and 14 years presenting to a general hospital outpatient department in Kathmandu Valley, Nepal, over two study periods (summer and winter. Blood from all patients was tested by conventional culture and by real-time PCR for Rickettsia typhi. RESULTS: Putative etiological agents for fever were identified in 164 (15% patients. Salmonella enterica serovar Typhi (S. Typhi was identified in 107 (10%, S. enterica serovar Paratyphi A (S. Paratyphi in 30 (3%, Streptococcus pneumoniae in 6 (0.6%, S. enterica serovar Typhimurium in 2 (0.2%, Haemophilus influenzae type b in 1 (0.1%, and Escherichia coli in 1 (0.1% patient. S. Typhi was the most common organism isolated from blood during both summer and winter. Twenty-two (2% patients were PCR positive for R. typhi. No significant demographic, clinical and laboratory features distinguished culture positive enteric fever and murine typhus. CONCLUSIONS: Salmonella infections are the leading cause of bloodstream infection among pediatric outpatients with fever in Kathmandu Valley. Extension of immunization programs against invasive bacterial disease to include the agents of enteric fever and pneumococcus could improve the health of children in Nepal.

  20. Bloodstream-To-Eye Infections Are Facilitated by Outer Blood-Retinal Barrier Dysfunction

    Science.gov (United States)

    Coburn, Phillip S.; Wiskur, Brandt J.; Miller, Frederick C.; LaGrow, Austin L.; Astley, Roger A.; Elliott, Michael H.; Callegan, Michelle C.

    2016-01-01

    The blood-retinal barrier (BRB) functions to maintain the immune privilege of the eye, which is necessary for normal vision. The outer BRB is formed by tightly-associated retinal pigment epithelial (RPE) cells which limit transport within the retinal environment, maintaining retinal function and viability. Retinal microvascular complications and RPE dysfunction resulting from diabetes and diabetic retinopathy cause permeability changes in the BRB that compromise barrier function. Diabetes is the major predisposing condition underlying endogenous bacterial endophthalmitis (EBE), a blinding intraocular infection resulting from bacterial invasion of the eye from the bloodstream. However, significant numbers of EBE cases occur in non-diabetics. In this work, we hypothesized that dysfunction of the outer BRB may be associated with EBE development. To disrupt the RPE component of the outer BRB in vivo, sodium iodate (NaIO3) was administered to C57BL/6J mice. NaIO3-treated and untreated mice were intravenously injected with 108 colony forming units (cfu) of Staphylococcus aureus or Klebsiella pneumoniae. At 4 and 6 days postinfection, EBE was observed in NaIO3-treated mice after infection with K. pneumoniae and S. aureus, although the incidence was higher following S. aureus infection. Invasion of the eye was observed in control mice following S. aureus infection, but not in control mice following K. pneumoniae infection. Immunohistochemistry and FITC-dextran conjugate transmigration assays of human RPE barriers after infection with an exoprotein-deficient agr/sar mutant of S. aureus suggested that S. aureus exoproteins may be required for the loss of the tight junction protein, ZO-1, and for permeability of this in vitro barrier. Our results support the clinical findings that for both pathogens, complications which result in BRB permeability increase the likelihood of bacterial transmigration from the bloodstream into the eye. For S. aureus, however, BRB permeability is

  1. Bloodstream-To-Eye Infections Are Facilitated by Outer Blood-Retinal Barrier Dysfunction.

    Science.gov (United States)

    Coburn, Phillip S; Wiskur, Brandt J; Miller, Frederick C; LaGrow, Austin L; Astley, Roger A; Elliott, Michael H; Callegan, Michelle C

    2016-01-01

    The blood-retinal barrier (BRB) functions to maintain the immune privilege of the eye, which is necessary for normal vision. The outer BRB is formed by tightly-associated retinal pigment epithelial (RPE) cells which limit transport within the retinal environment, maintaining retinal function and viability. Retinal microvascular complications and RPE dysfunction resulting from diabetes and diabetic retinopathy cause permeability changes in the BRB that compromise barrier function. Diabetes is the major predisposing condition underlying endogenous bacterial endophthalmitis (EBE), a blinding intraocular infection resulting from bacterial invasion of the eye from the bloodstream. However, significant numbers of EBE cases occur in non-diabetics. In this work, we hypothesized that dysfunction of the outer BRB may be associated with EBE development. To disrupt the RPE component of the outer BRB in vivo, sodium iodate (NaIO3) was administered to C57BL/6J mice. NaIO3-treated and untreated mice were intravenously injected with 108 colony forming units (cfu) of Staphylococcus aureus or Klebsiella pneumoniae. At 4 and 6 days postinfection, EBE was observed in NaIO3-treated mice after infection with K. pneumoniae and S. aureus, although the incidence was higher following S. aureus infection. Invasion of the eye was observed in control mice following S. aureus infection, but not in control mice following K. pneumoniae infection. Immunohistochemistry and FITC-dextran conjugate transmigration assays of human RPE barriers after infection with an exoprotein-deficient agr/sar mutant of S. aureus suggested that S. aureus exoproteins may be required for the loss of the tight junction protein, ZO-1, and for permeability of this in vitro barrier. Our results support the clinical findings that for both pathogens, complications which result in BRB permeability increase the likelihood of bacterial transmigration from the bloodstream into the eye. For S. aureus, however, BRB permeability is

  2. Bloodstream infections in intensive care unit patients: distribution and antibiotic resistance of bacteria

    Directory of Open Access Journals (Sweden)

    Russotto V

    2015-08-01

    Full Text Available Vincenzo Russotto,1 Andrea Cortegiani,1 Giorgio Graziano,2 Laura Saporito,2 Santi Maurizio Raineri,1 Caterina Mammina,2 Antonino Giarratano1 1Department of Biopathology and Medical Biotechnologies (DIBIMED, Section of Anaesthesia, Analgesia, Intensive Care and Emergency, Paolo Giaccone University Hospital, University of Palermo, Palermo, Italy; 2Department of Sciences for Health Promotion and Mother-Child Care, University of Palermo, Palermo, Italy Abstract: Bloodstream infections (BSIs are among the leading infections in critically ill patients. The case-fatality rate associated with BSIs in patients admitted to intensive care units (ICUs reaches 35%–50%. The emergence and diffusion of bacteria with resistance to antibiotics is a global health problem. Multidrug-resistant bacteria were detected in 50.7% of patients with BSIs in a recently published international observational study, with methicillin resistance detected in 48% of Staphylococcus aureus strains, carbapenem resistance detected in 69% of Acinetobacter spp., in 38% of Klebsiella pneumoniae, and in 37% of Pseudomonas spp. Prior hospitalization and antibiotic exposure have been identified as risk factors for infections caused by resistant bacteria in different studies. Patients with BSIs caused by resistant strains showed an increased risk of mortality, which may be explained by a higher incidence of inappropriate empirical therapy in different studies. The molecular genetic characterization of resistant bacteria allows the understanding of the most common mechanisms underlying their resistance and the adoption of surveillance measures. Knowledge of epidemiology, risk factors, mechanisms of resistance, and outcomes of BSIs caused by resistant bacteria may have a major influence on global management of ICU patients. The aim of this review is to provide the clinician an update on BSIs caused by resistant bacteria in ICU patients. Keywords: bloodstream infections, multidrug resistant

  3. Sexual transmission of Trypanosoma cruzi in murine model.

    Science.gov (United States)

    Ribeiro, Marcelle; Nitz, Nadjar; Santana, Camilla; Moraes, Aline; Hagström, Luciana; Andrade, Rafael; Rios, Adriano; Sousa, Alessandro; Dallago, Bruno; Gurgel-Gonçalves, Rodrigo; Hecht, Mariana

    2016-03-01

    Trypanosoma cruzi is mainly transmitted by blood-sucking triatomines, but other routes also have epidemiological importance, such as blood transfusion and congenital transmission. Although the possibility of sexual transmission of T. cruzi has been suggested since its discovery, few studies have been published on this subject. We investigated acquisition of T. cruzi by sexual intercourse in an experimental murine model. Male and female mice in the chronic phase of Chagas disease were mated with naive partners. Parasitological, serological and molecular tests demonstrated the parasites in tissues and blood of partners. These results confirm the sexual transmission of T. cruzi in mice.

  4. Trypanosoma cruzi screening in Texas blood donors, 2008-2012.

    Science.gov (United States)

    Garcia, M N; Woc-Colburn, L; Rossmann, S N; Townsend, R L; Stramer, S L; Bravo, M; Kamel, H; Beddard, R; Townsend, M; Oldham, R; Bottazzi, M E; Hotez, P J; Murray, K O

    2016-04-01

    Chagas disease is an important emerging disease in Texas that results in cardiomyopathy in about 30% of those infected with the parasite Trypanosoma cruzi. Between the years 2008 and 2012, about 1/6500 blood donors were T. cruzi antibody-confirmed positive. We found older persons and minority populations, particularly Hispanic, at highest risk for screening positive for T. cruzi antibodies during routine blood donation. Zip code analysis determined that T. cruzi is associated with poverty. Chagas disease has a significant disease burden and is a cause of substantial economic losses in Texas.

  5. Selective inhibition of 6-phosphogluconate dehydrogenase from Trypanosoma brucei

    Science.gov (United States)

    Bertelli, Massimo; El-Bastawissy, Eman; Knaggs, Michael H.; Barrett, Michael P.; Hanau, Stefania; Gilbert, Ian H.

    2001-05-01

    A number of triphenylmethane derivatives have been screened against 6-phosphogluconate dehydrogenase from Trypanosoma brucei and sheep liver. Some of these compounds show good inhibition of the enzymes and also selectivity towards the parasite enzyme. Modelling was undertaken to dock the compounds into the active sites of both enzymes. Using a combination of DOCK 3.5 and FLEXIDOCK a correlation was obtained between docking score and both activity for the enzymes and selectivity. Visualisation of the docked structures of the inhibitors in the active sites of the enzymes yielded a possible explanation of the selectivity for the parasite enzyme.

  6. High prevalence of Trypanosoma vegrandis in bats from Western Australia.

    Science.gov (United States)

    Austen, Jill M; O'Dea, Mark; Jackson, Bethany; Ryan, Una

    2015-12-15

    The present study describes the first report of Trypanosoma vegrandis in bats using morphology and sequence analysis of the 18S rRNA gene. The PCR prevalence of T. vegrandis in bats was 81.8% (18/22). The high prevalence of T. vegrandis in the present study suggests that bats may play an important role in the epidemiology of T. vegrandis in Australia. T. vegrandis appears to be geographically dispersed, has a wide distribution in Australia and low levels of host specificity.

  7. Transferability of Trypanosoma cruzi from mixed human host infection to Triatoma infestans and from insects to axenic culture.

    Science.gov (United States)

    Ortiz, Sylvia; Zulantay, Inés; Apt, Werner; Saavedra, Miguel; Solari, Aldo

    2015-02-01

    The etiologic agent of Chagas disease is Trypanosoma cruzi, a protozoan whose life cycle involves obligatory passage through vertebrate and invertebrate hosts in a series of stages. The aim of this study was to explore the transferability of mixed discrete typing units (DTUs) of T. cruzi present in chronic chagasic patients when passed through an invertebrate host during xenodiagnosis (XD) and then when transferred to axenic cultures to obtain T. cruzi isolates. DTUs of T. cruzi present in these two hosts and axenic cultures were identified by kDNA PCR amplification and subsequent hybridization with DTU-specific probes. Mixtures of Tc I, Tc II, Tc V and Tc VI DTUs were detected in blood samples. However as a result of XD and axenic cultures it was possible to identify mostly Tc V. We conclude that the transferability of an isolate of T.cruzi derived from mixed DTUs present in human blood depends upon the starved invertebrate host used for xenodiagnosis.

  8. Improved Method for In Vitro Secondary Amastigogenesis of Trypanosoma cruzi: Morphometrical and Molecular Analysis of Intermediate Developmental Forms

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    L. A. Hernández-Osorio

    2010-01-01

    Full Text Available Trypanosoma cruzi undergoes a biphasic life cycle that consists of four alternate developmental stages. In vitro conditions to obtain a synchronic transformation and efficient rates of pure intermediate forms (IFs, which are indispensable for further biochemical, biological, and molecular studies, have not been reported. In the present study, we established an improved method to obtain IFs from secondary amastigogenesis. During the transformation kinetics, we observed progressive decreases in the size of the parasite body, undulating membrane and flagellum that were concomitant with nucleus remodeling and kinetoplast displacement. In addition, a gradual reduction in parasite movement and acquisition of the amastigote-specific Ssp4 antigen were observed. Therefore, our results showed that the in vitro conditions used obtained large quantities of highly synchronous and pure IFs that were clearly distinguished by morphometrical and molecular analyses. Obtaining these IFs represents the first step towards an understanding of the molecular mechanisms involved in amastigogenesis.

  9. Trypanosoma b. rhodesiense (WRATat Serodeme): Purification and Characterization of Surface Antigens for the Vaccine Development Program.

    Science.gov (United States)

    1980-04-01

    149-151 (1975). 7. Snary, D. and L. Hudson: Trypanosoma cruzi cell surface proteins: Identifi- cation of one major glycoprotein. FEBS Lett. 100: 166...7 AD-A095 616 COLORADO STATE UNIV FORT COLLINS COLL OF VETERINARY --ETC F/B 6/5 TRYPANOSOMA B. RHODESIENSE (WRATAT SERODEME): PURIFICATION AND --ETC...U) APR 0 A R KEILMAN DAMD17-79-C-9117 UNCLASSIFIED NL ///I///I//IIf~f~fllf EEEEEL 1LE.. AD_ REPORT NUMBER 1 Trypanosoma b. rhodesiense (WRATat

  10. Trypanosoma (Herpetosoma) leeuwenhoeki in Choloepus hoffmanni and Didelphis marsupialis of the Pacific Coast of Colombia.

    Science.gov (United States)

    Travi, B L; Zea, A; D'Alessandro, A

    1989-04-01

    Trypanosoma (Herpetosoma) leeuwenhoeki, originally described in Panamanian sloths, was isolated from Didelphis marsupialis (Marsupialia) and Choloepus hoffmanni (Edentata) inhabiting the Pacific coast of Colombia. Trypanosomes were characterized by their large blood forms (total length 51-53 microns), poor infectivity for mice, and lack of development in Rhodnius prolixus. Isoenzyme studies, with either strains or clones, revealed homogeneous profiles clearly distinct from Trypanosoma cruzi and Trypanosoma rangeli reference strains. The present report extends the geographical distribution of T. leeuwenhoeki to South America and broadens its known host range to another order of mammals.

  11. Trypanosoma cruzi: Transporte de metabolitos esenciales obtenidos del hospedador Trypanosoma cruzi: Transport of essential metabolites acquired from the host

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    Claudio A. Pereira

    2008-10-01

    Full Text Available El Trypanosoma cruzi es el agente causal de la enfermedad de Chagas, endémica en Argentina y en toda América Latina. Presenta numerosas características metabólicas diferenciales respecto a sus hospedadores insectos y mamíferos. Algunas de estas diferencias fueron consecuencia de millones de años de adaptación al parasitismo en los cuales estos organismos protozoarios reemplazaron, a lo largo de su evolución, muchas rutas metabólicas de biosíntesis por sistemas de transporte de metabolitos desde el hospedador. En esta revisión se describen los avances en el conocimiento de los sistemas de transporte tanto bioquímicos como también de las moléculas involucradas en dichos procesos. Se aborda con especial énfasis los transportadores de aminoácidos y poliaminas de T. cruzi de la familia AAAP (Amino Acid/Auxin Permeases ya que parece ser exclusiva de los tripanosomátidos. Teniendo en cuenta que estas moléculas se encuentran completamente ausentes en mamíferos podrían ser consideradas como potenciales blancos contra el Trypanosoma cruzi.Trypanosoma cruzi is the etiological agent of Chagas disease, a disease endemic not only in Argentina but also in all of Latinamerica. T. cruzi presents several metabolic characteristics which are completely absent in its insect vectors and in mammalian hosts. Some of these differences were acquired after millions of years of adaptation to parasitism, during which this protozoan replaced many biosynthetic routes for transport systems. In the present review, we describe the advances in the knowledge of T. cruzi transport processes and the molecules involved. In particular, we focus on aminoacid and polyamine transporters from the AAAP family (Amino Acid/Auxin Permeases, because they seem to be exclusive transporters from trypanosomatids. Taking into account that these permeases are completely absent in mammals, they could be considered as a potential target against Trypanosoma cruzi.

  12. Delay in the administration of appropriate antimicrobial therapy in Staphylococcus aureus bloodstream infection : A prospective multicenter hospital-based cohort study

    NARCIS (Netherlands)

    Kaasch, A. J.; Rieg, S.; Kuetscher, J.; Brodt, H. -R.; Widmann, T.; Herrmann, M.; Meyer, C.; Welte, T.; Kern, P.; Haars, U.; Reuter, S.; Huebner, I.; Strauss, R.; Sinha, B.; Brunkhorst, F. M.; Hellmich, M.; Faetkenheuer, G.; Kern, W. V.; Seifert, H.

    2013-01-01

    Early broad-spectrum antimicrobial treatment reduces mortality in patients with septic shock. In a multicenter, prospective observational study, we explored whether delayed appropriate antimicrobial therapy (AAT) influences outcome in Staphylococcus aureus bloodstream infection (SAB). Two hundred an

  13. The short non-coding transcriptome of the protozoan parasite Trypanosoma cruzi.

    Directory of Open Access Journals (Sweden)

    Oscar Franzén

    2011-08-01

    Full Text Available The pathway for RNA interference is widespread in metazoans and participates in numerous cellular tasks, from gene silencing to chromatin remodeling and protection against retrotransposition. The unicellular eukaryote Trypanosoma cruzi is missing the canonical RNAi pathway and is unable to induce RNAi-related processes. To further understand alternative RNA pathways operating in this organism, we have performed deep sequencing and genome-wide analyses of a size-fractioned cDNA library (16-61 nt from the epimastigote life stage. Deep sequencing generated 582,243 short sequences of which 91% could be aligned with the genome sequence. About 95-98% of the aligned data (depending on the haplotype corresponded to small RNAs derived from tRNAs, rRNAs, snRNAs and snoRNAs. The largest class consisted of tRNA-derived small RNAs which primarily originated from the 3' end of tRNAs, followed by small RNAs derived from rRNA. The remaining sequences revealed the presence of 92 novel transcribed loci, of which 79 did not show homology to known RNA classes.

  14. Simultaneous stable expression of neomycin phosphotransferase and green fluorescence protein genes in Trypanosoma cruzi.

    Science.gov (United States)

    dos Santos, W G; Buck, G A

    2000-12-01

    The ribosomal RNA (rRNA) gene promoter was used to construct plasmid vectors that simultaneously express multiple exogenous genes in Trypanosoma cruzi. Vector pBSPANEO expresses neomycin phosphotransferase, and pPAGFPAN expresses both green fluorescent protein and neomycin phosphotransferase from a single promoter. Both vectors require the presence of the rRNA promoter for stable transfection; epimastigotes transfected with pPAGFPAN strongly fluoresced due to green fluorescent protein expression. Intact plasmids were rescued from the T. cruzi-transfected population after >8 mo of culture, indicating stable replication of these vectors. Vectors were integrated into the rRNA locus by homologous recombination and into other loci, presumably by illegitimate recombination. Parasites bearing tandem concatamers of plasmids were also found among the transfectants. Transfectants expressing green fluorescent protein showed a bright green fluorescence distributed throughout the cell. Fluorescence was also detected in amastigotes after infection of mammalian cells with transfected parasites, indicating that the rRNA promoter can drive efficient expression of these reporter genes in multiple life-cycle stages of the parasite. Expression of the heterologous genes was detected after passage in mice or in the insect vector. These vectors will be useful for the genetic dissection of T. cruzi biology and pathogenesis.

  15. Immunocytochemical and biochemical detection of alpha-L-fucosidase in Trypanosoma cruzi

    Directory of Open Access Journals (Sweden)

    Miletti L.C.

    2003-01-01

    Full Text Available The aim of the present study was to demonstrate the presence of alpha-L-fucosidase in Trypanosoma cruzi. Immunocytochemical and biochemical techniques were used to localize and characterize a membrane-associated, neutral-pH-optimum, alpha-L-fucosidase from the parasite. Light and electron microscopy localized the alpha-L-fucosidase specifically on the surface of the parasite and on membranes in the posterior region of the epimastigote stage. Although much less intense, labeling was also detected on the surface of trypomastigotes. At least 50% of the alpha-L-fucosidase activity was associated with epimastigote membrane solubilized with 1 M NaCl or 1% Triton X-100, suggesting that alpha-L-fucosidase is peripherally associated with membranes. The enzyme from epimastigotes had a neutral pH optimum (near 7 but displayed low specific activity when p-nitrophenyl-alpha-L-fucoside was employed as substrate (0.028 U/mg protein for epimastigotes and 0.015 U/mg protein for tissue culture-derived trypomastigotes. Polyacrylamide gel electrophoresis and Western blotting analysis both showed an expected 50-kDa polypeptide which was immunoreactive with anti-alpha-L-fucosidase antibodies.

  16. Identification of Protein Complex Associated with LYT1 of Trypanosoma cruzi

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    C. Lugo-Caballero

    2013-01-01

    Full Text Available To carry out the intracellular phase of its life cycle, Trypanosoma cruzi must infect a host cell. Although a few molecules have been reported to participate in this process, one known protein is LYT1, which promotes lysis under acidic conditions and is involved in parasite infection and development. Alternative transcripts from a single LYT1 gene generate two proteins with differential functions and compartmentalization. Single-gene products targeted to more than one location can interact with disparate proteins that might affect their function and targeting properties. The aim of this work was to study the LYT1 interaction map using coimmunoprecipitation assays with transgenic parasites expressing LYT1 products fused to GFP. We detected several proteins of sizes from 8 to 150 kDa that bind to LYT1 with different binding strengths. By MS-MS analysis, we identified proteins involved in parasite infectivity (trans-sialidase, development (kDSPs and histones H2A and H2B, and motility and protein traffic (dynein and α- and β-tubulin, as well as protein-protein interactions (TPR-protein and kDSPs and several hypothetical proteins. Our approach led us to identify the LYT1 interaction profile, thereby providing insights into the molecular mechanisms that contribute to parasite stage development and pathogenesis of T. cruzi infection.

  17. Evaluation of clinical signs, parasitemia, hematologic and biochemical changes in cattle experimentally infected with Trypanosoma vivax.

    Science.gov (United States)

    Fidelis Junior, Otavio Luiz; Sampaio, Paulo Henrique; Machado, Rosangela Zacarias; André, Marcos Rogério; Marques, Luiz Carlos; Cadioli, Fabiano Antonio

    2016-01-01

    Infections by Trypanosoma vivax cause great losses to livestock in Africa and Central and South Americas. Outbreaks due this parasite have been occurred with increasing frequency in Brazil. Knowledge of changes caused by T. vivax during the course of this disease can be of great diagnostic value. Thus, clinical signs, parasitemia, hematologic and biochemical changes of cattle experimentally infected by this hemoparasite were evaluated. Two distinct phases were verified during the infection - an acute phase where circulating parasites were seen and then a chronic phase where fluctuations in parasitemia were detected including aparasitemic periods. A constant reduction in erythrocytes, hemoglobin and packed cell volume (PVC) were observed. White blood cells (WBC) showed pronounced changes such as severe neutropenia and lymphopenia during the acute phase of the illness. Decreases in cholesterol, albumin, aspartate aminotransferase (AST), lactate dehydrogenase (LDH), and increases in glucose, globulin, protein, and alkaline phosphatase (ALP) were observed. The "Lins" isolate of T. vivax showed pathogenicity for cattle, and intense parasitemia was detected in the early stages of infection. Circulating parasites were detected for about two months. The most evident laboratory abnormalities were found in WBC parameters, including thrombocytopenia.

  18. Evaluation of clinical signs, parasitemia, hematologic and biochemical changes in cattle experimentally infected with Trypanosoma vivax

    Directory of Open Access Journals (Sweden)

    Otavio Luiz Fidelis Junior

    2016-03-01

    Full Text Available Abstract Infections by Trypanosoma vivax cause great losses to livestock in Africa and Central and South Americas. Outbreaks due this parasite have been occurred with increasing frequency in Brazil. Knowledge of changes caused byT. vivax during the course of this disease can be of great diagnostic value. Thus, clinical signs, parasitemia, hematologic and biochemical changes of cattle experimentally infected by this hemoparasite were evaluated. Two distinct phases were verified during the infection – an acute phase where circulating parasites were seen and then a chronic phase where fluctuations in parasitemia were detected including aparasitemic periods. A constant reduction in erythrocytes, hemoglobin and packed cell volume (PVC were observed. White blood cells (WBC showed pronounced changes such as severe neutropenia and lymphopenia during the acute phase of the illness. Decreases in cholesterol, albumin, aspartate aminotransferase (AST, lactate dehydrogenase (LDH, and increases in glucose, globulin, protein, and alkaline phosphatase (ALP were observed. The “Lins” isolate of T. vivax showed pathogenicity for cattle, and intense parasitemia was detected in the early stages of infection. Circulating parasites were detected for about two months. The most evident laboratory abnormalities were found in WBC parameters, including thrombocytopenia.

  19. Polypeptide profiles of South Indian isolate of Trypanosoma evansi.

    Science.gov (United States)

    Sivajothi, S; Rayulu, V C; Bhaskar Reddy, B V; Malakondaiah, P; Sreenivasulu, D; Sudhakara Reddy, B

    2016-09-01

    The field isolates of Trypanosoma evansi was collected from the infected cattle and it was propagated in rats. Trypanosoma evansi parasites were separated from the blood of infected rats by using diethylaminoethyl cellulose column chromatography. Whole cell lysate antigen (WCL) was prepared from purified trypanosomes by ultrasonication and centrifugation. The prepared WCL antigen was further purified by 50 % ammonium sulphate precipitation. Protein concentration of WCL antigen of T. evansi was 60 mg/ml. Protein concentration was adjusted to 1.0 mg/ml in PBS, pH 8.0 and stored at -20(0) C.   Polypeptide profiles of WCL antigen of T. evansi was determined by sodium dodecyl sulphate polyacrylamide gel electrophoresis. A total of eight polypeptide bands of the size ranging from 25 to 85 kDa in WCL antigen of T. evansi were obtained. Five prominent bands with molecular weight of 74, 60, 53, 42 and 37 kDa and three light bands with molecular weight of 85, 34 and 25 kDa were observed.

  20. Refractory hypoglycaemia in a dog infected with Trypanosoma congolense.

    Science.gov (United States)

    Deschamps, Jack-Yves; Desquesnes, Marc; Dorso, Laetitia; Ravel, Sophie; Bossard, Géraldine; Charbonneau, Morgane; Garand, Annabelle; Roux, Françoise A

    2016-01-01

    A 20 kg German shepherd dog was presented to a French veterinary teaching hospital for seizures and hyperthermia. The dog had returned 1 month previously from a six-month stay in Senegal and sub-Saharan Africa. Biochemistry and haematology showed severe hypoglycaemia (0.12 g/L), anaemia and thrombocytopenia. Despite administration of large amounts of glucose (30 mL of 30% glucose IV and 10 mL of 70% sucrose by gavage tube hourly), 26 consecutive blood glucose measurements were below 0.25 g/L (except one). Routine cytological examination of blood smears revealed numerous free extracytoplasmic protozoa consistent with Trypanosoma congolense. PCR confirmed a Trypanosoma congolense forest-type infection. Treatment consisted of six injections of pentamidine at 48-hour intervals. Trypanosomes had disappeared from the blood smears four days following the first injection. Clinical improvement was correlated with the normalization of laboratory values. The infection relapsed twice and the dog was treated again; clinical signs and parasites disappeared and the dog was considered cured; however, 6 years after this incident, serological examination by ELISA T. congolense was positive. The status of this dog (infected or non-infected) remains unclear. Hypoglycaemia was the most notable clinical feature in this case. It was spectacular in its severity and in its refractory nature; glucose administration seemed only to feed the trypanosomes, indicating that treatment of hypoglycaemia may in fact have been detrimental.

  1. Refractory hypoglycaemia in a dog infected with Trypanosoma congolense

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    Deschamps Jack-Yves

    2016-01-01

    Full Text Available A 20 kg German shepherd dog was presented to a French veterinary teaching hospital for seizures and hyperthermia. The dog had returned 1 month previously from a six-month stay in Senegal and sub-Saharan Africa. Biochemistry and haematology showed severe hypoglycaemia (0.12 g/L, anaemia and thrombocytopenia. Despite administration of large amounts of glucose (30 mL of 30% glucose IV and 10 mL of 70% sucrose by gavage tube hourly, 26 consecutive blood glucose measurements were below 0.25 g/L (except one. Routine cytological examination of blood smears revealed numerous free extracytoplasmic protozoa consistent with Trypanosoma congolense. PCR confirmed a Trypanosoma congolense forest-type infection. Treatment consisted of six injections of pentamidine at 48-hour intervals. Trypanosomes had disappeared from the blood smears four days following the first injection. Clinical improvement was correlated with the normalization of laboratory values. The infection relapsed twice and the dog was treated again; clinical signs and parasites disappeared and the dog was considered cured; however, 6 years after this incident, serological examination by ELISA T. congolense was positive. The status of this dog (infected or non-infected remains unclear. Hypoglycaemia was the most notable clinical feature in this case. It was spectacular in its severity and in its refractory nature; glucose administration seemed only to feed the trypanosomes, indicating that treatment of hypoglycaemia may in fact have been detrimental.

  2. Kinetic and biochemical characterization of Trypanosoma evansi nucleoside triphosphate diphosphohydrolase.

    Science.gov (United States)

    Weiss, Paulo Henrique Exterchoter; Batista, Franciane; Wagner, Glauber; Magalhães, Maria de Lourdes Borba; Miletti, Luiz Claudio

    2015-06-01

    Nucleoside triphosphate diphospho-hydrolases (NTPDases) catalyze the hydrolysis of several nucleosides tri and diphosphate playing major roles in eukaryotes including purinergic signaling, inflammation, hemostasis, purine salvage and host-pathogen interactions. These enzymes have been recently described in parasites where several evidences indicated their involvement in virulence and infection. Here, we have investigated the presence of NTPDase in the genome of Trypanosoma evansi. Based on the genomic sequence from Trypanosoma brucei, we have amplified an 1812 gene fragment corresponding to the T. evansi NTPDase gene. The protein was expressed in the soluble form and purified to homogeneity and enzymatic assays were performed confirming the enzyme identity. Kinetic parameters and substrate specificity were determined. The dependence of cations on enzymatic activity was investigated indicating the enzyme is stimulated by divalent cations and carbohydrates but inhibited by sodium. Bioinformatic analysis indicates the enzyme is a membrane bound protein facing the extracellular side of the cell with 98% identity to the T. brucei homologous NTPDase gene.

  3. A rapid method for testing in vivo the susceptibility of different strains of Trypanosoma cruzi to active chemotherapeutic agents

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    Leny S. Filardi

    1984-06-01

    Full Text Available A method is described which permits to determine in vivo an in a short period of time (4-6 hours the sensitivity of T. cruzo strains to known active chemotherapeutic agents. By using resistant- and sensitive T. cruzi stains a fairly good correlation was observed between the results obtained with this rapid method (which detects activity against the circulating blood forms and those obtained with long-term schedules which involve drug adminstration for at least 20 consecutive days and a prolonged period of assessment. This method may be used to characterize susceptibility to active drugs used clinically, provide infomation on the specific action against circulating trypomastigotes and screen active compounds. Differences in the natural susceptibility of Trypanosoma cruzi strains to active drugs have been already reported using different criteria, mostly demanding long-term study of the animal (Hauschka, 1949; Bock, Gonnert & Haberkorn, 1969; Brener, Costa & Chiari, 1976; Andrade & Figueira, 1977; Schlemper, 1982. In this paper we report a method which detects in 4-6 hours the effect of drugs on bloodstream forms in mice with established T. cruzi infections. The results obtained with this method show a fairly good correlation with those obtained by prolonged treatment schedules used to assess the action of drugs in experimental Chagas' disease and may be used to study the sensitivity of T. cruzi strains to active drugs.No presente trabalho descreve-se um metodo que permite determinar in vivo e em curto espaço de tempo (4-6 horas a sensibilidade de cepas de T. cruzi a agentes terapeuticos ativos na doença de Chagas. Usando-se cepas sensíveis e resistentes aos medicamentos foi possível observar uma boa correlação entre os resultados obtidos com o método rápido (que detecta atividade contra as formas circulantes do parasita e aqueles obtidos com esquema de acao prolongada que envolve a administração da droga por 20 dias e posterior avalia

  4. Pathogens and antimicrobial susceptibility profiles in critically ill patients with bloodstream infections: a descriptive study

    Science.gov (United States)

    Savage, Rachel D.; Fowler, Robert A.; Rishu, Asgar H.; Bagshaw, Sean M.; Cook, Deborah; Dodek, Peter; Hall, Richard; Kumar, Anand; Lamontagne, François; Lauzier, François; Marshall, John; Martin, Claudio M.; McIntyre, Lauralyn; Muscedere, John; Reynolds, Steven; Stelfox, Henry T.; Daneman, Nick

    2016-01-01

    Background: Surveillance of antimicrobial resistance is vital to guiding empirical treatment of infections. Collating and reporting routine data on clinical isolate testing may offer more timely information about resistance patterns than traditional surveillance network methods. Methods: Using routine microbiology testing data collected from the Bacteremia Antibiotic Length Actually Needed for Clinical Effectiveness retrospective cohort study, we conducted a descriptive secondary analysis among critically ill patients in whom bloodstream infections had been diagnosed in 14 intensive care units (ICUs) in Canada. The participating sites were located within tertiary care teaching hospitals and represented 6 provinces and 10 cities. More than 80% of the study population was accrued from 2011-2013. We assessed the epidemiologic features of the infections and corresponding antimicrobial susceptibility profiles. Susceptibility testing was done according to Clinical Laboratory Standards Institute guidelines at accredited laboratories. Results: A total of 1416 pathogens were isolated from 1202 patients. The most common organisms were Escherichia coli (217 isolates [15.3%]), Staphylococcus aureus (175 [12.4%]), coagulase-negative staphylococci (117 [8.3%]), Klebsiella pneumoniae (86 [6.1%]) and Streptococcus pneumoniae (85 [6.0%]). The contribution of individual pathogens varied by site. For 13 ICUs, gram-negative susceptibility rates were high for carbapenems (95.4%), tobramycin (91.2%) and piperacillin-tazobactam (90.0%); however, the proportion of specimens susceptible to these agents ranged from 75.0%-100%, 66.7%-100% and 75.0%-100%, respectively, across sites. Fewer gram-negative bacteria were susceptible to fluoroquinolones (84.5% [range 64.1%-97.2%]). A total of 145 patients (12.1%) had infections caused by highly resistant microorganisms, with significant intersite variation (range 2.6%-24.0%, χ2 = 57.50, p < 0.001). Interpretation: We assessed the epidemiologic

  5. Clinical and microbiological characterization of carbapenem-resistant Acinetobacter baumannii bloodstream infections.

    Science.gov (United States)

    Song, Joon Young; Cheong, Hee Jin; Choi, Won Suk; Heo, Jung Yeon; Noh, Ji Yun; Kim, Woo Joo

    2011-05-01

    The incidence of carbapenem-resistant Acinetobacter baumannii infection is increasing, which might be associated with high morbidity and mortality among critically ill patients with limited therapeutic options. This study was conducted to evaluate the clinical and microbiological features of carbapenem-resistant A. baumannii bacteraemia. The medical records of 28 adult patients with this bacteraemia admitted to Korea University Guro Hospital, from January 2005 through December 2010, were reviewed. Using the 28 bloodstream isolates, we intended to detect genes encoding carbapenemases, and investigate the inoculum effect on each of the antimicrobial agents rifampicin, imipenem, colistin and tigecycline. With one blood isolate from a patient with pneumonia, rifampicin-inducible resistance was examined using the experimental mouse pneumonia model. Out of 28 carbapenem-resistant A. baumannii bloodstream infections (BIs), the most common primary focus was the central venous catheter (35.7 %) and then the lung (32.1 %). The 30 day overall mortality was 53.6 %; in most cases (80 %) the patients died within 10 days after the onset of the bacteraemia. By univariate analysis, inappropriate antimicrobial therapy (73.3 vs 30.8 %, P = 0.02), mechanical ventilation (53.3 vs 15.4 %, P = 0.04) and a high Pitt bacteraemia score (4.9±1.9 vs 2.2±1.2, P<0.01) were statistically significant risk factors for mortality, while only a high Pitt bacteraemia score (odds ratio 2.6; 95 % confidence interval 1.1-6.5) was independently associated with 30 day mortality by multivariate analysis. All 28 isolates had the bla(OXA-51)-like gene with upstream ISAbaI, 2 of which additionally had the bla(OXA-58)-like gene and the bla(OXA-23)-like gene. Inoculum effect and rifampicin inducible resistance were not detected. Considering the rapid progression to death in carbapenem-resistant A. baumannii BIs, early empirical antibiotic therapy would be warranted based on the local

  6. Staging atmospheres

    DEFF Research Database (Denmark)

    Bille, Mikkel; Bjerregaard, Peter; Sørensen, Tim Flohr

    2015-01-01

    The article introduces the special issue on staging atmospheres by surveying the philosophical, political and anthropological literature on atmosphere, and explores the relationship between atmosphere, material culture, subjectivity and affect. Atmosphere seems to occupy one of the classic...... localities of tensions between matter and the immaterial, the practical and the ideal, and subject and object. In the colloquial language there can, moreover, often seem to be something authentic or genuine about atmosphere, juxtaposing it to staging, which is implied to be something simulated or artificial....... This introduction seeks to outline how a number of scholars have addressed the relationship between staged atmospheres and experience, and thus highlight both the philosophical, social and political aspects of atmospheres...

  7. In vitro evaluation of the activity of aromatic nitrocompounds against Trypanosoma cruzi

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    Renata B Oliveira

    2003-01-01

    Full Text Available Fourteen compounds were evaluated for their activity against Trypanosoma cruzi blood stream forms at the concentration of 500 µg/ml. Six compounds were active and re-tested at lower concentrations.

  8. Trypanosoma cruzi-Trypanosoma rangeli co-infection ameliorates negative effects of single trypanosome infections in experimentally infected Rhodnius prolixus.

    Science.gov (United States)

    Peterson, Jennifer K; Graham, Andrea L; Elliott, Ryan J; Dobson, Andrew P; Triana Chávez, Omar

    2016-08-01

    Trypanosoma cruzi, causative agent of Chagas disease, co-infects its triatomine vector with its sister species Trypanosoma rangeli, which shares 60% of its antigens with T. cruzi. Additionally, T. rangeli has been observed to be pathogenic in some of its vector species. Although T. cruzi-T. rangeli co-infections are common, their effect on the vector has rarely been investigated. Therefore, we measured the fitness (survival and reproduction) of triatomine species Rhodnius prolixus infected with just T. cruzi, just T. rangeli, or both T. cruzi and T. rangeli. We found that survival (as estimated by survival probability and hazard ratios) was significantly different between treatments, with the T. cruzi treatment group having lower survival than the co-infected treatment. Reproduction and total fitness estimates in the T. cruzi and T. rangeli treatments were significantly lower than in the co-infected and control groups. The T. cruzi and T. rangeli treatment group fitness estimates were not significantly different from each other. Additionally, co-infected insects appeared to tolerate higher doses of parasites than insects with single-species infections. Our results suggest that T. cruzi-T. rangeli co-infection could ameliorate negative effects of single infections of either parasite on R. prolixus and potentially help it to tolerate higher parasite doses.

  9. Partial Purification of Integral Membrane Antigenic Proteins from Trypanosoma evansi That Display Immunological Cross-Reactivity with Trypanosoma vivax

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    Norma P. Velásquez

    2014-01-01

    Full Text Available Trypanosoma evansi and Trypanosoma vivax, which are the major causative agents of animal trypanosomosis in Venezuela, have shown a very high immunological cross-reactivity. Since the production of T. vivax antigens is a limiting factor as this parasite is difficult to propagate in experimental animal models, our goal has been to identify and isolate antigens from T. evansi that cross-react with T. vivax. Here, we used the Venezuelan T. evansi TEVA1 isolate to prepare the total parasite lysate and its corresponding cytosolic and membranous fractions. In order to extract the T. evansi integral membrane proteins, the particulate portion was further extracted first with Triton X-100, and then with sodium dodecyl sulfate. After discarding the cytosolic and Triton X-100 solubilized proteins, we employed sedimentation by centrifugation on linear sucrose gradients to partially purify the sodium dodecyl sulfate-solubilized proteins from the Triton X-100 resistant particulate fraction of T. evansi. We obtained enriched pools containing polypeptide bands with apparent molecular masses of 27 kDa, 31 kDa, and 53 kDa, which were recognized by anti-T. vivax antibodies from experimentally and naturally infected bovines.

  10. Experimental transmission of the parasitic flagellates Trypanosoma cruzi and Trypanosoma rangeli between triatomine bugs or mice and captive neotropical bats

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    Maurice E Thomas

    2007-08-01

    Full Text Available Trypanosoma cruzi and Trypanosoma rangeli-like trypanosomes have been found in a variety of neotropical bat species. In this study, bats (Artibeus lituratus, Carollia perspicillata, Desmodus rotundus, Glossophaga soricina, Molossus molossus, Phyllostomus hastatus were maintained under controlled conditions, and experiments were conducted to determine how they might become infected naturally with trypanosomes. All bats were first screened for existing infections by hemoculture and the examination of blood smears, and only apparently uninfected animals were then used in the experiments. Proof was obtained that the triatomine bug Rhodnius prolixus would readily feed upon some of the bats, and two species became infected after being bitten by bugs infected with T. rangeli. Some bats also became infected by ingesting R. prolixus carrying T. cruzi, or following subcutaneous or intragastic inoculation with fecal suspensions of R. prolixus containing T. cruzi. P. hastatus became infected after ingesting mice carrying T. cruzi. All of the bats studied inhabit roosts that may be occupied by triatomine bugs and, with the exception of D. rotundus, all also feed to at least some extent upon insects. These findings provide further evidence of how bats may play significant roles in the epidemiology of T. cruzi and T. rangeli in the New World tropics.

  11. Parasite-based screening and proteome profiling reveal orlistat, an FDA-approved drug, as a potential anti Trypanosoma brucei agent.

    Science.gov (United States)

    Yang, Peng-Yu; Wang, Min; Liu, Kai; Ngai, Mun Hong; Sheriff, Omar; Lear, Martin J; Sze, Siu Kwan; He, Cynthia Y; Yao, Shao Q

    2012-07-02

    Trypanosoma brucei is a parasite that causes African sleeping sickness in humans and nagana in livestock and is transmitted by the tsetse fly. There is an urgent need for the development of new drugs against African trypanosomiasis due to the lack of vaccines and effective drugs. Orlistat (also called tetrahydrolipstatin or THL) is an FDA-approved antiobesity drug targeting primarily the pancreatic and gastric lipases within the gastrointestinal tract. It shows potential activities against tumors, mycobacteria, and parasites. Herein, we report the synthesis and evaluation of an expanded set of orlistat-like compounds, some of which showed highly potent trypanocidal activities in both the bloodstream form (BSF) and the procyclic form (PCF) of T. brucei. Subsequent in situ parasite-based proteome profiling was carried out to elucidate potential cellular targets of the drug in both forms. Some newly identified targets were further validated by the labeling of recombinantly expressed enzymes in Escherichia coli lysates. Bioimaging experiments with a selected compound were carried out to study the cellular uptake of the drug in T. brucei. Results indicated that orlistat is much more efficiently taken up by the BSF than the PCF of T. brucei and has clear effects on the morphology of mitochondria, glycosomes, and the endoplasmic reticulum in both BSF and PCF cells. These results support specific effects of orlistat on these organelles and correlate well with our in situ proteome profiling. Given the economic challenges of de novo drug development for neglected diseases, we hope that our findings will stimulate further research towards the conversion of orlistat-like compounds into new trypanocidal drugs.

  12. Trypanosoma cruzi: identification of specific epimastigote antigens by human immune sera

    OpenAIRE

    1989-01-01

    Soluble antigens from epimastigotes of Trypanosoma cruzi were analyzed by western blot in terms of their reactivity with sera from patients with Chagas' disease. In addition, sera from patients with visceral (AVL) and tegumentar leishmaniasis (ATL) were also tested in order to identify cross-reactivities with Trypanosoma cruzy antigens. Twenty eight polypeptides with molecular weights ranging from 14 kDa to 113 kDa were identified with sera from Chagas' disease patients. An extensive cross-re...

  13. The miRNA and mRNA Signatures of Peripheral Blood Cells in Humans Infected with Trypanosoma brucei gambiense.

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    Smiths Lueong

    Full Text Available Simple, reliable tools for diagnosis of human African Trypanosomiases could ease field surveillance and enhance patient care. In particular, current methods to distinguish patients with (stage II and without (stage I brain involvement require samples of cerebrospinal fluid. We describe here an exploratory study to find out whether miRNAs from peripheral blood leukocytes might be useful in diagnosis of human trypanosomiasis, or for determining the stage of the disease. Using microarrays, we measured miRNAs in samples from Trypanosoma brucei gambiense-infected patients (9 stage I, 10 stage II, 8 seronegative parasite-negative controls and 12 seropositive, but parasite-negative subjects. 8 miRNAs (out of 1205 tested showed significantly lower expression in patients than in seronegative, parasite-negative controls, and 1 showed increased expression. There were no clear differences in miRNAs between patients in different disease stages. The miRNA profiles could not distinguish seropositive, but parasitologically negative samples from controls and results within this group did not correlate with those from the trypanolysis test. Some of the regulated miRNAs, or their predicted mRNA targets, were previously reported changed during other infectious diseases or cancer. We conclude that the changes in miRNA profiles of peripheral blood lymphocytes in human African trypanosomiasis are related to immune activation or inflammation, are probably disease-non-specific, and cannot be used to determine the disease stage. The approach has little promise for diagnostics but might yield information about disease pathology.

  14. Bacillus Cereus catheter related bloodstream infection in a patient in a patient with acute lymphblastic leukemia

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    Lütfiye Öksüz

    2012-01-01

    Full Text Available Bacillus cereus infection is rarely associated with actual infection and for this reason single positive blood culture is usually regarded as contamination . However it may cause a number of infections, such catheter-related blood stream infections. Significant catheter-related bloodstream infections (CRBSI caused by Bacillus spp. are mainly due to B.cereus and have been predominantly reported in immunocompromised hosts1 . Catheter removal is generally advised for management of infection. In this report, catheter-related bacteremia caused by B.cereus in a patient with acute lymphoblastıc leukemia (ALL in Istanbul Medical Faculty was presented.A 44-year old man presented with fatigue, weight loss, epistaxis and high fever. A double-lumen Hickman–catheter (Bard 12.0 Fr, Round Dual Lumen was inserted by surgical cut-down to access the right subclavian vein which would be necessary for allogeneic stem cell transplantation. Three weeks later the patient presented with high fever and headache. Bacillus spp. was isolated from the cathether while blood culture obtained from the peripheral vein remained negative. The bacterial identification was confirmed as B.cereus using VITEK identification system It has been reported Bacillus cereus septicemia may be fatal in immunocompromised hosts despite broad-spectrum appropriate treatment10. Catheter removal is essential for prevention of recurrent bacteremia. Long-term cathater salvage should be reserved for appropriate patient group.

  15. Molecular differentiation and antifungal susceptibilities of Candida parapsilosis isolated from patients with bloodstream infections.

    Science.gov (United States)

    Tay, Sun Tee; Na, Shiang Ling; Chong, Jennifer

    2009-02-01

    The genetic heterogeneity and antifungal susceptibility patterns of Candida parapsilosis isolated from blood cultures of patients were investigated in this study. Randomly amplified polymorphic DNA (RAPD) analysis generated 5 unique profiles from 42 isolates. Based on the major DNA fragments of the RAPD profiles, the isolates were identified as RAPD type P1 (29 isolates), P2 (6 isolates), P3 (4 isolates), P4 (2 isolates) and P5 (1 isolate). Sequence analysis of the internal transcribed spacer (ITS) gene of the isolates identified RAPD type P1 as C. parapsilosis, P2 and P3 as Candida orthopsilosis, P4 as Candida metapsilosis, and P5 as Lodderomyces elongisporus. Nucleotide variations in ITS gene sequences of C. orthopsilosis and C. metapsilosis were detected. Antifungal susceptibility testing using Etests showed that all isolates tested in this study were susceptible to amphotericin B, fluconazole, ketoconazole, itraconazole and voriconazole. C. parapsilosis isolates exhibited higher MIC(50) values than those of C. orthopsilosis for all of the drugs tested in this study; however, no significant difference in the MICs for these two Candida species was observed. The fact that C. orthopsilosis and C. metapsilosis were responsible for 23.8 and 4.8 % of the cases attributed to C. parapsilosis bloodstream infections, respectively, indicates the clinical relevance of these newly described yeasts. Further investigations of the ecological niche, mode of transmission and virulence of these species are thus essential.

  16. Effectiveness of oral antibiotics for definitive therapy of Gram-negative bloodstream infections.

    Science.gov (United States)

    Kutob, Leila F; Justo, Julie Ann; Bookstaver, P Brandon; Kohn, Joseph; Albrecht, Helmut; Al-Hasan, Majdi N

    2016-11-01

    There is paucity of data evaluating intravenous-to-oral antibiotic switch options for Gram-negative bloodstream infections (BSIs). This retrospective cohort study examined the effectiveness of oral antibiotics for definitive treatment of Gram-negative BSI. Patients with Gram-negative BSI hospitalised for antibiotics were included in this study. The cohort was stratified into three groups based on bioavailability of oral antibiotics prescribed (high, ≥95%; moderate, 75-94%; and low, antibiotics were prescribed to 106, 179 and 77 patients, respectively, for definitive therapy of Gram-negative BSI. Mean patient age was 63 years, 217 (59.9%) were women and 254 (70.2%) had a urinary source of infection. Treatment failure rates were 2%, 12% and 14% in patients receiving oral antibiotics with high, moderate and low bioavailability, respectively (P = 0.02). Risk of treatment failure in the multivariate Cox model was higher in patients receiving antibiotics with moderate [adjusted hazard ratio (aHR) = 5.9, 95% CI 1.6-38.5; P = 0.005] and low bioavailability (aHR = 7.7, 95% CI 1.9-51.5; P = 0.003) compared with those receiving oral antimicrobial agents with high bioavailability. These data demonstrate the effectiveness of oral antibiotics with high bioavailability for definitive therapy of Gram-negative BSI. Risk of treatment failure increases as bioavailability of the oral regimen declines.

  17. Changes in plasma protein levels as an early indication of a bloodstream infection

    Science.gov (United States)

    Joenväärä, Sakari; Kaartinen, Johanna; Järvinen, Asko; Renkonen, Risto

    2017-01-01

    Blood culture is the primary diagnostic test performed in a suspicion of bloodstream infection to detect the presence of microorganisms and direct the treatment. However, blood culture is slow and time consuming method to detect blood stream infections or separate septic and/or bacteremic patients from others with less serious febrile disease. Plasma proteomics, despite its challenges, remains an important source for early biomarkers for systemic diseases and might show changes before direct evidence from bacteria can be obtained. We have performed a plasma proteomic analysis, simultaneously at the time of blood culture sampling from ten blood culture positive and ten blood culture negative patients, and quantified 172 proteins with two or more unique peptides. Principal components analysis, Orthogonal Projections to Latent Structures Discriminant Analysis (OPLS-DA) and ROC curve analysis were performed to select protein(s) features which can classify the two groups of samples. We propose a number of candidates which qualify as potential biomarkers to select the blood culture positive cases from negative ones. Pathway analysis by two methods revealed complement activation, phagocytosis pathway and alterations in lipid metabolism as enriched pathways which are relevant for the condition. Data are available via ProteomeXchange with identifier PXD005022. PMID:28235076

  18. Nosocomial bloodstream infections: organisms, risk factors and resistant phenotypes in the Brazilian University Hospital

    Directory of Open Access Journals (Sweden)

    Rosineide M. Ribas

    2007-06-01

    Full Text Available Bacteremia is one of the most frequent and challenging hospital-acquired infection and it is associated with high attributable morbidity and mortality and additional use of healthcare resources. The objective of this work was to determine the frequencies of its occurrence, organisms and resistance phenotypes associated to nosocomial acquired bloodstream infections. A total number of 51 nosocomial bacteremia by Gram-negative and 99 by Gram-positive were evaluated and compared during a 15-month period. The risk factors associated with these bacteremias were analyzed and antibiotic use and surgery were associated with bacteremia by Gram-negative and > 2 invasive devices with Gram-positive. The resistance phenotypes ESBL (extended-spectrum beta-lactamases (23.5% and AmpC/others (17.6% correspond to 41.2 % with predominance of E. agglomerans among AmpC (44.4% and K. pneumoniae among ESBLs (38.5%. Among S. aureus bacteremia, approximately 40% were associated to MRSA (methicillin-resistant Staphylococcus aureus.

  19. Risk Factors and Outcomes for Bloodstream Infections Secondary to Clostridium difficile Infection.

    Science.gov (United States)

    Falcone, Marco; Russo, Alessandro; Iraci, Federica; Carfagna, Paolo; Goldoni, Paola; Vullo, Vincenzo; Venditti, Mario

    2015-10-19

    We determined the incidence, risk factors, and outcomes of bloodstream infections (BSI) subsequent to Clostridium difficile infection (CDI). We performed a retrospective study of all patients with definite diagnosis of CDI admitted from January 2014 to December 2014 in two large hospitals in Rome. Two groups of patients were analyzed: those with CDI and subsequent BSI (CDI/BSI(+)) and those with CDI and no evidence of primary BSI (CDI/BSI(-)). Data about clinical features, microbiology, treatments, and mortality were obtained. Overall, 393 cases of CDI were included in the final analysis: 72 developed a primary nosocomial BSI, while 321 had CDI without microbiological and clinical evidence of BSI. Etiologic agents of BSI were Candida species (47.3%), Enterobacteriaceae (19.4%), enterococci (13.9%), and mixed infections (19.4%). In multivariate analysis, ribotype 027 status (odds ratio [OR], 6.5), CDI recurrence (OR, 5.5), severe CDI infection (OR, 8.3), and oral vancomycin at >500 mg/day (OR, 3.1) were recognized as factors independently associated with the development of nosocomial BSI. Thirty-day mortality from CDI diagnosis was higher for patients of the CDI/BSI(+) group than for the controls (38.9 versus 13.1%; P nosocomial BSI. Candida species and enteric bacteria appear to be the leading causative pathogens and are associated with poor outcomes.

  20. Haemodialysis catheter-related bloodstream infections: current treatment options and strategies for prevention.

    Science.gov (United States)

    Saxena, Anil K; Panhotra, Bodh R

    2005-03-05

    Regardless of the repeated reservations raised by countless researchers with reference to the use of catheters as vascular access for haemodialysis (HD), central venous catheters (CVCs) remain irreplaceable tools of the modern dialysis delivery system as a reliable option for the clinical situations requiring instant access to circulation, for various reasons. Patients on long-term haemodialysis are therefore at a significantly high risk for catheterrelated bloodstream infections (CRBSI) and ensuing serious complications. Although early systemic antibiotic treatment should include the coverage for Staphylococcus aureus, the pathogen with most devastating consequences including bacterial endocarditis; optimal treatment of CRBSI while preserving the catheter site, remains contentious. Nonetheless, catheter exchange over a guide wire and antimicrobial-anticoagulant "locks" have shown promising results as novel access salvage techniques. Despite the fact that a number of novel potentially useful strategies for the prevention of CRBSI are in the pipeline; equally essential however, remains the role of rigorous implementation of standard infection control measures for hygiene and aseptic handling of CVCs in long-term HD patients. The policy of increasing the AVF (arteriovenous fistula) prevalence beyond 50% while minimising the use of CVCs, dependent largely upon the timely referrals and prudently implemented pre-ESRD program - ought to have a positive impact on long-term HD outcomes.

  1. Seasonal trend and clinical presentation of Bacillus cereus bloodstream infection: association with summer and indwelling catheter.

    Science.gov (United States)

    Kato, K; Matsumura, Y; Yamamoto, M; Nagao, M; Ito, Y; Takakura, S; Ichiyama, S

    2014-08-01

    Bacillus cereus, an opportunistic pathogen, can cause fatal infection. However, B. cereus bloodstream infections (BSIs) have not been well characterised. From 2008 to 2013, B. cereus isolates from all of the specimens and patients with B. cereus BSIs were identified. Environmental samples were collected to detect B. cereus contamination. We also characterised the clinical presentation of B. cereus BSI through analyses of risk factors for BSI and mortality. A total of 217 clinical B. cereus isolates was detected. Fifty-one patients with nosocomial infections were diagnosed as B. cereus BSI, and 37 had contaminated blood cultures. The number of B. cereus isolates and BSI patients was significantly greater from June to September than from January to April (4.9 vs. 1.5 per month and 1.2 vs. 0.2, respectively). All BSIs were nosocomial and related to central or peripheral vascular catheter. Urinary catheter [odds ratio (OR) 6.93, 95% confidence interval (CI) 2.40-20.0] was the independent risk factor associated with BSI patients when compared to patients regarded as contaminated. In-hospital mortality among BSI patients was 20% and was associated with urinary catheter (OR 34.7, 95 % CI 1.89-63.6) and higher Charlson index (OR 1.99, 95 % CI 1.26-3.12). The number of B. cereus isolates and BSI increased during summer. Inpatients with indwelling vascular or urinary catheters should be carefully monitored for potential B. cereus BSIs.

  2. Risk factors and outcomes of imipenem-resistant Acinetobacter bloodstream infection in North-eastern Malaysia

    Institute of Scientific and Technical Information of China (English)

    Zakuan Zainy Deris; Mohd Nazri Shafei; Azian Harun

    2011-01-01

    Objective: To determine the risk factors and outcomes of imipenem-resistant Acinetobacterbaumannii (IRAB) bloodstream infection (BSI) cases, since there is very little publication on Acinetobacter baumannii infections from Malaysia. Methods: A cross sectional study of 41 cases (73.2%) of imipenem-sensitive Acinetobacter baumanii (ISAB) and 15 cases (26.8%) of IRAB was conducted in a teaching hospital which was located at North-Eastern state of Malaysia. Results:There was no independent risk factor for IRAB BSI identified but IRAB BSI was significantly associated with longer bacteraemic days [OR 1.23 (95% CI 1.01, 1.50)]. Although prior use of carbepenems and cephalosporin were higher among IRAB than ISAB group, statistically they were not significant. There was no significant difference in term of outcomes between the two groups. Conclusions: Although statistically not significant, this analysis compliments previous publication highlighting the importance of appropriate empiric antibiotic usage in hospital especially carbepenems and need further evaluation with bigger subjects.

  3. Species distribution and antifungal susceptibility profile of Candida spp. bloodstream isolates from Latin American hospitals

    Directory of Open Access Journals (Sweden)

    Godoy Patrício

    2003-01-01

    Full Text Available From March 1999 to March 2000, we conducted a prospective multicenter study of candidemia involving five tertiary care hospitals from four countries in Latin America. Yeast isolates were identified by classical methods and the antifungal susceptibility profile was determined according to the National Committee for Clinical Laboratory Standards microbroth assay method. During a 12 month-period we were able to collect a total of 103 bloodstream isolates of Candida spp. C. albicans was the most frequently isolated species accounting for 42% of all isolates. Non-albicans Candida species strains accounted for 58% of all episodes of candidemia and were mostly represented by C. tropicalis (24.2% and C. parapsilosis (21.3%. It is noteworthy that we were able to identify two cases of C. lusitaniae from different institutions. In our casuistic, non-albicans Candida species isolates related to candidemic episodes were susceptible to fluconazole. Continuously surveillance programs are needed in order to identify possible changes in the species distribution and antifungal susceptibility patterns of yeasts that may occurs after increasing the use of azoles in Latin American hospitals.

  4. Central venous catheter-related bloodstream infection caused by Staphylococcus aureus: microbiology and risk factors

    Directory of Open Access Journals (Sweden)

    Geraldo Sadoyma

    2006-04-01

    Full Text Available Although central vascular catheters (CVC are indispensable in modern medicine, they are an important risk factor for primary bacteremias. We examined the incidence and risk factors associated with catheter-related bloodstream infection (CR-BSI caused by Staphylococcus aureus in surgical patients. A prospective study was carried out in the Hospital das Clínicas da Universidade Federal de Uberlândia (HC-UFU from September 2000 to December 2002. The skin insertion site, catheter tip, and blood were microbiologically analyzed. Demographics and risk factors were recorded for each patient, and cultures were identified phenotypically. Staphylococcus aureus was the most frequent pathogen, with an incidence rate of 4.9 episodes of CR-BSIs per 1,000 catheter/days. Based on logistic regression, the independent risk factors were: colonization on the insertion site =200 colony forming units (CFU/20 cm² (p=0.03; odds ratio (OR =6.89 and catheter tip (p=0.01; OR=7.95. The CR-BSI rate was high; it was mainly associated with S. aureus, and skin colonization at the insertion site and on the catheter tip were important risk factors for CR-BSI.

  5. Magnet® Hospital Recognition Linked to Lower Central Line-Associated Bloodstream Infection Rates.

    Science.gov (United States)

    Barnes, Hilary; Rearden, Jessica; McHugh, Matthew D

    2016-04-01

    Central-line-associated bloodstream infections (CLABSI) are among the deadliest heathcare-associated infections, with an estimated 12-25% mortality rate. In 2014, the Centers for Medicare and Medicaid Services (CMS) began to penalize hospitals for poor performance with respect to selected hospital-acquired conditions, including CLABSI. A structural factor associated with high-quality nursing care and better patient outcomes is The Magnet Recognition Program®. The purpose of this study was to explore the relationship between Magnet status and hospital CLABSI rates. We used propensity score matching to match Magnet and non-Magnet hospitals with similar hospital characteristics. In a matched sample of 291 Magnet hospitals and 291 non-Magnet hospitals, logistic regression models were used to examine whether there was a link between Magnet status and CLABSI rates. Both before and after matching, Magnet hospital status was associated with better (lower than the national average) CLABSI rates (OR = 1.60, 95%CI: 1.10, 2.33 after matching). While established programs such as Magnet recognition are consistently correlated with high-quality nursing work environments and positive patient outcomes, additional research is needed to determine whether Magnet designation produces positive patient outcomes or rewards existing excellence.

  6. Antifungal susceptibility survey of 2,000 bloodstream Candida isolates in the United States.

    Science.gov (United States)

    Ostrosky-Zeichner, Luis; Rex, John H; Pappas, Peter G; Hamill, Richard J; Larsen, Robert A; Horowitz, Harold W; Powderly, William G; Hyslop, Newton; Kauffman, Carol A; Cleary, John; Mangino, Julie E; Lee, Jeannette

    2003-10-01

    Candida bloodstream isolates (n = 2,000) from two multicenter clinical trials carried out by the National Institute of Allergy and Infectious Diseases Mycoses Study Group between 1995 and 1999 were tested against amphotericin B (AMB), flucytosine (5FC), fluconazole (FLU), itraconazole (ITR), voriconazole (VOR), posaconazole (POS), caspofungin (CFG), micafungin (MFG), and anidulafungin (AFG) using the NCCLS M27-A2 microdilution method. All drugs were tested in the NCCLS-specified RPMI 1640 medium except for AMB, which was tested in antibiotic medium 3. A sample of isolates was also tested in RPMI 1640 supplemented to 2% glucose and by using the diluent polyethylene glycol (PEG) in lieu of dimethyl sulfoxide for those drugs insoluble in water. Glucose supplementation tended to elevate the MIC, whereas using PEG tended to decrease the MIC. Trailing growth occurred frequently with azoles. Isolates were generally susceptible to AMB, 5FC, and FLU. Rates of resistance to ITR approached 20%. Although no established interpretative breakpoints are available for the candins (CFG, MFG, and AFG) and the new azoles (VOR and POS), they all exhibited excellent antifungal activity, even for those strains resistant to the other aforementioned agents.

  7. Agglutination of Trypanosoma cruzi in infected cells treated with serum from chronically infected mice.

    Science.gov (United States)

    Wendelken, Jennifer L; Rowland, Edwin C

    2009-04-01

    The protozoan parasite Trypanosoma cruzi is the causative agent of Chagas disease. The chronic stage of infection is characterized by a production of neutralizing antibodies in the vertebrate host. A polyclonal antibody, anti-egressin, has been found to inhibit egress of parasites from the host cell late in the intracellular cycle, after the parasites have transformed from the replicative amastigote into the trypomastigote. It has also been found that BALB/c mouse fibroblasts in the late stages of parasite infection become permeable to molecules as large as antibodies, leading to the possibility that anti-egressin affects the intracellular parasites. This project addresses the fate of the intracellular trypomastigotes that have been inhibited from egressing the host cell. Extended cultures of infected fibroblasts treated with chronic mouse serum reduced parasite egress at all time points measured. Parasites released from infected fibroblasts treated with chronic serum had a reduced ability to infect fibroblasts in culture, yet did not lose infectivity entirely. Absorption of chronic serum with living trypomastigotes removed the anti-egressin effect. The possibility that the target of anti-egressin is a parasite surface component is further indicated by the agglutination of extracellular trypomastigotes by chronic serum. The possibility that cross-linking by antibody occurs intracellularly, thus inhibiting egress, was reinforced by cleaving purified IgG into Fab fragments, which did not inhibit egress when added to infected cultures. From this work, it is proposed that the current, best explanation of the mechanism of egress inhibition by anti-egressin is intracellular agglutination, preventing normal parasite-driven egress.

  8. Functional characterization of enzymes involved in cysteine biosynthesis and H(2)S production in Trypanosoma cruzi.

    Science.gov (United States)

    Marciano, Daniela; Santana, Marianela; Nowicki, Cristina

    2012-10-01

    Trypanosoma cruzi is expected to synthetize de novo cysteine by different routes, among which the two-step pathway involving serine acetyltransferase and cysteine synthase (CS) is comprised. Also, cystathionine β synthase (CBS) might contribute to the de novo generation of cysteine in addition to catalyze the first step of the reverse transsulfuration route producing cystathionine. However, neither the functionality of CS nor that of cystathionine γ lyase (CGL) has been assessed. Our results show that T. cruzi CS could participate notably more actively than CBS in the de novo synthesis of cysteine. Interestingly, at the protein level T. cruzi CS is more abundant in amastigotes than in epimastigotes. Unlike the mammalian homologues, T. cruzi CGL specifically cleaves cystathionine into cysteine and is unable to produce H(2)S. The expression pattern of T. cruzi CGL parallels that of CBS, which unexpectedly suggests that in addition to the de novo synthesis of cysteine, the reverse transsulfuration pathway could be operative in the mammalian and insect stages. Besides, T. cruzi CBS produces H(2)S by decomposing cysteine or via condensation of cysteine with homocysteine. The latter reaction leads to cystathionine production, and is catalyzed remarkably more efficiently than the breakdown of cysteine. In T. cruzi like in other organisms, H(2)S could exert regulatory effects on varied metabolic processes. Notably, T. cruzi seems to count on stage-specific routes involved in cysteine production, the multiple cysteine-processing alternatives could presumably reflect this parasite's high needs of reducing power for detoxification of reactive oxygen species.

  9. Development of an aptamer-based concentration method for the detection of Trypanosoma cruzi in blood.

    Directory of Open Access Journals (Sweden)

    Rana Nagarkatti

    Full Text Available Trypanosoma cruzi, a blood-borne parasite, is the etiological agent of Chagas disease. T. cruzi trypomastigotes, the infectious life cycle stage, can be detected in blood of infected individuals using PCR-based methods. However, soon after a natural infection, or during the chronic phase of Chagas disease, the number of parasites in blood may be very low and thus difficult to detect by PCR. To facilitate PCR-based detection methods, a parasite concentration approach was explored. A whole cell SELEX strategy was utilized to develop serum stable RNA aptamers that bind to live T. cruzi trypomastigotes. These aptamers bound to the parasite with high affinities (8-25 nM range. The highest affinity aptamer, Apt68, also demonstrated high specificity as it did not interact with the insect stage epimastigotes of T. cruzi nor with other related trypanosomatid parasites, L. donovani and T. brucei, suggesting that the target of Apt68 was expressed only on T. cruzi trypomastigotes. Biotinylated Apt68, immobilized on a solid phase, was able to capture live parasites. These captured parasites were visible microscopically, as large motile aggregates, formed when the aptamer coated paramagnetic beads bound to the surface of the trypomastigotes. Additionally, Apt68 was also able to capture and aggregate trypomastigotes from several isolates of the two major genotypes of the parasite. Using a magnet, these parasite-bead aggregates could be purified from parasite-spiked whole blood samples, even at concentrations as low as 5 parasites in 15 ml of whole blood, as detected by a real-time PCR assay. Our results show that aptamers can be used as pathogen specific ligands to capture and facilitate PCR-based detection of T. cruzi in blood.

  10. Development of an Aptamer-Based Concentration Method for the Detection of Trypanosoma cruzi in Blood

    Science.gov (United States)

    Nagarkatti, Rana; Bist, Vaibhav; Sun, Sirena; Fortes de Araujo, Fernanda; Nakhasi, Hira L.; Debrabant, Alain

    2012-01-01

    Trypanosoma cruzi, a blood-borne parasite, is the etiological agent of Chagas disease. T. cruzi trypomastigotes, the infectious life cycle stage, can be detected in blood of infected individuals using PCR-based methods. However, soon after a natural infection, or during the chronic phase of Chagas disease, the number of parasites in blood may be very low and thus difficult to detect by PCR. To facilitate PCR-based detection methods, a parasite concentration approach was explored. A whole cell SELEX strategy was utilized to develop serum stable RNA aptamers that bind to live T. cruzi trypomastigotes. These aptamers bound to the parasite with high affinities (8–25 nM range). The highest affinity aptamer, Apt68, also demonstrated high specificity as it did not interact with the insect stage epimastigotes of T. cruzi nor with other related trypanosomatid parasites, L. donovani and T. brucei, suggesting that the target of Apt68 was expressed only on T. cruzi trypomastigotes. Biotinylated Apt68, immobilized on a solid phase, was able to capture live parasites. These captured parasites were visible microscopically, as large motile aggregates, formed when the aptamer coated paramagnetic beads bound to the surface of the trypomastigotes. Additionally, Apt68 was also able to capture and aggregate trypomastigotes from several isolates of the two major genotypes of the parasite. Using a magnet, these parasite-bead aggregates could be purified from parasite-spiked whole blood samples, even at concentrations as low as 5 parasites in 15 ml of whole blood, as detected by a real-time PCR assay. Our results show that aptamers can be used as pathogen specific ligands to capture and facilitate PCR-based detection of T. cruzi in blood. PMID:22927983

  11. Anti-Trypanosoma cruzi and anti-laminin antibodies in chagasic patients after specific treatment.

    Science.gov (United States)

    Gazzinelli, R T; Galvão, L M; Cardoso, J E; Cançado, J R; Krettli, A U; Brener, Z; Gazzinelli, G

    1988-01-01

    The antibody response to Trypanosoma cruzi epimastigote and trypomastigote stages, as well as to laminin, was studied in several groups of chagasic patients. In six patients who were cured of the parasite, the serum antibody titers as revealed by indirect immunofluorescence and hemagglutination tests against epimastigotes (conventional serology) and a complement-mediated lysis test with living trypomastigotes did not differ from those of normal individuals. In seven presumably cured patients, although the complement-mediated lysis test turned negative, conventional serology remained positive. Sera from this group of so-called "dissociated" patients presented significant lower mean antibody titers against epimastigote but not trypomastigote stages than did sera from 14 untreated patients (P less than 0.01). Most of the antibodies against trypomastigotes, including the residual levels found in cured patients, were absorbed by mouse laminin. In fact, significantly higher titers of anti-laminin antibodies were observed in sera from untreated chagasic patients (1.131 +/- 0.458) and cured patients (1.103 +/- 0.572) than in sera from eight normal individuals (0.459 +/- 0.402) (P less than 0.01). The anti-laminin titers were higher in sera of patients of blood group A or O than in those of patients of group B or AB. In Western blotting (immunoblotting) analysis against trypomastigotes, sera from chronic untreated patients recognized many polypeptide bands ranging from 26 to 160 kilodaltons, whereas no protein bands were observed with sera from cured patients. Only faint bands of parasite proteins were observed with sera of dissociated patients. In conjunction, the above data suggest that the anti-trypomastigote antibodies which persist after parasitological cure of patients with Chagas' disease are due mainly to cross-reactive epitopes from mouse laminin. PMID:3141467

  12. Implication of CA repeated tracts on post-transcriptional regulation in Trypanosoma cruzi.

    Science.gov (United States)

    Pastro, Lucía; Smircich, Pablo; Pérez-Díaz, Leticia; Duhagon, María Ana; Garat, Beatriz

    2013-08-01

    In Trypanosoma cruzi gene expression regulation mainly relays on post-transcriptional events. Nevertheless, little is known about the signals which control mRNA abundance and functionality. We have previously found that CA repeated tracts (polyCA) are abundant in the vicinity of open reading frames and constitute specific targets for single stranded binding proteins from T. cruzi epimastigote. Given the reported examples of the involvement of polyCA motifs in gene expression regulation, we decided to further study their role in T. cruzi. Using an in silico genome-wide analysis, we identify the genes that contain polyCA within their predicted UTRs. We found that about 10% of T. cruzi genes carry polyCA therein. Strikingly, they are frequently concurrent with GT repeated tracts (polyGT), favoring the formation of a secondary structure exhibiting the complementary polydinucleotides in a double stranded helix. This feature is found in the species-specific family of genes coding for mucine associated proteins (MASPs) and other genes. For those polyCA-containing UTRs that lack polyGT, the polyCA is mainly predicted to adopt a single stranded structure. We further analyzed the functional role of such element using a reporter approach in T. cruzi. We found out that the insertion of polyCA at the 3' UTR of a reporter gene in the pTEX vector modulates its expression along the parasite's life cycle. While no significant change of the mRNA steady state of the reporter gene could be detected at the trypomastigote stage, significant increase in the epimastigote and reduction in the amastigote stage were observed. Altogether, these results suggest the involvement of polyCA as a signal in gene expression regulation in T. cruzi.

  13. Transcriptome Remodeling in Trypanosoma cruzi and Human Cells during Intracellular Infection.

    Directory of Open Access Journals (Sweden)

    Yuan Li

    2016-04-01

    Full Text Available Intracellular colonization and persistent infection by the kinetoplastid protozoan parasite, Trypanosoma cruzi, underlie the pathogenesis of human Chagas disease. To obtain global insights into the T. cruzi infective process, transcriptome dynamics were simultaneously captured in the parasite and host cells in an infection time course of human fibroblasts. Extensive remodeling of the T. cruzi transcriptome was observed during the early establishment of intracellular infection, coincident with a major developmental transition in the parasite. Contrasting this early response, few additional changes in steady state mRNA levels were detected once mature T. cruzi amastigotes were formed. Our findings suggest that transcriptome remodeling is required to establish a modified template to guide developmental transitions in the parasite, whereas homeostatic functions are regulated independently of transcriptomic changes, similar to that reported in related trypanosomatids. Despite complex mechanisms for regulation of phenotypic expression in T. cruzi, transcriptomic signatures derived from distinct developmental stages mirror known or projected characteristics of T. cruzi biology. Focusing on energy metabolism, we were able to validate predictions forecast in the mRNA expression profiles. We demonstrate measurable differences in the bioenergetic properties of the different mammalian-infective stages of T. cruzi and present additional findings that underscore the importance of mitochondrial electron transport in T. cruzi amastigote growth and survival. Consequences of T. cruzi colonization for the host include dynamic expression of immune response genes and cell cycle regulators with upregulation of host cholesterol and lipid synthesis pathways, which may serve to fuel intracellular T. cruzi growth. Thus, in addition to the biological inferences gained from gene ontology and functional enrichment analysis of differentially expressed genes in parasite and

  14. Infection of C57BL/6 mice by Trypanosoma musculi modulates host immune responses during Brucella abortus cocolonization.

    Science.gov (United States)

    Lowry, Jake E; Leonhardt, Jack A; Yao, Chaoqun; Belden, E Lee; Andrews, Gerard P

    2014-01-01

    Brucellosis, which results in fetal abortions in domestic and wildlife animal populations, is of major concern in the US and throughout much of the world. The disease, caused by Brucella abortus, poses an economic threat to agriculture-based communities. A moderately efficacious live attenuated vaccine (B. abortus strain RB51) exists. However, even with vaccine use, outbreaks occur. Evidence suggests that elk (Cervus canadensis), a wild host reservoir, are the source of recent outbreaks in domestic cattle herds in Wyoming, USA. Brucella abortus establishes a chronic, persistent infection in elk. The molecular mechanisms allowing the establishment of this persistent infective state are currently unknown. A potential mechanism could be that concurrent pathogen burdens contribute to persistence. In Wyoming, elk are chronically infected with Trypanosoma cervi, which may modulate host responses in a similar manner to that documented for other trypanosomes. To identify any synergistic relationship between the two pathogens, we simulated coinfection in the well-established murine brucellosis model using Trypanosoma musculi and B. abortus S19. Groups of C57BL/6 mice (Mus musculus) were infected with either B. abortus strain 19 (S19) or T. musculi or both. Sera were collected weekly; spleens from euthanized mice were tested to determine bacterial load near the end of normal brucellosis infection. Although changes in bacterial load were observed during the later stages of brucellosis in those mice coinfected with T. musculi, the most significant finding was the suppression of gamma interferon early during the infection along with an increase in interleukin-10 secretion compared with mice infected with either pathogen alone. These results suggest that immune modulatory events occur in the mouse during coinfection and that further experiments are warranted to determine if T. cervi impacts Brucella infection in elk.

  15. Staging Mobilities

    DEFF Research Database (Denmark)

    Jensen, Ole B.

    In recent years, the social sciences have taken a “mobilities turn.” There has been a developing realisation that mobilities do not “just happen.” Mobilities are carefully and meticulously designed, planned and staged (from above). However, they are equally importantly acted out, performed...... that mobility is more than movement between point A and B. It explores how the movement of people, goods, information, and signs influences human understandings of self, other and the built environment. Moving towards a new understanding of the relationship between movement, interaction and environments......, the book asks: what are the physical, social, technical, and cultural conditions to the staging of contemporary urban mobilities?...

  16. Proteomic Analysis of Trypanosoma cruzi Epimastigotes Subjected to Heat Shock

    Directory of Open Access Journals (Sweden)

    Deyanira Pérez-Morales

    2012-01-01

    Full Text Available Trypanosoma cruzi is exposed to sudden temperature changes during its life cycle. Adaptation to these variations is crucial for parasite survival, reproduction, and transmission. Some of these conditions may change the pattern of genetic expression of proteins involved in homeostasis in the course of stress treatment. In the present study, the proteome of T. cruzi epimastigotes subjected to heat shock and epimastigotes grow normally was compared by two-dimensional gel electrophoresis followed by mass spectrometry for protein identification. Twenty-four spots differing in abundance were identified. Of the twenty-four changed spots, nineteen showed a greater intensity and five a lower intensity relative to the control. Several functional categories of the identified proteins were determined: metabolism, cell defense, hypothetical proteins, protein fate, protein synthesis, cellular transport, and cell cycle. Proteins involved in the interaction with the cellular environment were also identified, and the implications of these changes are discussed.

  17. Periurban Trypanosoma cruzi-infected Triatoma infestans, Arequipa, Peru.

    Science.gov (United States)

    Levy, Michael Zachary; Bowman, Natalie M; Kawai, Vivian; Waller, Lance A; Cornejo del Carpio, Juan Geny; Cordova Benzaquen, Eleazar; Gilman, Robert H; Bern, Caryn

    2006-09-01

    In Arequipa, Peru, vectorborne transmission of Chagas disease by Triatoma infestans has become an urban problem. We conducted an entomologic survey in a periurban community of Arequipa to identify risk factors for triatomine infestation and determinants of vector population densities. Of 374 households surveyed, triatomines were collected from 194 (52%), and Trypanosoma cruzi-carrying triatomines were collected from 72 (19.3%). Guinea pig pens were more likely than other animal enclosures to be infested and harbored 2.38x as many triatomines. Stacked brick and adobe enclosures were more likely to have triatomines, while wire mesh enclosures were protected against infestation. In human dwellings, only fully stuccoed rooms were protected against infestation. Spatially, households with triatomines were scattered, while households with T. cruzi-infected triatomines were clustered. Keeping small animals in wire mesh cages could facilitate control of T. infestans in this densely populated urban environment.

  18. Bed Bugs (Cimex lectularius) as Vectors of Trypanosoma cruzi

    Science.gov (United States)

    Salazar, Renzo; Castillo-Neyra, Ricardo; Tustin, Aaron W.; Borrini-Mayorí, Katty; Náquira, César; Levy, Michael Z.

    2015-01-01

    Populations of the common bed bug, Cimex lectularius, have recently undergone explosive growth. Bed bugs share many important traits with triatomine insects, but it remains unclear whether these similarities include the ability to transmit Trypanosoma cruzi, the etiologic agent of Chagas disease. Here, we show efficient and bidirectional transmission of T. cruzi between hosts and bed bugs in a laboratory environment. Most bed bugs that fed on experimentally infected mice acquired the parasite. A majority of previously uninfected mice became infected after a period of cohabitation with exposed bed bugs. T. cruzi was also transmitted to mice after the feces of infected bed bugs were applied directly to broken host skin. Quantitative bed bug defecation measures were similar to those of important triatomine vectors. Our findings suggest that the common bed bug may be a competent vector of T. cruzi and could pose a risk for vector-borne transmission of Chagas disease. PMID:25404068

  19. Electron Microscopy Analysis of the Nucleolus of Trypanosoma cruzi

    Science.gov (United States)

    López-Velázquez, Gabriel; Hernández, Roberto; López-Villaseñor, Imelda; Reyes-Vivas, Horacio; Segura-Valdez, María De L.; Jiménez-García, Luis F.

    2005-08-01

    The nucleolus is the main site for synthesis and processing of ribosomal RNA in eukaryotes. In mammals, plants, and yeast the nucleolus has been extensively characterized by electron microscopy, but in the majority of the unicellular eukaryotes no such studies have been performed. Here we used ultrastructural cytochemical and immunocytochemical techniques as well as three-dimensional reconstruction to analyze the nucleolus of Trypanosoma cruzi, which is an early divergent eukaryote of medical importance. In T. cruzi epimastigotes the nucleolus is a spherical intranuclear ribonucleoprotein organelle localized in a relatively central position within the nucleus. Dense fibrillar and granular components but not fibrillar centers were observed. In addition, nuclear bodies resembling Cajal bodies were observed associated to the nucleolus in the surrounding nucleoplasm. Our results provide additional morphological data to better understand the synthesis and processing of the ribosomal RNA in kinetoplastids.

  20. Predominance of Trypanosoma cruzi I among Panamanian sylvatic isolates.

    Science.gov (United States)

    Samudio, Franklyn; Ortega-Barría, Eduardo; Saldaña, Azael; Calzada, Jose

    2007-02-01

    Trypanosoma cruzi is throughout Panama, which is in agreement with the widespread of the sylvatic vectors implicated in the transmission. Eco-epidemiological changes in some regions of the country have led to a successful dissemination of the palm-tree Attalea butyracea and a possible adaptation of the primary vector of Chagas' disease to human settlements. These facts might increase both vector-human contact and human infection with different potentials T. cruzi genotypes and make therefore necessary a study to disclose Panamanian T. cruzi make-up. In this study, 71 T. cruzi isolates from Rhodnius pallescens were analyzed using mini-exon gene and sequence-characterized amplified region markers. The analyzed strains were T. cruzi lineage I. This finding along with prior results indicates that T. cruzi I is the principal genotype circulating in both sylvatic and domestic/peridomestic cycles and consequently responsible for the disease in the country.

  1. Proteomic analysis of two Trypanosoma cruzi zymodeme 3 strains.

    Science.gov (United States)

    Kikuchi, Simone A; Sodré, Cátia L; Kalume, Dário E; Elias, Camila G R; Santos, André L S; de Nazaré Soeiro, Maria; Meuser, Marcus; Chapeaurouge, Alex; Perales, Jonas; Fernandes, Octavio

    2010-12-01

    Two Trypanosoma cruzi Z3 strains, designated as 3663 and 4167, were previously isolated from insect vectors captured in the Brazilian Amazon region. These strains exhibited different infection patterns in Vero, C6/36, RAW 264.7 and HEp-2 cell lineages, in which 3663 trypomastigote form was much less infective than 4167 ones. A proteomic approach was applied to investigate the differences in the global patterns of protein expression in these two Z3 strains. Two-dimensional (2D) protein maps were generated and certain spots were identified by mass spectrometry (MS). Our analyses revealed a significant difference in the expression profile of different proteins between strains 3663 and 4167. Among them, cruzipain, an important regulator of infectivity. This data was corroborated by flow cytometry analysis using anti-cruzipain antibody. This difference could contribute to the infectivity profiles observed for each strain by in vitro assay using different cell lines.

  2. Parasite Genome Projects and the Trypanosoma cruzi Genome Initiative

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    Wim Degrave

    1997-11-01

    Full Text Available Since the start of the human genome project, a great number of genome projects on other "model" organism have been initiated, some of them already completed. Several initiatives have also been started on parasite genomes, mainly through support from WHO/TDR, involving North-South and South-South collaborations, and great hopes are vested in that these initiatives will lead to new tools for disease control and prevention, as well as to the establishment of genomic research technology in developing countries. The Trypanosoma cruzi genome project, using the clone CL-Brener as starting point, has made considerable progress through the concerted action of more than 20 laboratories, most of them in the South. A brief overview of the current state of the project is given

  3. Bed bugs (Cimex lectularius) as vectors of Trypanosoma cruzi.

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    Salazar, Renzo; Castillo-Neyra, Ricardo; Tustin, Aaron W; Borrini-Mayorí, Katty; Náquira, César; Levy, Michael Z

    2015-02-01

    Populations of the common bed bug, Cimex lectularius, have recently undergone explosive growth. Bed bugs share many important traits with triatomine insects, but it remains unclear whether these similarities include the ability to transmit Trypanosoma cruzi, the etiologic agent of Chagas disease. Here, we show efficient and bidirectional transmission of T. cruzi between hosts and bed bugs in a laboratory environment. Most bed bugs that fed on experimentally infected mice acquired the parasite. A majority of previously uninfected mice became infected after a period of cohabitation with exposed bed bugs. T. cruzi was also transmitted to mice after the feces of infected bed bugs were applied directly to broken host skin. Quantitative bed bug defecation measures were similar to those of important triatomine vectors. Our findings suggest that the common bed bug may be a competent vector of T. cruzi and could pose a risk for vector-borne transmission of Chagas disease.

  4. Novo transmissor silvestre do "Trypanosoma (Schizotrypanum cruzi" (Chagas, 1909

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    F. Nery Guimarães

    1943-06-01

    Full Text Available In nests of opossum (Didelphis aurita, localized in palm-trees of the species Attalea indaya Dr., the authors found a new tritatoma, the description of which is being made by Dr. H. LENT. They verified that this triatoma had been naturally infected by Trypanosoma (Schizotrhypanum cruzy. Two guinea-pigs were subsequently infected by peritoneal inoculation of excrements of this new triatoma. The xenodiagnosis of these guinea-pigs, made with normal nymphas of. T. megistus and T. infestans resulted positive after 25 days. Evidence was obtained of being the opossum (Didelphis one of the sources infection of the new vector, because several specimens of them were found infected, and also a specimen of D. aurita, which contained trypanosomes with the morphology of T. (S. cruzy in the peripheral blood.

  5. Mosaic VSGs and the scale of Trypanosoma brucei antigenic variation.

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    James P J Hall

    Full Text Available A main determinant of prolonged Trypanosoma brucei infection and transmission and success of the parasite is the interplay between host acquired immunity and antigenic variation of the parasite variant surface glycoprotein (VSG coat. About 0.1% of trypanosome divisions produce a switch to a different VSG through differential expression of an archive of hundreds of silent VSG genes and pseudogenes, but the patterns and extent of the trypanosome diversity phenotype, particularly in chronic infection, are unclear. We applied longitudinal VSG cDNA sequencing to estimate variant richness and test whether pseudogenes contribute to antigenic variation. We show that individual growth peaks can contain at least 15 distinct variants, are estimated computationally to comprise many more, and that antigenically distinct 'mosaic' VSGs arise from segmental gene conversion between donor VSG genes or pseudogenes. The potential for trypanosome antigenic variation is probably much greater than VSG archive size; mosaic VSGs are core to antigenic variation and chronic infection.

  6. Trypanosoma cruzi: orchiectomy and dehydroepiandrosterone therapy in infected rats.

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    Filipin, Marina Del Vecchio; Brazão, Vânia; Caetano, Leony Cristina; Santello, Fabricia Helena; Toldo, Míriam Paula Alonso; Caetano, Luana Naiara; do Prado, José Clóvis

    2008-11-01

    The ability of gonadal hormones to influence and induce diverse immunological functions during the course of a number of parasitic infections has been extensively studied in the latest decades. Dehydroepiandrosterone and its sulfate are the most abundant steroid hormones secreted by the human adrenal cortex and are considered potent immune-activators. The effects of orchiectomy on the course of Trypanosoma cruzi infection in rats, treated and untreated with DHEA were examined, by comparing blood and cardiac parasitism, macrophage numbers, nitric oxide and IFN-gamma levels. Orchiectomy enhanced resistance against infection with elevated numbers of macrophages, enhanced concentrations of NO and IFN-gamma and reduced amastigote burdens in heart when compared to control animals. DHEA replacement exerted a synergistic effect, up-modulating the immune response. Male sex steroids appear to play fundamental role in determining the outcome of disease, through the regulation and modulation of the activity of the immune response.

  7. Isolation and characterization of paraflagellar proteins from Trypanosoma cruzi.

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    Saborio, J L; Manuel Hernandez, J; Narayanswami, S; Wrightsman, R; Palmer, E; Manning, J

    1989-03-05

    Two different Trypanosoma cruzi polypeptides, with masses of 70 and 68 kDa were purified and characterized in this work. These two polypeptides designated PAR 1 and PAR 2, respectively, co-purified during each step of the isolation procedure and were found to be located exclusively in T. cruzi flagella by indirect immunofluorescence. A pre-embedding immunoelectron microscopy procedure, with a gold-tagged secondary antibody, permitted direct identification of PAR 2 as a component of the T. cruzi paraflagellar rod. PAR 1 and PAR 2 were found to be immunologically distinct and showed no cross-reactivity with actin, tubulin, intermediate filament proteins, or other proteins present in mammalian cells. The results presented indicate that PAR 1 and PAR 2 are the major components of T. cruzi paraflagellar filaments, and that these filaments have no counterpart in mammalian cells.

  8. Pathogenesis of anemia in Trypanosoma brucei-infected mice.

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    Amole, B O; Clarkson, A B; Shear, H L

    1982-01-01

    The pathogenesis of anemia was studied in trypanosome-infected mice. A strain of Trypanosoma brucei, TREU 667, was used which first produces an acute phase marked by waves of parasitemia. Erythrocytes from infected animals were coated with immunoglobulin M during or just before the waves of anemia and parasitological crises. Erythrocytes from normal animals could be sensitized with "precrisis" sera presumably containing antigen and antibody. These data suggest that anemia during the acute phase is due to sensitization of erythrocytes with immunoglobulin M-antigen complexes. The anemia is partially compensated by a strong erythropoietic response. The acute phase is followed by a chronic phase marked by a constant high parasitemia and immunosuppression. The less marked anemia occurring during this latter phase is due to hemodilution and perhaps a low but significant immune response to the parasites, which causes continuing erythrocyte sensitization by immunoglobulin M-antigen complexes. PMID:7201455

  9. Trypanosoma brucei: a putative RNA polymerase II promoter.

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    Bayele, Henry K

    2009-12-01

    RNA polymerase II (pol II) promoters are rare in the African trypanosome Trypanosoma brucei because gene regulation in the parasite is complex and polycistronic. Here, we describe a putative pol II promoter and its structure-function relationship. The promoter has features of an archetypal eukaryotic pol II promoter including putative canonical CCAAT and TATA boxes, and an initiator element. However, the spatial arrangement of these elements is only similar to yeast pol II promoters. Deletion mapping and transcription assays enabled delineation of a minimal promoter that could drive orientation-independent reporter gene expression suggesting that it may be a bidirectional promoter. In vitro transcription in a heterologous nuclear extract revealed that the promoter can be recognized by the basal eukaryotic transcription complex. This suggests that the transcription machinery in the parasite may be very similar to those of other eukaryotes.

  10. Amastigotes forms of Trypanosoma cruzi detected in a renal allograft

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    CARVALHO Maria Fernanda C.

    1997-01-01

    Full Text Available Trypanosoma cruzi, the causative agent of Chagas?disease assumes two distinct forms in vertebrate hosts: circulating trypomastigote and tissular amastigote. This latter form infects predominantly the myocardium, smooth and skeletal muscle, and central nervous system. The present work describes for the first time the detection of amastigote forms of T. cruzi in the renal parenchyma of a kidney graft recipient one month after transplantation. The patient was serologically negative for Chagas?disease and received no blood transfusion prior to transplant. The cadaver donor was from an endemic area for Chagas?disease. The recipient developed the acute form of the disease with detection of amastigote forms of T. cruzi in the renal allograft biopsy and circulating trypomastigote forms. The present report demonstrates that T. cruzi can infect the renal parenchyma. This mode of transmission warrants in endemic areas of Chagas?disease

  11. In vitro antitrypanosomal activity of plant terpenes against Trypanosoma brucei.

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    Otoguro, Kazuhiko; Iwatsuki, Masato; Ishiyama, Aki; Namatame, Miyuki; Nishihara-Tukashima, Aki; Kiyohara, Hiroaki; Hashimoto, Toshihiro; Asakawa, Yoshinori; Omura, Satoshi; Yamada, Haruki

    2011-11-01

    During the course of screening to discover antitrypanosomal compounds, 24 known plant terpenes (6 sesquiterpenes, 14 sesquiterpene lactones and 4 diterpenes) were evaluated for in vitro antitrypanosomal activity against Trypanosoma brucei brucei. Among them, 22 terpenes exhibited antitrypanosomal activity. In particular, α-eudesmol, hinesol, nardosinone and 4-peroxy-1,2,4,5-tetrahydro-α-santonin all exhibited selective and potent antitrypanosomal activities in vitro. Detailed here in an in vitro antitrypanosomal properties and cytotoxicities of the 24 terpenes compared with two therapeutic antitrypanosomal drugs (eflornithine and suramin). This finding represents the first report of promising trypanocidal activity of these terpenes. Present results also provide some valuable insight with regard to structure-activity relationships and the possible mode of action of the compounds.

  12. Mother-to-Child Transmission of Trypanosoma cruzi.

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    Gebrekristos, Hirut T; Buekens, Pierre

    2014-09-01

    Among the world's most neglected tropical diseases, Chagas disease is vector-borne and caused by Trypanosoma cruzi. T cruzi infection is endemic to South and Central America as well as Mexico. Due to population migration, T cruzi is increasingly becoming a public health problem in nonendemic settings. Success with vector control strategies has led to a relative increase in the burden attributable to congenital transmission of T cruzi. In endemic settings, approximately 5% of infected pregnant women transmit to their offspring. Congenital T cruzi infection is generally asymptomatic and parasitological and serological testing is required for diagnosis. This review highlights research gaps with a focus on (1) improving screening, diagnostic, and treatment options and (2) designing epidemiologic studies to understand risk factors for congenital T cruzi.

  13. An iron-binding Trypanosoma cruzi urinary antigen.

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    Corral, R S; Bertot, G M; Petray, P B; Altcheh, J M; Singh, M; Orn, A; Rapoport, M F; Grinstein, S

    1995-12-01

    An 80-kDa Trypanosoma cruzi urinary antigen (UAg) was affinity-purified from the urine of infected dogs. We demonstrated that UAg is structurally and functionally related to proteins belonging to the transferrin family, as shown by amino acid sequence and iron binding experiments. Nevertheless, monoclonal antibodies raised against UAg specifically and selectively recognized this parasite's circulating antigen. The existence of an 80-kDa T. cruzi antigen co-migrating with UAg could be confirmed when epimastigotes were metabolically labelled with [35S] methionine and then immunoprecipitated with the above mentioned antibodies. We conclude that UAg is an iron-binding T. cruzi component eliminated in the urine of the infected host.

  14. Mechanisms controlling anaemia in Trypanosoma congolense infected mice.

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    Harry A Noyes

    Full Text Available BACKGROUND: Trypanosoma congolense are extracellular protozoan parasites of the blood stream of artiodactyls and are one of the main constraints on cattle production in Africa. In cattle, anaemia is the key feature of disease and persists after parasitaemia has declined to low or undetectable levels, but treatment to clear the parasites usually resolves the anaemia. METHODOLOGY/PRINCIPAL FINDINGS: The progress of anaemia after Trypanosoma congolense infection was followed in three mouse strains. Anaemia developed rapidly in all three strains until the peak of the first wave of parasitaemia. This was followed by a second phase, characterized by slower progress to severe anaemia in C57BL/6, by slow recovery in surviving A/J and a rapid recovery in BALB/c. There was no association between parasitaemia and severity of anaemia. Furthermore, functional T lymphocytes are not required for the induction of anaemia, since suppression of T cell activity with Cyclosporin A had neither an effect on the course of infection nor on anaemia. Expression of genes involved in erythropoiesis and iron metabolism was followed in spleen, liver and kidney tissues in the three strains of mice using microarrays. There was no evidence for a response to erythropoietin, consistent with anaemia of chronic disease, which is erythropoietin insensitive. However, the expression of transcription factors and genes involved in erythropoiesis and haemolysis did correlate with the expression of the inflammatory cytokines Il6 and Ifng. CONCLUSIONS/SIGNIFICANCE: The innate immune response appears to be the major contributor to the inflammation associated with anaemia since suppression of T cells with CsA had no observable effect. Several transcription factors regulating haematopoiesis, Tal1, Gata1, Zfpm1 and Klf1 were expressed at consistently lower levels in C57BL/6 mice suggesting that these mice have a lower haematopoietic capacity and therefore less ability to recover from

  15. INFECÇÃO EXPERIMENTAL POR Trypanosoma evansi EM COELHOS EXPERIMENTAL INFECTION BY Trypanosoma evansi IN RABBITS

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    Silvia Gonzalez Monteiro

    2008-07-01

    Full Text Available

    Avaliou-se o período pré-patente e a evolução da parasitemia de coelhos infectados experimentalmente por Trypanosoma evansi. Dividiram-se coelhos machos, com idade entre dois a seis meses, em quatro grupos (A, B, C, D com três animais cada. Inocularam-se os coelhos dos grupos A, B e C com T. evansi e procedeu-se à avaliação diária, mediante esfregaços sangüíneos periféricos, observando-se que o período pré-patente nos grupos infectados (A, B e C foi de seis, sete e onze dias, respectivamente. Infectaram-se os animais do grupo A com T. evansi e Eimeria spp. e eles tiveram uma longevidade de 56 dias após o início do trabalho. Já os animais do grupo B, infectados com T. evansi e tratados com coccidiostático, apresentaram longevidade semelhante ao grupo C, que continha coelhos parasitados somente por T. evansi. Nesses dois grupos a parasitemia manteve-se com picos irregulares em variações de zero a cinco tripomastigotas/campo por até 89 dias após a inoculação, desaparecendo da circulação após 90 dias. Os coelhos do grupo D, grupo-controle, não foram infectados. Os animais dos grupos B, C e D foram sacrificados após 120 dias do início do experimento.

     

    Palavras-chaveS: Coelho, parasitemia, Trypanosoma evansi.

    This work had the aim to evaluate the pre-patent period and the evolution of the parasitemy of rabbits experimentally infected by Trypanosoma evansi. Male rabbits from two to six months were divided in four groups of three animals each (A, B, C, D. Group A, B and C rabbits were infected with T. evansi and evaluated daily through peripheral blood smears. The daily analyses showed a pre-patent period in the infected groups (A, B and C of six, seven and 11 days, respectively. Animals from group A, infected by T. evansi

  16. TbFlabarin, a flagellar protein of Trypanosoma brucei, highlights differences between Leishmania and Trypanosoma flagellar-targeting signals.

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    Tetaud, Emmanuel; Lefebvre, Michèle; M'Bang-Benet, Diane-Ethna; Crobu, Lucien; Blancard, Corinne; Sterkers, Yvon; Pages, Michel; Bastien, Patrick; Merlin, Gilles

    2016-07-01

    TbFlabarin is the Trypanosoma brucei orthologue of the Leishmania flagellar protein LdFlabarin but its sequence is 33% shorter than LdFlabarin, as it lacks a C-terminal domain that is indispensable for LdFlabarin to localize to the Leishmania flagellum. TbFlabarin is mainly expressed in the procyclic forms of the parasite and localized to the flagellum, but only when two palmitoylable cysteines at positions 3 and 4 are present. TbFlabarin is more strongly attached to the membrane fraction than its Leishmania counterpart, as it resists complete solubilization with as much as 0.5% NP-40. Expression ablation by RNA interference did not change parasite growth in culture, its morphology or apparent motility. Heterologous expression showed that neither TbFlabarin in L. amazonensis nor LdFlabarin in T. brucei localized to the flagellum, revealing non-cross-reacting targeting signals between the two species.

  17. Valores de transaminasas en cabras criollas infectadas con Trypanosoma vivax Transaminases values in Creole goats infected with Trypanosoma vivax

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    Emir Espinoza

    2002-01-01

    Full Text Available La presente comunicación reporta los valores de las enzimas transaminasas, Aspartatoaminotransferasa (AST y Alaninaaminotransferasa (ALT encontrados en sueros de cabras infectadas con la cepa de Trypanosoma vivax Stock (TvIIV y sus controles. Las determinaciones se realizaron durante un lapso experimental de diez semanas, divididos en dos períodos iguales (pre y post-infección por intermedio de un método colorimétrico, utilizando kits comerciales. Los datos fueron analizados mediante la prueba t Student's. En el caso de la AST, la comparación de las medias parciales de ambos grupos infectado y control, no indicó diferencias estadísticas. Con respecto a la ALT, la contrastación de las medias parciales de pre y post-infección del grupo de cabras infectadas, señaló diferencias significativas (PThe present communication reports the transaminases enzymes values Aspartatoaminotransferase (AST and Alaninaaminotransferase (ALT in serum from goats infected with the Trypanosoma vivax Stock (TvIIV. The determinations were realized during a ten week experimental period divided into two equal periods (pre- and post-infection by colorimetric method, using commercial kits. The dates were analyzed through the t Student's test. In the AST case, the comparison between partial means of infected and control groups did not show any statistical differences. In relation to ALT, the contrast of partial means to pre- and post-infection from infected goats group indicated significant differences (P<0.01.

  18. Molecular demonstration of Trypanosoma evansi and Trypanosoma lewisi DNA in wild rodents from Cambodia, Lao PDR and Thailand.

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    Milocco, C; Kamyingkird, K; Desquesnes, M; Jittapalapong, S; Herbreteau, V; Chaval, Y; Douangboupha, B; Morand, S

    2013-02-01

    In this study, we investigated the molecular evidence of Trypanosoma evansi in wild rodents from Cambodia, Lao PDR and Thailand. Between November 2007 and June 2009, 1664 rodents were trapped at eight sites representative of various ecological habitats. Of those animals, 94 were tested by direct microscopic blood examination, 633 using the Card Agglutination Test for Trypanosomes (CATT/T. evansi) and 145 by Polymerase Chain Reaction (PCR) with two sets of primers: TRYP1 (amplifying ITS1 of ribosomal DNA of all trypanosomes) and TBR (amplifying satellite genomic DNA of Trypanozoon parasites). Using TRYP1, based on the size of the PCR products, 15 samples from the three countries were positive for Trypanosoma lewisi (two were confirmed by sequencing), and three were positive for Trypanozoon (one was confirmed by sequencing and three by TBR primers); the specificity of the primers failed as rodent DNA was amplified in some cases. Using TBR, six samples were positive for Trypanozoon (one was confirmed by sequencing); as T. evansi is the only species of the Trypanozoon sub-genus possibly present in Asian rodents, these results confirmed its presence in rodents from Thailand (Rattus tanezumi) and Cambodia (R. tanezumi, Niviventer fulvescens & Maxomys surifer). Further investigations are necessary to establish the situation in Lao PDR. None of the 16 samples most strongly positive to the CATT proved to be positive for Trypanozoon by PCR. The merits of the CATT for such studies were not confirmed. Studying the urban and rural circulation of these parasites in rodents will enable an evaluation of human exposure and infection risk, as human infections by T. evansi were recently described in India and by T. lewisi in India and Thailand. As sequencing PCR products is expensive, the development of new molecular and serological tools for rodents would be very useful.

  19. Genotyping of Trypanosoma cruzi DTUs and Trypanosoma rangeli genetic groups in experimentally infected Rhodnius prolixus by PCR-RFLP.

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    Sá, Amanda R N; Dias, Greicy B M; Kimoto, Karen Y; Steindel, Mário; Grisard, Edmundo C; Toledo, Max Jean O; Gomes, Mônica L

    2016-04-01

    The specific detection and genetic typing of trypanosomes that infect humans, mammalian reservoirs, and vectors is crucial for diagnosis and epidemiology. We utilized a PCR-RFLP assay that targeted subunit II of cytochrome oxidase and 24Sα-rDNA to simultaneously detect and discriminate six Trypanosoma cruzi discrete typing units (DTUs) and two genetic groups of Trypanosoma rangeli (KP1+/KP1-) in intestinal contents of experimentally infected Rhodnius prolixus. The PCR assays showed that in 23 of 29 (79.4%) mixed infections with the six T. cruzi DTUs and mixed infections with individual DTUs and/or groups KP1+ and KP1-, both parasites were successfully detected. In six mixed infections that involved TcIII, the TcI, TcII, TcV, and TcVI DTUs predominated to the detriment of TcIII, indicating the selection of genetic groups. Interactions between different genetic groups and vectors may lead to genetic selection over TcIII. The elimination of this DTU by the immune system of the vector appears unlikely because TcIII was present in other mixed infections (TcIII/TcIV and TcIII/KP1+). Both molecular markers used in this study were sensitive and specific, demonstrating their usefulness in a wide geographical area where distinct genotypes of these two species are sympatric. Although the cellular and molecular mechanisms that are involved in parasite-vector interactions are still poorly understood, our results indicate a dynamic selection toward specific T. cruzi DTUs in R. prolixus during mixed genotype infections.

  20. Closed Catheter Access System Implementation in Reducing Bloodstream Infection Rate in Low Birth Weight Preterm Infants

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    Lily eRundjan

    2015-03-01

    Full Text Available Background Bloodstream infection (BSI is one of the significant causes of morbidity and mortality encountered in a neonatal intensive care unit (NICU, especially in developing countries. Despite the implementation of infection control practices, such as strict hand hygiene, the BSI rate in our hospital is still high. The use of a closed catheter access system to reduce BSI related to intravascular catheter has hitherto never been evaluated in our hospital. Objective To determine the effects of closed catheter access system implementation in reducing the BSI rate in preterm neonates with low birth weight.Methods Randomized clinical trial was conducted on 60 low birth weight preterm infants hospitalized in the neonatal unit at Cipto Mangunkusumo Hospital, Jakarta, Indonesia from June to September, 2013. Randomized subjects either received a closed or non-closed catheter access system. Subjects were monitored for 2 weeks for the development of BSI based on clinical signs, abnormal infection parameters, and blood culture. Results Closed catheter access system implementation gave a protective effect towards the occurrence of culture-proven BSI (relative risk 0.095, 95% CI 0.011 to 0.85, p=0.026. Risk of culture-proven BSI in the control group was 10.545 (95% CI 1.227 to 90.662, p=0.026. BSI occurred in 75% of neonates without risk factors of infection in the control group compared to none in the study group.Conclusions The use of a closed catheter access system reduced the BSI in low birth weight preterm infants. Choosing the right device design, proper disinfection of device and appropriate frequency of connector change should be done simultaneously.

  1. Bacillus Cereus catheter related bloodstream infection in a patient in a patient with acute lymphblastic leukemia

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    Lütfiye Öksüz

    2012-01-01

    Full Text Available

    Bacillus cereus infection is rarely associated with actual infection and for this reason single positive blood culture is usually regarded as contamination . However it may cause a number of infections, such catheter-related blood stream infections. Significant catheter-related bloodstream infections (CRBSI caused by Bacillus spp. are mainly due to B.cereus and have been predominantly reported in immunocompromised hosts1 . Catheter removal is generally advised for management of infection. In this report, catheter-related bacteremia caused by B.cereus in a patient with acute lymphoblastıc leukemia (ALL in Istanbul Medical Faculty was presented.A 44-year old man presented with fatigue, weight loss, epistaxis and high fever. A double-lumen Hickman–catheter (Bard 12.0 Fr, Round Dual Lumen was inserted by surgical cut-down to access the right subclavian vein which would be necessary for allogeneic stem cell transplantation. Three weeks later the patient presented with high fever and headache. Bacillus spp. was isolated from the cathether while blood culture obtained from the peripheral vein remained negative. The bacterial identification was confirmed as B.cereus using VITEK identification system

    It has been reported Bacillus cereus septicemia may be fatal in immunocompromised hosts despite broad-spectrum appropriate treatment10. Catheter removal is essential for prevention of recurrent bacteremia. Long-term cathater salvage should be reserved for appropriate patient group.

  2. Molecular epidemiology and antimicrobial resistance of methicillin-resistant Staphylococcus aureus bloodstream isolates in Taiwan, 2010.

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    Chih-Jung Chen

    Full Text Available The information of molecular characteristics and antimicrobial susceptibility pattern of methicillin-resistant Staphylococcus aureus (MRSA is essential for control and treatment of diseases caused by this medically important pathogen. A total of 577 clinical MRSA bloodstream isolates from six major hospitals in Taiwan were determined for molecular types, carriage of Panton-Valentine leukocidin (PVL and sasX genes and susceptibilities to 9 non-beta-lactam antimicrobial agents. A total of 17 genotypes were identified in 577 strains by pulsotyping. Five major pulsotypes, which included type A (26.2%, belonging to sequence type (ST 239, carrying type III staphylococcal chromosomal cassette mec (SCCmec, type F (18.9%, ST5-SCCmecII, type C (18.5%, ST59-SCCmecIV, type B (12.0%, ST239-SCCmecIII and type D (10.9%, ST59-SCCmecVT/IV, prevailed in each of the six sampled hospitals. PVL and sasX genes were respectively carried by ST59-type D strains and ST239 strains with high frequencies (93.7% and 99.1%, respectively but rarely detected in strains of other genotypes. Isolates of different genotypes and from different hospitals exhibited distinct antibiograms. Multi-resistance to ≥3 non-beta-lactams was more common in ST239 isolates (100% than in ST5 isolates (97.2%, P = 0.0347 and ST59 isolates (8.2%, P<0.0001. Multivariate analysis further indicated that the genotype, but not the hospital, was an independent factor associated with muti-resistance of the MRSA strains. In conclusion, five common MRSA clones with distinct antibiograms prevailed in the major hospitals in Taiwan in 2010. The antimicrobial susceptibility pattern of invasive MRSA was mainly determined by the clonal distribution.

  3. Prevalence and Antimicrobial Resistance of Microbes Causing Bloodstream Infections in Unguja, Zanzibar.

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    Annette Onken

    Full Text Available Bloodstream infections (BSI are frequent and cause high case-fatality rates. Urgent antibiotic treatment can save patients' lives, but antibiotic resistance can render antibiotic therapy futile. This study is the first to collect epidemiological data on BSI from Unguja, Zanzibar.Clinical data and blood for culturing and susceptibility testing of isolated microbes were obtained from 469 consecutively enrolled neonates, children and adults presenting with signs of systemic infections at Mnazi Mmoja Hospital (MMH, Zanzibar.Pathogenic bacteria were recovered from the blood of 14% of the patients (66/469. The most frequently isolated microbes were Klebsiella pneumoniae, Escherichia coli, Acinetobacter spp. and Staphylococcus aureus. Infections were community-acquired in 56 patients (85% and hospital-acquired in 8 (12% (data missing for 2 patients. BSI caused by extended-spectrum beta-lactamase (ESBL producing Enterobacteriaceae (E. coli, K. pneumoniae was found in 5 cases, of which 3 were community-acquired and 2 hospital-acquired. Three of these patients died. Six of 7 Salmonella Typhi isolates were multidrug resistant. Streptococcus pneumoniae was found in one patient only.This is the first report of ESBL-producing bacteria causing BSI from the Zanzibar archipelago. Our finding of community-acquired BSI caused by ESBL-producing bacteria is alarming, as it implies that these difficult-to-treat bacteria have already spread in the society. In the local setting these infections are virtually impossible to cure. The findings call for increased awareness of rational antibiotic use, infection control and surveillance to counteract the problem of emerging antimicrobial resistance.

  4. Outbreak of Pantoea agglomerans Bloodstream Infections at an Oncology Clinic-Illinois, 2012-2013.

    Science.gov (United States)

    Yablon, Brian R; Dantes, Raymund; Tsai, Victoria; Lim, Rachel; Moulton-Meissner, Heather; Arduino, Matthew; Jensen, Bette; Patel, Megan Toth; Vernon, Michael O; Grant-Greene, Yoran; Christiansen, Demian; Conover, Craig; Kallen, Alexander; Guh, Alice Y

    2017-03-01

    OBJECTIVE To determine the source of a healthcare-associated outbreak of Pantoea agglomerans bloodstream infections. DESIGN Epidemiologic investigation of the outbreak. SETTING Oncology clinic (clinic A). METHODS Cases were defined as Pantoea isolation from blood or catheter tip cultures of clinic A patients during July 2012-May 2013. Clinic A medical charts and laboratory records were reviewed; infection prevention practices and the facility's water system were evaluated. Environmental samples were collected for culture. Clinical and environmental P. agglomerans isolates were compared using pulsed-field gel electrophoresis. RESULTS Twelve cases were identified; median (range) age was 65 (41-78) years. All patients had malignant tumors and had received infusions at clinic A. Deficiencies in parenteral medication preparation and handling were identified (eg, placing infusates near sinks with potential for splash-back contamination). Facility inspection revealed substantial dead-end water piping and inadequate chlorine residual in tap water from multiple sinks, including the pharmacy clean room sink. P. agglomerans was isolated from composite surface swabs of 7 sinks and an ice machine; the pharmacy clean room sink isolate was indistinguishable by pulsed-field gel electrophoresis from 7 of 9 available patient isolates. CONCLUSIONS Exposure of locally prepared infusates to a contaminated pharmacy sink caused the outbreak. Improvements in parenteral medication preparation, including moving chemotherapy preparation offsite, along with terminal sink cleaning and water system remediation ended the outbreak. Greater awareness of recommended medication preparation and handling practices as well as further efforts to better define the contribution of contaminated sinks and plumbing deficiencies to healthcare-associated infections are needed. Infect Control Hosp Epidemiol 2017;38:314-319.

  5. Incidence, Clinical Characteristics and Attributable Mortality of Persistent Bloodstream Infection in the Neonatal Intensive Care Unit

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    Hsu, Jen-Fu; Chu, Shih-Ming; Lee, Chiang-Wen; Yang, Pong-Hong; Lien, Reyin; Chiang, Ming-Chou; Fu, Ren-Huei; Huang, Hsuan-Rong; Tsai, Ming-Horng

    2015-01-01

    Background An atypical pattern of neonatal sepsis, characterized by persistent positive blood culture despite effective antimicrobial therapy, has been correlated with adverse outcomes. However, previous studies focused only on coagulate-negative staphylococcus infection. Methods All episodes of persistent bloodstream infection (BSI), defined as 3 or more consecutive positive blood cultures with the same bacterial species, at least two of them 48 hours apart, during a single sepsis episode, were enrolled over an 8-year period in a tertiary level neonatal intensive care unit. These cases were compared with all non-persistent BSI during the same period. Results We identified 81 episodes of persistent BSI (8.5% of all neonatal late-onset sepsis) in 74 infants, caused by gram-positive pathogens (n=38, 46.9%), gram-negative pathogens (n=21, 25.9%), fungus (n=20, 24.7%) and polymicrobial bacteremia (n=2, 2.5%). Persistent BSI does not differ from non-persistent BSI in most clinical characteristics and patient demographics, but tends to have a prolonged septic course, longer duration of feeding intolerance and more frequent requirement of blood transfusions. No difference was observed for death attributable to infection (9.8% vs. 6.5%), but neonates with persistent BSI had significantly higher rates of infectious complications (29.6% vs. 9.2%, P < 0.001), death from all causes (21.6% vs. 11.7%, P = 0.025), and duration of hospitalization among survivors [median (interquartile range): 80.0 (52.5-117.5) vs. 64.0 (40.0-96.0) days, P = 0.005] than those without persistent BSI. Conclusions Although persistent BSI does not contribute directly to increased mortality, the associated morbidities, infectious complications and prolonged septic courses highlight the importance of aggressive treatment to optimize outcomes. PMID:25875677

  6. Using real time process measurements to reduce catheter related bloodstream infections in the intensive care unit

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    Wall, R; Ely, E; Elasy, T; Dittus, R; Foss, J; Wilkerson, K; Speroff, T

    2005-01-01

    

Problem: Measuring a process of care in real time is essential for continuous quality improvement (CQI). Our inability to measure the process of central venous catheter (CVC) care in real time prevented CQI efforts aimed at reducing catheter related bloodstream infections (CR-BSIs) from these devices. Design: A system was developed for measuring the process of CVC care in real time. We used these new process measurements to continuously monitor the system, guide CQI activities, and deliver performance feedback to providers. Setting: Adult medical intensive care unit (MICU). Key measures for improvement: Measured process of CVC care in real time; CR-BSI rate and time between CR-BSI events; and performance feedback to staff. Strategies for change: An interdisciplinary team developed a standardized, user friendly nursing checklist for CVC insertion. Infection control practitioners scanned the completed checklists into a computerized database, thereby generating real time measurements for the process of CVC insertion. Armed with these new process measurements, the team optimized the impact of a multifaceted intervention aimed at reducing CR-BSIs. Effects of change: The new checklist immediately provided real time measurements for the process of CVC insertion. These process measures allowed the team to directly monitor adherence to evidence-based guidelines. Through continuous process measurement, the team successfully overcame barriers to change, reduced the CR-BSI rate, and improved patient safety. Two years after the introduction of the checklist the CR-BSI rate remained at a historic low. Lessons learnt: Measuring the process of CVC care in real time is feasible in the ICU. When trying to improve care, real time process measurements are an excellent tool for overcoming barriers to change and enhancing the sustainability of efforts. To continually improve patient safety, healthcare organizations should continually measure their key clinical processes in real

  7. Clinical and microbiological characteristics of bloodstream infections in hematological cancer patients

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    V. N. Chebotkevich

    2016-01-01

    Full Text Available Introduction. Bloodstream infections (BSI are life-threatening illness for immunocompromised patients with hematological malignancies.The aim of the study was to compare epidemiology, causative pathogens and outcome of hospital-acquired BSI and clarifying the role of herpes group of viruses in their development.Materials and methods. During the period 1991–2013 438 bacterial strains obtained from 360 patients (pts with hematological malignancies wеre studied. All blood cultures were incubated in the continuous monitoring system for 7 days before discard. The real-time PCR was used for human herpesviruses DNA detection: Herpes simplex viruses types 1 and 2 (HSV 1, 2, Cytomegalovirus (CMV, Epstein–Barr virus(EBV and Herpesvirus 6 (HHV-6. In this study 64 hematological cancer patients with infectious complications who fulfilled criteria of systemic inflammatory response syndrome with positive peripheral blood cultures were investigated. All pts received empirical anti-infectious therapy with subsequent correction based on the bacteriological, virological and mycological analyses.Results and discussion. A total Gram-positive (G+ accounted for 69.2 % of BSI, Gram-negative (G– for 30.8 %. Among G+ BSI Coagulase Negative Staphylococci and Staphylococcus aureus were the most frequent pathogens (58.8 %, among G– BSI Escherichia coli (13.0 % was predominant. It is shown that the development of bacteremia were significantly more frequently occurs in the case of cytomegalovirusand the Epstein–Barr virus detection.Conclusion. Further epidemiological surveillance is warranted in order emerging resistant strains and related mortality. Reactivation of CMV and EBV is significantly associated with higher incidence of bacterial BSI.

  8. Hospital-wide multidisciplinary, multimodal intervention programme to reduce central venous catheter-associated bloodstream infection.

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    Walter Zingg

    Full Text Available Central line-associated bloodstream infection (CLABSI is the major complication of central venous catheters (CVC. The aim of the study was to test the effectiveness of a hospital-wide strategy on CLABSI reduction. Between 2008 and 2011, all CVCs were observed individually and hospital-wide at a large university-affiliated, tertiary care hospital. CVC insertion training started from the 3rd quarter and a total of 146 physicians employed or newly entering the hospital were trained in simulator workshops. CVC care started from quarter 7 and a total of 1274 nurses were trained by their supervisors using a web-based, modular, e-learning programme. The study included 3952 patients with 6353 CVCs accumulating 61,366 catheter-days. Hospital-wide, 106 patients had 114 CLABSIs with a cumulative incidence of 1.79 infections per 100 catheters. We observed a significant quarterly reduction of the incidence density (incidence rate ratios [95% confidence interval]: 0.92 [0.88-0.96]; P<0.001 after adjusting for multiple confounders. The incidence densities (n/1000 catheter-days in the first and last study year were 2.3/1000 and 0.7/1000 hospital-wide, 1.7/1000 and 0.4/1000 in the intensive care units, and 2.7/1000 and 0.9/1000 in non-intensive care settings, respectively. Median time-to-infection was 15 days (Interquartile range, 8-22. Our findings suggest that clinically relevant reduction of hospital-wide CLABSI was reached with a comprehensive, multidisciplinary and multimodal quality improvement programme including aspects of behavioural change and key principles of good implementation practice. This is one of the first multimodal, multidisciplinary, hospital-wide training strategies successfully reducing CLABSI.

  9. Bloodstream infection following 217 consecutive systemic-enteric drained pancreas transplants

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    Mark Walter

    2006-08-01

    Full Text Available Abstract Background Combined kidney pancreas transplantation (PTx evolved as excellent treatment for diabetic nephropathy. Infections remain common and serious complications. Methods 217 consecutive enteric drained PTxs performed from 1997 to 2004 were retrospectively analyzed with regard to bloodstream infection. Immunosuppression consisted of antithymocyteglobuline induction, tacrolimus, mycophenolic acid and steroids for the majority of cases. Standard perioperative antimicrobial prophylaxis consisted of pipercillin/tazobactam in combination with ciprofloxacin and fluconazole. Results One year patient, pancreas and kidney graft survival were 96.4%, 88.5% and 94.8%, surgical complication rate was 35%, rejection rate 30% and rate of infection 59%. In total 46 sepsis episodes were diagnosed in 35 patients (16% with a median onset on day 12 (range 1–45 post transplant. Sepsis source was intraabdominal infection (IAI (n = 21, a contaminated central venous line (n = 10, wound infection (n = 5, urinary tract infection (n = 2 and graft transmitted (n = 2. Nine patients (4% experienced multiple episodes of sepsis. Overall 65 pathogens (IAI sepsis 39, line sepsis 15, others 11 were isolated from blood. Gram positive cocci accounted for 50 isolates (77%: Coagulase negative staphylococci (n = 28, i.e. 43% (nine multi-resistant, Staphylococcus aureus (n = 11, i.e. 17% (four multi-resistant, enterococci (n = 9, i.e. 14% (one E. faecium. Gram negative rods were cultured in twelve cases (18%. Patients with blood borne infection had a two year pancreas graft survival of 76.5% versus 89.4% for those without sepsis (p = 0.036, patient survival was not affected. Conclusion Sepsis remains a serious complication after PTx with significantly reduced pancreas graft, but not patient survival. The most common source is IAI.

  10. Patterns and trends of pediatric bloodstream infections: a 7-year surveillance study.

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    Buetti, N; Atkinson, A; Kottanattu, L; Bielicki, J; Marschall, J; Kronenberg, A

    2017-03-01

    We characterize the epidemiology of pediatric bloodstream infections (BSIs) in Switzerland. We analyzed pathogen distribution and resistance patterns in monomicrobial and polymicrobial BSIs in children from 2008 to 2014 using data from the Swiss antibiotic resistance centre (ANRESIS). A confirmatory statistical analysis was performed comparing pathogens and resistance across 20 acute care hospitals. We identified 3,067 bacteremia episodes, of which 1,823 (59 %) were considered true BSI episodes. Overall, S. aureus (16.5 %, 300) was the most frequent pathogen, followed by E. coli (15.1 %, 276), coagulase-negative staphylococci (CoNS, 12.9 %, 235), S. pneumoniae (11.1 %, 202) and non-E. coli Enterobacteriaceae (8.7 %, 159). S. aureus and E. coli showed similar frequencies in all of the variables analyzed (e.g., hospital acquisition, hospital type, medical specialty). The proportion of these microorganisms did not change over time, resistance rates remained low (4.3 % methicillin resistance in S. aureus; 7.3 % third-/fourth-generation cephalosporin resistance in E. coli), and no significant resistance trends were observed. We observed a 50 % increase of CoNS BSIs from 2008 (9.8 %, 27) to 2014 (15.2 %, 46, p value for trend = 0.03). S. pneumoniae decreased from 17.5 % (48) to 6.6 % (20) during that timeframe (p for trend = 0.007). S. aureus and E. coli remained the most significant pathogens among pediatric BSIs in Switzerland, exhibiting low resistance rates. CoNS accounted for a greater proportion of BSIs over time. The decrease in bacteremic pneumococcal infections can likely be attributed to the introduction of the 13-valent conjugate vaccine in 2011.

  11. The Changing Epidemiology of Bloodstream Infections and Resistance in Hematopoietic Stem Cell Transplantation Recipients

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    Mücahit Yemişen

    2016-08-01

    Full Text Available Objective: Patients receiving hematopoietic stem cell transplantation (HSCT are exposed to highly immunosuppressive conditions and bloodstream infections (BSIs are one of the most common major complications within this period. Our aim, in this study, was to evaluate the epidemiology of BSIs in these patients retrospectively. Materials and Methods: The epidemiological properties of 312 patients with HSCT were retrospectively evaluated. Results: A total of 312 patients, followed between 2000 and 2011, who underwent autologous (62% and allogeneic (38% HSCT were included in the study. The most common underlying malignancies were multiple myeloma (28% and Hodgkin lymphoma (21.5%. A total of 142 (45% patients developed at least 1 episode of BSI and 193 separate pathogens were isolated from the blood cultures. There was a trend of increase in the numbers of BSIs in 2005-2008 and a relative increase in the proportion of gram-positive infections in recent years (2009-2011, and central venous catheter-related BSI was found to be most common source. Coagulase-negative staphylococci (49.2% and Acinetobacter baumannii (8.8% were the most common pathogens. Extended-spectrum beta-lactamase-producing strains were 23% and 22% among Escherichia coli and Klebsiella spp. isolates, respectively. Quinolone resistance was detected in 10% of Enterobacteriaceae. Resistance to carbapenems was not detected in Enterobacteriaceae, while it was seen at 11.1% and 23.5% in Pseudomonas and Acinetobacter strains, respectively. Conclusion: A shift was detected from gram-negative bacteria to gram-positive in the etiology over the years and central lines were the most common sources of BSIs.

  12. Risk factors and outcomes for patients with bloodstream infection due to Acinetobacter baumannii-calcoaceticus complex.

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    Chopra, Teena; Marchaim, Dror; Johnson, Paul C; Awali, Reda A; Doshi, Hardik; Chalana, Indu; Davis, Naomi; Zhao, Jing J; Pogue, Jason M; Parmar, Sapna; Kaye, Keith S

    2014-08-01

    Identifying patients at risk for bloodstream infection (BSI) due to Acinetobacter baumannii-Acinetobacter calcoaceticus complex (ABC) and providing early appropriate therapy are critical for improving patient outcomes. A retrospective matched case-control study was conducted to investigate the risk factors for BSI due to ABC in patients admitted to the Detroit Medical Center (DMC) between January 2006 and April 2009. The cases were patients with BSI due to ABC; the controls were patients not infected with ABC. Potential risk factors were collected 30 days prior to the ABC-positive culture date for the cases and 30 days prior to admission for the controls. A total of 245 case patients were matched with 245 control patients. Independent risk factors associated with BSI due to ABC included a Charlson's comorbidity score of ≥ 3 (odds ratio [OR], 2.34; P = 0.001), a direct admission from another health care facility (OR, 4.63; P < 0.0001), a prior hospitalization (OR, 3.11; P < 0.0001), the presence of an indwelling central venous line (OR, 2.75; P = 0.011), the receipt of total parenteral nutrition (OR, 21.2; P < 0.0001), the prior receipt of β-lactams (OR, 3.58; P < 0.0001), the prior receipt of carbapenems (OR, 3.18; P = 0.006), and the prior receipt of chemotherapy (OR, 15.42; P < 0.0001). The median time from the ABC-positive culture date to the initiation of the appropriate antimicrobial therapy was 2 days (interquartile range [IQR], 1 to 3 days). The in-hospital mortality rate was significantly higher among case patients than among control patients (OR, 3.40; P < 0.0001). BSIs due to ABC are more common among critically ill and debilitated institutionalized patients, who are heavily exposed to health care settings and invasive devices.

  13. Trypanosome Motion Represents an Adaptation to the Crowded Environment of the Vertebrate Bloodstream

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    Heddergott, Niko; Krüger, Timothy; Babu, Sujin B.; Wei, Ai; Stellamanns, Erik; Uppaluri, Sravanti; Pfohl, Thomas; Stark, Holger; Engstler, Markus

    2012-01-01

    Blood is a remarkable habitat: it is highly viscous, contains a dense packaging of cells and perpetually flows at velocities varying over three orders of magnitude. Only few pathogens endure the harsh physical conditions within the vertebrate bloodstream and prosper despite being constantly attacked by host antibodies. African trypanosomes are strictly extracellular blood parasites, which evade the immune response through a system of antigenic variation and incessant motility. How the flagellates actually swim in blood remains to be elucidated. Here, we show that the mode and dynamics of trypanosome locomotion are a trait of life within a crowded environment. Using high-speed fluorescence microscopy and ordered micro-pillar arrays we show that the parasites mode of motility is adapted to the density of cells in blood. Trypanosomes are pulled forward by the planar beat of the single flagellum. Hydrodynamic flow across the asymmetrically shaped cell body translates into its rotational movement. Importantly, the presence of particles with the shape, size and spacing of blood cells is required and sufficient for trypanosomes to reach maximum forward velocity. If the density of obstacles, however, is further increased to resemble collagen networks or tissue spaces, the parasites reverse their flagellar beat and consequently swim backwards, in this way avoiding getting trapped. In the absence of obstacles, this flagellar beat reversal occurs randomly resulting in irregular waveforms and apparent cell tumbling. Thus, the swimming behavior of trypanosomes is a surprising example of micro-adaptation to life at low Reynolds numbers. For a precise physical interpretation, we compare our high-resolution microscopic data to results from a simulation technique that combines the method of multi-particle collision dynamics with a triangulated surface model. The simulation produces a rotating cell body and a helical swimming path, providing a functioning simulation method for a

  14. A multicentre analysis of epidemiology of the nosocomial bloodstream infections in Japanese university hospitals.

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    Nagao, M

    2013-09-01

    Nosocomial bloodstream infections (BSIs) are an important cause of morbidity and mortality. The current study analysed data from a concurrent surveillance programme to examine the current epidemiological trends for nosocomial BSIs at 22 Japanese university hospitals from 1 April 2008 to 31 March 2012. The number of blood culture sets taken, the rate of multiple blood culture sets and the rates of antibiotic-resistant isolates among six major nosocomial BSI pathogens (Staphylococcus aureus, Enterococcus spp., Escherichia coli, Klebsiella spp., Pseudomonas aeruginosa, and Candida spp.) not including coagulase-negative staphylococci, were evaluated. The clinical characteristics of nosocomial BSIs caused by these pathogens were also collected for 2941 patients. The number of blood culture sets taken per bed increased during the 4-year study period (from 4.07 in 2008 to 5.37 in 2011), and the rates of multiple blood culture sets also increased (from 29.9% in 2008 to 50.0% in 2011). Methicillin resistance was detected in 50.2% of S. aureus isolates. The prevalence rates of extended-spectrum beta-lactamase-producing E. coli and Klebsiella spp. isolates increased annually during the study period, and the average prevalence rates were 12.3% and 5.8%, respectively. The overall crude mortality of nosocomial BSIs due to the six pathogens evaluated was 24.5% (43.2% in ICU settings and 20.5% in non-ICU settings). Thus, our multicentre study evaluated the current epidemiological trends for nosocomial BSIs, and we found that further efforts are needed to increase the use of multiple blood culture sets and improve the prognosis of nosocomial BSIs in Japanese university hospitals.

  15. Staphylococcus species and their Methicillin-Resistance in 7424 Blood Cultures for Suspected Bloodstream Infections

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    Ariana ALMAŞ

    2011-06-01

    Full Text Available Objectives: The aim of this study was to evaluate the distribution of Staphylococcus species in bloodstream infections and to assess their susceptibility to methicillin. Material and Methods: Between January 1st 2008 - December 31st 2010, 7424 blood culture sets were submitted to the Laboratory Department of the Hospital for Clinical Infectious Diseases in Cluj-Napoca, Romania. The blood cultures were performed using BacT/Alert until January 2010 and BacT/Alert 3D automated system (bioMérieux after that date. The blood culture bottles were incubated at 37°C in a continuously monitoring system for up to 7 days. The strain identifications were performed by conventional methods, ApiStaph galleries and Vitek 2 Compact system. Susceptibility to methicillin was determined by disk diffusion method with cefoxitin disk and by using Vitek 2 Compact system. Results: From the total number of performed blood cultures, 568 were positive with Staphylococcus species. From 168 bacteriemic episodes 103 were with Staphylococcus aureus. Among 65 coagulase-negative staphylococci isolates, Staphylococcus epidermidis was the most frequently isolated species (34, followed by Staphylococcus hominis (15, Staphylococcus haemolyticus (8, Staphylococcus saprophyticus (3, Staphylococcus cohnii (1, Staphylococcus auricularis (1, and 3 strains that were not identified at species level. Methicillin resistance was encountered in 53.40% of Staphylococcus aureus strains and in 80% of coagulase-negative staphylococci. Conclusions: An important percentage of blood cultures were contaminated with Staphylococcus species. The main species identified in true bacteriemia cases were Staphylococcus aureus and Staphylococcus epidermidis. The percentage of methicillin-resistance, proved to be high not only for coagulase-negative staphylococci but also for Staphylococcus aureus.

  16. Classification of positive blood cultures: computer algorithms versus physicians' assessment - development of tools for surveillance of bloodstream infection prognosis using population-based laboratory databases

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    Gradel Kim O

    2012-09-01

    Full Text Available Abstract Background Information from blood cultures is utilized for infection control, public health surveillance, and clinical outcome research. This information can be enriched by physicians’ assessments of positive blood cultures, which are, however, often available from selected patient groups or pathogens only. The aim of this work was to determine whether patients with positive blood cultures can be classified effectively for outcome research in epidemiological studies by the use of administrative data and computer algorithms, taking physicians’ assessments as reference. Methods Physicians’ assessments of positive blood cultures were routinely recorded at two Danish hospitals from 2006 through 2008. The physicians’ assessments classified positive blood cultures as: a contamination or bloodstream infection; b bloodstream infection as mono- or polymicrobial; c bloodstream infection as community- or hospital-onset; d community-onset bloodstream infection as healthcare-associated or not. We applied the computer algorithms to data from laboratory databases and the Danish National Patient Registry to classify the same groups and compared these with the physicians’ assessments as reference episodes. For each classification, we tabulated episodes derived by the physicians’ assessment and the computer algorithm and compared 30-day mortality between concordant and discrepant groups with adjustment for age, gender, and comorbidity. Results Physicians derived 9,482 reference episodes from 21,705 positive blood cultures. The agreement between computer algorithms and physicians’ assessments was high for contamination vs. bloodstream infection (8,966/9,482 reference episodes [96.6%], Kappa = 0.83 and mono- vs. polymicrobial bloodstream infection (6,932/7,288 reference episodes [95.2%], Kappa = 0.76, but lower for community- vs. hospital-onset bloodstream infection (6,056/7,288 reference episodes [83.1%], Kappa = 0.57 and

  17. Serological survey of Leishmania infantum and Trypanosoma cruzi in dogs from urban areas of Brazil and Colombia

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    Leishmania infantum and Trypanosoma cruzi are zoonotic parasites that are endemic throughout many parts of Latin America. Infected dogs play an important role in transmission of both parasites to humans. A serological survey of Leishmania and Trypanosoma infection was conducted on 365 dogs from São ...

  18. In vivo observation of the hypo-echoic "black hole" phenomenon in rat arterial bloodstream: a preliminary Study.

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    Nam, Kweon-Ho; Paeng, Dong-Guk

    2014-07-01

    The "black hole," a hypo-echoic hole at the center of the bloodstream surrounded by a hyper-echoic zone in cross-sectional views, has been observed in ultrasound backscattering measurements of blood with red blood cell aggregation in in vitro studies. We investigated whether the phenomenon occurs in the in vivo arterial bloodstream of rats using a high-frequency ultrasound imaging system. Longitudinal and cross-sectional ultrasound images of the rat common carotid artery (CCA) and abdominal aorta were obtained using a 40-MHz ultrasound system. A high-frame-rate retrospective imaging mode was employed to precisely examine the dynamic changes in blood echogenicity in the arteries. When the imaging was performed with non-invasive scanning, blood echogenicity was very low in the CCA as compared with the surrounding tissues, exhibiting no hypo-echoic zone at the center of the vessel. Invasive imaging of the CCA by incising the skin and subcutaneous tissues at the imaging area provided clearer and brighter blood echo images, showing the "black hole" phenomenon near the center of the vessel in longitudinal view. The "black hole" was also observed in the abdominal aorta under direct imaging after laparotomy. The aortic "black hole" was clearly observed in both longitudinal and cross-sectional views. Although the "black hole" was always observed near the center of the arteries during the diastolic phase, it dissipated or was off-center along with the asymmetric arterial wall dilation at systole. In conclusion, we report the first in vivo observation of the hypo-echoic "black hole" caused by the radial variation of red blood cell aggregation in arterial bloodstream.

  19. Biofilm formation is a risk factor for mortality in patients with Candida albicans bloodstream infection—Scotland, 2012–2013

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    Rajendran, R.; Sherry, L.; Nile, C.J.; Sherriff, A.; Johnson, E.M.; Hanson, M.F.; Williams, C.; Munro, C.A.; Jones, B.J.; Ramage, G.

    2016-01-01

    Bloodstream infections caused by Candida species remain a significant cause of morbidity and mortality in hospitalized patients. Biofilm formation by Candida species is an important virulence factor for disease pathogenesis. A prospective analysis of patients with Candida bloodstream infection (n = 217) in Scotland (2012–2013) was performed to assess the risk factors associated with patient mortality, in particular the impact of biofilm formation. Candida bloodstream isolates (n = 280) and clinical records for 157 patients were collected through 11 different health boards across Scotland. Biofilm formation by clinical isolates was assessed in vitro with standard biomass assays. The role of biofilm phenotype on treatment efficacy was also evaluated in vitro by treating preformed biofilms with fixed concentrations of different classes of antifungal. Available mortality data for 134 patients showed that the 30-day candidaemia case mortality rate was 41%, with predisposing factors including patient age and catheter removal. Multivariate Cox regression survival analysis for 42 patients showed a significantly higher mortality rate for Candida albicans infection than for Candida glabrata infection. Biofilm-forming ability was significantly associated with C. albicans mortality (34 patients). Finally, in vitro antifungal sensitivity testing showed that low biofilm formers and high biofilm formers were differentially affected by azoles and echinocandins, but not by polyenes. This study provides further evidence that the biofilm phenotype represents a significant clinical entity, and that isolates with this phenotype differentially respond to antifungal therapy in vitro. Collectively, these findings show that greater clinical understanding is required with respect to Candida biofilm infections, and the implications of isolate heterogeneity. PMID:26432192

  20. The changing epidemiology of Acinetobacter spp. producing OXA carbapenemases causing bloodstream infections in Brazil: a BrasNet report.

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    Vasconcelos, Ana Tereza R; Barth, Afonso L; Zavascki, Alexandre P; Gales, Ana C; Levin, Anna S; Lucarevschi, Bianca R; Cabral, Blenda G; Brasiliense, Danielle M; Rossi, Flavia; Furtado, Guilherme H C; Carneiro, Irna Carla R S; da Silva, Juliana O; Ribeiro, Julival; Lima, Karla V B; Correa, Luci; Britto, Maria H; Silva, Mariama T; da Conceição, Marília L; Moreira, Marina; Martino, Marinês D V; de Freitas, Marise R; Oliveira, Maura S; Dalben, Mirian F; Guzman, Ricardo D; Cayô, Rodrigo; Morais, Rosângela; Santos, Sânia A; Martins, Willames M B S

    2015-12-01

    We evaluated the epidemiology of Acinetobacter spp. recovered from patients diagnosed with bloodstream infections in 9 tertiary hospitals located in all Brazilian geographic regions between April and August 2014. Although OXA-23-producing Acinetobacter baumannii clones were disseminated in most hospitals, it was observed for the first time the spread of OXA-72 among clonally related A. baumannii isolated from distinct hospitals. Interestingly, Acinetobacter pittii was the most frequent species found in a Northern region hospital. Contrasting with the multisusceptible profile displayed by A. pittii isolates, the tetracyclines and polymyxins were the only antimicrobials active against all A. baumannii isolates.

  1. Inhibition of HIV-1 replication in human monocyte-derived macrophages by parasite Trypanosoma cruzi.

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    Guadalupe Andreani

    Full Text Available BACKGROUND: Cells of monocyte/macrophage lineage are one of the major targets of HIV-1 infection and serve as reservoirs for viral persistence in vivo. These cells are also the target of the protozoa Trypanosoma cruzi, the causative agent of Chagas disease, being one of the most important endemic protozoonoses in Latin America. It has been demonstrated in vitro that co-infection with other pathogens can modulate HIV replication. However, no studies at cellular level have suggested an interaction between T. cruzi and HIV-1 to date. METHODOLOGY/PRINCIPAL FINDINGS: By using a fully replicative wild-type virus, our study showed that T. cruzi inhibits HIV-1 antigen production by nearly 100% (p99% being stronger than HIV-T. cruzi (approximately 90% for BaL and approximately 85% for VSV-G infection. In MDM with established HIV-1 infection, T. cruzi significantly inhibited luciferate activity (p<0.01. By quantifying R-U5 and U5-gag transcripts by real time PCR, our study showed the expression of both transcripts significantly diminished in the presence of trypomastigotes (p<0.05. Thus, T. cruzi inhibits viral post-integration steps, early post-entry steps and entry into MDM. Trypomastigotes also caused a approximately 60-70% decrease of surface CCR5 expression on MDM. Multiplication of T. cruzi inside the MDM does not seem to be required for inhibiting HIV-1 replication since soluble factors secreted by trypomastigotes have shown similar effects. Moreover, the major parasite antigen cruzipain, which is secreted by the trypomastigote form, was able to inhibit viral production in MDM over 90% (p<0.01. CONCLUSIONS/SIGNIFICANCE: Our study showed that T. cruzi inhibits HIV-1 replication at several replication stages in macrophages, a major cell target for both pathogens.

  2. Trypanocidal Effect of Isotretinoin through the Inhibition of Polyamine and Amino Acid Transporters in Trypanosoma cruzi

    Science.gov (United States)

    Reigada, Chantal; Valera-Vera, Edward A.; Sayé, Melisa; Errasti, Andrea E.; Avila, Carla C.; Miranda, Mariana R.; Pereira, Claudio A.

    2017-01-01

    Polyamines are essential compounds to all living organisms and in the specific case of Trypanosoma cruzi, the causative agent of Chagas disease, they are exclusively obtained through transport processes since this parasite is auxotrophic for polyamines. Previous works reported that retinol acetate inhibits Leishmania growth and decreases its intracellular polyamine concentration. The present work describes a combined strategy of drug repositioning by virtual screening followed by in vitro assays to find drugs able to inhibit TcPAT12, the only polyamine transporter described in T. cruzi. After a screening of 3000 FDA-approved drugs, 7 retinoids with medical use were retrieved and used for molecular docking assays with TcPAT12. From the docked molecules, isotretinoin, a well-known drug used for acne treatment, showed the best interaction score with TcPAT12 and was selected for further in vitro studies. Isotretinoin inhibited the polyamine transport, as well as other amino acid transporters from the same protein family (TcAAAP), with calculated IC50 values in the range of 4.6–10.3 μM. It also showed a strong inhibition of trypomastigote burst from infected cells, with calculated IC50 of 130 nM (SI = 920) being significantly less effective on the epimastigote stage (IC50 = 30.6 μM). The effect of isotretinoin on the parasites plasma membrane permeability and on mammalian cell viability was tested, and no change was observed. Autophagosomes and apoptotic bodies were detected as part of the mechanisms of isotretinoin-induced death indicating that the inhibition of transporters by isotretinoin causes nutrient starvation that triggers autophagic and apoptotic processes. In conclusion, isotretinoin is a promising trypanocidal drug since it is a multi-target inhibitor of essential metabolites transporters, in addition to being an FDA-approved drug largely used in humans, which could reduce significantly the requirements for its possible application in the treatment of

  3. Biochemical characterization of trans-sialidase TS1 variants from Trypanosoma congolense

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    Dietz Frank

    2011-07-01

    Full Text Available Abstract Background Animal African trypanosomiasis, sleeping sickness in humans and Nagana in cattle, is a resurgent disease in Africa caused by Trypanosoma parasites. Trans-sialidases expressed by trypanosomes play an important role in the infection cycle of insects and mammals. Whereas trans-sialidases of other trypanosomes like the American T. cruzi are well investigated, relatively little research has been done on these enzymes of T. congolense. Results Based on a partial sequence and an open reading frame in the WTSI database, DNA sequences encoding for eleven T. congolense trans-sialidase 1 variants with 96.3% overall amino acid identity were amplified. Trans-sialidase 1 variants were expressed as recombinant proteins, isolated and assayed for trans-sialylation activity. The purified proteins produced α2,3-sialyllactose from lactose by desialylating fetuin, clearly demonstrating their trans-sialidase activity. Using an HPLC-based assay, substrate specificities and kinetic parameters of two variants were characterized in detail indicating differences in substrate specificities for lactose, fetuin and synthetic substrates. Both enzymes were able to sialylate asialofetuin to an extent, which was sufficient to reconstitute binding sites for Siglec-4. A mass spectrometric analysis of the sialylation pattern of glycopeptides from fetuin revealed clear but generally similar changes in the sialylation pattern of the N-glycans on fetuin catalyzed by the trans-sialidases investigated. Conclusions The identification and characterization of a trans-sialidase gene family of the African parasite T. congolense has opened new perspectives for investigating the biological role of these enzymes in Nagana and sleeping sickness. Based on this study it will be interesting to address the expression pattern of these genes and their activities in the different stages of the parasite in its infection cycle. Furthermore, these trans-sialidases have the

  4. A monoclonal antibody marker for the exclusion-zone filaments of Trypanosoma brucei

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    Decossas Marion

    2008-07-01

    Full Text Available Abstract Background Trypanosoma brucei is a haemoflagellate pathogen of man, wild animals and domesticated livestock in central and southern Africa. In all life cycle stages this parasite has a single mitochondrion that contains a uniquely organised genome that is condensed into a flat disk-like structure called the kinetoplast. The kinetoplast is essential for insect form procyclic cells and therefore is a potential drug target. The kinetoplast is unique in nature because it consists of novel structural proteins and thousands of circular, interlocking, DNA molecules (kDNA. Secondly, kDNA replication is critically timed to coincide with nuclear S phase and new flagellum biogenesis. Thirdly, the kinetoplast is physically attached to the flagellum basal bodies via a structure called the tripartite attachment complex (TAC. The TAC consists of unilateral filaments (within the mitochondrion matrix, differentiated mitochondrial membranes and exclusion-zone filaments that extend from the distal end of the basal bodies. To date only one protein, p166, has been identified to be a component of the TAC. Results In the work presented here we provide data based on a novel EM technique developed to label and characterise cytoskeleton structures in permeabilised cells without extraction of mitochondrion membranes. We use this protocol to provide data on a new monoclonal antibody reagent (Mab 22 and illustrate the precise localisation of basal body-mitochondrial linker proteins. Mab 22 binds to these linker proteins (exclusion-zone filaments and provides a new tool for the characterisation of cytoskeleton mediated kinetoplast segregation. Conclusion The antigen(s recognised by Mab 22 are cytoskeletal, insensitive to extraction by high concentrations of non-ionic detergent, extend from the proximal region of basal bodies and bind to the outer mitochondrial membrane. This protein(s is the first component of the TAC exclusion-zone fibres to be identified. Mab 22

  5. Signal transduction induced in Trypanosoma cruzi metacyclic trypomastigotes during the invasion of mammalian cells

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    N. Yoshida

    2000-03-01

    Full Text Available Penetration of Trypanosoma cruzi into mammalian cells depends on the activation of the parasite's protein tyrosine kinase and on the increase in cytosolic Ca2+ concentration. We used metacyclic trypomastigotes, the T. cruzi developmental forms that initiate infection in mammalian hosts, to investigate the association of these two events and to identify the various components of the parasite signal transduction pathway involved in host cell invasion. We have found that i both the protein tyrosine kinase activation, as measured by phosphorylation of a 175-kDa protein (p175, and Ca2+ mobilization were induced in the metacyclic forms by the HeLa cell extract but not by the extract of T. cruzi-resistant K562 cells; ii treatment of parasites with the tyrosine kinase inhibitor genistein blocked both p175 phosphorylation and the increase in cytosolic Ca2+ concentration; iii the recombinant protein J18, which contains the full-length sequence of gp82, a metacyclic stage surface glycoprotein involved in target cell invasion, interfered with tyrosine kinase and Ca2+ responses, whereas the monoclonal antibody 3F6 directed at gp82 induced parasite p175 phosphorylation and Ca2+ mobilization; iv treatment of metacyclic forms with phospholipase C inhibitor U73122 blocked Ca2+ signaling and impaired the ability of the parasites to enter HeLa cells, and v drugs such as heparin, a competitive IP3-receptor blocker, caffeine, which affects Ca2+ release from IP3-sensitive stores, in addition to thapsigargin, which depletes intracellular Ca2+ compartments and lithium ion, reduced the parasite infectivity. Taken together, these data suggest that protein tyrosine kinase, phospholipase C and IP3 are involved in the signaling cascade that is initiated on the parasite cell surface by gp82 and leads to Ca2+ mobilization required for target cell invasion.

  6. Limited Ability of Posaconazole To Cure both Acute and Chronic Trypanosoma cruzi Infections Revealed by Highly Sensitive In Vivo Imaging.

    Science.gov (United States)

    Francisco, Amanda Fortes; Lewis, Michael D; Jayawardhana, Shiromani; Taylor, Martin C; Chatelain, Eric; Kelly, John M

    2015-08-01

    The antifungal drug posaconazole has shown significant activity against Trypanosoma cruzi in vitro and in experimental murine models. Despite this, in a recent clinical trial it displayed limited curative potential. Drug testing is problematic in experimental Chagas disease because of difficulties in demonstrating sterile cure, particularly during the chronic stage of infection when parasite burden is extremely low and tissue distribution is ill defined. To better assess posaconazole efficacy against acute and chronic Chagas disease, we have exploited a highly sensitive bioluminescence imaging system which generates data with greater accuracy than other methods, including PCR-based approaches. Mice inoculated with bioluminescent T. cruzi were assessed by in vivo and ex vivo imaging, with cyclophosphamide-induced immunosuppression used to enhance the detection of relapse. Posaconazole was found to be significantly inferior to benznidazole as a treatment for both acute and chronic T. cruzi infections. Whereas 20 days treatment with benznidazole was 100% successful in achieving sterile cure, posaconazole failed in almost all cases. Treatment of chronic infections with posaconazole did however significantly reduce infection-induced splenomegaly, even in the absence of parasitological cure. The imaging-based screening system also revealed that adipose tissue is a major site of recrudescence in mice treated with posaconazole in the acute, but not the chronic stage of infection. This in vivo screening model for Chagas disease is predictive, reproducible and adaptable to diverse treatment schedules. It should provide greater assurance that drugs are not advanced prematurely into clinical trial.

  7. Two pathways of homologous recombination in Trypanosoma brucei.

    Science.gov (United States)

    Conway, Colin; Proudfoot, Chris; Burton, Peter; Barry, J David; McCulloch, Richard

    2002-09-01

    African trypanosomes are unicellular parasites that use DNA recombination to evade the mammalian immune response. They do this in a process called antigenic variation, in which the parasites periodically switch the expression of VSG genes that encode distinct Variant Surface Glycoprotein coats. Recombination is used to move new VSG genes into specialised bloodstream VSG transcription sites. Genetic and molecular evidence has suggested that antigenic variation uses homologous recombination, but the detailed reaction pathways are not understood. In this study, we examine the recombination pathways used by trypanosomes to integrate transformed DNA into their genome, and show that they possess at least two pathways of homologous recombination. The primary mechanism is dependent upon RAD51, but a subsidiary pathway exists that is RAD51-independent. Both pathways contribute to antigenic variation. We show that the RAD51-independent pathway is capable of recombining DNA substrates with very short lengths of sequence homology and in some cases aberrant recombination reactions can be detected using such microhomologies.

  8. Kinetics of Trypanosoma cruzi destruction in the mouse spleen

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    Zulmira M.S. Cordeiro

    1997-02-01

    Full Text Available Massive destruction of parasitized splenic macrophages was histologically observed at the height of a virulent infection caused by Trypanosoma cruzi (Y strain in the mouse. This was coincident with a sudden drop in parasitemic curve. Most of the animals died at this point, probably due to the liberation of toxic products, such as TNF, following the massive destruction of parasitized cells. However, parasitized-cell destruction indicated the transition from susceptibility to resistance. Although it has been extensively studied in vitro, this study contributes with the morphological counterpart observed in vivo by optical and electron microscopy. When infected animals were specifically treated during early infection transition to chronic phase was immediately observed without splenic parasitism. Animals that apparently recovered from massive cell-destruction in the spleen showed evidences of a rapid restoration of splenic architecture.Um estudo histologico sequenciado mostrou que o parasitismo dos macrófagos esplênicos por uma cepa virulenta (cepa Y do Trypanosoma cruzi tem um curso progressivo, mas chega até um ponto em que todos as células parasitadas são subitamente destruídas. Tal achado coincidiu com uma quéda brusca da curva parasitêmica e com a morte da maioria dos animais, provavelmente devido à liberação de produtos tóxicos (como o TNF pelas células desintegradas. O achado foi interpretado como o auge da transição entre uma fase de susceptibilidade e outra de resistência. Embora esta transição tenha sido bem estudada in vitro, este estudo contribui com os dados do substrato morfológico observados in vivo, através da microscopia ótica e eletrônica. O tratamento específico e supressivo feito na fase inicial da infecção acarreta uma transição imediata para a fase crônica e aí o parasitismo esplênico desaparece completamente. Os animais que aparentemente se recuperaram expontaneamente após a fase de destrui

  9. Trypanosoma brucei Invasion and T-Cell Infiltration of the Brain Parenchyma in Experimental Sleeping Sickness: Timing and Correlation with Functional Changes

    Science.gov (United States)

    Laperchia, Claudia; Palomba, Maria; Seke Etet, Paul F.; Rodgers, Jean; Bradley, Barbara; Montague, Paul; Grassi-Zucconi, Gigliola; Bentivoglio, Marina

    2016-01-01

    Background The timing of Trypanosoma brucei entry into the brain parenchyma to initiate the second, meningoencephalitic stage of human African trypanosomiasis or sleeping sickness is currently debated and even parasite invasion of the neuropil has been recently questioned. Furthermore, the relationship between neurological features and disease stage are unclear, despite the important diagnostic and therapeutic implications. Methodology Using a rat model of chronic Trypanosoma brucei brucei infection we determined the timing of parasite and T-cell neuropil infiltration and its correlation with functional changes. Parasite DNA was detected using trypanosome-specific PCR. Body weight and sleep structure alterations represented by sleep-onset rapid eye movement (SOREM) periods, reported in human and experimental African trypanosomiasis, were monitored. The presence of parasites, as well as CD4+ and CD8+ T-cells in the neuropil was assessed over time in the brain of the same animals by immunocytochemistry and quantitative analyses. Principal findings Trypanosome DNA was present in the brain at day 6 post-infection and increased more than 15-fold by day 21. Parasites and T-cells were observed in the parenchyma from day 9 onwards. Parasites traversing blood vessel walls were observed in the hypothalamus and other brain regions. Body weight gain was reduced from day 7 onwards. SOREM episodes started in most cases early after infection, with an increase in number and duration after parasite neuroinvasion. Conclusion These findings demonstrate invasion of the neuropil over time, after an initial interval, by parasites and lymphocytes crossing the blood-brain barrier, and show that neurological features can precede this event. The data thus challenge the current clinical and cerebrospinal fluid criteria of disease staging. PMID:28002454

  10. Molecular Identification and Echinocandin Susceptibility of Candida parapsilosis Complex Bloodstream Isolates in Italy, 2007-2014.

    Science.gov (United States)

    Lovero, Grazia; Borghi, Elisa; Balbino, Stella; Cirasola, Daniela; De Giglio, Osvalda; Perdoni, Federica; Caggiano, Giuseppina; Morace, Giulia; Montagna, Maria Teresa

    2016-01-01

    The Candida parapsilosis group encompasses three species: C. parapsilosis, C. orthopsilosis, and C. metapsilosis. Here, we describe the incidence and echinocandin susceptibility profile of bloodstream isolates of these three species collected from patients admitted to an Italian university hospital from 2007 to 2014. Molecular identification of cryptic species of the C. parapsilosis complex was performed using polymerase chain reaction amplification of the gene encoding secondary alcohol dehydrogenase, followed by digestion with the restriction enzyme BanI. Minimum inhibitory concentrations were determined using the broth microdilution method according to European Committee for Antimicrobial Susceptibility Testing (EUCAST EDef 7.2) and Clinical Laboratory Standards Institute (CLSI M27-A3) guidelines, and the results were compared with those obtained using the E-test and Sensititre methods. Of the 163 C. parapsilosis complex isolates, 136 (83.4%) were identified as C. parapsilosis, and 27 (16.6%) as C. orthopsilosis. The species-specific incidences were 2.9/10,000 admissions for C. parapsilosis and 0.6/10,000 admissions for C. orthopsilosis. No resistance to echinocandins was detected with any of the methods. The percent essential agreement (EA) between the EUCAST and E-test/Sensititre methods for anidulafungin, caspofungin, and micafungin susceptibility was, respectively, as follows: C. parapsilosis, 95.6/97.8, 98.5/88.2, and 93.4/96.3; C. orthopsilosis, 92.6/92.6, 96.3/77.8, and 63.0/66.7. The EA between the CLSI and E-test/Sensititre methods was, respectively, as follows: C. parapsilosis, 99.3/100, 98.5/89.0, and 96.3/98.5; C. orthopsilosis, 96.3/92.6, 100/81.5, and 92.6/88.9. Only minor discrepancies, ranging from 16.9% (C. parapsilosis) to 11.1% (C. orthopsilosis), were observed between the CLSI and E-test/Sensititre methods. In conclusion, this epidemiologic study shows a typical C. parapsilosis complex species distribution, no echinocandin resistance, and it

  11. Molecular Identification and Echinocandin Susceptibility of Candida parapsilosis Complex Bloodstream Isolates in Italy, 2007–2014

    Science.gov (United States)

    Lovero, Grazia; Borghi, Elisa; Balbino, Stella; Cirasola, Daniela; De Giglio, Osvalda; Perdoni, Federica; Caggiano, Giuseppina; Morace, Giulia; Montagna, Maria Teresa

    2016-01-01

    The Candida parapsilosis group encompasses three species: C. parapsilosis, C. orthopsilosis, and C. metapsilosis. Here, we describe the incidence and echinocandin susceptibility profile of bloodstream isolates of these three species collected from patients admitted to an Italian university hospital from 2007 to 2014. Molecular identification of cryptic species of the C. parapsilosis complex was performed using polymerase chain reaction amplification of the gene encoding secondary alcohol dehydrogenase, followed by digestion with the restriction enzyme BanI. Minimum inhibitory concentrations were determined using the broth microdilution method according to European Committee for Antimicrobial Susceptibility Testing (EUCAST EDef 7.2) and Clinical Laboratory Standards Institute (CLSI M27-A3) guidelines, and the results were compared with those obtained using the E-test and Sensititre methods. Of the 163 C. parapsilosis complex isolates, 136 (83.4%) were identified as C. parapsilosis, and 27 (16.6%) as C. orthopsilosis. The species-specific incidences were 2.9/10,000 admissions for C. parapsilosis and 0.6/10,000 admissions for C. orthopsilosis. No resistance to echinocandins was detected with any of the methods. The percent essential agreement (EA) between the EUCAST and E-test/Sensititre methods for anidulafungin, caspofungin, and micafungin susceptibility was, respectively, as follows: C. parapsilosis, 95.6/97.8, 98.5/88.2, and 93.4/96.3; C. orthopsilosis, 92.6/92.6, 96.3/77.8, and 63.0/66.7. The EA between the CLSI and E-test/Sensititre methods was, respectively, as follows: C. parapsilosis, 99.3/100, 98.5/89.0, and 96.3/98.5; C. orthopsilosis, 96.3/92.6, 100/81.5, and 92.6/88.9. Only minor discrepancies, ranging from 16.9% (C. parapsilosis) to 11.1% (C. orthopsilosis), were observed between the CLSI and E-test/Sensititre methods. In conclusion, this epidemiologic study shows a typical C. parapsilosis complex species distribution, no echinocandin resistance, and it

  12. Molecular Identification and Echinocandin Susceptibility of Candida parapsilosis Complex Bloodstream Isolates in Italy, 2007-2014.

    Directory of Open Access Journals (Sweden)

    Grazia Lovero

    Full Text Available The Candida parapsilosis group encompasses three species: C. parapsilosis, C. orthopsilosis, and C. metapsilosis. Here, we describe the incidence and echinocandin susceptibility profile of bloodstream isolates of these three species collected from patients admitted to an Italian university hospital from 2007 to 2014. Molecular identification of cryptic species of the C. parapsilosis complex was performed using polymerase chain reaction amplification of the gene encoding secondary alcohol dehydrogenase, followed by digestion with the restriction enzyme BanI. Minimum inhibitory concentrations were determined using the broth microdilution method according to European Committee for Antimicrobial Susceptibility Testing (EUCAST EDef 7.2 and Clinical Laboratory Standards Institute (CLSI M27-A3 guidelines, and the results were compared with those obtained using the E-test and Sensititre methods. Of the 163 C. parapsilosis complex isolates, 136 (83.4% were identified as C. parapsilosis, and 27 (16.6% as C. orthopsilosis. The species-specific incidences were 2.9/10,000 admissions for C. parapsilosis and 0.6/10,000 admissions for C. orthopsilosis. No resistance to echinocandins was detected with any of the methods. The percent essential agreement (EA between the EUCAST and E-test/Sensititre methods for anidulafungin, caspofungin, and micafungin susceptibility was, respectively, as follows: C. parapsilosis, 95.6/97.8, 98.5/88.2, and 93.4/96.3; C. orthopsilosis, 92.6/92.6, 96.3/77.8, and 63.0/66.7. The EA between the CLSI and E-test/Sensititre methods was, respectively, as follows: C. parapsilosis, 99.3/100, 98.5/89.0, and 96.3/98.5; C. orthopsilosis, 96.3/92.6, 100/81.5, and 92.6/88.9. Only minor discrepancies, ranging from 16.9% (C. parapsilosis to 11.1% (C. orthopsilosis, were observed between the CLSI and E-test/Sensititre methods. In conclusion, this epidemiologic study shows a typical C. parapsilosis complex species distribution, no echinocandin

  13. Comparison of the systemic inflammatory response syndrome between monomicrobial and polymicrobial Pseudomonas aeruginosa nosocomial bloodstream infections

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    Wenzel Richard P

    2005-10-01

    Full Text Available Abstract Background Some studies of nosocomial bloodstream infection (nBSI have demonstrated a higher mortality for polymicrobial bacteremia when compared to monomicrobial nBSI. The purpose of this study was to compare differences in systemic inflammatory response and mortality between monomicrobial and polymicrobial nBSI with Pseudomonas aeruginosa. Methods We performed a historical cohort study on 98 adults with P. aeruginosa (Pa nBSI. SIRS scores were determined 2 days prior to the first positive blood culture through 14 days afterwards. Monomicrobial (n = 77 and polymicrobial BSIs (n = 21 were compared. Results 78.6% of BSIs were caused by monomicrobial P. aeruginosa infection (MPa and 21.4% by polymicrobial P. aeruginosa infection (PPa. Median APACHE II score on the day of BSI was 22 for MPa and 23 for PPa BSIs. Septic shock occurred in 33.3% of PPa and in 39.0% of MPa (p = 0.64. Progression to septic shock was associated with death more frequently in PPa (OR 38.5, CI95 2.9–508.5 than MPa (OR 4.5, CI95 1.7–12.1. Maximal SIR (severe sepsis, septic shock or death was seen on day 0 for PPa BSI vs. day 1 for MPa. No significant difference was noted in the incidence of organ failure, 7-day or overall mortality between the two groups. Univariate analysis revealed that APACHE II score ≥20 at BSI onset, Charlson weighted comorbidity index ≥3, burn injury and respiratory, cardiovascular, renal and hematologic failure were associated with death, while age, malignant disease, diabetes mellitus, hepatic failure, gastrointestinal complications, inappropriate antimicrobial therapy, infection with imipenem resistant P. aeruginosa and polymicrobial nBSI were not. Multivariate analysis revealed that hematologic failure (p Conclusion In this historical cohort study of nBSI with P. aeruginosa, the incidence of septic shock and organ failure was high in both groups. Additionally, patients with PPa BSI were not more acutely ill, as judged by APACHE II

  14. A soluble 3-hydroxy-3-methylglutaryl-CoA reductase in the protozoan Trypanosoma cruzi

    DEFF Research Database (Denmark)

    Pena Diaz, Javier; Montalvetti, A; Camacho, A;

    1997-01-01

    We report the isolation and characterization of a genomic clone containing the open reading frame sequence for 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase from Trypanosoma cruzi, the causative agent of Chagas' disease. The protozoan gene encoded for a smaller polypeptide than the rest...... sensitive to proteolytic inactivation. Furthermore the enzyme can be efficiently overexpressed in a highly active form by using the expression vector pET-11c. Thus Trypanosoma cruzi HMG-CoA reductase is unique in the sense that it totally lacks the membrane-spanning sequences present in all eukaryotic HMG...... cellular distribution of enzymic activity was investigated after differential centrifugation of Trypanosoma cell extracts. Reductase activity was primarily associated with the cellular soluble fraction because 95% of the total cellular activity was recovered in the supernatant and was particularly...

  15. Host stress physiology and Trypanosoma haemoparasite infection influence innate immunity in the woylie (Bettongia penicillata).

    Science.gov (United States)

    Hing, Stephanie; Currie, Andrew; Broomfield, Steven; Keatley, Sarah; Jones, Krista; Thompson, R C Andrew; Narayan, Edward; Godfrey, Stephanie S

    2016-06-01

    Understanding immune function is critical to conserving wildlife in view of infectious disease threats, particularly in threatened species vulnerable to stress, immunocompromise and infection. However, few studies examine stress, immune function and infection in wildlife. We used a flow cytometry protocol developed for human infants to assess phagocytosis, a key component of innate immunity, in a critically endangered marsupial, the woylie (Bettongia penicillata). The effects of stress physiology and Trypanosoma infection on phagocytosis were investigated. Blood and faecal samples were collected from woylies in a captive facility over three months. Trypanosoma status was determined using PCR. Faecal cortisol metabolites (FCM) were quantified by enzyme-immunoassay. Mean phagocytosis measured was >90%. An interaction between sex and FCM influenced the percentage of phagocytosing leukocytes, possibly reflecting the influence of sex hormones and glucocorticoids. An interaction between Trypanosoma status and FCM influenced phagocytosis index, suggesting that stress physiology and infection status influence innate immunity.

  16. Evaluation of Real-time PCR and Pyrosequencing for Screening Incubating Blood Culture Bottles from Adults with Suspected Bloodstream Infection

    Science.gov (United States)

    McCann, Chase D.; Moore, Miranda S.; May, Larissa S.; McCarroll, Matthew; Jordan, Jeanne A.

    2015-01-01

    Several molecular platforms can identify bacteria associated with bloodstream infections, but require positive culture bottles as starting material. Here we describe results of screening 1140 blood cultures at 8 hours post-inoculation, from 918 eligible adults being evaluated for bloodstream infection. DNA was extracted and analyzed by 16S and/or 23S rRNA real-time PCR/Pyrosequencing. Compared to culture, PCR/Pyrosequencing displayed 90.9% sensitivity, 99.6% specificity, 95.7% PPV, and 99.1% NPV. Overall concordance rate was 98.9% (1127/1140). In four cases with molecular-positive/culture-negative results, medical chart reviews provided evidence of identical bacteria from subsequent blood or concomitant urine/sputum cultures. Nine culture-positive/molecular-negative cases were associated with either polymicrobial growth, grew only in the anaerobic bottle of the clinical pair, and/or were detected by PCR/Pyrosequencing after 8 hours. In summary, this approach accurately detected and identified bacteria in ~91% of culture-confirmed cases significantly sooner than the phenotypic identification was available, having the potential to improve antibiotic stewardship. PMID:25534615

  17. Evaluation of real-time PCR and pyrosequencing for screening incubating blood culture bottles from adults with suspected bloodstream infection.

    Science.gov (United States)

    McCann, Chase D; Moore, Miranda S; May, Larissa S; McCarroll, Matthew G; Jordan, Jeanne A

    2015-03-01

    Several molecular platforms can identify bacteria associated with bloodstream infections but require positive culture bottles as starting material. Here, we describe results of screening 1140 blood cultures at 8h postinoculation, from 918 eligible adults being evaluated for bloodstream infection. DNA was extracted and analyzed by 16S and/or 23S rRNA real-time PCR/pyrosequencing. Compared to culture, PCR/pyrosequencing displayed 90.9% sensitivity, 99.6% specificity, 95.7% positive predictive value, and 99.1% negative predictive value. Overall concordance rate was 98.9% (1127/1140). In 4 cases with molecular-positive/culture-negative results, medical chart reviews provided evidence of identical bacteria from subsequent blood or concomitant urine/sputum cultures. Nine culture-positive/molecular-negative cases were associated with either polymicrobial growth, grew only in the anaerobic bottle of the clinical pair, and/or were detected by PCR/pyrosequencing after 8h. In summary, this approach accurately detected and identified bacteria in ~91% of culture-confirmed cases significantly sooner than the phenotypic identification was available, having the potential to improve antibiotic stewardship.

  18. Clonal distribution of bone sialoprotein-binding protein gene among Staphylococcus aureus isolates associated with bloodstream infections.

    Science.gov (United States)

    Wiśniewska, Katarzyna; Piórkowska, Anna; Kasprzyk, Joanna; Bronk, Marek; Świeć, Krystyna

    2014-11-01

    Staphylococcus aureus is a leading cause of bloodstream infections (BSI) and diseases that may be caused by hematogenous spread. The staphylococcal adhesin, for which the association with the infections emerging as a complication of septicemia has been well documented, is a bone sialoprotein-binding protein (Bbp). The aim of the study was to assess the prevalence of a bbp gene in S. aureus bloodstream isolates associated with BSI and to investigate to what degree the distribution of this gene is linked to the clonality of the population. Spa typing, used in order to explore the genetic population structure of the isolates, yielded 29 types. Six spa clusters and seven singletons were identified. The most frequent was spa clonal complex CC021 associated with MLST CC30 (38%). The bbp gene was found in 47% of isolates. Almost all isolates (95%) clustered in spa clonal complex CC021 were positive for this gene. All isolates carrying the bbp gene were sensitive to methicillin, and if clustered in the spa CC021, belonged to agr group III. Our study shows that Bbp is not strictly associated with BSI. However, one may conclude that for clonally related S. aureus strains most commonly causing BSI, the risk of Bbp-mediated complications of septicemia is expected to be higher than for other strains.

  19. Host Characteristics and Bacterial Traits Predict Experimental Virulence for Escherichia coli Bloodstream Isolates From Patients With Urosepsis.

    Science.gov (United States)

    Johnson, James R; Porter, Stephen; Johnston, Brian; Kuskowski, Michael A; Spurbeck, Rachel R; Mobley, Harry L T; Williamson, Deborah A

    2015-09-01

    Background.  Extraintestinal Escherichia coli infections are common, costly, and potentially serious. A better understanding of their pathogenesis is needed. Methods.  Sixty-seven E coli bloodstream isolates from adults with urosepsis (Seattle, WA; 1980s) underwent extensive molecular characterization and virulence assessment in 2 infection models (murine subcutaneous sepsis and moth larval lethality). Statistical comparisons were made among host characteristics, bacterial traits, and experimental virulence. Results.  The 67 source patients were diverse for age, sex, and underlying medical and urological conditions. The corresponding E coli isolates exhibited diverse phylogenetic backgrounds and virulence profiles. Despite the E coli isolates' common bloodstream origin, they exhibited a broad range of experimental virulence in mice and moth larvae, in patterns that (for the murine model only) corresponded significantly with host characteristics and bacterial traits. The most highly mouse-lethal strains were enriched with classic "urovirulence" traits and typically were from younger women with anatomically and functionally normal urinary tracts. The 2 animal models corresponded poorly with one another. Conclusions.  Host compromise, including older age and urinary tract abnormalities, allows comparatively low-virulence E coli strains to cause urosepsis. Multiple E coli traits predict both experimental and epidemiological virulence. The larval lethality model cannot be a substitute for the murine sepsis model.

  20. Clonal population structure of Colombian sylvatic Trypanosoma cruzi.

    Science.gov (United States)

    Márquez, E; Arcos-Burgos, M; Triana, O; Moreno, J; Jaramillo, N

    1998-12-01

    Isoenzyme variability and evidence of genetic exchange were evaluated in 75 wild stocks of Trypanosoma cruzi obtained from different hosts from 5 geographical regions within the endemic area in Colombia. Cluster analysis of genetic variability was attempted. Thirty-three multilocus enzyme genotypes (clonets) were identified from 75 stocks, 27 of which clustered with zymodeme Z1 and 6 with zymodeme Z3. Two stocks isolated from human infections showed the potential risk to rural communities in Colombia. The stocks exhibited departures from Hardy-Weinberg expectations, including both fixed heterozygote and fixed homozygote demes, where both segregation and recombination were absent. To inspect for population subdivision that might falsely imply clonality in these stocks, Wright's F statistics were calculated. Theta values (Fst) were significantly different from 0 when 33 clonets, 27 Z1-like clonets, and 5 geographical subpopulations were compared; thus, a significant amount of divergence has occurred between and within them. In addition, linkage disequilibrium was detected for most possible pairwise comparisons of loci. In conclusion, the above results all support a scenario of long-term clonal evolution in Colombian sylvatic T. cruzi populations.

  1. Genetically attenuated Trypanosoma cruzi parasites as a potential vaccination tool.

    Science.gov (United States)

    Pérez Brandan, Cecilia; Basombrío, Miguel Ángel

    2012-01-01

    Chagas disease is the clinical manifestation of the infection produced by the parasite Trypanosoma cruzi. Currently there is no vaccine to prevent this disease and the protection attained with vaccines containing non-replicating parasites is limited. Genetically attenuated trypanosomatid parasites can be obtained by deletion of selected genes. Gene deletion takes advantage of the fact that this parasite can undergo homologous recombination between endogenous and foreign DNA sequences artificially introduced in the cells. This approach facilitated the discovery of several unknown gene functions, as well as allowing us to speculate about the potential for genetically attenuated live organisms as experimental immunogens. Vaccination with live attenuated parasites has been used effectively in mice to reduce parasitemia and histological damage, and in dogs, to prevent vector-delivered infection in the field. However, the use of live parasites as immunogens is controversial due to the risk of reversion to a virulent phenotype. Herein, we present our results from experiments on genetic manipulation of two T. cruzi strains to produce parasites with impaired replication and infectivity, and using the mutation of the dhfr-ts gene as a safety device against reversion to virulence.

  2. MIF Contributes to Trypanosoma brucei Associated Immunopathogenicity Development

    Science.gov (United States)

    Stijlemans, Benoît; Leng, Lin; Brys, Lea; Sparkes, Amanda; Vansintjan, Liese; Caljon, Guy; Raes, Geert; Van Den Abbeele, Jan; Van Ginderachter, Jo A.; Beschin, Alain

    2014-01-01

    African trypanosomiasis is a chronic debilitating disease affecting the health and economic well-being of many people in developing countries. The pathogenicity associated with this disease involves a persistent inflammatory response, whereby M1-type myeloid cells, including Ly6Chigh inflammatory monocytes, are centrally implicated. A comparative gene analysis between trypanosusceptible and trypanotolerant animals identified MIF (macrophage migrating inhibitory factor) as an important pathogenic candidate molecule. Using MIF-deficient mice and anti-MIF antibody treated mice, we show that MIF mediates the pathogenic inflammatory immune response and increases the recruitment of inflammatory monocytes and neutrophils to contribute to liver injury in Trypanosoma brucei infected mice. Moreover, neutrophil-derived MIF contributed more significantly than monocyte-derived MIF to increased pathogenic liver TNF production and liver injury during trypanosome infection. MIF deficient animals also featured limited anemia, coinciding with increased iron bio-availability, improved erythropoiesis and reduced RBC clearance during the chronic phase of infection. Our data suggest that MIF promotes the most prominent pathological features of experimental trypanosome infections (i.e. anemia and liver injury), and prompt considering MIF as a novel target for treatment of trypanosomiasis-associated immunopathogenicity. PMID:25255103

  3. MIF contributes to Trypanosoma brucei associated immunopathogenicity development.

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    Benoît Stijlemans

    2014-09-01

    Full Text Available African trypanosomiasis is a chronic debilitating disease affecting the health and economic well-being of many people in developing countries. The pathogenicity associated with this disease involves a persistent inflammatory response, whereby M1-type myeloid cells, including Ly6C(high inflammatory monocytes, are centrally implicated. A comparative gene analysis between trypanosusceptible and trypanotolerant animals identified MIF (macrophage migrating inhibitory factor as an important pathogenic candidate molecule. Using MIF-deficient mice and anti-MIF antibody treated mice, we show that MIF mediates the pathogenic inflammatory immune response and increases the recruitment of inflammatory monocytes and neutrophils to contribute to liver injury in Trypanosoma brucei infected mice. Moreover, neutrophil-derived MIF contributed more significantly than monocyte-derived MIF to increased pathogenic liver TNF production and liver injury during trypanosome infection. MIF deficient animals also featured limited anemia, coinciding with increased iron bio-availability, improved erythropoiesis and reduced RBC clearance during the chronic phase of infection. Our data suggest that MIF promotes the most prominent pathological features of experimental trypanosome infections (i.e. anemia and liver injury, and prompt considering MIF as a novel target for treatment of trypanosomiasis-associated immunopathogenicity.

  4. Phenolic Constituents of Medicinal Plants with Activity against Trypanosoma brucei

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    Ya Nan Sun

    2016-04-01

    Full Text Available Neglected tropical diseases (NTDs affect over one billion people all over the world. These diseases are classified as neglected because they impact populations in areas with poor financial conditions and hence do not attract sufficient research investment. Human African Trypanosomiasis (HAT or sleeping sickness, caused by the parasite Trypanosoma brucei, is one of the NTDs. The current therapeutic interventions for T. brucei infections often have toxic side effects or require hospitalization so that they are not available in the rural environments where HAT occurs. Furthermore, parasite resistance is increasing, so that there is an urgent need to identify novel lead compounds against this infection. Recognizing the wide structural diversity of natural products, we desired to explore and identify novel antitrypanosomal chemotypes from a collection of natural products obtained from plants. In this study, 440 pure compounds from various medicinal plants were tested against T. brucei by in a screening using whole cell in vitro assays. As the result, twenty-two phenolic compounds exhibited potent activity against cultures of T. brucei. Among them, eight compounds—4, 7, 11, 14, 15, 18, 20, and 21—showed inhibitory activity against T. brucei, with IC50 values below 5 µM, ranging from 0.52 to 4.70 μM. Based on these results, we attempt to establish some general trends with respect to structure-activity relationships, which indicate that further investigation and optimization of these derivatives might enable the preparation of potentially useful compounds for treating HAT.

  5. Heterogeneous infectiousness in guinea pigs experimentally infected with Trypanosoma cruzi.

    Science.gov (United States)

    Castillo-Neyra, Ricardo; Borrini Mayorí, Katty; Salazar Sánchez, Renzo; Ancca Suarez, Jenny; Xie, Sherrie; Náquira Velarde, Cesar; Levy, Michael Z

    2016-02-01

    Guinea pigs are important reservoirs of Trypanosoma cruzi, the causative parasite of Chagas disease, and in the Southern Cone of South America, transmission is mediated mainly by the vector Triatoma infestans. Interestingly, colonies of Triatoma infestans captured from guinea pig corrals sporadically have infection prevalence rates above 80%. Such high values are not consistent with the relatively short 7-8 week parasitemic period that has been reported for guinea pigs in the literature. We experimentally measured the infectious periods of a group of T. cruzi-infected guinea pigs by performing xenodiagnosis and direct microscopy each week for one year. Another group of infected guinea pigs received only direct microscopy to control for the effect that inoculation by triatomine saliva may have on parasitemia in the host. We observed infectious periods longer than those previously reported in a number of guinea pigs from both the xenodiagnosis and control groups. While some guinea pigs were infectious for a short time, other "super-shedders" were parasitemic up to 22 weeks after infection, and/or positive by xenodiagnosis for a year after infection. This heterogeneity in infectiousness has strong implications for T. cruzi transmission dynamics and control, as super-shedder guinea pigs may play a disproportionate role in pathogen spread.

  6. Early Trypanosoma cruzi Infection Reprograms Human Epithelial Cells

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    María Laura Chiribao

    2014-01-01

    Full Text Available Trypanosoma cruzi, the causative agent of Chagas disease, has the peculiarity, when compared with other intracellular parasites, that it is able to invade almost any type of cell. This property makes Chagas a complex parasitic disease in terms of prophylaxis and therapeutics. The identification of key host cellular factors that play a role in the T. cruzi invasion is important for the understanding of disease pathogenesis. In Chagas disease, most of the focus is on the response of macrophages and cardiomyocytes, since they are responsible for host defenses and cardiac lesions, respectively. In the present work, we studied the early response to infection of T. cruzi in human epithelial cells, which constitute the first barrier for establishment of infection. These studies identified up to 1700 significantly altered genes regulated by the immediate infection. The global analysis indicates that cells are literally reprogrammed by T. cruzi, which affects cellular stress responses (neutrophil chemotaxis, DNA damage response, a great number of transcription factors (including the majority of NFκB family members, and host metabolism (cholesterol, fatty acids, and phospholipids. These results raise the possibility that early host cell reprogramming is exploited by the parasite to establish the initial infection and posterior systemic dissemination.

  7. Dehydroepiandrosterone increases resistance to experimental infection by Trypanosoma cruzi.

    Science.gov (United States)

    Santos, Carla Domingues; Toldo, Míriam Paula Alonso; Santello, Fabrícia Helena; Filipin, Marina Del Vecchio; Brazão, Vânia; do Prado Júnior, José Clóvis

    2008-05-31

    Dehydroepiandrosterone (DHEA) enhances immune responses against a wide range of viral, bacterial, and parasitic pathogens. In a previous study, we reported that administration of DHEA significantly decreased the numbers of blood parasites in Trypanosoma cruzi experimental infection. The present study was undertaken to determine the effectiveness of DHEA in reducing the severity of acute phase T. cruzi infection of male and female Wistar rats. Animals were treated subcutaneously with 40 mg/kg body weight/day of DHEA. The concentration of nitric oxide (NO) was determined in spleen peritoneal cavity. Interleukin-2 (IL-2) and interferon-gamma (IFN-gamma) were determined in the sera of uninfected and infected animals. DHEA treatment augments NO production for both sexes after in vitro LPS treatment for uninfected animals. Infection triggered enhanced NO levels although not significant. IL-2 and IFN-gamma were detectable in higher concentrations in treated and infected rats of both genders when compared to untreated controls. These data suggest that DHEA may have a potent immunoregulatory function that can affect the course of T. cruzi infection.

  8. Exosome secretion affects social motility in Trypanosoma brucei

    Science.gov (United States)

    Shaked, Hadassa; Arvatz, Gil; Tkacz, Itai Dov; Binder, Lior; Waldman Ben-Asher, Hiba; Okalang, Uthman; Chikne, Vaibhav; Cohen-Chalamish, Smadar; Michaeli, Shulamit

    2017-01-01

    Extracellular vesicles (EV) secreted by pathogens function in a variety of biological processes. Here, we demonstrate that in the protozoan parasite Trypanosoma brucei, exosome secretion is induced by stress that affects trans-splicing. Following perturbations in biogenesis of spliced leader RNA, which donates its spliced leader (SL) exon to all mRNAs, or after heat-shock, the SL RNA is exported to the cytoplasm and forms distinct granules, which are then secreted by exosomes. The exosomes are formed in multivesicular bodies (MVB) utilizing the endosomal sorting complexes required for transport (ESCRT), through a mechanism similar to microRNA secretion in mammalian cells. Silencing of the ESCRT factor, Vps36, compromised exosome secretion but not the secretion of vesicles derived from nanotubes. The exosomes enter recipient trypanosome cells. Time-lapse microscopy demonstrated that cells secreting exosomes or purified intact exosomes affect social motility (SoMo). This study demonstrates that exosomes are delivered to trypanosome cells and can change their migration. Exosomes are used to transmit stress signals for communication between parasites. PMID:28257521

  9. Identification of novel cyclic nucleotide binding proteins in Trypanosoma cruzi.

    Science.gov (United States)

    Jäger, Adriana V; De Gaudenzi, Javier G; Mild, Jesica G; Mc Cormack, Bárbara; Pantano, Sergio; Altschuler, Daniel L; Edreira, Martin M

    2014-12-01

    Cyclic AMP has been implicated as second messenger in a wide range of cellular processes. In the protozoan parasite Trypanosoma cruzi, cAMP is involved in the development of the parasite's life cycle. While cAMP effectors have been widely studied in other eukaryotic cells, little is known about cAMP's mechanism of action in T. cruzi. To date, only a cAMP-dependent protein kinase A (PKA) has been cloned and characterised in this parasite; however experimental evidence indicates the existence of cAMP-dependent, PKA-independent events. In order to identify new cAMP binding proteins as potential cAMP effectors, we carried out in silico studies using the predicted T. cruzi proteome. Using a combination of search methods 27 proteins with putative cNMP binding domains (CBDs) were identified. Phylogenetic analysis of the CBDs presented a homogeneous distribution, with sequences segregated into two main branches: one containing kinases-like proteins and the other gathering hypothetical proteins with different function or no other known. Comparative modelling of the strongest candidates provides support for the hypothesis that these proteins may give rise to structurally viable cyclic nucleotide binding domains. Pull-down and nucleotide displacement assays strongly suggest that TcCLB.508523.80 could bind cAMP and eventually be a new putative PKA-independent cAMP effector in T. cruzi.

  10. Immunity and immune modulation in Trypanosoma cruzi infection.

    Science.gov (United States)

    Cardillo, Fabíola; de Pinho, Rosa Teixeira; Antas, Paulo Renato Zuquim; Mengel, José

    2015-12-01

    Chagas disease is caused by the protozoan Trypanosoma cruzi. The parasite reaches the secondary lymphoid organs, the heart, skeletal muscles, neurons in the intestine and esophagus among other tissues. The disease is characterized by mega syndromes, which may affect the esophagus, the colon and the heart, in about 30% of infected people. The clinical manifestations associated with T. cruzi infection during the chronic phase of the disease are dependent on complex interactions between the parasite and the host tissues, particularly the lymphoid system that may either result in a balanced relationship with no disease or in an unbalanced relationship that follows an inflammatory response to parasite antigens and associated tissues in some of the host organs and/or by an autoimmune response to host antigens. This review discusses the findings that support the notion of an integrated immune response, considering the innate and adaptive arms of the immune system in the control of parasite numbers and also the mechanisms proposed to regulate the immune response in order to tolerate the remaining parasite load, during the chronic phase of infection. This knowledge is fundamental to the understanding of the disease progression and is essential for the development of novel therapies and vaccine strategies.

  11. Structural characterization of NETNES glycopeptide from Trypanosoma cruzi.

    Science.gov (United States)

    Chiodi, Carla G; Verli, Hugo

    2013-05-24

    Trypanosoma cruzi is a protozoan, responsible for Chagas disease, that parasites triatomines and some vertebrates, mainly Homo sapiens. In 2010, nearly 10 million people in whole world, most from Latin America, had Chagas disease, which is an illness of high morbidity, low mortality, and serious problems of quality of life. The available treatment has high toxicity and low efficacy at chronic phase. Some of the protozoan antigenic or virulence factors include complex carbohydrate structures that, due to their uniqueness, may constitute potential selective targets for the development of new treatments. One example of such structures is NETNES, a low abundance T. cruzi glycopeptide, comprising 13 amino acid residues, one or two N-glycosylation chains, a GPI anchor and two P-glycosylations. In this context, the current work aims to obtain an atomic model for NETNES, including its glycan chains and membrane attachment, in order to contribute in the characterization of its structure and dynamics. Based on POPC and GPI models built in agreement with experimental data, our results indicate that, in the first third of the simulation, NETNES peptide is very flexible in solution, bending itself between asparagine residues and lying down on some carbohydrates and membrane, exposing amino acid residues and some other glycans, mainly terminal mannoses, to the extracellular medium, remaining in this position until the end of simulations.

  12. Experimental infections in Venezuelan lizards by Trypanosoma cruzi.

    Science.gov (United States)

    Urdaneta-Morales, S; McLure, I

    1981-06-01

    Virulent trypomastigotes of the Y strain of Trypanosoma cruzi were administered to Tropidurus hispidus, Ameiva ameiva, Cnemidophorus lemniscatus, Polychrus marmoratus, and Phyllodactylus ventralis (Sauria). Intraperitoneal and subcutaneous inoculations of lizards with mouse blood or with feces of infected Rhodnius prolixus (Reduviidae, Triatominae), as well as forced ingestion of triturated Rhodnius, produced no parasitaemias detectable either directly or by xenodiagnosis, while control mice became parasitized. Pretreatment with the immunosuppressive drug Fluocinolone acetonide led to establishing patent infections in inoculated lizards. Cryptic infections were established by inoculation of 1 X 10(6) parasites from Davis' medium, or by 95 X 10(3) parasites from lizard tissue culture. Parasites were not seen in tissues. Mice inoculated with blood or tissue homogenates from these lizards became parasitized. Parasites from Davis' medium inoculated into the peritoneal cavity of lizards were capable, to a very low degree, of penetrating the free peritoneal macrophages and changing into amastigotes. The factors possibly responsible for the natural resistance of poikilothermic vertebrates to T. cruzi are discussed.

  13. Proteomic selection of immunodiagnostic antigens for Trypanosoma congolense.

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    Jennifer R Fleming

    2014-06-01

    Full Text Available Animal African Trypanosomosis (AAT presents a severe problem for agricultural development in sub-Saharan Africa. It is caused by several trypanosome species and current means of diagnosis are expensive and impractical for field use. Our aim was to discover antigens for the detection of antibodies to Trypanosoma congolense, one of the main causative agents of AAT. We took a proteomic approach to identify potential immunodiagnostic parasite protein antigens. One hundred and thirteen proteins were identified which were selectively recognized by infected cattle sera. These were assessed for likelihood of recombinant protein expression in E. coli and fifteen were successfully expressed and assessed for their immunodiagnostic potential by ELISA using pooled pre- and post-infection cattle sera. Three proteins, members of the invariant surface glycoprotein (ISG family, performed favorably and were then assessed using individual cattle sera. One antigen, Tc38630, evaluated blind with 77 randomized cattle sera in an ELISA assay gave sensitivity and specificity performances of 87.2% and 97.4%, respectively. Cattle immunoreactivity to this antigen diminished significantly following drug-cure, a feature helpful for monitoring the efficacy of drug treatment.

  14. In vitro effects of citral on Trypanosoma cruzi metacyclogenesis

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    Josiane Cardoso

    2010-12-01

    Full Text Available Citral, the main constituent of lemongrass (Cymbopogon citratus essential oil, was added to Trypanosoma cruzi cultures grown in TAU3AAG medium to observe the effect on the epimastigote-to-trypomastigote differentiation process (metacyclogenesis. Our results showed that citral (20 μg/mL did not affect epimastigote viability or inhibit the differentiation process. Concentrations higher than 60 μg/mL, however, led to 100% cell death (both epimastigote and trypomastigote forms. Although epimastigotes incubated with 30 μg/mL citral were viable and able to adhere to the substrate, we observed around 50% inhibition in metacyclogenesis, with a calculated concentration that inhibited metacyclogenesis by 50% after 24 h (IC50/24 h of about 31 μg/mL. Treatment with 30 μg/mL citral did not hinder epimastigote multiplication because epimastigote growth resumed when treated cells were transferred to a drug-free liver infusion tryptose culture medium. Metacyclogenesis was almost totally abolished at 40 μg/mL after 24 h of incubation. Furthermore, the metacyclic trypomastigotes obtained in vitro were similarly susceptible to citral, with an IC50/24 h, concentration that killed 50% of the cells after 24 h, of about 24.5 μg/mL. Therefore, citral appears to be a good candidate as an inhibitory drug for further studies analyzing the T. cruzi metacyclogenesis process.

  15. Chronic experimental infection by Trypanosoma cruzi in Cebus apella monkeys

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    A. Riarte

    1995-12-01

    Full Text Available Twenty young male Cebus apella monkeys were infected with CAl Trypanosoma cruzi strain and reinfected with CA l or Tulahuen T.cruzi strains, with different doses and parasite source. Subpatent parasitemia was usually demonstrated in acute and chronic phases. Patent parasitemia was evident in one monkey in the acute phase and in four of them in the chronic phase after re-inoculations with high doses of CAl strain. Serological conversion was observed in all monkeys; titers were low, regardless of the methods used to investigate anti-T. cruzi specific antibodies. Higher titers were induced only when re-inoculations were perfomed with the virulent Tulahuén strain or high doses of CAl strain. Clinical electrocardiographic and ajmaline test evaluations did not reveal changes between infected and control monkeys. Histopathologically, cardiac lesions were always characterized by focal or multifocal mononuclear infiltrates and/or isolated fibrosis, as seen during the acute and chronic phases; neither amastigote nests nor active inflammation and fibrogenic processes characteristic of human acute and chronic myocarditis respectively, were observed. These morphological aspects more closely resemble those found in the "indeterminate phase" and contrast with the more diffuse and progressive pattern of the human chagasic myocarditis. All monkeys survived and no mortality was observed.

  16. Proteomics and the Trypanosoma brucei cytoskeleton: advances and opportunities.

    Science.gov (United States)

    Portman, Neil; Gull, Keith

    2012-08-01

    Trypanosoma brucei is the etiological agent of devastating parasitic disease in humans and livestock in sub-saharan Africa. The pathogenicity and growth of the parasite are intimately linked to its shape and form. This is in turn derived from a highly ordered microtubule cytoskeleton that forms a tightly arrayed cage directly beneath the pellicular membrane and numerous other cytoskeletal structures such as the flagellum. The parasite undergoes extreme changes in cellular morphology during its life cycle and cell cycles which require a high level of integration and coordination of cytoskeletal processes. In this review we will discuss the role that proteomics techniques have had in advancing our understanding of the molecular composition of the cytoskeleton and its functions. We then consider future opportunities for the application of these techniques in terms of addressing some of the unanswered questions of trypanosome cytoskeletal cell biology with particular focus on the differences in the composition and organisation of the cytoskeleton through the trypanosome life-cycle.

  17. Crystal Structure of Triosephosphate Isomerase from Trypanosoma cruzi in Hexane

    Science.gov (United States)

    Gao, Xiu-Gong; Maldonado, Ernesto; Perez-Montfort, Ruy; Garza-Ramos, Georgina; Tuena de Gomez-Puyou, Marietta; Gomez-Puyou, Armando; Rodriguez-Romero, Adela

    1999-08-01

    To gain insight into the mechanisms of enzyme catalysis in organic solvents, the x-ray structure of some monomeric enzymes in organic solvents was determined. However, it remained to be explored whether the structure of oligomeric proteins is also amenable to such analysis. The field acquired new perspectives when it was proposed that the x-ray structure of enzymes in nonaqueous media could reveal binding sites for organic solvents that in principle could represent the starting point for drug design. Here, a crystal of the dimeric enzyme triosephosphate isomerase from the pathogenic parasite Trypanosoma cruzi was soaked and diffracted in hexane and its structure solved at 2- angstrom resolution. Its overall structure and the dimer interface were not altered by hexane. However, there were differences in the orientation of the side chains of several amino acids, including that of the catalytic Glu-168 in one of the monomers. No hexane molecules were detected in the active site or in the dimer interface. However, three hexane molecules were identified on the surface of the protein at sites, which in the native crystal did not have water molecules. The number of water molecules in the hexane structure was higher than in the native crystal. Two hexanes localized at <4 angstrom from residues that form the dimer interface; they were in close proximity to a site that has been considered a potential target for drug design.

  18. Trypanosoma rangeli protein tyrosine phosphatase is associated with the parasite's flagellum

    OpenAIRE

    Elisa Beatriz Prestes; Ethel Bayer-Santos; Patrícia Hermes Stoco; Thaís Cristine Marques Sincero; Glauber Wagner; Adriana Umaki; Stenio Perdigão Fragoso; Juliano Bordignon; Mário Steindel; Edmundo Carlos Grisard

    2012-01-01

    Protein tyrosine phosphatases (PTPs) play an essential role in the regulation of cell differentiation in pathogenic trypanosomatids. In this study, we describe a PTP expressed by the non-pathogenic protozoan Trypanosoma rangeli (TrPTP2). The gene for this PTP is orthologous to the T. brucei TbPTP1 and Trypanosoma cruzi (TcPTP2) genes. Cloning and expression of the TrPTP2 and TcPTP2 proteins allowed anti-PTP2 monoclonal antibodies to be generated in BALB/c mice. When expressed by T. rangeli ep...

  19. The pentose phosphate pathway in Trypanosoma cruzi: a potential target for the chemotherapy of Chagas disease

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    Mariana Igoillo-Esteve

    2007-12-01

    Full Text Available Trypanosoma cruzi is highly sensitive to oxidative stress caused by reactive oxygen species. Trypanothione, the parasite's major protection against oxidative stress, is kept reduced by trypanothione reductase, using NADPH; the major source of the reduced coenzyme seems to be the pentose phosphate pathway. Its seven enzymes are present in the four major stages in the parasite's biological cycle; we have cloned and expressed them in Escherichia coli as active proteins. Glucose 6-phosphate dehydrogenase, which controls glucose flux through the pathway by its response to the NADP/NADPH ratio, is encoded by a number of genes per haploid genome, and is induced up to 46-fold by hydrogen peroxide in metacyclic trypomastigotes. The genes encoding 6-phosphogluconolactonase, 6-phosphogluconate dehydrogenase, transaldolase and transketolase are present in the CL Brener clone as a single copy per haploid genome. 6-phosphogluconate dehydrogenase is very unstable, but was stabilized introducing two salt bridges by site-directed mutagenesis. Ribose-5-phosphate isomerase belongs to Type B; genes encoding Type A enzymes, present in mammals, are absent. Ribulose-5-phosphate epimerase is encoded by two genes. The enzymes of the pathway have a major cytosolic component, although several of them have a secondary glycosomal localization, and also minor localizations in other organelles.Trypanosoma cruzi é altamente sensível ao estresse oxidativo causado por espécies reativas do oxigênio. Tripanotiona, o principal protetor do parasita contra o estresse oxidativo, é mantido reduzido pela tripanotiona redutase, pela presença deNADPH; a principal fonte da coenzima reduzida parece ser a via da pentose fosfato. As sete enzimas dessa via estão presentes nos quatro principais estágios do ciclo biológico do parasita; nós clonamos e expressamos as enzimas em Escherichia coli como proteínas ativas. Glucose 6-fosfato desidrogenase, que controla o fluxo da glucose da

  20. The use of yellow fluorescent hybrids to indicate mating in Trypanosoma brucei

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    Ferris Vanessa

    2008-02-01

    Full Text Available Abstract Background Trypanosoma brucei undergoes genetic exchange in its insect vector, the tsetse fly, by an unknown mechanism. The difficulties of working with this experimental system of genetic exchange have hampered investigation, particularly because the trypanosome life cycle stages involved cannot be cultured in vitro and therefore must be examined in the insect. Searching for small numbers of hybrid trypanosomes directly in the fly has become possible through the incorporation of fluorescent reporter genes, and we have previously carried out a successful cross using a reporter-repressor strategy. However, we could not be certain that all fluorescent trypanosomes observed in that cross were hybrids, due to mutations of the repressor leading to spontaneous fluorescence, and we have therefore developed an alternative strategy. Results To visualize the production of hybrids in the fly, parental trypanosome clones were transfected with a gene encoding Green Fluorescent Protein (GFP or Red Fluorescent Protein (RFP. Co-infection of flies with red and green fluorescent parental trypanosomes produced yellow fluorescent hybrids, which were easily visualized in the fly salivary glands. Yellow trypanosomes were not seen in midgut or proventricular samples and first appeared in the glands as epimastigotes as early as 13 days after fly infection. Cloned progeny originating from individual salivary glands had yellow, red, green or no fluorescence and were confirmed as hybrids by microsatellite, molecular karyotype and kinetoplast (mitochondrial DNA analyses. Hybrid clones showed biparental inheritance of both nuclear and kinetoplast genomes. While segregation and reassortment of the reporter genes and microsatellite alleles were consistent with Mendelian inheritance, flow cytometry measurement of DNA content revealed both diploid and polyploid trypanosomes among the hybrid progeny clones. Conclusion The strategy of using production of yellow hybrids

  1. Pentamidine exerts in vitro and in vivo anti Trypanosoma cruzi activity and inhibits the polyamine transport in Trypanosoma cruzi.

    Science.gov (United States)

    Díaz, María V; Miranda, Mariana R; Campos-Estrada, Carolina; Reigada, Chantal; Maya, Juan D; Pereira, Claudio A; López-Muñoz, Rodrigo

    2014-06-01

    Pentamidine is an antiprotozoal and fungicide drug used in the treatment of leishmaniasis and African trypanosomiasis. Despite its extensive use as antiparasitic drug, little evidence exists about the effect of pentamidine in Trypanosoma cruzi, the etiological agent of Chagas' disease. Recent studies have shown that pentamidine blocks a polyamine transporter present in Leishmania major; consequently, its might also block these transporters in T. cruzi. Considering that T. cruzi lacks the ability to synthesize putrescine de novo, the inhibition of polyamine transport can bring a new therapeutic target against the parasite. In this work, we show that pentamidine decreases, not only the viability of T. cruzi trypomastigotes, but also the parasite burden of infected cells. In T. cruzi-infected mice pentamidine decreases the inflammation and parasite burden in hearts from infected mice. The treatment also decreases parasitemia, resulting in an increased survival rate. In addition, pentamidine strongly inhibits the putrescine and spermidine transport in T. cruzi epimastigotes and amastigotes. Thus, this study points to reevaluate the utility of pentamidine and introduce evidence of a potential new action mechanism. In the quest of new therapeutic strategies against Chagas disease, the extensive use of pentamidine in human has led to a well-known clinical profile, which could be an advantage over newly synthesized molecules that require more comprehensive trials prior to their clinical use.

  2. Eleganolone, a Diterpene from the French Marine Alga Bifurcaria bifurcata Inhibits Growth of the Human Pathogens Trypanosoma brucei and Plasmodium falciparum

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    Anne-Marie Rusig

    2013-02-01

    Full Text Available Organic extracts of 20 species of French seaweed have been screened against Trypanosoma brucei rhodesiense trypomastigotes, the parasite responsible for sleeping sickness. These extracts have previously shown potent antiprotozoal activities in vitro against Plasmodium falciparum and Leishmania donovani. The selectivity of the extracts was also evaluated by testing cytotoxicity on a mammalian L6 cell line. The ethyl acetate extract of the brown seaweed, Bifurcaria bifurcata, showed strong trypanocidal activity with a mild selectivity index (IC50 = 0.53 µg/mL; selectivity index (SI = 11.6. Bio-guided fractionation led to the isolation of eleganolone, the main diterpenoid isolated from this species. Eleganolone contributes only mildly to the trypanocidal activity of the ethyl acetate extract (IC50 = 45.0 µM, SI = 4.0. However, a selective activity against P. falciparum erythrocytic stages in vitro has been highlighted (IC50 = 7.9 µM, SI = 21.6.

  3. Eleganolone, a Diterpene from the French Marine Alga Bifurcaria bifurcata Inhibits Growth of the Human Pathogens Trypanosoma brucei and Plasmodium falciparum

    Science.gov (United States)

    Gallé, Jean-Baptiste; Attioua, Barthélémy; Kaiser, Marcel; Rusig, Anne-Marie; Lobstein, Annelise; Vonthron-Sénécheau, Catherine

    2013-01-01

    Organic extracts of 20 species of French seaweed have been screened against Trypanosoma brucei rhodesiense trypomastigotes, the parasite responsible for sleeping sickness. These extracts have previously shown potent antiprotozoal activities in vitro against Plasmodium falciparum and Leishmania donovani. The selectivity of the extracts was also evaluated by testing cytotoxicity on a mammalian L6 cell line. The ethyl acetate extract of the brown seaweed, Bifurcaria bifurcata, showed strong trypanocidal activity with a mild selectivity index (IC50 = 0.53 µg/mL; selectivity index (SI) = 11.6). Bio-guided fractionation led to the isolation of eleganolone, the main diterpenoid isolated from this species. Eleganolone contributes only mildly to the trypanocidal activity of the ethyl acetate extract (IC50 = 45.0 µM, SI = 4.0). However, a selective activity against P. falciparum erythrocytic stages in vitro has been highlighted (IC50 = 7.9 µM, SI = 21.6). PMID:23442789

  4. [Assessment of diagnostic methods for the catheter-related bloodstream infections in intensive care units].

    Science.gov (United States)

    Ataman Hatipoğlu, Ciğdem; Ipekkan, Korhan; Oral, Behiç; Onde, Ufuk; Bulut, Cemal; Demiröz, Ali Pekcan

    2011-01-01

    The majority of catheter-related bloodstream infections (CR-BSI) are associated with central venous catheters (CVCs) and most of them develop in patients staying at intensive care units (ICUs). The aim of this study was to assess the performance of different methods for the diagnosis of CR-BSI in neurology and neurosurgery ICUs of our hospital. This prospective study was carried out between January 2007 and January 2008 and all of the patients were followed daily for CR-BSI after the insertion of CVCs. Blood cultures were taken simultaneously from the catheter lumen and from at least one peripheral vein when there was a suspicion of CR-BSI. Additionally, from patients whose CVCs were removed, catheter tip cultures were taken and from patients with exit site infection, cultures of the skin surrounding the catheter entrance were taken. Catheter tip cultures were done by using quantitative and semiquantitative culture methods. Blood cultures taken from the catheter lumen and peripheral vein were incubated in the BACTEC 9050 (Becton Dickinson, USA) automated blood culture system. Gram and acridine orange (AO) staining were used for the smears prepared from the catheter tips and blood cultures. To evaluate the value of culture and staining methods in the diagnosis of CR-BSI; sensitivity, specificity, positive and negative predictive values (PPV and NPV, respectively) of each method were determined. A total of 148 patients (66 male, 82 female; age range: 1-94 years, mean age: 58.7 ± 21.8 years) were included in the study, of whom 67 (45.3%) were from neurology and 81 (54.7%) were from neurosurgery ICUs. One hundred ninety-nine CVC application performed in 148 patients were evaluated. Mean duration of catheterization was 8.5 ± 5.2 days. Thirty-two episodes of CR-BSI among 199 catheterizations (16%) in 29 patients among a total of 148 patients (19.6%) were determined. The most frequently isolated microorganisms were methicillin-resistant coagulase-negative staphylococci

  5. Impact of a modified Broviac maintenance care bundle on bloodstream infections in paediatric cancer patients

    Directory of Open Access Journals (Sweden)

    Furtwängler, Rhoikos

    2015-11-01

    Full Text Available Background: During intensive chemotherapy, bloodstream infection (BSI represents an important complication in paediatric cancer patients. Most patients carry a long-term central venous access device (CVAD. Improved maintenance care of these vascular catheters may decrease the risk of BSI.Methods: Intervention study (adapted CVAD prevention protocol with two observation periods (P1: 09-2009 until 05-2011; P2: 09-2011 until 05-2013; prospective surveillance of all laboratory confirmed BSIs. In P2, ready to use sterile NaCl 0.9% syringes were used for CVAD flushing and octenidine/isopropanol for the disinfection of catheter hubs and 3-way stopcocks. Results: During P1, 84 patients were included versus 81 patients during P2. There were no significant differences between the two patient populations in terms of median age, gender, underlying malignancy or disease status (first illness or relapse. Nearly all CVADs were Broviac catheters. The median duration from implantation to removal of the CVAD was 192 days (Inter-quartile-range (IQR; 110–288 days in P1 and 191 days (IQR; 103–270 days in P2. 28 BSI were diagnosed in 22 patients in P1 (26% of all patients experienced at least one BSI and 15 BSI in 12 patients in P2 (15% of all patients. The corresponding results for incidence density (ID were 0.44 (CI95 0.29–0.62 for P1 vs. 0.34 (0.19–0.53 BSI per 100 inpatient days for P2 and for incidence rate (IR 7.76 (5.16–10.86 in P1 vs. 4.75 (2.66–7.43 BSI per 1,000 inpatient CVAD utilization days. In P1, 9 BSI were caused by CoNS vs. only 2 in P2 (IR 2.49; CI95 0.17–4.17 vs. 0.63; CI95 0.08–1.72. In P1 two BSI (7% lead to early removal of the device. During P2 one CVAD was prematurely removed due to a Broviac-related BSI (6.7%.Conclusion: The preventive protocol investigated in this study led to a reduction of BSI in paediatric cancer patients. This result was clinically relevant but – due to insufficient power in a single centre observation

  6. Bisphosphonates inhibit the growth of Trypanosoma brucei, Trypanosoma cruzi, Leishmania donovani, Toxoplasma gondii, and Plasmodium falciparum: a potential route to chemotherapy.

    Science.gov (United States)

    Martin, M B; Grimley, J S; Lewis, J C; Heath, H T; Bailey, B N; Kendrick, H; Yardley, V; Caldera, A; Lira, R; Urbina, J A; Moreno, S N; Docampo, R; Croft, S L; Oldfield, E

    2001-03-15

    We have investigated the effects in vitro of a series of bisphosphonates on the proliferation of Trypanosoma cruzi, Trypanosoma brucei rhodesiense, Leishmania donovani, Toxoplasma gondii, and Plasmodium falciparum. The results show that nitrogen-containing bisphosphonates of the type used in bone resorption therapy have significant activity against parasites, with the aromatic species having in some cases nanomolar or low-micromolar IC(50) activity values against parasite replication (e.g. o-risedronate, IC(50) = 220 nM for T. brucei rhodesiense; risedronate, IC(50) = 490 nM for T. gondii). In T. cruzi, the nitrogen-containing bisphosphonate risedronate is shown to inhibit sterol biosynthesis at a pre-squalene level, most likely by inhibiting farnesylpyrophosphate synthase. Bisphosphonates therefore appear to have potential in treating parasitic protozoan diseases.

  7. DNA microarray genotyping and virulence and antimicrobial resistance gene profiling of methicillin-resistant Staphylococcus aureus bloodstream isolates from renal patients.

    LENUS (Irish Health Repository)

    McNicholas, Sinead

    2012-02-01

    Thirty-six methicillin-resistant Staphylococcus aureus (MRSA) bloodstream isolates from renal patients were genetically characterized by DNA microarray analysis and spa typing. The isolates were highly clonal, belonging mainly to ST22-MRSA-IV. The immune evasion and enterotoxin gene clusters were found in 29\\/36 (80%) and 33\\/36 (92%) isolates, respectively.

  8. DNA microarray genotyping and virulence and antimicrobial resistance gene profiling of methicillin-resistant Staphylococcus aureus bloodstream isolates from renal patients.

    LENUS (Irish Health Repository)

    McNicholas, Sinead

    2011-12-01

    Thirty-six methicillin-resistant Staphylococcus aureus (MRSA) bloodstream isolates from renal patients were genetically characterized by DNA microarray analysis and spa typing. The isolates were highly clonal, belonging mainly to ST22-MRSA-IV. The immune evasion and enterotoxin gene clusters were found in 29\\/36 (80%) and 33\\/36 (92%) isolates, respectively.

  9. Evaluation of the MALDI-TOF VITEK MS™ system for the identification of Candida parapsilosis, C. orthopsilosis and C. metapsilosis from bloodstream infections.

    Science.gov (United States)

    Nobrega de Almeida Júnior, João; de Souza, Letícia Bonato; Motta, Adriana Lopes; Rossi, Flávia; Romano Di Gioia, Thais Sabato; Benard, Gil; Del Negro, Gilda Maria Barbaro

    2014-10-01

    Twenty-nine Candida parapsilosis, seventeen Candida orthopsilosis and two Candida metapsilosis bloodstream isolates were submitted for identification by VITEK-MS™ mass spectrometer. Four isolates, two C. orthopsilosis and two C. metapsilosis, were not identified. Inclusion of Superspectra of both species in this database is required to improve its discrimination power.

  10. Burden of antimicrobial resistance in European hospitals : excess mortality and length of hospital stay associated with bloodstream infections due to Escherichia coli resistant to third-generation cephalosporins

    NARCIS (Netherlands)

    de Kraker, M. E. A.; Wolkewitz, M.; Davey, P. G.; Koller, W.; Berger, J.; Nagler, J.; Icket, C.; Kalenic, S.; Horvatic, J.; Seifert, H.; Kaasch, A.; Paniara, O.; Argyropoulou, A.; Bompola, M.; Smyth, E.; Skally, M.; Raglio, A.; Dumpis, U.; Kelmere, A. Melbarde; Borg, M.; Xuereb, D.; Ghita, M. C.; Noble, M.; Kolman, J.; Grabljevec, S.; Turner, D.; Lansbury, L.; Grundmann, H.

    2011-01-01

    This study determined excess mortality and length of hospital stay (LOS) attributable to bloodstream infection (BSI) caused by third-generation-cephalosporin-resistant Escherichia coli in Europe. A prospective parallel matched cohort design was used. Cohort I consisted of patients with third-generat

  11. Etiology and epidemiology of catheter related bloodstream infections in patients receiving home parenteral nutrition in a gastromedical center at a tertiary hospital in denmark

    DEFF Research Database (Denmark)

    Nielsen, Xiaohui Chen; Chen, Ming; Hellesøe, Anne-Marie Blok

    2012-01-01

    We conducted a retrospective epidemiologic study of catheter related bloodstream infections (CRBSI) in patients receiving long-term home parenteral nutrition (HPN) from January 2002 to December 2005. Our results showed that coagulase negative staphylococci (CoNS) were the most prevalent pathogens...

  12. Taurolidine lock is highly effective in preventing catheter-related bloodstream infections in patients on home parenteral nutrition: a heparin-controlled prospective trial.

    NARCIS (Netherlands)

    Bisseling, T.M.; Willems, M.C.M.; Versleijen, M.W.J.; Hendriks, J.C.M.; Vissers, R.K.; Wanten, G.J.A.

    2010-01-01

    BACKGROUND & AIMS: Catheter-related bloodstream infections remain the major threat for Home Parenteral Nutrition programs. Taurolidine, a potent antimicrobial agent, holds promise as an effective catheter lock to prevent such infections. Aim of the present study was to compare taurolidine with hepar

  13. Absence of microbial adaptation to taurolidine in patients on home parenteral nutrition who develop catheter related bloodstream infections and use taurolidine locks

    NARCIS (Netherlands)

    Olthof, E.D.; Rentenaar, R.J.; Rijs, A.J.M.M.; Wanten, G.J.A.

    2013-01-01

    BACKGROUND & AIMS: Some home parenteral nutrition (HPN) patients develop catheter related bloodstream infections (CRBSI) despite using an anti-microbial catheter lock solution taurolidine. The aim of this study was to assess whether long-term use of taurolidine leads to selective growth of microorga

  14. Effect of a vascular access team on central line-associated bloodstream infections in infants admitted to a neonatal intensive care unit : a systematic review

    NARCIS (Netherlands)

    Legemaat, Monique M; Jongerden, IP; van Rens, Roland M F P T; Zielman, Marjanne; van den Hoogen, Agnes

    2015-01-01

    OBJECTIVE: To review the effect of a vascular access team on the incidence of central line-associated bloodstream infections in infants admitted to a neonatal intensive care unit. DATA SOURCES: MEDLINE, CINAHL, Embase, Web-of-Science and the Cochrane Library were searched until December 2013. STUDY

  15. Draft Genome Sequence of Extremely Drug-Resistant Pseudomonas aeruginosa (ST357) Strain CMC_VB_PA_B22862 Isolated from a Community-Acquired Bloodstream Infection

    Science.gov (United States)

    Pragasam, Agila Kumari; Yesurajan, Francis; Doss C, George Priya; George, Biju; Devanga Ragupathi, Naveen Kumar; Walia, Kamini

    2016-01-01

    Extremely drug-resistant Pseudomonas aeruginosa strains causing severe infections have become a serious concern across the world. Here, we report draft genome sequence of P. aeruginosa with an extremely drug-resistant profile isolated from a patient with community-acquired bloodstream infection in India.

  16. Bloodstream infections during the onset of necrotizing enterocolitis and their relation with the pro-inflammatory response, gut wall integrity and severity of disease in NEC

    NARCIS (Netherlands)

    Heida, F. H.; Hulscher, J. B. F.; Schurink, M.; van Vliet, M. J.; Kooi, E. M. W.; Kasper, D. C.; Pones, M.; Bos, A. F.; Benkoe, T. M.

    2015-01-01

    Introduction: Bacterial involvement is believed to play a pivotal role in the development and disease outcome of NEC. However, whether a bloodstream infection (BSI) predisposes to NEC (e.g. by activating the pro-inflammatory response) or result from the loss of gut wall integrity during NEC developm

  17. Bloodstream infections during the onset of necrotizing enterocolitis and their relation with the pro-inflammatory response, gut wall integrity and severity of disease in NEC

    NARCIS (Netherlands)

    Heida, F.H.; Hulscher, J.B.; Schurink, M.; Vliet, M.J. van; Kooi, E.M.; Kasper, D.C.; Pones, M.; Bos, A.F; Benkoe, T.M.

    2015-01-01

    INTRODUCTION: Bacterial involvement is believed to play a pivotal role in the development and disease outcome of NEC. However, whether a bloodstream infection (BSI) predisposes to NEC (e.g. by activating the pro-inflammatory response) or result from the loss of gut wall integrity during NEC developm

  18. Identification of three classes of heteroaromatic compounds with activity against intracellular Trypanosoma cruzi by chemical library screening.

    Directory of Open Access Journals (Sweden)

    Esther Bettiol

    Full Text Available The development of new drugs against Chagas disease is a priority since the currently available medicines have toxic effects, partial efficacy and are targeted against the acute phase of disease. At present, there is no drug to treat the chronic stage. In this study, we have optimized a whole cell-based assay for high throughput screening of compounds that inhibit infection of mammalian cells by Trypanosoma cruzi trypomastigotes. A 2000-compound chemical library was screened using a recombinant T. cruzi (Tulahuen strain expressing beta-galactosidase. Three hits were selected for their high activity against T. cruzi and low toxicity to host cells in vitro: PCH1, NT1 and CX1 (IC(50: 54, 190 and 23 nM, respectively. Each of these three compounds presents a different mechanism of action on intracellular proliferation of T. cruzi amastigotes. CX1 shows strong trypanocidal activity, an essential characteristic for the development of drugs against the chronic stage of Chagas disease where parasites are found intracellular in a quiescent stage. NT1 has a trypanostatic effect, while PCH1 affects parasite division. The three compounds also show high activity against intracellular T. cruzi from the Y strain and against the related kinetoplastid species Leishmania major and L. amazonensis. Characterization of the anti-T. cruzi activity of molecules chemically related to the three library hits allowed the selection of two compounds with IC(50 values of 2 nM (PCH6 and CX2. These values are approximately 100 times lower than those of the medicines used in patients against T. cruzi. These results provide new candidate molecules for the development of treatments against Chagas disease and leishmaniasis.

  19. Mining SNPs in extracellular vesicular transcriptome of Trypanosoma cruzi: a step closer to early diagnosis of neglected Chagas disease

    Science.gov (United States)

    Chaturvedi, Anoop

    2016-01-01

    One of the newest and strongest members of intercellular communicators, the Extracellular vesicles (EVs) and their enclosed RNAs; Extracellular RNAs (exRNAs) have been acknowledged as putative biomarkers and therapeutic targets for various diseases. Although a very deep insight has not been possible into the physiology of these vesicles, they are believed to be involved in cell-to-cell communication and host-pathogen interactions. EVs might be significantly helpful in discovering biomarkers for possible target identification as well as prognostics, diagnostics and developing vaccines. In recent studies, highly bioactive EVs have drawn attention of parasitologists for being able to communicate between different cells and having likeliness of reflecting both source and target environments. Next-generation sequencing (NGS) has eased the way to have a deeper insight into these vesicles and their roles in various diseases. This article arises from bioinformatics-based analysis and predictive data mining of transcriptomic (RNA-Seq) data of EVs, derived from different life stages of Trypanosoma cruzi; a causing agent of neglected Chagas disease. Variants (Single Nucleotide Polymorphisms (SNPs)) were mined from Extracellular vesicular transcriptomic data and functionally analyzed using different bioinformatics based approaches. Functional analysis showed the association of these variants with various important factors like Trans-Sialidase (TS), Alpha Tubulin, P-Type H+-ATPase, etc. which, in turn, are associated with disease in different ways. Some of the ‘candidate SNPs’ were found to be stage-specific, which strengthens the probability of finding stage-specific biomarkers. These results may lead to a better understanding of Chagas disease, and improved knowledge may provide further development of the biomarkers for prognosis, diagnosis and drug development for treating Chagas disease. PMID:27904804

  20. Eco-epidemiological aspects of Trypanosoma cruzi, Trypanosoma rangeli and their vector (Rhodnius pallescens in Panama Generalidades do Trypanosoma cruzi, do Trypanosoma rangeli e do seu vetor (Rhodnius pallescens no Panamá

    Directory of Open Access Journals (Sweden)

    Ana Maria de Vasquez

    2004-08-01

    Full Text Available The eco-epidemiology of T. cruzi infection was investigated in the Eastern border of the Panama Canal in Central Panama. Between 1999 and 2000, 1110 triatomines were collected: 1050 triatomines (94.6% from palm trees, 27 (2.4% from periurban habitats and 33 (3.0% inside houses. All specimens were identified as R. pallescens. There was no evidence of vector domiciliation. Salivary glands from 380 R. pallescens revealed a trypanosome natural infection rate of 7.6%, while rectal ampoule content from 373 triatomines was 45%. Isoenzyme profiles on isolated trypanosomes demonstrated that 85.4% (n = 88 were T. cruzi and 14.6% (n = 15 were T. rangeli. Blood meal analysis from 829 R. pallescens demonstrated a zoophilic vector behavior, with opossums as the preferential blood source. Seroprevalence in human samples from both study sites was less than 2%. Our results demonstrate that T. cruzi survives in the area in balanced association with R. pallescens, and with several different species of mammals in their natural niches. However, the area is an imminent risk of infection for its population, consequently it is important to implement a community educational program regarding disease knowledge and control measures.A epidemiologia da infecção do T. cruzi foi investigada na margem oriental do canal do Panamá, na região central da Republica do Panamá. A informação obtida durante o estudo avaliou fatores de risco da doença de Chagas nesta área. Entre 1999 e 2000, 1110 triatomíneos foram coletados: 1050 triatomíneos (94,6% em palmeiras, 27 (2,4% em habitats periurbanos e 33 (3,0% no interior de casas. Todos os espécimens foram identificados como R. pallescens. Não havia nenhuma evidência de domiciliação do vetor. O exame de glândulas salivares de 380 R. pallescens revelaram taxa de infecção natural por Trypanosoma de 7,6%, mas o conteúdo da ampola rectal de 373 triatomíneos mostrou 45% de positividade. Os perfis de isoenzimas em

  1. Molecular basis of mammalian cell invasion by Trypanosoma cruzi

    Directory of Open Access Journals (Sweden)

    Nobuko Yoshida

    2006-03-01

    Full Text Available Establishment of infection by Trypanosoma cruzi, the agent of Chagas' disease, depends on a series of events involving interactions of diverse parasite molecules with host components. Here we focus on the mechanisms of target cell invasion by metacyclic trypomastigotes (MT and mammalian tissue culture trypomastigotes (TCT. During MT or TCT internalization, signal transduction pathways are activated both in the parasite and the target cell, leading to Ca2+ mobilization. For cell adhesion, MT engage surface glycoproteins, such as gp82 and gp35/50, which are Ca2+ signal-inducing molecules. In T. cruzi isolates that enter host cells in gp82-mediated manner, parasite protein tyrosine kinase as well as phospholipase C are activated, and Ca2+ is released from I P3-sensitive stores, whereas in T. cruzi isolates that attach to target cells mainly through gp35/50, the signaling pathway involving adenylate cyclase appears to be stimulated, with Ca2+ release from acidocalciosomes. In addition, T. cruzi isolate-dependent inhibitory signals, mediated by MT-specific gp90, may be triggered both in the host cell and the parasite. The repertoire of TCT molecules implicated in cell invasion includes surface glycoproteins of gp85 family, with members containing binding sites for laminin and cytokeratin 18, enzymes such as cruzipain, trans-sialidase, and an oligopeptidase B that generates a Ca2+-agonist from a precursor molecule.O estabelecimento da infecção por Trypanosoma cruzi, o agente da doença de Chagas, depende de uma série de eventos envolvendo interações de diversas moléculas do parasita com componentes do hospedeiro. Focalizamos aqui os mecanismos de invasão celular por tripomastigotas metacíclicos (TM e por tripomastigotas de cultura de tecido (TCT. Durante a internalização de TM ou TCT, vias de transdução de sinal são ativadas tanto no parasita como na célula alvo, acarretando a mobilização de Ca2+. Para adesão, TM utiliza as glicoprote

  2. Neuronal changes caused by Trypanosoma cruzi: an experimental model

    Directory of Open Access Journals (Sweden)

    Neide M Moreira

    2011-06-01

    Full Text Available Define an experimental model by evaluating quantitative and morphometric changes in myenteric neurons of the colon of mice infected with Trypanosoma cruzi. Twenty-eight Swiss male mice were distributed into groups: control (CG, n=9 and inoculated with 100 (IG100, n=9 and 1000 (IG1000, n=10 blood trypomastigotes, Y strain-T. cruzi II. Parasitemia was evaluated from 3-25 days post inoculation (dpi with parasites peak of 7.7 × 10(6 and 8.4 × 10(6 trypomastigotes/mL at 8th dpi (p>0.05 in IG100 and IG1000, respectively. Chronic phase of the infection was obtained with two doses of 100mg/Kg/weight and one dose of 250mg/Kg/weight of Benznidazole on 11, 16 and 18 dpi. Three animals from each group were euthanized at 18, 30 and 75 dpi. The colon was stained with Giemsa. The quantitative and morphometric analysis of neurons revealed that the infection caused a decrease of neuronal density on 30th dpi (pDefinir um modelo experimental de avaliação de alterações quantitativas e morfométricas nos neurônios mientéricos do cólon de camundongos infectados pelo Trypanosoma cruzi. Vinte e oito camundongos Swiss machos foram distribuídos nos grupos: controle (GC, n=9 e infectados com 100 (IG100, n=9 e 1000 (IG1000, n=10 tripomastigotas sanguíneos, cepa Y-T. cruzi II. A parasitemia foi avaliada 3-25 dias pós inoculação (dpi, com pico de parasitos de 7,7 × 10(6 e 8,4 × 10(6 tripomastigotas/mL no 8º dpi (p>0,05 em IG100 e IG1000, respectivamente. A fase crônica da infecção foi obtida com duas doses de 100mg/Kg/weight e uma dose de 250mg/Kg/ weight do benznidazol, em 11, 16 e 18 dpi. Três animais de cada grupo foram sacrificados aos 18, 30 e 75 dpi. O cólon foi corado com Giemsa. A análise quantitativa e morfométrica de neurônios revelou que a infecção causou uma diminuição da densidade neuronal no 30º dpi (p<0,05 e 75 dpi (p<0,05 em IG100 e IG1000. A infecção causou morte e hipertrofia neuronal no 75º dpi em IG100 e IG1000 (p<0,05, p

  3. Etiological characteristics of 108 patients with secondary bloodstream infections%继发性血流感染108例病原学特点分析

    Institute of Scientific and Technical Information of China (English)

    黄仁刚; 杨兴祥; 喻华; 龙姗姗; 林健梅; 江南

    2015-01-01

    Objective To investigate the etiological characteristics of laboratory-confirmed bloodstream infections with identi-fied infective sources. Methods The data of the patients with laboratory-confirmed bloodstream infections and identified infective sources, who were treated at Sichuan Provincial People's Hospital from Jan. 2011 to Jun. 2013 were collected to analyze the etiological characteristics retrospectively. Results A total of 108 patients with identified infective sources were enrolled in this study, of whom 93 patients suffered from monomicrobial infection, and 15 patients suffered from polymicrobial infection. Bloodstream infections were com-monly found in urinary tract, abdominal cavity and respiratory tract. Infection with Escherichia coli. accounted for 75.8%and 42.4%in patients with bloodstream infections in urinary tract and abdominal cavity, respectively; Infection with Acinetobacter baumannii ac-counted for 62.5%in patients with bloodstream infections in respiratory tract, and Acinetobacter baumannii was resistant to carbapen-em antibiotics. The 30-day mortality of 108 patients with bloodstream infections was 19.4%. The patients with bloodstream infections in urinary tract had the lowest 30-day mortality rate (3.0%), while the patients with bloodstream infections in lower respiratory tract had the highest 30-day mortality rate (45.8%). The 30-day mortality rates of the patients with bloodstream infections with non-fermentation gram negative bacillus and fungi were 55.0%and 50.0%, respectively. Conclusions The pathogen distribution of the patients with different sources of bloodstream infections varies widely. Appropriate antibiotic therapy should take infective sources, types of bacteria and drug resistance into consideration.%目的 研究感染来源明确的血流感染患者的病原学特点. 方法 收集四川省人民医院2011年1月—2013年6月实验室确诊、感染来源明确的血流感染患者临床资料,回

  4. Functional characterization of 8-oxoguanine DNA glycosylase of Trypanosoma cruzi.

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    Carolina Furtado

    Full Text Available The oxidative lesion 8-oxoguanine (8-oxoG is removed during base excision repair by the 8-oxoguanine DNA glycosylase 1 (Ogg1. This lesion can erroneously pair with adenine, and the excision of this damaged base by Ogg1 enables the insertion of a guanine and prevents DNA mutation. In this report, we identified and characterized Ogg1 from the protozoan parasite Trypanosoma cruzi (TcOgg1, the causative agent of Chagas disease. Like most living organisms, T. cruzi is susceptible to oxidative stress, hence DNA repair is essential for its survival and improvement of infection. We verified that the TcOGG1 gene encodes an 8-oxoG DNA glycosylase by complementing an Ogg1-defective Saccharomyces cerevisiae strain. Heterologous expression of TcOGG1 reestablished the mutation frequency of the yeast mutant ogg1(-/- (CD138 to wild type levels. We also demonstrate that the overexpression of TcOGG1 increases T. cruzi sensitivity to hydrogen peroxide (H(2O(2. Analysis of DNA lesions using quantitative PCR suggests that the increased susceptibility to H(2O(2 of TcOGG1-overexpressor could be a consequence of uncoupled BER in abasic sites and/or strand breaks generated after TcOgg1 removes 8-oxoG, which are not rapidly repaired by the subsequent BER enzymes. This hypothesis is supported by the observation that TcOGG1-overexpressors have reduced levels of 8-oxoG both in the nucleus and in the parasite mitochondrion. The localization of TcOgg1 was examined in parasite transfected with a TcOgg1-GFP fusion, which confirmed that this enzyme is in both organelles. Taken together, our data indicate that T. cruzi has a functional Ogg1 ortholog that participates in nuclear and mitochondrial BER.

  5. Metalloproteases in Trypanosoma rangeli-infected Rhodnius prolixus

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    D Feder

    1999-11-01

    Full Text Available Protease activities in the haemolymph and fat body in a bloodsucking insect, Rhodnius prolixus, infected with Trypanosoma rangeli, were investigated. After SDS-polyacrylamide gel electrophoresis containing gelatin as substrate, analysis of zymograms performed on samples of different tissues of controls and insects inoculated or orally infected with short or long epimastigotes of T. rangeli, demonstrated distinct patterns of protease activities: (i proteases were detected in the haemolymph of insects which were fed on, or inoculated with, short epimastigotes of T. rangeli (39 kDa and 33 kDa, respectively, but they were not observed in the fat body taken from these insects; (ii protease was also presented in the fat bodies derived from naive insects or controls inoculated with sterile phosphate-saline buffer (49 kDa, but it was not detected in the haemolymph of these insects; (iii no protease activity was observed in both haemolymph and fat bodies taken from insects inoculated with, or fed on, long epimastigotes of T. rangeli. Furthermore, in short epimastigotes of T. rangeli extracts, three bands of the protease activities with apparent molecular weights of 297, 198 and 95 kDa were detected while long epimastigotes preparation presented only two bands of protease activities with molecular weights of 297 and 198 kDa. The proteases from the insect infected with T. rangeli and controls belong to the class of either metalloproteases or metal-activated enzymes since they are inhibited by 1,10-phenanthroline. The significance of these proteases in the insects infected with short epimastigotes of T. rangeli is discussed in relation to the success of the establishment of infection of these parasites in its vector, R. prolixus.

  6. Growth hormones therapy in immune response against Trypanosoma cruzi.

    Science.gov (United States)

    Frare, Eduardo Osório; Santello, Fabricia Helena; Caetano, Leony Cristina; Caldeira, Jerri C; Toldo, Míriam Paula Alonso; Prado, José Clóvis do

    2010-04-01

    Growth hormone (GH) is an important hypophyseal hormone that is primarily involved in body growth and metabolism. In mammals, control of Trypanosoma cruzi parasitism during the acute phase of infection is considered to be critically dependent on direct macrophage activation by cytokines. To explore the possibility that GH might be effective in the treatment of Chagas' disease, we investigated its effects on the course of T. cruzi infection in rats, focusing our analyses on its influences on parasitemia, NO, TNF-alpha and IFN-gamma concentration and on histopathological alterations and parasite burden in heart tissue. T. cruzi-infected male Wistar rats were intraperitoneally treated with 5 ng/10 g body weight/day of GH. Animals treated with GH showed a significant reduction in the number of blood trypomastigotes during the acute phase of infection compared with untreated animals (P<0.05). For all experimental days (7, 14 and 21 post infection) of the acute phase, infected and GH treated animals reached higher concentrations of TNF-alpha, IFN-gamma and nitric oxide as compared to untreated and infected counterparts (P<0.05) Histopathological observations of heart tissue revealed that GH administration also resulted in fewer and smaller amastigote burdens, and less inflammatory infiltrate and tissue disorganization, indicating a reduced parasitism of this tissue. These results show that GH can be considered as an immunomodulator substance for controlling parasite replication and combined with the current drug used may represent in the future a new therapeutic tool to reduce the harmful effects of Chagas' disease.

  7. Immunotherapy of Trypanosoma cruzi infection with DNA vaccines in mice.

    Science.gov (United States)

    Dumonteil, Eric; Escobedo-Ortegon, Javier; Reyes-Rodriguez, Norma; Arjona-Torres, Arletty; Ramirez-Sierra, Maria Jesus

    2004-01-01

    The mechanisms involved in the pathology of chronic chagasic cardiomyopathy are still debated, and the controversy has interfered with the development of new treatments and vaccines. Because of the potential of DNA vaccines for immunotherapy of chronic and infectious diseases, we tested if DNA vaccines could control an ongoing Trypanosoma cruzi infection. BALB/c mice were infected with a lethal dose (5 x 10(4) parasites) as a model of acute infection, and then they were treated with two injections of 100 microg of plasmid DNA 1 week apart, beginning on day 5 postinfection. Control mice had high levels of parasitemia and mortality and severe cardiac inflammation, while mice treated with plasmid DNA encoding trypomastigote surface antigen 1 or Tc24 had reduced parasitemia and mild cardiac inflammation and >70% survived the infection. The efficacy of the immunotherapy also was significant when it was delayed until days 10 and 15 after infection. Parasitological analysis of cardiac tissue of surviving mice indicated that most mice still contained detectable parasite kinetoplast DNA but fewer mice contained live parasites, suggesting that there was efficient but not complete parasite elimination. DNA vaccine immunotherapy was also evaluated in CD1 mice infected with a low dose (5 x 10(2) parasites) as a model of chronic infection. Immunotherapy was initiated on day 70 postinfection and resulted in improved survival and reduced cardiac tissue inflammation. These results suggest that DNA vaccines have strong potential for the immunotherapy of T. cruzi infection and may provide new alternatives for the control of Chagas' disease.

  8. Trypanosoma cruzi population dynamics in the Central Ecuadorian Coast.

    Science.gov (United States)

    Costales, Jaime A; Jara-Palacios, Miguel A; Llewellyn, Martin S; Messenger, Louisa A; Ocaña-Mayorga, Sofía; Villacís, Anita G; Tibayrenc, Michel; Grijalva, Mario J

    2015-11-01

    Chagas disease is the most important parasitic disease in Latin America. The causative agent, Trypanosoma cruzi, displays high genetic diversity and circulates in complex transmission cycles among domestic, peridomestic and sylvatic environments. In Ecuador, Rhodnius ecuadoriensis is known to be the major vector species implicated in T. cruzi transmission. However, across vast areas of Ecuador, little is known about T. cruzi genetic diversity in relation to different parasite transmission scenarios. Fifty-eight T. cruzi stocks from the central Ecuadorian coast, most of them derived from R. ecuadoriensis, were included in the study. All of them were genotyped as T. cruzi discrete typing unit I (DTU TcI). Analysis of 23 polymorphic microsatellite loci through neighbor joining and discriminant analysis of principal components yielded broadly congruent results and indicate genetic subdivision between sylvatic and peridomestic transmission cycles. However, both analyses also suggest that any barriers are imperfect and significant gene flow between parasite subpopulations in different habitats exists. Also consistent with moderate partition and residual gene flow between subpopulations, the fixation index (FST) was significant, but of low magnitude. Finally, the lack of private alleles in the domestic/peridomestic transmission cycle suggests the sylvatic strains constitute the ancestral population. The T. cruzi population in the central Ecuadorian coast shows moderate tendency to subdivision according to transmission cycle. However, connectivity between cycles exists and the sylvatic T. cruzi population harbored by R. ecuadoriensis vectors appears to constitute a source from which the parasite invades human domiciles and their surroundings in this region. We discuss the implications these findings have for the planning, implementation and evaluation of local Chagas disease control interventions.

  9. New Trypanosoma evansi Type B Isolates from Ethiopian Dromedary Camels.

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    Hadush Birhanu

    2016-04-01

    Full Text Available Trypanosoma (T. evansi is a dyskinetoplastic variant of T. brucei that has gained the ability to be transmitted by all sorts of biting flies. T. evansi can be divided into type A, which is the most abundant and found in Africa, Asia and Latin America and type B, which has so far been isolated only from Kenyan dromedary camels. This study aimed at the isolation and the genetic and phenotypic characterisation of type A and B T. evansi stocks from camels in Northern Ethiopia.T. evansi was isolated in mice by inoculation with the cryopreserved buffy coat of parasitologically confirmed animals. Fourteen stocks were thus isolated and subject to genotyping with PCRs targeting type-specific variant surface glycoprotein genes, mitochondrial minicircles and maxicircles, minisatellite markers and the F1-ATP synthase γ subunit gene. Nine stocks corresponded to type A, two stocks were type B and three stocks represented mixed infections between A and B, but not hybrids. One T. evansi type A stock was completely akinetoplastic. Five stocks were adapted to in vitro culture and subjected to a drug sensitivity assay with melarsomine dihydrochloride, diminazene diaceturate, isometamidium chloride and suramin. In vitro adaptation induced some loss of kinetoplasts within 60 days. No correlation between drug sensitivity and absence of the kinetoplast was observed. Sequencing the full coding sequence of the F1-ATP synthase γ subunit revealed new type-specific single nucleotide polymorphisms and deletions.This study addresses some limitations of current molecular markers for T. evansi genotyping. Polymorphism within the F1-ATP synthase γ subunit gene may provide new markers to identify the T. evansi type that do not rely on variant surface glycoprotein genes or kinetoplast DNA.

  10. Rab23 is a flagellar protein in Trypanosoma brucei

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    Field Mark C

    2011-06-01

    Full Text Available Abstract Background Rab small GTPases are important mediators of membrane transport, and orthologues frequently retain similar locations and functions, even between highly divergent taxa. In metazoan organisms Rab23 is an important negative regulator of Sonic hedgehog signaling and is crucial for correct development and differentiation of cellular lineages by virtue of an involvement in ciliary recycling. Previously, we reported that Trypanosoma brucei Rab23 localized to the nuclear envelope 1, which is clearly inconsistent with the mammalian location and function. As T. brucei is unicellular the potential that Rab23 has no role in cell signaling was possible. Here we sought to further investigate the role(s of Rab23 in T. brucei to determine if Rab23 was an example of a Rab protein with divergent function in distinct taxa. Methods/major findings The taxonomic distribution of Rab23 was examined and compared with the presence of flagella/cilia in representative taxa. Despite evidence for considerable secondary loss, we found a clear correlation between a conventional flagellar structure and the presence of a Rab23 orthologue in the genome. By epitope-tagging, Rab23 was localized and found to be present at the flagellum throughout the cell cycle. However, RNAi knockdown did not result in a flagellar defect, suggesting that Rab23 is not required for construction or maintenance of the flagellum. Conclusions The location of Rab23 at the flagellum is conserved between mammals and trypanosomes and the Rab23 gene is restricted to flagellated organisms. These data may suggest the presence of a Rab23-mediated signaling mechanism in trypanosomes.

  11. Optimized multilocus sequence typing (MLST scheme for Trypanosoma cruzi.

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    Patricio Diosque

    2014-08-01

    Full Text Available Trypanosoma cruzi, the aetiological agent of Chagas disease possess extensive genetic diversity. This has led to the development of a plethora of molecular typing methods for the identification of both the known major genetic lineages and for more fine scale characterization of different multilocus genotypes within these major lineages. Whole genome sequencing applied to large sample sizes is not currently viable and multilocus enzyme electrophoresis, the previous gold standard for T. cruzi typing, is laborious and time consuming. In the present work, we present an optimized Multilocus Sequence Typing (MLST scheme, based on the combined analysis of two recently proposed MLST approaches. Here, thirteen concatenated gene fragments were applied to a panel of T. cruzi reference strains encompassing all known genetic lineages. Concatenation of 13 fragments allowed assignment of all strains to the predicted Discrete Typing Units (DTUs, or near-clades, with the exception of one strain that was an outlier for TcV, due to apparent loss of heterozygosity in one fragment. Monophyly for all DTUs, along with robust bootstrap support, was restored when this fragment was subsequently excluded from the analysis. All possible combinations of loci were assessed against predefined criteria with the objective of selecting the most appropriate combination of between two and twelve fragments, for an optimized MLST scheme. The optimum combination consisted of 7 loci and discriminated between all reference strains in the panel, with the majority supported by robust bootstrap values. Additionally, a reduced panel of just 4 gene fragments displayed high bootstrap values for DTU assignment and discriminated 21 out of 25 genotypes. We propose that the seven-fragment MLST scheme could be used as a gold standard for T. cruzi typing, against which other typing approaches, particularly single locus approaches or systematic PCR assays based on amplicon size, could be compared.

  12. Sialic Acid Glycobiology Unveils Trypanosoma cruzi Trypomastigote Membrane Physiology.

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    Andrés B Lantos

    2016-04-01

    Full Text Available Trypanosoma cruzi, the flagellate protozoan agent of Chagas disease or American trypanosomiasis, is unable to synthesize sialic acids de novo. Mucins and trans-sialidase (TS are substrate and enzyme, respectively, of the glycobiological system that scavenges sialic acid from the host in a crucial interplay for T. cruzi life cycle. The acquisition of the sialyl residue allows the parasite to avoid lysis by serum factors and to interact with the host cell. A major drawback to studying the sialylation kinetics and turnover of the trypomastigote glycoconjugates is the difficulty to identify and follow the recently acquired sialyl residues. To tackle this issue, we followed an unnatural sugar approach as bioorthogonal chemical reporters, where the use of azidosialyl residues allowed identifying the acquired sugar. Advanced microscopy techniques, together with biochemical methods, were used to study the trypomastigote membrane from its glycobiological perspective. Main sialyl acceptors were identified as mucins by biochemical procedures and protein markers. Together with determining their shedding and turnover rates, we also report that several membrane proteins, including TS and its substrates, both glycosylphosphatidylinositol-anchored proteins, are separately distributed on parasite surface and contained in different and highly stable membrane microdomains. Notably, labeling for α(1,3Galactosyl residues only partially colocalize with sialylated mucins, indicating that two species of glycosylated mucins do exist, which are segregated at the parasite surface. Moreover, sialylated mucins were included in lipid-raft-domains, whereas TS molecules are not. The location of the surface-anchored TS resulted too far off as to be capable to sialylate mucins, a role played by the shed TS instead. Phosphatidylinositol-phospholipase-C activity is actually not present in trypomastigotes. Therefore, shedding of TS occurs via microvesicles instead of as a fully

  13. Aspirin treatment exacerbates oral infections by Trypanosoma cruzi.

    Science.gov (United States)

    Cossentini, Luana Aparecida; Da Silva, Rosiane Valeriano; Yamada-Ogatta, Sueli Fumie; Yamauchi, Lucy Megumi; De Almeida Araújo, Eduardo José; Pinge-Filho, Phileno

    2016-05-01

    Oral transmission of the protozoan parasite Trypanosoma cruzi, the etiological agent of Chagas disease, has been documented in Latin American countries. The reported cases of infection were due to the ingestion of contaminated fresh fruit, juices, or sugar cane juice. There have been few studies on the physiopathology of the disease in oral transmission cases. Gastritis is a common ailment that can be caused by poor dietary habits, intake of alcohol or other gastric irritants, bacterial infection, or by the widespread use of non-steroidal anti-inflammatory drugs (NSAIDs). This study investigated in a mouse model whether gastric mucosal injury, induced by aspirin, would affect the course of disease in animals infected with T. cruzi by the oral route. The CL14 and G strains of T. cruzi, both of low infectivity, were used. To this end, groups of BALB/c mice were treated during 5 days with aspirin (100 mg kg(-1)) before oral infection with T. cruzi metacyclic forms (4 × 10(5) or 5 × 10(7) parasites/mouse). Histological analysis and determination of nitric oxide and TNF-α were performed in gastric samples obtained 5 days after infection. Parasitemia was monitored from the thirteenth day after infection. The results indicate that aspirin treatment of mice injured their gastric mucosa and facilitated invasion by both CL14 and G strains of T. cruzi. Strain CL14 caused more severe infection compared to the G strain, as larger numbers of amastigote nests were found in the stomach and parasitemia levels were higher. Our study is novel in that it shows that gastric mucosal damage caused by aspirin, a commonly used NSAID, facilitates T. cruzi infection by the oral route.

  14. Trypanosoma cruzi as an effective cancer antigen delivery vector.

    Science.gov (United States)

    Junqueira, Caroline; Santos, Luara I; Galvão-Filho, Bruno; Teixeira, Santuza M; Rodrigues, Flávia G; DaRocha, Wanderson D; Chiari, Egler; Jungbluth, Achim A; Ritter, Gerd; Gnjatic, Sacha; Old, Lloyd J; Gazzinelli, Ricardo T

    2011-12-06

    One of the main challenges in cancer research is the development of vaccines that induce effective and long-lived protective immunity against tumors. Significant progress has been made in identifying members of the cancer testis antigen family as potential vaccine candidates. However, an ideal form for antigen delivery that induces robust and sustainable antigen-specific T-cell responses, and in particular of CD8(+) T lymphocytes, remains to be developed. Here we report the use of a recombinant nonpathogenic clone of Trypanosoma cruzi as a vaccine vector to induce vigorous and long-term T cell-mediated immunity. The rationale for using the highly attenuated T. cruzi clone was (i) the ability of the parasite to persist in host tissues and therefore to induce a long-term antigen-specific immune response; (ii) the existence of intrinsic parasite agonists for Toll-like receptors and consequent induction of highly polarized T helper cell type 1 responses; and (iii) the parasite replication in the host cell cytoplasm, leading to direct antigen presentation through the endogenous pathway and consequent induction of antigen-specific CD8(+) T cells. Importantly, we found that parasites expressing a cancer testis antigen (NY-ESO-1) were able to elicit human antigen-specific T-cell responses in vitro and solid protection against melanoma in a mouse model. Furthermore, in a therapeutic protocol, the parasites expressing NY-ESO-1 delayed the rate of tumor development in mice. We conclude that the T. cruzi vector is highly efficient in inducing T cell-mediated immunity and protection against cancer cells. More broadly, this strategy could be used to elicit a long-term T cell-mediated immunity and used for prophylaxis or therapy of chronic infectious diseases.

  15. The cell cycle regulated transcriptome of Trypanosoma brucei.

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    Stuart K Archer

    Full Text Available Progression of the eukaryotic cell cycle requires the regulation of hundreds of genes to ensure that they are expressed at the required times. Integral to cell cycle progression in yeast and animal cells are temporally controlled, progressive waves of transcription mediated by cell cycle-regulated transcription factors. However, in the kinetoplastids, a group of early-branching eukaryotes including many important pathogens, transcriptional regulation is almost completely absent, raising questions about the extent of cell-cycle regulation in these organisms and the mechanisms whereby regulation is achieved. Here, we analyse gene expression over the Trypanosoma brucei cell cycle, measuring changes in mRNA abundance on a transcriptome-wide scale. We developed a "double-cut" elutriation procedure to select unperturbed, highly synchronous cell populations from log-phase cultures, and compared this to synchronization by starvation. Transcriptome profiling over the cell cycle revealed the regulation of at least 430 genes. While only a minority were homologous to known cell cycle regulated transcripts in yeast or human, their functions correlated with the cellular processes occurring at the time of peak expression. We searched for potential target sites of RNA-binding proteins in these transcripts, which might earmark them for selective degradation or stabilization. Over-represented sequence motifs were found in several co-regulated transcript groups and were conserved in other kinetoplastids. Furthermore, we found evidence for cell-cycle regulation of a flagellar protein regulon with a highly conserved sequence motif, bearing similarity to consensus PUF-protein binding motifs. RNA sequence motifs that are functional in cell-cycle regulation were more widespread than previously expected and conserved within kinetoplastids. These findings highlight the central importance of post-transcriptional regulation in the proliferation of parasitic kinetoplastids.

  16. Novel trypanosome Trypanosoma gilletti sp. (Euglenozoa: Trypanosomatidae) and the extension of the host range of Trypanosoma copemani to include the koala ( Phascolarctos cinereus).

    Science.gov (United States)

    McInnes, L M; Hanger, J; Simmons, G; Reid, S A; Ryan, U M

    2011-01-01

    Trypanosoma irwini was previously described from koalas and we now report the finding of a second novel species, T. gilletti, as well as the extension of the host range of Trypanosoma copemani to include koalas. Phylogenetic analysis at the 18S rDNA and gGAPDH loci demonstrated that T. gilletti was genetically distinct with a genetic distance (± s.e.) at the 18S rDNA locus of 2.7 ± 0.5% from T. copemani (wombat). At the gGAPDH locus, the genetic distance (± s.e.) of T. gilletti was 8.7 ± 1.1% from T. copemani (wombat). Trypanosoma gilletti was detected using a nested trypanosome 18S rDNA PCR in 3/139 (∼2%) blood samples and in 2/29 (∼7%) spleen tissue samples from koalas whilst T. irwini was detected in 72/139 (∼52%) blood samples and T. copemani in 4/139 (∼3%) blood samples from koalas. In addition, naturally occurring mixed infections were noted in 2/139 (∼1.5%) of the koalas tested.

  17. Dynamic Modelling under Uncertainty : The Case of Trypanosoma brucei Energy Metabolism

    NARCIS (Netherlands)

    Achcar, Fiona; Kerkhoven, Eduard J.; Bakker, Barbara M.; Barrett, Michael P.; Breitling, Rainer; Papin, Jason A.

    2012-01-01

    Kinetic models of metabolism require detailed knowledge of kinetic parameters. However, due to measurement errors or lack of data this knowledge is often uncertain. The model of glycolysis in the parasitic protozoan Trypanosoma brucei is a particularly well analysed example of a quantitative metabol

  18. Handling Uncertainty in Dynamic Models : The Pentose Phosphate Pathway in Trypanosoma brucei

    NARCIS (Netherlands)

    Kerkhoven, Eduard J.; Achcar, Fiona; Alibu, Vincent P.; Burchmore, Richard J.; Gilbert, Ian H.; Trybilo, Maciej; Driessen, Nicole N.; Gilbert, David; Breitling, Rainer; Bakker, Barbara M.; Barrett, Michael P.

    2013-01-01

    Dynamic models of metabolism can be useful in identifying potential drug targets, especially in unicellular organisms. A model of glycolysis in the causative agent of human African trypanosomiasis, Trypanosoma brucei, has already shown the utility of this approach. Here we add the pentose phosphate

  19. Quantitative enzymatic production of sialylated galactooligosaccharides with an engineered sialidase from Trypanosoma rangeli

    DEFF Research Database (Denmark)

    Zeuner, Birgitte; Holck, Jesper; Perna, Valentina

    2016-01-01

    be produced from κ(kappa)-casein glycomacropeptide (CGMP), a sialylated side stream component from cheese-making, by sialidase-catalyzed transsialylation. Using a rationally designed mutant of the sialidase from Trypanosoma rangeli, Tr13, with enhanced transsialylation activity, six different GOS preparations...

  20. Mixed infection with Trypanoplasma borreli and Trypanosoma carassii induces protection: Involvement of cross-reactive antibodies

    NARCIS (Netherlands)

    Joerink, M.; Groeneveld, A.; Ducro, B.J.; Savelkoul, H.F.J.; Wiegertjes, G.F.

    2007-01-01

    Mixed infections with Trypanoplasma borreli and Trypanosoma carassii in common carp (Cyprinus carpio L.) are commonly found in nature. So far, in the laboratory, only mono-parasitic infections have been examined in more detail. We studied the influence of mixed rather than mono-parasitic infections

  1. The genome sequence of Trypanosoma cruzi, etiologic agent of Chagas disease

    NARCIS (Netherlands)

    El-Sayed, NM; Myler, PJ; Bartholomeu, DC; Nilsson, D; Aggarwal, G; Tran, AN; Ghedin, E; Worthey, EA; Delcher, AL; Blandin, G; Westenberger, SJ; Caler, E; Cerqueira, GC; Branche, C; Haas, B; Anupama, A; Arner, E; Aslund, L; Attipoe, P; Bontempi, E; Bringaud, F; Burton, P; Cadag, E; Campbell, DA; Carrington, M; Crabtree, J; Darban, H; da Silveira, JF; de Jong, P; Edwards, K; Englund, PT; Fazelina, G; Feldblyum, T; Ferella, M; Frasch, AC; Gull, K; Horn, D; Hou, LH; Huang, YT; Kindlund, E; Ktingbeil, M; Kluge, S; Koo, H; Lacerda, D; Levin, MJ; Lorenzi, H; Louie, T; Machado, CR; McCulloch, R; McKenna, A; Mizuno, Y; Mottram, JC; Nelson, S; Ochaya, S; Osoegawa, K; Pai, G; Parsons, M; Pentony, M; Pettersson, U; Pop, M; Ramirez, JL; Rinta, J; Robertson, L; Salzberg, SL; Sanchez, DO; Seyler, A; Sharma, R; Shetty, J; Simpson, AJ; Sisk, E; Tammi, MT; Tarteton, R; Teixeira, S; Van Aken, S; Vogt, C; Ward, PN; Wickstead, B; Wortman, J; White, O; Fraser, CM; Stuart, KD; Andersson, B

    2005-01-01

    Whole-genome sequencing of the protozoan pathogen Trypanosoma cruzi revealed that the diploid genome contains a predicted 22,570 proteins encoded by genes, of which 12,570 represent allelic pairs. Over 50% of the genome consists of repeated sequences, such as retrotransposons and genes for large, fa

  2. The effect of the diterpene 5-epi-icetexone on the cell cycle of Trypanosoma cruzi.

    NARCIS (Netherlands)

    Lozano, E.; Barrera, P.; Tonn, C.; Nieto, M.; Sartor, T.; Sosa, M.A.

    2012-01-01

    Numerous natural compounds have been used against Trypanosoma cruzi, the causative agent of Chagas' disease. Here, we studied the effect of the diterpene 5-epi-icetexone on growth and morphology of parasites synchronized with hydroxyurea, at different periods of time after removal of the nucleotide.

  3. Importation of Hybrid Human-Associated Trypanosoma cruzi Strains of Southern South American Origin, Colombia

    OpenAIRE

    2016-01-01

    We report the characterization of Trypanosoma cruzi of southern South American origin among humans, domestic vectors, and peridomestic hosts in Colombia using high-resolution nuclear and mitochondrial genotyping. Expanding our understanding of the geographic range of lineage TcVI, which is associated with severe Chagas disease, will help clarify risk of human infection for improved disease control.

  4. Molecular Confirmation of Trypanosoma evansi and Babesia bigemina in Cattle from Lower Egypt

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    Mahmoud M. Elhaig, Abdelfattah Selim, Mohamed M. Mahmoud and Eman K El-Gayar

    2016-11-01

    Full Text Available Trypanosomosis and babesiosis are economically important vector-borne diseases for animal health and productivity in developing countries. In Egypt, molecular epidemiological surveys on such diseases are scarce. In the present study, we examined 475 healthy and 25 clinically diagnosed cattle from three provinces in Lower Egypt, for Trypanosoma (T. and Babesia (B. infections using an ITS1 PCR assay that confirmed Trypanosoma species presence and an 18S rRNA assay that detected B. bigemina. Results confirmed Trypanosoma spp. and B. bigemina presence in 30.4% and 11% individuals, respectively, with eight animals (1.6% being co-infected with both hemoparasites. Subsequent type-specific PCRs revealed that all Trypanosoma PCR positive samples corresponded to T. evansi and that none of the animals harboured T. brucei gambiense or T. brucei rhodesiense. Nucleotide sequencing of the variable surface glycoprotein revealed the T. evansi cattle strain to be most closely related (99% nucleotide sequence identity to strains previously detected in dromedary camels in Egypt, while the 18S rRNA gene phylogeny confirmed the presence of a unique B. bigemina haplotype closely related to strains from Turkey and Brazil. Statistically significant differences in PCR prevalence were noted with respect to gender, clinical status and locality. These results confirm the presence of high numbers of carrier animals and signal the need for expanded surveillance and control efforts.

  5. Galactonolactone oxidoreductase from Trypanosoma cruzi employs a FAD cofactor for the synthesis of vitamin C.

    NARCIS (Netherlands)

    Kudryashova, E.V.; Leferink, N.G.H.; Slot, I.G.M.; Berkel, van W.J.H.

    2011-01-01

    Trypanosoma cruzi, the aetiological agent of Chagas' disease, is unable to salvage vitamin C (l-ascorbate) from its environment and relies on de novo synthesis for its survival. Because humans lack the capacity to synthesize ascorbate, the trypanosomal enzymes involved in ascorbate biosynthesis are

  6. Novel molecular mechanism for targeting the parasite Trypanosoma brucei with snake venom toxins

    DEFF Research Database (Denmark)

    Martos Esteban, Andrea; Laustsen, Andreas Hougaard; Carrington, Mark

    Trypanosoma brucei is a parasitic protozoan species capable to infecting insect vectors whose bite further produces African sleeping sickness inhuman beings. During parasites’extracellular lives in the mammalian host, its outer coat, mainly composedof Variable surface glycoproteins (VSGs)[2], und...

  7. Nucleic acid sequence-based amplification with oligochromatography for detection of Trypanosoma brucei in clinical samples

    NARCIS (Netherlands)

    C.M. Mugasa; T. Laurent; G.J. Schoone; P.A. Kager; G.W. Lubega; H.D.F.H. Schallig

    2009-01-01

    Molecular tools, such as real-time nucleic acid sequence-based amplification (NASBA) and PCR, have been developed to detect Trypanosoma brucei parasites in blood for the diagnosis of human African trypanosomiasis (HAT). Despite good sensitivity, these techniques are not implemented in HAT control pr

  8. Molecular detection of Trypanosoma from one-humped camels slaughtered in Najafabad slaughterhouse

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    Samaneh Mehrabiyan

    2014-07-01

    Full Text Available   Introduction: Trypanosomes infect a variety of animals and cause fever, weakness, and lethargy, which lead to weight loss and anemia. In livestock the disease is fatal unless treated and causes serious economic losses. Present study was conducted to detection of Trypanosoma from one-humped camels slaughtered in Najafabad slaughterhouse .   Materials and methods: 278 blood samples were taken at three different time periods from one-humped camels slaughtered in Najafabad slaughterhouse and were tested by polymerase chain reaction (PCR to detection of Trypanosoma .   Results: The overall infection rate of Trypanosoma was 1.07%. Positive samples (3.8% only found in spring and summer, and no positive sample was found in fall and winter .   Discussion and conclusion : Fortunately, this may suggest that the prevalence of Trypanosoma is very low in the region. It is suggested to prevent the disease by preventing livestock suspected of carrying the disease from entering the country. If it is not possible to test imported healthy livestock by molecular assays, it is recommended that simple and rapid tests such as agglutination tests that do not need an expert be assessed .

  9. Two new Trypanosoma species from African birds, with notes on the taxonomy of avian trypanosomes.

    Science.gov (United States)

    Valkiūnas, Gediminas; Iezhova, Tatjana A; Carlson, Jenny S; Sehgal, Ravinder N M

    2011-10-01

    Trypanosoma anguiformis n. sp. and Trypanosoma polygranularis n. sp. are described from the African olive sunbird, Cyanomitra olivacea, and Latham's forest francolin, Francolinus lathami, respectively, based on the morphology of their hematozoic trypomastigotes and partial sequences of the small subunit ribosomal RNA gene. Both new species belong to the group of small non-striated avian trypanosomes (Trypanosoma anguiformis can be readily distinguished from other small avian trypanosomes due to its markedly attenuated (snake-shaped) form of the hematozoic trypomastigotes and the dumbbell-shaped nucleus of the parasite. Trypanosoma polygranularis is readily distinguishable due to the markedly off-center (anteriorly) located nucleus, numerous azurophilic granules that are arranged in a line following the undulating membrane, and the large kinetoplast (with an area up to 1.7 µm(2) [1.1 µm(2) on average]). Illustrations of hematozoic trypomastigotes of the new species are given, and DNA lineages associated with these parasites are reported. The current situation in species taxonomy of avian trypanosomes is discussed. We call for the redescription of valid species of avian trypanosomes from their type vertebrate hosts and type localities by using morphological and polymerase chain reaction-based techniques as an initial essential step towards revising the species composition of avian trypanosomes and reconstructing the taxonomy of these organisms.

  10. Trypanosoma culicavium sp. nov., an avian trypanosome transmitted by Culex mosquitoes.

    Science.gov (United States)

    Votýpka, Jan; Szabová, Jana; Rádrová, Jana; Zídková, Lenka; Svobodová, Milena

    2012-03-01

    A novel avian trypanosome, Trypanosoma culicavium sp. nov., isolated from Culex mosquitoes, is described on the basis of naturally and experimentally infected vectors and bird hosts, localization in the vector, morphological characters and molecular data. This study provides the first comprehensive description of a trypanosome species transmitted by mosquitoes, in which parasites form plugs and rosettes on the stomodeal valve. Trypanosomes occurred as long epimastigotes and short trypomastigotes in vectors and culture and as long trypomastigotes in birds. Transmission of parasites to bird hosts was achieved exclusively by ingestion of experimentally infected Culex mosquito females by canaries (Serinus canaria), but not by Japanese quails (Coturnix japonica), nor by the bite of infected vectors, nor by ingestion of parasites from laboratory cultures. Transmission experiments and the identity of isolates from collared flycatchers (Ficedula albicollis) and Culex mosquitoes suggests that the natural hosts of T. culicavium are insectivorous songbirds (Passeriformes). Phylogenetic analyses of small-subunit rRNA and glycosomal glyceraldehyde-3-phosphate dehydrogenase gene sequences demonstrated that T. culicavium sp. nov. is more related to Trypanosoma corvi than to other avian trypanosomes (e.g. Trypanosoma avium and Trypanosoma bennetti).

  11. Maxi-circles and mini-circles in kinetoplast DNA from Trypanosoma cruzi.

    NARCIS (Netherlands)

    W. Leon; A.C.C. Frasch; J.H.J. Hoeijmakers (Jan); F. Fase-Fowler; P. Borst (Piet); F. Brunel; J. Davison

    1980-01-01

    textabstractMaxi-circles are a minor component of kinetoplast DNAs from all trypanosomatids studied, but they have not previously been found in Trypanosoma cruzi; We have spread intact kinetoplast DNA from the epimastigotes of strain Y in protein monolayers and analysed the mini-circle networks by e

  12. Anti-trypanosomal effect of Peristrophe bicalyculata extract on Trypanosoma brucei brucei-infected rats

    Institute of Scientific and Technical Information of China (English)

    Abdulazeez Mansurah Abimbola; Ibrahim Abdulrazak Baba; Edibo Zakari Yenusa; Sidali Joseph Omanibe; Idris Habeeb Oladimeji

    2013-01-01

    Objective: To investigate the in vitro and in vivo effect of whole plant extracts of Peristrophe bicalyculata on Trypanosoma brucei brucei-infected rats. Methods: The experiment was divided into two phases: In the first phase, the anti-trypanosomal activity of the hot water, cold water, methanol and butanol extracts of the whole plant were determined by incubating with Trypanosoma brucei brucei. The cold water extract was partially-purified and the anti-trypanosomal activity of the fractions determined. In the second phase, Trypanosoma brucei brucei-infected rats were treated with fraction 2c for nine days. Packed cell volume (PCV), high density lipoprotein (HDL), low density lipoprotein (LDL), total cholesterol (TC), triacylglycerol (TAG), aspartate aminotransferase, alanine aminotransferases (ALT), alkaline phosphatase (ALP), total and direct bilirubin levels were determined at the end of the experiment. Results:Cold water extract immobilized 90%of the parasites after 60 min of incubation, and fraction 2c completely immobilized the parasites after 35 min. It significantly increased PCV in Trypanosoma brucei brucei-infected rats. Decreased TC, TAG, HDL and LDL levels of infected rats increased significantly when rats were treated with the fraction, while elevated levels of total bilirubin and ALT also decreased. The difference in urea, direct bilirubin and ALP was not significant when infected rats were compared to rats in other groups. Conclusions:The ability of the plant to ameliorate the infection-induced biochemical changes calls for detailed investigation of the potentials of the plant for antitrypanosomiasis drug delivery.

  13. Comprehensive proteomic analysis of Trypanosoma cruzi epimastigote cell surface proteins by two complementary methods

    DEFF Research Database (Denmark)

    Queiroz, Rayner M L; Charneau, Sébastien; Motta, Flávia N;

    2013-01-01

    Trypanosoma cruzi is a protozoan that causes Chagas' disease, a neglected infectious illness that affects millions of people, mostly in Latin America. Here, the cell surface subproteome of the T. cruzi epimastigote life form was characterized. In order to prepare samples enriched in epimastigote...

  14. Molecular characterization of Trypanosoma cruzi Mexican strains and their behavior in the mouse experimental model

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    César Gómez-Hernández

    2011-12-01

    Full Text Available INTRODUCTION: For a long time, the importance of Chagas disease in Mexico, where many regarded it as an exotic malady, was questioned. Considering the great genetic diversity among isolates of Trypanosoma cruzi, the importance of this biological characterization, and the paucity of information on the clinical and biological aspects of Chagas disease in Mexico, this study aimed to identify the molecular and biological characterization of Trypanosoma cruzi isolates from different endemic areas of this country, especially of the State of Jalisco. METHODS: Eight Mexican Trypanosoma cruzi strains were biologically and genetically characterized (PCR specific for Trypanosoma cruzi, multiplex-PCR, amplification of space no transcript of the genes of the mini-exon, amplification of polymorphic regions of the mini-exon, classification by amplification of intergenic regions of the spliced leader genes, RAPD - (random amplified polymorphic DNA. RESULTS: Two profiles of parasitaemia were observed, patent (peak parasitaemia of 4.6×10(6 to 10(7 parasites/mL and subpatent. In addition, all isolates were able to infect 100% of the animals. The isolates mainly displayed tropism for striated (cardiac and skeletal muscle. PCR amplification of the mini-exon gene classified the eight strains as TcI. The RAPD technique revealed intraspecies variation among isolates, distinguishing strains isolated from humans and triatomines and according to geographic origin. CONCLUSIONS: The Mexican T. cruzi strains are myotrophic and belong to group TcI.

  15. The major transcripts of the kinetoplast Trypanosoma brucei are very small ribosomal RNA's.

    NARCIS (Netherlands)

    I.C. Eperon; J.W.G. Janssen; J.H.J. Hoeijmakers (Jan); P. Borst (Piet)

    1983-01-01

    textabstractThe nucleotide sequence has been determined of a 2.2 kb segment of kinetoplast DNA, which encodes the major mitochondrial transcripts (12S and 9S) of Trypanosoma brucei. The sequence shows that the 12S RNA is a large subunit rRNA, although sufficiently unusual for resistance to chloramph

  16. Kinetic properties and inhibition of Trypanosoma cruzi 3-hydroxy-3-methylglutaryl CoA reductase

    DEFF Research Database (Denmark)

    Hurtado-Guerrrero, Ramón; Pena Diaz, Javier; Montalvetti, Andrea;

    2002-01-01

    A detailed kinetic analysis of the recombinant soluble enzyme 3-hydroxy-3-methylglutaryl CoA reductase (HMGR) from Trypanosoma cruzi has been performed. The enzyme catalyzes the normal anabolic reaction and the reductant is NADPH. It also catalyzes the oxidation of mevalonate but at a lower propo...

  17. 1,4-Diamino-2-butanone, a putrescine analogue, promotes redox imbalance in Trypanosoma cruzi and mammalian cells.

    Science.gov (United States)

    Soares, Chrislaine O; Colli, Walter; Bechara, Etelvino J H; Alves, Maria Julia M

    2012-12-15

    The putrescine analogue 1,4-diamino-2-butanone (DAB) is highly toxic to various microorganisms, including Trypanosoma cruzi. Similar to other α-aminocarbonyl metabolites, DAB exhibits pro-oxidant properties. DAB undergoes metal-catalyzed oxidation yielding H(2)O(2), NH(4)(+) ion, and a highly toxic α-oxoaldehyde. In vitro, DAB decreases mammalian cell viability associated with changes in redox balance. Here, we aim to clarify the DAB pro-oxidant effects on trypomastigotes and on intracellular T. cruzi amastigotes. DAB (0.05-5 mM) exposure in trypomastigotes, the infective stage of T. cruzi, leads to a decline in parasite viability (IC(50)c.a. 0.2 mM DAB; 4 h incubation), changes in morphology, thiol redox imbalance, and increased TcSOD activity. Medium supplementation with catalase (2.5 μM) protects trypomastigotes against DAB toxicity, while host cell invasion by trypomastigotes is hampered by DAB. Additionally, intracellular amastigotes are susceptible to DAB toxicity. Furthermore, pre-treatment with 100-500 μM buthionine sulfoximine (BSO) of LLC-MK2 potentiates DAB cytotoxicity, whereas 5 mM N-acetyl-cysteine (NAC) protects cells from oxidative stress. Together, these data support the hypothesis that redox imbalance contributes to DAB cytotoxicity in both T. cruzi and mammalian host cells.

  18. Clinical forms of Trypanosoma cruzi infected individuals in the chronic phase of Chagas disease in Puebla, Mexico

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    María del Carmen Sánchez-Guillén

    2006-11-01

    Full Text Available In Mexico, despite the relatively high seroprevalence of Trypanosoma cruzi infection in humans in some areas, reported morbidity of Chagas disease is not clear. We determined clinical stage in 71 individuals seropositive to T. cruzi in the state of Puebla, Mexico, an area endemic for Chagas disease with a reported seroprevalence of 7.7%. Diagnosis of Chagas disease was made by two standardized serological tests (ELISA, IHA. Individuals were stratified according to clinical studies. All patients were submitted to EKG, barium swallow, and barium enema. Groups were identified as indeterminate form (IF asymptomatic individuals without evidence of abnormalities (n = 34 cases; those with gastrointestinal alterations (12 patients including symptoms of abnormal relaxation of the lower esophageal sphincter and absent peristalsis in the esophageal body, grade I megaesophagus, and/or megacolon; patients with clinical manifestations and documented changes of chronic Chagas heart disease who were subdivided as follows: mild (8 patients - mild electrocardiographic changes of ventricular repolarization, sinus bradychardia; moderate (6 patients - left bundle branch block, right bundle branch block associated with left anterior fascicular block; severe (8 patients - signs of cardiomegaly, dilated cardiomyopathy; and the associated form (3 cases that included presence of both cardiomyopathy and megaesophagus. These data highlight the importance of accurate evaluation of the prevalence and clinical course of Chagas disease in endemic and non-endemic areas of Mexico.

  19. Clinical forms of Trypanosoma cruzi infected individuals in the chronic phase of Chagas disease in Puebla, Mexico.

    Science.gov (United States)

    Sánchez-Guillén, María Del Carmen; López-Colombo, Aurelio; Ordóñez-Toquero, Guillermo; Gomez-Albino, Isidoro; Ramos-Jimenez, Judith; Torres-Rasgado, Enrique; Salgado-Rosas, Hilda; Romero-Díaz, Mónica; Pulido-Pérez, Patricia; Pérez-Fuentes, Ricardo

    2006-11-01

    In Mexico, despite the relatively high seroprevalence of Trypanosoma cruzi infection in humans in some areas, reported morbidity of Chagas disease is not clear. We determined clinical stage in 71 individuals seropositive to T. cruzi in the state of Puebla, Mexico, an area endemic for Chagas disease with a reported seroprevalence of 7.7%. Diagnosis of Chagas disease was made by two standardized serological tests (ELISA, IHA). Individuals were stratified according to clinical studies. All patients were submitted to EKG, barium swallow, and barium enema. Groups were identified as indeterminate form (IF) asymptomatic individuals without evidence of abnormalities (n = 34 cases); those with gastrointestinal alterations (12 patients) including symptoms of abnormal relaxation of the lower esophageal sphincter and absent peristalsis in the esophageal body, grade I megaesophagus, and/or megacolon; patients with clinical manifestations and documented changes of chronic Chagas heart disease who were subdivided as follows: mild (8 patients)--mild electrocardiographic changes of ventricular repolarization, sinus bradychardia); moderate (6 patients)--left bundle branch block, right bundle branch block associated with left anterior fascicular block); severe (8 patients)--signs of cardiomegaly, dilated cardiomyopathy); and the associated form (3 cases) that included presence of both cardiomyopathy and megaesophagus. These data highlight the importance of accurate evaluation of the prevalence and clinical course of Chagas disease in endemic and non-endemic areas of Mexico.

  20. The F(0F(1-ATP synthase complex contains novel subunits and is essential for procyclic Trypanosoma brucei.

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    Alena Zíková

    2009-05-01

    Full Text Available The mitochondrial F(0F(1 ATP synthase is an essential multi-subunit protein complex in the vast majority of eukaryotes but little is known about its composition and role in Trypanosoma brucei, an early diverged eukaryotic pathogen. We purified the F(0F(1 ATP synthase by a combination of affinity purification, immunoprecipitation and blue-native gel electrophoresis and characterized its composition and function. We identified 22 proteins of which five are related to F(1 subunits, three to F(0 subunits, and 14 which have no obvious homology to proteins outside the kinetoplastids. RNAi silencing of expression of the F(1 alpha subunit or either of the two novel proteins showed that they are each essential for the viability of procyclic (insect stage cells and are important for the structural integrity of the F(0F(1-ATP synthase complex. We also observed a dramatic decrease in ATP production by oxidative phosphorylation after silencing expression of each of these proteins while substrate phosphorylation was not severely affected. Our procyclic T. brucei cells were sensitive to the ATP synthase inhibitor oligomycin even in the presence of glucose contrary to earlier reports. Hence, the two novel proteins appear essential for the structural organization of the functional complex and regulation of mitochondrial energy generation in these organisms is more complicated than previously thought.

  1. Antisense Oligonucleotides Targeting Parasite Inositol 1,4,5-Trisphosphate Receptor Inhibits Mammalian Host Cell Invasion by Trypanosoma cruzi

    Science.gov (United States)

    Hashimoto, Muneaki; Nara, Takeshi; Hirawake, Hiroko; Morales, Jorge; Enomoto, Masahiro; Mikoshiba, Katsuhiko

    2014-02-01

    Chagas disease is caused by an intracellular parasitic protist, Trypanosoma cruzi. As there are no highly effective drugs against this agent that also demonstrate low toxicity, there is an urgent need for development of new drugs to treat Chagas disease. We have previously demonstrated that the parasite inositol 1,4,5-trisphosphate receptor (TcIP3R) is crucial for invasion of the mammalian host cell by T. cruzi. Here, we report that TcIP3R is a short-lived protein and that its expression is significantly suppressed in trypomastigotes. Treatment of trypomastigotes, an infective stage of T. cruzi, with antisense oligonucleotides specific to TcIP3R deceased TcIP3R protein levels and impaired trypomastigote invasion of host cells. Due to the resulting instability and very low expression level of TcIP3R in trypomastigotes indicates that TcIP3R is a promising target for antisense therapy in Chagas disease.

  2. Incidence of colonization and bloodstream infection with carbapenem-resistant Enterobacteriaceae in children receiving antineoplastic chemotherapy in Italy.

    Science.gov (United States)

    Caselli, Desiree; Cesaro, Simone; Fagioli, Franca; Carraro, Francesca; Ziino, Ottavio; Zanazzo, Giulio; Meazza, Cristina; Colombini, Antonella; Castagnola, Elio

    2016-02-01

    Few data are available on the incidence of carbapenemase-producing Enterobacteriaceae (CPE) infection or colonization in children receiving anticancer chemotherapy. We performed a nationwide survey among centers participating in the pediatric hematology-oncology cooperative study group (Associazione Italiana Ematologia Oncologia Pediatrica, AIEOP). During a 2-year observation period, we observed a threefold increase in the colonization rate, and a fourfold increase of bloodstream infection episodes, caused by CPE, with a 90-day mortality of 14%. This first nationwide Italian pediatric survey shows that the circulation of CPE strains in the pediatric hematology-oncology environment is increasing. Given the mortality rate, which is higher than for other bacterial strains, specific monitoring should be applied and the results should have implications for health-care practice in pediatric hematology-oncology.

  3. SURVEILLANCE OF TRYPANOSOMA SPP OF RODENTS AND STUDIES IN THEIR TRANSMISSION PROBABILITY BY FLEAS IN SOME RURAL EGYPTIAN AREAS.

    Science.gov (United States)

    Dahesh, Salwa M A; Mikhail, Micheal W

    2016-04-01

    A new public health problem arises from animal trypanosomes that afflict human by a disease called atypical human trypanosomiasis. Although humans have an innate protection against most Trypanosoma species, nineteen cases of atypical human trypanosomiasis caused by the animal trypanosome as T. b. brucei, T. vivax, T. congolense, T. evansi and T. lewisi have been recorded. Some of theserecorded cases were transient, six required trypanocidal treatments however two patients died. Rodent trypanosome, T. lewisi is transmitted via ingestion of fleas or their feces containing the infective stage, the metacyclic trypomastigote. Because of the high densities of various species of rodents and their distribution all over the country especially in rural areas, the present work aimed to evaluate the trypanosomiasis among rodents collected from November to March 2016 and study transmission probability by their fleas in some rural areas in Abu Alnomros Center, Giza. The overall trypanosomiasis prevalence among the different rodent species was (21 rats) 24.7%. All the infected rats belonged to Rattus r. spp where the prevalence of infection with Trypanosoma lewisi among that species was very high 51.2% while none of rats belonged to Rattus norvegicus were infected. That may be attributed to the solid immunity gained by the R. norvegicus where most of the collected norvegicus were aged and weighed more than 200 grams. There was an inverse significant correlation between the densities of parasites and the weights of the losts. The rat which recorded the highest parasite density (60,000 parasites/microliter) was a female Rattus r. captured indoor (inside house). As to sex of Rattus rattus spp no significant difference was found between males and females in trypanosomiasis. Also there was no significant correlation between the densities of parasites and the number of white blood cells among Rattus rattus spp. All positive rats were collected indoors (from houses) and all the rats

  4. Ten-year study of species distribution and antifungal susceptibilities of Candida bloodstream isolates at a Brazilian tertiary hospital.

    Science.gov (United States)

    Bonfietti, L X; Szeszs, M W; Chang, M R; Martins, M A; Pukinskas, S R B S; Nunes, M O; Pereira, G H; Paniago, A M M; Purisco, S U; Melhem, M S C

    2012-12-01

    To describe the incidence and susceptibility profile of Candida bloodstream infections in a tertiary-care hospital, we performed a retrospective observational study from 1998 to 2007. Comorbidities and risk factors were compiled from all cases. In vitro susceptibility testing to fluconazole, itraconazole, voriconazole, and amphotericin B was performed for 100 isolates, and caspofungin was tested for C. parapsilosis complex. In a ten-year evaluation of candidemias, 44 % were caused by C. albicans, and species of the C. parapsilosis complex were the second most frequent agents (37 %). Other species presented lower incidences (C. tropicalis, 13 %, C. glabrata, 5 %, and C. krusei, 1 %). Neither C. dubliniensis nor C. metapsilosis were observed in this study. C. orthopsilosis (3 %) and C. parapsilosis stricto sensu (34 %) were also found. Species distribution was independent of catheterization, mechanical ventilation, or previous use of antifungals or corticoids. Parenteral nutrition administration was strongly related to C. glabrata infection, and the highest mortality (80 %) was observed in patients infected by this species. All C. albicans isolates showed high susceptibility to all tested drugs. However, two C. parapsilosis stricto sensu isolates presented high minimum inhibitory concentration (MIC) (4 mg/L each) to fluconazole, and one exhibited voriconazole MIC of 0.25 mg/L, highlighting the cross-resistance to these azoles. All isolates of C. tropicalis and C. glabrata showed no resistance to any drug tested. No difference was noted between C. parapsilosis and C. orthopsilosis susceptibilities to caspofungin. Our results suggest that resistance to amphotericin B, fluconazole, voriconazole, itraconazole, and caspofungin in Brazilian Candida bloodstream isolates is still uncommon.

  5. Antimicrobial Resistance and Molecular Epidemiology of Escherichia coli Causing Bloodstream Infections in Three Hospitals in Shanghai, China.

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    Su Wang

    Full Text Available Escherichia coli (E. coli is one of the most frequent and lethal causes of bloodstream infections (BSIs. We carried out a retrospective multicenter study on antimicrobial resistance and phylogenetic background of clinical E. coli isolates recovered from bloodstream in three hospitals in Shanghai. E. coli isolates causing BSIs were consecutively collected between Sept 2013 and Sept 2014. Ninety isolates randomly selected (30 from each hospital were enrolled in the study. Antimicrobial susceptibility testing was performed by disk diffusion. PCR was used to detect antimicrobial resistance genes coding for β-lactamases (TEM, CTX-M, OXA, etc., carbapenemases (IMP, VIM, KPC, NDM-1 and OXA-48, and phylogenetic groups. eBURST was applied for analysis of multi-locus sequence typing (MLST. The resistance rates for penicillins, second-generation cephalosporins, fluoroquinolone and tetracyclines were high (>60%. Sixty-one of the 90 (67.8% strains enrolled produced ESBLs and no carbapenemases were found. Molecular analysis showed that CTX-M-15 (25/61, CTX-M-14 (18/61 and CTX-M-55 (9/61 were the most common ESBLs. Phylogenetic group B2 predominated (43.3% and exhibited the highest rates of ESBLs production. ST131 (20/90 was the most common sequence type and almost assigned to phylogenetic group B2 (19/20. The following sequence types were ST405 (8/90 and ST69 (5/90. Among 61 ESBL-producers isolates, B2 (26, 42.6% and ST131 (18, 29.5% were also the most common phylogenetic group and sequence type. Genetic diversity showed no evidence suggesting a spread of these antimicrobial resistant isolates in the three hospitals. In order to provide more comprehensive and reliable epidemiological information for preventing further dissemination, well-designed and continuous surveillance with more hospitals participating was important.

  6. Antimicrobial Resistance and Molecular Epidemiology of Escherichia coli Causing Bloodstream Infections in Three Hospitals in Shanghai, China.

    Science.gov (United States)

    Wang, Su; Zhao, Sheng-Yuan; Xiao, Shu-Zhen; Gu, Fei-Fei; Liu, Qing-Zhong; Tang, Jin; Guo, Xiao-Kui; Ni, Yu-Xing; Han, Li-Zhong

    2016-01-01

    Escherichia coli (E. coli) is one of the most frequent and lethal causes of bloodstream infections (BSIs). We carried out a retrospective multicenter study on antimicrobial resistance and phylogenetic background of clinical E. coli isolates recovered from bloodstream in three hospitals in Shanghai. E. coli isolates causing BSIs were consecutively collected between Sept 2013 and Sept 2014. Ninety isolates randomly selected (30 from each hospital) were enrolled in the study. Antimicrobial susceptibility testing was performed by disk diffusion. PCR was used to detect antimicrobial resistance genes coding for β-lactamases (TEM, CTX-M, OXA, etc.), carbapenemases (IMP, VIM, KPC, NDM-1 and OXA-48), and phylogenetic groups. eBURST was applied for analysis of multi-locus sequence typing (MLST). The resistance rates for penicillins, second-generation cephalosporins, fluoroquinolone and tetracyclines were high (>60%). Sixty-one of the 90 (67.8%) strains enrolled produced ESBLs and no carbapenemases were found. Molecular analysis showed that CTX-M-15 (25/61), CTX-M-14 (18/61) and CTX-M-55 (9/61) were the most common ESBLs. Phylogenetic group B2 predominated (43.3%) and exhibited the highest rates of ESBLs production. ST131 (20/90) was the most common sequence type and almost assigned to phylogenetic group B2 (19/20). The following sequence types were ST405 (8/90) and ST69 (5/90). Among 61 ESBL-producers isolates, B2 (26, 42.6%) and ST131 (18, 29.5%) were also the most common phylogenetic group and sequence type. Genetic diversity showed no evidence suggesting a spread of these antimicrobial resistant isolates in the three hospitals. In order to provide more comprehensive and reliable epidemiological information for preventing further dissemination, well-designed and continuous surveillance with more hospitals participating was important.

  7. Surveillance of Candida spp bloodstream infections: epidemiological trends and risk factors of death in two Mexican tertiary care hospitals.

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    Dora E Corzo-Leon

    Full Text Available INTRODUCTION: Larger populations at risk, broader use of antibiotics and longer hospital stays have impacted on the incidence of Candida sp. bloodstream infections (CBSI. OBJECTIVE: To determine clinical and epidemiologic characteristics of patients with CBSI in two tertiary care reference medical institutions in Mexico City. DESIGN: Prospective and observational laboratory-based surveillance study conducted from 07/2008 to 06/2010. METHODS: All patients with CBSI were included. Identification and antifungal susceptibility were performed using CLSI M27-A3 standard procedures. Frequencies, Mann-Whitney U test or T test were used as needed. Risk factors were determined with multivariable analysis and binary logistic regression analysis. RESULTS: CBSI represented 3.8% of nosocomial bloodstream infections. Cumulative incidence was 2.8 per 1000 discharges (incidence rate: 0.38 per 1000 patient-days. C. albicans was the predominant species (46%, followed by C. tropicalis (26%. C. glabrata was isolated from patients with diabetes (50%, and elderly patients. Sixty-four patients (86% received antifungals. Amphotericin-B deoxycholate (AmBD was the most commonly used agent (66%. Overall mortality rate reached 46%, and risk factors for death were APACHE II score ≥ 16 (OR = 6.94, CI95% = 2.34-20.58, p<0.0001, and liver disease (OR = 186.11, CI95% = 7.61-4550.20, p = 0.001. Full susceptibility to fluconazole, AmBD and echinocandins among C. albicans, C. tropicalis, and C. parapsilosis was observed. CONCLUSIONS: The cumulative incidence rate in these centers was higher than other reports from tertiary care hospitals from Latin America. Knowledge of local epidemiologic patterns permits the design of more specific strategies for prevention and preemptive therapy of CBSI.

  8. Surveillance of Candida spp Bloodstream Infections: Epidemiological Trends and Risk Factors of Death in Two Mexican Tertiary Care Hospitals

    Science.gov (United States)

    Corzo-Leon, Dora E.; Alvarado-Matute, Tito; Colombo, Arnaldo L.; Cornejo-Juarez, Patricia; Cortes, Jorge; Echevarria, Juan I.; Guzman-Blanco, Manuel; Macias, Alejandro E.; Nucci, Marcio; Ostrosky-Zeichner, Luis; Ponce-de-Leon, Alfredo; Queiroz-Telles, Flavio; Santolaya, Maria E.; Thompson-Moya, Luis; Tiraboschi, Iris N.; Zurita, Jeannete; Sifuentes-Osornio, Jose

    2014-01-01

    Introduction Larger populations at risk, broader use of antibiotics and longer hospital stays have impacted on the incidence of Candida sp. bloodstream infections (CBSI). Objective To determine clinical and epidemiologic characteristics of patients with CBSI in two tertiary care reference medical institutions in Mexico City. Design Prospective and observational laboratory-based surveillance study conducted from 07/2008 to 06/2010. Methods All patients with CBSI were included. Identification and antifungal susceptibility were performed using CLSI M27-A3 standard procedures. Frequencies, Mann-Whitney U test or T test were used as needed. Risk factors were determined with multivariable analysis and binary logistic regression analysis. Results CBSI represented 3.8% of nosocomial bloodstream infections. Cumulative incidence was 2.8 per 1000 discharges (incidence rate: 0.38 per 1000 patient-days). C. albicans was the predominant species (46%), followed by C. tropicalis (26%). C. glabrata was isolated from patients with diabetes (50%), and elderly patients. Sixty-four patients (86%) received antifungals. Amphotericin-B deoxycholate (AmBD) was the most commonly used agent (66%). Overall mortality rate reached 46%, and risk factors for death were APACHE II score ≥16 (OR = 6.94, CI95% = 2.34–20.58, p<0.0001), and liver disease (OR = 186.11, CI95% = 7.61–4550.20, p = 0.001). Full susceptibility to fluconazole, AmBD and echinocandins among C. albicans, C. tropicalis, and C. parapsilosis was observed. Conclusions The cumulative incidence rate in these centers was higher than other reports from tertiary care hospitals from Latin America. Knowledge of local epidemiologic patterns permits the design of more specific strategies for prevention and preemptive therapy of CBSI. PMID:24830654

  9. High prevalence of tuberculosis and serious bloodstream infections in ambulatory individuals presenting for antiretroviral therapy in Malawi.

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    Richard A Bedell

    Full Text Available BACKGROUND: Tuberculosis (TB and serious bloodstream infections (BSI may contribute to the high early mortality observed among patients qualifying for antiretroviral therapy (ART with unexplained weight loss, chronic fever or chronic diarrhea. METHODS AND FINDINGS: A prospective cohort study determined the prevalence of undiagnosed TB or BSI among ambulatory HIV-infected adults with unexplained weight loss and/or chronic fever, or diarrhea in two routine program settings in Malawi. Subjects with positive expectorated sputum smears for AFB were excluded. Investigations Bacterial and mycobacterial blood cultures, cryptococcal antigen test (CrAg, induced sputum (IS for TB microscopy and solid culture, full blood count and CD4 lymphocyte count. Among 469 subjects, 52 (11% had microbiological evidence of TB; 50 (11% had a positive (non-TB blood culture and/or positive CrAg. Sixty-five additional TB cases were diagnosed on clinical and radiological grounds. Nontyphoidal Salmonellae (NTS were the most common blood culture pathogens (29 cases; 6% of participants and 52% of bloodstream isolates. Multivariate analysis of baseline clinical and hematological characteristics found significant independent associations between oral candidiasis or lymphadenopathy and TB, marked CD4 lymphopenia and NTS infection, and severe anemia and either infection, but low positive likelihood ratios (<2 for all combinations. CONCLUSIONS: We observed a high prevalence of TB and serious BSI, particularly NTS, in a program cohort of chronically ill HIV-infected outpatients. Baseline clinical and hematological characteristics were inadequate predictors of infection. HIV clinics need better rapid screening tools for TB and BSI. Clinical trials to evaluate empiric TB or NTS treatment are required in similar populations.

  10. Regional variations in fluoroquinolone non-susceptibility among Escherichia coli bloodstream infections within the Veterans Healthcare Administration

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    Daniel J. Livorsi

    2016-10-01

    Full Text Available Abstract Objectives We sought to define regional variations in fluoroquinolone non-susceptibility (FQ-NS among bloodstream isolates of Escherichia coli across the Veterans Health Administration (VHA in the United States. Methods We analyzed a retrospective cohort of patients managed at 136 VHA hospitals who had a blood culture positive for E.coli between 2003 and 2013. Hospitals were classified based on US Census Divisions, and regional variations in FQ-NS were analyzed. Results Twenty-four thousand five hundred twenty-three unique E.coli bloodstream infections (BSIs were identified between 2003 and 2013. 53.9 % of these were community-acquired, 30.7 % were healthcare-associated, and 15.4 % were hospital-onset BSIs. The proportion of E.coli BSIs with FQ-NS significantly varied across US Census Divisions (p < 0.001. During 2003–2013, the proportion of E.coli BSIs with FQ-NS was highest in the West South-Central Division (32.7 % and lowest in the Mountain Division (20.0 %. Multivariable analysis showed that there were universal secular trends towards higher FQ-NS rates (p < 0.001 with significant variability of slopes across US Census Divisions (p < 0.001. Conclusion There has been a universal increase in FQ-NS among E.coli BSIs within VHA, but the rate of increase has significantly varied across Census Divisions. The reasons for this variability are unclear. These findings reinforce the importance of using local data to develop and update local antibiograms and antibiotic-prescribing guidelines.

  11. Does antimicrobial use density at the ward level influence monthly central line-associated bloodstream infection rates?

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    Yoshida J

    2014-12-01

    Full Text Available Junichi Yoshida, Yukiko Harada, Tetsuya Kikuchi, Ikuyo Asano, Takako Ueno, Nobuo Matsubara Infection Control Committee, Shimonoseki City Hospital, Shimonoseki, Japan Abstract: The aim of this study was to elucidate risk factors, including ward antimicrobial use density (AUD, for central line-associated bloodstream infection (CLABSI as defined by the Centers for Disease Control and Prevention in a 430-bed community hospital using central venous lines with closed-hub systems. We calculated AUD as (total dose/(defined daily dose × patient days ×1,000 for a total of 20 drugs, nine wards, and 24 months. Into each line day data, we inputed AUD and device utilization ratios, number of central line days, and CLABSI. The ratio of susceptible strains in isolates were subjected to correlation analysis with AUD. Of a total of 9,997 line days over 24 months, CLABSI was present in 33 cases (3.3 ‰, 14 (42.4% of which were on surgical wards out of nine wards. Of a total of 43 strains isolated, eight (18.6% were methicillin-resistant Staphylococcus aureus (MRSA; none of the MRSA-positive patients had received cefotiam before the onset of infection. Receiver-operating characteristic analysis showed that central line day 7 had the highest accuracy. Logistic regression analysis showed the central line day showed an odds ratio of 5.511 with a 95% confidence interval of 1.936–15.690 as did AUD of cefotiam showing an odds ratio of 0.220 with 95% confidence interval of 0.00527–0.922 (P=0.038. Susceptible strains ratio and AUD showed a negative correlation (R2=0.1897. Thus, CLABSI could be prevented by making the number of central line days as short as possible. The preventative role of AUD remains to be investigated. Keywords: bloodstream infection, central line, antimicrobial use density

  12. Clinical and Microbiological Characteristics of Heteroresistant and Vancomycin-Intermediate Staphylococcus aureus from Bloodstream Infections in a Brazilian Teaching Hospital

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    da Costa, Thaina Miranda; Morgado, Priscylla Guimarães Migueres; Cavalcante, Fernanda Sampaio; Damasco, Andreia Paredes; Nouér, Simone Aranha; dos Santos, Kátia Regina Netto

    2016-01-01

    This study analyzed clinical and microbiological characteristics of heteroresistant (hVISA) and vancomycin-intermediate Staphylococcus aureus (VISA) from bloodstream infections (BSI) in a Brazilian teaching hospital, between 2011 and 2013. Minimum inhibitory concentrations (MIC) of antimicrobials were determined by broth microdilution method and SCCmec was detected by PCR. Isolates with a vancomycin MIC ≥ 2mg/L were cultured on BHI agar with 3, 4 or 6 mg/L (BHIa3, BHIa4 or BHIa6) of vancomycin and BHIa4 with casein (BHIa4ca). Macromethod Etest® and Etest® Glicopeptides Resistance Detection were also used. VISA and hVISA isolates were confirmed by the population analysis profile then typed by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing. Medical data from the patients were obtained from their medical records. Among 110 consecutive isolates, 31 (28%) were MRSA and carried the SCCmec type II (15 isolates) or IV (16 isolates). Vancomycin MIC50 and MIC90 were 1 and 2 mg/L, respectively. MRSA isolates had increased non-susceptibility to daptomycin (p = 0.0003). Six (5%) isolates were VISA, four of which were MRSA, three SCCmec type II/USA100/ST5 and one type IV/USA800/ST3192. One MRSA SCCmec II isolate grew on agar BHIa3, BHIa4 and BHIa4ca, and it was confirmed as hVISA. Among the six VISA isolates, five (83%) grew on BHIa3 and three (50%) on BHI4ca. Four of the six VISA isolates and the one hVISA isolate were from patients who had undergone dialysis. Thus, a possible dissemination of the SCCmec II/USA100/ST5 lineage may have occurred in the hospital comprising the VISA, hVISA and daptomycin non-susceptible S. aureus Brazilian isolates from health care associated bloodstream infections. PMID:27575698

  13. Epidemiological characterization of Acinetobacter baumannii bloodstream isolates from a Chinese Burn Institute: A three-year study.

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    Huang, Guangtao; Yin, Supeng; Xiang, Lijuan; Gong, Yali; Sun, Kedai; Luo, Xiaoqiang; Zhang, Cheng; Yang, Zichen; Deng, Liuyang; Jiang, Bei; Jin, Shouguang; Chen, Jing; Peng, Yizhi

    2016-11-01

    Acinetobacter baumannii infection is a serious threat to burn patients. Bacteremia due to A. baumannii is becoming the most common cause of mortality following burn. However, the epidemiology of A. baumannii causing burn-related bloodstream infections has rarely been reported. We retrospectively collected 81 A. baumannii isolates from the bloodstream of burn patients over a three-year period. Antibiotic susceptibility tests, the prevalence of antibiotic-resistant genes and sequence typing (ST) were conducted to characterize these strains. Most of the isolates showed an extensive drug-resistant phenotype. The resistance frequencies to imipenem and meropenem were 94% and 91%, respectively. The blaOXA-23-like gene, AmpC, IS-AmpC, PER and SIM are the five most prevalent resistant genes, and their prevalence rates are 93% (75/81), 86% (70/81), 73% (59/81), 73% (59/81) and 52% (42/81), respectively. The 81 isolates were grouped into 10 known and 18 unknown ST types, with ST368 (38%) being the most prevalent. Except for ST457 and four new types (STn2, STn6, STn11 and STn14), the remaining 23 ST types belonged to one clonal complex 92, which is most common among clinical isolate in China. The above results indicated that ST368 isolates possessing both the blaOXA-23-like gene and ampC gene were the main culprits of the increasing nosocomial A. baumannii infection in this study. More attention should be paid to monitoring the molecular epidemiology of A. baumannii isolates from burn patients to prevent further distribution. Such information may help clinicians with therapeutic decisions and infection control in the Burns Institute.

  14. Ageing is not associated with an altered immune response during Trypanosoma cruzi infection: Ageing and Trypanosoma cruzi infection.

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    Colato, Rafaela Pravato; Brazão, Vânia; Santello, Fabricia Helena; Toldo, Míriam Paula Alonso; do Vale, Gabriel Tavares; Tirapelli, Carlos Renato; Pereira-da-Silva, Gabriela; do Prado, José Clóvis

    2017-01-25

    The aims of this work were to evaluate the influence of ageing on the magnitude of the immune response in male Wistar rats infected with the Y strain of Trypanosoma cruzi (T. cruzi). Infected young animals displayed enhanced CD4(+) T cells as compared to uninfected counterparts. Ageing also triggered a significant reduction in CD8(+) T cells compared to young and uninfected groups. The percentage of spleen NKT cells was reduced for all groups, regardless of the infection status. Significant decreased B-cells was noted in aged controls and infected animals as compared to young counterparts. A significant decrease in MHC class II (RT1B) expression in all aged animals was observed, whether infected or not. The highest and significant levels of Thiobarbituric Acid Reactive Substances (TBARS) were noted in the aged and infected animals as compared to young-infected ones (16day). Consequently superoxide dismutase (SOD) activity was reduced for both control and infected aged animals. Significant elevation of 8-isoprostane levels was found in aged control and infected animals. Plasma glutathione (GSH) concentration was reduced in aged control animals, as well as, in the young infected animals. NO production was increased in both infected and uninfected aged animals compared to young infected and uninfected animals. Corticosterone levels were elevated in aged animals, whether infected or not. Thus, our results are inedited since the immune response is not worsened by the simple fact of animals being older. Ageing by itself triggered a damaged immune response as well as enhanced reactive oxygen species, when compared to young counterparts, but it did not contribute to impair the immune response of T. cruzi infected and aged rats.

  15. Comparative pathogenicity in Swiss mice of Trypanosoma cruzi IV from northern Brazil and Trypanosoma cruzi II from southern Brazil.

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    Meza, Sheila Karina Lüders; Kaneshima, Edilson Nobuyoshi; Silva, Sueli de Oliveira; Gabriel, Maristela; de Araújo, Silvana Marques; Gomes, Mônica Lúcia; Monteiro, Wuelton Marcelo; Barbosa, Maria das Graças Vale; Toledo, Max Jean de Ornelas

    2014-11-01

    The geographical heterogeneity of Chagas disease (ChD) is mainly caused by genetic variability of the etiological agent Trypanosoma cruzi. Our hypothesis was that the pathogenicity for mice may vary with the genetic lineage (or Discrete Typing Unit - DTU) of the parasite. To test this hypothesis, parasitological and histopathological evaluations were performed in mice inoculated with strains belonging to the DTU T. cruzi IV (TcIV) from the State of Amazonas (northern Brazil), or the DTU T. cruzi II (TcII) from the State of Paraná (southern Brazil). Groups of 10 Swiss mice were inoculated with eight strains of TcIV obtained from acute cases (7) from two outbreaks of orally acquired ChD, and from the triatomine Rhodnius robustus (1) from Amazonas; and three strains of TcII obtained from chronic patients in Paraná. We evaluated the pre-patent period, patent period, maximum peak of parasitemia, day of maximum peak of parasitemia, area under the parasitemia curve, inflammatory process, and tissue parasitism in the acute phase. TcIV was less virulent than TcII, and showed significantly (p < 0.005) lower parasitemia levels. Although the levels of tissue parasitism did not differ statistically, mice infected with TcIV displayed significantly (p < 0.001) fewer inflammatory processes than mice infected with TcII. This supported the working hypothesis, since TcIV from Amazonas was less pathogenic than TcII from Paraná; and agreed with the lower severity of human cases of ChD in the Amazon region.

  16. Structural characterization of CYP51 from Trypanosoma cruzi and Trypanosoma brucei bound to the antifungal drugs posaconazole and fluconazole.

    Directory of Open Access Journals (Sweden)

    Chiung-Kuang Chen

    Full Text Available BACKGROUND: Chagas Disease is the leading cause of heart failure in Latin America. Current drug therapy is limited by issues of both efficacy and severe side effects. Trypansoma cruzi, the protozoan agent of Chagas Disease, is closely related to two other major global pathogens, Leishmania spp., responsible for leishmaniasis, and Trypansoma brucei, the causative agent of African Sleeping Sickness. Both T. cruzi and Leishmania parasites have an essential r