Science.gov

Sample records for infectado con trypanosoma

  1. Malleable mitochondrion of Trypanosoma brucei.

    PubMed

    Verner, Zdeněk; Basu, Somsuvro; Benz, Corinna; Dixit, Sameer; Dobáková, Eva; Faktorová, Drahomíra; Hashimi, Hassan; Horáková, Eva; Huang, Zhenqiu; Paris, Zdeněk; Peña-Diaz, Priscila; Ridlon, Lucie; Týč, Jiří; Wildridge, David; Zíková, Alena; Lukeš, Julius

    2015-01-01

    The importance of mitochondria for a typical aerobic eukaryotic cell is undeniable, as the list of necessary mitochondrial processes is steadily growing. Here, we summarize the current knowledge of mitochondrial biology of an early-branching parasitic protist, Trypanosoma brucei, a causative agent of serious human and cattle diseases. We present a comprehensive survey of its mitochondrial pathways including kinetoplast DNA replication and maintenance, gene expression, protein and metabolite import, major metabolic pathways, Fe-S cluster synthesis, ion homeostasis, organellar dynamics, and other processes. As we describe in this chapter, the single mitochondrion of T. brucei is everything but simple and as such rivals mitochondria of multicellular organisms.

  2. Lipid metabolism in Trypanosoma brucei

    PubMed Central

    Smith, Terry K.; Bütikofer, Peter

    2013-01-01

    Trypanosoma brucei membranes consist of all major eukaryotic glycerophospholipid and sphingolipid classes. These are de novo synthesized from precursors obtained either from the host or from catabolised endocytosed lipids. In recent years, substantial progress has been made in the molecular and biochemical characterisation of several of these lipid biosynthetic pathways, using gene knockout or RNA interference strategies or by enzymatic characterization of individual reactions. Together with the completed genome, these studies have highlighted several possible differences between mammalian and trypanosome lipid biosynthesis that could be exploited for the development of drugs against the diseases caused by these parasites. PMID:20382188

  3. Immunotherapy of Trypanosoma cruzi infections.

    PubMed

    Chamond, N; Coatnoan, N; Minoprio, P

    2002-10-01

    The protozoan parasite Trypanosoma cruzi, causative agent of Chagas' disease, is transmitted to man and other mammals by triatominae insects, or 'kissing bugs'. Since its discovery in 1909, by Carlos Chagas, this parasite has been the object of several publications in the domains of immunology, cellular biology and of control gene organization, regulation and expression. Although much progress has been made concerning prophylaxis of Chagas' disease, particularly vector eradication, additional cases of infection and disease development still occur every day throughout the world. Whilst infection was largely limited in the past to vector transmission in endemic areas of Latin America, its impact has increased in terms of congenital and blood transmission, transplants and recrudescence following immunosuppressive states. Reports on new insect vectors adapted to the parasite and domestic animals infected in more developed countries, emphasize the continuing worldwide public health issue. Therapy against this parasite is limited and cure is subjected to several criteria, such as susceptibility of the parasite strain, age of the host and stage of the disease. The ability of Trypanosoma cruzi to induce important and various host immune system dysfunctions makes the development of effective vaccines a laborious and complex task. These considerations strengthen the latent significance of Chagas' disease and encourage the search for new preventive procedures and the research on rational vaccines.

  4. Artemisinins Inhibit Trypanosoma cruzi and Trypanosoma brucei rhodesiense In Vitro Growth▿

    PubMed Central

    Mishina, Yuliya V.; Krishna, Sanjeev; Haynes, Richard K.; Meade, John C.

    2007-01-01

    Artemisinin compounds inhibit in vitro growth of cultured Trypanosoma cruzi and Trypanosoma brucei rhodesiense at concentrations in the low micromolar range. Artemisinin also inhibits calcium-dependent ATPase activity in T. cruzi membranes, suggesting a mode of action via membrane pumps. Artemisinins merit further investigation as chemotherapeutic options for these pathogens. PMID:17339374

  5. Characterization of Trypanosoma cruzi telomerase.

    PubMed

    Campelo, Riward; Galindo, Maria Mercedes; Ramirez, Jose Luis

    2011-12-01

    High telomerase activity is always associated with actively dividing cells, however the detection of this activity in dividing Leishmania and Trypanosoma cruzi cells has always been disappointingly low. Recently, we have found that Leishmania major telomerase activity can be activated by heat, which combined with dilutions of the nuclear extracts produced an increase in activity comparable to cancer cells. Here we examined whether T. cruzi telomerase shares the same physicochemical properties of primer specificity and overall features of the L. major. Our studies revealed that no telomerase inhibitory factors were present in the nuclear lysates of T. cruzi however the enzyme was activated by heat and was very resilient to heat denaturation. We also showed the extension primer specificity, susceptibility to RNase-A and RNase-H digestion, and the effect of telomerase inhibitors.

  6. Protective immunity against Trypanosoma cruzi.

    PubMed

    Parodi, Cecilia; Padilla, Angel Marcelo; Basombrío, Miguel Angel

    2009-07-01

    Upon infection, Trypanosoma cruzi triggers a strong immune response that has both protective and pathological consequences. In this work, several important questions regarding protective immunity are reviewed. Emphasis is placed on recent studies of the important protective role of CD8+ T cells and on previous studies of immunisation of domestic T. cruzi reservoirs that sought to address practical vaccination problems. Research on the maturation of memory cells and studies indicating that the prevalence of T. cruzi-specific T-cell responses and a high frequency of committed CD8+ T cells are associated with better clinical outcomes are also reviewed. Additionally, animal models in which protection was achieved without immunopathological consequences are discussed.

  7. Transcription of Trypanosoma brucei maxicircles

    SciTech Connect

    Michelotti, E.F.; Hajduk, S.L.

    1987-05-01

    Trypanosoma brucei is a protozoan parasite which developmentally regulates mitochondrial activity. In the mammal T. brucei produces ATP entirely by glycolysis while cytochrome mediated respiration resumes in the life-stage in the midgut of the insect vector. Using quantitative S1 nuclease protection assays two types of regulation of the steady state levels of the mitochondrial transcripts were found. Transcription of cytochrome b, cytochrome oxidase, and the rRNA genes is repressed in early bloodstream developmental stages, undergoes dramatic activation in later bloodstream stages, and finally a lesser activation in the insect developmental stage. Transcription of NADH dehydrogenase genes, however, is unregulated. Mitochondrial transcripts with a 5' triphosphate terminus, representing the site of transcription initiation, were capped using guanylyl transferase. The in vitro capped RNA hybridized to only one of eight mitochondrial restriction fragments on a Southern blot, however, hybridization of Southern blots with RNA from ..cap alpha..-/sup 32/P-UTP pulsed mitochondria labelled all restriction fragments equally. These results suggest that each DNA strand has a single promoter which directs the transcription of a full-length RNA which is subsequently processed. Different mitochondrial genes, despite being expressed on the same precursor RNA molecule, are independently regulated by both transcription initiation and RNA processing.

  8. What happens when Trypanosoma brucei leaves Africa.

    PubMed

    Jensen, Robert E; Simpson, Larry; Englund, Paul T

    2008-10-01

    Julius Lukes and co-workers evaluated the evolutionary origin of Trypanosoma equiperdum and Trypanosoma evansi, parasites that cause horse and camel diseases. Although similar to T. brucei, the sleeping-sickness parasite, these trypanosomes do not cycle through the tsetse fly and have been able to spread beyond Africa. Transmission occurs sexually, or via blood-sucking flies or vampire bats. They concluded that these parasites, which resemble yeast petite mutants, are T. brucei sub-species, which have evolved recently through changes in mitochondrial DNA.

  9. RNA turnover in Trypanosoma brucei.

    PubMed Central

    Ehlers, B; Czichos, J; Overath, P

    1987-01-01

    Regulation of variant surface glycoprotein (VSG) mRNA turnover in Trypanosoma brucei was studied in bloodstream forms, in procyclic cells, and during in vitro transformation of bloodstream forms to procyclic cells by approach-to-equilibrium labeling and pulse-chase experiments. Upon initiation of transformation at 27 degrees C in the presence of citrate-cis-aconitate, the half-life of VSG mRNA was reduced from 4.5 h in bloodstream forms to 1.2 h in transforming cells. Concomitantly, an approximately 25-fold decrease in the rate of transcription was observed, resulting in a 100-fold reduction in the steady-state level of de novo-synthesized VSG mRNA. This low level of expression was maintained for at least 7 h, finally decreasing to an undetectable level after 24 h. Transcription of the VSG gene in established procyclic cells was undetectable. For comparison, the turnover of polyadenylated and nonpolyadenylated RNA, beta-tubulin mRNA, and mini-exon-derived RNA (medRNA) was studied. For medRNA, no significant changes in the rate of transcription or stability were observed during differentiation. In contrast, while the rate of transcription of beta-tubulin mRNA in in vitro-cultured bloodstream forms, transforming cells, and established procyclic cells was similar, the half life was four to five times longer in procyclic cells (t1/2, 7 h) than in cultured bloodstream forms (t1/2, 1.4 h) or transforming cells (t1/2, 1.7 h). Inhibition of protein synthesis in bloodstream forms at 37 degrees Celsius caused a dramatic 20-fold decrease in the rate of VSG mRNA synthesis and a 6-fold decrease in half-life to 45 min, while beta-tubulin mRNA was stabilized 2- to 3-fold and mRNA stability remained unaffected. It is postulated that triggering transformation or inhibiting protein synthesis induces changes in the abundance of the same regulatory molecules which effect the shutoff of VSG gene transcription in addition to shortening the half-life of VSG mRNA. Images PMID:2436040

  10. Overview of DNA Repair in Trypanosoma cruzi, Trypanosoma brucei, and Leishmania major

    PubMed Central

    Passos-Silva, Danielle Gomes; Rajão, Matheus Andrade; Nascimento de Aguiar, Pedro Henrique; Vieira-da-Rocha, João Pedro; Machado, Carlos Renato; Furtado, Carolina

    2010-01-01

    A wide variety of DNA lesions arise due to environmental agents, normal cellular metabolism, or intrinsic weaknesses in the chemical bonds of DNA. Diverse cellular mechanisms have evolved to maintain genome stability, including mechanisms to repair damaged DNA, to avoid the incorporation of modified nucleotides, and to tolerate lesions (translesion synthesis). Studies of the mechanisms related to DNA metabolism in trypanosomatids have been very limited. Together with recent experimental studies, the genome sequencing of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania major, three related pathogens with different life cycles and disease pathology, has revealed interesting features of the DNA repair mechanism in these protozoan parasites, which will be reviewed here. PMID:20976268

  11. 21 CFR 866.3870 - Trypanosoma spp. serological reagents.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Trypanosoma spp. serological reagents. 866.3870 Section 866.3870 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3870 Trypanosoma...

  12. 21 CFR 866.3870 - Trypanosoma spp. serological reagents.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Trypanosoma spp. serological reagents. 866.3870 Section 866.3870 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3870 Trypanosoma...

  13. Activity of Bisnaphthalimidopropyl Derivatives against Trypanosoma brucei

    PubMed Central

    Graça, Nuno A. G.; Gaspar, Luis; Costa, David M.; Loureiro, Inês; Thoo-Lin, Paul Kong; Ramos, Isbaal; Roura, Meritxell; Pruvost, Alain; Pemberton, Ian K.; Loukil, Hadjer; MacDougall, Jane

    2016-01-01

    Current treatments for African trypanosomiasis are either toxic, costly, difficult to administer, or prone to elicit resistance. This study evaluated the activity of bisnaphthalimidopropyl (BNIP) derivatives against Trypanosoma brucei. BNIPDiaminobutane (BNIPDabut), the most active of these compounds, showed in vitro inhibition in the single-unit nanomolar range, similar to the activity in the reference drug pentamidine, and presented low toxicity and adequate metabolic stability. Additionally, using a murine model of acute infection and live imaging, a significant decrease in parasite load in BNIPDabut-treated mice was observed. However, cure was not achieved. BNIPDabut constitutes a new scaffold for antitrypanosomal drugs that deserves further consideration. PMID:26787703

  14. Species-specific markers for the differential diagnosis of Trypanosoma cruzi and Trypanosoma rangeli and polymorphisms detection in Trypanosoma rangeli.

    PubMed

    Ferreira, Keila Adriana Magalhães; Fajardo, Emanuella Francisco; Baptista, Rodrigo P; Macedo, Andrea Mara; Lages-Silva, Eliane; Ramírez, Luis Eduardo; Pedrosa, André Luiz

    2014-06-01

    Trypanosoma cruzi and Trypanosoma rangeli are kinetoplastid parasites which are able to infect humans in Central and South America. Misdiagnosis between these trypanosomes can be avoided by targeting barcoding sequences or genes of each organism. This work aims to analyze the feasibility of using species-specific markers for identification of intraspecific polymorphisms and as target for diagnostic methods by PCR. Accordingly, primers which are able to specifically detect T. cruzi or T. rangeli genomic DNA were characterized. The use of intergenic regions, generally divergent in the trypanosomatids, and the serine carboxypeptidase gene were successful. Using T. rangeli genomic sequences for the identification of group-specific polymorphisms and a polymorphic AT(n) dinucleotide repeat permitted the classification of the strains into two groups, which are entirely coincident with T. rangeli main lineages, KP1 (+) and KP1 (-), previously determined by kinetoplast DNA (kDNA) characterization. The sequences analyzed totalize 622 bp (382 bp represent a hypothetical protein sequence, and 240 bp represent an anonymous sequence), and of these, 581 (93.3%) are conserved sites and 41 bp (6.7%) are polymorphic, with 9 transitions (21.9%), 2 transversions (4.9%), and 30 (73.2%) insertion/deletion events. Taken together, the species-specific markers analyzed may be useful for the development of new strategies for the accurate diagnosis of infections. Furthermore, the identification of T. rangeli polymorphisms has a direct impact in the understanding of the population structure of this parasite.

  15. Immune Evasion Strategies of Trypanosoma cruzi

    PubMed Central

    Flávia Nardy, Ana; Freire-de-Lima, Célio Geraldo; Morrot, Alexandre

    2015-01-01

    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. PMID:26240832

  16. Differentiation of Trypanosoma cruzi and Trypanosoma rangeli of Colombia using minicircle hybridization tests.

    PubMed

    Botero, Adriana; Ortiz, Sylvia; Muñoz, Sergio; Triana, Omar; Solari, Aldo

    2010-11-01

    Although Trypanosoma rangeli is harmless for humans, it is a serious problem since it may be confused with diagnosis of Trypanosoma cruzi, the etiologic agent of Chagas disease. Both parasites overlap geographically, share antigenic protein, and are able to infect the same Triatominae vector and vertebrate host, including human. Our objective was to differentiate T. cruzi and T. rangeli isolates from Colombia based on polymerase chain reaction (PCR) amplification of the minicircles followed by appropriate hybridization tests with selected DNA probes and restriction fragment length polymorphism (RFLP) analysis. We worked with highly characterized T. cruzi and T. rangeli isolates from different biologic origins and geographic areas of Colombia, and they were analyzed by RFLP and PCR amplification of variable region of minicircles and Southern blot analysis. Our results and experimental conditions demonstrate the usefulness of PCR amplification of the minicircles followed by Southern blot analysis to differentiate T. cruzi from T. rangeli, which can be highly important to improve diagnosis of Chagas disease.

  17. Variant surface glycoprotein from Trypanosoma evansi is partially responsible for the cross-reaction between Trypanosoma evansi and Trypanosoma vivax.

    PubMed

    Uzcanga, Graciela L; Perrone, Trina; Noda, José Alfredo; Pérez-Pazos, Jacqueline; Medina, Rafael; Hoebeke, Johan; Bubis, José

    2004-01-27

    Salivarian trypanosomes use antigenic variation of their variant-specific surface glycoprotein (VSG) coat as a defense against the host immune system. Although about 1000 VSG and pseudo-VSG genes are scattered throughout the trypanosome genome, each trypanosome expresses only one VSG, while the rest of the genes are transcriptionally silent. A 64-kDa glycosylated cross-reacting antigen between Trypanosoma evansi and Trypanosoma vivax (p64), which was purified from the TEVA1 T. evansi Venezuelan isolate, was proven here to represent the soluble form of a VSG. Initially, a biochemical characterization of p64 was carried out. Gel filtration chromatography, sedimentation, and chemical cross-linking provided evidences of the dimeric nature of p64. The hydrodynamic parameters indicated that p64 is asymmetrical with a frictional ratio f/fo = 1.57. Isoelectric focusing and two-dimensional polyacrylamide gel electrophoresis revealed that p64 contained two isoforms with isoelectric points of 6.8-6.9 and 7.1-7.2. When p64 and three p64 Staphylococcus aureus V8 proteolytic fragments were sequenced, the same N-termini sequence was obtained: Ala-Pro-Ile-Thr-Asp-Ala-Asp-Leu-Gly-Pro-Ala-Gln-Ile-Ala-Asp, which displayed a significant homology with a putative Trypanosoma brucei VSG gene located on chromosome 4. Additionally, immunofluorescence microscopy on T. evansi and T. vivax established that p64 and its T. vivax homologue were confined to the surface of both parasites. An immunological characterization of this antigen was also carried out using several Venezuelan T. evansi isolates expressing different VSGs, which were obtained from naturally infected animals. Although sera from animals infected with the various T. evansi isolates recognized p64, only one isolate, besides TEVA1, contained polypeptides that were recognized by anti-p64 antibodies. All these results together with prior evidences [Uzcanga, G. et al. (2002) Parasitology 124, 287-299] confirmed that p64 is the

  18. In Vitro Uptake of Isometamidium and Diminazene by Trypanosoma brucei

    PubMed Central

    Girgis-Takla, Pamela; James, Dinah M.

    1974-01-01

    The cattle trypanocide, isometamidium, was readily taken up by inactivated Trypanosoma brucei, while its uptake by living parasites was reduced or inhibited by plasma. In both respects isometamidium differs from diminazene. PMID:15830490

  19. Trypanosoma cruzi meningoencephalitis in a patient with acquired immunodeficiency syndrome.

    PubMed

    Yasukawa, Kosuke; Patel, Shital M; Flash, Charlene A; Stager, Charles E; Goodman, Jerry C; Woc-Colburn, Laila

    2014-07-01

    As a result of global migration, a significant number of people with Trypanosoma cruzi infection now live in the United States, Canada, many countries in Europe, and other non-endemic countries. Trypanosoma cruzi meningoencephalitis is a rare cause of ring-enhancing lesions in patients with acquired immunodeficiency syndrome (AIDS) that can closely mimic central nervous system (CNS) toxoplasmosis. We report a case of CNS Chagas reactivation in an AIDS patient successfully treated with benznidazole and antiretroviral therapy in the United States.

  20. Immunization of mice with Trypanosoma cruzi polyribosomes.

    PubMed Central

    Leon, L L; Leon, W; Chaves, L; Costa, S C; Cruz, M Q; Brascher, H M; Lima, A O

    1980-01-01

    Studies were carried out with a polyribosomal fraction isolated from Trypanosoma cruzi Y epimastigotes, with the intention to determine both its immunogenic activity and the degree of protection it could induce against experimental T. cruzi infection. This fraction was assayed in four groups of mice by using different schedules of vaccination and varying the dose, intervals, and route of administration. Seven days after the last dose, the animals were sacrificed for immunological studies or subjected to challenge with T. cruzi trypomastigotes. The results obtained in all schedules showed that our polyribosomal fraction only induced a weak antibody response, but was capable of evoking an expressive cellular response. It was also shown that this fraction has the capacity of inducing a high degree of protection against T. cruzi infection, as determined by the decrease of parasitemia and the prolonged survival time of immunized animals. PMID:6987175

  1. Trypanosoma cruzi Infection and Host Lipid Metabolism

    PubMed Central

    Miao, Qianqian

    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 metabolic pathways. We reviewed the interaction of T. cruzi with each of the relevant host components, in order to further understand the roles of host lipid metabolism in T. cruzi infection. This review sheds light on the potential impact of T. cruzi infection on the status of host lipid homeostasis. PMID:25276058

  2. Trypanosoma cruzi, cancer and the Cold War.

    PubMed

    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.

  3. Trypanosoma cruzi: inhibition of metacyclogenesis by mannose.

    PubMed

    Barbieri, M A; Lammel, E M; Isola, E L; Bertini, F

    1992-08-31

    Metacyclogenesis of Trypanosoma cruzi epimastigotes was evaluated in a medium supplemented with Triatoma infestans intestinal homogenate in the presence of sugars and derivates as are mannose, galactose, fucose, N-acetylglucosamine, mannose 6-P, and fructose 1,6-P at a concentration of 25 mM. Only mannose significantly inhibited metacyclogenesis. Sodium metaperiodate and trypsin treatment of the intestinal homogenate also inhibited differentiation. In our opinion there exists a proteinic factor in the intestine of the vector that promotes metacyclogenesis and is incorporated by the parasite. Treatment of the intestinal homogenate with alkaline phosphatase had no effect. Instead, high ionic strength in the medium (0.4 M NaCl) strongly inhibited metacyclogenesis indicating that, in these conditions, the possible binding of the differentiation factor to the parasite surface was inhibited.

  4. Proteomics of Trypanosoma evansi Infection in Rodents

    PubMed Central

    Pallavi, Rani; Chakravarthy, Harshini; Chandran, Syama; Kumar, Rajender; Gupta, Ashok Kumar; Singh, Raj Kumar; Yadav, Suresh Chandra; Tatu, Utpal

    2010-01-01

    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 first ever

  5. Nanomolar Inhibitors of Trypanosoma brucei RNA Triphosphatase.

    PubMed

    Smith, Paul; Ho, C Kiong; Takagi, Yuko; Djaballah, Hakim; Shuman, Stewart

    2016-02-23

    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. The stark differences between the structure and mechanism of the RNA triphosphatase (RTPase) component of the mRNA capping apparatus in pathogenic protozoa, fungi, and viruses and those of their metazoan hosts highlight RTPase as a target for anti

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

    PubMed

    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.

  7. New Trypanosoma species, Trypanosoma gennarii sp. nov., from South American marsupial in Brazilian Cerrado.

    PubMed

    Ferreira, Juliana I G S; da Costa, Andréa P; Nunes, Pablo Henrique; Ramirez, Diego; Fournier, Gislene F R; Saraiva, Danilo; Tonhosolo, Renata; Marcili, Arlei

    2017-12-01

    Hundreds of trypanosome species have been described in all mammalian orders, on every continent, including with mixed infections. Trypanosomes circulate in the form of sylvatic enzootic infections transmitted by blood-sucking insects that are associated with the host mammals. Small wild mammals were caught in a fragment of Cerrado terrain on an island in the hydroelectric reservoir of Três Marias, in the central region of the state of Minas Gerais, using pitfall and Sherman traps with different means of attraction. DNA samples from these mammals were subjected to the conventional polymerase chain reaction (PCR) for the full-length genes SSU rDNA and gGAPDH. A total of 232 animals of the orders Didelphimorphia, Rodentia, Chiroptera and Cingulata were caught (total of 17 species). There were also four species of marsupials: Monodelphis domestica, Didelphis albiventris, Gralicinanus agilis and Micoureus paraguaianus. Among these, there were eight positive individuals of Monodelphis domestica. However, nine cultures were established, because one of them was parasitized by two species of trypanosomes: Trypanosoma cruzi and a new trypanosome species. The new species have a large epimastigote forms, and with a well-developed undulating membrane in trypomastigote forms. The new species Trypanosoma gennarii was described in Monodelphis domestica. Copyright © 2017 Elsevier B.V. All rights reserved.

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

    PubMed Central

    Habila, Nathan; Agbaji, Abel S.; Ladan, Zakari; Bello, Isaac A.; Haruna, Emmanuel; Dakare, Monday A.; Atolagbe, Taofiq O.

    2010-01-01

    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. PMID:20700425

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

    PubMed

    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 (p<0.0001-0.01) and a discrete lymphomonocytic infiltrate in cardiac and skeletal muscles was present. In the chronic phase, the histological view was normal. In contrast, control group revealed amastigote nests and typical histopathological alterations compatible with chagasic myocarditis, endocarditis and pericarditis. These results, together with previous works in our laboratory, show that T. rangeli induces immunoprotection in three species of animals: mice, guinea 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. Copyright © 2013 Elsevier B.V. All rights reserved.

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

    PubMed Central

    Moreno, S Andrea; Nava, Mayerly

    2015-01-01

    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. PMID:26061149

  11. Phospholipase A2 from Trypanosoma brucei gambiense and Trypanosoma brucei brucei: inhibition by organotins.

    PubMed

    Shuaibu, M N; Kanbara, H; Yanagi, T; Ameh, D A; Bonire, J J; Nok, A J

    2001-11-01

    Activity and kinetics of phospholipase A2 (PLA2) from Trypanosoma brucei gambiense (Wellcome strain) and Trypanosoma brucei brucei (GUTat 3.1) were examined using two different fluorescent substrates. The activity in the supernatants of sonicated parasites was Ca2+-independent, strongly stimulated by Triton X-100 with optimum activity at 37 degrees C and pH 6.5-8.5. To encourage a possible interaction between the parasite enzyme and organotin compounds, fatty acid derivatives of dibutyltin dichloride were synthesized and evaluated as potential inhibitors of PLA2. The enzyme from the two-trypanosome species differ with respect to kinetic parameters and are noncompetitively inhibited by the organotin compounds. The Michaelis constant (KM) for PLA2 from T. b. brucei is 63.87 and 30.90 microM while for T. b. gambiense it is 119.64 and 32.91 microM for the substrates 1,2-bis-(1-pyrenebutanoyl)-sn-glycero-3-phosphocholine (PBGPC) and 2-(12-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)dodecanoyl-1-hexadecanoyl-sn-glycero-3-phosphocholine (NBDC12-HPC), respectively.

  12. Troglitazone induces differentiation in Trypanosoma brucei

    SciTech Connect

    Denninger, Viola; Figarella, Katherine; Schoenfeld, Caroline; Brems, Stefanie; Busold, Christian; Lang, Florian; Hoheisel, Joerg; Duszenko, Michael . E-mail: michael.duszenko@uni-tuebingen.de

    2007-05-15

    Trypanosoma brucei, a protozoan parasite causing sleeping sickness, is transmitted by the tsetse fly and undergoes a complex lifecycle including several defined stages within the insect vector and its mammalian host. In the latter, differentiation from the long slender to the short stumpy form is induced by a yet unknown factor of trypanosomal origin. Here we describe that some thiazolidinediones are also able to induce differentiation. In higher eukaryotes, thiazolidinediones are involved in metabolism and differentiation processes mainly by binding to the intracellular receptor peroxisome proliferator activated receptor {gamma}. Our studies focus on the effects of troglitazone on bloodstream form trypanosomes. Differentiation was monitored using mitochondrial markers (membrane potential, succinate dehydrogenase activity, inhibition of oxygen uptake by KCN, amount of cytochrome transcripts), morphological changes (Transmission EM and light microscopy), and transformation experiments (loss of the Variant Surface Glycoprotein coat and increase of dihydroliponamide dehydrogenase activity). To further investigate the mechanisms responsible for these changes, microarray analyses were performed, showing an upregulation of expression site associated gene 8 (ESAG8), a potential differentiation regulator.

  13. Motility modes of the parasite Trypanosoma brucei

    NASA Astrophysics Data System (ADS)

    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.

  14. Immunocytochemical localization of neuraminidase in Trypanosoma cruzi.

    PubMed Central

    Souto-Padrón, T; Harth, G; de Souza, W

    1990-01-01

    A polyclonal antibody obtained against neuraminidase purified from Trypanosoma cruzi was used for the localization of the protein in whole cells by immunofluorescence microscopy and in thin sections of parasites (epimastigote, amastigote, and trypomastigote forms) embedded at a low temperature in Lowicryl K4M resin. The intensity of labeling, as evaluated by the number of gold particles associated with the parasite, varied according to the protozoan developmental stage. In the noninfective epimastigote forms, labeling of the cell surface was very weak. However, an intense labeling of some cytoplasmic vacuoles was observed. Labeling of the surfaces of most of the trypomastigote forms was weak, while gold particles were seen in association with the flagellar pockets of these forms, which suggests that the enzyme is secreted through this region. Intense labeling of the surfaces of many, but not all, transition forms between trypomastigote and amastigote forms was observed. Amastigote forms found in the supernatant of infected cell cultures had their surfaces intensely labeled, while few particles were seen on the surfaces of intracellular amastigotes. The results obtained are discussed in relation to the role played by T. cruzi neuraminidase in the process of parasite-host cell interaction. Images PMID:2407649

  15. The flagellar adenylate kinases of Trypanosoma cruzi.

    PubMed

    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. © The Author 2014. Published by Oxford University Press on behalf of FEMS. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  16. The Architecture of Trypanosoma brucei editosomes.

    PubMed

    McDermott, Suzanne M; Luo, Jie; Carnes, Jason; Ranish, Jeff A; Stuart, Kenneth

    2016-10-18

    Uridine insertion and deletion RNA editing generates functional mitochondrial mRNAs in Trypanosoma brucei Editing is catalyzed by three distinct ∼20S editosomes that have a common set of 12 proteins, but are typified by mutually exclusive RNase III endonucleases with distinct cleavage specificities and unique partner proteins. Previous studies identified a network of protein-protein interactions among a subset of common editosome proteins, but interactions among the endonucleases and their partner proteins, and their interactions with common subunits were not identified. Here, chemical cross-linking and mass spectrometry, comparative structural modeling, and genetic and biochemical analyses were used to define the molecular architecture and subunit organization of purified editosomes. We identified intra- and interprotein cross-links for all editosome subunits that are fully consistent with editosome protein structures and previously identified interactions, which we validated by genetic and biochemical studies. The results were used to create a highly detailed map of editosome protein domain proximities, leading to identification of molecular interactions between subunits, insights into the functions of noncatalytic editosome proteins, and a global understanding of editosome architecture.

  17. Crystal structure of a Trypanosoma brucei metacaspase.

    PubMed

    McLuskey, Karen; Rudolf, Jana; Proto, William R; Isaacs, Neil W; Coombs, Graham H; Moss, Catherine X; Mottram, Jeremy C

    2012-05-08

    Metacaspases are distantly related caspase-family cysteine peptidases implicated in programmed cell death in plants and lower eukaryotes. They differ significantly from caspases because they are calcium-activated, arginine-specific peptidases that do not require processing or dimerization for activity. To elucidate the basis of these differences and to determine the impact they might have on the control of cell death pathways in lower eukaryotes, the previously undescribed crystal structure of a metacaspase, an inactive mutant of metacaspase 2 (MCA2) from Trypanosoma brucei, has been determined to a resolution of 1.4 Å. The structure comprises a core caspase fold, but with an unusual eight-stranded β-sheet that stabilizes the protein as a monomer. Essential aspartic acid residues, in the predicted S1 binding pocket, delineate the arginine-specific substrate specificity. In addition, MCA2 possesses an unusual N terminus, which encircles the protein and traverses the catalytic dyad, with Y31 acting as a gatekeeper residue. The calcium-binding site is defined by samarium coordinated by four aspartic acid residues, whereas calcium binding itself induces an allosteric conformational change that could stabilize the active site in a fashion analogous to subunit processing in caspases. Collectively, these data give insights into the mechanistic basis of substrate specificity and mode of activation of MCA2 and provide a detailed framework for understanding the role of metacaspases in cell death pathways of lower eukaryotes.

  18. Phosphatidylinositol kinase activities in Trypanosoma cruzi epimastigotes.

    PubMed

    Gimenez, Alba Marina; Gesumaría, María Celeste; Schoijet, Alejandra C; Alonso, Guillermo D; Flawiá, Mirtha M; Racagni, Graciela E; Machado, Estela E

    2015-01-01

    Phosphatidylinositol (PtdIns) metabolism through phosphatidylinositol kinase (PIKs) activities plays a central role in different signaling pathways. In Trypanosoma cruzi, causative agent of Chagas disease, PIKs have been proposed as target for drug design in order to combat this pathogen. In this work, we studied the classes of PI4K, PIPK and PI3K that could participate in signaling pathways in T. cruzi epimastigote forms. For this reason, we analyzed their enzymatic parameters and detailed responses to avowed kinase inhibitors (adenosine, sodium deoxycholate, wortmannin and LY294002) and activators (Ca(2+), phosphatidic acid, spermine and heparin). Our results suggest the presence and activity of a class III PI4K, a class I PIPK, a class III PI3K previously described (TcVps34) and a class I PI3K. Class I PI3K enzyme, here named TcPI3K, was cloned and expressed in a bacterial system, and their product was tested for kinase activity. The possible participation of TcPI3K in central cellular events of the parasite is also discussed.

  19. Trypanosoma cruzi expresses diverse repetitive protein antigens.

    PubMed Central

    Hoft, D F; Kim, K S; Otsu, K; Moser, D R; Yost, W J; Blumin, J H; Donelson, J E; Kirchhoff, L V

    1989-01-01

    We screened a Trypanosoma cruzi cDNA expression library with human and rabbit anti-T. cruzi sera and identified cDNA clones that encode polypeptides containing tandemly arranged repeats which are 6 to 34 amino acids in length. The peptide repeats encoded by these cDNAs varied markedly in sequence, copy number, and location relative to the polyadenylation site of the mRNAs from which they were derived. The repeats were specific for T. cruzi, but in each case the sizes of the corresponding mRNAs and the total number of repeat copies encoded varied considerably among different isolates of the parasite. Expression of the peptide repeats was not stage specific. One of the peptide repeats occurred in a protein with an Mr of greater than 200,000 and one was in a protein of Mr 75,000 to 105,000. The frequent occurrence and diversity of these peptide repeats suggested that they may play a role in the ability of the parasite to evade immune destruction in its invertebrate and mammalian hosts, but the primary roles of these macromolecules may be unrelated to the host-parasite relationship. Images PMID:2659529

  20. Unveiling the Trypanosoma cruzi Nuclear Proteome

    PubMed Central

    dos Santos Júnior, Agenor de Castro Moreira; Kalume, Dário Eluan; Camargo, Ricardo; Gómez-Mendoza, Diana Paola; Correa, José Raimundo; Charneau, Sébastien; de Sousa, Marcelo Valle; de Lima, Beatriz Dolabela; Ricart, Carlos André Ornelas

    2015-01-01

    Replication of Trypanosoma cruzi, the etiological agent of Chagas disease, displays peculiar features, such as absence of chromosome condensation and closed mitosis. Although previous proteome and subproteome analyses of T. cruzi have been carried out, the nuclear subproteome of this protozoan has not been described. Here, we report, for the first time to the best of our knowledge, the isolation and proteome analysis of T. cruzi nuclear fraction. For that, T. cruzi epimastigote cells were lysed and subjected to cell fractionation using two steps of sucrose density gradient centrifugation. The purity of the nuclear fraction was confirmed by phase contrast and fluorescence microscopy. Liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) allowed the identification of 864 proteins. Among those, 272 proteins were annotated as putative uncharacterized, and 275 had not been previously reported on global T. cruzi proteome analysis. Additionally, to support our enrichment method, bioinformatics analysis in DAVID was carried out. It grouped the nuclear proteins in 65 gene clusters, wherein the clusters with the highest enrichment scores harbor members with chromatin organization and DNA binding functions. PMID:26383644

  1. Messenger RNA processing sites in Trypanosoma brucei.

    PubMed

    Benz, Corinna; Nilsson, Daniel; Andersson, Björn; Clayton, Christine; Guilbride, D Lys

    2005-10-01

    In Kinetoplastids, protein-coding genes are transcribed polycistronically by RNA polymerase II. Individual mature mRNAs are generated from polycistronic precursors by 5' trans splicing of a 39-nt capped leader RNA and 3' polyadenylation. It was previously known that trans splicing generally occurs at an AG dinucleotide downstream of a polypyrimidine tract, and that polyadenylation is coupled to downstream trans splicing. The few polyadenylation sites that had been examined were 100-400 nt upstream of the polypyrimidine tract which marked the adjacent trans splice site. We wished to define the sequence requirements for trypanosome mRNA processing more tightly and to generate a predictive algorithm. By scanning all available Trypanosoma brucei cDNAs for splicing and polyadenylation sites, we found that trans splicing generally occurs at the first AG following a polypyrimidine tract of 8-25 nt, giving rise to 5'-UTRs of a median length of 68 nt. We also found that in general, polyadenylation occurs at a position with one or more A residues located between 80 and 140 nt from the downstream polypyrimidine tract. These data were used to calibrate free parameters in a grammar model with distance constraints, enabling prediction of polyadenylation and trans splice sites for most protein-coding genes in the trypanosome genome. The data from the genome analysis and the program are available from: .

  2. A Protein Complex Map of Trypanosoma brucei

    PubMed Central

    Mehta, Vaibhav; Najafabadi, Hamed S.; Moshiri, Houtan; Jardim, Armando; Salavati, Reza

    2016-01-01

    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. PMID:26991453

  3. Subcellular proteomics of Trypanosoma cruzi reservosomes

    PubMed Central

    Sant’Anna, Celso; Nakayasu, Ernesto S.; Pereira, Miria G.; Lourenço, Daniela; de Souza, Wanderley; Almeida, Igor C.; Cunha-e-Silva, Narcisa L.

    2009-01-01

    Reservosomes are the endpoint of the endocytic pathway in Trypanosoma cruzi epimastigotes. These organelles have the particular ability to concentrate proteins and lipids obtained from medium together with the main proteolytic enzymes originated from the secretory pathway, being at the same time a storage organelle and the main site of protein degradation. Subcellular proteomics have been extensively used for profiling organelles in different cell types. Here, we combine cell fractionation and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis to identify reservosome-resident proteins. Starting from a purified reservosome fraction, we established a protocol to isolate reservosome membranes. Transmission electron microscopy was applied to confirm the purity of the fractions. To achieve a better coverage of identified proteins we analyzed the fractions separately and combined the results. LC-MS/MS analysis identified in total 709 T. cruzi-specific proteins; of these, 456 had predicted function and 253 were classified as hypothetical proteins. We could confirm the presence of most of the proteins validated by previous work and identify new proteins from different classes such as enzymes, proton pumps, transport proteins and others. The definition of the reservosome protein profile is a good tool to assess their molecular signature, identify molecular markers, and understand their relationship with different organelles. PMID:19288526

  4. Troglitazone induces differentiation in Trypanosoma brucei.

    PubMed

    Denninger, Viola; Figarella, Katherine; Schönfeld, Caroline; Brems, Stefanie; Busold, Christian; Lang, Florian; Hoheisel, Jörg; Duszenko, Michael

    2007-05-15

    Trypanosoma brucei, a protozoan parasite causing sleeping sickness, is transmitted by the tsetse fly and undergoes a complex lifecycle including several defined stages within the insect vector and its mammalian host. In the latter, differentiation from the long slender to the short stumpy form is induced by a yet unknown factor of trypanosomal origin. Here we describe that some thiazolidinediones are also able to induce differentiation. In higher eukaryotes, thiazolidinediones are involved in metabolism and differentiation processes mainly by binding to the intracellular receptor peroxisome proliferator activated receptor gamma. Our studies focus on the effects of troglitazone on bloodstream form trypanosomes. Differentiation was monitored using mitochondrial markers (membrane potential, succinate dehydrogenase activity, inhibition of oxygen uptake by KCN, amount of cytochrome transcripts), morphological changes (Transmission EM and light microscopy), and transformation experiments (loss of the Variant Surface Glycoprotein coat and increase of dihydroliponamide dehydrogenase activity). To further investigate the mechanisms responsible for these changes, microarray analyses were performed, showing an upregulation of expression site associated gene 8 (ESAG8), a potential differentiation regulator.

  5. Trypanosoma brucei: infection in murine diabetes.

    PubMed

    Amole, B O; Wittner, M; Hewlett, D; Tanowitz, H B

    1985-12-01

    The course of infection due to Trypanosoma brucei infection was observed in genetically diabetic and streptozotocin-induced diabetic mice. A strain of T. brucei, TREU 667, was used which produces a chronic infection in C57BL/6(B6) mice lasting greater than 60 days. Genetic diabetic mice (+db/+db) are obese, and have elevated blood glucose levels, normal levels of insulin, and impaired cell-mediated immunity. Their littermates (m+/m+, m+/+db) are of normal weight, and are normoglycemic and immunocompetent. The infected +db/+db mice lived significantly longer than the nondiabetic littermates. In contrast to this finding, streptozotocin-induced diabetic B6 mice developed higher parasitemia and had shorter survival times than control B6 mice. Continuous treatment with insulin of these streptozotocin-induced diabetic mice led to normalization of blood glucose and a significant reduction of parasitemia. While hyperglycemia may be associated with higher parasitemia and death in streptozotozin-induced diabetes, genetic factors may play an additional role in the genetic models.

  6. Clomipramine kills Trypanosoma brucei by apoptosis.

    PubMed

    de Silva Rodrigues, Jean Henrique; Stein, Jasmin; Strauss, Mariana; Rivarola, Héctor Walter; Ueda-Nakamura, Tânia; Nakamura, Celso Vataru; Duszenko, Michael

    2016-06-01

    Drug repositioning, i.e. use of existing medicals to treat a different illness, is especially rewarding for neglected tropical diseases (NTD), since in this field the pharmaceutical industry is rather reluctant to spend vast investments for drug development. NTDs afflict primarily poor populations in under-developed countries, which minimizes financial profit. Here we investigated the trypanocidal effect of clomipramine, a commercial antipsychotic drug, on Trypanosoma brucei. The data showed that this drug killed the parasite with an IC50 of about 5μM. Analysis of the involved cell death mechanism revealed furthermore an initial autophagic stress response and finally the induction of apoptosis. The latter was substantiated by a set of respective markers such as phosphatidylserine exposition, DNA degradation, loss of the inner mitochondrial membrane potential and characteristic morphological changes. Clomipramine was described as a trypanothione inhibitor, but as judged from our results it also showed DNA binding capacities and induced substantial morphological changes. We thus consider it likely that the drug induces a multifold adverse interaction with the parasite's physiology and induces stress in a way that trypanosomes cannot cope with.

  7. Trypanosoma brucei metabolism is under circadian control.

    PubMed

    Rijo-Ferreira, Filipa; Pinto-Neves, Daniel; Barbosa-Morais, Nuno L; Takahashi, Joseph S; Figueiredo, Luisa M

    2017-03-13

    The Earth's rotation forced life to evolve under cyclic day and night environmental changes. To anticipate such daily cycles, prokaryote and eukaryote free-living organisms evolved intrinsic clocks that regulate physiological and behavioural processes. Daily rhythms have been observed in organisms living within hosts, such as parasites. Whether parasites have intrinsic molecular clocks or whether they simply respond to host rhythmic physiological cues remains unknown. Here, we show that Trypanosoma brucei, the causative agent of human sleeping sickness, has an intrinsic circadian clock that regulates its metabolism in two different stages of the life cycle. We found that, in vitro, ∼10% of genes in T. brucei are expressed with a circadian rhythm. The maximum expression of these genes occurs at two different phases of the day and may depend on a post-transcriptional mechanism. Circadian genes are enriched in cellular metabolic pathways and coincide with two peaks of intracellular adenosine triphosphate concentration. Moreover, daily changes in the parasite population lead to differences in suramin sensitivity, a drug commonly used to treat this infection. These results demonstrate that parasites have an intrinsic circadian clock that is independent of the host, and which regulates parasite biology throughout the day.

  8. Trypanosoma cruzi Meningoencephalitis in a Patient with Acquired Immunodeficiency Syndrome

    PubMed Central

    Yasukawa, Kosuke; Patel, Shital M.; Flash, Charlene A.; Stager, Charles E.; Goodman, Jerry C.; Woc-Colburn, Laila

    2014-01-01

    As a result of global migration, a significant number of people with Trypanosoma cruzi infection now live in the United States, Canada, many countries in Europe, and other non-endemic countries. Trypanosoma cruzi meningoencephalitis is a rare cause of ring-enhancing lesions in patients with acquired immunodeficiency syndrome (AIDS) that can closely mimic central nervous system (CNS) toxoplasmosis. We report a case of CNS Chagas reactivation in an AIDS patient successfully treated with benznidazole and antiretroviral therapy in the United States. PMID:24891470

  9. Detection of Trypanosoma cruzi by Polymerase Chain Reaction.

    PubMed

    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.

  10. Biological action of inosine analogs in Leishmania and Trypanosoma spp.

    PubMed Central

    Marr, J J; Berens, R L; Cohn, N K; Nelson, D J; Klein, R S

    1984-01-01

    Previous investigations have suggested that inosine analogs would be good models for the development of chemotherapeutic agents active against pathogenic hemoflagellates. We have systematically modified the five-membered heterocyclic ring of six inosine analogs and tested them for their antiprotozoal activities and toxicity to a mammalian cell line. All six analogs were very active against the three protozoan pathogens Leishmania donovani, Trypanosoma cruzi, and Trypanosoma gambiense. Two of the six, 9-deazainosine and allopurinol ribonucleoside, had very little toxicity for mouse L cells and offer promise as potential chemotherapeutic agents. PMID:6712205

  11. Kinetics of methionine transport and metabolism by Trypanosoma brucei brucei and Trypanosoma brucei rhodesiense.

    PubMed

    Goldberg, B; Rattendi, D; Lloyd, D; Yarlett, N; Bacchi, C J

    2000-05-01

    Methionine is an essential amino acid for both prokaryotic and eukaryotic organisms; however, little is known concerning its utilization in African trypanosomes, protozoa of the Trypanosoma brucei group. This study explored the Michaelis-Menten kinetic constants for transport and pool formation as well as metabolic utilization of methionine by two divergent strains of African trypanosomes, Trypanosoma brucei brucei (a veterinary pathogen), highly sensitive to trypanocidal agents, and Trypanosoma brucei rhodesiense (a human pathogenic isolate), highly refractory to trypanocidal arsenicals. The Michaelis-Menten constants derived by Hanes-Woolf analysis for transport of methionine for T. b. brucei and T. b. rhodesiense, respectively, were as follows: K(M) values, 1. 15 and 1.75 mM; V(max) values, 3.97 x 10(-5) and 4.86 x 10(-5) mol/L/min. Very similar values were obtained by Lineweaver-Burk analysis (K(M), 0.25 and 1.0 mM; V(max), 1 x 10(-5) and 2.0 x 10(-5) mol/L/min, T. b. brucei and T. b. rhodesiense, respectively). Cooperativity analyses by Hill (log-log) plot gave Hill coefficients (n) of 6 and 2 for T. b. brucei and T. b. rhodesiense, respectively. Cytosolic accumulation of methionine after 10-min incubation with 25 mM exogenous methionine was 1.8-fold greater in T. b. rhodesiense than T. b. brucei (2.1 vs 1.1 mM, respectively). In African trypanosomes as in their mammalian host, S-adenosylmethionine (AdoMet) is the major product of methionine metabolism. Accumulation of AdoMet was measured by HPLC analysis of cytosolic extracts incubated in the presence of increasing cytosolic methionine. In trypanosomes incubated for 10 min with saturating methionine, both organisms accumulated similar amounts of AdoMet (approximately 23 microM), but the level of trans-sulfuration products (cystathionine and cysteine) in T. b. rhodesiense was double that of T. b. brucei. Methionine incorporation during protein synthesis in T. b. brucei was 2.5 times that of T. b. rhodesiense

  12. Nanomolar Inhibitors of Trypanosoma brucei RNA Triphosphatase

    PubMed Central

    Smith, Paul; Ho, C. Kiong; Takagi, Yuko; Djaballah, Hakim

    2016-01-01

    ABSTRACT 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. PMID:26908574

  13. Mapping of VSG similarities in Trypanosoma brucei.

    PubMed

    Weirather, Jason L; Wilson, Mary E; Donelson, John E

    2012-02-01

    The protozoan parasite Trypanosoma brucei switches its variant surface glycoprotein (VSG) to subvert its mammalian hosts' immune responses. The T. brucei genome contains as many as 1600 VSG genes (VSGs), but most are silent noncoding pseudogenes. Only one functional VSG, located in a telomere-linked expression site, is transcribed at a time. Silent VSGs are copied into a VSG expression site through gene conversion. Truncated gene conversion events can generate new mosaic VSGs with segments of sequence identity to other VSGs. To examine the VSG family sub-structure within which these events occur, we combined the available VSG sequences and annotations with scripted BLAST searches to map the relationships among VSGs in the T. brucei genome. Clusters of related VSGs were visualized in 2- and 3-dimensions for different N- and C-terminal regions. Five types of N-termini (N1-N5) were observed, within which gene recombinational events are likely to occur, often with fully-coding 'functional' or 'atypical'VSGs centrally located between more dissimilar VSGs. Members of types N1, N3 and N4 are most closely related in the middle of the N-terminal region, whereas type N2 members are more similar near the N-terminus. Some preference occurs in pairing between specific N- and C-terminal types. Statistical analyses indicated no overall tendency for more related VSGs to be located closer in the genome than less related VSGs, although exceptions were noted. Many potential mosaic gene formation events within each N-terminal type were identified, contrasted by only one possible mosaic gene formation between N-terminal types (N1 and N2). These data suggest that mosaic gene formation is a major contributor to the overall VSG diversity, even though gene recombinational events between members of different N-terminal types occur only rarely.

  14. Shelter Dogs as Sentinels for Trypanosoma cruzi Transmission across Texas

    PubMed Central

    Tenney, Trevor D.; Curtis-Robles, Rachel; Snowden, Karen F.

    2014-01-01

    Chagas disease, an infection with the parasite Trypanosoma cruzi, is increasingly diagnosed among humans in the southern United States. We assessed exposure of shelter dogs in Texas to T. cruzi; seroprevalence across diverse ecoregions was 8.8%. Canine serosurveillance is a useful tool for public health risk assessment. PMID:25062281

  15. Immunization of mice with Trypanosoma rhodesiense exposed to ultraviolet irradiation

    SciTech Connect

    Charoenvit, Y.; Campbell, G.H.

    1981-11-01

    Exposure time of Trypanosoma rhodesiense as short as 1 minute to ultraviolet (uv) light prevents the organisms from causing infection. Live trypanosome challenge of mice immunized with uv-irradiated trypanosomes results in sterile immunity. This allows a method for the induction of protective immunity to experimental trypanosomiasis which can be performed in most laboratories using uv germicidal lamps found in sterile hoods.

  16. Autochthonous Transmission of Trypanosoma Cruzi in Southern California

    PubMed Central

    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

  17. Bestatin Induces Specific Changes in Trypanosoma cruzi Dipeptide Pool

    PubMed Central

    Creek, Darren J.; Faral-Tello, Paula; Barrett, Michael P.

    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. PMID:25712359

  18. Differentiation between Trypanosoma cruzi and Trypanosoma rangeli using heat-shock protein 70 polymorphisms.

    PubMed

    Fraga, Jorge; Fernandez-Calienes, Aymé; Montalvo, Ana Margarita; Maes, Ilse; Dujardin, Jean-Claude; Van der Auwera, Gert

    2014-02-01

    Differential diagnosis of infection with Trypanosoma cruzi or T. rangeli is relevant for epidemiological studies and clinical practice as both species infect humans, but only T. cruzi causes Chagas' disease. Their common antigen determinants complicate the distinction between both species, while current PCR assays used for differentiation show some drawbacks. We developed and validated a generic PCR discriminating the species by restriction fragment length polymorphism (RFLP) analysis and a duplex PCR specifically amplifying a differently sized fragment of both species. The assays are based upon a partial region of the heat-shock protein 70 gene (hsp70). The analytical sensitivity and specificity were determined for both PCRs. The assays were analytically evaluated on a panel of six T. cruzi, one T. cruzi marinkellei and four T. rangeli strains, various other infectious pathogens, a panel of spiked samples of T. cruzi, and artificially mixed infections of T. cruzi and T. rangeli. Finally, the tools were applied on 36 additional isolates of Trypanosoma species. The detection limit of the PCRs was between 0.05 and 0.5 parasite genomes, and 1-10 parasites spiked in 200 μl blood. In artificial mixtures, PCR-RFLP picked up both species in ratios up to 10(2) and duplex PCR up to 10(4) . In the 36 isolates tested, both single and mixed infections were identified. All assays were shown to be specific. Our PCRs show high potential for the differential diagnosis of T. cruzi and T. rangeli, which in view of their sensitivity can aid in the confirmation of infection with these parasites in vectors, reservoirs and clinical samples. © 2013 John Wiley & Sons Ltd.

  19. The susceptibility of Trypanosoma congolense and Trypanosoma brucei to isometamidium chloride and its synthetic impurities.

    PubMed

    Sahin, Annelise; Asencio, Corinne; Izotte, Julien; Pillay, Davita; Coustou, Virginie; Karembe, Hamadi; Baltz, Théo

    2014-07-14

    Since the 1950s, the chemotherapy of animal African trypanosomosis in cattle has essentially relied on only two compounds: isometamidium chloride (ISM), a phenanthridine, and diminazene aceturate, an aromatic diamidine. The commercial formulations of ISM, including Veridium(®) and Samorin(®), are a mixture of different compounds: ISM is the major component, mixed with the red isomer, blue isomer and disubstituted compound. To investigate the pharmacological effects of these individual compounds ISM, the blue and red isomers and the disubstituted compound were synthesised and purified by HPLC. The activity of each compound was analysed both in vitro, and in mice in vivo. For the in vitro analysis, a drug sensitivity assay was developed in 96-well tissue culture plates to determine the effective concentration which killed 50% of trypanosome population within 48 h of drug exposure (IC50). All compounds tested in vitro possessed trypanocidal activity, and purified ISM was the most active. Veridium(®) and Samorin(®) had similar IC50 values to purified ISM for both Trypanosoma congolense and Trypanosoma brucei brucei. The disubstituted compound had the highest IC50 values whereas intermediate IC50 values were obtained for the blue and red isomers. In vivo, single-dose tests were used to evaluate the trypanocidal and prophylactic activity against T. congolense. Interestingly, the prophylactic effect two months post treatment was as efficient with ISM, Veridium(®), Samorin(®) and the disubstituted compound at the highest dose of 1mg/kg whereas the red and blue isomers both showed much lower prophylactic activity. This study on T. congolense implies that it is necessary to limit the quantity of the blue and red isomers in the commercial mixture. Finally, the in vitro sensitivity assay may be useful for screening new trypanocides but also for the testing and detection of resistant trypanosome isolates.

  20. Structure and transcription of the actin gene of Trypanosoma brucei

    SciTech Connect

    Ben Amar, M.F.; Pays, A.; Tebabi, P.; Dero, B.; Seebeck, T.; Steinert, M.; Pays, E.

    1988-05-01

    In Trypanosoma brucei, the actin gene is present in a cluster of two, three, or four tandemly linked copies, depending on the strain. Each cluster seems to exist in two allelic versions, as suggested by the polymorphism of both gene number and restriction fragment length in the DNA from cloned trypanosomes. The amplification of the gene copy number probably occurs through unequal sister chromatic exchange. The chromosomes harboring the actin genes belong to the large size class. The coding sequence was 1,128 nucleotides long and showed 60 to 70% homology to other eucaryotic actin genes. Surprisingly, this homology seemed weaker with Trypanosoma congolense, Trypanosoma cruzi, Trypanosoma vivax, Trypanosoma mega, or Leishmania acting-specific sequences. The mRNA was around 1.6 kilobases long and was synthesized at the same level in bloodstream and procyclic forms of the parasite. Large RNA precursors, up to 7.7 kilobases, were found in a pattern identical in strains containing either two or three gene copies. Probing of the flanking regions of the gene with either steady-state or in vitro transcripts, as well as S1 nuclease protection and primer extension experiments, allowed mapping of the 3' splice site of the actin mRNA, 38 nucleotides upstream from the translation initiation codon. A variably sized poly(dT) tract was found about 30 base pairs ahead of the splice site. The largest detected actin mRNA precursor seemed to give rise to at least two additional stable mRNAs. The RNA polymerase transcribing the actin gene exhibited the same sensitivity to inhibition by ..cap alpha..-amanitin as that transcribing both the spliced leader and the bulk of polyadenylated mRNAs.

  1. Adaptations of Trypanosoma brucei to gradual loss of kinetoplast DNA: Trypanosoma equiperdum and Trypanosoma evansi are petite mutants of T. brucei

    PubMed Central

    Lai, De-Hua; Hashimi, Hassan; Lun, Zhao-Rong; Ayala, Francisco J.; Lukeš, Julius

    2008-01-01

    Trypanosoma brucei is a kinetoplastid flagellate, the agent of human sleeping sickness and ruminant nagana in Africa. Kinetoplastid flagellates contain their eponym kinetoplast DNA (kDNA), consisting of two types of interlocked circular DNA molecules: scores of maxicircles and thousands of minicircles. Maxicircles have typical mitochondrial genes, most of which are translatable only after RNA editing. Minicircles encode guide RNAs, required for decrypting the maxicircle transcripts. The life cycle of T. brucei involves a bloodstream stage (BS) in vertebrates and a procyclic stage (PS) in the tsetse fly vector. Partial [dyskinetoplastidy (Dk)] or total [akinetoplastidy (Ak)] loss of kDNA locks the trypanosome in the BS form. Transmission between vertebrates becomes mechanical without PS and tsetse mediation, allowing the parasite to spread outside the African tsetse belt. Trypanosoma equiperdum and Trypanosoma evansi are agents of dourine and surra, diseases of horses, camels, and water buffaloes. We have characterized representative strains of T. equiperdum and T. evansi by numerous molecular and classical parasitological approaches. We show that both species are actually strains of T. brucei, which lost part (Dk) or all (Ak) of their kDNA. These trypanosomes are not monophyletic clades and do not qualify for species status. They should be considered two subspecies, respectively T. brucei equiperdum and T. brucei evansi, which spontaneously arose recently. Dk/Ak trypanosomes may potentially emerge repeatedly from T. brucei. PMID:18245376

  2. Adaptations of Trypanosoma brucei to gradual loss of kinetoplast DNA: Trypanosoma equiperdum and Trypanosoma evansi are petite mutants of T. brucei.

    PubMed

    Lai, De-Hua; Hashimi, Hassan; Lun, Zhao-Rong; Ayala, Francisco J; Lukes, Julius

    2008-02-12

    Trypanosoma brucei is a kinetoplastid flagellate, the agent of human sleeping sickness and ruminant nagana in Africa. Kinetoplastid flagellates contain their eponym kinetoplast DNA (kDNA), consisting of two types of interlocked circular DNA molecules: scores of maxicircles and thousands of minicircles. Maxicircles have typical mitochondrial genes, most of which are translatable only after RNA editing. Minicircles encode guide RNAs, required for decrypting the maxicircle transcripts. The life cycle of T. brucei involves a bloodstream stage (BS) in vertebrates and a procyclic stage (PS) in the tsetse fly vector. Partial [dyskinetoplastidy (Dk)] or total [akinetoplastidy (Ak)] loss of kDNA locks the trypanosome in the BS form. Transmission between vertebrates becomes mechanical without PS and tsetse mediation, allowing the parasite to spread outside the African tsetse belt. Trypanosoma equiperdum and Trypanosoma evansi are agents of dourine and surra, diseases of horses, camels, and water buffaloes. We have characterized representative strains of T. equiperdum and T. evansi by numerous molecular and classical parasitological approaches. We show that both species are actually strains of T. brucei, which lost part (Dk) or all (Ak) of their kDNA. These trypanosomes are not monophyletic clades and do not qualify for species status. They should be considered two subspecies, respectively T. brucei equiperdum and T. brucei evansi, which spontaneously arose recently. Dk/Ak trypanosomes may potentially emerge repeatedly from T. brucei.

  3. Protective efficacy of isometamidium chloride and diminazene aceturate against natural Trypanosoma brucei, Trypanosoma congolense and Trypanosoma vivax infections in cattle under a suppressed tsetse population in Uganda.

    PubMed

    Magona, J W; Mayende, J S P; Okiria, R; Okuna, N M

    2004-09-01

    The protective efficacy of isometamidium chloride (ISMM) and diminazene aceturate (DIM) against Trypanosoma brucei, Trypanosoma congolense and Trypanosoma vivax infections in cattle under a suppressed tsetse population was assessed in southeast Uganda. A total of 66 and 57 trypanosome-infected cattle were treated with ISMM and DIM, respectively together with 177 trypanosome-free animals not treated were followed for 12 months, checked every 4 weeks. There was no statistical difference in the mean time to infection with any trypanosome species in animals treated with ISMM or DIM. However, the mean time to trypanosome infection was significantly longer for treated animals than controls. The mean time to infection with each of the three trypanosome species differed significantly, with the average time to T. vivax infection the lowest, followed by T. congolense and then T. brucei. The protective efficacy of DIM was as good as that of ISMM; implying curative treatments against trypanosomosis are sufficient for combination with tsetse control. Isometamidium chloride or DIM had the highest impact on T. brucei and T. congolense infections in cattle.

  4. Protein geranylgeranyltransferase-I of Trypanosoma cruzi

    PubMed Central

    Yokoyama, Kohei; Gillespie, John R.; Van Voorhis, Wesley C.; Buckner, Frederick S.; Gelb, Michael H.

    2008-01-01

    Protein geranylgeranyltransferase type I (PGGT-I) and protein farnesyltransferase (PFT) occur in many eukaryotic cells. Both consist of two subunits, the common αsubunit and a distinct β subunit. In the gene database of protozoa Trypanosoma cruzi, the causative agent of Chagas' disease, a putative protein that consists of 401 amino acids with ∼20% amino acid sequence identity to the PGGT-I β of other species was identified, cloned, and characterized. Multiple sequence alignments show that the T. cruzi ortholog contains all three of the zinc-binding residues and several residues uniquely conserved in the β subunit of PGGT-I. Co-expression of this protein and the α subunit of T. cruzi PFT in Sf9 insect cells yielded a dimeric protein that forms a tight complex selectively with [3H]geranylgeranyl pyrophosphate, indicating a key characteristic of a functional PGGT-I. Recombinant T. cruzi PGGT-I ortholog showed geranylgeranyltransferase activity with distinct specificity toward the C-terminal CaaX motif of protein substrates compared to that of the mammalian PGGT-I and T. cruzi PFT. Most of the CaaX-containing proteins with X=Leu are good substrates of T. cruzi PGGT-I, and those with X=Met are substrates for both T. cruzi PFT and PGGT-I, whereas unlike mammalian PGGT-I, those with X=Phe are poor substrates for T. cruzi PGGT-I. Several candidates for T. cruzi PGGT-I or PFT substrates containing the C-terminal CaaX motif are found in the T. cruzi gene database. Among five C-terminal peptides of those tested, a peptide of a Ras-like protein ending with CVLL was selectively geranylgeranylated by T. cruzi PGGT-I. Other peptides with CTQQ (Tcj2 DNAJ protein), CAVM (TcPRL-1 protein tyrosine phosphatase), CHFM (a small GTPase like protein), and CQLF (TcRho1 GTPase) were specific substrates for T. cruzi PFT but not for PGGT-I. The mRNA and protein of the T. cruzi PGGT-I β ortholog were detected in three life-cycle stages of T. cruzi. Cytosol fractions from

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

    PubMed Central

    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 intracellular processes are poorly understood. As a part of an effort to understand parasite signaling functions, we report the results of a genome-wide analysis of protein kinases (PKs) of these three trypanosomatids. Results Bioinformatic searches of the trypanosomatid genomes for eukaryotic PKs (ePKs) and atypical PKs (aPKs) revealed a total of 176 PKs in T. brucei, 190 in T. cruzi and 199 in L. major, most of which are orthologous across the three species. This is approximately 30% of the number in the human host and double that of the malaria parasite, Plasmodium falciparum. The representation of various groups of ePKs differs significantly as compared to humans: trypanosomatids lack receptor-linked tyrosine and tyrosine kinase-like kinases, although they do possess dual-specificity kinases. A relative expansion of the CMGC, STE and NEK groups has occurred. A large number of unique ePKs show no strong affinity to any known group. The trypanosomatids possess few ePKs with predicted transmembrane domains, suggesting that receptor ePKs are rare. Accessory Pfam domains, which are frequently present in human ePKs, are uncommon in trypanosomatid ePKs. Conclusion Trypanosomatids possess a large set of PKs, comprising approximately 2% of each genome, suggesting a key role for phosphorylation in parasite biology. Whilst it was possible to place most of the trypanosomatid ePKs into the seven established groups using bioinformatic analyses, it has not been possible to ascribe function

  6. Cytochrome oxidase subunit 2 gene allows simultaneous detection and typing of Trypanosoma rangeli and Trypanosoma cruzi.

    PubMed

    de Sá, Amanda Regina Nichi; Steindel, Mário; Demeu, Lara Maria Kalempa; Lückemeyer, Débora Denardin; Grisard, Edmundo Carlos; Neto, Quirino Alves de Lima; de Araújo, Silvana Marques; Toledo, Max Jean de Ornelas; Gomes, Mônica Lúcia

    2013-12-23

    The parasites Trypanosoma rangeli and Trypanosoma cruzi share vectors and hosts over a wide geographical area in Latin America. In this study, we propose a single molecular approach for simultaneous detection and typing of T. rangeli and T. cruzi. A restriction fragment length polymorphism analysis of the mitochondrial cytochrome oxidase II gene (COII-RFLP) using enzyme AluI and different amounts of DNA from the major genetic groups of T. rangeli and T. cruzi (KP1+/KP1- and DTU-I/DTU-II) was carried out. The same marker was tested on the other T. cruzi DTUs (DTU-III to DTU-VI) and on DNA extracted from gut contents of experimentally infected triatomines. The COII PCR generates a ~400 bp fragment, which after digestion with AluI (COII-RFLP) can be used to distinguish T. rangeli from T. cruzi and simultaneously differentiate the major genetic groups of T. rangeli (KP1+ and KP1-) and T. cruzi (DTU-I and DTU-II). The COII-RFLP generated bands of ~120 bp and ~280 bp for KP1+, whereas for KP1- no amplicon cleavage was observed. For T. cruzi, digestion of COII revealed a ~300 bp band for DTU-I and a ~250 bp band for DTU-II. For DTU-III to DTU-VI, COII-RFLP generated bands ranging from ~310 to ~330 bp, but the differentiation of these DTUs was not as clear as the separation between DTU-I and DTU-II. After AluI digestion, a species-specific fragment of ~80 bp was observed for all DTUs of T. cruzi. No cross-amplification was observed for Leishmania spp., T. vivax or T. evansi. The COII-RFLP allowed simultaneous detection and typing of T. rangeli and T. cruzi strains according to their major genetic groups (KP1+/KP1- and DTU-I/DTU-II) in vitro and in vivo, providing a reliable and sensitive tool for epidemiological studies in areas where T. rangeli and T. cruzi coexist.

  7. Effective gene delivery to Trypanosoma cruzi epimastigotes through nucleofection.

    PubMed

    Pacheco-Lugo, Lisandro; Díaz-Olmos, Yirys; Sáenz-García, José; Probst, Christian Macagnan; DaRocha, Wanderson Duarte

    2017-06-01

    New opportunities have raised to study the gene function approaches of Trypanosoma cruzi after its genome sequencing in 2005. Functional genomic approaches in Trypanosoma cruzi are challenging due to the reduced tools available for genetic manipulation, as well as to the reduced efficiency of the transient transfection conducted through conventional methods. The Amaxa nucleofector device was systematically tested in the present study in order to improve the electroporation conditions in the epimastigote forms of T. cruzi. The transfection efficiency was quantified using the green fluorescent protein (GFP) as reporter gene followed by cell survival assessment. The herein used nucleofection parameters have increased the survival rates (>90%) and the transfection efficiency by approximately 35%. The small amount of epimastigotes and DNA required for the nucleofection can turn the method adopted here into an attractive tool for high throughput screening (HTS) applications, and for gene editing in parasites where genetic manipulation tools remain relatively scarce. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. [Trypanosoma cruzi: transport of essential metabolites acquired from the host].

    PubMed

    Pereira, Claudio A; Carrillo, Carolina; Miranda, Mariana R; Bouvier, León A; Cánepa, Gaspar E

    2008-01-01

    Trypanosoma cruzi is the etiological agent of Chagas disease, a disease endemic not only in Argentina but also in all of Latin America. 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 amino acid 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.

  9. Congenital Transmission of Trypanosoma cruzi Infection in Argentina

    PubMed Central

    Segura, Elsa L.; Cohen, Joel E.

    2003-01-01

    Trypanosoma cruzi, the causative agent of Chagas disease, infects 10–18 million people and may be transmitted to the newborn. Using various data sources, we estimated that nearly 850 congenital cases occurred in Argentina in 1993, or 6.3 expected cases per each reported case in 1994 and in 1994–2001. The congenital transmission of T. cruzi constitutes a sizeable public health problem in the region. PMID:12533278

  10. The detection of phosphonolipids in the protozoan Trypanosoma cruzi

    PubMed Central

    Ferguson, Michael A. J.; Allen, Anthony K.; Snary, David

    1982-01-01

    2-Aminoethylphosphonate was detected in the acid hydrolysates of the phosphonolipids and the lipopeptidophosphoglycan of Trypanosoma cruzi, the causative agent of Chagas' disease. This finding represents the first evidence of phosphonolipids in a zooflagellate. By comparison, no phosphonolipids were detected in Trypanosama brucei, indicating that phosphonolipids are not a ubiquitous feature of the Order Kinetoplastidia. ImagesFig. 1.Fig. 2.Fig. 3. PMID:6758765

  11. Human and sylvatic Trypanosoma cruzi infection in California.

    PubMed Central

    Navin, T R; Roberto, R R; Juranek, D D; Limpakarnjanarat, K; Mortenson, E W; Clover, J R; Yescott, R E; Taclindo, C; Steurer, F; Allain, D

    1985-01-01

    In August 1982, a 56-year-old woman from Lake Don Pedro, California, developed acute Chagas' disease (American trypanosomiasis). She had not traveled to areas outside the United States with endemic Chagas' disease, she had never received blood transfusions, and she did not use intravenous drugs. Trypanosoma cruzi cultured from the patient's blood had isoenzyme patterns and growth characteristics similar to T. cruzi belonging to zymodeme Z1. Triatoma protracta (a vector of Trypanosoma cruzi) infected with T. cruzi were found near the patient's home, a trypanosome resembling T. cruzi was cultured from the blood of two of 19 ground squirrels (Spermophilus beecheyi), and six of 10 dogs had antibody to T. cruzi. A serosurvey of three groups of California residents revealed antibody to T. cruzi by complement fixation in six of 237 (2.5 per cent) individuals living near the patient and in 12 of 1,706 (0.7 per cent) individuals living in a community 20 miles northeast of the patient's home, but in only one of 637 (0.2 per cent) blood donors from the San Francisco Bay area. This is the first case of indigenously acquired Chagas' disease reported from California and the first case recognized in the United States since 1955. This investigation suggests that transmission of sylvatic Trypanosoma cruzi infection to humans occurs in California but that Chagas' disease in humans is rare. PMID:3919598

  12. Characterization of a RAB5 homologue in Trypanosoma cruzi.

    PubMed

    Araripe, Júlia Rolão; Ramos, Fabiane Pereira; Cunha e Silva, Narcisa Leal; Urményi, Turán Péter; Silva, Rosane; Leite Fontes, Carlos Frederico; da Silveira, José Franco; Rondinelli, Edson

    2005-04-08

    RAB proteins are small GTPases involved in exocytic and endocytic pathways of eukaryotic cells, controlling vesicle docking and fusion. RABs show a remarkable specificity in subcellular localization, so they can be used as molecular markers for studying protein trafficking in Trypanosoma cruzi, the causal agent of Chagas' disease. RAB5 is a component of early endosomes. It has been identified in kinetoplastids such as Trypanosoma brucei and Leishmania donovani. In this work, we describe the characterization of the complete coding sequence of a RAB5 gene homologue in T. cruzi (TcRAB5, GenBank Accession No. AY730667). It is present as a single copy gene, located at chromosomal bands XIII and XIV. TcRAB5 shares the highest degrees of similarity (71%) and identity (63%) with Trypanosoma brucei rhodesiense RAB5a and contains all five characteristic RAB motifs. TcRAB5 is transcribed as a single 1.5kb mRNA in epimastigotes. Its transcript was also detected in the other two forms of the parasite, metacyclic trypomastigotes and spheromastigotes. The recombinant TcRAB5 protein was able to bind and hydrolyze GTP. The identification of proteins involved in T. cruzi endo- and exocytic pathways may generate cellular compartment markers, an invaluable tool to better understand the vesicular transport in this parasite.

  13. Regulation and spatial organization of PCNA in Trypanosoma brucei

    SciTech Connect

    Kaufmann, Doris; Gassen, Alwine; Maiser, Andreas; Leonhardt, Heinrich; Janzen, Christian J.

    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

    PubMed Central

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

    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. Domestic dogs, free from T. cruziinfection, received three immunisations with fixed T. rangeliepimastigotes. Dogs were not challenged with T. cruzi, but they were left in their environment. This immunisation induced antibodies againstT. cruzi for more than three years in dogs in their natural habitat, while control dogs remained serologically negative. PMID:27074257

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

    PubMed

    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.

  16. Effects of astaxanthin in mice acutely infected with Trypanosoma cruzi.

    PubMed

    Contreras-Ortiz, José María Eloy; Barbabosa-Pliego, Alberto; Oros-Pantoja, Rigoberto; Aparicio-Burgos, José Esteban; Zepeda-Escobar, José Antonio; Hassan-Moustafa, Wael Hegazy; Ochoa-García, Laucel; Uxúa Alonso-Fresan, María; Tenorio Borroto, Esvieta; Vázquez-Chagoyán, Juan Carlos

    2017-01-01

    During Trypanosoma cruzi infection, oxidative stress is considered a contributing factor for dilated cardiomyopathy development. In this study, the effects of astaxanthin (ASTX) were evaluated as an alternative drug treatment for Chagas disease in a mouse model during the acute infection phase, given its anti-inflammatory, immunomodulating, and anti-oxidative properties. ASTX was tested in vitro in parasites grown axenically and in co-culture with Vero cells. In vivo tests were performed in BALB/c mice (4-6 weeks old) infected with Trypanosoma cruzi and supplemented with ASTX (10 mg/kg/day) and/or nifurtimox (NFMX; 100 mg/kg/day). Results show that ASTX has some detrimental effects on axenically cultured parasites, but not when cultured with mammalian cell monolayers. In vivo, ASTX did not have any therapeutic value against acute Trypanosoma cruzi infection, used either alone or in combination with NFMX. Infected animals treated with NFMX or ASTX/NFMX survived the experimental period (60 days), while infected animals treated only with ASTX died before day 30 post-infection. ASTX did not show any effect on the control of parasitemia; however, it was associated with an increment in focal heart lymphoplasmacytic infiltration, a reduced number of amastigote nests in cardiac tissue, and less hyperplasic spleen follicles when compared to control groups. Unexpectedly, ASTX showed a negative effect in infected animals co-treated with NFMX. An increment in parasitemia duration was observed, possibly due to ASTX blocking of free radicals, an anti-parasitic mechanism of NFMX. In conclusion, astaxanthin is not recommended during the acute phase of Chagas disease, either alone or in combination with nifurtimox. © J.M.E. Contreras-Ortiz et al., published by EDP Sciences, 2017.

  17. [Immunosuppressor effect of Trypanosoma musculi (Mastigophora: Trypanosomatidae) on experimental toxoplasmosis].

    PubMed

    Piccolo-Johanning, Loretta; Kellerman-Guterman, Vivian; Valerio-Campos, Idalia; Chinchilla-Carmona, Misael

    2013-06-01

    The immunosuppression caused by species of the gender Trypanosoma has been widely documented. The influence over experimental infections with Toxoplasma gondii is evident when using Trypanosoma lewisi, a natural parasite of white rats. We decided to test the effect of Trypanosoma musculi from mice, an organism with very similar biological characteristics to T. lewisi, to see if this trypanosomatid could induce a similar effect. Four groups of Swiss mice were inoculated with T. musculi previously to infection with T. gondii, and we determined the survival of the animals, as well as the number of cysts developed in the brain of survivors. We isolated and tested different strains of T. gondii from different sources. In a first experiment, the animals were previously inoculated with T. musculi at different times prior to the infection with Toxoplasma; this allowed us to determine that the immunosuppression process resulted more evident when T. musculi inoculation was made four days before. In a second experiment, we used different inoculi dose and found that it did not influenced the process. Furthermore, the results were negative when evaluating if the amount of the inoculated trypomastigote influenced the process. In order to demonstrate if there were differences in the immnosuppressive effect, related to Toxoplasma strains, groups of mice were inoculated with brain cysts of TFC, TLP, TLW and TBT strains. Excluding the TLP strain, that resulted to be very pathogenic regardless the previous inoculation with T. lewisi, the other strains kept the same pattern of immunosuppression in mice, whose survival time was shorter as the presence of cysts in the brain was higher. These observations were in agreement with an experimental immunosuppression model, associated with immunosuppressive diseases, specially cancer and AIDS.

  18. Effects of astaxanthin in mice acutely infected with Trypanosoma cruzi

    PubMed Central

    Contreras-Ortiz, José María Eloy; Barbabosa-Pliego, Alberto; Oros-Pantoja, Rigoberto; Aparicio-Burgos, José Esteban; Zepeda-Escobar, José Antonio; Hassan-Moustafa, Wael Hegazy; Ochoa-García, Laucel; Uxúa Alonso-Fresan, María; Tenorio Borroto, Esvieta; Vázquez-Chagoyán, Juan Carlos

    2017-01-01

    During Trypanosoma cruzi infection, oxidative stress is considered a contributing factor for dilated cardiomyopathy development. In this study, the effects of astaxanthin (ASTX) were evaluated as an alternative drug treatment for Chagas disease in a mouse model during the acute infection phase, given its anti-inflammatory, immunomodulating, and anti-oxidative properties. ASTX was tested in vitro in parasites grown axenically and in co-culture with Vero cells. In vivo tests were performed in BALB/c mice (4–6 weeks old) infected with Trypanosoma cruzi and supplemented with ASTX (10 mg/kg/day) and/or nifurtimox (NFMX; 100 mg/kg/day). Results show that ASTX has some detrimental effects on axenically cultured parasites, but not when cultured with mammalian cell monolayers. In vivo, ASTX did not have any therapeutic value against acute Trypanosoma cruzi infection, used either alone or in combination with NFMX. Infected animals treated with NFMX or ASTX/NFMX survived the experimental period (60 days), while infected animals treated only with ASTX died before day 30 post-infection. ASTX did not show any effect on the control of parasitemia; however, it was associated with an increment in focal heart lymphoplasmacytic infiltration, a reduced number of amastigote nests in cardiac tissue, and less hyperplasic spleen follicles when compared to control groups. Unexpectedly, ASTX showed a negative effect in infected animals co-treated with NFMX. An increment in parasitemia duration was observed, possibly due to ASTX blocking of free radicals, an anti-parasitic mechanism of NFMX. In conclusion, astaxanthin is not recommended during the acute phase of Chagas disease, either alone or in combination with nifurtimox. PMID:28560955

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

    PubMed

    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. Virulence factors of Trypanosoma cruzi: who is who?

    PubMed

    Osorio, Luis; Ríos, Isabel; Gutiérrez, Bessy; González, Jorge

    2012-12-01

    The aim of this review is to gather the current knowledge of Trypanosoma cruzi's virulence factors described to date in an integrative way, relating these with the parasite's life cycle and trying to elucidate their importance in each process. Several aspects relevant for the parasite's survival, such as invasion, resistance to oxidative damage, escape from the phagolysosomal vacuole and differentiation, among others, will be discussed. However, there is still a lot to learn about what virulence really means in T. cruzi and which parasite molecules are absolutely required to make T. cruzi one of the most successful pathogens to invade, survive and persist in a mammalian host.

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

    NASA Astrophysics Data System (ADS)

    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.

  2. Trypanosoma cruzi infection in B-cell-deficient rats.

    PubMed Central

    Rodriguez, A M; Santoro, F; Afchain, D; Bazin, H; Capron, A

    1981-01-01

    The effect of neonatally initiated injections of anti-mu rabbit antiserum on immunity of rats against Trypanosoma cruzi infection was investigated in vivo. Anti-mu treatment resulted in a loss of immunoglobulin M (IgM) and IgG2a synthesis and, subsequently, of antibody production. These rats so treated were shown to be significantly more susceptible to the acute phase of the infection than the control rats treated with normal rabbit serum, as measured by increased parasitemia and mortality. These results indicate the essential role of antibodies, probably in association with complement or effector cells or both, in immunity to acute Chagas' disease. PMID:6783543

  3. Design and evaluation of Trypanosoma brucei metacaspase inhibitors.

    PubMed

    Berg, Maya; Van der Veken, Pieter; Joossens, Jurgen; Muthusamy, Venkatraj; Breugelmans, Matthias; Moss, Catherine X; Rudolf, Jana; Cos, Paul; Coombs, Graham H; Maes, Louis; Haemers, Achiel; Mottram, Jeremy C; Augustyns, Koen

    2010-03-15

    Metacaspase (MCA) is an important enzyme in Trypanosoma brucei, absent from humans and differing significantly from the orthologous human caspases. Therefore MCA constitutes a new attractive drug target for antiparasitic chemotherapeutics, which needs further characterization to support the discovery of innovative drug candidates. A first series of inhibitors has been prepared on the basis of known substrate specificity and the predicted catalytic mechanism of the enzyme. In this Letter we present the first inhibitors of TbMCA2 with low micromolar enzymatic and antiparasitic activity in vitro combined with low cytotoxicity.

  4. Non-natural acetogenin analogues as potent Trypanosoma brucei inhibitors

    PubMed Central

    Florence, Gordon J.; Fraser, Andrew L.; Gould, Eoin R.; King, Elizabeth F.; Menzies, Stefanie K.; Morris, Joanne C.; Tulloch, Lindsay B.; Smith, Terry K.

    2015-01-01

    A series of novel bis-tetrahydropyran 1,4-triazole analogues based on the acetogenin framework display low micromolar trypanocidal activities towards both bloodstream and insect forms of Trypanosoma brucei, the causative agent of African sleeping sickness. A divergent synthetic strategy was adopted for the synthesis of the key tetrahydropyran intermediates to enable rapid access to diastereochemical variation either side of the 1,4-triazole core. The resulting diastereomeric analogues displayed varying degrees of trypanocidal activity and selectivity in structure activity relationship studies. PMID:25145275

  5. Thermal-unfolding reaction of triosephosphate isomerase from Trypanosoma cruzi.

    PubMed

    Mixcoha-Hernández, Edgar; Moreno-Vargas, Liliana M; Rojo-Domínguez, Arturo; Benítez-Cardoza, Claudia G

    2007-10-01

    Thermal denaturation of triosephosphate isomerase from Trypanosoma cruzi was studied by circular dicrhoism and fluorescence spectroscopies. The unfolding transition was found to be highly irreversible even at the very early stages of the reaction. Kinetic studies, allowed us to identify consecutive reactions. Firstly, only the tryptophan environment is altered. Next, changes on the secondary structure and hydrophobic surface exposure measured by 1-anilino-8-naphthalenesulfonate (ANS) binding were observed. Further conformational changes imply additional modifications on the secondary and tertiary structures and release of the hydrophobic dye leading to the formation of the unfolded state that is prone to aggregate.

  6. Sexual transmission of Trypanosoma cruzi in murine model.

    PubMed

    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. Copyright © 2015 Elsevier Inc. All rights reserved.

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

    PubMed

    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.

  8. First Draft Genome Sequence of the Dourine Causative Agent: Trypanosoma Equiperdum Strain OVI

    PubMed Central

    Hébert, Laurent; Moumen, Bouziane; Madeline, Anthony; Steinbiss, Sascha; Lakhdar, Latifa; Van Reet, Nick; Büscher, Philippe; Laugier, Claire; Cauchard, Julien; Petry, Sandrine

    2017-01-01

    Trypanosoma equiperdum is the causative agent of dourine, a sexually-transmitted infection of horses. This parasite belongs to the subgenus Trypanozoon that also includes the agent of sleeping sickness (Trypanosoma brucei) and surra (Trypanosoma evansi). We herein report the genome sequence of a T. equiperdum strain OVI, isolated from a horse in South-Africa in 1976. This is the first genome sequence of the T. equiperdum species, and its availability will provide important insights for future studies on genetic classification of the subgenus Trypanozoon. PMID:28138343

  9. Characterization of the M32 metallocarboxypeptidase of Trypanosoma brucei: differences and similarities with its orthologue in Trypanosoma cruzi

    PubMed Central

    Frasch, Alejandra P.; Carmona, Adriana K.; Juliano, Luiz; Cazzulo, Juan J.; Niemirowicz, Gabriela T.

    2012-01-01

    Metallocarboxypeptidases (MCP) of the M32 family of peptidases have been identified in a number of prokaryotic organisms but they are absent from eukaryotic genomes with the remarkable exception of those of trypanosomatids. The genome of Trypanosoma brucei, the causative agent of Sleeping Sickness, encodes one such MCP which displays 72% identity to the characterized TcMCP-1 from Trypanosoma cruzi. As its orthologue, TcMCP-1, Trypanosoma brucei MCP is a cytosolic enzyme expressed in both major stages of the parasite. Purified recombinant TbMCP-1 exhibits a significant hydrolytic activity against the carboxypeptidase B substrate FA (furylacryloil)-Ala-Lys at pH 7.0–7.8 resembling the T. cruzi enzyme. S everal divalent cations had little effect on TbMCP-1 activity but increasing amounts of Co2+ inhibited the enzyme. Despite having similar tertiary structure, both protozoan MCPs display different substrate specificity with respect to P1 position. Thus, TcMCP-1 enzyme cleaved Abz-FVK-(Dnp)-OH substrate (where Abz: o-aminobenzoic acid and Dnp: 2,4-dinitrophenyl) whereas TbMCP-1 had no activity on this substrate. Comparative homology models and sequence alignments using TcMCP-1 as a template led us to map several residues that could explain this difference. To verify this hypothesis, site-directed mutagenesis was undertaken replacing the TbMCP-1 residues by those present in TcMCP-1. We found that the substitution A414M led TbMCP-1 to gain activity on Abz-FVK-(Dnp)-OH, thus showing that this residue is involved in specificity determination, probably being part of the S1 sub-site. Moreover, the activity of both protozoan MCPs was explored on two vasoactive compounds such as bradykinin and angiotensin I resulting in two different hydrolysis patterns. PMID:22575602

  10. In vitro drug susceptibility of two strains of the wildlife trypanosome, Trypanosoma copemani: A comparison with Trypanosoma cruzi.

    PubMed

    Botero, Adriana; Keatley, Sarah; Peacock, Christopher; Thompson, R C Andrew

    2017-04-01

    Trypanosomes are blood protozoan parasites that are capable of producing illness in the vertebrate host. Within Australia, several native Trypanosoma species have been described infecting wildlife. However, only Trypanosoma copemani has been associated with pathological lesions in wildlife hosts and more recently has been associated with the drastic decline of the critically endangered woylie (Bettongia penicillata). The impact that some trypanosomes have on the health of the vertebrate host has led to the development of numerous drug compounds that could inhibit the growth or kill the parasite. This study investigated and compared the in vitro susceptibility of two strains of T. copemani (G1 and G2) and one strain of Trypanosoma cruzi (10R26) against drugs that are known to show trypanocidal activity (benznidazole, posaconazole, miltefosine and melarsoprol) and against four lead compounds, two fenarimols and two pyridine derivatives (EPL-BS1937, EPL-BS2391, EPL-BS0967, and EPL-BS1246), that have been developed primarily against T.cruzi. The in vitro cytotoxicity of all drugs against L6 rat myoblast cells was also assessed. Results showed that both strains of T. copemani were more susceptible to all drugs and lead compounds than T. cruzi, with all IC50 values in the low and sub-μM range for both species. Melarsoprol and miltefosine exhibited the highest drug activity against both T. copemani and T. cruzi, but they also showed the highest toxicity in L6 cells. Interestingly, both fenarimol and pyridine derivative compounds were more active against T. copemani and T. cruzi than the reference drugs benznidazole and posaconazole. T. copemani strains exhibited differences in susceptibility to all drugs demonstrating once again considerable differences in their biological behaviour.

  11. Refractory hypoglycaemia in a dog infected with Trypanosoma congolense

    PubMed Central

    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. PMID:26795063

  12. Catabolism of proline by procyclic culture forms of Trypanosoma congolense.

    PubMed

    Obungu, V H; Kiaira, J K; Njogu, R M; Olembo, N K

    1999-05-01

    The effect of various metabolic inhibitors on the rate of oxygen consumption by procyclic culture forms of Trypanosoma congolense utilizing proline as substrate was investigated. Cyanide inhibited the rate of oxygen consumption by 81.0 +/- 6.7%, malonate inhibited the rate by 51.6 +/- 1.6% and Antimycin A by 73.1 +/- 5.9%. A combination of cyanide and malonate inhibited the rate of oxygen consumption by 84.9 +/- 6.7% while a combination of antimycin A and malonate inhibited the rate by 81.6 +/- 7.6%. Rotenone had no effect on the rate of respiration except when the intact cells were first permeabilized by digitonin after which rotenone decreased the rate of respiration by 20-30%. Salicylhydroxamate (SHAM) did not have any effect on the rate of oxygen consumption. Enzymes involved in the catabolism of proline with high activities were: proline dehydrogenase, alpha-ketoglutarate dehydrogenase, succinate dehydrogenase, fumarase, NADP-linked malic enzyme, alanine aminotransferase and malate dehydrogenase. Activities of 1-pyrroline-5 carboxylate dehydrogenase, glutamate dehydrogenase, aspartate aminotransferase and NAD-linked malic enzyme were detectable but lower. The end products of proline catabolism were alanine and glutamate. Unlike the case in Trypanosoma brucei brucei aspartate was not detected. Possible pathways of proline catabolism in procyclic culture forms of T. congolense and of electron transfer are proposed.

  13. Trypanosoma cruzi antibodies in blood donors in Yucatan state, Mexico.

    PubMed

    García-Montalvo, Beatriz

    2011-01-01

    Blood transfusion is the second most frequent way of Trypanosoma cruzi (T. cruzi) transmission in Latin American countries. Few data exists on the geographic distribution and prevalence of T. cruzi seropositive blood donors in Mexico. The objective was to document T. cruzi antibody distribution, and identify the regions with the highest prevalence of seropositive blood donors. the analyzed data was collected over a six-year period during blood donations made at the Central Blood Bank and at the transfusion services and donation modules of the Instituto Mexicano del Seguro Social (IMSS) located in the Yucatan state. Trypanosoma cruzi antibody reactivity was determined in 86343 blood donors. Overall seroprevalence was 0.70 % (607/86 343). Since 2002 to 2004, the majority (58 %) of seropositive donors were rural residents, but since 2005 to 2007 the majority (56.6 %) were urban residents. The two highest seroprevalences by region were in the Metropolitan area (0.42 %) and in rural south Yucatan (0.09 %). Most seropositive donors resided in the municipality of Merida (60.3 %). seroprevalence distribution was heterogeneous during the study period but urban transmission has apparently surpassed rural transmission in recent years.

  14. Inositolphosphoceramide Metabolism in Trypanosoma cruzi as Compared to other Trypanosomatids

    PubMed Central

    DE LEDERKREMER, ROSA M.; AGUSTI, ROSALÍA; DOCAMPO, ROBERTO

    2012-01-01

    Chagas disease is caused by Trypanosoma cruzi and is endemic to North, Central and South American countries. Current therapy against this disease is only partially effective and produces adverse side effects. Studies on the metabolic pathways of T. cruzi, in particular those with no equivalent in mammalian cells, might identify targets for the development of new drugs. Ceramide is metabolized to inositolphosphoceramide (IPC) in T. cruzi and other kinetoplastid protists whereas in mammals it is mainly incorporated into sphingomyelin. In T. cruzi, in contrast to Trypanosoma brucei and Leishmania spp., IPC functions as lipid anchor constituent of glycoproteins and free glycosylinositolphospholipids (GIPLs). Inhibition of IPC and GIPLs biosynthesis impairs differentiation of trypomastigotes into the intracellular amastigote forms. The gene encoding IPC synthase in T. cruzi has been identified and the enzyme has been expressed in a cell-free system. The enzyme involved in IPC degradation and the remodelases responsible for the incorporation of ceramide into free GIPLs or into the glycosylphosphatidyl inositols (GPIs) anchoring glycoproteins, and in fatty acid modifications of these molecules of T. cruzi have been understudied. IPC metabolism and remodeling could be exploited as targets for Chagas disease chemotherapy. PMID:21332877

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

    PubMed

    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.

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

    PubMed

    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.

  17. New Imidazole-Based Compounds Active Against Trypanosoma cruzi.

    PubMed

    Adeyemi, Oluyomi Stephen; Molina, María Teresa; Eseola, Abiodun Omokehinde; Fonseca-Berzal, Cristina; Gómez-Barrio, Alicia

    2017-01-01

    Current drugs available for the treatment of Chagas disease are fraught with several challenges including severe toxicity and limited efficacy. These factors coupled with the absence of effective drugs for treating the chronic stage of the disease have rendered the development of new drugs against Chagas disease a priority. This study screened several imidazole-based compounds for anti-Trypanosoma potential. Using an in vitro experimental infection model, several imidazole-based compounds were screened for anti-proliferative effect on Trypanosoma cruzi epimastigotes. Additionally, all test compounds were evaluated for unspecific cytotoxicity on L929 murine fibroblasts. Benznidazole (BZN) served as reference drug. All test compounds demonstrated interesting trypanocidal potential with IC50 values in the μM range (1< 1C50 <8 μM). The activities of the test compounds compared favorably with BZN, which had an IC50 value ca. 30 μM. Conversely, most of the test compounds were highly cytotoxic, resulting in selectivity lower than that of BZN (SI > 9.42). We provide evidence which implicate the imidazole-based compounds as potential prototypes for the development of anti-parasitic agents. Findings have far-reaching relevance to drug discovery efforts for trypanosomiasis. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  18. Quercetin, a fluorescent bioflavanoid, inhibits Trypanosoma brucei hexokinase 1

    PubMed Central

    Dodson, Heidi C.; Lyda, Todd L.; Chambers, Jeremy W.; Morris, Meredith T.; Christensen, Kenneth A.; Morris, James C.

    2010-01-01

    Hexokinases from the African trypanosome, Trypanosoma brucei, are attractive targets for the development of anti-parasitic drugs, in part because the parasite utilizes glycolysis exclusively for ATP production during the mammalian infection. Here, we have demonstrated that the bioflavanoid quercetin (QCN), a known trypanocide, is a mixed inhibitor of Trypanosoma brucei hexokinase 1 (TbHK1) (IC50 = 4.1 ± 0.8 μM). Spectroscopic analysis of QCN binding to TbHK1, taking advantage of the intrinsically fluorescent single tryptophan (Trp177) in TbHK1, revealed that QCN quenches emission of Trp177, which is located near the hinge region of the enzyme. ATP similarly quenched Trp177 emission, while glucose had no impact on fluorescence. Supporting the possibility that QCN toxicity is a consequence of inhibition of the essential hexokinase, in live parasites QCN fluorescence localizes to glycosomes, the subcellular home of TbHK1. Additionally, RNAi-mediated silencing of TbHK1 expression expedited QCN induced death, while over-expressing TbHK1 protected trypanosomes from the compound. In summary, these observations support the suggestion that QCN toxicity is in part attributable to inhibition of the essential TbHK1. PMID:20971104

  19. A nested PCR for the ssrRNA gene detects Trypanosoma binneyi in the platypus and Trypanosoma sp. in wombats and kangaroos in Australia.

    PubMed

    Noyes, H A; Stevens, J R; Teixeira, M; Phelan, J; Holz, P

    1999-02-01

    Trypanosome infections in their natural hosts are frequently difficult to detect by microscopy, and culture methods are unreliable and not suitable for all species of Trypanosoma. A nested PCR strategy for detecting and identifying Trypanosoma species, suitable for detecting both known and unknown trypanosomes, is presented. Thirty-two blood samples from 23 species of Australian birds and mammals were screened by a nested PCR for the presence of Trypanosoma sp. ssrRNA. Three infections were detected, one in an eastern grey kangaroo (Macropus giganteus), one in a common wombat (Vombatus ursinus) and one in a platypus (Ornithorhynchus anatinus). The kangaroo and wombat are new host records for Trypanosoma sp.; the platypus parasite was Trypanosoma hinneyi. The three parasites could be distinguished by restriction fragment length polymorphisms of the amplified fragment of the ssrRNA gene. The kangaroo and wombat parasites were also isolated in a semi-solid blood agar medium. The culture forms of the kangaroo trypanosome had an expanded flagellar sheath in which structures similar to hemidesmosomes were detected by EM. The nested PCR was at least as sensitive as culture, and analysis of the PCR products gave parasite-specific fingerprints. Therefore this method could be suitable for rapidly screening host animals for the presence of trypanosomes and identifying the infecting strain.

  20. First Report of Trypanosoma sp. in Spectacled Caiman (Caiman crocodilus): Morphological and Phylogenetic Relationships

    PubMed Central

    da Costa, Andrea P.; Acosta, Igor C. L.; de Lima, Julia T. R.; Minervino, Antonio H. H.; Gennari, Solange M.

    2013-01-01

    In Crocodylidae family three trypanosomes species were described, T. grayi in African crocodilian and T. cecili and Trypanosoma sp. in Caimans species from Brazil. T. grayi was transmitted by tsetse flies and the vector of Brazilian caimans trypanosomes is unknown. We characterized first Brazilian trypanosome isolated in spectacled caiman (Caiman crocodilus) from Mato Grosso State in Brazil. Morphological findings in epimastigotes forms from axenic culture showed high similarity with Trypanosoma sp. described in Caiman yacare from Brazilian Pantanal. Phylogenetic studies performed with SSU rDNA and gGAPDH (glyceraldehydes-3-phosphato dehydrogenase glycosomal) clustering in T. grayi Clade and together to genotype Cay 01 from Trypanosoma unnamed species isolated in C. yacare. This is the first isolate of Trypanosoma sp. from C. crocodilus and the phylogenetic position with isolates in C. yacare from Pantanal region and demonstrates the low host specificity of cayman trypanosomes in Brazil. PMID:27335853

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

    PubMed

    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.

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

    PubMed Central

    Velásquez, Norma P.; Camargo, Rocío E.; Uzcanga, Graciela L.; Bubis, José

    2014-01-01

    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. PMID:24757558

  3. Effects of Infection by Trypanosoma cruzi and Trypanosoma rangeli on the Reproductive Performance of the Vector Rhodnius prolixus

    PubMed Central

    Fellet, Maria Raquel; Lorenzo, Marcelo Gustavo; Elliot, Simon Luke; Carrasco, David; Guarneri, Alessandra Aparecida

    2014-01-01

    The insect Rhodnius prolixus is responsible for the transmission of Trypanosoma cruzi, which is the etiological agent of Chagas disease in areas of Central and South America. Besides this, it can be infected by other trypanosomes such as Trypanosoma rangeli. The effects of these parasites on vectors are poorly understood and are often controversial so here we focussed on possible negative effects of these parasites on the reproductive performance of R. prolixus, specifically comparing infected and uninfected couples. While T. cruzi infection did not delay pre-oviposition time of infected couples at either temperature tested (25 and 30°C) it did, at 25°C, increase the e-value in the second reproductive cycle, as well as hatching rates. Meanwhile, at 30°C, T. cruzi infection decreased the e-value of insects during the first cycle and also the fertility of older insects. When couples were instead infected with T. rangeli, pre-oviposition time was delayed, while reductions in the e-value and hatching rate were observed in the second and third cycles. We conclude that both T. cruzi and T. rangeli can impair reproductive performance of R. prolixus, although for T. cruzi, this is dependent on rearing temperature and insect age. We discuss these reproductive costs in terms of potential consequences on triatomine behavior and survival. PMID:25136800

  4. Infestation of Mauritia flexuosa palms by triatomines (Hemiptera: Reduviidae), vectors of Trypanosoma cruzi and Trypanosoma rangeli in the Brazilian savanna.

    PubMed

    Gurgel-Gonçalves, Rodrigo; Cura, Carolina; Schijman, Alejandro G; Cuba, César A Cuba

    2012-02-01

    To determine the infestation and trypanosome infection of triatomines captured in Mauritia flexuosa palm trees across its geographic distribution in the Brazilian savanna (Cerrado), we sampled 42 localities in eight states and in the Federal District, Brazil, between July 2005 and January 2010. Overall, 2154 specimens of the species Rhodnius neglectus, Psammolestes tertius, Triatoma sordida, and Microtriatoma borbai, were collected. Among the 341 palms sampled, 182 (53.3%) were infested with R. neglectus, which resulted in the capture of 1639 specimens (9.0 insects per infested palm). P. tertius occurred in 26 palms (8%), which resulted in the capture of 484 specimens (19 insects per infested palm). T. sordida (n=30) and M. borbai (n=1) occurred in only one location. From 537 R. neglectus examined, 44 were infected (8%) with Trypanosoma rangeli and/or Trypanosoma cruzi (Tc Id). M. flexuosa was previously recognized as a suitable breeding ecotope for R. neglectus in the Brazilian states of Minas Gerais, Goiás, Tocantins and the Federal District. Our results expand this distribution to other states (São Paulo, Bahia, Mato Grosso, Maranhão and Piauí), and also show that this particular palm tree harbors other triatomine species. Finally, we show that R. neglectus plays an important role in maintaining the enzootic circulation of T. cruzi and T. rangeli in the Brazilian savanna.

  5. Periurban Trypanosoma cruzi–infected Triatoma infestans, Arequipa, Peru

    PubMed Central

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

    2006-01-01

    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 2.38× 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. PMID:17073082

  6. Bed Bugs (Cimex lectularius) as Vectors of Trypanosoma cruzi

    PubMed Central

    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

  7. In vitro antitrypanosomal activity of plant terpenes against Trypanosoma brucei.

    PubMed

    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.

  8. Human Trypanosoma cruzi infection and seropositivity in dogs, Mexico.

    PubMed

    Estrada-Franco, Jose G; Bhatia, Vandanajay; Diaz-Albiter, Hector; Ochoa-Garcia, Laucel; Barbabosa, Alberto; Vazquez-Chagoyan, Juan C; Martinez-Perez, Miguel A; Guzman-Bracho, Carmen; Garg, Nisha

    2006-04-01

    We used 5 diagnostic tests in a cross-sectional investigation of the prevalence of Trypanosoma cruzi in Tejupilco municipality, State of Mexico, Mexico. Our findings showed a substantial prevalence of immunoglobulin G (IgG) and IgM antibodies to T. cruzi in human (n = 293, IgG 2.05%, IgM 5.5%, both 7.1%) and dog (n = 114, IgG 15.8%, IgM 11.4%, both 21%) populations. We also found antibodies to T. cruzi (n = 80, IgG 10%, IgM 15%, both 17.5%) in dogs from Toluca, an area previously considered free of T. cruzi. Our data demonstrate the need for active epidemiologic surveillance programs in these regions. A direct correlation (r2 = 0.955) of seropositivity between humans and dogs suggests that seroanalysis in dogs may help identify the human prevalence of T. cruzi infection in these areas.

  9. The inositol phosphate/diacylglycerol signalling pathway in Trypanosoma cruzi.

    PubMed Central

    Docampo, R; Pignataro, O P

    1991-01-01

    Using [32P]Pi and [3H]inositol as precursors, we have detected the presence of phosphatidylinositol, phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate, and their derivatives inositol phosphate, inositol 1,4-bisphosphate and inositol 1,4,5-trisphosphate respectively, in Trypanosoma cruzi epimastigotes. Using digitonin-permeabilized cells it was possible to detect a stimulation in the formation of inositol 1,4,5-trisphosphate and inositol 1,4-bisphosphate as well as an increased generation of diacylglycerol in the presence of 1 mM-CaCl2. These results are consistent with the operation of a functional inositol phosphate/diacylglycerol pathway in T. cruzi, and constitute the first demonstration of the presence and activation of this pathway in a parasitic protozoan. These results also indicate that this pathway is conserved during evolution from lower to higher eukaryotic organisms. Images Fig. 1. PMID:2025225

  10. [Congenital transmission of Trypanosoma cruzi infection in Argentina].

    PubMed

    Sosa-Estani, Sergio

    2005-01-01

    Congenital transmission of Trypanosoma cruzi infection in Argentina has being increasing its relative importance with control of vectorial and transfusional transmission growth. It is for this reason that vertical transmission is seen, in the future, as a continuous source of infected newborns, even with vectorial and transfusional transmission completely controlled. Preventing vertical transmission of T.cruzi is not possible, but it can be precociously detected, permitting mother and child to be incorporated into the medical attention system, and so allowing the newbornś treatment with practically 100% efficacy. It is estimated that between 800 and 1700 children infected with T. cruzi by congenital transmission are born in Argentina, per year. The implementation of an early strategy of detection for an effective and opportune treatment acquires great relevance as a Public Health measure.

  11. Editing the Trypanosoma cruzi genome with zinc finger nucleases.

    PubMed

    Burle-Caldas, Gabriela Assis; Grazielle-Silva, Viviane; Soares-Simões, Melissa; Schumann Burkard, Gabriela; Roditi, Isabel; DaRocha, Wanderson Duarte; Teixeira, Santuza M

    2017-03-01

    Gene function studies in Trypanosoma cruzi, the protozoan parasite that causes Chagas disease, have been hindered by the lack of efficient genetic manipulation protocols. In most organisms, insertion and deletion of DNA fragments in the genome are dependent on the generation of double-stranded DNA break (DSB) and repair. By inducing a site-specific DSB, zinc finger nucleases (ZFNs) have proven to be useful to enhance gene editing in many cell types. Using a pair of ZFNs targeted to the T. cruzi gp72 gene, we were able to generate gp72 knockout parasites with improved efficiency compared to the conventional gene knockout protocol. We also provide evidence that, in T. cruzi, repair of DSBs generated by ZFNs occurs primarily by the homologous recombination pathway.

  12. Predominance of Trypanosoma cruzi I among Panamanian sylvatic isolates.

    PubMed

    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.

  13. The ins and outs of phosphatidylethanolamine synthesis in Trypanosoma brucei.

    PubMed

    Farine, Luce; Bütikofer, Peter

    2013-03-01

    Phospholipids are not only major building blocks of biological membranes but fulfill a wide range of critical functions that are often widely unrecognized. In this review, we focus on phosphatidylethanolamine, a major glycerophospholipid class in eukaryotes and bacteria, which is involved in many unexpected biological processes. We describe (i) the ins, i.e. the substrate sources and biochemical reactions involved in phosphatidylethanolamine synthesis, and (ii) the outs, i.e. the different roles of phosphatidylethanolamine and its involvement in various cellular events. We discuss how the protozoan parasite, Trypanosoma brucei, has contributed and may contribute in the future as eukaryotic model organism to our understanding of phosphatidylethanolamine homeostasis. This article is part of a Special Issue entitled Phospholipids and Phospholipid Metabolism. Copyright © 2012 Elsevier B.V. All rights reserved.

  14. Bed bugs (Cimex lectularius) as vectors of Trypanosoma cruzi.

    PubMed

    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.

  15. Trypanosoma cruzi in Persons without Serologic Evidence of Disease, Argentina

    PubMed Central

    Basquiera, Ana L.; Sembaj, Adela; Aguerri, Ana M.; Reyes, María E.; Omelianuk, Mirtha; Fernández, Ruth A.; Enders, Julio; Palma, Atilio; Barral, José Moreno; Madoery, Roberto J.

    2003-01-01

    Current diagnosis of chronic Chagas disease relies on serologic detection of specific immunoglobulin G against Trypanosoma cruzi. However, the presence of parasites detected by polymerase chain reaction (PCR) in patients without positive conventional serologic testing has been observed. We determined the prevalence and clinical characteristics of persons with seronegative results for T. cruzi DNA detected by PCR in a population at high risk for chronic American trypanosomiasis. We studied a total of 194 persons from two different populations: 110 patients were recruited from an urban cardiology clinic, and 84 persons were nonselected citizens from a highly disease-endemic area. Eighty (41%) of persons had negative serologic findings; 12 (15%) had a positive PCR. Three patients with negative serologic findings and positive PCR results had clinical signs and symptoms that suggested Chagas cardiomyopathy. This finding challenges the current recommendations for Chagas disease diagnosis, therapy, and blood transfusion policies. PMID:14720396

  16. The role of Trypanosoma brucei MRPA in melarsoprol susceptibility.

    PubMed

    Alibu, Vincent P; Richter, Christina; Voncken, Frank; Marti, Gabriela; Shahi, Sanjay; Renggli, Christina Kunz; Seebeck, Thomas; Brun, Reto; Clayton, Christine

    2006-03-01

    We previously showed that over-expression of Trypanosoma brucei MRPA, a member of the multidrug resistance protein family in T. brucei, reproducibly resulted in resistance to the anti-trypanosomal drug melarsoprol in vitro. MRPA is predicted to mediate efflux of melarsoprol as a conjugate with trypanothione, a glutathione-spermidine conjugate which is the major small thiol in trypanosomes. Here, we show that depletion of MRPA by RNA interference resulted in moderate hypersensitivity to both melarsoprol and melarsen oxide. Over-expression of MRPA alone is not sufficient to cause melarsoprol resistance in vivo, although it is sufficient in vitro. This discrepancy is not an effect of drug metabolism since over-expression of MRPA alone conferred resistance to melarsoprol and its principle metabolite, melarsen oxide, in vitro. Over-expression of MRPA was not detected in four melarsoprol-resistant trypanosome isolates from sleeping sickness patients.

  17. Interaction of Trypanosoma cruzi adenylate cyclase with liver regulatory factors.

    PubMed Central

    Eisenschlos, C; Flawiá, M M; Torruella, M; Torres, H N

    1986-01-01

    Trypanosoma cruzi adenylate cyclase catalytic subunits may interact with regulatory factors from rat liver membranes, reconstituting heterologous systems which are catalytically active in assay mixtures containing MgATP. The systems show stimulatory responses to glucagon and guanosine 5'-[beta gamma-imido]triphosphate (p[NH]ppG) or fluoride. Reconstitution was obtained by three different methods: fusion of rat liver membranes (pretreated with N-ethylmaleimide) to T. cruzi membranes; interaction of detergent extracts of rat liver membranes with T. cruzi membranes; or interaction of purified preparations of T. cruzi adenylate cyclase and of liver membrane factors in phospholipid vesicles. The liver factors responsible for the guanine nucleotide effect were characterized as the NS protein. Data also indicate that reconstitution requires the presence of a membrane substrate. PMID:2947568

  18. Effect of isometamidium hydrochloride on Trypanosoma evansi infections in rats.

    PubMed

    Kumar, Upendra; Jas, Ruma; Ghosh, J D

    2009-12-01

    Effect of Isometamidium treatment of rats infected with Trypanosoma evansi was studied under experimental condition. The infection resulted in 100 per cent mortality by 7 day post treatment (DPT) in the infected untreated rats (Gr.II) and Isometamidium-HCl exerted 90 percent anti-trypanosomal efficacy. Haematological studies of all the groups on 3 day post infection (DPI) and on 7 DPT showed that T. evansi infection significantly decreased haematological values, however Isometamidium treatment significantly (P < 0.05) improved Hb and PCV without any significant changes in TEC and TLC values. Serum AST and ALT activities were significantly (P < 0.01) higher in the treated rats compared to healthy rats. Treatment with Isometamidium-HCl significantly improved blood glucose level but decreased creatinine level compared to pretreatment values. There was no significant change in BUN level of treated rats but the value was higher (P < 0.01) compared to healthy rats.

  19. Kinetic modelling of isometamidium chloride (Samorin) uptake by Trypanosoma congolense.

    PubMed

    Sutherland, I A; Mounsey, A; Eisler, M; Holmes, P H

    1992-08-01

    Clones of Trypanosoma congolense which express resistance to the widely used trypanocide isometamidium chloride accumulate less of the drug than clones which are sensitive to drug treatment. A mathematical model has been developed which was able to predict theoretical lines representing the uptake kinetics in trypanosomes which were sensitive to isometamidium, as well as for resistant trypanosomes in which reduced accumulation was a result of either reduced uptake or enhanced efflux of the drug. Data from drug uptake experiments were then fitted to these theoretical lines. While the value for drug efflux could not be separated from the dissociation constant of the trypanosomes for isometamidium, it was demonstrated that reduced accumulation is not a result of reduced uptake of isometamidium by drug-resistant trypanosomes.

  20. Structure of Trypanosoma brucei flagellum accounts for its bihelical motion

    PubMed Central

    Koyfman, Alexey Y.; Schmid, Michael F.; Gheiratmand, Ladan; Fu, Caroline J.; Khant, Htet A.; Huang, Dandan; He, Cynthia Y.; Chiu, Wah

    2011-01-01

    Trypanosoma brucei is a parasitic protozoan that causes African sleeping sickness. It contains a flagellum required for locomotion and viability. In addition to a microtubular axoneme, the flagellum contains a crystalline paraflagellar rod (PFR) and connecting proteins. We show here, by cryoelectron tomography, the structure of the flagellum in three bending states. The PFR lattice in straight flagella repeats every 56 nm along the length of the axoneme, matching the spacing of the connecting proteins. During flagellar bending, the PFR crystallographic unit cell lengths remain constant while the interaxial angles vary, similar to a jackscrew. The axoneme drives the expansion and compression of the PFR lattice. We propose that the PFR modifies the in-plane axoneme motion to produce the characteristic trypanosome bihelical motility as captured by high-speed light microscope videography. PMID:21690369

  1. Trypanosoma cruzi: orchiectomy and dehydroepiandrosterone therapy in infected rats.

    PubMed

    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.

  2. Human Trypanosoma cruzi Infection and Seropositivity in Dogs, Mexico

    PubMed Central

    Estrada-Franco, Jose G.; Bhatia, Vandanajay; Diaz-Albiter, Hector; Ochoa-Garcia, Laucel; Barbabosa, Alberto; Vazquez-Chagoyan, Juan C.; Martinez-Perez, Miguel A.; Guzman-Bracho, Carmen

    2006-01-01

    We used 5 diagnostic tests in a cross-sectional investigation of the prevalence of Trypanosoma cruzi in Tejupilco municipality, State of Mexico, Mexico. Our findings showed a substantial prevalence of immunoglobulin G (IgG) and IgM antibodies to T. cruzi in human (n = 293, IgG 2.05%, IgM 5.5%, both 7.1%) and dog (n = 114, IgG 15.8%, IgM 11.4%, both 21%) populations. We also found antibodies to T. cruzi (n = 80, IgG 10%, IgM 15%, both 17.5%) in dogs from Toluca, an area previously considered free of T. cruzi. Our data demonstrate the need for active epidemiologic surveillance programs in these regions. A direct correlation (r2 = 0.955) of seropositivity between humans and dogs suggests that seroanalysis in dogs may help identify the human prevalence of T. cruzi infection in these areas. PMID:16704811

  3. Biochemical changes in cats infected with Trypanosoma evansi.

    PubMed

    Da Silva, Aleksandro S; Wolkmer, Patricia; Costa, Márcio M; Tonin, Alexandre A; Eilers, Tiago L; Gressler, Lucas T; Otto, Mateus A; Zanette, Régis A; Santurio, Janio M; Lopes, Sonia T A; Monteiro, Silvia G

    2010-07-15

    This study aimed at evaluating biochemical changes of cats (Felis catus) experimentally infected with Trypanosoma evansi. Seven animals were infected with 10(8) blood trypomastigotes per animal and six were used as controls. Blood smears were performed daily for 56 days and the hepatic, renal and muscular parameters in blood serum were evaluated at days 0, 7, 21, 35 and 49. The protozoan was found in the bloodstream 24-48 h post-inoculation (PI) and irregular peaks of parasitemia were observed throughout the experiment. Muscular enzymatic activities (aspartate aminotransferase and creatine kinase) were increased in infected cats compared to controls. Increased concentrations of total proteins and globulins and decreased levels of albumin and albumin/globulin ratio were observed in infected group versus the controls values (P<0.05). No alteration in serum activity of alanine aminotransferase, gamma-glutamyltransferase, creatinine and urea was observed in both groups. Copyright (c) 2010 Elsevier B.V. All rights reserved.

  4. First report of Trypanosoma vegrandis in koalas (Phascolarctos cinereus).

    PubMed

    Barbosa, Amanda; Austen, Jill; Gillett, Amber; Warren, Kristin; Paparini, Andrea; Irwin, Peter; Ryan, Una

    2016-08-01

    The present study describes the first report of Trypanosoma vegrandis in koalas using morphology and sequence analysis of the 18S rRNA gene. The prevalence of T. vegrandis in koalas was 13.6% (6/44). It is likely that the small size of T. vegrandis (<10μm in length), coupled with the difficulties in amplifying DNA of this parasite in mixed infections using trypanosome generic primers, are the reason why this organism has not been identified in koalas until now. This study highlights the importance of further research comprising a larger sample size to determine the prevalence of T. vegrandis in koalas as well as its potential impacts upon this marsupial species' health. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  5. Pathogenesis of anemia in Trypanosoma brucei-infected mice.

    PubMed Central

    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

  6. Cations in body fluids of Trypanosoma brucei in infected rabbits.

    PubMed

    Amole, B O; Thomas, K D; Asemota, D O

    1990-01-01

    Serum copper, magnesium, zinc, calcium and ionized calcium (Ca++) concentrations were compared in 6 rabbits infected with Trypanosoma brucei brucei and 5 uninfected rabbits. There was a significant depletion of Mg and Zn and a significant increase in Cu from about day 10 of infection to the end. There was no change in plasma total calcium or free diffusible calcium. There was a development of kidney damage as shown clinically by proteinuria and urinary loss of magnesium and zinc, and histologically by the observation of hypercellularity in the glomeruli and tubular degeneration. Our findings thus indicate that trypanosomiasis causes kidney damage which may be responsible for the depletion of the cations seen in the study. Some of the clinical manifestations associated with African trypanosomiasis such as convulsions, anaemia, electrocardiographic changes and splenomegaly may therefore be related to these cation changes.

  7. Pathogenesis of anemia in Trypanosoma brucei-infected mice.

    PubMed

    Amole, B O; Clarkson, A B; Shear, H L

    1982-06-01

    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.

  8. Lysophosphatidylcholine: A Novel Modulator of Trypanosoma cruzi Transmission

    PubMed Central

    Silva-Neto, Mário A. C.; Carneiro, Alan B.; Silva-Cardoso, Livia; Atella, Georgia C.

    2012-01-01

    Lysophosphatidylcholine is a bioactive lipid that regulates a large number of cellular processes and is especially present during the deposition and infiltration of inflammatory cells and deposition of atheromatous plaque. Such molecule is also present in saliva and feces of the hematophagous organism Rhodnius prolixus, a triatominae bug vector of Chagas disease. We have recently demonstrated that LPC is a modulator of Trypanosoma cruzi transmission. It acts as a powerful chemoattractant for inflammatory cells at the site of the insect bite, which will provide a concentrated population of cells available for parasite infection. Also, LPC increases macrophage intracellular calcium concentrations that ultimately enhance parasite invasion. Finally, LPC inhibits NO production by macrophages stimulated by live T. cruzi, and thus interferes with the immune system of the vertebrate host. In the present paper, we discuss the main signaling mechanisms that are likely used by such molecule and their eventual use as targets to block parasite transmission and the pathogenesis of Chagas disease. PMID:22132309

  9. Mosaic VSGs and the scale of Trypanosoma brucei antigenic variation.

    PubMed

    Hall, James P J; Wang, Huanhuan; Barry, J David

    2013-01-01

    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.

  10. Trypanosoma cruzi: susceptibility in mice carrying mutant gene lpr (lymphoproliferation).

    PubMed

    Boyer, M H; Hoff, R; Kipnis, T L; Murphy, E D; Roths, J B

    1983-03-01

    There is evidence that autoimmune aberrations may contribute to the immunopathological consequences of Chagas' disease and because of this we sought to determine whether four inbred strains of mice bearing the single autosomal recessive gene, lpr (lymphoproliferation), which controls certain autoimmune manifestations, are particularly susceptible to acute infection with the Y strain of Trypanosoma cruzi. MRL/MpJ-lpr/lpr, C57Bl/6J-lpr/lpr, AKR/J-lpr/lpr, C3H/HeJ-lpr/lpr showed parasitaemias 2-10 times higher when compared to their congenic partners. Mortality was significantly higher in three of the four lpr strains. The results indicate that a single autosomal recessive gene which is associated with autoimmunity can influence susceptibility to acute T. cruzi infection in mice.

  11. Mosaic VSGs and the Scale of Trypanosoma brucei Antigenic Variation

    PubMed Central

    Hall, James P. J.; Wang, Huanhuan; Barry, J. David

    2013-01-01

    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. PMID:23853603

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

    PubMed

    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.

  13. The Oral Antimalarial Drug Tafenoquine Shows Activity against Trypanosoma brucei

    PubMed Central

    Carvalho, Luis; Martínez-García, Marta; Pérez-Victoria, Ignacio; Manzano, José Ignacio; Yardley, Vanessa

    2015-01-01

    The protozoan parasite Trypanosoma brucei causes human African trypanosomiasis, or sleeping sickness, a neglected tropical disease that requires new, safer, and more effective treatments. Repurposing oral drugs could reduce both the time and cost involved in sleeping sickness drug discovery. Tafenoquine (TFQ) is an oral antimalarial drug belonging to the 8-aminoquinoline family which is currently in clinical phase III. We show here that TFQ efficiently kills different T. brucei spp. in the submicromolar concentration range. Our results suggest that TFQ accumulates into acidic compartments and induces a necrotic process involving cell membrane disintegration and loss of cytoplasmic content, leading to parasite death. Cell lysis is preceded by a wide and multitarget drug action, affecting the lysosome, mitochondria, and acidocalcisomes and inducing a depolarization of the mitochondrial membrane potential, elevation of intracellular Ca2+, and production of reactive oxygen species. This is the first report of an 8-aminoquinoline demonstrating significant in vitro activity against T. brucei. PMID:26195527

  14. [Cellular components and placental alkaline phosphatase in Trypanosoma cruzi infection ].

    PubMed

    Sartori, Maria José; Mezzano, Luciana; Lin, Susana; Repossi, Gastón; Fabro, Sofía P

    2005-01-01

    Trypanosoma cruzi induces changes in the protein pattern of human placenta syncytiotrophoblast. Placental alkaline phosphatase (PLAP) is a glycoenzyme anchored to the membrane by a glycosyl-phosphatidylinositol molecule. PLAP activity and its presence was altered by the parasite in cultures of human placental villi and HEp2 cells with T.cruzi. The cells treated before the cultures with agents which affect PILAP or glycosyl-phosphatidylinositol (antibodies, PL-C, genistein, lithium) presented less parasitic invasion than the control ones. It was also observed a modification in the pattern of actine filaments of the host cells infected. We concluded that PLAP would participate in the process of T. cruzi invasion into placental syncitiotrophoblast cells, by a mechanism that involves hydrolysis of the glycosyl-phosphatidylinositol molecules, the activation of tyrosine kinase proteins, the increase of cytosolic calcium and the rearrangement of actine filaments of the host cells.

  15. Polymorphisms in Trypanosoma cruzi: evidence of genetic recombination.

    PubMed

    Bogliolo, A R; Lauria-Pires, L; Gibson, W C

    1996-03-01

    The ploidy of Trypanosoma cruzi is until now undetermined although analysis of isoenzymes, molecular karyotype and DNA content suggest diploidy in a very plastic genome. Also, there has been no convincing demonstration of genetic exchange and it has been proposed that reproduction is clonal. We have compared 18 T cruzi stocks and clones from the same area or host by means of isoenzyme analysis (12 loci) and restriction site polymorphisms in and around three glycolytic genes (glyceraldehyde-3-phosphate dehydrogenase, aldolase and glucosephosphate isomerase). The analysis demonstrated the presence of homozygotes and heterozygotes and is compatible with diploidy for these housekeeping genes. This strongly supports the hypothesis of genetic exchange in T cruzi and further elucidates the genetic diversity within natural T cruzi populations.

  16. Stage specific kinetoplast DNA-binding proteins in Trypanosoma cruzi.

    PubMed

    Zavala-Castro, J E; Acosta-Viana, K; Guzmán-Marín, E; Rosado-Barrera, M E; Rosales-Encina, J L

    2000-09-18

    Knowledge regarding kinetoplast DNA organization in all members of the Trypanosomatid family is incomplete. Recently, the presence of kinetoplast-associated proteins in condensing kDNA networks in Crithidia fasciculata has been described and a role for these proteins in the maintenance of these complex structures was suggested. To investigate the presence of protein components in Trypanosoma cruzi kinetoplast, we previously described seven epimastigote kinetoplast-associated proteins. We report here the existence of kinetoplast binding proteins in amastigote and trypomastigote stages of T. cruzi, which could bind both mini and maxicircles components with a stage specific elements for every infective form of the parasite. We propose three major classes of kinetoplast-associated proteins related to the basic processes of this intricate disc structure and suggest a possible function of these binding proteins in the T. cruzi mitochondrial DNA organization.

  17. Beta-interferon inhibits cell infection by Trypanosoma cruzi

    NASA Technical Reports Server (NTRS)

    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.

  18. The Trypanosoma cruzi Surface, a Nanoscale Patchwork Quilt.

    PubMed

    Mucci, Juan; Lantos, Andrés B; Buscaglia, Carlos A; Leguizamón, María Susana; Campetella, Oscar

    2017-02-01

    The Trypanosoma cruzi trypomastigote membrane provides a major protective role against mammalian host-derived defense mechanisms while allowing the parasite to interact with different cell types and trigger pathogenesis. This surface has been historically appreciated as a rather unstructured 'coat', mainly consisting of a continuous layer of glycolipids and heavily O-glycosylated mucins, occasionally intercalated with different developmentally regulated molecules displaying adhesive and/or enzymatic properties. Recent findings, however, indicate that the trypomastigote membrane is made up of multiple, densely packed and discrete 10-150nm lipid-driven domains bearing different protein composition; hence resembling a highly organized 'patchwork quilt' design. Here, we discuss different aspects underlying the biogenesis, assembly, and dynamics of this cutting-edge fashion outfit, as well as its functional implications. Copyright © 2016 Elsevier Ltd. All rights reserved.

  19. Electron Microscopy Analysis of the Nucleolus of Trypanosoma cruzi

    NASA Astrophysics Data System (ADS)

    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. Trypanosoma evansi: pharmacological evidence of a nicotinic acetylcholine receptor.

    PubMed

    Portillo, R; Bruges, G; Delgado, D; Betancourt, M; Mijares, A

    2010-06-01

    The role of calcium and its relevance have been deeply revised with respect to trypanosomatids, as the mechanism by which calcium enters trypanosomes was, until now, not well understood. There is evidence supporting the presence of a nAChR in another member of the trypanosomatidae family, Trypanosoma cruzi, these receptors being one entry path to calcium ions. The aims of this work were to determine if there was a nicotinic acetylcholine receptor (nAChR) in Trypanosoma evansi, and to subsequently perform a partial pharmacological characterization of this receptor. After being loaded with FURA-2AM, individual cells of T. evansi, were exposed to cholinergic compounds, and the cells displayed a dose-dependent response to carbachol. This observation indicated that a cholinergic receptor may be present in T. evansi. Although a dose-dependent response to muscarine could not be demonstrated, nicotine could promote an incremental dose-dependent response. The relative potency of this specific agonist of nAChR is in agreement with previous reports. The estimated affinity values were a Kd1 value of 29.6+/-5.72 nM and a Kd2 value of 315.9+/-26.6 nM, which is similar to the Kd value reported for the alpha4 nicotinic receptor. The Hill coefficients were determined to be an n1 of 1.2+/-0.3 and an n2 of 4.2+/-1.3. Finally, our calculations indicated that there are about 1020 receptors in each T. evansi parasite, which is approximately 15-fold lower than the number reported in Torpedo californica electric cells. These results suggest the presence of a nAChR in T. evansi, which is able to bind nicotinic ligands and induce calcium signals. Copyright (c) 2010 Elsevier Inc. All rights reserved.

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

    PubMed

    Fajardo, Emanuella Francisco; Cabrine-Santos, Marlene; Ferreira, Keila Adriana Magalhães; Lages-Silva, Eliane; Ramírez, Luis Eduardo; Pedrosa, André Luiz

    2016-01-01

    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. 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). SSL-HU medium improved clone isolation. PCR revealed that T. cruzi strains predominate on mixed-strain plates. PFGE confirmed that isolated parasites share the same molecular karyotype as parental cell lines. SSL-HU medium constitutes a novel tool for obtaining T. cruzi and T. rangeli clonal lineages.

  2. Comparative haematological study of single and mixed infections of mongrel dogs with Trypanosoma congolense and Trypanosoma brucei brucei.

    PubMed

    Ezeokonkwo, Romanus C; Ezeh, I O; Onunkwo, J I; Obi, P O; Onyenwe, I W; Agu, W E

    2010-10-11

    The haematological effects of single and mixed infections of Trypanosoma congolense and Trypanosoma brucei brucei were compared in experimentally infected mongrel dogs. Twenty mongrel dogs of both sexes aged between 3 and 6 months, and weighing between 2.5 and 5.9 kg were used for the study. The dogs were kept in clean metal cages in a fly-proof house and were adequately fed and given water ad libitum. The twenty dogs were divided into four groups of five dogs each. Group I dogs were uninfected control, group II dogs were infected with T. congolense, group III dogs were infected with T. brucei brucei and group IV dogs were infected with both T. congolense and T. brucei brucei. Parasitaemia occurred in the infected dogs in groups II, III, and IV; 10-13 days post-infection (PI) with the mean pre-patent period (PPP) of 12, 10, and 11 days respectively. Mixed infection persisted throughout the duration of the experiment. T. brucei predominated T. congolense in the mixed infection constituting about 70% of the trypanosomes. The significant (P<0.05) decrease in the mean haemoglobin concentration (Hb) and packed cell volume (PCV) caused by the infection did not differ significantly (P>0.05) between the infected groups. Also the significant (P<0.05) reduction in the total white blood cell count (TWBC) caused by the infection did not differ significantly (P>0.05) between the infected groups. The decline in the total WBC count was due primarily to significant (P<0.05) reduction in the lymphocyte counts of the infected dogs. It was thus concluded that single or mixed infection of mongrel dogs with T. congolense and T. brucei brucei resulted in anaemia and leucopenia which did not differ significantly (P>0.05) among the infected groups. Copyright © 2010 Elsevier B.V. All rights reserved.

  3. Trypanosoma livingstonei: a new species from African bats supports the bat seeding hypothesis for the Trypanosoma cruzi clade

    PubMed Central

    2013-01-01

    Background Bat trypanosomes have been implicated in the evolutionary history of the T. cruzi clade, which comprises species from a wide geographic and host range in South America, Africa and Europe, including bat-restricted species and the generalist agents of human American trypanosomosis T. cruzi and T. rangeli. Methods Trypanosomes from bats (Rhinolophus landeri and Hipposideros caffer) captured in Mozambique, southeast Africa, were isolated by hemoculture. Barcoding was carried out through the V7V8 region of Small Subunit (SSU) rRNA and Fluorescent Fragment Length barcoding (FFLB). Phylogenetic inferences were based on SSU rRNA, glyceraldehyde phosphate dehydrogenase (gGAPDH) and Spliced Leader (SL) genes. Morphological characterization included light, scanning and transmission electron microscopy. Results New trypanosomes from bats clustered together forming a clade basal to a larger assemblage called the T. cruzi clade. Barcoding, phylogenetic analyses and genetic distances based on SSU rRNA and gGAPDH supported these trypanosomes as a new species, which we named Trypanosoma livingstonei n. sp. The large and highly polymorphic SL gene repeats of this species showed a copy of the 5S ribosomal RNA into the intergenic region. Unique morphological (large and broad blood trypomastigotes compatible to species of the subgenus Megatrypanum and cultures showing highly pleomorphic epimastigotes and long and slender trypomastigotes) and ultrastructural (cytostome and reservosomes) features and growth behaviour (when co-cultivated with HeLa cells at 37°C differentiated into trypomastigotes resembling the blood forms and do not invaded the cells) complemented the description of this species. Conclusion Phylogenetic inferences supported the hypothesis that Trypanosoma livingstonei n. sp. diverged from a common ancestral bat trypanosome that evolved exclusively in Chiroptera or switched at independent opportunities to mammals of several orders forming the clade T. cruzi

  4. Genotyping of Trypanosoma cruzi DTUs and Trypanosoma rangeli genetic groups in experimentally infected Rhodnius prolixus by PCR-RFLP.

    PubMed

    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. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Molecular demonstration of Trypanosoma evansi and Trypanosoma lewisi DNA in wild rodents from Cambodia, Lao PDR and Thailand.

    PubMed

    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.

  6. Trypanosoma livingstonei: a new species from African bats supports the bat seeding hypothesis for the Trypanosoma cruzi clade.

    PubMed

    Lima, Luciana; Espinosa-Álvarez, Oneida; Hamilton, Patrick B; Neves, Luis; Takata, Carmen S A; Campaner, Marta; Attias, Márcia; de Souza, Wanderley; Camargo, Erney P; Teixeira, Marta M G

    2013-08-03

    Bat trypanosomes have been implicated in the evolutionary history of the T. cruzi clade, which comprises species from a wide geographic and host range in South America, Africa and Europe, including bat-restricted species and the generalist agents of human American trypanosomosis T. cruzi and T. rangeli. Trypanosomes from bats (Rhinolophus landeri and Hipposideros caffer) captured in Mozambique, southeast Africa, were isolated by hemoculture. Barcoding was carried out through the V7V8 region of Small Subunit (SSU) rRNA and Fluorescent Fragment Length barcoding (FFLB). Phylogenetic inferences were based on SSU rRNA, glyceraldehyde phosphate dehydrogenase (gGAPDH) and Spliced Leader (SL) genes. Morphological characterization included light, scanning and transmission electron microscopy. New trypanosomes from bats clustered together forming a clade basal to a larger assemblage called the T. cruzi clade. Barcoding, phylogenetic analyses and genetic distances based on SSU rRNA and gGAPDH supported these trypanosomes as a new species, which we named Trypanosoma livingstonei n. sp. The large and highly polymorphic SL gene repeats of this species showed a copy of the 5S ribosomal RNA into the intergenic region. Unique morphological (large and broad blood trypomastigotes compatible to species of the subgenus Megatrypanum and cultures showing highly pleomorphic epimastigotes and long and slender trypomastigotes) and ultrastructural (cytostome and reservosomes) features and growth behaviour (when co-cultivated with HeLa cells at 37°C differentiated into trypomastigotes resembling the blood forms and do not invaded the cells) complemented the description of this species. Phylogenetic inferences supported the hypothesis that Trypanosoma livingstonei n. sp. diverged from a common ancestral bat trypanosome that evolved exclusively in Chiroptera or switched at independent opportunities to mammals of several orders forming the clade T. cruzi, hence, providing further support for

  7. Toxicity and anti-trypanosomal effects of ethanolic extract of Butyrospermum paradoxum (Sapotaceae) stem bark in rats infected with Trypanosoma brucei and Trypanosoma congolense.

    PubMed

    Mbaya, Albert W; Nwosu, Chukwunyere O; Onyeyili, Patrick A

    2007-05-22

    The ethanolic extract of Butyrospermum paradoxum stem bark, commonly used in the traditional treatment of various diseases including animal and human trypanosomosis in north-eastern Nigeria, was tested for toxicity and anti-trypanosomal efficacy in rats infected with Trypanosoma congolense and Trypanosoma brucei. Following intra-peritoneal administration, the extract induced behavioural changes, morbidity and mortality in the rats. The symptoms observed included anorexia, dehydration, depression, prostration, coma and death. These symptoms were noted at high doses (> 800 mg/kg) only. At necropsy, the pathological lesions were mainly congestion and oedema of the lungs, bronchi, bronchioles and the kidney, hepatomegally and focal necrosis of the liver cells. The severity of the symptoms and lesions were dose related. The intra-peritoneal LD50 of the extract was 820 mg/kg. The extract produced anti-trypanosomal effect through the complete suppression or delay in parasite establishment with reduction in the level of parasitaemia and the severity of the attendant disease as well as enhanced survival of the rats infected with Trypanosoma congolense and Trypanosoma brucei. The results suggest that the folkloric medicinal application of the extracts of Butyrospermum paradoxum has a pharmacological basis.

  8. FIRST REPORT OF TRYPANOSOMA EVANSI INFECTION (SURRA) IN A PUMA (FELIS CONCOLOR) OF LAHORE ZOO, PAKISTAN.

    PubMed

    Rashid, Imran; Akbar, Haroon; Gharbi, Mohamed; Riaz, Farooq; Islam, Saher; Saleem, Muhammad Baber; Shahzad, Sammuel; Shehzad, Wasim; Rouatbi, Mariem; Ashraf, Kamran

    2017-09-01

    The blood protozoan Trypanosoma evansi, which is transmitted by biting flies, is frequently neglected due to subclinical infections. This report describes a case of trypanosomiasis due to T. evansi in a 9-yr-old male puma (Felis concolor) housed at the Lahore Zoo in Pakistan. Early in January 2015, this male puma presented with chronic lethargy, weight loss, incoordination, hyperthermia, anorexia, sunken eyes, and unthriftiness. Microscopic examination of Giemsa-stained blood smears showed numerous Trypanosoma parasites. The puma was treated with diminazene aceturate subcutaneously twice. A few days later, a blood smear examination showed absence of trypanosomes. Five months later the cat presented with acute epistaxis and died. Postmortem examination showed emaciation, pale liver and kidneys, and hemorrhages on the spleen. Examination of a blood smear taken at the time of death showed numerous Trypanosoma parasites. PCR testing confirmed the presence of Trypanosoma DNA. DNA sequencing of two amplicons confirmed the presence of Trypanosoma in the blood smears with a 98-99% identity with the previously identified GenBank sequences. A phylogenetic tree was then constructed. Further studies are needed to improve our knowledge about the epidemiology and pathogenesis of T. evansi infection in wild animal species.

  9. Glycoproteins, antigens, and regulation of complement activation on the surface of the protozoan parasite Trypanosoma lewisi: implications for immune evasion

    SciTech Connect

    Sturtevant, J.E.

    1985-01-01

    The surface antigens and glycoproteins of the rat parasitic protozoan, Trypanosoma lewisi were characterized. Radioiodination with /sup 125/I identified 10 out of more 40 polypeptides separated on sodium dodecyl sulfate polyacrylamide gel electrophoresis. All of these components were identified as glycoproteins by peroxidase-conjugated Conconavalin A (HR-Con A) lectin affinoblotting. This analysis detected that quantitative but not qualitative changes occurred during infection. Localization of most of the reactive determinants was indicated by immunoblotting extracts of radioiodinated T. lewisi. Changes in the antigenicity as related to survival in the host are discussed. The presence of IgG and IgM on the surface of T. lewisi isolated from intact and ..gamma..-irradiated rats (irr.) and that determinants bind Ig from uninfected rat sera (NRS) was indicated by flow cytometric analysis. Immunoblotting identified the major NRS IgG binding component as the 74 kd surface glycoprotein. Complement component C3 deposition during infection was indicated by flow cytometric analysis and immunoblotting. Incubation of intact T. lewisi with normal human sera indicated that C3, C5, and factor B deposition was Mg/sup 2 +/ dependent, Ca/sup 2 +/ independent and deposited C3 was rapidly processed to hemolytically inactive fragments. Radioiodination of intact and protease T. lewisi after cultivation identified three components which correlate with resistance to lysis. This suggests that surface moieties on intact T. lewisi modulate host complement activity by restricting C3/C5 convertase activity.

  10. ConSCRIPT

    PubMed Central

    Mottarella, Scott E.; Rosa, Mario; Bangura, Abdul; Bernstein, Herbert J.; Craig, Paul A.

    2011-01-01

    The aim of the Structural Biology Extensible Visualization Scripting Language (SBEVSL) project is to allow users who are experts in one scripting language to use that language in a second molecular visualization environment without requiring the user to learn a new scripting language. ConSCRIPT, the first SBEVSL release, is a plug-in for PyMOL that accepts RasMol scripting commands either as premade scripts or as line-by-line entries from PyMOL's own command line. The plug-in is available for download at http://sourceforge.net/projects/sbevsl/files in the ConSCRIPT folder. PMID:21567873

  11. Comparative serum biochemical changes in mongrel dogs following single and mixed infections of Trypanosoma congolense and Trypanosoma brucei brucei.

    PubMed

    Ezeokonkwo, R C; Ezeh, I O; Onunkwo, J I; Onyenwe, I W; Iheagwam, C N; Agu, W E

    2012-11-23

    The serum activities of alkaline phosphatase (AP), alanine aminotransferase (ALT), aspartate aminotransferase (AST) and the serum levels of conjugated bilirubin (CB), blood urea nitrogen (BUN) and creatinine were studied following single and mixed infections of mongrel dogs with Trypanosoma congolense and Trypanosoma brucei brucei. Twenty mongrel dogs of both sexes aged between 3 and 6 months, and weighing between 2.5 and 5.9 kg were used for the study. The dogs were kept in clean metal cages in a fly-proof house and were fed and given water ad libitum. The twenty dogs were divided into four groups of five dogs each. Group I dogs were uninfected control, group II were infected with T. congolense, group III were infected with T. brucei brucei and group IV were infected with both T. congolense and T. brucei brucei. Each dog in the infected groups II and III was inoculated intraperitonealy (i/p) with 1.0 ml of PBS diluted blood containing 1.0×10(6) trypanosomes whereas each infected dog in group IV (mixed infection) was inoculated with 0.5 ml of the PBS diluted blood containing 0.5×10(6)T. congolense and 0.5 ml of the PBS diluted blood containing 0.5×10(6)T. brucei brucei i/p. Parasites were detectable in the blood of the infected dogs in groups II, III, and IV 10-13 days post infection (PI) with the mean pre-patent period (PP) of 12, 10, and 11 days respectively. Trypanosome infection caused a significant (P<0.05) increase in the serum activities of AP, ALT, AST and the serum levels of creatinine, CB, and BUN. The significant increases in the serum levels of CB, BUN, and creatinine and serum activities of AP and AST became noticeable from day seven PI in all the infected groups whereas that of ALT became noticeable from day 14 PI and increased continuously until the experiment was terminated. These increases however did not differ significantly (P>0.05) between the infected groups in most cases. It was thus concluded that single or mixed infection of mongrel dogs

  12. Rhodnius prolixus: from physiology by Wigglesworth to recent studies of immune system modulation by Trypanosoma cruzi and Trypanosoma rangeli.

    PubMed

    Azambuja, P; Garcia, E S; Waniek, P J; Vieira, C S; Figueiredo, M B; Gonzalez, M S; Mello, C B; Castro, D P; Ratcliffe, N A

    This review is dedicated to the memory of Professor Sir Vincent B. Wigglesworth (VW) in recognition of his many pioneering contributions to insect physiology which, even today, form the basis of modern-day research in this field. Insects not only make vital contributions to our everyday lives by their roles in pollination, balancing eco-systems and provision of honey and silk products, but they are also outstanding models for studying the pathogenicity of microorganisms and the functioning of innate immunity in humans. In this overview, the immune system of the triatomine bug, Rhodnius prolixus, is considered which is most appropriate to this dedication as this insect species was the favourite subject of VW's research. Herein are described recent developments in knowledge of the functioning of the R. prolixus immune system. Thus, the roles of the cellular defences, such as phagocytosis and nodule formation, as well as the role of eicosanoids, ecdysone, antimicrobial peptides, reactive oxygen and nitrogen radicals, and the gut microbiota in the immune response of R. prolixus are described. The details of many of these were unknown to VW although his work gives indications of his awareness of the importance to R. prolixus of cellular immunity, antibacterial activity, prophenoloxidase and the gut microbiota. This description of R. prolixus immunity forms a backdrop to studies on the interaction of the parasitic flagellates, Trypanosoma cruzi and Trypanosoma rangeli, with the host defences of this important insect vector. These parasites remarkably utilize different strategies to avoid/modulate the triatomine immune response in order to survive in the extremely hostile host environments present in the vector gut and haemocoel. Much recent information has also been gleaned on the remarkable diversity of the immune system in the R. prolixus gut and its interaction with trypanosome parasites. This new data is reviewed and gaps in our knowledge of R. prolixus immunity are

  13. Testicular pathology, gonadal and epididymal sperm reserves of Yankasa rams infected with experimental Trypanosoma brucei brucei and Trypanosoma evansi

    PubMed Central

    Wada, Yunusa A.; Oniye, Sonnie J.; Rekwot, Peter I.; Okubanjo, Oluyinka O.

    2016-01-01

    Aim: The study was conducted to evaluate the pathological effects of trypanosomosis on the testes, gonadal, and epididymal sperm reserves of Yankasa rams for 98 days. Materials and Methods: A total of 16 Yankasa rams, aged between 24 and 30 months and weighed between 22 and 25 kg, were acclimatized for a period of 2-months in a clean fly proof house and were adequately fed and given water ad-libitum. Of the 16 rams, 12 that were clinically fit for the experiment at the end of the acclimatization period were randomly divided into four groups: Groups I, II, III, and IV, each having 3 rams. Groups I and II were each challenged singly with experimental Trypanosoma brucei brucei (Federer strain) and Trypanosoma evansi (Sokoto strain), respectively, while Group III was challenged with mixed T. brucei brucei and T. evansi parasites (50% of each species in the infective inoculum) and Group IV was left as an uninfected control. Each infected ram received 2 mL of the infected blood containing 2×106 trypomastigotes via the jugular vein, while the control group received 2 mL each, normal saline. Results: All the infected rams developed clinical signs typical of trypanosomosis at varying pre-patent periods. The gross lesions observed in the infected rams in Group II were moderate and more severe in those of Groups I and III. Histological sections of the testes of infected rams (Groups I, II, and III) showed moderate (T. evansi-infected group) to severe (mixed and T. brucei brucei-infected groups) testicular degenerations with reduction in number of spermatogenic cell layers, degenerated seminiferous tubules, congested interlobular spaces, loss of tissue architecture with significant (p<0.01) depletion, and loss of gonadal and epididymal sperm reserves in Groups I and III in comparison to Group II and the control Group IV. No observable clinical signs and histopathological lesions were found in those rams of the control Group IV. Conclusion: The study concluded that

  14. MIF contributes to Trypanosoma brucei associated immunopathogenicity development.

    PubMed

    Stijlemans, Benoît; Leng, Lin; Brys, Lea; Sparkes, Amanda; Vansintjan, Liese; Caljon, Guy; Raes, Geert; Van Den Abbeele, Jan; Van Ginderachter, Jo A; Beschin, Alain; Bucala, Richard; De Baetselier, Patrick

    2014-09-01

    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 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.

  15. Antimicrobial activity of synthetic bornyl benzoates against Trypanosoma cruzi

    PubMed Central

    Corrêa, P R C; Miranda, R R S; Duarte, L P; Silva, G D F; Filho, S A Vieira; Okuma, A A; Carazza, F; Morgado-Díaz, J A; Pinge-Filho, P; Yamauchi, L M; Nakamura, C V; Yamada-Ogatta, S F

    2012-01-01

    We report here for the first time the in vitro effects of (1S,2R,4S)-1,7,7-trimethyl-bicyclo[2.2.1]heptan-2-yl-3′,4′,5′-trimethoxy benzoate (1) and (1S,2R,4S)-1,7,7-trimethyl-bicyclo[2.2.1]heptan-2-yl benzoate (2) on the growth and ultrastructure of Trypanosoma cruzi. These two synthetic compounds exerted an antiproliferative effect on the epimastigote forms of the parasite. The ICs50/72h of two synthetic L-bornyl benzoates, 1 and 2, was 10.1 and 12.8 μg/ml, respectively. Both compounds were more selective against epimastigotes than HEp-2 cells. Ultrastructural analysis revealed intense cytoplasmic vacuolization and the appearance of cytoplasmic materials surrounded by membranes. The treatment of peritoneal macrophages with compounds 1 and 2 caused a significant decrease in the number of T. cruzi-infected cells. L-Bornyl benzoate derivatives may serve as a potential source for the development of more effective and safer chemotherapeutic agents against T. cruzi infections. PMID:22943546

  16. mRNA localization mechanisms in Trypanosoma cruzi.

    PubMed

    Alves, Lysangela R; Guerra-Slompo, Eloise P; de Oliveira, Arthur V; Malgarin, Juliane S; Goldenberg, Samuel; Dallagiovanna, Bruno

    2013-01-01

    Asymmetric mRNA localization is a sophisticated tool for regulating and optimizing protein synthesis and maintaining cell polarity. Molecular mechanisms involved in the regulated localization of transcripts are widespread in higher eukaryotes and fungi, but not in protozoa. Trypanosomes are ancient eukaryotes that branched off early in eukaryote evolution. We hypothesized that these organisms would have basic mechanisms of mRNA localization. FISH assays with probes against transcripts coding for proteins with restricted distributions showed a discrete localization of the mRNAs in the cytoplasm. Moreover, cruzipain mRNA was found inside reservosomes suggesting new unexpected functions for this vacuolar organelle. Individual mRNAs were also mobilized to RNA granules in response to nutritional stress. The cytoplasmic distribution of these transcripts changed with cell differentiation, suggesting that localization mechanisms might be involved in the regulation of stage-specific protein expression. Transfection assays with reporter genes showed that, as in higher eukaryotes, 3'UTRs were responsible for guiding mRNAs to their final location. Our results strongly suggest that Trypanosoma cruzi have a core, basic mechanism of mRNA localization. This kind of controlled mRNA transport is ancient, dating back to early eukaryote evolution.

  17. Comparative genomics of drug resistance in Trypanosoma brucei rhodesiense.

    PubMed

    Graf, Fabrice E; Ludin, Philipp; Arquint, Christian; Schmidt, Remo S; Schaub, Nadia; Kunz Renggli, Christina; Munday, Jane C; Krezdorn, Jessica; Baker, Nicola; Horn, David; Balmer, Oliver; Caccone, Adalgisa; de Koning, Harry P; Mäser, Pascal

    2016-09-01

    Trypanosoma brucei rhodesiense is one of the causative agents of human sleeping sickness, a fatal disease that is transmitted by tsetse flies and restricted to Sub-Saharan Africa. Here we investigate two independent lines of T. b. rhodesiense that have been selected with the drugs melarsoprol and pentamidine over the course of 2 years, until they exhibited stable cross-resistance to an unprecedented degree. We apply comparative genomics and transcriptomics to identify the underlying mutations. Only few mutations have become fixed during selection. Three genes were affected by mutations in both lines: the aminopurine transporter AT1, the aquaporin AQP2, and the RNA-binding protein UBP1. The melarsoprol-selected line carried a large deletion including the adenosine transporter gene AT1, whereas the pentamidine-selected line carried a heterozygous point mutation in AT1, G430R, which rendered the transporter non-functional. Both resistant lines had lost AQP2, and both lines carried the same point mutation, R131L, in the RNA-binding motif of UBP1. The finding that concomitant deletion of the known resistance genes AT1 and AQP2 in T. b. brucei failed to phenocopy the high levels of resistance of the T. b. rhodesiense mutants indicated a possible role of UBP1 in melarsoprol-pentamidine cross-resistance. However, homozygous in situ expression of UBP1-Leu(131) in T. b. brucei did not affect the sensitivity to melarsoprol or pentamidine.

  18. Characterisation of the fumarate hydratase repertoire in Trypanosoma cruzi.

    PubMed

    de Pádua, Ricardo A P; Kia, Ali Martin; Costa-Filho, Antonio J; Wilkinson, Shane R; Nonato, M Cristina

    2017-09-01

    Nifurtimox and benznidazole represent the only treatments options available targeting Chagas disease, the most important parasitic infection in the Americas. However, use of these is problematic as they are toxic and ineffective against the more severe stages of the disease. In this work, we used a multidisciplinary approach to characterise the fumarases from Trypanosoma cruzi, the causative agent of Chagas Disease. We showed this trypanosome expresses cytosolic and mitochondrial fumarases that via an iron-sulfur cluster mediate the reversible conversion of fumarate to S-malate. Based on sequence, biochemical properties and co-factor binding, both T. cruzi proteins share characteristics with class I fumarases, enzymes found in bacteria and some other protozoa but absent from humans, that possess class II isoforms instead. Gene disruption suggested that although the cytosolic or mitochondrial fumarase activities are individually dispensable their combined activity is essential for parasite viability. Finally, based on the mechanistic differences with the human (host) fumarase, we designed and validated a selective inhibitor targeting the parasite enzyme. This study showed that T. cruzi fumarases should be exploited as targets for the development of new chemotherapeutic interventions against Chagas disease. Copyright © 2017 Elsevier B.V. All rights reserved.

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

    PubMed

    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. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  20. Single molecule analysis of Trypanosoma brucei DNA replication dynamics

    PubMed Central

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

    2015-01-01

    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. PMID:25690894

  1. Geographical Distribution of Trypanosoma cruzi Genotypes in Venezuela

    PubMed Central

    Carrasco, Hernán J.; Segovia, Maikell; Llewellyn, Martin S.; Morocoima, Antonio; Urdaneta-Morales, Servio; Martínez, Cinda; Martínez, Clara E.; Garcia, Carlos; Rodríguez, Marlenes; Espinosa, Raul; de Noya, Belkisyolé A.; Díaz-Bello, Zoraida; Herrera, Leidi; Fitzpatrick, Sinead; Yeo, Matthew; Miles, Michael A.; Feliciangeli, M. Dora

    2012-01-01

    Chagas disease is an endemic zoonosis native to the Americas and is caused by the kinetoplastid protozoan parasite Trypanosoma cruzi. The parasite is also highly genetically diverse, with six discrete typing units (DTUs) reported TcI – TcVI. These DTUs broadly correlate with several epidemiogical, ecological and pathological features of Chagas disease. In this manuscript we report the most comprehensive evaluation to date of the genetic diversity of T. cruzi in Venezuela. The dataset includes 778 samples collected and genotyped over the last twelve years from multiple hosts and vectors, including nine wild and domestic mammalian host species, and seven species of triatomine bug, as well as from human sources. Most isolates (732) can be assigned to the TcI clade (94.1%); 24 to the TcIV group (3.1%) and 22 to TcIII (2.8%). Importantly, among the 95 isolates genotyped from human disease cases, 79% belonged to TcI - a DTU common in the Americas, however, 21% belonged to TcIV- a little known genotype previously thought to be rare in humans. Furthermore, were able to assign multiple oral Chagas diseases cases to TcI in the area around the capital, Caracas. We discuss our findings in the context of T. cruzi DTU distributions elsewhere in the Americas, and evaluate the impact they have on the future of Chagas disease control in Venezuela. PMID:22745843

  2. Interaction of Trypanosoma evansi with the plasminogen-plasmin system.

    PubMed

    Acosta, Héctor; Rondón-Mercado, Rocío; Avilán, Luisana; Concepción, Juan Luis

    2016-08-15

    Trypanosoma evansi is a widely-distributed haemoflagellated parasite of veterinary importance that infects a variety of mammals including horses, mules, camels, buffalos, cattle and deer. It is the causal agent of a trypanosomiasis known as Surra which produces epidemics of great economic importance in Africa, Asia and South America. The main pathology includes an enlarged spleen with hypertrophy of lymphoid follicles, congested lungs, neuronal degeneration and meningoencephalitis, where migration of the parasites from the blood to the tissues is essential. Most cells, including pathogenic cells, use diverse strategies for tissue invasion, such as the expression of surface receptors to bind plasminogen or plasmin. In this work, we show that T. evansi is able to bind plasminogen and plasmin on its surface. The analysis of this binding revealed a high affinity dissociation constant (Kd of 0.080±0.009μM) and 1×10(5) plasminogen binding sites per cell. Also a second population of receptors with a Kd of 0.255±0.070μM and 3.2×10(4) plasminogen binding sites per cell was determined. Several proteins with molecular masses between ∼18 and ∼70kDa are responsible for this binding. This parasite-plasminogen interaction may be important in the establishment of the infection in the vertebrate host, where the physiological concentration of available plasminogen is around 2μM. Copyright © 2016 Elsevier B.V. All rights reserved.

  3. Trypanosoma cruzi: modification of macrophage function during infection

    PubMed Central

    1977-01-01

    Infection of mice with Trypanosoma cruzi and subsequent intraperitoneal challenge with heat-killed trypanosomes elicits peritoneal macrophages which display in vitro microbicidal activity against trypomastigotes of T. cruzi. These cells also display other activated properties including rapid spreading, intense membrane activity, secretion of high levels of plasminogen activator, and ingestion mediated by the C3 receptor. An intravenous infection with BCG, followed by an intraperitoneal challenge with mycobacterial antigens brings about macrophages with similar properties. These criteria of macrophage activation were compared in normal and BCG- or T. cruzi-immune mice, with or without an intraperitoneal challenge with specific or unrelated antigens. Trypanocidal activity is displayed by both BCG- and T. cruzi-immune macrophages after intraperitoneal challenge with either antigen. Resident-immune macrophages from both T. cruzi- and BCG-infected mice show a trypanostatic, rather than trypanocidal activity. Macrophages from noninfected mice, challenged with the same antigens, show neither trypanostatic nor trypanocidal activity. Increased secretion of plasminogen activator shows a definite immunological specificity. Challenge with the specific antigen induces the appearance of macrophages secreting high levels of plasminogen activator, while unrelated antigens induce much smaller levels. Noninfected mice challenged with the same antigens do not display any enchancement in secretion. In contrast, increased spreading and phagocytosis mediated by the complement receptor are also displayed by cells from noninfected mice challenged with any of the agents tested. PMID:327012

  4. Ca2+/H+ exchange in acidic vacuoles of Trypanosoma brucei.

    PubMed Central

    Vercesi, A E; Moreno, S N; Docampo, R

    1994-01-01

    The use of digitonin to permeabilize the plasma membrane of Trypanosoma brucei procyclic and bloodstream trypomastigotes allowed the identification of a non-mitochondrial nigericin-sensitive Ca2+ compartment. The proton ionophore carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP) was able to cause Ca2+ release from this compartment, which was also sensitive to sodium orthovanadate. Preincubation of the cells with the vacuolar H(+)-ATPase inhibitor bafilomycin A1 greatly reduced the nigericin-sensitive Ca2+ compartment. Bafilomycin A1 inhibited the initial rate of ATP-dependent non-mitochondrial Ca2+ uptake and stimulated the initial rate of nigericin-induced Ca2+ release by permeabilized procyclic trypomastigotes. ATP-dependent and bafilomycin A1- and 7-chloro-4-nitrobenz-2-oxa-1,3-diazole (NBD-Cl)-sensitive Acridine Orange uptake was demonstrated in permeabilized cells. Under these conditions Acridine Orange was concentrated in abundant cytoplasmic round vacuoles by a process inhibited by bafilomycin A1, NBD-Cl, nigericin, and Ca2+. Vanadate or EGTA significantly increased Acridine Orange uptake, while Ca2+ released Acridine Orange from these preparations, thus suggesting that the dye and Ca2+ were being accumulated in the same acidic vacuole. Acridine Orange uptake was reversed by nigericin, bafilomycin A1 and NH4Cl. The results are consistent with the presence of a Ca2+/H(+)-ATPase system pumping Ca2+ into an acidic vacuole, that we tentatively named the acidocalcisome. Images Figure 5 PMID:7998937

  5. Gene Discovery through Expressed Sequence Tag Sequencing in Trypanosoma cruzi

    PubMed Central

    Verdun, Ramiro E.; Di Paolo, Nelson; Urmenyi, Turan P.; Rondinelli, Edson; Frasch, Alberto C. C.; Sanchez, Daniel O.

    1998-01-01

    Analysis of expressed sequence tags (ESTs) constitutes a useful approach for gene identification that, in the case of human pathogens, might result in the identification of new targets for chemotherapy and vaccine development. As part of the Trypanosoma cruzi genome project, we have partially sequenced the 5′ ends of 1,949 clones to generate ESTs. The clones were randomly selected from a normalized CL Brener epimastigote cDNA library. A total of 14.6% of the clones were homologous to previously identified T. cruzi genes, while 18.4% had significant matches to genes from other organisms in the database. A total of 67% of the ESTs had no matches in the database, and thus, some of them might be T. cruzi-specific genes. Functional groups of those sequences with matches in the database were constructed according to their putative biological functions. The two largest categories were protein synthesis (23.3%) and cell surface molecules (10.8%). The information reported in this paper should be useful for researchers in the field to analyze genes and proteins of their own interest. PMID:9784549

  6. Oxidative stress fuels Trypanosoma cruzi infection in mice

    PubMed Central

    Paiva, Claudia N.; Feijó, Daniel F.; Dutra, Fabianno F.; Carneiro, Vitor C.; Freitas, Guilherme B.; Alves, Letícia S.; Mesquita, Jacilene; Fortes, Guilherme B.; Figueiredo, Rodrigo T.; Souza, Heitor S.P.; Fantappié, Marcelo R.; Lannes-Vieira, Joseli; Bozza, Marcelo T.

    2012-01-01

    Oxidative damage contributes to microbe elimination during macrophage respiratory burst. Nuclear factor, erythroid-derived 2, like 2 (NRF2) orchestrates antioxidant defenses, including the expression of heme-oxygenase–1 (HO-1). Unexpectedly, the activation of NRF2 and HO-1 reduces infection by a number of pathogens, although the mechanism responsible for this effect is largely unknown. We studied Trypanosoma cruzi infection in mice in which NRF2/HO-1 was induced with cobalt protoporphyrin (CoPP). CoPP reduced parasitemia and tissue parasitism, while an inhibitor of HO-1 activity increased T. cruzi parasitemia in blood. CoPP-induced effects did not depend on the adaptive immunity, nor were parasites directly targeted. We also found that CoPP reduced macrophage parasitism, which depended on NRF2 expression but not on classical mechanisms such as apoptosis of infected cells, induction of type I IFN, or NO. We found that exogenous expression of NRF2 or HO-1 also reduced macrophage parasitism. Several antioxidants, including NRF2 activators, reduced macrophage parasite burden, while pro-oxidants promoted it. Reducing the intracellular labile iron pool decreased parasitism, and antioxidants increased the expression of ferritin and ferroportin in infected macrophages. Ferrous sulfate reversed the CoPP-induced decrease in macrophage parasite burden and, given in vivo, reversed their protective effects. Our results indicate that oxidative stress contributes to parasite persistence in host tissues and open a new avenue for the development of anti–T. cruzi drugs. PMID:22728935

  7. Characterization of the late endosomal ESCRT machinery in Trypanosoma brucei.

    PubMed

    Silverman, Jason S; Muratore, Katherine A; Bangs, James D

    2013-10-01

    The multivesicular body (MVB) is a specialized Rab7+ late endosome (LE) containing multiple intralumenal vesicles that function in targeting ubiquitinylated cell surface proteins to the lysosome for degradation. African trypanosomes lack a morphologically well-defined MVB, but contain orthologs of the ESCRT (Endosomal Sorting Complex Required for Transport) machinery that mediates MVB formation. We investigate the role of TbVps23, an early ESCRT component, and TbVps4, the terminal ESCRT ATPase, in lysosomal trafficking in bloodstream form trypanosomes. Both localize to the TbRab7+ LE and RNAi silencing of each rapidly blocks growth. TbVps4 silencing results in approximately threefold accumulation of TbVps23 at the LE, consistent with blocking terminal ESCRT disassembly. Trafficking of endocytic and biosynthetic cargo, but not default lysosomal reporters, is also negatively affected. Others reported that TbVps23 mediates ubiquitin-dependent lysosomal degradation of invariant surface glycoproteins (ISG65) (Leung et al., Traffic 2008;9:1698-1716). In contrast, we find that TbVps23 ablation does not affect ISG65 turnover, while TbVps4 silencing markedly enhances lysosomal degradation. We propose several models to accommodate these results, including that the ESCRT machinery actually retrieves ISG65 from the LE to earlier endocytic compartments, and in its absence ISG65 traffics more efficiently to the lysosome. Overall, these results confirm that the ESCRT machinery is essential in Trypanosoma brucei and plays important and novel role(s) in LE function in trypanosomes.

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

    PubMed

    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.

  9. Trypanosoma cruzi and Chagas' Disease in the United States.

    PubMed

    Bern, Caryn; Kjos, Sonia; Yabsley, Michael J; Montgomery, Susan P

    2011-10-01

    Chagas' disease is caused by the protozoan parasite Trypanosoma cruzi and causes potentially life-threatening disease of the heart and gastrointestinal tract. The southern half of the United States contains enzootic cycles of T. cruzi, involving 11 recognized triatomine vector species. The greatest vector diversity and density occur in the western United States, where woodrats are the most common reservoir; other rodents, raccoons, skunks, and coyotes are also infected with T. cruzi. In the eastern United States, the prevalence of T. cruzi is highest in raccoons, opossums, armadillos, and skunks. A total of 7 autochthonous vector-borne human infections have been reported in Texas, California, Tennessee, and Louisiana; many others are thought to go unrecognized. Nevertheless, most T. cruzi-infected individuals in the United States are immigrants from areas of endemicity in Latin America. Seven transfusion-associated and 6 organ donor-derived T. cruzi infections have been documented in the United States and Canada. As improved control of vector- and blood-borne T. cruzi transmission decreases the burden in countries where the disease is historically endemic and imported Chagas' disease is increasingly recognized outside Latin America, the United States can play an important role in addressing the altered epidemiology of Chagas' disease in the 21st century.

  10. An in vivo role for Trypanosoma cruzi calreticulin in antiangiogenesis.

    PubMed

    Molina, María C; Ferreira, Viviana; Valck, Carolina; Aguilar, Lorena; Orellana, Juana; Rojas, Alvaro; Ramirez, Galia; Billetta, Rosario; Schwaeble, Wilhelm; Lemus, David; Ferreira, Arturo

    2005-04-01

    Angiogenesis leads to neovascularization from existing blood vessels. It is associated with tumor growth and metastasis and is regulated by pro- and antiangiogenic molecules, some of them currently under clinical trials for cancer treatment. During the last few years we have cloned, sequenced and expressed a Trypanosoma cruzi calreticulin gene (TcCRT). Its product, TcCRT, a 45 kDa protein, is more than 50% identical to human CRT (HuCRT). TcCRT, present on the surface of trypomastigotes, binds both C1q and mannan binding lectin and inhibits the classical activation pathway of human complement. Since TcCRT is highly homologous to a functional antiangiogenic fragment from HuCRT (aa 120-180), recombinant (r) and native (n) TcCRT were tested in their antiangiogenic effects, in the chick embryonic chorioallantoid membrane (CAM) assay. Both proteins mediated highly significant antiangiogenic effects in the in vivo CAM assay. This effect was further substantiated in experiments showing that the plasmid construct pSecTag/TcCRT also displayed significant antiangiogenic properties, as compared to the empty vector. Most likely, the fact that antiangiogenic substances act preferentially on growing neoplasic tissues, but not on already established tumors, is due to their effects on emerging blood vessels. The results shown here indicate that TcCRT, like its human counterpart, has antiangiogenic properties. These properties may explain, at least partly, the reported antineoplasic effect of experimental T. cruzi infection.

  11. Retrospective distribution of Trypanosoma cruzi I genotypes in Colombia.

    PubMed

    León, Cielo M; Hernández, Carolina; Montilla, Marleny; Ramírez, Juan David

    2015-05-01

    Trypanosoma cruzi is the aetiological agent of Chagas disease, which affects approximately eight million people in the Americas. This parasite exhibits genetic variability, with at least six discrete typing units broadly distributed in the American continent. T. cruzi I (TcI) shows remarkable genetic diversity; a genotype linked to human infections and a domestic cycle of transmission have recently been identified, hence, this strain was named TcIDom. The aim of this work was to describe the spatiotemporal distribution of TcI subpopulations across humans, insect vectors and mammalian reservoirs in Colombia by means of molecular typing targeting the spliced leader intergenic region of mini-exon gene. We analysed 101 TcI isolates and observed a distribution of sylvatic TcI in 70% and TcIDom in 30%. In humans, the ratio was sylvatic TcI in 60% and TcIDom in 40%. In mammal reservoirs, the distribution corresponded to sylvatic TcI in 96% and TcIDom in 4%. Among insect vectors, sylvatic TcI was observed in 48% and TcIDom in 52%. In conclusion, the circulation of TcIDom is emerging in Colombia and this genotype is still adapting to the domestic cycle of transmission. The epidemiological and clinical implications of these findings are discussed herein.

  12. Exosome secretion affects social motility in Trypanosoma brucei

    PubMed Central

    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

  13. Trypanosoma evansi: hematologic changes in experimentally infected cats.

    PubMed

    Da Silva, Aleksandro Schafer; Costa, Márcio Machado; Wolkmer, Patrícia; Zanette, Régis Adriel; Faccio, Luciana; Gressler, Lucas Trevisan; Dorneles, Tagor Eduardo Andreolla; Santurio, Janio Morais; Lopes, Sonia Terezinha dos Anjos; Monteiro, Silvia Gonzalez

    2009-09-01

    This study aimed at evaluating hemogram and erythropoietic changes in cats experimentally infected with Trypanosoma evansi. Thirteen adult female non-breeding Felix catus were separated into two groups: seven animals were infected with 10(8) trypomastigotes each, and six animals were used as negative controls. Animals were kept in air-conditioned rooms and blood smears were performed daily for 49 days. Blood samples were collected from the jugular vein at days 0, 7, 21, 35 and 49 and stored in blood-collecting tubes containing anticoagulant. Bone marrow was collected from the proximal epiphysis of the right femur at days 14 and 42 post-inoculation (PI). Total erythrocyte count, hematocrit and hemoglobin showed statistical differences among groups from the seventh day PI onwards (P<0.05). The mean corpuscular volume and the mean corpuscular hemoglobin concentration remained normal, characterizing a normocytic-normochromic anemia. Reticulocyte count increased in the infected group from the 21st day onwards, but remained near normal values suggesting a mild regenerative anemia. Moreover, the myeloid:erythroid ratio significantly reduced at day 42 PI, evidencing a bone marrow hematopoietic response. Based on these results we conclude that cats infected with T. evansi have normocytic, normochromic, regenerative anemia.

  14. Crystal Structure of Triosephosphate Isomerase from Trypanosoma cruzi in Hexane

    NASA Astrophysics Data System (ADS)

    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.

  15. Interferon-Gamma Promotes Infection of Astrocytes by Trypanosoma cruzi

    PubMed Central

    Silva, Rafael Rodrigues; Mariante, Rafael M.; Silva, Andrea Alice; dos Santos, Ana Luiza Barbosa; Roffê, Ester; Santiago, Helton; Gazzinelli, Ricardo Tostes; Lannes-Vieira, Joseli

    2015-01-01

    The inflammatory cytokine interferon-gamma (IFNγ) is crucial for immunity against intracellular pathogens such as the protozoan parasite Trypanosoma cruzi, the causative agent of Chagas disease (CD). IFNγ is a pleiotropic cytokine which regulates activation of immune and non-immune cells; however, the effect of IFNγ in the central nervous system (CNS) and astrocytes during CD is unknown. Here we show that parasite persists in the CNS of C3H/He mice chronically infected with the Colombian T. cruzi strain despite the increased expression of IFNγ mRNA. Furthermore, most of the T. cruzi-bearing cells were astrocytes located near IFNγ+ cells. Surprisingly, in vitro experiments revealed that pretreatment with IFNγ promoted the infection of astrocytes by T. cruzi increasing uptake and proliferation of intracellular forms, despite inducing increased production of nitric oxide (NO). Importantly, the effect of IFNγ on T. cruzi uptake and growth is completely blocked by the anti-tumor necrosis factor (TNF) antibody Infliximab and partially blocked by the inhibitor of nitric oxide synthesis L-NAME. These data support that IFNγ fuels astrocyte infection by T. cruzi and critically implicate IFNγ-stimulated T. cruzi-infected astrocytes as sources of TNF and NO, which may contribute to parasite persistence and CNS pathology in CD. PMID:25695249

  16. Structural analysis of inositol phospholipids from Trypanosoma cruzi epimastigote forms.

    PubMed Central

    Bertello, L E; Gonçalvez, M F; Colli, W; de Lederkremer, R M

    1995-01-01

    Inositol phospholipids (IPL) from epimastigote forms of Trypanosoma cruzi have been investigated by metabolic labelling with [3H]palmitic acid and by GLC-MS analysis of the lipids obtained from non-labelled parasites. The IPL fraction was separated into phosphatidylinositol (PI) and inositol-phosphoceramide subfractions, the latter accounting for 80-85% of the total IPL. The neutral lipids released from the IPLs by PI-specific phospholipase C (PI-PLC) from Bacillus thuringiensis were analysed by silica-gel and reverse-phase TLC for the radioactive lipids and by GLC-MS for the non-radioactive samples. Ceramides containing dihydrosphingosine and sphingosine with C16:0 and C18:0 fatty acids were identified. The main component in the [3H]palmitic acid-labelled ceramides was palmitoyldihydrospingosine, while in the non-labelled sample the ceramides contained mainly sphingosine. This could reflect partial uptake of phospholipid from the medium. The PI contain both alkylacyl- and diacyl-glycerol lipids, with the ether lipid being more abundant. The latter was identified as 1-O-hexadecylglycerol esterified by C18:2 and C18:1 fatty acids. Interestingly, the same lipid had been identified in the anchor of the 1G7 glycoprotein of T. cruzi metacyclic forms. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 7 PMID:7646454

  17. Characterization of inositolphospholipids in Trypanosoma cruzi trypomastigote forms.

    PubMed

    Uhrig, M L; Couto, A S; Colli, W; de Lederkremer, R M

    1996-05-20

    In vivo labeling experiments with [3H]palmitic acid, [3H]inositol, and [3H]glucose allowed the identification of two main classes of inositolphospholipids (IPLs) from the trypomastigote stage of Trypanosoma cruzi. Purification of these compounds was achieved by ion-exchange chromatography, high performance liquid chromatography and thin layer chromatography. Specific phosphatidyl-inositol phospholipase C digestion, dephosphorylation and acid methanolysis showed a ceramide structure for the lower migrating IPL1. Palmitoyldihydrosphingosine and palmitoylsphingosine were detected by reverse-phase thin-layer chromatography. On the other hand, IPL2 showed to be a mixture of diacylglycero- and alkylacylglycero-phospholipids in a 1:1 ratio. After PI-PLC digestion, the lipids were separated by preparative TLC and individually analysed. The diacylglycerol contained mainly C18:0 fatty acid together with a low amount of C16:0. Hexadecylglycerol esterified with the C18:0 fatty acid was the only alkylacylglycerol detected. The C18:2 and C18:1 fatty acids, preponderant in the PI molecules of epimastigote forms, were not detected in trypomastigote forms. This is the first report on inositol phospholipids, putative precursors of lipid anchors in the infective stage of T. cruzi.

  18. Mouse infection and pathogenesis by Trypanosoma brucei motility mutants.

    PubMed

    Kisalu, Neville K; Langousis, Gerasimos; Bentolila, Laurent A; Ralston, Katherine S; Hill, Kent L

    2014-06-01

    The flagellum of Trypanosoma brucei is an essential and multifunctional organelle that drives parasite motility and is receiving increased attention as a potential drug target. In the mammalian host, parasite motility is suspected to contribute to infection and disease pathogenesis. However, it has not been possible to test this hypothesis owing to lack of motility mutants that are viable in the bloodstream life cycle stage that infects the mammalian host. We recently identified a bloodstream-form motility mutant in 427-derived T. brucei in which point mutations in the LC1 dynein subunit disrupt propulsive motility but do not affect viability. These mutants have an actively beating flagellum, but cannot translocate. Here we demonstrate that the LC1 point mutant fails to show enhanced cell motility upon increasing viscosity of the surrounding medium, which is a hallmark of wild type T. brucei, thus indicating that motility of the mutant is fundamentally altered compared with wild type cells. We next used the LC1 point mutant to assess the influence of trypanosome motility on infection in mice. Wesurprisingly found that disrupting parasite motility has no discernible effect on T. brucei bloodstream infection. Infection time-course, maximum parasitaemia, number of waves of parasitaemia, clinical features and disease outcome are indistinguishable between motility mutant and control parasites. Our studies provide an important step toward understanding the contribution of parasite motility to infection and a foundation for future investigations of T. brucei interaction with the mammalian host.

  19. Cancer in the parasitic protozoans Trypanosoma brucei and Toxoplasma gondii

    PubMed Central

    Lun, Zhao-Rong; Lai, De-Hua; Wen, Yan-Zi; Zheng, Ling-Ling; Shen, Ji-Long; Yang, Ting-Bo; Zhou, Wen-Liang; Qu, Liang-Hu; Hide, Geoff; Ayala, Francisco J.

    2015-01-01

    Cancer is a general name for more than 100 malignant diseases. It is postulated that all cancers start from a single abnormal cell that grows out of control. Untreated cancers can cause serious consequences and deaths. Great progress has been made in cancer research that has significantly improved our knowledge and understanding of the nature and mechanisms of the disease, but the origins of cancer are far from being well understood due to the limitations of suitable model systems and to the complexities of the disease. In view of the fact that cancers are found in various species of vertebrates and other metazoa, here, we suggest that cancer also occurs in parasitic protozoans such as Trypanosoma brucei, a blood parasite, and Toxoplasma gondii, an obligate intracellular pathogen. Without treatment, these protozoan cancers may cause severe disease and death in mammals, including humans. The simpler genomes of these single-cell organisms, in combination with their complex life cycles and fascinating life cycle differentiation processes, may help us to better understand the origins of cancers and, in particular, leukemias. PMID:26195778

  20. Cancer in the parasitic protozoans Trypanosoma brucei and Toxoplasma gondii.

    PubMed

    Lun, Zhao-Rong; Lai, De-Hua; Wen, Yan-Zi; Zheng, Ling-Ling; Shen, Ji-Long; Yang, Ting-Bo; Zhou, Wen-Liang; Qu, Liang-Hu; Hide, Geoff; Ayala, Francisco J

    2015-07-21

    Cancer is a general name for more than 100 malignant diseases. It is postulated that all cancers start from a single abnormal cell that grows out of control. Untreated cancers can cause serious consequences and deaths. Great progress has been made in cancer research that has significantly improved our knowledge and understanding of the nature and mechanisms of the disease, but the origins of cancer are far from being well understood due to the limitations of suitable model systems and to the complexities of the disease. In view of the fact that cancers are found in various species of vertebrates and other metazoa, here, we suggest that cancer also occurs in parasitic protozoans such as Trypanosoma brucei, a blood parasite, and Toxoplasma gondii, an obligate intracellular pathogen. Without treatment, these protozoan cancers may cause severe disease and death in mammals, including humans. The simpler genomes of these single-cell organisms, in combination with their complex life cycles and fascinating life cycle differentiation processes, may help us to better understand the origins of cancers and, in particular, leukemias.

  1. Trypanosoma cruzi: Inhibition of infection of human monocytes by aspirin.

    PubMed

    Carvalho de Freitas, Rafael; Lonien, Sandra Cristina Heim; Malvezi, Aparecida Donizette; Silveira, Guilherme Ferreira; Wowk, Pryscilla Fanini; da Silva, Rosiane Valeriano; Yamauchi, Lucy Megumi; Yamada-Ogatta, Sueli Fumie; Rizzo, Luiz Vicente; Bordignon, Juliano; Pinge-Filho, Phileno

    2017-09-19

    Cell invasion by Trypanosoma cruzi and its intracellular replication are essential for progression of the parasite life cycle and development of Chagas disease. Prostaglandin E2 (PGE2) and other eicosanoids potently modulate host response and contribute to Chagas disease progression. In this study, we evaluated the effect of aspirin (ASA), a non-selective cyclooxygenase (COX) inhibitor on the T. cruzi invasion and its influence on nitric oxide and cytokine production in human monocytes. The pretreatment of monocytes with ASA or SQ 22536 (adenylate-cyclase inhibitor) induced a marked inhibition of T. cruzi infection. On the other hand, the treatment of monocytes with SQ 22536 after ASA restored the invasiveness of T. cruzi. This reestablishment was associated with a decrease in nitric oxide and PGE2 production, and also an increase of interleukin-10 and interleukin-12 by cells pre-treated with ASA. Altogether, these results reinforce the idea that the cyclooxygenase pathway plays a fundamental role in the process of parasite invasion in an in vitro model of T. cruzi infection. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Distinct phosphatase activity profiles in two strains of Trypanosoma cruzi.

    PubMed

    Morales-Neto, R; Hulshof, L; Ferreira, C V; Gadelha, F R

    2009-12-01

    Phosphorylation of parasite proteins plays a key role in the process of cell invasion by Trypanosoma cruzi, the etiologic agent of Chagas' disease. In this sense, characterization of parasite kinases and phosphatases could open new possibilities for the rational design of chemotherapeutic agents for the treatment of Chagas' disease. In this work, we analyzed phosphatase activities in T. cruzi homogenates from 2 strains belonging to different lineages and with different resistance to oxidative stress. Tulahuen 2 cells (Lineage I) showed higher phosphatase activities and specificity constants when compared to the Y strain (Lineage II). Tulahuen 2 had an optimum phosphatase activity at pH 4.0 and the Y strain at pH 7.0. In both cases, neutral–basic, but not acid, phosphatase activities were increased in the presence of Mg2+. Although calcium had an inhibitory effect at a pH of 7.0 and 8.0 in the Y strain, this inhibition was restricted to pH 8.0 in the other strain. Different substrates and acid phosphotyrosine and alkaline phosphatase inhibitors exhibited distinct effects on the phosphatase activity of both strains. Our results provide a better understanding of T. cruzi phosphatases and reinforce the notion of heterogeneity among T. cruzi populations.

  3. A multigene family encoding surface glycoproteins in Trypanosoma congolense

    PubMed Central

    Thonnus, Magali; Guérin, Amandine; Rivière, Loïc

    2017-01-01

    Trypanosoma congolense, the causative agent of the most important livestock disease in Africa, expresses specific surface proteins involved in its parasitic lifestyle. Unfortunately, the complete repertoire of such molecules is far from being deciphered. As these membrane components are exposed to the host environment, they could be used as therapeutic or diagnostic targets. By mining the T. congolense genome database, we identified a novel family of lectin-like glycoproteins (TcoClecs). These molecules are predicted to have a transmembrane domain, a tandem repeat amino acid motif, a signal peptide and a C-type lectin-like domain (CTLD). This paper depicts several experimental arguments in favor of a surface localization in bloodstream forms of T. congolense. A TcoClec gene was heterologously expressed in U-2 OS cells and the product could be partially found at the plasma membrane. TcoClecs were also localized at the surface of T. congolense bloodstream forms. The signal was suppressed when the cells were treated with a detergent to remove the plasma membrane or with trypsin to « shave » the parasites and remove their external proteins. This suggests that TcoClecs could be potential diagnostic or therapeutic antigens of African animal trypanosomiasis. The potential role of these proteins in T. congolense as well as in other trypanosomatids is discussed. PMID:28357394

  4. Transcriptional and phenotypical heterogeneity of Trypanosoma cruzi cell populations

    PubMed Central

    Seco-Hidalgo, Víctor; De Pablos, Luis Miguel; Osuna, Antonio

    2015-01-01

    Trypanosoma cruzi has a complex life cycle comprising pools of cell populations which circulate among humans, vectors, sylvatic reservoirs and domestic animals. Recent experimental evidence has demonstrated the importance of clonal variations for parasite population dynamics, survival and evolution. By limiting dilution assays, we have isolated seven isogenic clonal cell lines derived from the Pan4 strain of T. cruzi. Applying different molecular techniques, we have been able to provide a comprehensive characterization of the expression heterogeneity in the mucin-associated surface protein (MASP) gene family, where all the clonal isogenic populations were transcriptionally different. Hierarchical cluster analysis and sequence comparison among different MASP cDNA libraries showed that, despite the great variability in MASP expression, some members of the transcriptome (including MASP pseudogenes) are conserved, not only in the life-cycle stages but also among different strains of T. cruzi. Finally, other important aspects for the parasite, such as growth, spontaneous metacyclogenesis or excretion of different catabolites, were also compared among the clones, demonstrating that T. cruzi populations of cells are also phenotypically heterogeneous. Although the evolutionary strategy that sustains the MASP expression polymorphism remains unknown, we suggest that MASP clonal variability and phenotypic heterogeneities found in this study might provide an advantage, allowing a rapid response to environmental pressure or changes during the life cycle of T. cruzi. PMID:26674416

  5. Trypanosoma cruzi and Chagas' Disease in the United States

    PubMed Central

    Bern, Caryn; Kjos, Sonia; Yabsley, Michael J.; Montgomery, Susan P.

    2011-01-01

    Summary: Chagas' disease is caused by the protozoan parasite Trypanosoma cruzi and causes potentially life-threatening disease of the heart and gastrointestinal tract. The southern half of the United States contains enzootic cycles of T. cruzi, involving 11 recognized triatomine vector species. The greatest vector diversity and density occur in the western United States, where woodrats are the most common reservoir; other rodents, raccoons, skunks, and coyotes are also infected with T. cruzi. In the eastern United States, the prevalence of T. cruzi is highest in raccoons, opossums, armadillos, and skunks. A total of 7 autochthonous vector-borne human infections have been reported in Texas, California, Tennessee, and Louisiana; many others are thought to go unrecognized. Nevertheless, most T. cruzi-infected individuals in the United States are immigrants from areas of endemicity in Latin America. Seven transfusion-associated and 6 organ donor-derived T. cruzi infections have been documented in the United States and Canada. As improved control of vector- and blood-borne T. cruzi transmission decreases the burden in countries where the disease is historically endemic and imported Chagas' disease is increasingly recognized outside Latin America, the United States can play an important role in addressing the altered epidemiology of Chagas' disease in the 21st century. PMID:21976603

  6. A Primate APOL1 Variant That Kills Trypanosoma brucei gambiense

    PubMed Central

    Cooper, Anneli; Capewell, Paul; Clucas, Caroline; Veitch, Nicola; Weir, William; Thomson, Russell; Raper, Jayne; MacLeod, Annette

    2016-01-01

    Humans are protected against infection from most African trypanosomes by lipoprotein complexes present in serum that contain the trypanolytic pore-forming protein, Apolipoprotein L1 (APOL1). The human-infective trypanosomes, Trypanosoma brucei rhodesiense in East Africa and T. b. gambiense in West Africa have separately evolved mechanisms that allow them to resist APOL1-mediated lysis and cause human African trypanosomiasis, or sleeping sickness, in man. Recently, APOL1 variants were identified from a subset of Old World monkeys, that are able to lyse East African T. b. rhodesiense, by virtue of C-terminal polymorphisms in the APOL1 protein that hinder that parasite’s resistance mechanism. Such variants have been proposed as candidates for developing therapeutic alternatives to the unsatisfactory anti-trypanosomal drugs currently in use. Here we demonstrate the in vitro lytic ability of serum and purified recombinant protein of an APOL1 ortholog from the West African Guinea baboon (Papio papio), which is able to lyse examples of all sub-species of T. brucei including T. b. gambiense group 1 parasites, the most common agent of human African trypanosomiasis. The identification of a variant of APOL1 with trypanolytic ability for both human-infective T. brucei sub-species could be a candidate for universal APOL1-based therapeutic strategies, targeted against all pathogenic African trypanosomes. PMID:27494254

  7. Heterogeneous infectiousness in guinea pigs experimentally infected with Trypanosoma cruzi.

    PubMed

    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. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  8. Modes of flagellar assembly in Chlamydomonas reinhardtii and Trypanosoma brucei

    PubMed Central

    Höög, Johanna L; Lacomble, Sylvain; O’Toole, Eileen T; Hoenger, Andreas; McIntosh, J Richard; Gull, Keith

    2014-01-01

    Defects in flagella growth are related to a number of human diseases. Central to flagellar growth is the organization of microtubules that polymerize from basal bodies to form the axoneme, which consists of hundreds of proteins. Flagella exist in all eukaryotic phyla, but neither the mechanism by which flagella grow nor the conservation of this process in evolution are known. Here, we study how protein complexes assemble onto the growing axoneme tip using (cryo) electron tomography. In Chlamydomonas reinhardtii microtubules and associated proteins are added simultaneously. However, in Trypanosoma brucei, disorganized arrays of microtubules are arranged into the axoneme structure by the later addition of preformed protein complexes. Post assembly, the T. brucei transition zone alters structure and its association with the central pair loosens. We conclude that there are multiple ways to form a flagellum and that species-specific structural knowledge is critical before evaluating flagellar defects. DOI: http://dx.doi.org/10.7554/eLife.01479.001 PMID:24448408

  9. Trypanosoma cruzi: adaptation to its vectors and its hosts

    PubMed Central

    Noireau, François; Diosque, Patricio; Jansen, Ana Maria

    2009-01-01

    American trypanosomiasis is a parasitic zoonosis that occurs throughout Latin America. The etiological agent, Trypanosoma cruzi, is able to infect almost all tissues of its mammalian hosts and spreads in the environment in multifarious transmission cycles that may or not be connected. This biological plasticity, which is probably the result of the considerable heterogeneity of the taxon, exemplifies a successful adaptation of a parasite resulting in distinct outcomes of infection and a complex epidemiological pattern. In the 1990s, most endemic countries strengthened national control programs to interrupt the transmission of this parasite to humans. However, many obstacles remain to the effective control of the disease. Current knowledge of the different components involved in elaborate system that is American trypanosomiasis (the protozoan parasite T. cruzi, vectors Triatominae and the many reservoirs of infection), as well as the interactions existing within the system, is still incomplete. The Triatominae probably evolve from predatory reduvids in response to the availability of vertebrate food source. However, the basic mechanisms of adaptation of some of them to artificial ecotopes remain poorly understood. Nevertheless, these adaptations seem to be associated with a behavioral plasticity, a reduction in the genetic repertoire and increasing developmental instability. PMID:19250627

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

    PubMed Central

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

    2016-01-01

    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. PMID:27502971

  11. Perspectives on Trypanosoma cruzi-induced heart disease (Chagas disease)

    PubMed Central

    Tanowitz, Herbert B.; Machado, Fabiana S.; Jelicks, Linda A.; Shirani, Jamshid; Campos de Carvalho, Antonio C.; Spray, David C.; Factor, Stephen M.; Kirchhoff, Louis V.; Weiss, Louis M.

    2009-01-01

    Chagas disease is caused by the parasite Trypanosoma cruzi it is the most common cause of heart disease in endemic areas of Latin America. The year 2009 marks the 100th anniversary of the discovery of T. cruzi infection and Chagas disease by the Brazilian physician Carlos Chagas. Chagasic cardiomyopathy develops in from 10 to 30 percent of persons who are chronically infected with this parasite. Echocardiography and magnetic resonance imaging are important modalities in the evaluation and prognosis of individuals with chagasic heart disease. The etiology of chagasic heart disease likely is multifactorial. Parasite persistence, autoimmunity, and microvascular abnormalities have been studied extensively as possible pathogenic mechanisms. Experimental studies suggest that alterations in cardiac gap junctions may be etiologic in the pathogenesis of conduction abnormalities. The diagnosis of chronic Chagas disease is made by serology. The treatment of this infection has shortcomings that need to be addressed. Cardiac transplantation and bone marrow stem cell therapy for persons with Chagas disease have received increasing research attention in recent years. PMID:19410685

  12. Geographical distribution of Trypanosoma cruzi genotypes in Venezuela.

    PubMed

    Carrasco, Hernán J; Segovia, Maikell; Llewellyn, Martin S; Morocoima, Antonio; Urdaneta-Morales, Servio; Martínez, Cinda; Martínez, Clara E; Garcia, Carlos; Rodríguez, Marlenes; Espinosa, Raul; de Noya, Belkisyolé A; Díaz-Bello, Zoraida; Herrera, Leidi; Fitzpatrick, Sinead; Yeo, Matthew; Miles, Michael A; Feliciangeli, M Dora

    2012-01-01

    Chagas disease is an endemic zoonosis native to the Americas and is caused by the kinetoplastid protozoan parasite Trypanosoma cruzi. The parasite is also highly genetically diverse, with six discrete typing units (DTUs) reported TcI - TcVI. These DTUs broadly correlate with several epidemiogical, ecological and pathological features of Chagas disease. In this manuscript we report the most comprehensive evaluation to date of the genetic diversity of T. cruzi in Venezuela. The dataset includes 778 samples collected and genotyped over the last twelve years from multiple hosts and vectors, including nine wild and domestic mammalian host species, and seven species of triatomine bug, as well as from human sources. Most isolates (732) can be assigned to the TcI clade (94.1%); 24 to the TcIV group (3.1%) and 22 to TcIII (2.8%). Importantly, among the 95 isolates genotyped from human disease cases, 79% belonged to TcI - a DTU common in the Americas, however, 21% belonged to TcIV- a little known genotype previously thought to be rare in humans. Furthermore, were able to assign multiple oral Chagas diseases cases to TcI in the area around the capital, Caracas. We discuss our findings in the context of T. cruzi DTU distributions elsewhere in the Americas, and evaluate the impact they have on the future of Chagas disease control in Venezuela.

  13. Molecular mechanisms of Trypanosoma cruzi infection by oral route.

    PubMed

    Yoshida, Nobuko

    2009-07-01

    Frequent reports on outbreaks of acute Chagas' disease by ingestion of food contaminated with parasites from triatomine insects illustrate the importance of this mode of transmission. Studies on oral Trypanosoma cruzi infection in mice have indicated that metacyclic trypomastigotes invade the gastric mucosal epithelium. A key molecule in this process is gp82, a stage-specific surface glycoprotein that binds to both gastric mucin and to target epithelial cells. By triggering Ca2+ signalling, gp82 promotes parasite internalisation. Gp82 is relatively resistant to peptic digestion at acidic pH, thus preserving the properties critical for oral infection. The infection process is also influenced by gp90, a metacyclic stage-specific molecule that negatively regulates the invasion process. T. cruzi strains expressing high gp90 levels invade cells poorly in vitro. However, their infectivity by oral route varies considerably due to varying susceptibilities of different gp90 isoforms to peptic digestion. Parasites expressing pepsin-susceptible gp90 become highly invasive against target cells upon contact with gastric juice. Such is the case of a T. cruzi isolate from an acute case of orally acquired Chagas' disease; the gp90 from this strain is extensively degraded upon short period of parasite permanence in the gastric milieu. If such an exacerbation of infectivity occurs in humans, it may be responsible for the severity of Chagas' disease reported in outbreaks of oral infection.

  14. Cyclic AMP-dependent protein kinase activity in Trypanosoma cruzi.

    PubMed Central

    Ulloa, R M; Mesri, E; Esteva, M; Torres, H N; Téllez-Iñón, M T

    1988-01-01

    A cyclic AMP-dependent protein kinase activity from epimastigote forms of Trypanosoma cruzi was characterized. Cytosolic extracts were chromatographed on DEAE-cellulose columns, giving two peaks of kinase activity, which were eluted at 0.15 M- and 0.32 M-NaCl respectively. The second activity peak was stimulated by nanomolar concentrations of cyclic AMP. In addition, a cyclic AMP-binding protein co-eluted with the second kinase activity peak. Cyclic AMP-dependent protein kinase activity was further purified by gel filtration, affinity chromatography on histone-agarose and cyclic AMP-agarose, as well as by chromatography on CM-Sephadex. The enzyme ('holoenzyme') could be partially dissociated into two different components: 'catalytic' and 'regulatory'. The 'regulatory' component had specific binding for cyclic AMP, and it inhibited phosphotransferase activity of the homologous 'catalytic component' or of the 'catalytic subunit' from bovine heart. Cyclic AMP reversed these inhibitions. A 'holoenzyme preparation' was phosphorylated in the absence of exogenous phosphate acceptor and analysed by polyacrylamide-gel electrophoresis. A 56 kDa band was phosphorylated. The same preparation was analysed by Western blotting, by using polyclonal antibodies to the regulatory subunits of protein kinases type I or II. Both antibodies reacted with the 56 kDa band. Images Fig. 7. Fig. 8. PMID:2848508

  15. Altered Cardiomyocyte Function and Trypanosoma cruzi Persistence in Chagas Disease.

    PubMed

    Cruz, Jader Santos; Santos-Miranda, Artur; Sales-Junior, Policarpo Ademar; Monti-Rocha, Renata; Campos, Paula Peixoto; Machado, Fabiana Simão; Roman-Campos, Danilo

    2016-05-04

    Chagas disease, caused by the triatominae Trypanosoma cruzi, is one of the leading causes of heart malfunctioning in Latin America. The cardiac phenotype is observed in 20-30% of infected people 10-40 years after their primary infection. The cardiac complications during Chagas disease range from cardiac arrhythmias to heart failure, with important involvement of the right ventricle. Interestingly, no studies have evaluated the electrical properties of right ventricle myocytes during Chagas disease and correlated them to parasite persistence. Taking advantage of a murine model of Chagas disease, we studied the histological and electrical properties of right ventricle in acute (30 days postinfection [dpi]) and chronic phases (90 dpi) of infected mice with the Colombian strain of T. cruzi and their correlation to parasite persistence. We observed an increase in collagen deposition and inflammatory infiltrate at both 30 and 90 dpi. Furthermore, using reverse transcriptase polymerase chain reaction, we detected parasites at 90 dpi in right and left ventricles. In addition, we observed action potential prolongation and reduced transient outward K(+) current and L-type Ca(2+) current at 30 and 90 dpi. Taking together, our results demonstrate that T. cruzi infection leads to important modifications in electrical properties associated with inflammatory infiltrate and parasite persistence in mice right ventricle, suggesting a causal role between inflammation, parasite persistence, and altered cardiomyocyte function in Chagas disease. Thus, arrhythmias observed in Chagas disease may be partially related to altered electrical function in right ventricle. © The American Society of Tropical Medicine and Hygiene.

  16. High Throughput Screening for Anti–Trypanosoma cruzi Drug Discovery

    PubMed Central

    Alonso-Padilla, Julio; Rodríguez, Ana

    2014-01-01

    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. PMID:25474364

  17. Characterization of Protein Kinase CK2 from Trypanosoma brucei

    PubMed Central

    Jensen, Bryan C; Kifer, Charles T; Brekken, Deirdre L; Randall, Amber C; Wang, Qin; Drees, Becky L.; Parsons, Marilyn

    2007-01-01

    CK2 is a ubiquitous but enigmatic kinase. The difficulty in assigning a role to CK2 centers on the fact that, to date, no biologically relevant modulator of its function has been identified. One common theme revolves around a constellation of known substrates involved in growth control, compatible with its concentration in the nucleus and nucleolus. We had previously described the identification of two catalytic subunits of CK2 in Trypanosoma brucei and characterized one of them. Here we report the characterization of the second catalytic subunit, CK2α’, and the identification and characterization of the regulatory subunit CK2β. All three subunits are primarily localized to the nucleolus in T. brucei. We also show that CK2β interacts with the nucleolar protein NOG1, adding to the interaction map which previously linked CK2α to the nucleolar protein NOPP44/46, which in turn associates with the rRNA binding protein p37. CK2 activity has four distinctive features: near equal affinity for GTP and ATP, heparin sensitivity, and stimulation by polyamines and polybasic peptides. Sequence comparison shows that the parasite orthologues have mutations in residues previously mapped as important in specifying affinity for GTP and stimulation by both polyamines and polybasic peptides. Studies of the enzymatic activity of the T. brucei CK2s show that both the affinity for GTP and stimulation by polyamines have been lost and only the features of heparin inhibition and stimulation by polybasic peptides are conserved. PMID:17097160

  18. Cell-substrate adhesion during Trypanosoma cruzi differentiation

    PubMed Central

    1988-01-01

    The transformation of Trypanosoma cruzi epimastigotes to the mammal infective metacyclic trypomastigotes (metacyclogenesis) can be performed in vitro under chemically defined conditions. Under these conditions, differentiating epimastigotes adhere to a surface before their transformation into metacyclic trypomastigotes. Scanning and transmission electron microscopy of adhered and non-adhered parasites during the metacyclogenesis process show that only epimastigotes and few transition forms are found in the first population, whereas metacyclic trypomastigotes are exclusively found in the cell culture supernatant. PAGE analysis of the [35S]methionine metabolic labeling products of adhered and non-adhered parasites shows that although most of the polypeptides are conserved, adhered parasites express specifically four polypeptides in the range of 45-50 kD with an isoelectric point of 4.8. These proteins might be involved in the adhesion process and are recognized by an antiserum against total adhered parasite proteins. This antiserum also recognized a group of 45- 50 kD in the iodine-radiolabeled surface proteins of differentiating cells, providing direct evidence that these components are indeed surface antigens. The results suggest that epimastigotes must adhere to a substrate before their transformation to metacyclic trypomastigotes, being released to the medium as the metacyclogenesis process is accomplished. This could correspond to the process naturally occurring within the triatomine invertebrate host. PMID:3283152

  19. Structural characterization of NETNES glycopeptide from Trypanosoma cruzi.

    PubMed

    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.

  20. Trypanosoma cruzi: histopathology of endocrine system in immunocompromised mice.

    PubMed Central

    Calabrese, K. S.; Lagrange, P. H.; da Costa, S. C.

    1994-01-01

    Naturally immunocompromised athymic mice, neonatal mice and adult outbred OFI mice treated with the immunosuppressive agents cyclophosphamide (CY), dexamethasone (DM) and indomethacin (IM) were infected with trypomastigotes of Trypanosoma cruzi Y and CL strains. 10(4) parasites were used, except in the case of IM treatment, where mice received 10(3) trypomastigotes in one group and 10(5) in another. The course of parasitaemia, tissue distribution of amastigotes and time of mortality were compared with an infected thymus intact control group. Neonate and indomethacin treated mice presented the same pattern of parasitaemia. Death occurred as early as 9-10 days after infection. A single dose of CY 200 mg/kg given 5 days after infection enhanced the parasitaemia and increased the number of parasites in the tissues. All groups were similar in terms of colonization of the endocrine system by parasites and the adrenals showed the highest density of amastigotes nests. The thyroid gland (analysed only in neonates) showed intense amastigote accumulation. Colonization of the ovary was observed with amastigotes in both the theca interna and in the stroma. The testes (also examined only in the neonate) showed that the interstitial cells, the tunica albuginea of the seminiferous tubules and the loose connective tissue were infected. Athymic nude mice showed the most intense parasite colonization of the islets of Langerhans. Images Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 PMID:7734334

  1. Fluorescence analysis of the interaction of isometamidium with Trypanosoma congolense.

    PubMed Central

    Zilberstein, D; Wilkes, J; Hirumi, H; Peregrine, A S

    1993-01-01

    Isometamidium chloride (Samorin) is the only compound recommended for prophylaxis against bovine trypanosomiasis in sub-Saharan Africa. The fluorescence property of this compound was used to investigate the interaction of the molecule with in vitro-derived bloodstream forms of Trypanosoma congolense IL 1180. Incubation of isometamidium with trypanosomes at 37 degrees C for 180 min resulted in a gradual alteration of the lambda max. with time (from 600 to 584 nm) and an increase in the intensity of trypanosome-associated fluorescence of approx. 2-fold. The alteration in fluorescence was temperature-dependent and inhibited by the addition of N-ethylmaleimide. In contrast, with intact cells addition of digitonin caused a rapid increase in fluorescence intensity to approximately four times that observed with intact cells. Uptake of isometamidium was also determined using radiolabelled drug; the results indicated that the time course of the uptake process resembled the fluorescence profile and was temperature-dependent. The results therefore indicate that the alteration of fluorescence is due to interaction of isometamidium with an intracellular component(s) and that isometamidium is transported across the plasma membrane via a protein carrier. The data also indicate that the described fluorescence technique can be used to investigate the role of membrane transport in resistance to isometamidium. PMID:8503859

  2. Fluorescence analysis of the interaction of isometamidium with Trypanosoma congolense.

    PubMed

    Zilberstein, D; Wilkes, J; Hirumi, H; Peregrine, A S

    1993-05-15

    Isometamidium chloride (Samorin) is the only compound recommended for prophylaxis against bovine trypanosomiasis in sub-Saharan Africa. The fluorescence property of this compound was used to investigate the interaction of the molecule with in vitro-derived bloodstream forms of Trypanosoma congolense IL 1180. Incubation of isometamidium with trypanosomes at 37 degrees C for 180 min resulted in a gradual alteration of the lambda max. with time (from 600 to 584 nm) and an increase in the intensity of trypanosome-associated fluorescence of approx. 2-fold. The alteration in fluorescence was temperature-dependent and inhibited by the addition of N-ethylmaleimide. In contrast, with intact cells addition of digitonin caused a rapid increase in fluorescence intensity to approximately four times that observed with intact cells. Uptake of isometamidium was also determined using radiolabelled drug; the results indicated that the time course of the uptake process resembled the fluorescence profile and was temperature-dependent. The results therefore indicate that the alteration of fluorescence is due to interaction of isometamidium with an intracellular component(s) and that isometamidium is transported across the plasma membrane via a protein carrier. The data also indicate that the described fluorescence technique can be used to investigate the role of membrane transport in resistance to isometamidium.

  3. Specific antibodies induce apoptosis in Trypanosoma cruzi epimastigotes.

    PubMed

    Fernández-Presas, Ana María; Tato, Patricia; Becker, Ingeborg; Solano, Sandra; Copitin, Natalia; Kopitin, Natalia; Berzunza, Miriam; Willms, Kaethe; Hernández, Joselin; Molinari, José Luis

    2010-05-01

    The susceptibility of Trypanosoma cruzi epimastigotes to lysis by normal or immune sera in a complement-dependent reaction has been reported. Mouse immune sera depleted complement-induced damage in epimastigotes characterized by morphological changes and death. The purpose of this work was to study the mechanism of death in epimastigotes exposed to decomplemented mouse immune serum. Epimastigotes were maintained in RPMI medium. Immune sera were prepared in mice by immunization with whole crude epimastigote extracts. Viable epimastigotes were incubated with decomplemented normal or immune sera at 37 degrees C. By electron microscopy, agglutinated parasites showed characteristic patterns of membrane fusion between two or more parasites; this fusion also produced interdigitation of the subpellicular microtubules. Apoptosis was determined by flow cytometry using terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and annexin V assays. Nuclear features were examined by 4'-,6-diamidino-2'-phenylindole diHCI cytochemistry that demonstrated apoptotic nuclear condensation. Caspase activity was also measured. TUNEL results showed that parasites incubated with decomplemented immune sera took up 26% of specific fluorescence as compared to 1.3% in parasites incubated with decomplemented normal sera. The Annexin-V-Fluos staining kit revealed that epimastigotes incubated with decomplemented immune sera exposed phosphatidylserine on the external leaflet of the plasma membrane. The incubation of parasites with immune sera showed caspase 3 activity. We conclude that specific antibodies are able to induce agglutination and apoptosis in epimastigotes, although the pathway is not elucidated.

  4. VSG gene expression site control in insect form Trypanosoma brucei.

    PubMed

    Rudenko, G; Blundell, P A; Taylor, M C; Kieft, R; Borst, P

    1994-11-15

    When the African trypanosome Trypanosoma brucei is taken up from mammals by a tse-tse fly, it replaces its variant surface glycoprotein (VSG) coat by a procyclin coat. Transcription of VSG genes stops in the fly, but transcription of sequences derived from the promoter area of the VSG expression site(s) remains high. Whether this is due to continuing high activity of one promoter or to low activity of many promoters was unclear. We have used the small differences between the sequences of different expression sites to show that multiple expression site promoters are active in insect form trypanosomes. This is confirmed by the low expression of single copy marker genes introduced into the transcribed area. However, if the expression site promoter is removed from the genomic location of the expression site and inserted in the non-transcribed spacer of the ribosomal DNA (rDNA), it is derepressed. Derepression of transcription can also be accomplished by replacing the promoter of an expression site by an rDNA promoter. We conclude that the down-regulation of VSG gene expression site promoters in insect form trypanosomes is affected by both the DNA sequence of the promoter and the genomic context in which it resides.

  5. Surface proteins, ERAD and antigenic variation in Trypanosoma brucei.

    PubMed

    Tiengwe, Calvin; Muratore, Katherine A; Bangs, James D

    2016-11-01

    Variant surface glycoprotein (VSG) is central to antigenic variation in African trypanosomes. Although much prior work documents that VSG is efficiently synthesized and exported to the cell surface, it was recently claimed that 2-3 fold more is synthesized than required, the excess being eliminated by ER-Associated Degradation (ERAD) (Field et al., ). We now reinvestigate VSG turnover and find no evidence for rapid degradation, consistent with a model whereby VSG synthesis is precisely regulated to match requirements for a functional surface coat on each daughter cell. However, using a mutated version of the ESAG7 subunit of the transferrin receptor (E7:Ty) we confirm functional ERAD in trypanosomes. E7:Ty fails to assemble into transferrin receptors and accumulates in the ER, consistent with retention of misfolded protein, and its turnover is selectively rescued by the proteasomal inhibitor MG132. We also show that ER accumulation of E7:Ty does not induce an unfolded protein response. These data, along with the presence of ERAD orthologues in the Trypanosoma brucei genome, confirm ERAD in trypanosomes. We discuss scenarios in which ERAD could be critical to bloodstream parasites, and how these may have contributed to the evolution of antigenic variation in trypanosomes.

  6. Pharmacological Validation of Trypanosoma brucei Phosphodiesterases as Novel Drug Targets

    PubMed Central

    de Koning, Harry P.; Gould, Matthew K.; Sterk, Geert Jan; Tenor, Hermann; Kunz, Stefan; Luginbuehl, Edith; Seebeck, Thomas

    2012-01-01

    The development of drugs for neglected infectious diseases often uses parasite-specific enzymes as targets. We here demonstrate that parasite enzymes with highly conserved human homologs may represent a promising reservoir of new potential drug targets. The cyclic nucleotide-specific phosphodiesterases (PDEs) of Trypanosoma brucei, causative agent of the fatal human sleeping sickness, are essential for the parasite. The highly conserved human homologs are well-established drug targets. We here describe what is to our knowledge the first pharmacological validation of trypanosomal PDEs as drug targets. High-throughput screening of a proprietary compound library identified a number of potent hits. One compound, the tetrahydrophthalazinone compound A (Cpd A), was further characterized. It causes a dramatic increase of intracellular cyclic adenosine monophosphate (cAMP). Short-term cell viability is not affected, but cell proliferation is inhibited immediately, and cell death occurs within 3 days. Cpd A prevents cytokinesis, resulting in multinucleated, multiflagellated cells that eventually lyse. These observations pharmacologically validate the highly conserved trypanosomal PDEs as potential drug targets. PMID:22291195

  7. Dehydroepiandrosterone increases resistance to experimental infection by Trypanosoma cruzi.

    PubMed

    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. Trypanosoma cruzi Calreticulin Topographical Variations in Parasites Infecting Murine Macrophages.

    PubMed

    González, Andrea; Valck, Carolina; Sánchez, Gittith; Härtel, Steffen; Mansilla, Jorge; Ramírez, Galia; Fernández, María Soledad; Arias, José Luis; Galanti, Norbel; Ferreira, Arturo

    2015-05-01

    Trypanosoma cruzi calreticulin (TcCRT), a 47-kDa chaperone, translocates from the endoplasmic reticulum to the area of flagellum emergence. There, it binds to complement components C1 and mannan-binding lectin (MBL), thus acting as a main virulence factor, and inhibits the classical and lectin pathways. The localization and functions of TcCRT, once the parasite is inside the host cell, are unknown. In parasites infecting murine macrophages, polyclonal anti-TcCRT antibodies detected TcCRT mainly in the parasite nucleus and kinetoplast. However, with a monoclonal antibody (E2G7), the resolution and specificity of the label markedly improved, and TcCRT was detected mainly in the parasite kinetoplast. Gold particles, bound to the respective antibodies, were used as probes in electron microscopy. This organelle may represent a stopover and accumulation site for TcCRT, previous its translocation to the area of flagellum emergence. Finally, early during T. cruzi infection and by unknown mechanisms, an important decrease in the number of MHC-I positive host cells was observed. © The American Society of Tropical Medicine and Hygiene.

  9. Trypanosoma cruzi Calreticulin Topographical Variations in Parasites Infecting Murine Macrophages

    PubMed Central

    González, Andrea; Valck, Carolina; Sánchez, Gittith; Härtel, Steffen; Mansilla, Jorge; Ramírez, Galia; Fernández, María Soledad; Arias, José Luis; Galanti, Norbel; Ferreira, Arturo

    2015-01-01

    Trypanosoma cruzi calreticulin (TcCRT), a 47-kDa chaperone, translocates from the endoplasmic reticulum to the area of flagellum emergence. There, it binds to complement components C1 and mannan-binding lectin (MBL), thus acting as a main virulence factor, and inhibits the classical and lectin pathways. The localization and functions of TcCRT, once the parasite is inside the host cell, are unknown. In parasites infecting murine macrophages, polyclonal anti-TcCRT antibodies detected TcCRT mainly in the parasite nucleus and kinetoplast. However, with a monoclonal antibody (E2G7), the resolution and specificity of the label markedly improved, and TcCRT was detected mainly in the parasite kinetoplast. Gold particles, bound to the respective antibodies, were used as probes in electron microscopy. This organelle may represent a stopover and accumulation site for TcCRT, previous its translocation to the area of flagellum emergence. Finally, early during T. cruzi infection and by unknown mechanisms, an important decrease in the number of MHC-I positive host cells was observed. PMID:25758653

  10. Tracking autophagy during proliferation and differentiation of Trypanosoma brucei

    PubMed Central

    Proto, William R.; Jones, Nathaniel G.; Coombs, Graham H.; Mottram, Jeremy C.

    2014-01-01

    Autophagy is a lysosome-dependent degradation mechanism that sequesters target cargo into autophagosomal vesicles. The Trypanosoma brucei genome contains apparent orthologues of several autophagy-related proteins including an ATG8 family. These ubiquitin-like proteins are required for autophagosome membrane formation, but our studies show that ATG8.3 is atypical. To investigate the function of other ATG proteins, RNAi compatible T. brucei were modified to function as autophagy reporter lines by expressing only either YFP-ATG8.1 or YFP-ATG8.2. In the insect procyclic lifecycle stage, independent RNAi down-regulation of ATG3 or ATG7 generated autophagy-defective mutants and confirmed a pro-survival role for autophagy in the procyclic form nutrient starvation response. Similarly, RNAi depletion of ATG5 or ATG7 in the bloodstream form disrupted autophagy, but did not impede proliferation. Further characterisation showed bloodstream form autophagy mutants retain the capacity to undergo the complex cellular remodelling that occurs during differentiation to the procyclic form and are equally susceptible to dihydroxyacetone-induced cell death as wild type parasites, not supporting a role for autophagy in this cell death mechanism. The RNAi reporter system developed, which also identified TOR1 as a negative regulator controlling YFP-ATG8.2 but not YFP-ATG8.1 autophagosome formation, will enable further targeted analysis of the mechanisms and function of autophagy in the medically relevant bloodstream form of T. brucei. PMID:28357206

  11. Potent Anti-Trypanosoma cruzi Activities of Oxidosqualene Cyclase Inhibitors

    PubMed Central

    Buckner, Frederick S.; Griffin, John H.; Wilson, Aaron J.; Van Voorhis, Wesley C.

    2001-01-01

    Trypanosoma cruzi is the protozoan agent that causes Chagas' disease, a major health problem in Latin America. Better drugs are needed to treat infected individuals. The sterol biosynthesis pathway is a potentially excellent target for drug therapy against T. cruzi. In this study, we investigated the antitrypanosomal activities of a series of compounds designed to inhibit a key enzyme in sterol biosynthesis, oxidosqualene cyclase. This enzyme converts 2,3-oxidosqualene to the tetracyclic product, lanosterol. The lead compound, N-(4E,8E)-5,9, 13-trimethyl-4,8, 12-tetradecatrien-1-ylpyridinium, is an electron-poor aromatic mimic of a monocyclized transition state or high-energy intermediate formed from oxidosqualene. This compound and 27 related compounds were tested against mammalian-stage T. cruzi, and 12 inhibited growth by 50% at concentrations below 25 nM. The lead compound was shown to cause an accumulation of oxidosqualene and decreased production of lanosterol and ergosterol, consistent with specific inhibition of the oxidosqualene cyclase. The data demonstrate potent anti-T. cruzi activity associated with inhibition of oxidosqualene cyclase. PMID:11257036

  12. Phospholipid and glycolipid composition of acidocalcisomes of Trypanosoma cruzi.

    PubMed

    Salto, María Laura; Kuhlenschmidt, Theresa; Kuhlenschmidt, Mark; de Lederkremer, Rosa M; Docampo, Roberto

    2008-04-01

    Highly purified acidocalcisomes from Trypanosoma cruzi epimastigotes were obtained by differential centrifugation and iodixanol gradient ultracentrifugation. Lipid analysis of acidocalcisomes revealed the presence of low amounts of 3beta-hydroxysterols and predominance of phospholipids. Alkylacyl phosphatidylinositol (16:0/18:2), diacyl phosphatidylinositol (18:0/18:2), diacyl phosphatidylcholine (16:0/18:2; 16:1/18:2; 16:2/18:2; 18:1/18:2 and 18:2/18:2), and diacyl phosphatidylethanolamine (16:0/18:2 and 16:1/18:2) were the only phospholipids characterized by electrospray ionization-mass spectrometry (ESI-MS). Incubation of epimastigotes with [(3)H]-mannose and isolation of acidocalcisomes allowed the detection of a glycoinositolphospholipid (GIPL) in these organelles. The sugar content of the acidocalcisomal GIPL was similar to that of the GIPL present in a microsomal fraction but the amount of galactofuranose and inositol with respect to the other monosaccharides was lower, suggesting a different chemical structure. Taken together, these results indicate that acidocalcisomes of T. cruzi have a distinct lipid and carbohydrate composition.

  13. Trypanocidal activity of free and nanoencapsulated curcumin on Trypanosoma evansi.

    PubMed

    Gressler, L T; Oliveira, C B; Coradini, K; Dalla Rosa, L; Grando, T H; Baldissera, M D; Zimmermann, C E; Da Silva, A S; Almeida, T C; Hermes, C L; Wolkmer, P; Silva, C B; Moreira, K L S; Beck, R C R; Moresco, R N; Da Veiga, M L; Stefani, L M; Monteiro, S G

    2015-03-01

    This study aimed to evaluate in vitro and in vivo trypanocidal activity of free and nanoencapsulated curcumin against Trypanosoma evansi. In vitro efficacy of free curcumin (CURC) and curcumin-loaded in lipid-core nanocapsules (C-LNCs) was evaluated to verify their lethal effect on T. evansi. To perform the in vivo tests, T. evansi-infected animals were treated with CURC (10 and 100 mg kg(-1), intraperitoneally [i.p.]) and C-LNCs (10 mg kg(-1), i.p.) during 6 days, with the results showing that these treatments significantly attenuated the parasitaemia. Infected untreated rats showed protein peroxidation and an increase of nitrites/nitrates, whereas animals treated with curcumin showed a reduction on these variables. As a result, the activity of antioxidant enzymes (superoxide dismutase and catalase) differs between groups (P<0.05). Infected animals and treated with CURC exhibited a reduction in the levels of alanine aminotransferase and creatinine, when compared with the positive control group. The use of curcumin in vitro resulted in a better parasitaemia control, an antioxidant activity and a protective effect on liver and kidney functions of T. evansi-infected adult male Wistar rats.

  14. Imidazolium compounds are active against all stages of Trypanosoma cruzi.

    PubMed

    Faral-Tello, Paula; Liang, Mary; Mahler, Graciela; Wipf, Peter; Robello, Carlos

    2014-03-01

    Imidazolium salts are best known for their applications in organic synthesis as room-temperature ionic liquids, or as precursors of stable carbenes, but they also show important biological properties such as anti-oxidative effects, induction of mitochondrial membrane permeabilisation and inhibition of the infection cycle of Plasmodium falciparum. For these reasons, and since chemotherapy for Chagas disease is inefficient, the aim of this study was to test the use of imidazolium compounds against the kinetoplastid haemoflagellate aetiological agent for this disease, namely Trypanosoma cruzi. The results show that five of the tested compounds are more effective than the reference drug benznidazole against the epimastigote and trypomastigote forms of T. cruzi. Moreover, intracellular amastigotes were also affected by the compounds, which showed lower toxicity in host cells. Transmission electron microscopy analysis demonstrated that the tested agents induced alterations of the kinetoplast and particularly of the mitochondria, leading to extraordinary swelling of the organelle. These results further demonstrate that the test agents with the best profile are those bearing symmetrical bulky substituents at N(1) and N(3), displaying promising activity against all forms of T. cruzi, interesting selectivity indexes and exceptional activity at low doses. Accordingly, these agents represent promising candidates for the treatment of Chagas disease.

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

    PubMed Central

    Brandan, Cecilia Pérez; 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. PMID:22705838

  16. Phospholipid and glycolipid composition of acidocalcisomes of Trypanosoma cruzi

    PubMed Central

    Salto, María Laura; Kuhlenschmidt, Theresa; Kuhlenschmidt, Mark; de Lederkremer, Rosa M.; Docampo, Roberto

    2008-01-01

    Highly purified acidocalcisomes from Trypanosoma cruzi epimastigotes were obtained by differential centrifugation and iodixanol gradient ultracentrifugation. Lipid analysis of acidocalcisomes revealed the presence of low amounts of 3β-hydroxysterols and predominance of phospholipids. Alkylacyl phosphatidylinositol (16:0/18:2), diacyl phosphatidylinositol (18:0/18:2), diacyl phosphatidylcholine (16:0/18:2; 16:1/18:2; 16:2/18:2, 18:1/18:2, and 18:2/18:2), and diacyl phosphatidylethanolamine (16:0/18:2 and 16:1/18:2) were the only phospholipids characterized by electrospray ionization-mass spectrometry (ESI-MS). Incubation of epimastigotes with [3H]-mannose and isolation of acidocalcisomes allowed the detection of a glycoinositolphospholipid (GIPL) in these organelles. The sugar content of the acidocalcisomal GIPL was similar to that of the GIPL present in a microsomal fraction but the amount of galactofuranose and inositol with respect to the other monosaccharides was lower, suggesting a different chemical structure. Taken together, these results indicate that acidocalcisomes of T. cruzi have a distinct lipid and carbohydrate composition. PMID:18207579

  17. Targeting Trypanosoma cruzi Sterol 14α-Demethylase (CYP51)

    PubMed Central

    Lepesheva, Galina I.; Villalta, Fernando; Waterman, Michael R.

    2012-01-01

    There are at least two obvious features that must be considered upon targeting specific metabolic pathways/enzymes for drug development: the pathway must be essential and the enzyme must allow the design of pharmacologically useful inhibitors. Here, we describe Trypanosoma cruzi sterol 14α-demethylase as a promising target for anti-Chagasic chemotherapy. The use of anti-fungal azoles, which block sterol biosynthesis and therefore membrane formation in fungi, against the protozoan parasite has turned out to be highly successful: a broad spectrum anti-fungal drug, the triazole compound posaconazole, is now entering phase II clinical trials for treatment of Chagas disease. This review summarizes comparative information on anti-fungal azoles and novel inhibitory scaffolds selective for Trypanosomatidae sterol 14α-demethylase through the lens of recent structure/functional characterization of the target enzyme. We believe our studies open wide opportunities for rational design of novel, pathogen-specific and therefore more potent and efficient anti-trypanosomal drugs. PMID:21820552

  18. Molecular characterisation of Trypanosoma brucei alkyl dihydroxyacetone-phosphate synthase.

    PubMed

    Zomer, A W; Michels, P A; Opperdoes, F R

    1999-10-25

    Alkyl dihydroxyacetone-phosphate synthase is the second enzyme of the ether-lipid biosynthetic pathway which is responsible for the introduction of the ether linkage between a fatty alcohol and a glycerol present in a subclass of phospholipids, the plasmalogens and possibly in glycolipid membrane anchors. In this study the gene coding for alkyl dihydroxyacetone-phosphate synthase was isolated from Trypanosoma brucei. Southern blot analysis of total genomic DNA suggested the presence of a single copy gene. The analysis, together with sequencing of different cDNA clones showed that the two alleles of the gene differ in only one nucleotide. The gene encodes a protein of 612 amino acids with a calculated molecular mass of 68,891, not counting the initiator methionine. It carries a type-1 peroxisomal targeting signal (a C-terminal tripeptide--AHL) and a calculated overall positive charge of +10. The gene was expressed in a bacterial system and the corresponding protein carrying a His-tag was purified. The recombinant alkyl dihydroxyacetone-phosphate synthase and the enzyme isolated directly from the glycosomes of bloodstream-form trypanosomes have comparable kinetics. The Km for hexadecanol was 42 microM, while approximately 100 microM of palmitoyl dihydroxyacetone phosphate (DHAP) was necessary for optimal activity. Sodium chloride inhibited both the His-tagged protein and the enzyme isolated from the glycosomes of bloodstream-form and insect stage T. brucei.

  19. Trypanosoma cruzi Gene Expression in Response to Gamma Radiation

    PubMed Central

    Grynberg, Priscila; Passos-Silva, Danielle Gomes; Mourão, Marina de Moraes; Hirata Jr, Roberto; Macedo, Andrea Mara; Machado, Carlos Renato; Bartholomeu, Daniella Castanheira; Franco, Glória Regina

    2012-01-01

    Trypanosoma cruzi is an organism highly resistant to ionizing radiation. Following a dose of 500 Gy of gamma radiation, the fragmented genomic DNA is gradually reconstructed and the pattern of chromosomal bands is restored in less than 48 hours. Cell growth arrests after irradiation but, while DNA is completely fragmented, RNA maintains its integrity. In this work we compared the transcriptional profiles of irradiated and non-irradiated epimastigotes at different time points after irradiation using microarray. In total, 273 genes were differentially expressed; from these, 160 were up-regulated and 113 down-regulated. We found that genes with predicted functions are the most prevalent in the down-regulated gene category. Translation and protein metabolic processes, as well as generation of precursor of metabolites and energy pathways were affected. In contrast, the up-regulated category was mainly composed of obsolete sequences (which included some genes of the kinetoplast DNA), genes coding for hypothetical proteins, and Retrotransposon Hot Spot genes. Finally, the tyrosyl-DNA phosphodiesterase 1, a gene involved in double-strand DNA break repair process, was up-regulated. Our study demonstrated the peculiar response to ionizing radiation, raising questions about how this organism changes its gene expression to manage such a harmful stress. PMID:22247781

  20. The Oral Antimalarial Drug Tafenoquine Shows Activity against Trypanosoma brucei.

    PubMed

    Carvalho, Luis; Martínez-García, Marta; Pérez-Victoria, Ignacio; Manzano, José Ignacio; Yardley, Vanessa; Gamarro, Francisco; Pérez-Victoria, José M

    2015-10-01

    The protozoan parasite Trypanosoma brucei causes human African trypanosomiasis, or sleeping sickness, a neglected tropical disease that requires new, safer, and more effective treatments. Repurposing oral drugs could reduce both the time and cost involved in sleeping sickness drug discovery. Tafenoquine (TFQ) is an oral antimalarial drug belonging to the 8-aminoquinoline family which is currently in clinical phase III. We show here that TFQ efficiently kills different T. brucei spp. in the submicromolar concentration range. Our results suggest that TFQ accumulates into acidic compartments and induces a necrotic process involving cell membrane disintegration and loss of cytoplasmic content, leading to parasite death. Cell lysis is preceded by a wide and multitarget drug action, affecting the lysosome, mitochondria, and acidocalcisomes and inducing a depolarization of the mitochondrial membrane potential, elevation of intracellular Ca(2+), and production of reactive oxygen species. This is the first report of an 8-aminoquinoline demonstrating significant in vitro activity against T. brucei. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  1. Resistance to drug by different isolates Trypanosoma evansi in China.

    PubMed

    Zhou, Jinlin; Shen, Jie; Liao, Dangjin; Zhou, Yongzhi; Lin, Jiaojiao

    2004-05-01

    In order to understand drug resistance in Chinese Trypanosoma evansi isolates, the in vitro growth inhibition test was carried out to detect the sensitivities to suramin and antrycide of 12 T. evansi isolates. For suramin, 50% inhibitory concentrations (IC(50)) ranged from 0.041 to 0.752 microg/ml; for antrycide, the IC(50) values ranged from 0.00151 to 0.06694 microg/ml. In vivo experiments in mice with selected isolates showed that the isolate most resistant to suramin was not cured at a dosage of 10 mg/kg, while the isolate that was most resistant to antrycide showed only 50% cure rate at a dose rate of 5 mg/kg. The data presented here indicate that T. evansi isolates from China had differences in drug resistance, with some isolates exhibiting low sensitivity to suramin or antrycide, while a few isolates showed complete drug resistance to curative dosages of suramin or antrycide. No drug cross-resistance was observed between suramin and antrycide.

  2. Trypanosoma cruzi Antioxidant Enzymes As Virulence Factors in Chagas Disease

    PubMed Central

    Piacenza, Lucía; Peluffo, Gonzalo; Alvarez, María Noel; Martínez, Alejandra

    2013-01-01

    Abstract Significance: Chagas disease (CD) affects several million people in Latin America and is spreading beyond its classical boundaries due to the migration of infected host and insect vectors, HIV co-infection, and blood transfusion. The current therapy is not adequate for treatment of the chronic phase of CD, and new drugs are warranted. Recent Advances: Trypanosoma cruzi is equipped with a specialized and complex network of antioxidant enzymes that are located at different subcellular compartments which defend the parasite against host oxidative assaults. Recently, strong evidence has emerged which indicates that enzyme components of the T. cruzi antioxidant network (cytosolic and mitochondrial peroxiredoxins and trypanothione synthetase) in naturally occurring strains act as a virulence factor for CD. This precept is recapitulated with the observed increased resistance of T. cruzi peroxirredoxins overexpressers to in vivo or in vitro nitroxidative stress conditions. In addition, the modulation of mitochondrial superoxide radical levels by iron superoxide dismutase (FeSODA) influences parasite programmed cell death, underscoring the role of this enzyme in parasite survival. Critical Issues: The unraveling of the biological significance of FeSODs in T. cruzi programmed cell death in the context of chronic infection in CD is still under examination. Future Directions: The role of the antioxidant enzymes in the pathogenesis of CD, including parasite virulence and persistence, and their feasibility as pharmacological targets justifies further investigation. Antioxid. Redox Signal. 19, 723–734. PMID:22458250

  3. The effect of placental subfractions on Trypanosoma cruzi.

    PubMed

    Frank, F; Sartori, M J; Asteggiano, C; Lin, S; de Fabro, S P; Fretes, R E

    2000-10-01

    Five subfractions were collected from six term placentas by mincing and differential centrifugation: homogenate, nuclear, mitochondrial, lysosomal, and supernatant. The effect of each subfraction on Trypanosoma cruzi was assessed by trypan blue exclusion, relative infectivity of mice, and penetration of susceptible cultured VERO cells. Ultrastructural changes in trypomastigotes were identified after high cell mortality was shown by dye exclusion following treatment with lysosomal and supernatant fractions. Trypomastigotes treated with other subfractions or preheated subfractions, those recovered from infected VERO cells, and controls remained unaffected. This was confirmed by the ability of treated trypomastigotes to infect mice or to penetrate susceptible cultured VERO cells. There were a 48% decrease in parasitemia and fewer myocardial lesions in Balb/c mice following treatment with the lysosomal subfraction compared to homogenate and controls. VERO cells were invaded about half as often after lysosomal treatment compared to controls (P < 0. 05); an 11% decrease in cell invasion following homogenate treatment was not significant. Placental lysosomal enzyme activity was unaffected by trypomastigotes. Human placentas contain one or more heat-labile substances in lysosomal and supernatant subfractions which inhibit or injure trypomastigotes of T. cruzi in cell-free systems. Copyright 2000 Academic Press.

  4. Trypanosoma cruzi infection induces microfilament depletion in human placenta syncytiotrophoblast.

    PubMed

    Sartori, M J; Pons, P; Mezzano, L; Lin, S; de Fabro, S P

    2003-08-01

    Congenital Chagas disease, endemic in Latin America, is associated with premature labour, miscarriage, and placentitis. Metacyclic trypomastigotes adhere to specific receptors on the outer membrane of host cells as a prelude to intracellular invasion, causing calcium ion mobilization, rearrangement of host cell microfilaments, recruitment of lysosomes and parasite internalization. The actin cytoskeleton plays an important role in many cellular processes including the parasite invasion into mammalian cells. In order to observe if placental cytoskeleton is altered in the process of parasite invasion into placental villi, actin microfilaments were studied. Using immunohistochemical techniques, it was observed that the presence of actin in the syncytiotrophoblast was intense throughout the brush border in control placentae belonging to non-chagasic women. But after culture with the trypomastigote, this labelling disappeared, indicating that the parasite induced disassembly of the cortical actin cytoskeleton when the placenta was infected. As a control, placentae from chagasic women were studied, and no actin was found. The same results were obtained by the electron microscope. We confirmed that cortical actin rearrangements may be an early step in the Trypanosoma cruzi invasion mechanism into placental cells, in order to allow lysosomes access to the plasma membrane, and formation of the parasitophorous vacuole. The recruitment of lysosomes occurs directly beneath the invasion site, and this process is required for parasite internalization.

  5. [Digestive tract dilation in mice infected with Trypanosoma cruzi].

    PubMed

    Guillén-Pernía, B; Lugo-Yarbuh, A; Moreno, E

    2001-09-01

    This paper will analyze alterations in the digestive tract (DT) of mice with chronic Chagas' disease infection produced by Trypanosoma cruzi from different sources. X-rays of the DT of 18 mice infected with T. cruzi and 6 control mice were compared after the ingestion of a barium sulfate solution over a period of 6 hours. 120 days post-infection (pi) the X-rays of the DT of the 5 mice of group 1A infected with trypanosomes DMI isolated from the opossum Didelphis marsupialis, and 4 mice in group 2A infected with the isolate EP taken from a patient with acute Chagas' disease, showed swelling of the stomach and the colon (C). 180 days pi, the X-rays of the DT of the 5 mice of group 1B infected with isolated DMI and the 4 mice in group 2B infected with isolate EP, showed an even greater swelling of the C. Histological examination of the DT of all infected mice showed extensive changes of the intestinal muscle layer, such as the diminution of the muscular and mucous layers and the loss of colonic folds and myoenteric plexus. These results suggest that T. cruzi populations caused severe alterations in the digestive system of the mice used in the experiment, and that the same alterations could occur in the digestive organs of humans, especially those living in areas where Chagas' disease is endemic, but where these abnormalities have not yet been reported.

  6. Interferon-gamma promotes infection of astrocytes by Trypanosoma cruzi.

    PubMed

    Silva, Rafael Rodrigues; Mariante, Rafael M; Silva, Andrea Alice; dos Santos, Ana Luiza Barbosa; Roffê, Ester; Santiago, Helton; Gazzinelli, Ricardo Tostes; Lannes-Vieira, Joseli

    2015-01-01

    The inflammatory cytokine interferon-gamma (IFNγ) is crucial for immunity against intracellular pathogens such as the protozoan parasite Trypanosoma cruzi, the causative agent of Chagas disease (CD). IFNγ is a pleiotropic cytokine which regulates activation of immune and non-immune cells; however, the effect of IFNγ in the central nervous system (CNS) and astrocytes during CD is unknown. Here we show that parasite persists in the CNS of C3H/He mice chronically infected with the Colombian T. cruzi strain despite the increased expression of IFNγ mRNA. Furthermore, most of the T. cruzi-bearing cells were astrocytes located near IFNγ+ cells. Surprisingly, in vitro experiments revealed that pretreatment with IFNγ promoted the infection of astrocytes by T. cruzi increasing uptake and proliferation of intracellular forms, despite inducing increased production of nitric oxide (NO). Importantly, the effect of IFNγ on T. cruzi uptake and growth is completely blocked by the anti-tumor necrosis factor (TNF) antibody Infliximab and partially blocked by the inhibitor of nitric oxide synthesis L-NAME. These data support that IFNγ fuels astrocyte infection by T. cruzi and critically implicate IFNγ-stimulated T. cruzi-infected astrocytes as sources of TNF and NO, which may contribute to parasite persistence and CNS pathology in CD.

  7. [Trypanosoma cruzi in triatomines from Nuevo Leon, Mexico].

    PubMed

    Molina-Garza, Zinnia Judith; Rosales-Encina, José Luis; Galaviz-Silva, Lucio; Molina-Garza, Daniel

    2007-01-01

    To determine the prevalence of Trypanosoma cruzi in triatomines from Nuevo León using the standardization of an improved enzyme-linked immunosorbent assay test. From July to September 2005, 52 triatomines were captured in General Terán, a municipality located in Nuevo León. They were analyzed using optical microscopy (OM) and a polymerase chain reaction (PCR), as standards of reference, to develop a technique for detecting the parasite using enzyme-linked immunosorbent assay (ELISA). Using OM and PCR, 31 triatomines were found to be positive and 21 negative. Using ELISA, 27 samples were identified as positive and 25 negative (specificity 100%, sensitivity 87%, negative predictive value 84%, and positive predictive value 100%). The prevalence of infected triatomines was 59.61% with OM and PCR, and 51.92% with ELISA. Our data confirm that the ELISA assay in triatomines is a fast, reliable and useful tool. Since it was possible to simultaneously analyze a large number of samples with high sensibility and specificity values, the ELISA test proves to be useful for new epidemiologic studies having a high number of vectors. It is also less expensive than PCR. It is therefore recommended for epidemiological and preventive surveillance programs as a first screening test before conducting a confirmatory test using PCR.

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

    PubMed

    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.

  9. Crystal structure of triosephosphate isomerase from Trypanosoma cruzi in hexane

    PubMed Central

    Gao, Xiu-Gong; Maldonado, Ernesto; Pérez-Montfort, Ruy; Garza-Ramos, Georgina; de Gómez-Puyou, Marietta Tuena; Gómez-Puyou, Armando; Rodríguez-Romero, Adela

    1999-01-01

    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-Å 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 Å 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. PMID:10468562

  10. Murine models susceptibility to distinct Trypanosoma cruzi I genotypes infection.

    PubMed

    León, Cielo M; Montilla, Marleny; Vanegas, Ricardo; Castillo, Maria; Parra, Edgar; Ramírez, Juan David

    2017-04-01

    Chagas disease is a complex zoonosis that affects around 8 million people worldwide. This pathology is caused by Trypanosoma cruzi, a kinetoplastid parasite that shows tremendous genetic diversity evinced in six distinct Discrete Typing Units (TcI-TcVI) including a recent genotype named as TcBat and associated with anthropogenic bats. TcI presents a broad geographical distribution and has been associated with chronic cardiomyopathy. Recent phylogenetic studies suggest the existence of two genotypes (Domestic (TcIDom) and sylvatic TcI) within TcI. The understanding of the course of the infection in different mouse models by these two genotypes is not yet known. Therefore, we infected 126 animals (ICR-CD1, National Institute of Health (NIH) and Balb/c) with two TcIDom strains and one sylvatic strain for a follow-up period of 60 days. We quantified the parasitaemia, immune response and histopathology observing that the maximum day of parasitaemia was achieved at day 21 post-infection. Domestic strains showed higher parasitaemia than the sylvatic strain in the three mouse models; however in the survival curves Balb/c mice were less susceptible to infection compared with NIH and ICR-CD1. Our results suggest that the genetic background plays a fundamental role in the natural history of the infection and the sympatric TcI genotypes have relevant implications in disease pathogenesis.

  11. Trypanosoma cruzi infection: a review with emphasis on cutaneous manifestations

    PubMed Central

    Hemmige, Vagish; Tanowitz, Herbert; Sethi, Aisha

    2013-01-01

    Chagas disease, an infection caused by the protozoan Trypanosoma cruzi and transmitted by the Reduuvid insect vector, remains a major cause of morbidity in Central and South America over a century after its discovery in 1909. Though major advances in preventing the spread of this disease have been made in recent decades, millions of individuals remain chronically infected due to prior exposure to T. cruzi and are at risk for future complications from the disease. Dermatologic manifestations of acute infection may include localized swelling at the site of inoculation (chagoma), conjunctivitis (Romaña’s sign), and a generalized morbilliform eruption (schizotrypanides). Reactivation of quiescent infection in immunocompromised hosts due to the acquired immunodeficiency syndrome or organ transplantation can present with fever and skin lesions including panniculitis. The wide-spread emigration of chronic carriers of T. cruzi to North America, Europe, and Australia makes it imperative that dermatologists worldwide be familiar with this entity to ensure proper diagnosis and treatment. PMID:22515575

  12. Synthesis and anti-Trypanosoma cruzi activity of diaryldiazepines.

    PubMed

    Menezes, Júlio César L; Vaz, Luana Beatriz A; de Abreu Vieira, Paula Melo; da Silva Fonseca, Kátia; Carneiro, Cláudia Martins; Taylor, Jason G

    2014-12-23

    Chagas disease is a so-called "neglected disease" and endemic to Latin America. Nifurtimox and benznidazole are drugs that have considerable efficacy in the treatment of the acute phase of the disease but cause many significant side effects. Furthermore, in the Chronic Phase its efficiency is reduced and their therapeutic effectiveness is dependent on the type of T. cruzi strain. For this reason, the present work aims to drive basic research towards the discovery of new chemical entities to treat Chagas disease. Differently substituted 5,7-diaryl-2,3-dihydro-1,4-diazepines were synthesized by cyclocondensation of substituted flavones with ethylenediamine and tested as anti-Trypanosoma cruzi candidates. Epimastigotes of the Y strain from T. cruzi were used in this study and the number of parasites was determined in a Neubauer chamber. The most potent diaryldiazepine that reduced epimastigote proliferation exhibited an IC50 value of 0.25 μM, which is significantly more active than benznidazole.

  13. Blood viscosity changes in experimentally Trypanosoma cruzi-infected rats.

    PubMed

    Berra, H H; Piaggio, E; Revelli, S S; Luquita, A

    2005-01-01

    Microcirculatory alterations would explain focal lesions found in Chagas' cardiomyopathy. Trypanosoma cruzi (T. cruzi) infection induces host blood properties modifications and defensive responses capable of producing blood hyperviscosity, an ischemic risk factor able to affect microvascular blood flow. We studied whole blood viscosity (eta(b)) and plasmatic and cellular factors influencing it in rats, 7 and 14 days after experimental infection with T. cruzi. Increased plasma viscosity (eta(p)) was found in infected versus control rats and it was correlated with high blood parasite levels at 7 days and enhanced gamma-globulin fraction concentration at 14 days. The hematocrit, mean corpuscular volume (MCV) and eta(b) were higher in 14 days infected rats vs. 7 days and control animals. Also, electron microscopy observation showed morphological changes in red blood cells (RBC) at 7 and 14 days post-infection, with increased proportion of echinocyte and stomatocyte shapes transformation. In our rat model of Chagas' disease, BPL, increased plasmatic protein concentration, enhanced MCV and RBC shapes transformation would determine blood hyperviscosity, cause of microvascular blood flow abnormalities.

  14. Trypanosoma avium of raptors (Falconiformes): phylogeny and identification of vectors.

    PubMed

    Votýpka, J; Oborník, M; Volf, P; Svobodová, M; Lukes, J

    2002-09-01

    Avian trypanosomes are widespread parasites of birds, the transmission of which remains mostly unclear, with various blood-sucking insects mentioned as possible vectors. A search for vectors of trypanosomes of sparrowhawk (Accipiter nisus), buzzard (Buteo buteo), lesser-spotted eagle (Aquila pomarina) and kestrel (Falco tinnunculus) was performed in Czech and Slovak Republics. Black flies (Eusimulium spp.), hippoboscid flies (Ornithomyia avicularia), mosquitoes (Culex pipiens pipiens) and biting midges (Culicoides spp.), trapped while attempting to feed on raptor nestlings, were found to contain trypanosomatids in their intestine. Trypanosomes from the raptors and blood-sucking insects were isolated, and their 18S rRNA sequences were used for species identification and for the inference of intra- and interspecific relationships. Together with the trypanosome isolated from a black fly, the bird trypanosomes formed a well-supported Trypanosoma avium clade. The isolates derived from hippoboscid flies and mosquitoes are most likely also avian trypanosomes infecting birds other than the studied raptors. Analysis of the kinetoplast, that has features characteristic for the avian trypanosomes (minicircle size; dimensions of the kinetoplast disc), provided further evidence for the identification of vectors. It is suggested that all trypanosomes isolated from raptors included in this study belong to the T. avium complex and are transmitted by the ornithophilic simuliids such as Eusimulium securiforme.

  15. Retrospective distribution of Trypanosoma cruzi I genotypes in Colombia

    PubMed Central

    León, Cielo M; Hernández, Carolina; Montilla, Marleny; Ramírez, Juan David

    2015-01-01

    Trypanosoma cruzi is the aetiological agent of Chagas disease, which affects approximately eight million people in the Americas. This parasite exhibits genetic variability, with at least six discrete typing units broadly distributed in the American continent. T. cruzi I (TcI) shows remarkable genetic diversity; a genotype linked to human infections and a domestic cycle of transmission have recently been identified, hence, this strain was named TcIDom. The aim of this work was to describe the spatiotemporal distribution of TcI subpopulations across humans, insect vectors and mammalian reservoirs in Colombia by means of molecular typing targeting the spliced leader intergenic region of mini-exon gene. We analysed 101 TcI isolates and observed a distribution of sylvatic TcI in 70% and TcIDom in 30%. In humans, the ratio was sylvatic TcI in 60% and TcIDom in 40%. In mammal reservoirs, the distribution corresponded to sylvatic TcI in 96% and TcIDom in 4%. Among insect vectors, sylvatic TcI was observed in 48% and TcIDom in 52%. In conclusion, the circulation of TcIDom is emerging in Colombia and this genotype is still adapting to the domestic cycle of transmission. The epidemiological and clinical implications of these findings are discussed herein. PMID:25946157

  16. Clonal population structure of Colombian sylvatic Trypanosoma cruzi.

    PubMed

    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.

  17. The Con Test

    ERIC Educational Resources Information Center

    Fletcher, Michael

    2009-01-01

    In this article, the author describes the format of the Con Test, an Australian television game show which followed the same general rules and game play as the UK show PokerFace. At the end of each round a contestant needs to decide whether or not he or she should fold. A contestant needs to know how likely it is that he or she is in last place.…

  18. Trypanosoma brucei prenylated-protein carboxyl methyltransferase prefers farnesylated substrates.

    PubMed Central

    Buckner, Frederick S; Kateete, David P; Lubega, George W; Van Voorhis, Wesley C; Yokoyama, Kohei

    2002-01-01

    Carboxyl methylation of the C-terminal prenylated cysteine, which occurs in most farnesylated and geranylgeranylated proteins, is a reversible step and is implicated in the regulation of membrane binding and cellular functions of prenylated proteins such as GTPases. The gene coding for prenylated-protein carboxyl methyltransferase (PPMT) of the protozoan parasite Trypanosoma brucei has been cloned and expressed in the baculovirus/Sf9 cell system. The protein of 245 amino acids has 24-28% sequence identity to the orthologues from other species including human and Saccharomyces cerevisiae. Methyltransferase activity was detected in the membrane fraction from Sf9 cells infected with the recombinant baculovirus using N -acetyl- S -farnesylcysteine (AFC) and S -adenosyl[ methyl -(3)H]methionine ([(3)H]AdoMet) as substrates. Recombinant T. brucei PPMT prefers AFC to N -acetyl- S -geranylgeranylcysteine (AGGC) by 10-50-fold based on the V (max)/ K (m) values. Native PPMT activity detected in the membrane fraction from T. brucei procyclics displays similar substrate specificity ( approximately 40-fold preference for AFC over AGGC). In contrast, mouse liver PPMT utilizes both AFC and AGGC as substrates with similar catalytic efficiencies. Several cellular proteins of the T. brucei bloodstream form were shown to be carboxyl methylated in a cell-free system. Incorporation of [(3)H]methyl group from [(3)H]AdoMet into most of the proteins was significantly inhibited by AFC but not AGGC at 20 microM, suggesting that T. brucei PPMT acts on farnesylated proteins in the cell. Cells of the T. brucei bloodstream form show higher sensitivity to AFC and AGGC (EC(50)=70-80 microM) compared with mouse 3T3 cells (EC(50)>150 microM). PMID:12141948

  19. Arginine and Lysine Transporters Are Essential for Trypanosoma brucei

    PubMed Central

    Hürlimann, Daniel; Wirdnam, Corina; Haindrich, Alexander C.; Suter Grotemeyer, Marianne; González-Salgado, Amaia; Schmidt, Remo S.; Inbar, Ehud; Mäser, Pascal; Bütikofer, Peter; Zilberstein, Dan; Rentsch, Doris

    2017-01-01

    For Trypanosoma brucei arginine and lysine are essential amino acids and therefore have to be imported from the host. Heterologous expression in Saccharomyces cerevisiae mutants identified cationic amino acid transporters among members of the T. brucei AAAP (amino acid/auxin permease) family. TbAAT5-3 showed high affinity arginine uptake (Km 3.6 ± 0.4 μM) and high selectivity for L-arginine. L-arginine transport was reduced by a 10-times excess of L-arginine, homo-arginine, canavanine or arginine-β-naphthylamide, while lysine was inhibitory only at 100-times excess, and histidine or ornithine did not reduce arginine uptake rates significantly. TbAAT16-1 is a high affinity (Km 4.3 ± 0.5 μM) and highly selective L-lysine transporter and of the compounds tested, only L-lysine and thialysine were competing for L-lysine uptake. TbAAT5-3 and TbAAT16-1 are expressed in both procyclic and bloodstream form T. brucei and cMyc-tagged proteins indicate localization at the plasma membrane. RNAi-mediated down-regulation of TbAAT5 and TbAAT16 in bloodstream form trypanosomes resulted in growth arrest, demonstrating that TbAAT5-mediated arginine and TbAAT16-mediated lysine transport are essential for T. brucei. Growth of induced RNAi lines could partially be rescued by supplementing a surplus of arginine or lysine, respectively, while addition of both amino acids was less efficient. Single and double RNAi lines indicate that additional low affinity uptake systems for arginine and lysine are present in T. brucei. PMID:28045943

  20. OBSERVATIONS ON THE RESPIRATION OF TRYPANOSOMA CRUZI IN CULTURE

    PubMed Central

    von Brand, Theodor; Johnson, Eleanor M.; Rees, Charles W.

    1946-01-01

    1. The oxygen consumption of cultural forms of Trypanosoma cruzi decreases in intensity with increasing age of the cultures; no correlation with any other factor studied could be established. 2. The respiratory quotient was high for the first 10 days, i.e. as long as the population increased; with the onset of a decline in numbers, the R.Q. began to drop. It is believed that the flagellates consume in the beginning predominantly sugar and later predominantly protein. Observations on the pH of the cultures bear out this view. 3. The oxygen consumption was independent of the oxygen tension over a wide range of tensions. 4. The oxygen consumption increased in the temperature range 13° to 40°C., while a temperature of 44°C. proved to be lethal. Upon application of Arrhenius' equation, two straight lines, intersecting at about 28°C., resulted. The µ values were 23,980 and 5275 for the lower and higher temperature range respectively. 5. Of the oxidase inhibitors tested, strong inhibition of the oxygen consumption was achieved with azide, cyanide, and hydrogen sulfide. Pyrophosphate had no influence at all. There is some probability that cytochrome oxidase is the chief oxidase present. 6. The strongest inhibitory influence due to dehydrogenase inhibitors was observed with propyl carbamate and high concentrations of ethyl carbamate. 7. A small fraction of the oxygen consumption, about 10 per cent, may be due to substances with sulfhydryl groups, as indicated by a slight but distinct inhibition due to dilute iodoacetate and to arsenite. PMID:19873484

  1. Trypanosoma cruzi contains two galactokinases; molecular and biochemical characterization.

    PubMed

    Lobo-Rojas, Ángel E; González-Marcano, Eglys B; Valera-Vera, Edward A; Acosta, Héctor R; Quiñones, Wilfredo A; Burchmore, Richard J S; Concepción, Juan L; Cáceres, Ana J

    2016-10-01

    Two different putative galactokinase genes, found in the genome database of Trypanosoma cruzi were cloned and sequenced. Expression of the genes in Escherichia coli resulted for TcGALK-1 in the synthesis of a soluble and active enzyme, and in the case of TcGALK-2 gene a less soluble protein, with predicted molecular masses of 51.9kDa and 51.3kDa, respectively. The Km values determined for the recombinant proteins were for galactose 0.108mM (TcGALK-1) and 0.091mM (TcGALK-2) and for ATP 0.36mM (TcGALK-1) and 0.1mM (TcGALK-2). Substrate inhibition by ATP (Ki 0.414mM) was only observed for TcGALK-2. Gel-filtration chromatography showed that natural TcGALKs and recombinant TcGALK-1 are monomeric. In agreement with the possession of a type-1 peroxisome-targeting signal by both TcGALKs, they were found to be present inside glycosomes using two different methods of subcellular fractionation in conjunction with mass spectrometry. Both genes are expressed in epimastigote and trypomastigote stages since the respective proteins were immunodetected by western blotting. The T. cruzi galactokinases present their highest (52-47%) sequence identity with their counterpart from Leishmania spp., followed by prokaryotic galactokinases such as those from E. coli and Lactococcus lactis (26-23%). In a phylogenetic analysis, the trypanosomatid galactokinases form a separate cluster, showing an affiliation with bacteria. Epimastigotes of T. cruzi can grow in glucose-depleted LIT-medium supplemented with 20mM of galactose, suggesting that this hexose, upon phosphorylation by a TcGALK, could be used in the synthesis of UDP-galactose and also as a possible carbon and energy source. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  2. Biology of Trypanosoma (Trypanozoon) evansi in experimental heterologous mammalian hosts.

    PubMed

    Misra, K K; Roy, S; Choudhury, A

    2016-09-01

    Trypanosoma (Trypanozoon) evansi is a causative agent of the dreadful mammalian disease trypanosomiasis or 'Surra' and carried as a latent parasite in domestic cattle but occasionally proves fatal when transmitted to horses and camel. Sporadic outbreak of 'Surra' to different animals (beside their natural hosts) reminds that T. evansi may be zoonotic, as their close relative cause sleeping sickness to human being. This haemoflagellate is mechanically transmitted by horse fly and its effect on different host varies depending on certain factors including the effectiveness of transmission by mechanical vector, the suitability and susceptibility of the host as well as most importantly the ability of the disease establishment of parasite to adapt itself to the host's resistance, etc. The course of the disease caused by T. evansi is similar to that of human sleeping sickness caused by T. (T.) brucei gambiense. The target organs and symptoms show close similarity. T. evansi can successfully be transmitted among unnatural hosts i.e., other classes of vertebrates, like chicken. In transmission experiments, the unnatural hosts may sometimes induce profound changes in the biology of trypanosomes. Hence, in present study the observations are the biology of different morphological changes of T. evansi as well as its ability of disease formation within some heterologous mammal viz., albino rat, guineapig, bandicoot, mongoose, domestic cat and common monkey. Blood smears of infected albino rats, bandicoot, and mongoose revealed only monomorphic form. Interestingly, blood smears of infected cat and monkey, T. evansi shows slender trypomastigote form and short intermediate form whereas organ smears shows other two forms of haemoflagellate viz., sphaeromastigote and amastigote form. The haemoflagellate maintains a common reproductive cycle in all the experimental heterologous hosts whereas disease symptoms differ. T. evansi infected cat and monkey shows nervous symptoms. Infected

  3. Seroprevalence of Trypanosoma cruzi in raccoons from Tennessee.

    PubMed

    Maloney, Jenny; Newsome, Anthony; Huang, Junjun; Kirby, Jordona; Kranz, Melissa; Wateska, Angela; Dunlap, Brett; Yabsley, Michael J; Dunn, John R; Jones, Timothy F; Moncayo, Abelardo C

    2010-04-01

    Trypanosoma cruzi is the etiologic agent of Chagas' disease. Autochthonous human and canine transmission of T. cruzi has been documented in Tennessee, but little is known about its ecology, including the prevalence of T. cruzi among wildlife in Tennessee. Serum samples from 706 raccoons (Procyon lotor) from 10 counties in the Ridge and Valley and Blue Ridge Mountains ecoregions of eastern Tennessee were tested for antibodies reactive with T. cruzi using the indirect fluorescent antibody assay. Two hundred six (29.2%) samples were seropositive, with 9 counties yielding positive samples (range 14.6-63.6%). Significantly more raccoons from rural habitats (35.1%) were found positive for T. cruzi exposure than were those from suburban habitats (23.1%, P < 0.001). Land cover class was not associated with seropositivity status (P = 0.441), even though deciduous forest was the most common site from where raccoons were trapped and the most common site of positive raccoons in rural areas (42%). Interestingly, age was positively associated with seropositivity. Raccoons older than 1 yr (adults) were 40.1% seropositive compared to 12.2% of those less than 1 yr (juveniles; P < 0.001). Female adults were significantly more likely to be exposed to T. cruzi than were male adult raccoons (P < 0.001). No significant seroprevalence difference was seen among male and female juveniles. This study contributes to understanding the dynamics of T. cruzi exposure within raccoon populations in Tennessee. The importance of habitat (rural vs. suburban) and microhabitat (dens) in risk of exposure to these populations is also discussed.

  4. KINETOPLAST DEOXYRIBONUCLEIC ACID OF THE HEMOFLAGELLATE TRYPANOSOMA LEWISI

    PubMed Central

    Renger, Hartmut C.; Wolstenholme, David R.

    1970-01-01

    Cesium chloride centrifugation of DNA extracted from cells of blood strain Trypanosoma lewisi revealed a main band, ρ = 1.707, a light satellite, ρ = 1.699, and a heavy satellite, ρ = 1.721. Culture strain T. lewisi DNA comprised only a main band, ρ = 1.711, and a light satellite, ρ = 1.699. DNA isolated from DNase-treated kinetoplast fractions of both the blood and culture strains consisted of only the light satellite DNA. Electron microscope examination of rotary shadowed preparations of lysates revealed that DNA from kinetoplast fractions was mainly in the form of single 0.4 µ circular molecules and large masses of 0.4 µ interlocked circles with which longer, often noncircular molecules were associated. The 0.4 µ circular molecules were mainly in the covalently closed form: they showed a high degree of resistance to thermal denaturation which was lost following sonication; and they banded at a greater density than linear DNA in cesium chloride-ethidium bromide gradients. Interpretation of the large masses of DNA as comprising interlocked covalently closed 0.4 µ circles was supported by the findings that they banded with single circular molecules in cesium chloride-ethidium bromide gradients, and following breakage of some circles by mild sonication, they disappeared and were replaced by molecules made up of low numbers of apparently interlocked 0.4 µ circles. When culture strain cells were grown in the presence of either ethidium bromide or acriflavin, there was a loss of stainable kinetoplast DNA in cytological preparations. There was a parallel loss of light satellite and of circular molecules from DNA extracted from these cells. PMID:5497546

  5. Trypanosoma cruzi as an effective cancer antigen delivery vector

    PubMed Central

    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.

    2011-01-01

    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. PMID:22114198

  6. Aspirin treatment exacerbates oral infections by Trypanosoma cruzi.

    PubMed

    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. Copyright © 2016 Elsevier Inc. All rights reserved.

  7. Trypanosoma cruzi maxicircle heterogeneity in Chagas disease patients from Brazil.

    PubMed

    Carranza, Julio César; Valadares, Helder M S; D'Avila, Daniella A; Baptista, Rodrigo P; Moreno, Margoth; Galvão, Lúcia M C; Chiari, Egler; Sturm, Nancy R; Gontijo, Eliane D; Macedo, Andrea M; Zingales, Bianca

    2009-07-15

    The majority of individuals in the chronic phase of Chagas disease are asymptomatic (indeterminate form, IF). Each year, approximately 3% of them develop lesions in the heart or gastrointestinal tract. Cardiomyopathy (CCHD) is the most severe manifestation of Chagas disease. The factors that determine the outcome of the infection are unknown, but certainly depend on complex interactions amongst the genetic make-up of the parasite, the host immunogenetic background and environment. In a previous study we verified that the maxicircle gene NADH dehydrogenase (mitochondrial complex I) subunit 7 (ND7) from IF isolates had a 455 bp deletion compared with the wild type (WT) ND7 gene from CCHD strains. We proposed that ND7 could constitute a valuable target for PCR assays in the differential diagnosis of the infective strain. In the present study we evaluated this hypothesis by examination of ND7 structure in parasites from 75 patients with defined pathologies, from Southeast Brazil. We also analysed the structure of additional mitochondrial genes (ND4/CR4, COIII and COII) since the maxicircle is used for clustering Trypanosoma cruzi strains into three clades/haplogroups. We conclude that maxicircle genes do not discriminate parasite populations which induce IF or CCHD forms. Interestingly, the great majority of the analysed isolates belong to T. cruzi II (discrete typing unit, (DTU) IIb) genotype. This scenario is at variance with the prevalence of hybrid (DTU IId) human isolates in Bolivia, Chile and Argentina. The distribution of WT and deleted ND7 and ND4 genes in T. cruzi strains suggests that mutations in the two genes occurred in different ancestrals in the T. cruzi II cluster, allowing the identification of at least three mitochondrial sub-lineages within this group. The observation that T. cruzi strains accumulate mutations in several genes coding for complex I subunits favours the hypothesis that complex I may have a limited activity in this parasite.

  8. Antiparasitic Effect of Vitamin B12 on Trypanosoma cruzi

    PubMed Central

    Ciccarelli, Alejandra B.; Frank, Fernanda M.; Puente, Vanesa; Malchiodi, Emilio L.

    2012-01-01

    A nutritional characteristic of trypanosomatid protozoa is that they need a heme compound as a growth factor. Because of the cytotoxic activity of heme and its structural similarity to cobalamins, we have investigated the in vitro and in vivo effect of vitamin B12 (or cyanocobalamin) on the different forms of Trypanosoma cruzi. Cyanocobalamin showed a marked antiparasitic activity against epimastigotes (50% inhibitory concentration [IC50], 2.42 μM), amastigotes (IC50, 10.69 μM), and trypomastigotes (IC50, 9.46 μM). Anti-epimastigote and -trypomastigote values were 1.7 to 4 times lower than those obtained with the reference drug benznidazole (Bnz). We also found that B12 and hemin do not interact with each other in their modes of action. Our results show that B12 increases intracellular oxidative activity and stimulates both superoxide dismutase (50%) and ascorbate peroxidase (20%) activities, while the activity of trypanothione reductase was not modified. In addition, we found that the antioxidants dithiothreitol and ascorbic acid increase the susceptibility of the parasite to the cytotoxic action of B12. We propose that vitamin B12 exerts its growth-inhibitory effect through the generation of reactive oxygen species. In an in vivo assay, a significant reduction in the number of circulating parasites was found in T. cruzi-infected mice treated with cyanocobalamin and ascorbic acid. The reduction of parasitemia in benznidazole-treated mice was improved by the addition of these vitamins. According to our results, a combination of B12 and Bnz should be further investigated due to its potential as a new therapeutic modality for the treatment of Chagas' disease. PMID:22869565

  9. Anti-Trypanosoma brucei activity of nonprimate zoo sera.

    PubMed

    Black, S J; Wang, Q; Makadzange, T; Li, Y L; Van Praagh, A; Loomis, M; Seed, J R

    1999-02-01

    Constitutive anti-Trypanosoma brucei subsp. brucei S 427 clone 1 and 22 activities were evaluated in sera from 22 species of nonprimate mammals. The sera fell into 5 categories. Sera from Cape buffalo, giraffe, and greater kudu showed a concentration-dependent inhibition of replication of the 2 clones of organisms, which was dependent on the presence of xanthine oxidase. Sera from warthog and springbok also severely limited trypanosome replication but lacked xanthine oxidase. Their antitrypanosome activity was inactivated by heating at 56 C for 30 min but not affected by absorbing with trypanosomes at 4 C. Sera from lion and leopard showed a concentration-dependent inhibition of the growth of T. brucei S427 clone 1 organisms, but not clone 22 organisms. These sera lacked xanthine oxidase. Their anti-T. brucei S 427 clone 1 activity was inactivated by heating at 56 C for 30 min but not removed by absorbing with trypanosomes. Serum from Grant's gazelle prevented replication of both T. brucei clones, lacked xanthine oxidase, and was not affected by heating at 56 C. Sera from waterbuck, Thompson's gazelle, sitatunga, Cape hartebeeste, gerenuk, Grant's zebra, cow, several cat, cougar, bobcat, and domestic cat were fully supportive of trypanosome replication irrespective of concentration tested up to a maximum of 48% v/v in culture medium. Sera from different individuals of the same mammal species had similar effects on trypanosomes, and samples collected from the same individual at different times also had similar activities indicating species-specific stable expression, or lack thereof, of constitutive serum antitrypanosome components.

  10. Ca2+ Regulation of Trypanosoma brucei Phosphoinositide Phospholipase C.

    PubMed

    King-Keller, Sharon; Moore, Christina A; Docampo, Roberto; Moreno, Silvia N J

    2015-05-01

    We characterized a phosphoinositide phospholipase C (PI-PLC) from the procyclic form (PCF) of Trypanosoma brucei. The protein contains a domain organization characteristic of typical PI-PLCs, such as X and Y catalytic domains, an EF-hand calcium-binding motif, and a C2 domain, but it lacks a pleckstrin homology (PH) domain. In addition, the T. brucei PI-PLC (TbPI-PLC) contains an N-terminal myristoylation consensus sequence found only in trypanosomatid PI-PLCs. A peptide containing this N-terminal domain fused to green fluorescent protein (GFP) was targeted to the plasma membrane. TbPI-PLC enzymatic activity was stimulated by Ca(2+) concentrations below the cytosolic levels in the parasite, suggesting that the enzyme is constitutively active. TbPI-PLC hydrolyzes both phosphatidylinositol (PI) and phosphatidylinositol 4,5-bisphosphate (PIP2), with a higher affinity for PIP2. We found that modification of a single amino acid in the EF-hand motif greatly affected the protein's Ca(2+) sensitivity and substrate preference, demonstrating the role of this motif in Ca(2+) regulation of TbPI-PLC. Endogenous TbPI-PLC localizes to intracellular vesicles and might be using an intracellular source of PIP2. Knockdown of TbPI-PLC expression by RNA interference (RNAi) did not result in growth inhibition, although enzymatic activity was still present in parasites, resulting in hydrolysis of PIP2 and a contribution to the inositol 1,4,5-trisphosphate (IP3)/diacylglycerol (DAG) pathway. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  11. Succinate-dependent metabolism in Trypanosoma cruzi epimastigotes.

    PubMed

    Denicola-Seoane, A; Rubbo, H; Prodanov, E; Turrens, J F

    1992-08-01

    Trypanosoma cruzi epimastigotes permeabilized with digitonin (65 micrograms (mg protein)-1) to measure mitochondrial respiration were exposed to different substrates. Although none of the NADH-dependent substrates stimulated respiration, succinate supported not only oxygen consumption but also oxidative phosphorylation (respiratory control ratio of 1.9 +/- 0.3) indicating that the mitochondria were coupled. The rate of NADH-dependent oxygen consumption by membrane fractions (9.4 +/- 0.7 nmol min-1 (mg protein)-1) was reduced by 50% upon addition of catalase indicating that the electrons from NADH oxidation reduced oxygen to H2O2. NADH-dependent H2O2 production (16 +/- 1 nmol min-1 (mg protein)-1) was confirmed using cytochrome c peroxidase. This activity was inhibited by fumarate by 70%, suggesting a competition between fumarate and oxygen for the electrons from NADH, probably at the fumarate reductase level. The respiratory chain inhibitor antimycin blocked both respiration by intact cells and succinate-dependent cytochrome c by isolated membranes. No inhibition by antimycin was observed when NADH replaced succinate as an electron donor, indicating that the electrons from NADH oxidation reduced cytochrome c through a different route. Malonate blocked not only succinate-cytochrome c reductase and fumarate reductase, but also intact cell motility. These results suggest that succinate has a central role in the intermediate metabolism of i. cruzi, as it may be used for respiration or excreted to the extracellular space under anaerobic conditions. In addition, 2 potential sources of H2O2 were tentatively identified as: (a) the enzyme fumarate reductase; and (b) a succinate-dependent site, which may be the semiquinone form of Coenzyme Q9, as in mammalian mitochondria.

  12. Quantitative Proteomic and Phosphoproteomic Analysis of Trypanosoma cruzi Amastigogenesis*

    PubMed Central

    Queiroz, Rayner M. L.; Charneau, Sébastien; Mandacaru, Samuel C.; Schwämmle, Veit; Lima, Beatriz D.; Roepstorff, Peter; Ricart, Carlos A. O.

    2014-01-01

    Chagas disease is a tropical neglected disease endemic in Latin America caused by the protozoan Trypanosoma cruzi. The parasite has four major life stages: epimastigote, metacyclic trypomastigote, bloodstream trypomastigote, and amastigote. The differentiation from infective trypomastigotes into replicative amastigotes, called amastigogenesis, takes place in vivo inside mammalian host cells after a period of incubation in an acidic phagolysosome. This differentiation process can be mimicked in vitro by incubating tissue-culture-derived trypomastigotes in acidic DMEM. Here we used this well-established differentiation protocol to perform a comprehensive quantitative proteomic and phosphoproteomic analysis of T. cruzi amastigogenesis. Samples from fully differentiated forms and two biologically relevant intermediate time points were Lys-C/trypsin digested, iTRAQ-labeled, and multiplexed. Subsequently, phosphopeptides were enriched using a TiO2 matrix. Non-phosphorylated peptides were fractionated via hydrophilic interaction liquid chromatography prior to LC-MS/MS analysis. LC-MS/MS and bioinformatics procedures were used for protein and phosphopeptide quantitation, identification, and phosphorylation site assignment. We were able to identify regulated proteins and pathways involved in coordinating amastigogenesis. We also observed that a significant proportion of the regulated proteins were membrane proteins. Modulated phosphorylation events coordinated by protein kinases and phosphatases that are part of the signaling cascade induced by incubation in acidic medium were also evinced. To our knowledge, this work is the most comprehensive quantitative proteomics study of T. cruzi amastigogenesis, and these data will serve as a trustworthy basis for future studies, and possibly for new potential drug targets. PMID:25225356

  13. Role of iron in Trypanosoma cruzi infection of mice.

    PubMed Central

    Lalonde, R G; Holbein, B E

    1984-01-01

    The role of iron in experimental infection of mice with Trypanosoma cruzi was investigated. B6 mice had a transient parasitemia and a transient anemia, both of maximal intensity 28 d after the inoculation of T. cruzi. There was a biphasic hypoferremic host response to infection with T. cruzi with the peak hypoferremia also occurring 28 d after inoculation of the parasite. The mortality rate from infection was increased from 23% in phosphate-buffered saline-treated B6 mice to 50% in a group of B6 mice receiving iron-dextran (P less than or equal to 0.025), whereas depletion of iron stores with the iron chelator desferrioxamine B and an iron-deficient diet provided complete protection of B6 mice (P less than or equal to 0.05). The mortality rate in the highly susceptible C3H strain was reduced from 100% in the control group to 45% (P less than or equal to 0.025) in the iron-depleted group. The tissue iron stores were altered in mice receiving either iron-dextran or desferrioxamine B and an iron-deficient diet. In vitro, T. cruzi was shown to require both a heme and a nonheme iron source for an optimal growth rate. The effects of iron excess or depletion on the outcome of infection with T. cruzi correlated both with the growth requirements of the parasite for iron and with the availability of intracellular iron. Thus, it was suggested that the hypoferremic response, by sequestering iron within intracellular stores, potentially enhanced the pathogenicity of the intracellular parasites. Furthermore, the in vivo effects of iron excess and depletion correlated with an effect of iron on the growth rate and pathogenicity of the parasite. PMID:6421877

  14. Ancestral Genomes, Sex, and the Population Structure of Trypanosoma cruzi

    PubMed Central

    Bastos-Rodrigues, Luciana; Gonçalves, Vanessa F; Teixeira, Santuza M. R; Chiari, Egler; Junqueira, Ângela C. V; Fernandes, Octavio; Macedo, Andréa M; Machado, Carlos Renato; Pena, Sérgio D. J

    2006-01-01

    Acquisition of detailed knowledge of the structure and evolution of Trypanosoma cruzi populations is essential for control of Chagas disease. We profiled 75 strains of the parasite with five nuclear microsatellite loci, 24Sα RNA genes, and sequence polymorphisms in the mitochondrial cytochrome oxidase subunit II gene. We also used sequences available in GenBank for the mitochondrial genes cytochrome B and NADH dehydrogenase subunit 1. A multidimensional scaling plot (MDS) based in microsatellite data divided the parasites into four clusters corresponding to T. cruzi I (MDS-cluster A), T. cruzi II (MDS-cluster C), a third group of T. cruzi strains (MDS-cluster B), and hybrid strains (MDS-cluster BH). The first two clusters matched respectively mitochondrial clades A and C, while the other two belonged to mitochondrial clade B. The 24Sα rDNA and microsatellite profiling data were combined into multilocus genotypes that were analyzed by the haplotype reconstruction program PHASE. We identified 141 haplotypes that were clearly distributed into three haplogroups (X, Y, and Z). All strains belonging to T. cruzi I (MDS-cluster A) were Z/Z, the T. cruzi II strains (MDS-cluster C) were Y/Y, and those belonging to MDS-cluster B (unclassified T. cruzi) had X/X haplogroup genotypes. The strains grouped in the MDS-cluster BH were X/Y, confirming their hybrid character. Based on these results we propose the following minimal scenario for T. cruzi evolution. In a distant past there were at a minimum three ancestral lineages that we may call, respectively, T. cruzi I, T. cruzi II, and T. cruzi III. At least two hybridization events involving T. cruzi II and T. cruzi III produced evolutionarily viable progeny. In both events, the mitochondrial recipient (as identified by the mitochondrial clade of the hybrid strains) was T. cruzi II and the mitochondrial donor was T. cruzi III. PMID:16609729

  15. Cynaropicrin targets the trypanothione redox system in Trypanosoma brucei.

    PubMed

    Zimmermann, Stefanie; Oufir, Mouhssin; Leroux, Alejandro; Krauth-Siegel, R Luise; Becker, Katja; Kaiser, Marcel; Brun, Reto; Hamburger, Matthias; Adams, Michael

    2013-11-15

    In mice cynaropicrin (CYN) potently inhibits the proliferation of Trypanosoma brucei-the causative agent of Human African Trypanosomiasis-by a so far unknown mechanism. We hypothesized that CYNs α,β-unsaturated methylene moieties act as Michael acceptors for glutathione (GSH) and trypanothione (T(SH)2), the main low molecular mass thiols essential for unique redox metabolism of these parasites. The analysis of this putative mechanism and the effects of CYN on enzymes of the T(SH)2 redox metabolism including trypanothione reductase, trypanothione synthetase, glutathione-S-transferase, and ornithine decarboxylase are shown. A two step extraction protocol with subsequent UPLC-MS/MS analysis was established to quantify intra-cellular CYN, T(SH)2, GSH, as well as GS-CYN and T(S-CYN)2 adducts in intact T. b. rhodesiense cells. Within minutes of exposure to CYN, the cellular GSH and T(SH)2 pools were entirely depleted, and the parasites entered an apoptotic stage and died. CYN also showed inhibition of the ornithine decarboxylase similar to the positive control eflornithine. Significant interactions with the other enzymes involved in the T(SH)2 redox metabolism were not observed. Alongside many other biological activities sesquiterpene lactones including CYN have shown antitrypanosomal effects, which have been postulated to be linked to formation of Michael adducts with cellular nucleophiles. Here the interaction of CYN with biological thiols in a cellular system in general, and with trypanosomal T(SH)2 redox metabolism in particular, thus offering a molecular explanation for the antitrypanosomal activity is demonstrated. At the same time, the study provides a novel extraction and analysis protocol for components of the trypanosomal thiol metabolism. Copyright © 2013 Elsevier Ltd. All rights reserved.

  16. Secretome analysis of Trypanosoma cruzi by proteomics studies.

    PubMed

    Brossas, Jean-Yves; Gulin, Julián Ernesto Nicolás; Bisio, Margarita Maria Catalina; Chapelle, Manuel; Marinach-Patrice, Carine; Bordessoules, Mallaury; Palazon Ruiz, George; Vion, Jeremy; Paris, Luc; Altcheh, Jaime; Mazier, Dominique

    2017-01-01

    Chagas disease is a debilitating often fatal disease resulting from infection by the protozoan parasite Trypanosoma cruzi. Chagas disease is endemic in 21 countries of the Americas, and it is an emerging disease in other countries as a result of migration. Given the chronic nature of the infection where intracellular parasites persist for years, the diagnosis of T. cruzi by direct detection is difficult, whereas serologic tests though sensitive may yield false-positive results. The development of new rapid test based on the identification of soluble parasitic antigens in serum would be a real innovation in the diagnosis of Chagas disease. To identify new soluble biomarkers that may improve diagnostic tests, we investigated the proteins secreted by T. cruzi using mass spectrometric analyses of conditioned culture media devoid of serum collected during the emergence of trypomastigotes from infected Vero cells. In addition, we compared the secretomes of two T. cruzi strains from DTU Tc VI (VD and CL Brener). Analysis of the secretome collected during the emergence of trypomastigotes from Vero cells led to the identification of 591 T. cruzi proteins. Three hundred sixty three proteins are common to both strains and most belong to different multigenic super families (i.e. TcS, GP63, MASP, and DGF1). Ultimately we have established a list of 94 secreted proteins, common to both DTU Tc VI strains that do not belong to members of multigene families. This study provides the first comparative analysis of the secretomes from two distinct T. cruzi strains of DTU TcVI. This led us to identify a subset of common secreted proteins that could potentially serve as serum markers for T. cruzi infection. Their potential could now be evaluated, with specific antibodies using sera collected from patients and residents from endemic regions.

  17. Role of placental barrier integrity in infection by Trypanosoma cruzi.

    PubMed

    Díaz-Luján, C; Triquell, M F; Castillo, C; Hardisson, D; Kemmerling, U; Fretes, R E

    2016-12-01

    American trypanosomiasis has long been a neglected disease endemic in LatinAmerica, but congenital transmission has now spread Chagas disease to cause a global health problem. As the early stages of the infection of placental tissue and the vertical transmission by Trypanosoma cruzi are still not well understood, it is important to investigate the relevance of the first structure of the placental barrier in chorionic villi infection by T. cruzi during the initial stage of the infection. Explants of human chorionic villi from healthy pregnant women at term were denuded of their syncytiotrophoblast and co-cultured for 3h, 24h and 96h with 800,000 trypomastigotes (simulating acute infection). T. cruzi infected cells were identified by immunohistochemistry for cytokeratin-7 (+cytotrophoblast) and CD68 (+macrophages), and the infection was quantified. In placental tissue, the parasite load was analyzed by qPCR and microscopy, and the motile trypomastigotes were quantified in culture supernatant. In denuded chorionic villous, the total area occupied by the parasite (451.23μm(2), 1.33%) and parasite load (RQ: 87) was significantly higher (p<0.05) than in the entire villous (control) (5.98μm(2), 0.016%) (RQ:1) and with smaller concentration of nitric oxide. Stromal non-macrophage cells were infected as well as cytotrophoblasts and some macrophages, but with significant differences being observed. The parasite quantity in the culture supernatant was significantly higher (p<0.05) in denuded culture explants from 96h of culture. Although the human complete chorionic villi limited the infection, the detachment of the first structure of the placenta barrier (syncytiotrophoblast) increased both the infection of the villous stroma and the living trypomastigotes in the culture supernatant. Therefore structural and functional alterations to chorionic villi placental barrier reduce placental defenses and may contribute to the vertical transmission of Chagas.

  18. Targeted Screening Strategies to Detect Trypanosoma cruzi Infection in Children

    PubMed Central

    Levy, Michael Z.; Kawai, Vivian; Bowman, Natalie M.; Waller, Lance A.; Cabrera, Lilia; Pinedo-Cancino, Viviana V.; Seitz, Amy E.; Steurer, Frank J.; Cornejo del Carpio, Juan G.; Cordova-Benzaquen, Eleazar; Maguire, James H.; Gilman, Robert H.; Bern, Caryn

    2007-01-01

    Background Millions of people are infected with Trypanosoma cruzi, the causative agent of Chagas disease in Latin America. Anti-trypanosomal drug therapy can cure infected individuals, but treatment efficacy is highest early in infection. Vector control campaigns disrupt transmission of T. cruzi, but without timely diagnosis, children infected prior to vector control often miss the window of opportunity for effective chemotherapy. Methods and Findings We performed a serological survey in children 2–18 years old living in a peri-urban community of Arequipa, Peru, and linked the results to entomologic, spatial and census data gathered during a vector control campaign. 23 of 433 (5.3% [95% CI 3.4–7.9]) children were confirmed seropositive for T. cruzi infection by two methods. Spatial analysis revealed that households with infected children were very tightly clustered within looser clusters of households with parasite-infected vectors. Bayesian hierarchical mixed models, which controlled for clustering of infection, showed that a child's risk of being seropositive increased by 20% per year of age and 4% per vector captured within the child's house. Receiver operator characteristic (ROC) plots of best-fit models suggest that more than 83% of infected children could be identified while testing only 22% of eligible children. Conclusions We found evidence of spatially-focal vector-borne T. cruzi transmission in peri-urban Arequipa. Ongoing vector control campaigns, in addition to preventing further parasite transmission, facilitate the collection of data essential to identifying children at high risk of T. cruzi infection. Targeted screening strategies could make integration of diagnosis and treatment of children into Chagas disease control programs feasible in lower-resource settings. PMID:18160979

  19. A bioinformatic analysis of the RAB genes of Trypanosoma brucei.

    PubMed

    Ackers, John P; Dhir, Vivek; Field, Mark C

    2005-05-01

    RAB proteins are small GTPases with vital roles in eukaryotic intracellular transport; orthologous RABs appear to fulfil similar functions in diverse organisms. Trypanosoma brucei spp., the causative organisms of Old World trypanosomiasis of humans and domestic animals, have extremely effective endocytic and exocytic mechanisms that are likely to be involved in maintenance of infection, making study of these systems of importance. Taking advantage of the essential completion of the T. brucei genome, we have re-examined the T. brucei RABs (TbRABs) so far described and identified a total of 16. BLAST searches and phylogenetic analysis show that nine of the TbRABs can confidently be assigned as orthologues or homologues of known RAB proteins from higher eukaryotes, and four more with reasonable probability. The core endocytic pathway is probably similar in complexity to yeast, whilst the early exocytic pathway appears to be more complex than in yeast. Two of the TbRAB family (RAB23 and 28) with clear mammalian orthologues appear to be unusual, and may be involved in nuclear processes and are described in more detail in an accompanying paper. Three TbRABs appear, however, to have no close homologues and may fulfil specialised functions in this organism. The availability of a complete set of TbRABs--which includes orthologues of the RABs responsible for control of the core of the endomembrane system (i.e. RAB1, 2, 4-7 and 11)--provides a first overview of the trafficking complexity that is present within a kinetoplastid parasite. Based on these homologies we suggest a systematic nomenclature for the TbRABs to reflect their functional homologies. This information is of importance both from the perspective of understanding the evolution and diversity of eukaryotic trafficking, but also in providing a framework by which to understand protein processing, trafficking, endocytosis and other related processes in these parasites.

  20. Proteomic Analysis of Trypanosoma cruzi Response to Ionizing Radiation Stress

    PubMed Central

    Vieira, Helaine Graziele Santos; Grynberg, Priscila; Bitar, Mainá; Pires, Simone da Fonseca; Hilário, Heron Oliveira; Macedo, Andrea Mara; Machado, Carlos Renato; de Andrade, Hélida Monteiro; Franco, Glória Regina

    2014-01-01

    Trypanosoma cruzi, the causative agent of Chagas disease, is extremely resistant to ionizing radiation, enduring up to 1.5 kGy of gamma rays. Ionizing radiation can damage the DNA molecule both directly, resulting in double-strand breaks, and indirectly, as a consequence of reactive oxygen species production. After a dose of 500 Gy of gamma rays, the parasite genome is fragmented, but the chromosomal bands are restored within 48 hours. Under such conditions, cell growth arrests for up to 120 hours and the parasites resume normal growth after this period. To better understand the parasite response to ionizing radiation, we analyzed the proteome of irradiated (4, 24, and 96 hours after irradiation) and non-irradiated T. cruzi using two-dimensional differential gel electrophoresis followed by mass spectrometry for protein identification. A total of 543 spots were found to be differentially expressed, from which 215 were identified. These identified protein spots represent different isoforms of only 53 proteins. We observed a tendency for overexpression of proteins with molecular weights below predicted, indicating that these may be processed, yielding shorter polypeptides. The presence of shorter protein isoforms after irradiation suggests the occurrence of post-translational modifications and/or processing in response to gamma radiation stress. Our results also indicate that active translation is essential for the recovery of parasites from ionizing radiation damage. This study therefore reveals the peculiar response of T. cruzi to ionizing radiation, raising questions about how this organism can change its protein expression to survive such a harmful stress. PMID:24842666

  1. Trypanosoma cruzi as an effective cancer antigen delivery vector.

    PubMed

    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.

  2. Biochemical Diversity in the Trypanosoma congolense Trans-sialidase Family

    PubMed Central

    Gbem, Thaddeus T.; Waespy, Mario; Hesse, Bettina; Dietz, Frank; Smith, Joel; Chechet, Gloria D.; Nok, Jonathan A.; Kelm, Sørge

    2013-01-01

    Trans-sialidases are key enzymes in the life cycle of African trypanosomes in both, mammalian host and insect vector and have been associated with the disease trypanosomiasis, namely sleeping sickness and nagana. Besides the previously reported TconTS1, we have identified three additional active trans-sialidases, TconTS2, TconTS3 and TconTS4, and three trans-sialidase like genes in Trypanosoma congolense. At least TconTS1, TconTS2 and TconTS4 are found in the bloodstream of infected animals. We have characterised the enzymatic properties of recombinant proteins expressed in eukaryotic fibroblasts using fetuin as model blood glycoprotein donor substrate. One of the recombinant trans-sialidases, TconTS2, had the highest specific activity reported thus far with very low sialidase activity. The active trans-sialidases share all the amino acids critical for the catalytic reaction with few variations in the predicted binding site for the leaving or acceptor glycan. However, these differences cannot explain the orders of magnitudes between their transfer activities, which must be due to other unidentified structural features of the proteins or substrates selectivity. Interestingly, the phylogenetic relationships between the lectin domains correlate with their specific trans-sialylation activities. This raises the question whether and how the lectin domains regulate the trans-sialidase reaction. The identification and enzymatic characterisation of the trans-sialidase family in T. congolense will contribute significantly towards the understanding of the roles of these enzymes in the pathogenesis of Animal African Trypanosomiasis. PMID:24340108

  3. New Trypanosoma evansi Type B Isolates from Ethiopian Dromedary Camels.

    PubMed

    Birhanu, Hadush; Gebrehiwot, Tadesse; Goddeeris, Bruno Maria; Büscher, Philippe; Van Reet, Nick

    2016-04-01

    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.

  4. Structural features affecting variant surface glycoprotein expression in Trypanosoma brucei.

    PubMed

    Wang, Jun; Böhme, Ulrike; Cross, George A M

    2003-05-01

    The glycosylphosphatidylinositol (GPI)-anchored variant surface glycoprotein (VSG) of Trypanosoma brucei is the most abundant GPI-anchored protein expressed on any cell, and is an essential virulence factor. To determine what structural features affect efficient expression of VSG, we made a series of mutations in two VSGs. Inserting 18 amino acids, between the amino- and carboxy-terminal domains, reduced the expression of VSG 221 to about 3% of the wild-type level. When this insertion was combined with deletion of the single carboxy-terminal subdomain, expression was reduced a further three-fold. In VSG 117, which contains two carboxy-terminal subdomains, point mutation of the intervening N-glycosylation site reduced expression about 15-fold. Deleting the most carboxy-terminal subdomain and intervening region, including the N-glycosylation site, reduced expression to 15-20% of wild type VSG, and deletion of both subdomains reduced expression to <1%. Despite their low abundance, all VSG mutants were GPI anchored on the cell surface. Our results suggest that, for a protein to be efficiently displayed on the surface of bloodstream-form T. brucei, it is essential that it contains the conserved structural motifs of a T. brucei VSG. Serum resistance-associated protein (SRA), which confers human infectivity on T. brucei, strongly resembles a VSG deletion mutant. Expression of three epitope-tagged versions of SRA in T. brucei conferred total resistance to human serum. SRA possesses a canonical GPI signal sequence, but we were unable to obtain unequivocal evidence for the presence of a GPI anchor. SRA was not released during osmotic lysis, indicating that it is not GPI anchored on the cell surface.

  5. Growth hormones therapy in immune response against Trypanosoma cruzi.

    PubMed

    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.

  6. Sialic Acid Glycobiology Unveils Trypanosoma cruzi Trypomastigote Membrane Physiology

    PubMed Central

    Lantos, Andrés B.; Carlevaro, Giannina; Araoz, Beatriz; Ruiz Diaz, Pablo; Camara, María de los Milagros; Buscaglia, Carlos A.; Bossi, Mariano; Yu, Hai; Chen, Xi; Bertozzi, Carolyn R.; Mucci, Juan; Campetella, Oscar

    2016-01-01

    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(1,3)Galactosyl 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 soluble form. PMID

  7. Induction of Resistance to Azole Drugs in Trypanosoma cruzi

    PubMed Central

    Buckner, Frederick S.; Wilson, Aaron J.; White, Theodore C.; Van Voorhis, Wesley C.

    1998-01-01

    Trypanosoma cruzi is the protozoan parasite that causes Chagas’ disease, a frequently fatal illness affecting the heart and gastrointestinal systems. An estimated 16 million to 18 million people in Latin America and 50,000 to 100,000 people in the United States are infected with this pathogen. Treatment options for T. cruzi infections are suboptimal due to the toxicities and limited effectiveness of the available drugs. Azole antimicrobial agents have been discovered to have antitrypanosomal activity by inhibition of ergosterol synthesis. The triazole itraconazole was recently shown to produce a parasitologic cure rate of 53% in chronically infected patients (W. Apt et al., Am. J. Trop. Med. Hyg. 59:133–138, 1998), a result which may lead to more use of this family of drugs for the treatment of T. cruzi infections. In the experiments reported on here, resistance to azoles was induced in vitro by serial passage of mammalian-stage parasites in the presence of fluconazole for 4 months. These parasites were cross resistant to the other azoles, ketoconazole, miconazole, and itraconazole. They remained susceptible to benznidazole and amphotericin B. The azole-resistant phenotype was stable for more than 2 months of in vitro serial passage without fluconazole. In addition, the parasites resisted treatment in mice receiving ketoconazole. The rapid development of azole resistance in T. cruzi in vitro suggests that resistance to azole drugs has the potential to occur in patients and may pose an impediment to the progress being made in the treatment of T. cruzi infection. PMID:9835521

  8. Regulation of the cell division cycle in Trypanosoma brucei.

    PubMed

    Li, Ziyin

    2012-10-01

    The cell division cycle is tightly regulated by the activation and inactivation of a series of proteins that control the replication and segregation of organelles to the daughter cells. During the past decade, we have witnessed significant advances in our understanding of the cell cycle in Trypanosoma brucei and how the cycle is regulated by various regulatory proteins. However, many other regulators, especially those unique to trypanosomes, remain to be identified, and we are just beginning to delineate the signaling pathways that drive the transitions through different cell cycle stages, such as the G(1)/S transition, G(2)/M transition, and mitosis-cytokinesis transition. Trypanosomes appear to employ both evolutionarily conserved and trypanosome-specific molecules to regulate the various stages of its cell cycle, including DNA replication initiation, spindle assembly, chromosome segregation, and cytokinesis initiation and completion. Strikingly, trypanosomes lack some crucial regulators that are well conserved across evolution, such as Cdc6 and Cdt1, which are involved in DNA replication licensing, the spindle motor kinesin-5, which is required for spindle assembly, the central spindlin complex, which has been implicated in cytokinesis initiation, and the actomyosin contractile ring, which is located at the cleavage furrow. Conversely, trypanosomes possess certain regulators, such as cyclins, cyclin-dependent kinases, and mitotic centromere-associated kinesins, that are greatly expanded and likely play diverse cellular functions. Overall, trypanosomes apparently have integrated unique regulators into the evolutionarily conserved pathways to compensate for the absence of those conserved molecules and, additionally, have evolved certain cell cycle regulatory pathways that are either different from its human host or distinct between its own life cycle forms.

  9. Trypanosoma cruzi population dynamics in the Central Ecuadorian Coast.

    PubMed

    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. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights

  10. Biochemical diversity in the Trypanosoma congolense trans-sialidase family.

    PubMed

    Gbem, Thaddeus T; Waespy, Mario; Hesse, Bettina; Dietz, Frank; Smith, Joel; Chechet, Gloria D; Nok, Jonathan A; Kelm, Sørge

    2013-01-01

    Trans-sialidases are key enzymes in the life cycle of African trypanosomes in both, mammalian host and insect vector and have been associated with the disease trypanosomiasis, namely sleeping sickness and nagana. Besides the previously reported TconTS1, we have identified three additional active trans-sialidases, TconTS2, TconTS3 and TconTS4, and three trans-sialidase like genes in Trypanosoma congolense. At least TconTS1, TconTS2 and TconTS4 are found in the bloodstream of infected animals. We have characterised the enzymatic properties of recombinant proteins expressed in eukaryotic fibroblasts using fetuin as model blood glycoprotein donor substrate. One of the recombinant trans-sialidases, TconTS2, had the highest specific activity reported thus far with very low sialidase activity. The active trans-sialidases share all the amino acids critical for the catalytic reaction with few variations in the predicted binding site for the leaving or acceptor glycan. However, these differences cannot explain the orders of magnitudes between their transfer activities, which must be due to other unidentified structural features of the proteins or substrates selectivity. Interestingly, the phylogenetic relationships between the lectin domains correlate with their specific trans-sialylation activities. This raises the question whether and how the lectin domains regulate the trans-sialidase reaction. The identification and enzymatic characterisation of the trans-sialidase family in T. congolense will contribute significantly towards the understanding of the roles of these enzymes in the pathogenesis of Animal African Trypanosomiasis.

  11. Congenital Trypanosoma cruzi Transmission in Santa Cruz, Bolivia

    PubMed Central

    Bern, Caryn; Verastegui, Manuela; Gilman, Robert H.; LaFuente, Carlos; Galdos-Cardenas, Gerson; Calderon, Maritza; Pacori, Juan; Abastoflor, Maria del Carmen; Aparicio, Hugo; Brady, Mark F.; Ferrufino, Lisbeth; Angulo, Noelia; Marcus, Sarah; Sterling, Charles; Maguire, James H.

    2017-01-01

    Background We conducted a study of congenital Trypanosoma cruzi infection in Santa Cruz, Bolivia. Our objective was to apply new tools to identify weak points in current screening algorithms, and find ways to improve them. Methods Women presenting for delivery were screened by rapid and conventional serological tests. For infants of infected mothers, blood specimens obtained on days 0, 7, 21, 30, 90, 180, and 270 were concentrated and examined microscopically; serological tests were performed for the day 90, 180, and 270 specimens. Maternal and infant specimens, including umbilical tissue, were tested by polymerase chain reaction (PCR) targeting the kinetoplast minicircle and by quantitative PCR. Results Of 530 women, 154 (29%) were seropositive. Ten infants had congenital T. cruzi infection. Only 4 infants had positive results of microscopy evaluation in the first month, and none had positive cord blood microscopy results. PCR results were positive for 6 (67%) of 9 cord blood and 7 (87.5%) of 8 umbilical tissue specimens. PCR-positive women were more likely to transmit T. cruzi than were seropositive women with negative PCR results (P < .05). Parasite loads determined by quantitative PCR were higher for mothers of infected infants than for seropositive mothers of uninfected infants (P < .01). Despite intensive efforts, only 58% of at-risk infants had a month 9 specimen collected. Conclusions On the basis of the low sensitivity of microscopy in cord blood and high rate of loss to follow-up, we estimate that current screening programs miss one-half of all infected infants. Molecular techniques may improve early detection. PMID:19877966

  12. Vertical transmission of Trypanosoma evansi in experimentally infected rats.

    PubMed

    Campigotto, Gabriela; Volpato, Andréia; Galli, Gabriela M; Glombowsky, Patrícia; Baldissera, Matheus D; Miletti, Luiz Claudio; Jaguezeski, Antonise M; Stefani, Lenita M; Da Silva, Aleksandro S

    2017-03-01

    Many reproductive problems has been described in male and female animals infected by Trypanosoma evansi. Thus, the aim of this study was to evaluate the occurrence of vertical (Experiment I) and venereal (Experiment II) transmission of T. evansi in rats experimentally infected. In the experiment I, eight female Wistar rats were used: three animals as negative controls, and five rats were infected by T. evansi on day ten of gestation. Out of these eight females, half puppies were used for molecular analysis (polymerase chain reaction - PCR) for T. evansi. Two infected females showed delivery problems, such as stillbirth, and fetal death that also led to female death. Three female rats infected had normal delivery of stunted offspring at term that died 2 days after birth. Rats from the control group had normal delivery of healthy offspring. T. evansi PCR was positive for 80% (12/15) of pups in the infected group. For the experiment II, five male rats were infected by T. evansi, and monitored by blood smears to check the parasitemia level. When the male rats showed parasitemia between 2 and 5 parasites per field, they were individually housed with one female adult rat. After approximately 21 days, the females delivered their offspring. Blood sample was collected from the females for blood smears and T. evansi PCR tests, which revealed negative results. Therefore, we were able to prove the occurrence of transplacental transmission of T. evansi and its negative effect on female rats and their offspring.

  13. Identification of Contractile Vacuole Proteins in Trypanosoma cruzi

    PubMed Central

    Park, Miyoung; Martins, Vicente P.; Atwood, James; Moles, Kristen; Collins, Dalis; Rohloff, Peter; Tarleton, Rick; Moreno, Silvia N. J.; Orlando, Ron; Docampo, Roberto

    2011-01-01

    Contractile vacuole complexes are critical components of cell volume regulation and have been shown to have other functional roles in several free-living protists. However, very little is known about the functions of the contractile vacuole complex of the parasite Trypanosoma cruzi, the etiologic agent of Chagas disease, other than a role in osmoregulation. Identification of the protein composition of these organelles is important for understanding their physiological roles. We applied a combined proteomic and bioinfomatic approach to identify proteins localized to the contractile vacuole. Proteomic analysis of a T. cruzi fraction enriched for contractile vacuoles and analyzed by one-dimensional gel electrophoresis and LC-MS/MS resulted in the addition of 109 newly detected proteins to the group of expressed proteins of epimastigotes. We also identified different peptides that map to at least 39 members of the dispersed gene family 1 (DGF-1) providing evidence that many members of this family are simultaneously expressed in epimastigotes. Of the proteins present in the fraction we selected several homologues with known localizations in contractile vacuoles of other organisms and others that we expected to be present in these vacuoles on the basis of their potential roles. We determined the localization of each by expression as GFP-fusion proteins or with specific antibodies. Six of these putative proteins (Rab11, Rab32, AP180, ATPase subunit B, VAMP1, and phosphate transporter) predominantly localized to the vacuole bladder. TcSNARE2.1, TcSNARE2.2, and calmodulin localized to the spongiome. Calmodulin was also cytosolic. Our results demonstrate the utility of combining subcellular fractionation, proteomic analysis, and bioinformatic approaches for localization of organellar proteins that are difficult to detect with whole cell methodologies. The CV localization of the proteins investigated revealed potential novel roles of these organelles in phosphate metabolism

  14. Polyamine depletion inhibits the autophagic response modulating Trypanosoma cruzi infectivity.

    PubMed

    Vanrell, María C; Cueto, Juan A; Barclay, Jeremías J; Carrillo, Carolina; Colombo, María I; Gottlieb, Roberta A; Romano, Patricia S

    2013-07-01

    Autophagy is a cell process that in normal conditions serves to recycle cytoplasmic components and aged or damaged organelles. The autophagic pathway has been implicated in many physiological and pathological situations, even during the course of infection by intracellular pathogens. Many compounds are currently used to positively or negatively modulate the autophagic response. Recently it was demonstrated that the polyamine spermidine is a physiological inducer of autophagy in eukaryotic cells. We have previously shown that the etiological agent of Chagas disease, the protozoan parasite Trypanosoma cruzi, interacts with autophagic compartments during host cell invasion and that preactivation of autophagy significantly increases host cell colonization by this parasite. In the present report we have analyzed the effect of polyamine depletion on the autophagic response of the host cell and on T. cruzi infectivity. Our data showed that depleting intracellular polyamines by inhibiting the biosynthetic enzyme ornithine decarboxylase with difluoromethylornithine (DFMO) suppressed the induction of autophagy in response to starvation or rapamycin treatment in two cell lines. This effect was associated with a decrease in the levels of LC3 and ATG5, two proteins required for autophagosome formation. As a consequence of inhibiting host cell autophagy, DFMO impaired T. cruzi colonization, indicating that polyamines and autophagy facilitate parasite infection. Thus, our results point to DFMO as a novel autophagy inhibitor. While other autophagy inhibitors such as wortmannin and 3-methyladenine are nonspecific and potentially toxic, DFMO is an FDA-approved drug that may have value in limiting autophagy and the spread of the infection in Chagas disease and possibly other pathological settings.

  15. Polyamine depletion inhibits the autophagic response modulating Trypanosoma cruzi infectivity

    PubMed Central

    Vanrell, María C.; Cueto, Juan A.; Barclay, Jeremías J.; Carrillo, Carolina; Colombo, María I.; Gottlieb, Roberta A.; Romano, Patricia S.

    2013-01-01

    Autophagy is a cell process that in normal conditions serves to recycle cytoplasmic components and aged or damaged organelles. The autophagic pathway has been implicated in many physiological and pathological situations, even during the course of infection by intracellular pathogens. Many compounds are currently used to positively or negatively modulate the autophagic response. Recently it was demonstrated that the polyamine spermidine is a physiological inducer of autophagy in eukaryotic cells. We have previously shown that the etiological agent of Chagas disease, the protozoan parasite Trypanosoma cruzi, interacts with autophagic compartments during host cell invasion and that preactivation of autophagy significantly increases host cell colonization by this parasite. In the present report we have analyzed the effect of polyamine depletion on the autophagic response of the host cell and on T. cruzi infectivity. Our data showed that depleting intracellular polyamines by inhibiting the biosynthetic enzyme ornithine decarboxylase with difluoromethylornithine (DFMO) suppressed the induction of autophagy in response to starvation or rapamycin treatment in two cell lines. This effect was associated with a decrease in the levels of LC3 and ATG5, two proteins required for autophagosome formation. As a consequence of inhibiting host cell autophagy, DFMO impaired T. cruzi colonization, indicating that polyamines and autophagy facilitate parasite infection. Thus, our results point to DFMO as a novel autophagy inhibitor. While other autophagy inhibitors such as wortmannin and 3-methyladenine are nonspecific and potentially toxic, DFMO is an FDA-approved drug that may have value in limiting autophagy and the spread of the infection in Chagas disease and possibly other pathological settings. PMID:23697944

  16. Optimized Multilocus Sequence Typing (MLST) Scheme for Trypanosoma cruzi

    PubMed Central

    Diosque, Patricio; Tomasini, Nicolás; Lauthier, Juan José; Messenger, Louisa Alexandra; Monje Rumi, María Mercedes; Ragone, Paula Gabriela; Alberti-D'Amato, Anahí Maitén; Pérez Brandán, Cecilia; Barnabé, Christian; Tibayrenc, Michel; Lewis, Michael David; Llewellyn, Martin Stephen; Miles, Michael Alexander; Yeo, Matthew

    2014-01-01

    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. PMID:25167160

  17. Optimized multilocus sequence typing (MLST) scheme for Trypanosoma cruzi.

    PubMed

    Diosque, Patricio; Tomasini, Nicolás; Lauthier, Juan José; Messenger, Louisa Alexandra; Monje Rumi, María Mercedes; Ragone, Paula Gabriela; Alberti-D'Amato, Anahí Maitén; Pérez Brandán, Cecilia; Barnabé, Christian; Tibayrenc, Michel; Lewis, Michael David; Llewellyn, Martin Stephen; Miles, Michael Alexander; Yeo, Matthew

    2014-08-01

    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.

  18. New Trypanosoma evansi Type B Isolates from Ethiopian Dromedary Camels

    PubMed Central

    Birhanu, Hadush; Gebrehiwot, Tadesse; Goddeeris, Bruno Maria; Büscher, Philippe; Van Reet, Nick

    2016-01-01

    Background 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. Methodology/principal findings 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. Conclusions/significance 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. PMID:27035661

  19. Novel trypanosome Trypanosoma gilletti sp. (Euglenozoa: Trypanosomatidae) and the extension of the host range of Trypanosoma copemani to include the koala ( Phascolarctos cinereus).

    PubMed

    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.

  20. TESA-blot for the diagnosis of Chagas disease in dogs from co-endemic regions for Trypanosoma cruzi, Trypanosoma evansi and Leishmania chagasi.

    PubMed

    Umezawa, E S; Souza, A I; Pinedo-Cancino, V; Marcondes, M; Marcili, A; Camargo, L M A; Camacho, A A; Stolf, A M S; Teixeira, M M G

    2009-07-01

    We standardized serodiagnosis of dogs infected with Trypanosoma cruzi using TESA (trypomastigote excreted-secreted antigen)-blot developed for human Chagas disease. TESA-blot showed 100% sensitivity and specificity. In contrast, ELISA using TESA (TESA-ELISA) or epimastigotes (epi-ELISA) as antigen yielded 100% sensitivity but specificity of 94.1% and 49.4%, respectively. When used in field studies in an endemic region for Chagas disease, visceral leishmaniasis and Trypanosoma evansi (Mato Grosso do Sul state, Central Brazil), positivities were 9.3% for TESA-blot, 10.7% for TESA-ELISA and 32% for epi-ELISA. Dogs from a non-endemic region for these infections (Rondonia state, western Amazonia) where T. cruzi is enzootic showed positivity of 4.5% for TESA-blot and epi-ELISA and 6.8% for TESA-ELISA. Sera from urban dogs from Santos, São Paulo, where these diseases are absent, yielded negative results. TESA-blot was the only method that distinguished dogs infected with T. cruzi from those infected with Leishmania chagasi and/or Trypanosoma evansi.

  1. Trypanosomiasis:goats as a possible reservoir of Trypanosoma congolense in the Republic of the Sudan.

    PubMed

    Mahmoud, M M; Elmalik, K H

    1977-08-01

    Experimental Trypanosoma congolense infections of goats and calves were compared. Goats developed a chronic form of trypanosomiasis, often recovering spontaneously from a strain which caused an acute fatal disease in calves. Goats may be important in the maintenace of T. congolense in nature in the Sudan.

  2. Anti-trypanosomal effect of Peristrophe bicalyculata extract on Trypanosoma brucei brucei-infected rats

    PubMed Central

    Abimbola, Abdulazeez Mansurah; Baba, Ibrahim Abdulrazak; Yenusa, Edibo Zakari; Omanibe, Sidali Joseph; Oladimeji, Idris Habeeb

    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. PMID:23835905

  3. Two new Trypanosoma species from African birds, with notes on the taxonomy of avian trypanosomes.

    PubMed

    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 (<30 µm in length on average) with the kinetoplast situated close to the posterior end of the body. 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.

  4. Likely Autochthonous Transmission of Trypanosoma cruzi to Humans, South Central Texas, USA

    PubMed Central

    Gunter, Sarah M.; Murray, Kristy O.; Gorchakov, Rodion; Beddard, Rachel; Rossmann, Susan N.; Montgomery, Susan P.; Rivera, Hilda; Brown, Eric L.; Aguilar, David; Widman, Lawrence E.

    2017-01-01

    Chagas disease, caused by Trypanosoma cruzi, is a major neglected tropical disease affecting the Americas. The epidemiology of this disease in the United States is incomplete. We report evidence of likely autochthonous vectorborne transmission of T. cruzi and health outcomes in T. cruzi–seropositive blood donors in south central Texas, USA. PMID:28221110

  5. Molecular characterization of Trypanosoma cruzi Mexican strains and their behavior in the mouse experimental model.

    PubMed

    Gómez-Hernández, César; Rezende-Oliveira, Karine; Nascentes, Gabriel Antônio Nogueira; Batista, Lara Rocha; Kappel, Henrique Borges; Martinez-Ibarra, José Alejandro; Trujillo Contreras, Francisco; Lages-Silva, Eliane; Ramírez, Luis Eduardo

    2011-01-01

    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. 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). 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. The Mexican T. cruzi strains are myotrophic and belong to group TcI.

  6. Ageing is not associated with an altered immune response during Trypanosoma cruzi infection: Ageing and Trypanosoma cruzi infection.

    PubMed

    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.

  7. Trypanosoma from rodents as potential source of infection in human-shaped landscapes of South-East Asia.

    PubMed

    Pumhom, Pornpan; Morand, Serge; Tran, Annelise; Jittapalapong, Sathaporn; Desquesnes, Marc

    2015-03-15

    Reports of atypical human cases of Trypanosoma lewisi or T. lewisi-like and Trypanosoma evansi infections have increased in South-East Asia, urging to investigate the possible links between humans, animal reservoirs and habitats. We tested how habitat structure affects the infection by Trypanosoma species of common murine rodents, inhabiting human-dominated landscapes in South East Asia. For this, we used geo-referenced data of rodents investigated for Trypanosoma infection and land cover maps produced for seven study sites in Thailand, Cambodia and Lao PDR. High prevalence of infection by T. lewisi was observed in rodents living near human settlement and in areas with high cover of built-up habitat, while the infection of rodents by T. evansi was explained by increased landscape patchiness and high cover of rain-fed agriculture lands. These results suggest a likely role of wild rodents as reservoir and possible source of atypical human infection by animal trypanosomes.

  8. Redescription of Trypanosoma ophiocephali Chen 1964 (Kinetoplastida: Trypanosomatina: Trypanosomatidae) and first record from the blood of dark sleeper (Odontobutis obscura Temminck and Schlegel) in China.

    PubMed

    Gu, Zemao; Wang, Jianguo; Zhang, Jinyong; Gong, Xiaoning

    2006-12-01

    During the parasite fauna investigation within 2004 and 2005, the freshwater fish trypanosomes were isolated from the blood of dark sleeper (Odontobutis obscura Temminck and Schlegel) and snakehead fish (Ophiocephalus argus Cantor) from Niushan Lake, Hubei Province, China. Blood trypomastigotes were used for light microscopy investigations. The detailed descriptions of three morphological groups of the genus Trypanosoma: Trypanosoma sp. I and Trypanosoma sp. II found in blood of O. obscura, and Trypanosoma sp. III found in blood of O. argus were provided. Morphological features and host species show Trypanosoma sp. III belong to Trypanosoma ophiocephali Chen 1964, an incompletely described species. Infection with trypanosomes of O. obscura was recorded for the first time. According to the size and appearance, the trypanosomes in O. obscura were also tentatively identified as T. ophiocephali Chen 1964.

  9. Nam Con Son Basin

    SciTech Connect

    Tin, N.T.; Ty, N.D.; Hung, L.T.

    1994-07-01

    The Nam Con Son basin is the largest oil and gas bearing basin in Vietnam, and has a number of producing fields. The history of studies in the basin can be divided into four periods: Pre-1975, 1976-1980, 1981-1989, and 1990-present. A number of oil companies have carried out geological and geophysical studies and conducted drilling activities in the basin. These include ONGC, Enterprise Oil, BP, Shell, Petro-Canada, IPL, Lasmo, etc. Pre-Tertiary formations comprise quartz diorites, granodiorites, and metamorphic rocks of Mesozoic age. Cenozoic rocks include those of the Cau Formation (Oligocene and older), Dua Formation (lower Miocene), Thong-Mang Cau Formation (middle Miocene), Nam Con Son Formation (upper Miocene) and Bien Dong Formation (Pliocene-Quaternary). The basement is composed of pre-Cenozoic formations. Three fault systems are evident in the basin: north-south fault system, northeast-southwest fault system, and east-west fault system. Four tectonic zones can also be distinguished: western differentiated zone, northern differentiated zone, Dua-Natuna high zone, and eastern trough zone.

  10. Transmission of Donor-Derived Trypanosoma cruzi and Subsequent Development of Chagas Disease in a Lung Transplant Recipient

    PubMed Central

    Sonetti, D.; Maloney, J. D.; Montgomery, S. P.; Rademacher, B. L.; Taylor, L. J.; Smith, J. A.; Striker, R.

    2017-01-01

    Donor infection status should be considered when accepting an organ for transplant. Here we present a case of Chagas disease developing after a lung transplant where the donor was known to be Trypanosoma cruzi antibody positive. The recipient developed acute Trypanosoma cruzi infection with reactivation after treatment. Chagas disease-positive donors are likely to be encountered in the United States; donor targeted screening is needed to guide decisions regarding organ transplant and posttransplant monitoring.

  11. Nitric oxide-releasing polymeric nanoparticles against Trypanosoma cruzi

    NASA Astrophysics Data System (ADS)

    Seabra, A. B.; Kitice, N. A.; Pelegrino, M. T.; Lancheros, C. A. C.; Yamauchi, L. M.; Pinge-Filho, P.; Yamada-Ogatta, S. F.

    2015-05-01

    Chagas disease, also known as American trypanosomiasis, is a potentially life-threatening illness caused by the protozoan parasite, Trypanosoma cruzi (T. cruzi), and the disease remains a major health problem in many Latin American countries. Several papers report that the killing of the parasite is dependent on the production of nitric oxide (NO). The endogenous free radical NO is an important cellular signalling molecule that plays a key role in the defense against pathogens, including T. cruzi. As T. cruzi is able to compromise host macrophages decreasing endogenous NO production, the administration of exogenous NO donors represents an interesting strategy to combat Chagas disease. Thus, the aims of this study were to prepare and evaluate the antimicrobial activity of NO-releasing polymeric nanoparticles against T. cruzi. Biocompatible polymeric nanoparticles composed of chitosan/sodium tripolyphosphate(TPP) were prepared and used to encapsulate mercaptosuccinic acid (MSA), which is a thiol-containing molecule. Nitrosation of free thiols (SH) groups of MSA were performed by the addition of equimolar amount of sodium nitrite (NaNO2), leading to the formation of S-nitroso-MSA-containing nanoparticles. These polymeric nanoparticles act as spontaneous NO donors, with free NO release. The results show the formation of nanoparticles with average hydrodynamic diameter ranging from 270 to 500 nm, average of polydispersity index of 0.35, and encapsulation efficiency in the range of 99%. The NO release kinetics from the S-nitroso-MSA-containing nanoparticles showed sustained and controlled NO release over several hours. The microbicidal activity of S-nitroso-MSA-containing nanoparticles was evaluated by incubating NO-releasing nanoparticles (200 - 600 μg/mL) with replicative and non-infective epimastigote, and non-replicative and infective trypomastigote forms of T. cruzi. In addition, a significant decrease in the percentage of macrophage-infected (with amastigotes) and

  12. Landscape ecology of Trypanosoma cruzi in the southern Yucatan Peninsula.

    PubMed

    López-Cancino, Sury Antonio; Tun-Ku, Ezequiel; De la Cruz-Felix, Himmler Keynes; Ibarra-Cerdeña, Carlos Napoleón; Izeta-Alberdi, Amaia; Pech-May, Angélica; Mazariegos-Hidalgo, Carlos Jesús; Valdez-Tah, Alba; Ramsey, Janine M

    2015-11-01

    Landscape interactions of Trypanosoma cruzi (Tc) with Triatoma dimidiata (Td) depend on the presence and relative abundance of mammal hosts. This study analyzed a landscape adjacent to the Calakmul Biosphere Reserve, composed of conserved areas, crop and farming areas, and the human community of Zoh Laguna with reported Chagas disease cases. Sylvatic mammals of the Chiroptera, Rodentia, and Marsupialia orders were captured, and livestock and pets were sampled along with T. dimidiata in all habitats. Infection by T. cruzi was analyzed using mtDNA markers, while lineage and DTU was analyzed using the mini-exon. 303 sylvatic specimens were collected, corresponding to 19 species during the rainy season and 114 specimens of 18 species during dry season. Five bats Artibeus jamaicensis, Artibeus lituratus, Sturnira lilium, Sturnira ludovici, Dermanura phaeotis (Dp) and one rodent Heteromys gaumeri were collected in the three habitats. All but Dp, and including Carollia brevicauda and Myotis keaysi, were infected with predominately TcI in the sylvatic habitat and TcII in the ecotone. Sigmodon hispidus was the rodent with the highest prevalence of infection by T. cruzi I and II in ecotone and domestic habitats. Didelphis viginiana was infected only with TcI in both domestic and sylvatic habitats; the only two genotyped human cases were TcII. Two main clades of T. cruzi, lineages I (DTU Ia) and II (DTU VI), were found to be sympatric (all habitats and seasons) in the Zoh-Laguna landscape, suggesting that no species-specific interactions occur between the parasite and any mammal host, in any habitat. We have also found mixed infections of the two principal T. cruzi clades in individuals across modified habitats, particularly in livestock and pets, and in both haplogroups of T. dimidiata. Results are contradictory to the dilution hypothesis, although we did find that most resilient species had an important role as T. cruzi hosts. Our study detected some complex trends in

  13. The Complement System: A Prey of Trypanosoma cruzi

    PubMed Central

    Lidani, Kárita C. F.; Bavia, Lorena; Ambrosio, Altair R.; de Messias-Reason, Iara J.

    2017-01-01

    Trypanosoma cruzi is a protozoan parasite known to cause Chagas disease (CD), a neglected sickness that affects around 6–8 million people worldwide. Originally, CD was mainly found in Latin America but more recently, it has been spread to countries in North America, Asia, and Europe due the international migration from endemic areas. Thus, at present CD represents an important concern of global public health. Most of individuals that are infected by T. cruzi may remain in asymptomatic form all lifelong, but up to 40% of them will develop cardiomyopathy, digestive mega syndromes, or both. The interaction between the T. cruzi infective forms and host-related immune factors represents a key point for a better understanding of the physiopathology of CD. In this context, the complement, as one of the first line of host defense against infection was shown to play an important role in recognizing T. cruzi metacyclic trypomastigotes and in controlling parasite invasion. The complement consists of at least 35 or more plasma proteins and cell surface receptors/regulators, which can be activated by three pathways: classical (CP), lectin (LP), and alternative (AP). The CP and LP are mainly initiated by immune complexes or pathogen-associated molecular patterns (PAMPs), respectively, whereas AP is spontaneously activated by hydrolysis of C3. Once activated, several relevant complement functions are generated which include opsonization and phagocytosis of particles or microorganisms and cell lysis. An important step during T. cruzi infection is when intracellular trypomastigotes are release to bloodstream where they may be target by complement. Nevertheless, the parasite uses a sequence of events in order to escape from complement-mediated lysis. In fact, several T. cruzi molecules are known to interfere in the initiation of all three pathways and in the assembly of C3 convertase, a key step in the activation of complement. Moreover, T. cruzi promotes secretion of plasma

  14. Prevention of transfusional Trypanosoma cruzi infection in Latin America.

    PubMed

    Schmunis, G A

    1999-01-01

    Trypanosoma cruzi is a protozoan infection widely spread in Latin America, from Mexico in the north to Argentina and Chile in the south. The second most important way of acquiring the infection is by blood transfusion. Even if most countries of Latin America have law/decree/norms, that make mandatory the screening of blood donors for infectious diseases, including T. cruzi (El Salvador and Nicaragua do not have laws on the subject), there is usually no enforcement or it is very lax. Analysis of published serologic surveys of T. cruzi antibodies in blood donors done in 1993, indicating the number of donors and screening coverage for T. cruzi in ten countries of Central and South America indicated that the probability of receiving a potentially infected transfusion unit in each country varied from 1,096 per 10,000 transfusions in Bolivia, the highest, to 13.02 or 13.86 per 10,000 transfusions in Honduras and Venezuela respectively, where screening coverage was 100%. On the other hand the probability of transmitting a T. cruzi infected unit was 219/10,000 in Bolivia, 24/10,000 in Colombia, 17/10,000 in El Salvador, and around 2-12/10,000 for the seven other countries. Infectivity risks defined as the likelihood of being infected when receiving an infected transfusion unit were assumed to be 20% for T. cruzi. Based on this, estimates of the absolute number of infections induced by transfusion indicated that they were 832, 236, and 875 in Bolivia, Chile and Colombia respectively. In all the other countries varied from seven in Honduras to 85 in El Salvador. Since 1993, the situation has improved. At that time only Honduras and Venezuela screened 100% of donors, while seven countries, Argentina, Colombia, El Salvador, Honduras, Paraguay, Uruguay and Venezuela, did the same in 1996. In Central America, without information from Guatemala, the screening of donors for T. cruzi prevented the transfusion of 1,481 infected units and the potential infection of 300 individuals in

  15. A genomic scale map of genetic diversity in Trypanosoma cruzi

    PubMed Central

    2012-01-01

    Background Trypanosoma cruzi, the causal agent of Chagas Disease, affects more than 16 million people in Latin America. The clinical outcome of the disease results from a complex interplay between environmental factors and the genetic background of both the human host and the parasite. However, knowledge of the genetic diversity of the parasite, is currently limited to a number of highly studied loci. The availability of a number of genomes from different evolutionary lineages of T. cruzi provides an unprecedented opportunity to look at the genetic diversity of the parasite at a genomic scale. Results Using a bioinformatic strategy, we have clustered T. cruzi sequence data available in the public domain and obtained multiple sequence alignments in which one or two alleles from the reference CL-Brener were included. These data covers 4 major evolutionary lineages (DTUs): TcI, TcII, TcIII, and the hybrid TcVI. Using these set of alignments we have identified 288,957 high quality single nucleotide polymorphisms and 1,480 indels. In a reduced re-sequencing study we were able to validate ~ 97% of high-quality SNPs identified in 47 loci. Analysis of how these changes affect encoded protein products showed a 0.77 ratio of synonymous to non-synonymous changes in the T. cruzi genome. We observed 113 changes that introduce or remove a stop codon, some causing significant functional changes, and a number of tri-allelic and tetra-allelic SNPs that could be exploited in strain typing assays. Based on an analysis of the observed nucleotide diversity we show that the T. cruzi genome contains a core set of genes that are under apparent purifying selection. Interestingly, orthologs of known druggable targets show statistically significant lower nucleotide diversity values. Conclusions This study provides the first look at the genetic diversity of T. cruzi at a genomic scale. The analysis covers an estimated ~ 60% of the genetic diversity present in the population, providing an

  16. A genomic scale map of genetic diversity in Trypanosoma cruzi.

    PubMed

    Ackermann, Alejandro A; Panunzi, Leonardo G; Cosentino, Raul O; Sánchez, Daniel O; Agüero, Fernán

    2012-12-27

    Trypanosoma cruzi, the causal agent of Chagas Disease, affects more than 16 million people in Latin America. The clinical outcome of the disease results from a complex interplay between environmental factors and the genetic background of both the human host and the parasite. However, knowledge of the genetic diversity of the parasite, is currently limited to a number of highly studied loci. The availability of a number of genomes from different evolutionary lineages of T. cruzi provides an unprecedented opportunity to look at the genetic diversity of the parasite at a genomic scale. Using a bioinformatic strategy, we have clustered T. cruzi sequence data available in the public domain and obtained multiple sequence alignments in which one or two alleles from the reference CL-Brener were included. These data covers 4 major evolutionary lineages (DTUs): TcI, TcII, TcIII, and the hybrid TcVI. Using these set of alignments we have identified 288,957 high quality single nucleotide polymorphisms and 1,480 indels. In a reduced re-sequencing study we were able to validate ~ 97% of high-quality SNPs identified in 47 loci. Analysis of how these changes affect encoded protein products showed a 0.77 ratio of synonymous to non-synonymous changes in the T. cruzi genome. We observed 113 changes that introduce or remove a stop codon, some causing significant functional changes, and a number of tri-allelic and tetra-allelic SNPs that could be exploited in strain typing assays. Based on an analysis of the observed nucleotide diversity we show that the T. cruzi genome contains a core set of genes that are under apparent purifying selection. Interestingly, orthologs of known druggable targets show statistically significant lower nucleotide diversity values. This study provides the first look at the genetic diversity of T. cruzi at a genomic scale. The analysis covers an estimated ~ 60% of the genetic diversity present in the population, providing an essential resource for future

  17. Heme modulates Trypanosoma cruzi bioenergetics inducing mitochondrial ROS production.

    PubMed

    Nogueira, Natália P; Saraiva, Francis M S; Oliveira, Matheus P; Mendonça, Ana Paula M; Inacio, Job D F; Almeida-Amaral, Elmo E; Menna-Barreto, Rubem F; Laranja, Gustavo A T; Torres, Eduardo J Lopes; Oliveira, Marcus F; Paes, Marcia C

    2017-03-29

    Trypanosoma cruzi is the causative agent of Chagas disease and has a single mitochondrion, an organelle responsible for ATP production and the main site for the formation of reactive oxygen species (ROS). T. cruzi is an obligate intracellular parasite with a complex life cycle that alternates between vertebrate and invertebrate hosts, therefore the development of survival strategies and morphogenetic adaptations to deal with the various environments is mandatory. Over the years our group has been studying the vector-parasite interactions using heme as a physiological oxidant molecule that triggered epimastigote proliferation however, the source of ROS induced by heme remained unknown. In the present study we demonstrate the involvement of heme in the parasite mitochondrial metabolism, decreasing oxygen consumption leading to increased mitochondrial ROS and membrane potential. First, we incubated epimastigotes with carbonyl cyanide p-(trifluoromethoxy) phenylhydrazone (FCCP), an uncoupler of oxidative phosphorylation, which led to decreased ROS formation and parasite proliferation, even in the presence of heme, correlating mitochondrial ROS and T. cruzi survival. This hypothesis was confirmed after the mitochondria-targeted antioxidant ((2-(2,2,6,6 Tetramethylpiperidin-1-oxyl-4-ylamino)-2-oxoethyl) triphenylphosphonium chloride (MitoTEMPO) decreased both heme-induced ROS and epimastigote proliferation. Furthermore, heme increased the percentage of tetramethylrhodamine methyl ester (TMRM) positive parasites tremendously-indicating the hyperpolarization and increase of potential of the mitochondrial membrane (ΔΨm). Assessing the mitochondrial functional metabolism, we observed that in comparison to untreated parasites, heme-treated epimastigotes decreased their oxygen consumption, and increased the complex II-III activity. These changes allowed the electron flow into the electron transport system, even though the complex IV (cytochrome c oxidase) activity decreased

  18. Sequence diversity of the Trypanosoma cruzi complement regulatory protein family.

    PubMed

    Beucher, M; Norris, K A

    2008-02-01

    As a central component of innate immunity, complement activation is a critical mechanism of containment and clearance of microbial pathogens in advance of the development of acquired immunity. Several pathogens restrict complement activation through the acquisition of host proteins that regulate complement activation or through the production of their own complement regulatory molecules (M. K. Liszewski, M. K. Leung, R. Hauhart, R. M. Buller, P. Bertram, X. Wang, A. M. Rosengard, G. J. Kotwal, and J. P. Atkinson, J. Immunol. 176:3725-3734, 2006; J. Lubinski, L. Wang, D. Mastellos, A. Sahu, J. D. Lambris, and H. M. Friedman, J. Exp. Med. 190:1637-1646, 1999). The infectious stage of the protozoan parasite Trypanosoma cruzi produces a surface-anchored complement regulatory protein (CRP) that functions to inhibit alternative and classical pathway complement activation (K. A. Norris, B. Bradt, N. R. Cooper, and M. So, J. Immunol. 147:2240-2247, 1991). This study addresses the genomic complexity of the T. cruzi CRP and its relationship to the T. cruzi supergene family comprising active trans-sialidase (TS) and TS-like proteins. The TS superfamily consists of several functionally distinct subfamilies that share a characteristic sialidase domain at their amino termini. These TS families include active TS, adhesions, CRPs, and proteins of unknown functions (G. A. Cross and G. B. Takle, Annu. Rev. Microbiol. 47:385-411, 1993). A sequence comparison search of GenBank using BLASTP revealed several full-length paralogs of CRP. These proteins share significant homology at their amino termini and a strong spatial conservation of cysteine residues. Alternative pathway complement regulation was confirmed for CRP paralogs with 58% (low) and 83% (high) identity to AAB49414. CRPs are functionally similar to the microbial and mammalian proteins that regulate complement activation. Sequence alignment of mammalian complement control proteins to CRP showed that these sequences are

  19. Intracellular Ca2+ storage in acidocalcisomes of Trypanosoma cruzi.

    PubMed Central

    Docampo, R; Scott, D A; Vercesi, A E; Moreno, S N

    1995-01-01

    The use of digitonin to permeabilize the plasma membrane of Trypanosoma cruzi allowed the identification of a non-mitochondrial nigericin- or bafilomycin A1-sensitive Ca(2+)-uptake mechanism. Proton uptake, as detected by ATP-dependent Acridine Orange accumulation, was also demonstrated in these permeabilized cells. Under these conditions Acridine Orange was concentrated in abundant cytoplasmic round vacuoles. This latter process was inhibited (and reversed) by bafilomycin A1, nigericin and NH4Cl in different stages of T. cruzi. Ca2+ released Acridine Orange from permeabilized cells, suggesting that the dye and Ca2+ were being accumulated in the same acidic compartment and that Ca2+ was taken up in exchange for protons. Addition of bafilomycin A1 (5 microM), nigericin (1 microM) or carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP; 1 microM) to fura 2-loaded epimastigotes increased their intracellular Ca2+ concentration ([Ca2+]i). Although this effect was more noticeable in the presence of extracellular Ca2+, it was also observed in its absence. Addition of NH4Cl (10-40 mM) to different stages of T. cruzi, in the nominal absence of extracellular Ca2+ to preclude Ca2+ entry, increased both [Ca2+]i in fura 2-loaded cells, and intracellular pH (pHi) in 2',7'-bis-(2-carboxyethyl)-5-(and -6)-carboxyfluorescein acetoxymethyl ester (BCECF)-loaded cells. Treatment of the cells with the Ca2+ ionophore ionomycin under similar conditions (nominal absence of extracellular Ca2+) resulted in an increase in [Ca2+]i and a significantly higher increase in [Ca2+]i after addition of NH4Cl, nigericin or bafilomycin A1, all agents which increase the pH of acidic compartments and make ionomycin more effective as a Ca(2+)-releasing ionophore. Similar results were obtained when the order of additions was reversed. Taking into account the relative importance of the ionomycin-releasable and the ionomycin plus NH4Cl-releasable Ca2+ pools, it is apparent that most of the Ca2+ stored in

  20. The potential of canine sentinels for reemerging Trypanosoma cruzi transmission

    PubMed Central

    Neyra, Ricardo Castillo; Chu, Lily Chou; Quispe-Machaca, Victor; Ancca-Juarez, Jenny; Malaga Chavez, Fernando S.; Mazuelos, Milagros Bastos; Naquira, Cesar; Bern, Caryn; Gilman, Robert H.; Levy, Michael Z.

    2015-01-01

    Background Chagas disease, a vector-borne disease transmitted by triatomine bugs and caused by the parasite Trypanosoma cruzi, affects millions of people in the Americas. In Arequipa, Peru, indoor residual insecticide spraying campaigns are routinely conducted to eliminate Triatoma infestans, the only vector in this area. Following insecticide spraying, there is risk of vector return and reinitiation of parasite transmission. Dogs are important reservoirs of T. cruzi and may play a role in reinitiating transmission in previously sprayed areas. Dogs may also serve as indicators of reemerging transmission. Methods We conducted a cross-sectional serological screening to detect T. cruzi antibodies in dogs, in conjunction with an entomological vector collection survey at the household level, in a disease endemic area that had been treated with insecticide 13 years prior. Spatial clustering of infected animals and vectors was assessed using Ripley’s K statistic, and the odds of being seropositive for dogs proximate to infected colonies was estimated with multivariate logistic regression. Results There were 106 triatomine-infested houses (41.1%), and 45 houses infested with T. cruzi-infected triatomine insects (17.4%). Canine seroprevalence in the area was 12.3% (n=154); all seropositive dogs were 9 months old or older. We observed clustering of vectors carrying the parasite, but no clustering of seropositive dogs. The age- and sex-adjusted odds ratio between seropositivity to T. cruzi and proximity to an infected triatomine (≤50m) was 5.67 (95% CI: 1.12 – 28.74; p=0.036). Conclusions Targeted control of reemerging transmission can be achieved by improved understanding of T. cruzi in canine populations. Our results suggest that dogs may be useful sentinels to detect re-initiation of transmission following insecticide treatment. Integration of canine T. cruzi blood sampling into existing interventions for zoonotic disease control (e.g. rabies vaccination programs

  1. Recent, independent and anthropogenic origins of Trypanosoma cruzi hybrids.

    PubMed

    Lewis, Michael D; Llewellyn, Martin S; Yeo, Matthew; Acosta, Nidia; Gaunt, Michael W; Miles, Michael A

    2011-10-01

    The single celled eukaryote Trypanosoma cruzi, a parasite transmitted by numerous species of triatomine bug in the Americas, causes Chagas disease in humans. T. cruzi generally reproduces asexually and appears to have a clonal population structure. However, two of the six major circulating genetic lineages, TcV and TcVI, are TcII-TcIII inter-lineage hybrids that are frequently isolated from humans in regions where chronic Chagas disease is particularly severe. Nevertheless, a prevalent view is that hybridisation events in T. cruzi were evolutionarily ancient and that active recombination is of little epidemiological importance. We analysed genotypes of hybrid and non-hybrid T. cruzi strains for markers representing three distinct evolutionary rates: nuclear GPI sequences (n = 88), mitochondrial COII-ND1 sequences (n = 107) and 28 polymorphic microsatellite loci (n = 35). Using Maximum Likelihood and Bayesian phylogenetic approaches we dated key evolutionary events in the T. cruzi clade including the emergence of hybrid lineages TcV and TcVI, which we estimated to have occurred within the last 60,000 years. We also found evidence for recent genetic exchange between TcIII and TcIV and between TcI and TcIV. These findings show that evolution of novel recombinants remains a potential epidemiological risk. The clearly distinguishable microsatellite genotypes of TcV and TcVI were highly heterozygous and displayed minimal intra-lineage diversity indicative of even earlier origins than sequence-based estimates. Natural hybrid genotypes resembled typical meiotic F1 progeny, however, evidence for mitochondrial introgression, absence of haploid forms and previous experimental crosses indicate that sexual reproduction in T. cruzi may involve alternatives to canonical meiosis. Overall, the data support two independent hybridisation events between TcII and TcIII and a recent, rapid spread of the hybrid progeny in domestic transmission cycles concomitant with, or as a

  2. Preparation, crystallization and preliminary crystallographic analysis of old yellow enzyme from Trypanosoma cruzi

    SciTech Connect

    Sugiyama, Shigeru; Tokuoka, Keiji; Uchiyama, Nahoko; Okamoto, Naoki; Okano, Yousuke; Matsumura, Hiroyoshi; Inaka, Koji; Urade, Yoshihiro; Inoue, Tsuyoshi

    2007-10-01

    Old yellow enzyme from Trypanosoma cruzi, has been crystallized using the hanging-drop vapour-diffusion method. Old yellow enzyme (OYE) is an NADPH oxidoreductase that contains a flavin mononucleotide as a prosthetic group. The OYE from Trypanosoma cruzi, which produces prostaglandin F{sub 2α}, a potent mediator of various physiological and pathological processes, from prostaglandin H2. The protein was recombinantly expressed and purified from Escherichia coli and was crystallized using the hanging-drop vapour-diffusion method. The crystal belongs to the monoclinic space group P2{sub 1}, with unit-cell parameters a = 56.3, b = 78.8, c = 78.8 Å, β = 93.4° and two molecules per asymmetric unit. The crystals were suitable for X-ray crystallographic studies and diffracted to 1.70 Å resolution. A Patterson search method is in progress using the structure of OYE from Pseudomonas putida as a starting model.

  3. Polymerase Chain Reaction Detection of Trypanosoma cruzi in Macaca fascicularis Using Archived Tissues

    PubMed Central

    Williams, Jeff T.; Mubiru, James N.; Schlabritz-Loutsevitch, Natalia E.; Rubicz, Rohina C.; VandeBerg, John L.; Dick, Edward J.; Hubbard, Gene B.

    2009-01-01

    This study describes conventional and real-time polymerase chain reaction (PCR) methods developed to detect and quantify Trypanosoma cruzi DNA in cynomolgus monkeys (Macaca fascicularis) using formalin-fixed paraffin-embedded blocks archived for periods of up to 6 years. The highest concentration of T. cruzi DNA was found in the myocardium, urinary bladder, stomach, lymph node, adrenal gland, and colon. The concentration of T. cruzi DNA detected in cardiac tissues was 10–100-fold greater than found elsewhere; the mean concentrations of T. cruzi DNA in non-cardiac tissues were otherwise comparable. Trypanosoma cruzi DNA was amplified from cerebrum but not cerebellum or kidney. Successful use of DNA from formalin-fixed, paraffin-embedded blocks is important because most pathology laboratories routinely archive wax blocks. This archived resource can be used for further studies on the prevalence of this disease. PMID:19635875

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

    PubMed

    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.

  5. Structural and Functional Highlights of Vacuolar Soluble Protein 1 from Pathogen Trypanosoma brucei brucei*

    PubMed Central

    Jamwal, Abhishek; Round, Adam R.; Bannwarth, Ludovic; Venien-Bryan, Catherine; Belrhali, Hassan; Yogavel, Manickam; Sharma, Amit

    2015-01-01

    Trypanosoma brucei (T. brucei) is responsible for the fatal human disease called African trypanosomiasis, or sleeping sickness. The causative parasite, Trypanosoma, encodes soluble versions of inorganic pyrophosphatases (PPase), also called vacuolar soluble proteins (VSPs), which are localized to its acidocalcisomes. The latter are acidic membrane-enclosed organelles rich in polyphosphate chains and divalent cations whose significance in these parasites remains unclear. We here report the crystal structure of T. brucei brucei acidocalcisomal PPases in a ternary complex with Mg2+ and imidodiphosphate. The crystal structure reveals a novel structural architecture distinct from known class I PPases in its tetrameric oligomeric state in which a fused EF hand domain arranges around the catalytic PPase domain. This unprecedented assembly evident from TbbVSP1 crystal structure is further confirmed by SAXS and TEM data. SAXS data suggest structural flexibility in EF hand domains indicative of conformational plasticity within TbbVSP1. PMID:26494625

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

    USGS Publications Warehouse

    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.

  7. Seroprevalence of Trypanosoma cruzi in stray and pet dogs in Grenada, West Indies.

    PubMed

    Chikweto, A; Kumthekar, S; Chawla, P; Tiwari, K P; Perea, L M; Paterson, T; Sharma, R N

    2014-06-01

    American trypanosomiasis (Chagas disease) caused by the protozoan parasite Trypanosoma cruzi is endemic to parts of South America and the Caribbean. Infected dogs are important in the epidemiology of the parasite as they can play a role in the transmission of the parasite to humans. A total of 399 dog sera (242 stray and 157 pet dogs) were examined for T. cruzi infection; using a qualitative immunochromatographic dipstick test, based on recombinant antigens specific for American trypanosomiasis (Trypanosoma detect rapid test; InBios international, Inc., Seattle, Washington). Overall seroprevalence for T. cruzi was estimated at 10.5% (95% confidence interval: 7.5% to 13.5%); with stray dogs being significantly more affected (p<0.05, χ2). Results from this study indicate that dogs in Grenada are moderately exposed to T. cruzi compared to other areas in the region.

  8. Mammalian cell invasion by closely related Trypanosoma species T. dionisii and T. cruzi.

    PubMed

    Maeda, Fernando Yukio; Cortez, Cristian; Alves, Renan Melatto; Yoshida, Nobuko

    2012-02-01

    Protozoan parasites of the genus Trypanosoma can infect virtually all mammalian species. Within this genus, Trypanosoma dionisii from bats and Trypanosoma cruzi that causes Chagas' disease, belonging to the subgenus Schizotrypanum, can invade mammalian cells. The mechanisms of cell invasion by T. dionisii are poorly understood. To address that question, metacyclic trypomastigotes (MT) and human epithelial HeLa cells were used. Similarly to genetically divergent T. cruzi strains G (TcI) and CL (TcVI), associated, respectively with marsupial and human infections, T. dionisii infectivity increased under nutritional stress, a condition that induces host cell lysosome exocytosis required for parasite internalization. For efficient internalization, T. dionisii depended on MT protein tyrosine kinase (PTK) and Ca(2+) mobilization from acidocalcisomes, whereas T. cruzi strains also relied on phosphatidylinositol 3-kinase (PI3K), protein kinase C (PKC) and Ca(2+) released from thapsigargin-sensitive compartments. T. dionisii-induced signaling in host cells implicated PKC and Ca(2+) mobilized from thapsigargin-sensitive stores, like T. cruzi, but without PI3K involvement. Unlike T. cruzi, T. dionisii metacyclic forms did not use l-proline as source of energy required for internalization. Molecules related to T. cruzi surface glycoproteins involved in MT-host cell interaction were undetectable in T. dionisii. The difference in the surface profile of the two species was also inferred from the susceptibility of T. dionisii metacyclic forms to complement-mediated lysis, as opposed to complete resistance of T. cruzi. In summary, the two Trypanosoma species display distinct surface profiles but invade host cells through a common mechanism involving lysosome mobilization to the site of parasite entry.

  9. Influence of saccharides and sodium chloride on growth and differentiation of Trypanosoma cruzi.

    PubMed

    Adroher, F J; Lupiáñez, J A; Osuna, A

    1988-01-01

    The influence of saccharides, especially glucose and fructose, on the metacyclogenesis and growth of Trypanosoma cruzi has been investigated. In the absence of glucose and fructose in the media, both the percentage of metacyclic forms and the growth increased significantly. Furthermore, the addition of NaCl to the medium without monosaccharides strongly increased the formation of metacyclic forms. Presence of NaCl and absence of monosaccharides showed a synergic effect on differentiation of T. cruzi.

  10. Farnesyl Diphosphate Synthase Localizes to the Cytoplasm of Trypanosoma cruzi and T.brucei

    PubMed Central

    Ferella, Marcela; Li, Zhu-Hong; Andersson, Björn; Docampo, Roberto

    2008-01-01

    The farnesyl diphosphate synthase (FPPS) has previously been characterized in trypanosomes as an essential enzyme for their survival and as the target for bisphosphonates, drugs that are effective both in vitro and in vivo against these parasites. Enzymes from the isoprenoid pathway have been assigned to different compartments in eukaryotes, including trypanosomatids. We here report that FPPS localizes to the cytoplasm of both Trypanosoma cruzi and T. brucei, and is not present in other organelles such as the mitochondria and glycosomes. PMID:18406406

  11. An evaluation of Minor Groove Binders as anti-Trypanosoma brucei brucei therapeutics.

    PubMed

    Scott, Fraser J; Khalaf, Abedawn I; Giordani, Federica; Wong, Pui Ee; Duffy, Sandra; Barrett, Michael; Avery, Vicky M; Suckling, Colin J

    2016-06-30

    A series of 32 structurally diverse MGBs, derived from the natural product distamycin, was evaluated for activity against Trypanosoma brucei brucei. Four compounds have been found to possess significant activity, in the nanomolar range, and represent hits for further optimisation towards novel treatments for Human and Animal African Trypanosomiases. Moreover, SAR indicates that the head group linking moiety is a significant modulator of biological activity.

  12. Digestion of human immunoglobulin G by the major cysteine proteinase (cruzipain) from Trypanosoma cruzi.

    PubMed

    Bontempi, E; Cazzulo, J J

    1990-08-01

    The major cysteine proteinase (cruzipain) from Trypanosoma cruzi was able to digest human IgG, as shown by polyacrylamide gel electrophoresis in the presence of SDS, and by gel filtration on a Superose 12 column, in a FPLC system. The Fab fragment of IgG was only slightly degraded, but Fc was extensively hydrolyzed to small peptides. The results suggest that cruzipain might be involved in the defense mechanisms of the parasite against the immune response of the host.

  13. Trypanosoma humboldti n. sp. from the Chilean catshark, Schroederichthys chilensis (Guichenot, 1848).

    PubMed

    Morillas, J; George-Nascimento, M; Valeria, H; Khan, R A

    1987-08-01

    The morphology of Trypanosoma humboldti n. sp. is described from living and stained specimens obtained from the blood of a catshark, Schroederichthys chilensis. This represents the first report of a trypanosome in fish from the eastern Pacific Ocean. It is distinguished by its size and apparent lack of pleomorphism. The presence of a leech, Branchellion ravenellii, attached to the catshark, raises the possibility that it can act as a vector. Additionally, this leech is recorded for the first time from the Pacific Ocean.

  14. First case of natural infection in pigs. Review of Trypanosoma cruzi reservoirs in Mexico.

    PubMed

    Salazar-Schettino, P M; Bucio, M I; Cabrera, M; Bautista, J

    1997-01-01

    An epidemiological research project was performed in the State of Morelos including collection of samples for blood smears and culture, serological tests, and xenodiagnoses from a total of 76 domestic and peridomestic mammals. Two strains of Trypanosoma cruzi were isolated by haemocultures; one from a pig (Sus scrofa), the first case of natural infection reported in Mexico, and the other from a dog (Canis familiaris). This study summarizes current information in Mexico concerning confirmed reservoirs of T. cruzi.

  15. Rapid Detection of Trypanosoma cruzi in Human Serum by Use of an Immunochromatographic Dipstick Test ▿

    PubMed Central

    Reithinger, Richard; Grijalva, Mario J.; Chiriboga, Rosa F.; Alarcón de Noya, Belkisyolé; Torres, Jaime R.; Pavia-Ruz, Norma; Manrique-Saide, Pablo; Cardinal, Marta V.; Gürtler, Ricardo E.

    2010-01-01

    We evaluated a commercially available immunochromatographic dipstick test to detect Trypanosoma cruzi infection in 366 human serum samples with known serological results from Argentina, Ecuador, Mexico, and Venezuela. One hundred forty-nine of 366 (40.7%) and 171/366 (46.7%) samples tested positive by dipstick and serology, respectively. Dipstick sensitivity was calculated to be 84.8% (range between countries, 77.5 to 95%), and specificity was 97.9% (95.9 to 100%). PMID:20534801

  16. Cross-resistance associated with development of resistance to isometamidium in a clone of Trypanosoma congolense.

    PubMed

    Peregrine, A S; Gray, M A; Moloo, S K

    1997-07-01

    Resistance to isometamidium was increased 94-fold in a clone of Trypanosoma congolense (clone IL 1180) by repeated subcurative treatment of infected mice for 11 months. This was associated with 3.4-, 33-, and 4.2-fold increases in resistance to diminazene, homidium, and quinapyramine, respectively. Both T. congolense IL 1180 and the resistant derivative were able to undergo cyclical development in Glossina morsitans centralis tsetse flies, producing hypopharyngeal infection rates of 40.0 and 39.8%, respectively.

  17. Evidence for Trypanosoma cruzi in adipose tissue in human chronic Chagas disease

    PubMed Central

    Ferreira, Adaliene Versiani Matos; Segatto, Marcela; Menezes, Zélia; Macedo, Andréa Mara; Gelape, Cláudio; de Oliveira Andrade, Luciana; Nagajyothi, Fnu; Scherer, Philipp E.; Teixeira, Mauro Martins; Tanowitz, Herbert B.

    2013-01-01

    Trypanosoma cruzi the cause of Chagas disease persists in tissues of infected experimental animals and humans. Here we demonstrate the persistence of the parasite in adipose tissue from of three of 10 elderly seropositive patients with chronic chagasic heart disease. Nine control patients had no parasites in the fat. We also demonstrate that T. cruzi parasitizes primary adipocytes in vitro. Thus, in humans as in mice the parasite may persist in adipose tissue for decades and become a reservoir of infection. PMID:21726660

  18. Evidence for Trypanosoma cruzi in adipose tissue in human chronic Chagas disease.

    PubMed

    Ferreira, Adaliene Versiani Matos; Segatto, Marcela; Menezes, Zélia; Macedo, Andréa Mara; Gelape, Cláudio; de Oliveira Andrade, Luciana; Nagajyothi, Fnu; Scherer, Philipp E; Teixeira, Mauro Martins; Tanowitz, Herbert B

    2011-11-01

    Trypanosoma cruzi the cause of Chagas disease persists in tissues of infected experimental animals and humans. Here we demonstrate the persistence of the parasite in adipose tissue from of three of 10 elderly seropositive patients with chronic chagasic heart disease. Nine control patients had no parasites in the fat. We also demonstrate that T. cruzi parasitizes primary adipocytes in vitro. Thus, in humans as in mice the parasite may persist in adipose tissue for decades and become a reservoir of infection.

  19. Trypanosoma cruzi modulates gene expression of plasma membrane repair-related proteins.

    PubMed

    Brígido, Rebecca Tavares E Silva; Tavares, Paula Cristina Brígido; Santos, Marlus Alves Dos; Santos, Júlia de Gouveia; Souza, Maria Aparecida de; Goulart, Isabela Maria Bernardes; Silva, Claudio Vieira da

    2017-10-01

    Plasma membrane injury and repair is particularly prevalent in muscle cells. Here, we aimed to verify dysferlin, acid sphingomyelinase and transcriptional factor EB gene expression during Trypanosoma cruzi infection in vitro and in vivo. Our results showed that the parasite modulates gene expression of these proteins in a way dependent on the number of plasma membrane interacting parasites and in a rapamycin-sensitive manner. Copyright © 2016 Elsevier B.V. All rights reserved.

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

    DTIC Science & Technology

    1980-04-01

    of Trypanosoma b. rhodesiense (WRATat Serodeme) .... 20 Figure 4: PAGE gel of VSSA2...respectively. Gel permeation chromatography using HPLC and a TSK 3,000-SW column was performed on WRATat-12 glycoorotein. Molecular weight analyses of...LECTIN*! Mac lura porrif-era Seeds lomoCjenize w/ blendor / 4 C Extract wl PBS - 24 h/ 4 C Filter Extract ’* i ue (di scird) I Filtrate (save) Add 1.1

  1. The diversity and expansion of the trans-sialidase gene family is a common feature in Trypanosoma cruzi clade members.

    PubMed

    Chiurillo, Miguel Angel; Cortez, Danielle R; Lima, Fábio M; Cortez, Caroline; Ramírez, José Luis; Martins, Andre G; Serrano, Myrna G; Teixeira, Marta M G; da Silveira, José Franco

    2016-01-01

    Trans-sialidase (TS) is a polymorphic protein superfamily described in members of the protozoan genus Trypanosoma. Of the eight TS groups recently described, TS group I proteins (some of which have catalytic activity) are present in the distantly related Trypanosoma brucei and Trypanosoma cruzi phylogenetic clades, whereas other TS groups have only been described in some species belonging to the T. cruzi clade. In the present study we analyzed the repertoire, distribution and phylogenetic relationships of TS genes among species of the T. cruzi clade based on sequence similarity, multiple sequence alignment and tree-reconstruction approaches using TS sequences obtained with the aid of PCR-based strategies or retrieved from genome databases. We included the following representative isolates of the T. cruzi clade from South America: T. cruzi, T. cruzi Tcbat, Trypanosoma cruzi marinkellei, Trypanosoma dionisii, Trypanosoma rangeli and Trypanosoma conorhini. The cloned sequences encoded conserved TS protein motifs Asp-box and VTVxNVxLYNR but lacked the FRIP motif (conserved in TS group I). The T. conorhini sequences were the most divergent. The hybridization patterns of TS probes with chromosomal bands confirmed the abundance of these sequences in species in the T. cruzi clade. Divergence and relationship analysis placed most of the TS sequences in the groups defined in T. cruzi. Further examination of members of TS group II, which includes T. cruzi surface glycoproteins implicated in host cell attachment and invasion, showed that sequences of T. cruzi Tcbat grouped with those of T. cruzi genotype TcI. Our analysis indicates that different members of the T. cruzi clade, with different vertebrate hosts, vectors and pathogenicity, share the extensive expansion and sequence diversification of the TS gene family. Altogether, our results are congruent with the evolutionary history of the T. cruzi clade and represent a contribution to the understanding of the molecular

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

    SciTech Connect

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

    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.

  3. Exploring the Trypanosoma brucei Hsp83 potential as a target for structure guided drug design.

    PubMed

    Pizarro, Juan Carlos; Hills, Tanya; Senisterra, Guillermo; Wernimont, Amy K; Mackenzie, Claire; Norcross, Neil R; Ferguson, Michael A J; Wyatt, Paul G; Gilbert, Ian H; Hui, Raymond

    2013-01-01

    Human African trypanosomiasis is a neglected parasitic disease that is fatal if untreated. The current drugs available to eliminate the causative agent Trypanosoma brucei have multiple liabilities, including toxicity, increasing problems due to treatment failure and limited efficacy. There are two approaches to discover novel antimicrobial drugs--whole-cell screening and target-based discovery. In the latter case, there is a need to identify and validate novel drug targets in Trypanosoma parasites. The heat shock proteins (Hsp), while best known as cancer targets with a number of drug candidates in clinical development, are a family of emerging targets for infectious diseases. In this paper, we report the exploration of T. brucei Hsp83--a homolog of human Hsp90--as a drug target using multiple biophysical and biochemical techniques. Our approach included the characterization of the chemical sensitivity of the parasitic chaperone against a library of known Hsp90 inhibitors by means of differential scanning fluorimetry (DSF). Several compounds identified by this screening procedure were further studied using isothermal titration calorimetry (ITC) and X-ray crystallography, as well as tested in parasite growth inhibitions assays. These experiments led us to the identification of a benzamide derivative compound capable of interacting with TbHsp83 more strongly than with its human homologs and structural rationalization of this selectivity. The results highlight the opportunities created by subtle structural differences to develop new series of compounds to selectively target the Trypanosoma brucei chaperone and effectively kill the sleeping sickness parasite.

  4. Apoptotic-like activity of staurosporine in axenic cultures of Trypanosoma evansi.

    PubMed

    Bruges, Gustavo; Betancourt, Meyerling; March, Mariana; Sanchez, Evangelina; Mijares, Alfredo

    2012-01-01

    Trypanosoma evansi is a blood protozoan parasite of the genus Trypanosoma which is responsible for surra (Trypanosomosis) in domestic and wild animals. This study addressed apoptotic-like features in Trypanosoma evansi in vitro. The mechanism of parasite death was investigated using staurosporine as an inducing agent. We evaluated its effects through several cytoplasmic features of apoptosis, including cell shrinkage, phosphatidylserine exposure, maintenance of plasma membrane integrity, and mitochondrial trans-membrane potential. For access to these features we have used the flow cytometry and fluorescence microscopy with cultures in the stationary phase and adjusted to a density of 10(6) cells/mL. The apoptotic effect of staurosporine in T. evansi was evaluated at 20 nM final concentration. There was an increase of phosphatidylserine exposure, whereas mitochondrial potential was decreased. Moreover, no evidence of cell permeability increasing with staurosporine was observed in this study, suggesting the absence of a necrotic process. Additional studies are needed to elucidate the possible pathways associated with this form of cell death in this hemoparasite.

  5. Developmental and Ultrastructural Characterization and Phylogenetic Analysis of Trypanosoma herthameyeri n. sp. of Brazilian Leptodactilydae Frogs.

    PubMed

    Attias, Márcia; Sato, Lyslaine H; Ferreira, Robson C; Takata, Carmen S A; Campaner, Marta; Camargo, Erney P; Teixeira, Marta M G; de Souza, Wanderley

    2016-09-01

    We described the phylogenetic affiliation, development in cultures and ultrastructural features of a trypanosome of Leptodacylus chaquensis from the Pantanal biome of Brazil. In the inferred phylogeny, this trypanosome nested into the Anura clade of the basal Aquatic clade of Trypanosoma, but was separate from all known species within this clade. This finding enabled us to describe it as Trypanosoma herthameyeri n. sp., which also infects other Leptodacylus species from the Pantanal and Caatinga biomes. Trypanosoma herthameyeri multiplies as small rounded forms clumped together and evolving into multiple-fission forms and rosettes of epimastigotes released as long forms with long flagella; scarce trypomastigotes and glove-like forms are common in stationary-phase cultures. For the first time, a trypanosome from an amphibian was observed by field emission scanning electron microscopy, revealing a cytostome opening, well-developed flagellar lamella, and many grooves in pumpkin-like forms. Transmission electron microscopy showed highly developed Golgi complexes, relaxed catenation of KDNA, and a rich set of spongiome tubules in a regular parallel arrangement to the flagellar pocket as confirmed by electron tomography. Considering the basal position in the phylogenetic tree, developmental and ultrastructural data of T. herthameyeri are valuable for evolutionary studies of trypanosome architecture and cell biology. © 2016 The Author(s) Journal of Eukaryotic Microbiology © 2016 International Society of Protistologists.

  6. Molecular characterization of Trypanosoma spp. infecting cattle (Bos taurus), white-tailed deer (Odocoileus virginianus), and elk (Cervus elaphus canadensis) in the United States

    USDA-ARS?s Scientific Manuscript database

    The benign trypanosomes of cattle and wild ungulates in the United States are designated Trypanosoma theileri and Trypanosoma cervi, respectively. Historically these parasites have been identified based on morphology, host, and vector, if known. No molecular characterization has been reported for T....

  7. Applicability of plant-based products in the treatment of Trypanosoma cruzi and Trypanosoma brucei infections: a systematic review of preclinical in vivo evidence.

    PubMed

    Pereira, Rodrigo M; Greco, Glícia M Z; Moreira, Andreia M; Chagas, Pablo F; Caldas, Ivo S; Gonçalves, Reggiani V; Novaes, Rômulo D

    2017-09-01

    Chagas disease and sleeping sickness are neglected tropical diseases closely related to poverty, for which the development of plant-derived treatments has not been a promising prospect. Thus, we systematicaly review the preclinical in vivo evidence on the applicability of plant-based products in the treatment of Trypanosoma cruzi and Trypanosoma brucei infections. Characteristics such as disease models, treatments, toxicological safety and methodological bias were analysed. We recovered 66 full text articles from 16 countries investigating 91 plant species. The disease models and treatments were highly variable. Most studies used native (n = 36, 54·54%) or exotic (n = 30, 45·46%) plants with ethnodirected indication (n = 45, 68·18%) for trypanosomiasis treatment. Complete phytochemical screening and toxicity assays were reported in only 15 (22·73%) and 32 (48·49%) studies, respectively. The currently available preclinical evidence is at high risk of bias. The absence of or incomplete characterization of animal models, treatment protocols, and phytochemical/toxicity analyses impaired the internal validity of the individual studies. Contradictory results of a same plant species compromise the external validity of the evidence, making it difficult determine the effectiveness, safety and biotechnological potential of plant-derived products in the development of new anti-infective agents to treat T. cruzi and T. brucei infections.

  8. Study of optically trapped living Trypanosoma cruzi/Trypanosoma rangeli - Rhodnius prolixus interactions by real time confocal images using CdSe quantum dots

    NASA Astrophysics Data System (ADS)

    de Thomaz, A. A.; Almeida, D. B.; Faustino, W. M.; Jacob, G. J.; Fontes, A.; Barbosa, L. C.; Cesar, C. L.; Stahl, C. V.; Santos-Mallet, J. R.; Gomes, S. A. O.; Feder, D.

    2008-08-01

    One of the fundamental goals in biology is to understand the interplay between biomolecules of different cells. This happen, for example, in the first moments of the infection of a vector by a parasite that results in the adherence to the cell walls. To observe this kind of event we used an integrated Optical Tweezers and Confocal Microscopy tool. This tool allow us to use the Optical Tweezers to trigger the adhesion of the Trypanosoma cruzi and Trypanosoma rangeli parasite to the intestine wall cells and salivary gland of the Rhodnius prolixus vector and to, subsequently observe the sequence of events by confocal fluorescence microscopy under optical forces stresses. We kept the microorganism and vector cells alive using CdSe quantum dot staining. Besides the fact that Quantum Dots are bright vital fluorescent markers, the absence of photobleaching allow us to follow the events in time for an extended period. By zooming to the region of interested we have been able to acquire confocal images at the 2 to 3 frames per second rate.

  9. Occurrence of Trypanosoma caninum in areas overlapping with leishmaniasis in Brazil: what is the real impact of canine leishmaniasis control?

    PubMed

    Barros, J H S; Almeida, A B P F; Figueiredo, F B; Sousa, V R F; Fagundes, A; Pinto, A G S; Baptista, C; Madeira, M F

    2012-07-01

    Trypanosoma caninum is a parasite of the Trypanosoma genus recently described in the natural infection of dogs in the municipality of Rio de Janeiro, Brazil. Suspecting the existence of a natural cycle and the circulation of this new species, the objective of this study was the taxonomic identification of samples of Trypanosoma spp. isolated from dogs in different Brazilian regions. Parasites were solely obtained from skin fragments culture and characterized by nested-PCR targeting the partial sequence of 18S rRNA gene and PCR products were sequenced. Thirty-three samples, obtained in São Paulo, Minas Gerais, Goiás, Mato Grosso and Rio de Janeiro states were analyzed. PCR and sequencing showed that the isolates were genetically identical or closely similar and confirmed T. caninum identity. This report broadens the geographical distribution of T. caninum in Brazil and discusses the impact of the presence of this parasite in areas of canine leishmaniasis occurrence.

  10. Variant surface glycoproteins from Venezuelan trypanosome isolates are recognized by sera from animals infected with either Trypanosoma evansi or Trypanosoma vivax.

    PubMed

    Camargo, Rocío; Izquier, Adriana; Uzcanga, Graciela L; Perrone, Trina; Acosta-Serrano, Alvaro; Carrasquel, Liomary; Arias, Laura P; Escalona, José L; Cardozo, Vanessa; Bubis, José

    2015-01-15

    Salivarian trypanosomes sequentially express only one variant surface glycoprotein (VSG) on their cell surface from a large repertoire of VSG genes. Seven cryopreserved animal trypanosome isolates known as TeAp-ElFrio01, TEVA1 (or TeAp-N/D1), TeGu-N/D1, TeAp-Mantecal01, TeGu-TerecayTrino, TeGu-Terecay03 and TeGu-Terecay323, which had been isolated from different hosts identified in several geographical areas of Venezuela were expanded using adult albino rats. Soluble forms of predominant VSGs expressed during the early infection stages were purified and corresponded to concanavalin A-binding proteins with molecular masses of 48-67 kDa by sodium dodecyl sulfate-polyacrylamide gel electropohoresis, and pI values between 6.1 and 7.5. The biochemical characterization of all purified soluble VSGs revealed that they were dimers in their native form and represented different gene products. Sequencing of some of these proteins yielded peptides homologous to VSGs from Trypanosoma (Trypanozoon) brucei and Trypanosoma (Trypanozoon) evansi and established that they most likely are mosaics generated by homologous recombination. Western blot analysis showed that all purified VSGs were cross-reacting antigens that were recognized by sera from animals infected with either T. evansi or Trypanosoma (Dutonella) vivax. The VSG glycosyl-phosphatidylinositol cross-reacting determinant epitope was only partially responsible for the cross-reactivity of the purified proteins, and antibodies appeared to recognize cross-reacting conformational epitopes from the various soluble VSGs. ELISA experiments were performed using infected bovine sera collected from cattle in a Venezuelan trypanosome-endemic area. In particular, soluble VSGs from two trypanosome isolates, TeGu-N/D1 and TeGu-TeracayTrino, were recognized by 93.38% and 73.55% of naturally T. vivax-infected bovine sera, respectively. However, approximately 70% of the sera samples did not recognize all seven purified proteins. Hence, the

  11. Variant surface glycoproteins from Venezuelan trypanosome isolates are recognized by sera from animals infected with either Trypanosoma evansi or Trypanosoma vivax

    PubMed Central

    Camargo, Rocío; Izquier, Adriana; Uzcanga, Graciela L.; Perrone, Trina; Acosta-Serrano, Alvaro; Carrasquel, Liomary; Arias, Laura P.; Escalona, José L.; Cardozo, Vanessa; Bubis, José

    2015-01-01

    Salivarian trypanosomes sequentially express only one variant surface glycoprotein (VSG) on their cell surface from a large repertoire of VSG genes. Seven cryopreserved animal trypanosome isolates known as TeAp-ElFrio01, TEVA1 (or TeAp-N/D1), TeGu-N/D1, TeAp-Mantecal01, TeGu-TerecayTrino, TeGu-Terecay03 and TeGu-Terecay323, which had been isolated from different hosts identified in several geographical areas of Venezuela were expanded using adult albino rats. Soluble forms of predominant VSGs expressed during the early infection stages were purified and corresponded to concanavalin A-binding proteins with molecular masses of 48–67 kDa by sodium dodecyl sulfate-polyacrylamide gel electropohoresis, and pI values between 6.1 and 7.5. The biochemical characterization of all purified soluble VSGs revealed that they were dimers in their native form and represented different gene products. Sequencing of some of these proteins yielded peptides homologous to VSGs from Trypanosoma (Trypanozoon) brucei and Trypanosoma (Trypanozoon) evansi and established that they most likely are mosaics generated by homologous recombination. Western blot analysis showed that all purified VSGs were cross-reacting antigens that were recognized by sera from animals infected with either T. evansi or Trypanosoma (Dutonella) vivax. The VSG glycosyl-phosphatidylinositol cross-reacting determinant epitope was only partially responsible for the cross-reactivity of the purified proteins, and antibodies appeared to recognize cross-reacting conformational epitopes from the various soluble VSGs. ELISA experiments were performed using infected bovine sera collected from cattle in a Venezuelan trypanosome-endemic area. In particular, soluble VSGs from two trypanosome isolates, TeGu-N/D1 and TeGu-TeracayTrino, were recognized by 93.38% and 73.55% of naturally T. vivax-infected bovine sera, respectively. However, approximately 70% of the sera samples did not recognize all seven purified proteins. Hence

  12. Origins of Chagas disease: Didelphis species are natural hosts of Trypanosoma cruzi I and armadillos hosts of Trypanosoma cruzi II, including hybrids.

    PubMed

    Yeo, Matthew; Acosta, Nidia; Llewellyn, Martin; Sánchez, Humberto; Adamson, Susie; Miles, Graham A J; López, Elsa; González, Nilsa; Patterson, James S; Gaunt, Michael W; de Arias, Antonieta Rojas; Miles, Michael A

    2005-02-01

    Trypanosoma cruzi, the causative agent of Chagas disease, has at least two principal intraspecific subdivisions, T. cruzi I (TCI) and T. cruzi II (TCII), the latter containing up to five subgroups (a-e). Whilst it is known that TCI predominates from the Amazon basin northwards and TCII to the South, where the disease is considered to be clinically more severe, the precise clinical and evolutionary significance of these divisions remains enigmatic. Here, we present compelling evidence of an association between TCI and opossums (Didelphis), and TCII and armadillos, on the basis of key new findings from the Paraguayan Chaco region, together with a comprehensive analysis of historical data. We suggest that the distinct arboreal and terrestrial ecologies, respectively, of these mammal hosts provide a persuasive explanation for the extant T. cruzi intraspecific diversity in South America, and for separate origins of Chagas disease in northern South America and in the southern cone countries.

  13. Trypanosoma brucei adenine-phosphoribosyltransferases mediate adenine salvage and aminopurinol susceptibility but not adenine toxicity.

    PubMed

    Lüscher, Alexandra; Lamprea-Burgunder, Estelle; Graf, Fabrice E; de Koning, Harry P; Mäser, Pascal

    2014-04-01

    African trypanosomes, like all obligate parasitic protozoa, cannot synthesize purines de novo and import purines from their hosts to build nucleic acids. The purine salvage pathways of Trypanosoma brucei being redundant, none of the involved enzymes is likely to be essential. Nevertheless they can be of pharmacological interest due to their role in activation of purine nucleobase or nucleoside analogues, which only become toxic when converted to nucleotides. Aminopurine antimetabolites, in particular, are potent trypanocides and even adenine itself is toxic to trypanosomes at elevated concentrations. Here we report on the T. brucei adenine phosphoribosyltransferases TbAPRT1 and TbAPRT2, encoded by the two genes Tb927.7.1780 and Tb927.7.1790, located in tandem on chromosome seven. The duplication is syntenic in all available Trypanosoma genomes but not in Leishmania. While TbAPRT1 is cytosolic, TbAPRT2 possesses a glycosomal targeting signal and co-localizes with the glycosomal marker aldolase. Interestingly, the distribution of glycosomal targeting signals among trypanosomatid adenine phosphoribosyltransferases is not consistent with their phylogeny, indicating that the acquisition of adenine salvage to the glycosome happened after the radiation of Trypanosoma. Double null mutant T. brucei Δtbaprt1,2 exhibited no growth phenotype but no longer incorporated exogenous adenine into the nucleotide pool. This, however, did not reduce their sensitivity to adenine. The Δtbaprt1,2 trypanosomes were resistant to the adenine isomer aminopurinol, indicating that it is activated by phosphoribosyl transfer. Aminopurinol was about 1000-fold more toxic to bloodstream-form T. brucei than the corresponding hypoxanthine isomer allopurinol. Aminopurinol uptake was not dependent on the aminopurine permease P2 that has been implicated in drug resistance.

  14. Mode of Action of the Sesquiterpene Lactones Psilostachyin and Psilostachyin C on Trypanosoma cruzi

    PubMed Central

    Papademetrio, Daniela; Batlle, Alcira; Martino, Virginia S.; Frank, Fernanda M.; Lombardo, María E.

    2016-01-01

    Trypanosoma cruzi is the causative agent of Chagas’ disease, which is a major endemic disease in Latin America and is recognized by the WHO as one of the 17 neglected tropical diseases in the world. Psilostachyin and psilostachyin C, two sesquiterpene lactones isolated from Ambrosia spp., have been demonstrated to have trypanocidal activity. Considering both the potential therapeutic targets present in the parasite, and the several mechanisms of action proposed for sesquiterpene lactones, the aim of this work was to characterize the mode of action of psilostachyin and psilostachyin C on Trypanosoma cruzi and to identify the possible targets for these molecules. Psilostachyin and psilostachyin C were isolated from Ambrosia tenuifolia and Ambrosia scabra, respectively. Interaction of sesquiterpene lactones with hemin, the induction of oxidative stress, the inhibition of cruzipain and trypanothione reductase and their ability to inhibit sterol biosynthesis were evaluated. The induction of cell death by apoptosis was also evaluated by analyzing phosphatidylserine exposure detected using annexin-V/propidium iodide, decreased mitochondrial membrane potential, assessed with Rhodamine 123 and nuclear DNA fragmentation evaluated by the TUNEL assay. Both STLs were capable of interacting with hemin. Psilostachyin increased about 5 times the generation of reactive oxygen species in Trypanosoma cruzi after a 4h treatment, unlike psilostachyin C which induced an increase in reactive oxygen species levels of only 1.5 times. Only psilostachyin C was able to inhibit the biosynthesis of ergosterol, causing an accumulation of squalene. Both sesquiterpene lactones induced parasite death by apoptosis. Upon evaluating the combination of both compounds, and additive trypanocidal effect was observed. Despite their structural similarity, both sesquiterpene lactones exerted their anti-T. cruzi activity through interaction with different targets. Psilostachyin accomplished its

  15. Subcellular localization of glycolytic enzymes and characterization of intermediary metabolism of Trypanosoma rangeli.

    PubMed

    Rondón-Mercado, Rocío; Acosta, Héctor; Cáceres, Ana J; Quiñones, Wilfredo; Concepción, Juan Luis

    2017-09-01

    Trypanosoma rangeli is a hemoflagellate protist that infects wild and domestic mammals as well as humans in Central and South America. Although this parasite is not pathogenic for human, it is being studied because it shares with Trypanosoma cruzi, the etiological agent of Chagas' disease, biological characteristics, geographic distribution, vectors and vertebrate hosts. Several metabolic studies have been performed with T. cruzi epimastigotes, however little is known about the metabolism of T. rangeli. In this work we present the subcellular distribution of the T. rangeli enzymes responsible for the conversion of glucose to pyruvate, as determined by epifluorescense immunomicroscopy and subcellular fractionation involving either selective membrane permeabilization with digitonin or differential and isopycnic centrifugation. We found that in T. rangeli epimastigotes the first six enzymes of the glycolytic pathway, involved in the conversion of glucose to 1,3-bisphosphoglycerate are located within glycosomes, while the last four steps occur in the cytosol. In contrast with T. cruzi, where three isoenzymes (one cytosolic and two glycosomal) of phosphoglycerate kinase are expressed simultaneously, only one enzyme with this activity is detected in T. rangeli epimastigotes, in the cytosol. Consistent with this latter result, we found enzymes involved in auxiliary pathways to glycolysis needed to maintain adenine nucleotide and redox balances within glycosomes such as phosphoenolpyruvate carboxykinase, malate dehydrogenase, fumarate reductase, pyruvate phosphate dikinase and glycerol-3-phosphate dehydrogenase. Glucokinase, galactokinase and the first enzyme of the pentose-phosphate pathway, glucose-6-phosphate dehydrogenase, were also located inside glycosomes. Furthermore, we demonstrate that T. rangeli epimastigotes growing in LIT medium only consume glucose and do not excrete ammonium; moreover, they are unable to survive in partially-depleted glucose medium. The

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

    PubMed

    Planer, Joseph D; Hulverson, Matthew A; Arif, Jennifer A; Ranade, Ranae M; Don, Robert; Buckner, Frederick S

    2014-07-01

    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.

  17. Mode of Action of the Sesquiterpene Lactones Psilostachyin and Psilostachyin C on Trypanosoma cruzi.

    PubMed

    Sülsen, Valeria P; Puente, Vanesa; Papademetrio, Daniela; Batlle, Alcira; Martino, Virginia S; Frank, Fernanda M; Lombardo, María E

    2016-01-01

    Trypanosoma cruzi is the causative agent of Chagas' disease, which is a major endemic disease in Latin America and is recognized by the WHO as one of the 17 neglected tropical diseases in the world. Psilostachyin and psilostachyin C, two sesquiterpene lactones isolated from Ambrosia spp., have been demonstrated to have trypanocidal activity. Considering both the potential therapeutic targets present in the parasite, and the several mechanisms of action proposed for sesquiterpene lactones, the aim of this work was to characterize the mode of action of psilostachyin and psilostachyin C on Trypanosoma cruzi and to identify the possible targets for these molecules. Psilostachyin and psilostachyin C were isolated from Ambrosia tenuifolia and Ambrosia scabra, respectively. Interaction of sesquiterpene lactones with hemin, the induction of oxidative stress, the inhibition of cruzipain and trypanothione reductase and their ability to inhibit sterol biosynthesis were evaluated. The induction of cell death by apoptosis was also evaluated by analyzing phosphatidylserine exposure detected using annexin-V/propidium iodide, decreased mitochondrial membrane potential, assessed with Rhodamine 123 and nuclear DNA fragmentation evaluated by the TUNEL assay. Both STLs were capable of interacting with hemin. Psilostachyin increased about 5 times the generation of reactive oxygen species in Trypanosoma cruzi after a 4h treatment, unlike psilostachyin C which induced an increase in reactive oxygen species levels of only 1.5 times. Only psilostachyin C was able to inhibit the biosynthesis of ergosterol, causing an accumulation of squalene. Both sesquiterpene lactones induced parasite death by apoptosis. Upon evaluating the combination of both compounds, and additive trypanocidal effect was observed. Despite their structural similarity, both sesquiterpene lactones exerted their anti-T. cruzi activity through interaction with different targets. Psilostachyin accomplished its antiparasitic

  18. Vector-borne transmission of Trypanosoma cruzi among captive Neotropical primates in a Brazilian zoo.

    PubMed

    Minuzzi-Souza, Thaís Tâmara Castro; Nitz, Nadjar; Knox, Monique Britto; Reis, Filipe; Hagström, Luciana; Cuba, César A Cuba; Hecht, Mariana Machado; Gurgel-Gonçalves, Rodrigo

    2016-01-26

    Neotropical primates are important sylvatic hosts of Trypanosoma cruzi, the etiological agent of Chagas disease. Infection is often subclinical, but severe disease has been described in both free-ranging and captive primates. Panstrongylus megistus, a major T. cruzi vector, was found infesting a small-primate unit at Brasília zoo (ZooB), Brazil. ZooB lies close to a gallery-forest patch where T. cruzi circulates naturally. Here, we combine parasitological and molecular methods to investigate a focus of T. cruzi infection involving triatomine bugs and Neotropical primates at a zoo located in the Brazilian Savannah. We assessed T. cruzi infection in vectors using optical microscopy (n = 34) and nested PCR (n = 50). We used quantitative PCR (qPCR) to examine blood samples from 26 primates and necropsy samples from two primates that died during the study. We determined parasite lineages in five vectors and two primates by comparing glucose-6-phosphate isomerase (G6pi) gene sequences. Trypanosoma cruzi was found in 44 vectors and 17 primates (six genera and eight species); one Mico chrysoleucus and one Saguinus niger had high parasitaemias. Trypanosoma cruzi DNA was detected in three primates born to qPCR-negative mothers at ZooB and in the two dead specimens. One Callithrix geoffroyi became qPCR-positive over a two-year follow-up. All G6pi sequences matched T. cruzi lineage TcI. Our findings strongly suggest vector-borne T. cruzi transmission within a small-primate unit at ZooB - with vectors, and perhaps also parasites, presumably coming from nearby gallery forest. Periodic checks for vectors and parasites would help eliminate T. cruzi transmission foci in captive-animal facilities. This should be of special importance for captive-breeding programs involving endangered mammals, and would reduce the risk of accidental T. cruzi transmission to keepers and veterinarians.

  19. Erythrina abyssinica prevents meningoencephalitis in chronic Trypanosoma brucei brucei mouse model.

    PubMed

    Nasimolo, Johnson; Kiama, Stephen Gitahi; Gathumbi, Peter Karuri; Makanya, Andrew Ndegwa; Kagira, John Maina

    2014-06-01

    Human African trypanosomiasis is prevalent in Sub-sahara African countries that lie between 14° North and 29° south of the equator. Sixty million people are at risk of infection. Trypanosoma brucei gambesience occurs in West and Central Africa while Trypanosoma brucei rhodesience occurs in East and Southern Africa. The neurological stage of the disease is characterized by neuroinflammation. About 10% of patients treated with the recommended drug, melarsoprol develop post treatment reactive encephalopathy, which is fatal in 50% of these patients, thus melarsoprol is fatal in 5% of all treated patients. This study was aimed at establishing the potential activity of Erythrina abyssinica in reducing neuroinflammation following infection with Trypanosoma brucei brucei. Swiss white mice were divided into ten groups, two control groups and eight infected groups. Infected mice received either methanol or water extract of Erythrina abyssinica at 12.5, 25, 50 or 100 mg/kg body weight. Parasite counts were monitored in peripheral circulation from the third day post infection up to the end of the study. Brains were processed for histology, immunohistochemistry scanning and transmission electron microscopy. Following infection, trypanosomes were observed in circulation 3 days post-infection, with the parasitaemia occurring in waves. In the cerebrum, typical brain pathology of chronic trypanosomiasis was reproduced. This was exhibited as astrocytosis, perivascular cuffing and infiltration of inflammatory cells into the neuropil. However, mice treated with Erythrina abyssinica water extract exhibited significant reduction in perivascular cuffing, lymphocytic infiltration and astrocytosis in the cerebrum. The methanol extract did not have a significant difference compared to the non-treated group. This study provides evidence of anti-inflammatory properties of Erythrina abyssinica and may support its wide use as a medicinal plant by various communities in Kenya.

  20. Role of cytokines in Trypanosoma brucei-induced anaemia: A review of the literature.

    PubMed

    Musaya, J; Matovu, E; Nyirenda, M; Chisi, J

    2015-06-01

    Anaemia is an important complication of trypanosomiasis. The mechanisms through which trypanosomal infection leads to anaemia are poorly defined. A number of studies have implicated inflammatory cytokines, but these data are limited and inconsistent. In this article, we reviewed the published literature on cytokines associated with Trypanosoma brucei infections and their role in the immunopathology leading to anaemia. Articles were searched in PubMed through screening of titles and abstracts with no limitation on date of publishing and study design. Articles in English were searched using keywords "African trypanosomiasis", "sleeping sickness", "Trypanosoma brucei", in all possible combinations with "anaemia" and/or "cytokines". Twelve articles examining cytokines and their role in trypanosomeinduced anaemia were identified out of 1095 originally retrieved from PubMed. None of the articles identified were from human-based studies. A total of eight cytokines were implicated, with four cytokines (IFN-γ, IL-10, TNF-α, IL-12) showing an association with anaemia. These articles reported that mice lacking TNF-α were able to control anaemia, and that IFN-γ was linked to severe anaemia given its capacity to suppress erythropoiesis, while IL-10 was shown to regulate IFN-γ and TNF-α, providing a balance that was associated with severity of anaemia. IFN-γ and TNF-α have also been reported to work in concert with other factors such as nitric oxide and iron in order to induce anaemia. IFN-γ, IL-10, and TNF-α were the three major cytokines identified to be heavily involved in anaemia caused by Trypanosoma brucei infection. The anti-inflammatory cytokine, IL-10, was shown to counter the effects of proinflammatory cytokines in order to balance the severity of anaemia. The mechanism of anaemia is multifactorial and therefore requires further, more elaborate research. Data from human subjects would also shed more light.

  1. Micro RNA expression profiles in peripheral blood cells of rats that were experimentally infected with Trypanosoma congolense and different Trypanosoma brucei subspecies.

    PubMed

    Simo, Gustave; Lueong, Smiths; Grebaut, Pascal; Guny, Gerard; Hoheisel, Jörg D

    2015-08-01

    To identify miRNAs whose expression are differentially regulated during trypanosome infections a microarray targeting more than 600 rat miRNA was used to analyze the miRNA expression profiles between uninfected rats and animals infected by Trypanosoma congolense and Trypanosoma brucei s.l. The potential targets of dysregulated miRNAs as well as their biological pathways and functions were predicted using several bioinformatics software tools. Irrespective of the infecting trypanosome species, eight miRNAs (seven up- and one down-regulated) were dysregulated during infections. Moreover, other miRNAs were differentially regulated in rats infected by specific trypanosome species. Functional analyses of differentially regulated miRNAs indicated their involvement in diverse biological processes. Among these, transcription repressor activity, gene expression control as well as protein transporter activity were predominant. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis of dysregulated miRNAs revealed their involvement in several biological pathways and disease conditions. This suggests possible modulation of such pathways following trypanosome infection; for example, the MAPK signaling pathway which is known to play vital roles in apoptosis, innate immune response and response to viral infections was highly affected. Axon guidance was equally highly impacted and may indicate a cross reactivity between pathogen proteins and guidance molecules representing one pathological mechanism as it has been observed with influenza HA. Furthermore, Ingenuity pathway analyses of dysregulated miRNAs and potential targets indicated strong association with inflammatory responses, cell death and survival as well as infectious diseases. The data generated here provide valuable information to understand the regulatory function of miRNAs during trypanosome infections. They improved our knowledge on host-parasite cross-talks and provide a framework for investigations to

  2. Trypanosoma cruzi: effect and mode of action of nitroimidazole and nitrofuran derivatives.

    PubMed

    Maya, Juan Diego; Bollo, Soledad; Nuñez-Vergara, Luis J; Squella, Juan A; Repetto, Yolanda; Morello, Antonio; Périé, Jacques; Chauvière, Gérard

    2003-03-15

    With the aim of determining the actual target(s) of nitro-group bearing compounds considered as possible leads for the development of drugs against Chagas' disease, we studied in parallel nitrofurans and nitroimidazoles. We investigated nine representative compounds for the following properties: efficacy on different Trypanosoma cruzi strains, redox cyclers, inhibition of respiration, production of corresponding nitroso derivatives and intracellular thiol scavengers. Our results indicate that nifurtimox and related compounds act as redox cyclers, whereas the most active in the series, the 5-nitroimidazole megazol essentially acts as thiol scavenger particularly for trypanothione, the cofactor for trypanothione reductase, an essential enzyme in the detoxification process.

  3. Trypanothione Reductase: A Target for the Development of Anti-Trypanosoma cruzi drugs.

    PubMed

    Vázquez, Karina; Paulino, Margot; Salas, Cristian O; Zarate-Ramos, Juan J; Vera, Brenda; Rivera, Gildardo

    2017-03-15

    Chagas disease or American trypanosomiasis is a major parasitic disease in Latin America with treatment available via two drugs: nifurtimox and benznidazole. These two treatments are ineffective in the chronic phase of the disease. Therefore, there is a need for the development of new, efficient and safe drugs for the treatment of these diseases. With this goal, one of the promising targets proposed is the trypanothione reductase (TR), a key enzyme important in the metabolism of Trypanosoma cruzi. In this review, we analyze the importance of TR as a drug target, as well as their compounds inhibitors reported in the last decade as potential therapeutic agents for Chagas disease.

  4. High Trypanosoma cruzi (Kinetoplastida: Trypanosomatidae) Prevalence in Triatoma sanguisuga (Hemiptera: Redviidae) in Southeastern Louisiana

    PubMed Central

    CESA, K.; CAILLOUËT, K. A.; DORN, P. L.; WESSON, D. M.

    2012-01-01

    From May through November 2007, intensive weekly surveys at the site of a previously reported autochthonous human case of Chagas parasite infection resulted in the collection of 298 Triatoma sanguisuga (Leconte) specimens, of which 60.4% (180) were polymerase chain reaction positive for Trypanosoma cruzi Chagas. All were adults, in a ratio of ≈ 1:1 female to male, indicating that the domicile was not colonized, but was a destination for these host-seeking adults. We report on seasonal activity pattern, T. cruzi prevalence in T. sanguisuga, and attempts at insect exclusion and control at the case residence. PMID:21936329

  5. Influence of electrolytes and non-electrolytes on growth and differentiation of Trypanosoma cruzi.

    PubMed

    Osuna, A; Adroher, F J; Lupiáñez, J A

    1990-05-01

    The influence of electrolytes and non-electrolytes, especially NaCl and sorbitol, on the metacyclogenesis and growth of Trypanosoma cruzi has been studied. The addition of 50 or 100 mEq/l NaCl to the culture media significantly increased the development of metacyclic forms. Other electrolytes and non-electrolytes had no effect on epimastigote-metacyclic differentiation. The growth rate was never modified to any extent. The influence of sodium concentration, osmotic pressure, among other factors, are discussed. Electrophoresis showed proteins bands which could be related either to the adaptation of T. cruzi to the new culture media or to the initiation of differentiation processes.

  6. Long-term preservation of blood samples for diagnosis of Trypanosoma cruzi infection.

    PubMed

    Pérez, A C; Cura, E; Subías, E; Lansetti, J C; Segura, E L

    1990-03-01

    Feasability and suitability for field research of a whole-blood preservation method was evaluated through the screening of anti-Trypanosoma cruzi antibodies in 1209 samples under different conditions. Antibody reactivity of paired samples from preserved capillary blood (CBP) and sera from venous blood (VBS) were studied by specific techniques. Over 96% concordance was found on indoor studies carried out with samples without storage or after 15 or 30 days preservation of CBP at 37 degrees C and VBS at -20 degrees C. Outdoor studies performed at field conditions, achieved a 92.1% concordance.

  7. Membrane traffic and synaptic cross-talk during host cell entry by Trypanosoma cruzi

    PubMed Central

    Butler, Claire E; Tyler, Kevin M

    2012-01-01

    It is widely accepted that Trypanosoma cruzi can exploit the natural exocytic response of the host to cell damage, utilizing host cell lysosomes as important effectors. It is, though, increasingly clear that the parasite also exploits endocytic mechanisms which allow for incorporation of plasma membrane into the parasitophorous vacuole. Further, that these endocytic mechanisms are involved in cross-talk with the exocytic machinery, in the recycling of vesicles and in the manipulation of the cytoskeleton. Here we review the mechanisms by which T. cruzi exploits features of the exocytic and endocytic pathways in epithelial and endothelial cells and the evidence for cross-talk between these pathways. PMID:22646288

  8. Metacaspase 2 of Trypanosoma brucei is a calcium-dependent cysteine peptidase active without processing.

    PubMed

    Moss, Catherine X; Westrop, Gareth D; Juliano, Luiz; Coombs, Graham H; Mottram, Jeremy C

    2007-12-11

    Metacaspases are cysteine peptidases that are distantly related to the caspases, for which proteolytic processing is central to their activation. Here, we show that recombinant metacaspase 2 (MCA2) from Trypanosoma brucei has arginine/lysine-specific, Ca(2+)-dependent proteolytic activity. Autocatalytic processing of MCA2 occurred after Lys55 and Lys268; however, this was shown not to be required for the enzyme to be proteolytically active. The necessity of Ca(2+), but not processing, for MCA2 enzymatic activity clearly distinguishes MCA2 from the caspases and would be consistent with different physiological roles.

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

    PubMed Central

    Zingales, Bianca; Miles, Michael A; Moraes, Carolina B; Luquetti, Alejandro; Guhl, Felipe; Schijman, Alejandro G; Ribeiro, Isabela

    2014-01-01

    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. PMID:25317712

  10. 2-Pyridyl thiazoles as novel anti-Trypanosoma cruzi agents: structural design, synthesis and pharmacological evaluation.

    PubMed

    Cardoso, Marcos Veríssimo de Oliveira; de Siqueira, Lucianna Rabelo Pessoa; da Silva, Elany Barbosa; Costa, Lívia Bandeira; Hernandes, Marcelo Zaldini; Rabello, Marcelo Montenegro; Ferreira, Rafaela Salgado; da Cruz, Luana Faria; Moreira, Diogo Rodrigo Magalhães; Pereira, Valéria Rêgo Alves; de Castro, Maria Carolina Accioly Brelaz; Bernhardt, Paul V; Leite, Ana Cristina Lima

    2014-10-30

    The present work reports on the synthesis, anti-Trypanosoma cruzi activities and docking studies of a novel series of 2-(pyridin-2-yl)-1,3-thiazoles derived from 2-pyridine thiosemicarbazone. The majority of these compounds are potent cruzain inhibitors and showed excellent inhibition on the trypomastigote form of the parasite, and the resulting structure-activity relationships are discussed. Together, these data present a novel series of thiazolyl hydrazones with potential effects against Chagas disease and they could be important leads in continuing development against Chagas disease. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  11. Trypanosoma vivax: a simplified protocol for in vivo growth, isolation and cryopreservation.

    PubMed

    Ndao, M; Magnus, E; Büscher, P; Geerts, S

    2004-03-01

    A rodent adapted clone of Trypanosoma vivax was used to infect cyclophosphamide treated mice and rats. Fresh blood containing trypanosomes, was centrifuged in a density gradient of three Percoll solutions, 1.07, 1.06, 1.05 g/ml, respectively, carefully layered on top of each other. The yields of this simple procedure for trypanosome purification were about six times higher than those obtained with the conventional anion-exchange columns. Cryopreservation of trypanosomes using glycerol yielded 90% viable parasites, whereas using dimethylsulfoxide, a more commonly used cryoprotectant, the viability was only 35%.

  12. High Trypanosoma cruzi (Kinetoplastida: Trypanosomatidae) prevalence in Triatoma sanguisuga (Hemiptera: Redviidae) in southeastern Louisiana.

    PubMed

    Cesa, K; Caillouët, K A; Dorn, P L; Wesson, D M

    2011-09-01

    From May through November 2007, intensive weekly surveys at the site of a previously reported autochthonous human case of Chagas parasite infection resulted in the collection of 298 Triatoma sanguisuga (Leconte) specimens, of which 60.4% (180) were polymerase chain reaction positive for Trypanosoma cruzi Chagas. All were adults, in a ratio of approximately 1:1 female to male, indicating that the domicile was not colonized, but was a destination for these host-seeking adults. We report on seasonal activity pattern, T. cruzi prevalence in T. sanguisuga, and attempts at insect exclusion and control at the case residence.

  13. Structures of aspartate aminotransferases from Trypanosoma brucei, Leishmania major and Giardia lamblia

    PubMed Central

    Abendroth, Jan; Choi, Ryan; Wall, Abigail; Clifton, Matthew C.; Lukacs, Christine M.; Staker, Bart L.; Van Voorhis, Wesley; Myler, Peter; Lorimer, Don D.; Edwards, Thomas E.

    2015-01-01

    The structures of three aspartate aminotransferases (AATs) from eukaryotic pathogens were solved within the Seattle Structural Genomics Center for Infectious Disease (SSGCID). Both the open and closed conformations of AAT were observed. Pyridoxal phosphate was bound to the active site via a Schiff base to a conserved lysine. An active-site mutant showed that Trypanosoma brucei AAT still binds pyridoxal phosphate even in the absence of the tethering lysine. The structures highlight the challenges for the structure-based design of inhibitors targeting the active site, while showing options for inhibitor design targeting the N-terminal arm. PMID:25945710

  14. Structures of aspartate aminotransferases from Trypanosoma brucei, Leishmania major and Giardia lamblia.

    PubMed

    Abendroth, Jan; Choi, Ryan; Wall, Abigail; Clifton, Matthew C; Lukacs, Christine M; Staker, Bart L; Van Voorhis, Wesley; Myler, Peter; Lorimer, Don D; Edwards, Thomas E

    2015-05-01

    The structures of three aspartate aminotransferases (AATs) from eukaryotic pathogens were solved within the Seattle Structural Genomics Center for Infectious Disease (SSGCID). Both the open and closed conformations of AAT were observed. Pyridoxal phosphate was bound to the active site via a Schiff base to a conserved lysine. An active-site mutant showed that Trypanosoma brucei AAT still binds pyridoxal phosphate even in the absence of the tethering lysine. The structures highlight the challenges for the structure-based design of inhibitors targeting the active site, while showing options for inhibitor design targeting the N-terminal arm.

  15. Synthesis, cytotoxicity, and anti-Trypanosoma cruzi activity of new chalcones.

    PubMed

    Aponte, José C; Verástegui, Manuela; Málaga, Edith; Zimic, Mirko; Quiliano, Miguel; Vaisberg, Abraham J; Gilman, Robert H; Hammond, Gerald B

    2008-10-09

    Synthesis of a cytotoxic dihydrochalcone, first isolated from a traditional Amazonian medicinal plant Iryanthera juruensis Warb (Myristicaceae), followed by a comprehensive SAR analysis of saturated and unsaturated chalcone synthetic intermediates, led to the identification of analogues with selective and significant in vitro anti- Trypanosoma cruzi activity. Further SAR studies were undertaken with the synthesis of 21 new chalcones containing two allyloxy moieties that resulted in the discovery of 2',4'-diallyloxy-6'-methoxy chalcones with improved selectivity against this parasite at concentrations below 25 microM, four of which exhibited a selectivity index greater than 12.

  16. Evidence for the existence of an Ns-type regulatory protein in Trypanosoma cruzi membranes.

    PubMed Central

    Eisenschlos, C D; Paladini, A A; Molina y Vedia, L; Torres, H N; Flawiá, M M

    1986-01-01

    The existence of a GTP-binding protein of the Ns type in Trypanosoma cruzi was explored. Epimastigote membranes were labelled by cholera toxin in the presence of [adenine-14C]NAD+. After SDS/polyacrylamide-gel electrophoresis of extracted membrane proteins, a single labelled polypeptide band of apparent Mr approx. 45,000 was detected. Epimastigote cells were treated with N-ethylmaleimide and electrofused to lymphoma S49 cells lacking the Ns protein. Evidence indicates that in such electrofusion-generated cell hybrids a heterologous adenylate cyclase system was reconstituted with the Ns protein provided by T. cruzi epimastigotes. Images Fig. 2. PMID:3099761

  17. Active penetration of Trypanosoma cruzi into host cells: historical considerations and current concepts

    PubMed Central

    de Souza, Wanderley; de Carvalho, Tecia M. Ulisses

    2013-01-01

    In the present short review, we analyze past experiments that addressed the interactions of intracellular pathogenic protozoa (Trypanosoma cruzi, Toxoplasma gondii, and Plasmodium) with host cells and the initial use of the term active penetration to indicate that a protozoan “crossed the host cell membrane, penetrating into the cytoplasm.” However, the subsequent use of transmission electron microscopy showed that, for all of the protozoans and cell types examined, endocytosis, classically defined as involving the formation of a membrane-bound vacuole, took place during the interaction process. As a consequence, the recently penetrated parasites are always within a vacuole, designated the parasitophorous vacuole (PV). PMID:23355838

  18. Cross-resistance associated with development of resistance to isometamidium in a clone of Trypanosoma congolense.

    PubMed Central

    Peregrine, A S; Gray, M A; Moloo, S K

    1997-01-01

    Resistance to isometamidium was increased 94-fold in a clone of Trypanosoma congolense (clone IL 1180) by repeated subcurative treatment of infected mice for 11 months. This was associated with 3.4-, 33-, and 4.2-fold increases in resistance to diminazene, homidium, and quinapyramine, respectively. Both T. congolense IL 1180 and the resistant derivative were able to undergo cyclical development in Glossina morsitans centralis tsetse flies, producing hypopharyngeal infection rates of 40.0 and 39.8%, respectively. PMID:9210695

  19. 5-Nitro-2-furyl derivative actives against Trypanosoma cruzi: preliminary in vivo studies.

    PubMed

    Cabrera, Eliana; Murguiondo, Mariana González; Arias, Marelina González; Arredondo, Carolina; Pintos, Cristina; Aguirre, Gabriela; Fernández, Marcelo; Basmadjián, Yester; Rosa, Raquel; Pacheco, José Pedro; Raymondo, Stella; Di Maio, Rossanna; González, Mercedes; Cerecetto, Hugo

    2009-10-01

    Ten 5-nitro-2-furyl derivatives, with good to excellent in vitro anti-Trypanosoma cruzi activity, and nifurtimox were tested oral and intraperitoneally on healthy animals for its acute toxicity on murine models. According to animals' survival percentage, organ histological results, biochemical and haematological findings, three new derivatives, with toxicity like nifurtimox, were selected to test in vivo as antichagasic agents. Clearly, dependences between chemical structure and both acute toxicity and in vivo anti-T. cruzi activity were observed. 4-Hexyl-1-[3-(5-nitro-2-furyl)-2-propenylidene]semicarbazide displayed good profile as anti-T. cruzi agent and better acute toxicity profile than nifurtimox.

  20. Selective delivery of 2-hydroxy APA to Trypanosoma brucei using the melamine motif

    PubMed Central

    Klee, Nina; Wong, Pui Ee; Baragaña, Beatriz; Mazouni, Farah El; Phillips, Margaret A.; Barrett, Michael P.; Gilbert, Ian H.

    2010-01-01

    Trypanosoma brucei, the parasite that causes human African trypanosomiasis, is auxotrophic for purines and has specialist nucleoside transporters to import these metabolites. In particular, the P2 aminopurine transporter can also selectively accumulate melamine derivatives. In this Letter, we report the coupling of the melamine moiety to 2-hydroxy APA, a potent ornithine decarboxylase inhibitor, with the aim of selectively delivering this compound to the parasite. The best compound described here shows an increased in vitro trypanocidal activity compared with the parent. PMID:20615694

  1. Trypanosoma cruzi: TcRAB7 protein is localized at the Golgi apparatus in epimastigotes.

    PubMed

    Araripe, Júlia R; Cunha e Silva, Narcisa L; Leal, Simone T; de Souza, Wanderley; Rondinelli, Edson

    2004-08-20

    In mammalian cells, the Rab7 protein is a key element of late endocytic membrane traffic. Several results suggest that it is involved in the transport from early to late endosome or from late endosome to lysosome. We have previously characterized a Rab7 gene homologue (TcRAB7) in Trypanosoma cruzi. Now, using an affinity-purified antibody specific to TcRAB7 protein we have determined that it is localized at the Golgi apparatus of the parasite. Our results indicate that the T. cruzi Rab7 homologue may function in a different route than its counterparts in mammalian cells.

  2. Studies on the virulence and attenuation of Trypanosoma cruzi using immunodeficient animals.

    PubMed

    Basombrío, M A; Segura, M A; Gómez, L; Padilla, M

    2000-01-01

    Tissue invasion and pathology by Trypanosoma cruzi result from an interaction between parasite virulence and host immunity. Successive in vivo generations of the parasite select populations with increasing ability to invade the host. Conversely, prolonged in vitro selection of the parasite produces attenuated sublines with low infectivity for mammals. One such subline (TCC clone) has been extensively used in our laboratory as experimental vaccine and tested in comparative experiments with its virulent ancestor (TUL). The experiments here reviewed aimed at the use of immunodeficient mice for testing the infectivity of TCC parasites. It has not been possible to obtain virulent, revertant sublines by prolonged passaged in such mice.

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

    PubMed

    Zingales, Bianca; Miles, Michael A; Moraes, Carolina B; Luquetti, Alejandro; Guhl, Felipe; Schijman, Alejandro G; Ribeiro, Isabela

    2014-09-01

    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.

  4. Evolutionary insights from bat trypanosomes: morphological, developmental and phylogenetic evidence of a new species, Trypanosoma (Schizotrypanum) erneyi sp. nov., in African bats closely related to Trypanosoma (Schizotrypanum) cruzi and allied species.

    PubMed

    Lima, Luciana; Silva, Flávia Maia da; Neves, Luis; Attias, Márcia; Takata, Carmen S A; Campaner, Marta; de Souza, Wanderley; Hamilton, Patrick B; Teixeira, Marta M G

    2012-11-01

    Parasites of the genus Trypanosoma are common in bats and those of the subgenus Schizotrypanum are restricted to bats throughout the world, with the exception of Trypanosoma (Schizotrypanum) cruzi that also infects other mammals and is restricted to the American Continent. We have characterized trypanosome isolates from Molossidae bats captured in Mozambique, Africa. Morphology and behaviour in culture, supported by phylogenetic inferences using SSU (small subunit) rRNA, gGAPDH (glycosomal glyceraldehyde 3-phosphate dehydrogenase) and Cyt b (cytochrome b) genes, allowed to classify the isolates as a new Schizotrypanum species named Trypanosoma (Schizotrypanum) erneyi sp. nov. This is the first report of a Schizotrypanum species from African bats cultured, characterized morphologically and biologically, and positioned in phylogenetic trees. The unprecedented finding of a new species of the subgenus Schizotrypanum from Africa that is closest related to the America-restricted Trypanosoma (Schizotrypanum) cruzi marinkellei and T. cruzi provides new insights into the origin and evolutionary history of T. cruzi and closely related bat trypanosomes. Altogether, data from our study support the hypothesis of an ancestor trypanosome parasite of bats evolving to infect other mammals, even humans, and adapted to transmission by triatomine bugs in the evolutionary history of T. cruzi in the New World. Copyright © 2012 Elsevier GmbH. All rights reserved.

  5. Morphological forms of Trypanosoma evansi from blood of Arabian camel (Camelus dromedarius) in the Riyadh metropolitan areas.

    PubMed

    Amoudi, Mikky A; Al-Yousif, Mohamed S; Al-Shawa, Yaser

    2011-04-01

    Trypanosoma evansi is commonly referred to as haemoflagellate infesting camels and other vertebrates. It is causative agent of number of diseases. T. evansi has several morphological forms. We have detected three forms in the blood of camels, slender, transitional and intermediate. The present study is the first investigation on the morphological forms of T. evansi in Arabian camels in Saudi Arabia.

  6. Screening North American plant extracts in vitro against Trypanosoma brucei, the causative agent for Human African Trypanosomiasis

    USDA-ARS?s Scientific Manuscript database

    Natural products extracts from 522 plants collected from different parts of the North America were screened in vitro against trypamastigote forms of Trypanosoma brucei. The active extracts(150)with >90% inhibition at 20ug/mL concentrations from the plants namely, Alnus rubra, Hoita macrostachya, S...

  7. Molecular survey of rodent-borne Trypanosoma in Niger with special emphasis on T. lewisi imported by invasive black rats.

    PubMed

    Dobigny, Gauthier; Poirier, Philippe; Hima, Karmadine; Cabaret, Odile; Gauthier, Philippe; Tatard, Caroline; Costa, Jean-Marc; Bretagne, Stéphane

    2011-03-01

    Invading rodent species can harbor parasites with potential transmission to native rodents and/or humans. To investigate trypanosomes prevalence in rodents, the spleen of 76 rodents from Niger identified by their karyotype was used as a DNA source for Trypanosoma detection using a newly developed qPCR assay. Of the invasive black rat, Rattus rattus, 71% (10/14) were PCR positive as well as 6% (4/62) of native African rodents. Sequences of ~400bp of the SSU rDNA gene identified phylogenetically close Trypanosoma lineages. Trypanosoma lewisi was present in all positive black rats and the sequences displayed 100% similarity with T. lewisi-infected humans in Senegal. T. lewisi was also detected in one Acomys johannis, suggesting a possible transmission to native species. In addition to improved knowledge of Trypanosoma diversity in rodents, our data underscore the introduction of the potentially pathogenic T. lewisi kinetoplastid through the human-mediated invasion of black rats all over West Africa.

  8. Rodent-borne Trypanosoma from cities and villages of Niger and Nigeria: A special role for the invasive genus Rattus?

    PubMed

    Tatard, C; Garba, M; Gauthier, P; Hima, K; Artige, E; Dossou, D K H J; Gagaré, S; Genson, G; Truc, P; Dobigny, G

    2017-07-01

    Although they are known to sometimes infect humans, atypical trypanosomes are very poorly documented, especially in Africa where one lethal case has yet been described. Here we conducted a survey of rodent-borne Trypanosoma in 19 towns and villages of Niger and Nigeria, with a special emphasis on Niamey, the capital city of Niger. The 1298 rodents that were captured yielded 189 qPCR-positive animals from 14 localities, thus corresponding to a 14.6% overall prevalence. Rats, especially black rats, displayed particularly elevated prevalence (27.4%), with some well sampled sites showing 40-50% and up to 68.8% of Trypanosoma-carrying individuals. Rattus were also characterized by significantly lower Ct values than in the other non-Rattus species. DNA sequences could be obtained for 43 rodent-borne Trypanosoma and corresponded to 41 T. lewisi (all from Rattus) and 2 T. microti (from Cricetomys gambianus). These results, together with data compiled from the available literature, suggest that Rattus may play a particular role for the maintaining and circulation of Trypanosoma, especially T. lewisi, in Africa. Taken into account its strong abilities to invade coastal and inland regions of the continent, we believe that this genus deserves a particular attention in regards to potentially under-looked but emerging atypical trypanosome-related diseases. Copyright © 2017 Elsevier B.V. All rights reserved.

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

    PubMed

    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.

  10. Treatment and follow-up of the first case of human trypanosomiasis caused by Trypanosoma evansi in India.

    PubMed

    Joshi, P P; Chaudhari, A; Shegokar, V R; Powar, R M; Dani, V S; Somalwar, A M; Jannin, J; Truc, P

    2006-10-01

    The first reported human case of trypanosomiasis caused by Trypanosoma evansi was treated using suramin. Patient follow-up indicates that the drug and specific regimen used were well tolerated. Clinical, serological and parasitological investigations at 6 months indicate complete cure of the patient. Suramin should be considered in the treatment of other cases of human T. evansi infection, if they occur.

  11. Seroprevalence of Trypanosoma cruzi Infection in Schoolchildren and in Pregnant Women from an Amazonian Region in Orellana Province, Ecuador

    PubMed Central

    Vargas, Caty Carrera; Narváez, Alberto Orlando; Aroca, Jenny Muzzio; Shiguango, Gonzalo; Robles, Luiggi Martini; Herrera, Claudia; Dumonteil, Eric

    2015-01-01

    Chagas disease is a parasitic disease caused by the protozoan parasite Trypanosoma cruzi and about 230,000 persons are estimated to be infected in Ecuador. However, limited studies have been performed in the Amazon region, on the eastern side of the country. We evaluated here the seroprevalence of Trypanosoma cruzi infection in 12 rural villages of the Loreto canton, Orellana Province in schoolchildren aged 5–15 years and in pregnant women. A total of 1,649 blood samples were tested for Trypanosoma cruzi antibodies by enzyme-linked immunosorbent assay and indirect hemaglutination, and discordant samples were tested by indirect immunofluorescence assay. We detected a seroprevalence of anti-Trypanosoma cruzi antibodies of 1.3% in schoolchildren aged 5–15 years, indicating the persistence of a constant and active vectorial transmission in the Loreto County and confirming the need of the implementation of nonconventional vector control. We also observed a seroprevalence of 3.8% in pregnant women, indicating a clear risk of congenital transmission. Further studies should help define this risk more precisely and implement current international guidelines for the diagnosis, treatment, and care of these cases. PMID:26283751

  12. Seroprevalence of Trypanosoma cruzi Infection in Schoolchildren and in Pregnant Women from an Amazonian Region in Orellana Province, Ecuador.

    PubMed

    Carrera Vargas, Caty; Narváez, Alberto Orlando; Muzzio Aroca, Jenny; Shiguango, Gonzalo; Robles, Luiggi Martini; Herrera, Claudia; Dumonteil, Eric

    2015-10-01

    Chagas disease is a parasitic disease caused by the protozoan parasite Trypanosoma cruzi and about 230,000 persons are estimated to be infected in Ecuador. However, limited studies have been performed in the Amazon region, on the eastern side of the country. We evaluated here the seroprevalence of Trypanosoma cruzi infection in 12 rural villages of the Loreto canton, Orellana Province in schoolchildren aged 5-15 years and in pregnant women. A total of 1,649 blood samples were tested for Trypanosoma cruzi antibodies by enzyme-linked immunosorbent assay and indirect hemaglutination, and discordant samples were tested by indirect immunofluorescence assay. We detected a seroprevalence of anti-Trypanosoma cruzi antibodies of 1.3% in schoolchildren aged 5-15 years, indicating the persistence of a constant and active vectorial transmission in the Loreto County and confirming the need of the implementation of nonconventional vector control. We also observed a seroprevalence of 3.8% in pregnant women, indicating a clear risk of congenital transmission. Further studies should help define this risk more precisely and implement current international guidelines for the diagnosis, treatment, and care of these cases.

  13. Insight into the Exoproteome of the Tissue-Derived Trypomastigote form of Trypanosoma cruzi

    PubMed Central

    Queiroz, Rayner M. L.; Ricart, Carlos A. O.; Machado, Mara O.; Bastos, Izabela M. D.; de Santana, Jaime M.; de Sousa, Marcelo V.; Roepstorff, Peter; Charneau, Sébastien

    2016-01-01

    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 3 h 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 Trypanosoma 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 defenses. PMID:27872839

  14. Nitrofuran drugs as common subversive substrates of Trypanosoma cruzi lipoamide dehydrogenase and trypanothione reductase.

    PubMed

    Blumenstiel, K; Schöneck, R; Yardley, V; Croft, S L; Krauth-Siegel, R L

    1999-12-01

    Lipoamide dehydrogenase (LipDH), trypanothione reductase (TR), and glutathione reductase (GR) catalyze the NAD(P)H-dependent reduction of disulfide substrates. TR occurs exclusively in trypanosomatids which lack a GR. Besides their physiological reactions, the flavoenzymes catalyze the single-electron reduction of nitrofurans with the concomitant generation of superoxide anions. Here, we report on the interaction of clinically used antimicrobial nitrofurans with LipDH and TR from Trypanosoma cruzi, the causative agent of Chagas' disease (South American trypanosomiasis), in comparison to mammalian LipDH and GR. The compounds were studied as inhibitors and as subversive substrates of the enzymes. None of the nitrofurans inhibited LipDH, although they did interfere with the disulfide reduction of TR and GR. When the compounds were studied as substrates, T. cruzi LipDH showed a high rate of nitrofuran reduction and was even more efficient than its mammalian counterpart. Several derivatives were also effective subversive substrates of TR, but the respective reaction with human GR was negligible. Nifuroxazide, nifuroxime, and nifurprazine proved to be the most promising derivatives since they were redox-cycled by both T. cruzi LipDH and TR and had pronounced antiparasitic effects in cultures of T. cruzi and Trypanosoma brucei. The results suggest that those nitrofuran derivatives which interact with both parasite flavoenzymes should be revisited as trypanocidal drugs.

  15. Expression, purification and crystallization of Trypanosoma cruzi dihydroorotate dehydrogenase complexed with orotate

    SciTech Connect

    Inaoka, Daniel Ken; Takashima, Eizo; Osanai, Arihiro; Shimizu, Hironari; Nara, Takeshi; Aoki, Takashi; Harada, Shigeharu; Kita, Kiyoshi

    2005-10-01

    The Trypanosoma cruzi dihydroorotate dehydrogenase, a key enzyme in pyrimidine de novo biosynthesis and redox homeostasis, was crystallized in complex with its first reaction product, orotate. Dihydroorotate dehydrogenase (DHOD) catalyzes the oxidation of dihydroorotate to orotate, the fourth step and the only redox reaction in the de novo biosynthesis of pyrimidine. DHOD from Trypanosoma cruzi (TcDHOD) has been expressed as a recombinant protein in Escherichia coli and purified to homogeneity. Crystals of the TcDHOD–orotate complex were grown at 277 K by the sitting-drop vapour-diffusion technique using polyethylene glycol 3350 as a precipitant. The crystals diffract to better than 1.8 Å resolution using synchrotron radiation (λ = 0.900 Å). X-ray diffraction data were collected at 100 K and processed to 1.9 Å resolution with 98.2% completeness and an overall R{sub merge} of 7.8%. The TcDHOD crystals belong to the orthorhombic space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 67.87, b = 71.89, c = 123.27 Å. The presence of two molecules in the asymmetric unit (2 × 34 kDa) gives a crystal volume per protein weight (V{sub M}) of 2.2 Å{sup 3} Da{sup −1} and a solvent content of 44%.

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

    PubMed

    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. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. Differential behaviour of glucose 6-phosphate dehydrogenase in two morphological forms of Trypanosoma cruzi.

    PubMed

    Lupiañez, J A; Adroher, F J; Vargas, A M; Osuna, A

    1987-01-01

    1. Glucose 6-phosphate dehydrogenase activity (EC 1.1.1.49) of two morphological forms of Trypanosoma cruzi, epimastigotes and metacyclics, are reported. 2. The kinetic behaviour and some of the kinetic parameters of the enzyme in both forms were studied. The enzymes showed a simple Michaelis-Menten kinetic. 3. The activity in epimastigote forms was alway higher than the metacyclic ones. At subsaturating concentrations of substrate was almost 10-fold higher, whereas at saturating concentrations was about 2-fold higher. 4. In epimastigote forms the specific activity and Km values, at pH 7.5 and 37 degrees C, was found to be 142 mUnits x mg-1 of protein and 0.23 mM, respectively. 5. In the same conditions, the specific activity and Km values in metacyclic forms was 75 mUnits x mg-1 of protein and 1.06 mM, respectively. 6. A possible role in the carbohydrate metabolism of glucose 6-phosphate dehydrogenase in both forms of Trypanosoma cruzi is discussed.

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

    PubMed Central

    Cardinal, Marta V.; Lauricella, Marta A.; Ceballos, Leonardo A.; Lanati, Leonardo; Marcet, Paula L.; Levin, Mariano J.; Kitron, Uriel; Gürtler, Ricardo E.; Schijman, Alejandro G.

    2011-01-01

    Genetic diversity of Trypanosoma cruzi populations and parasite transmission dynamics have been well documented throughout the Americas, but few studies have been conducted in the Gran Chaco ecoregion, one of the most highly endemic areas for Chagas disease, caused by T. cruzi. In this study we assessed the distribution of T. cruzi lineages (identified by PCR strategies) in Triatoma infestans, domestic dogs, cats, humans and sylvatic mammals from two neighboring rural areas with different histories of transmission and vector control in northern Argentina. Lineage II predominated among the 99 isolates characterized and lineage I among the six isolates obtained from sylvatic mammals. Trypanosoma cruzi lineage IIe predominated in domestic habitats; it was found in 87% of 54 isolates from Tr. infestans, in 82% of 33 isolates from dogs, and in the four cats found infected. Domestic and sylvatic cycles overlapped in the study area in the late 1980s, when intense domestic transmission occurred, and still overlap marginally. The introduction of T. cruzi from sylvatic into domestic habitats is likely to occur very rarely in the current epidemiological context. The household distribution of T. cruzi lineages showed that Tr. infestans, dogs and cats from a given house compound shared the same parasite lineage in most cases. Based on molecular evidence, this result lends further support to the importance of dogs and cats as domestic reservoir hosts of T. cruzi. We believe that in Argentina, this is the first time that lineage IIc has been isolated from naturally-infected domestic dogs and Tr. infestans. PMID:18585717

  19. Partial biochemical characterization of a metalloproteinase from the bloodstream forms of Trypanosoma brucei brucei parasites.

    PubMed

    de Sousa, Karina Pires; Atouguia, Jorge; Silva, Marcelo Sousa

    2010-05-01

    Metalloproteinases (MMP) belong to the family of cation dependent endopeptidases that degrade matrices at physiological pH and to cleave extracellular matrix proteins. They play an important role in diverse physiological and pathological processes; not only there diverse types of MMP differ in structure and functionally, but also their enzymatic activity is regulated at multiple levels. Trying to shed some light over the processes that govern the pathology of African Trypanosomiasis, the aim of the present study was to examine the proteolytic activity of the crude trypanosome protein extract obtained from the bloodstream forms of Trypanosoma brucei brucei parasites. We hereby report the partial biochemical characterization of a neutral Trypanosoma brucei-metalloproteinase that displays marked proteolytic activities on gelatin and casein, with a molecular mass of approximately 40 kDa, whose activity is strongly dependent of pH and temperature. Furthermore, we show that this activity can be inhibited by classical MMP inhibitors such as EDTA, EGTA, phenantroline, and also by tetracycline and derivatives. This study has a relevant role in the search for new therapeutical targets, for the use of metalloproteinases inhibitors as treatment strategies, or as enhancement to trypanocidal drugs used in the treatment of the disease.

  20. Effects of dietary polyamine deficiency on Trypanosoma gambiense infection in rats.

    PubMed

    Nishimura, K; Araki, N; Ohnishi, Y; Kozaki, S

    2001-02-01

    Nishimura, K., Araki, N., Ohnishi, Y., and Kozaki, S. 2001. Effects of dietary polyamine deficiency on Trypanosoma gambiense infection in rats. Experimental Parasitology 97, 95-101. A diet deficient in polyamines decreases the availability of dietary polyamines. We used rats infected with the Wellcome strain of Trypanosoma gambiense to examine the effects of polyamine-deficient chow (PDC) on trypanosome proliferation and symptoms of infection. Rats fed PDC showed limited increase of trypanosome and symptoms of infection and limited loss of body weight and anemia. Survival in these rats was prolonged. Before infection, the heparinized plasma concentration of spermidine in the PDC-fed rats was lower than that in control rats fed with standard chow. After infection, the content of spermidine in red blood cells increased in the control rats, but was only slightly increased in PDC-fed rats. The content of spermidine in the trypanosomes after infection was low in the PDC-fed rats. Decreases in the polyamine content of trypanosomes limited their increase. These observations suggest that a reduction in dietary polyamines may help in the regulation of trypanosome infection. Copyright 2001 Academic Press.

  1. Identification of a glycosylphosphatidylinositol anchor-modifying β1-3 galactosyltransferase in Trypanosoma brucei

    PubMed Central

    Izquierdo, Luis; Acosta-Serrano, Alvaro; Mehlert, Angela; Ferguson, Michael AJ

    2015-01-01

    Trypanosoma brucei is the causative agent of human African sleeping sickness and the cattle disease nagana.  Trypanosoma brucei is dependent on glycoproteins for its survival and infectivity throughout its life cycle. Here we report the functional characterization of TbGT3, a glycosyltransferase expressed in the bloodstream and procyclic form of the parasite. Bloodstream and procyclic form TbGT3 conditional null mutants were created and both exhibited normal growth under permissive and nonpermissive conditions. Under nonpermissive conditions, the normal glycosylation of the major glycoprotein of bloodstream form T. brucei, the variant surface glycoprotein and the absence of major alterations in lectin binding to other glycoproteins suggested that the major function of TbGT3 occurs in the procyclic form of the parasite. Consistent with this, the major surface glycoprotein of the procyclic form, procyclin, exhibited a marked reduction in molecular weight due to changes in glycosylphosphatidylinositol (GPI) anchor side chains. Structural analysis of the mutant procyclin GPI anchors indicated that TbGT3 encodes a UDP-Gal: β-GlcNAc-GPI β1-3 Gal transferase. Despite the alterations in GPI anchor side chains, TbGT3 conditional null mutants remained infectious to tsetse flies under nonpermissive conditions. PMID:25467966

  2. Trypanosoma cf. varani in an imported ball python (Python reginus) from Ghana.

    PubMed

    Sato, Hiroshi; Takano, Ai; Kawabata, Hiroki; Une, Yumi; Watanabe, Haruo; Mukhtar, Maowia M

    2009-08-01

    Peripheral blood from a ball python (Python reginus) imported from Ghana was cultured in Barbour-Stoenner-Kelly (BSK) medium for Borrelia spp. isolation, resulting in the prominent appearance of free, and clusters of, trypanosomes in a variety of morphological forms. The molecular phylogenetic characterization of these cultured trypanosomes, using the small subunit rDNA, indicated that this python was infected with a species closely related to Trypanosoma varani Wenyon, 1908, originally described in the Nile monitor lizard (Varanus niloticus) from Sudan. Furthermore, nucleotide sequences of glycosomal glyceraldehyde-3-phosphate dehydrogenase gene of both isolates showed few differences. Giemsa-stained blood smears, prepared from the infected python 8 mo after the initial observation of trypanosomes in hemoculture, contained trypomastigotes with a broad body and a short, free flagellum; these most closely resembled the original description of T. varani, or T. voltariae Macfie, 1919 recorded in a black-necked spitting cobra (Naja nigricollis) from Ghana. It is highly possible that lizards and snakes could naturally share an identical trypanosome species. Alternatively, lizards and snakes in the same region might have closely related, but distinct, Trypanosoma species as a result of sympatric speciation. From multiple viewpoints, including molecular phylogenetic analyses, reappraisal of trypanosome species from a wide range of reptiles in Africa is needed to clarify the relationship of recorded species, or to unmask unrecorded species.

  3. Trypanosoma cruzi Polyamine Transporter: Its Role on Parasite Growth and Survival Under Stress Conditions.

    PubMed

    Reigada, Chantal; Sayé, Melisa; Vera, Edward Valera; Balcazar, Darío; Fraccaroli, Laura; Carrillo, Carolina; Miranda, Mariana R; Pereira, Claudio A

    2016-08-01

    Trypanosoma cruzi is the etiological agent of Chagas disease, a major health problem in Latin America. Polyamines are polycationic compounds that play a critical role as regulators of cell growth and differentiation. In contrast with other protozoa, T. cruzi is auxotrophic for polyamines because of its inability to synthesize putrescine due to the lack of both, arginine and ornithine decarboxylase; therefore, the intracellular availability of polyamines depends exclusively on transport processes. In this work, the polyamine transporter TcPAT12 was overexpressed in T. cruzi epimastigotes demonstrating that growth rates at different concentrations of polyamines strongly depend on the regulation of the polyamine transport. In addition, parasites overexpressing TcPAT12 showed a highly increased resistance to hydrogen peroxide and the trypanocidal drugs nifurtimox and benznidazole, which act by oxidative stress and interfering the synthesis of polyamine derivatives, respectively. Finally, the presence of putative polyamine transporters was analyzed in T. cruzi, Trypanosoma brucei, and Leishmania major genomes identifying 3-6 genes in these trypanosomatids.

  4. Digital holographic microscopy for detection of Trypanosoma cruzi parasites in fresh blood mounts

    NASA Astrophysics Data System (ADS)

    Romero, G. G.; Monaldi, A. C.; Alanís, E. E.

    2012-03-01

    An off-axis holographic microscope, in a transmission mode, calibrated to automatically detect the presence of Trypanosoma cruzi in blood is developed as an alternative diagnosis tool for Chagas disease. Movements of the microorganisms are detected by measuring the phase shift they produce on the transmitted wave front. A thin layer of blood infected by Trypanosoma cruzi parasites is examined in the holographic microscope, the images of the visual field being registered with a CCD camera. Two consecutive holograms of the same visual field are subtracted point by point and a phase contrast image of the resulting hologram is reconstructed by means of the angular spectrum propagation algorithm. This method enables the measurement of phase distributions corresponding to temporal differences between digital holograms in order to detect whether parasites are present or not. Experimental results obtained using this technique show that it is an efficient alternative that can be incorporated successfully as a part of a fully automatic system for detection and counting of this type of microorganisms.

  5. Antiparasitic evaluation of betulinic acid derivatives reveals effective and selective anti-Trypanosoma cruzi inhibitors.

    PubMed

    Meira, Cássio Santana; Barbosa-Filho, José Maria; Lanfredi-Rangel, Adriana; Guimarães, Elisalva Teixeira; Moreira, Diogo Rodrigo Magalhães; Soares, Milena Botelho Pereira

    2016-07-01

    Betulinic acid is a pentacyclic triterpenoid with several biological properties already described, including antiparasitic activity. Here, the anti-Trypanosoma cruzi activity of betulinic acid and its semi-synthetic amide derivatives (BA1-BA8) was investigated. The anti-Trypanosoma cruzi activity and selectivity were enhanced in semi-synthetic derivatives, specially on derivatives BA5, BA6 and BA8. To understand the mechanism of action underlying betulinic acid anti-T. cruzi activity, we investigated ultrastructural changes by electron microscopy. Ultrastructural studies showed that trypomastigotes incubated with BA5 had membrane blebling, flagella retraction, atypical cytoplasmic vacuoles and Golgi cisternae dilatation. Flow cytometry analysis showed that parasite death is mainly caused by necrosis. Treatment with derivatives BA5, BA6 or BA8 reduced the invasion process, as well as intracellular parasite development in host cells, with a potency and selectivity similar to that observed in benznidazole-treated cells. More importantly, the combination of BA5 and benznidazole revealed synergistic effects on trypomastigote and amastigote forms of T. cruzi. In conclusion, we demonstrated that BA5 compound is an effective and selective anti-T. cruzi agent. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. The Role of the Trypanosoma cruzi TcNRBD1 Protein in Translation.

    PubMed

    Oliveira, Camila; Carvalho, Paulo Costa; Alves, Lysangela Ronalte; Goldenberg, Samuel

    2016-01-01

    The regulation of gene expression in trypanosomatids occurs mainly at the post-transcriptional level. Despite the importance of this type of control in Trypanosoma cruzi, few RNA binding proteins have been characterized. The RRM domain (RNA Recognition Motif) is one of the most abundant domains found in RNA-binding proteins in higher eukaryotes. Proteins containing the RRM domain are involved in the majority of post-transcriptional processes regulating gene expression. In this work, we aimed to characterize the protein TcNRBD1 from T. cruzi. TcNRBD1 is an RNA-binding protein that contains 2 RRM domains and is the ortholog of the P34 and P37 proteins from Trypanosoma brucei. The TcNRBD1 protein is expressed in all developmental stages of T. cruzi, and its localization pattern is concentrated at the perinuclear region. TcNRBD1 is associated with polysomes and with the 80S monosomes. Furthermore, sequencing of the mRNAs bound to TcNRBD1 allowed the identification of several transcripts that encode ribosomal proteins. Immunoprecipitation assays followed by mass spectrometry showed that the protein complexes with several ribosomal proteins from both the 40S and 60S subunits. In summary, the results indicate that TcNRBD1 is associated with different parts of the translation process, either by regulating mRNAs that encode ribosomal proteins or by acting in some step of ribosome assembly in T. cruzi.

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

    PubMed

    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.

  8. Structural role of the active-site metal in the conformation of Trypanosoma brucei phosphoglycerate mutase.

    PubMed

    Mercaldi, Gustavo F; Pereira, Humberto M; Cordeiro, Artur T; Michels, Paul A M; Thiemann, Otavio H

    2012-06-01

    Phosphoglycerate mutases (PGAMs) participate in both the glycolytic and the gluconeogenic pathways in reversible isomerization of 3-phosphoglycerate and 2-phosphoglycerate. PGAMs are members of two distinct protein families: enzymes that are dependent on or independent of the 2,3-bisphosphoglycerate cofactor. We determined the X-ray structure of the monomeric Trypanosoma brucei independent PGAM (TbiPGAM) in its apoenzyme form, and confirmed this observation by small angle X-ray scattering data. Comparing the TbiPGAM structure with the Leishmania mexicana independent PGAM structure, previously reported with a phosphoglycerate molecule bound to the active site, revealed the domain movement resulting from active site occupation. The structure reported here shows the interaction between Asp319 and the metal bound to the active site, and its contribution to the domain movement. Substitution of the metal-binding residue Asp319 by Ala resulted in complete loss of independent PGAM activity, and showed for the first time its involvement in the enzyme's function. As TbiPGAM is an attractive molecular target for drug development, the apoenzyme conformation described here provides opportunities for its use in structure-based drug design approaches. Database Structural data for the Trypanosoma brucei 2,3-bisphosphoglycerate-independent phosphoglycerate mutase (iPGAM) has been deposited with the Research Collaboratory for Structural Bioinformatics (RCSB) Protein Data Bank under code 3NVL.

  9. Insight into the Exoproteome of the Tissue-Derived Trypomastigote form of Trypanosoma cruzi.

    PubMed

    Queiroz, Rayner M L; Ricart, Carlos A O; Machado, Mara O; Bastos, Izabela M D; de Santana, Jaime M; de Sousa, Marcelo V; Roepstorff, Peter; Charneau, Sébastien

    2016-01-01

    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 3 h 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 Trypanosoma 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 defenses.

  10. Trypanosoma evansi infection and major risk factors for Iranian one-humped camels (Camelus dromedarius).

    PubMed

    Mirshekar, Freshteh; Yakhchali, Mohammad; Shariati-Sharifi, Fariborz

    2017-09-01

    Trypanosoma evansi is a cosmopolitan protozoan which affects camelids and may cause illness and economic losses. The present study was aimed to determine the prevalence and geographic distribution of T. evansi in Iranian one-humped camels of South East Iran. A total of 369 camels were randomly examined from three parts of southeastern Iran from March to September 2015. Blood samples were taken from jugular vein and examined by using thin blood smear and mercuric chloride test. Ninety-five out of 369 examined camels (25.75%) with clinical signs (15.8%, 58/369) were found to be infected with T. evansi. The prevalence was significantly higher in camels more than 4 years-old (23.3%) with clinical signs (8.8%) than the other ones. There was no significant difference between the prevalence and sex (6.23% male and 19.51% female). Trypanosoma evansi infection was geographically found in all investigated regions with the highest prevalence in North (17.61%). The results indicated that T. evansi infection was prevalent in Iranian one-humped camels which could be useful finding to lunch control programs in the region.

  11. Molecular epidemiology of Trypanosoma cruzi and Triatoma dimidiata in costal Ecuador.

    PubMed

    Wong, Yim Yan; Sornosa Macias, Karen Jeniffer; Guale Martínez, Doris; Solorzano, Luis F; Ramirez-Sierra, Maria Jesus; Herrera, Claudia; Dumonteil, Eric

    2016-07-01

    Chagas disease is a neglected tropical disease caused by the protozoan parasite Trypanosoma cruzi. In Ecuador, Triatoma dimidiata and Rhodnius ecuadoriensis are the main vector species, responsible for over half of the cases of T. cruzi infection in the country. T. dimidiata is believed to have been introduced in Ecuador during colonial times, and its elimination from the country is thus believed to be feasible. We investigated here the molecular ecology of T. dimidiata and T. cruzi in costal Ecuador to further guide control efforts. Analysis of the Internal Transcribed Spacer 2 (ITS-2) of 23 specimens from Progreso, Guayas, unambiguously supported the likely importation of T. dimidiata from Central America to Ecuador. The observation of a very high parasite infection rate (54%) and frequent feeding on humans (3/5) confirmed a continued risk of transmission to humans. All genotyped parasites corresponded to TcI DTU and Trypanosoma rangeli was not detected in T. dimidiata. TcI subgroups corresponded to TcIa (25%), and mixed infections with TcIa and TcId (75%). Further studies should help clarify T. cruzi genetic structure in the country, and the possible impact of the introduction of T. dimidiata on the circulating parasite strains. The elevated risk posed by this species warrants continuing efforts for its control, but its apparent mobility between peridomestic and domestic habitats may favor reinfestation following insecticide spraying.

  12. Molecular profiles of Venezuelan isolates of Trypanosoma sp. by random amplified polymorphic DNA method.

    PubMed

    Perrone, T M; Gonzatti, M I; Villamizar, G; Escalante, A; Aso, P M

    2009-05-12

    Nine Trypanosoma sp. Venezuelan isolates, initially presumed to be T. evansi, were collected from three different hosts, capybara (Apure state), horse (Apure state) and donkey (Guarico state) and compared by the random amplification polymorphic DNA technique (RAPD). Thirty-one to 46 reproducible fragments were obtained with 12 of the 40 primers that were used. Most of the primers detected molecular profiles with few polymorphisms between the seven horse, capybara and donkey isolates. Quantitative analyses of the RAPD profiles of these isolates revealed a high degree of genetic conservation with similarity coefficients between 85.7% and 98.5%. Ten of the primers generated polymorphic RAPD profiles with two of the three Trypanosoma sp. horse isolates, namely TeAp-N/D1 and TeGu-N/D1. The similarity coefficient between these two isolates and the rest, ranged from 57.9% to 68.4% and the corresponding dendrogram clustered TeAp-N/D1 and Te Gu-N/D1 in a genetically distinct group.

  13. Variable sensitivity to complement-mediated lysis among Trypanosoma rangeli reference strains.

    PubMed

    de Sousa, Maria Auxiliadora; Dos Santos Pereira, Sheila Medeiros; Dos Santos Faissal, Barbara Neves

    2012-02-01

    Six reference strains of Trypanosoma rangeli from different days of growth in axenic cultures were assayed for susceptibility to complement-mediated lysis by non-immune guinea-pig serum. Their authenticity was also confirmed by isoenzyme analyses. Parasites were incubated with 25% active or 68°C-inactivated serum (37°C, 30 min) for all tests; thereafter the lysis rates were determined. Highly variable lysis percentages were observed among T. rangeli strains and in the same stock at different growing days. In a few assays, three strains (Macias, R-1625 and Choachi) presented total or very high resistance. The others (H-14, San Agustín and SC-58) were generally most susceptible, and could reach lysis rates as high as Trypanosoma cruzi. After incubation with active sera, the epimastigotes were usually the predominant stages, being followed by spheromastigotes and/or transitional forms. Those stages and trypomastigotes could also be partially susceptible. In four strains, the short epimastigotes were more resistant to lysis than the long ones. Experiments with C3-deficient serum displayed total or partial participation of the alternative-complement pathway in T. rangeli lysis. This study confirmed the variable complement sensitivity of T. rangeli, which can be related to its intraspecific heterogeneity, to the remarkable complexity of its life-cycle stages, and to the methodology employed.

  14. The Role of the Trypanosoma cruzi TcNRBD1 Protein in Translation

    PubMed Central

    Oliveira, Camila; Carvalho, Paulo Costa; Goldenberg, Samuel

    2016-01-01

    The regulation of gene expression in trypanosomatids occurs mainly at the post-transcriptional level. Despite the importance of this type of control in Trypanosoma cruzi, few RNA binding proteins have been characterized. The RRM domain (RNA Recognition Motif) is one of the most abundant domains found in RNA-binding proteins in higher eukaryotes. Proteins containing the RRM domain are involved in the majority of post-transcriptional processes regulating gene expression. In this work, we aimed to characterize the protein TcNRBD1 from T. cruzi. TcNRBD1 is an RNA-binding protein that contains 2 RRM domains and is the ortholog of the P34 and P37 proteins from Trypanosoma brucei. The TcNRBD1 protein is expressed in all developmental stages of T. cruzi, and its localization pattern is concentrated at the perinuclear region. TcNRBD1 is associated with polysomes and with the 80S monosomes. Furthermore, sequencing of the mRNAs bound to TcNRBD1 allowed the identification of several transcripts that encode ribosomal proteins. Immunoprecipitation assays followed by mass spectrometry showed that the protein complexes with several ribosomal proteins from both the 40S and 60S subunits. In summary, the results indicate that TcNRBD1 is associated with different parts of the translation process, either by regulating mRNAs that encode ribosomal proteins or by acting in some step of ribosome assembly in T. cruzi. PMID:27760165

  15. Recombinant SSP4 protein from Trypanosoma cruzi amastigotes regulates nitric oxide production by macrophages.

    PubMed

    Ramos-Ligonio, A; López-Monteon, A; Talamás-Rohana, P; Rosales-Encina, J L

    2004-10-01

    Acute infection with Trypanosoma cruzi is characterized by immunosuppression mediated by T cells and macrophages (Mphis). Nitric oxide (NO) production during the initial phase of acute infection might participate in the clearance of parasites by Mphis, whereas its overproduction during the late phase of acute infection would account for the immunosuppression observed. Trypanosoma cruzi molecules that might regulate the host responses have not been fully identified. Here, we demonstrate that active immunization with MBP::SSP4, a recombinant protein derived from a surface antigen specific of T. cruzi amastigotes (TcSSP4), was able to stimulate Ab production (IgG1, IgG2a, and IgG2b). On the other hand, MBP::SSP4 was able to stimulate NO production by peritoneal Mphis from BALB/c mice and Mphis from the J774 cell line. This effect was also observed at the level of inducible nitric oxide synthase (iNOS) detected by Western Blot. Furthermore, MBP::SSP4 was also shown to induce the expression of IL-1alpha, IL-6, IL-12, IFN-gamma, and TNF-alpha in normal animals, and IL-10 in immunized animals. In addition the protein MBP::SSP4 was able to bind to the surface of PMphis and J774 Mphis. These results suggest that TcSSP4 could modulate Mphi NO production and this may represent a mechanism participating in the immunoregulatory processes during Chagas' disease.

  16. Genetic Engineering of Trypanosoma (Dutonella) vivax and In Vitro Differentiation under Axenic Conditions

    PubMed Central

    D'Archivio, Simon; Medina, Mathieu; Cosson, Alain; Chamond, Nathalie; Rotureau, Brice; Minoprio, Paola; Goyard, Sophie

    2011-01-01

    Trypanosoma vivax is one of the most common parasites responsible for animal trypanosomosis, and although this disease is widespread in Africa and Latin America, very few studies have been conducted on the parasite's biology. This is in part due to the fact that no reproducible experimental methods had been developed to maintain the different evolutive forms of this trypanosome under laboratory conditions. Appropriate protocols were developed in the 1990s for the axenic maintenance of three major animal Trypanosoma species: T. b. brucei, T. congolense and T. vivax. These pioneer studies rapidly led to the successful genetic manipulation of T. b. brucei and T. congolense. Advances were made in the understanding of these parasites' biology and virulence, and new drug targets were identified. By contrast, challenging in vitro conditions have been developed for T. vivax in the past, and this per se has contributed to defer both its genetic manipulation and subsequent gene function studies. Here we report on the optimization of non-infective T. vivax epimastigote axenic cultures and on the process of parasite in vitro differentiation into metacyclic infective forms. We have also constructed the first T. vivax specific expression vector that drives constitutive expression of the luciferase reporter gene. This vector was then used to establish and optimize epimastigote transfection. We then developed highly reproducible conditions that can be used to obtain and select stably transfected mutants that continue metacyclogenesis and are infectious in immunocompetent rodents. PMID:22216367

  17. The challenge of Trypanosoma brucei gambiense sleeping sickness diagnosis outside Africa.

    PubMed

    Lejon, V; Boelaert, M; Jannin, J; Moore, A; Büscher, P

    2003-12-01

    Sleeping sickness is a lethal African disease caused by parasites of the Trypanosoma brucei subspecies, which is transmitted by tsetse flies. Occasionally, patients are reported outside Africa. Diagnosis of such imported cases can be problematic when the infection is due to Trypanosoma brucei gambiense, the chronic form of sleeping sickness found in west and central Africa. The low number of trypanosomes in the blood and the non-specific, variable symptoms make the diagnosis difficult, particularly when the index of suspicion is low. When the trypanosomes have penetrated into the central nervous system, neuropathological signs become apparent but even at this stage, misdiagnosis is frequent. Rapid and correct diagnosis of sleeping sickness can avoid inappropriate or delayed treatment and even death of the patient. In this article, an overview on diagnosis of imported cases of T b gambiense sleeping sickness is given, and possible pitfalls in the diagnostic process are highlighted. Bioclinical parameters that should raise the suspicion of sleeping sickness in a patient who has been in west or central Africa are discussed. Techniques for diagnosis are reviewed. A clinician suspecting sleeping sickness should contact a national reference centre for tropical medicine in his or her country, or the WHO, Geneva, Switzerland, or the Centers for Disease Control and Prevention (CDC), Atlanta, GA, USA, for clinical consultation and provision of specific diagnostic tests. Appropriate drugs for sleeping sickness treatment are also provided by WHO and the CDC.

  18. Trypanosoma cruzi: variability of stocks from Colombia determined by molecular karyotype and minicircle Southern blot analysis.

    PubMed

    Triana, Omar; Ortiz, Sylvia; Dujardin, Jean-Claude; Solari, Aldo

    2006-05-01

    Nineteen Trypanosoma cruzi stocks, most of them of wild origin, and four Trypanosoma rangeli stocks from Colombia were analysed by molecular karyotype analysis with cloned DNA cruzipain as the probe. Another 27 cloned stocks of T. cruzi from different geographic areas of South America were used as reference for T. cruzi lineages. Phenetic analysis of chromosome size polymorphism demonstrated a great variability of Colombian T. cruzi stocks, suggesting that most belong to lineage I, although two of them belong to lineage II. The 2 lineage II T. cruzi, 17 T. cruzi lineage I, and 3 T. rangeli stocks from Colombia were studied further by Southern blot analysis with a panel of kinetoplast DNA minicircle probes. Hybridisation results indicate that the two T. cruzi II stocks are genetically distant from each other and from T. cruzi lineages IIb, IId, and IIe from Chile. Finally, T. cruzi minicircle probes do not cross-hybridise in any stringency condition tested with T. rangeli minicircles, a clear indication that these parasites can be easily distinguished by this method.

  19. The Dialogue of the Host-Parasite Relationship: Leishmania spp. and Trypanosoma cruzi Infection

    PubMed Central

    de Morais, Carlos Gustavo Vieira; Castro Lima, Ana Karina; dos Santos, Rosiane Freire; Da-Silva, Silvia Amaral Gonçalves; Dutra, Patrícia Maria Lourenço

    2015-01-01

    The intracellular protozoa Leishmania spp. and Trypanosoma cruzi and the causative agents of Leishmaniasis and Chagas disease, respectively, belong to the Trypanosomatidae family. Together, these two neglected tropical diseases affect approximately 25 million people worldwide. Whether the host can control the infection or develops disease depends on the complex interaction between parasite and host. Parasite surface and secreted molecules are involved in triggering specific signaling pathways essential for parasite entry and intracellular survival. The recognition of the parasite antigens by host immune cells generates a specific immune response. Leishmania spp. and T. cruzi have a multifaceted repertoire of strategies to evade or subvert the immune system by interfering with a range of signal transduction pathways in host cells, which causes the inhibition of the protective response and contributes to their persistence in the host. The current therapeutic strategies in leishmaniasis and trypanosomiasis are very limited. Efficacy is variable, toxicity is high, and the emergence of resistance is increasingly common. In this review, we discuss the molecular basis of the host-parasite interaction of Leishmania and Trypanosoma cruzi infection and their mechanisms of subverting the immune response and how this knowledge can be used as a tool for the development of new drugs. PMID:26090399

  20. Trypanosoma teixeirae: A new species belonging to the T. cruzi clade causing trypanosomosis in an Australian little red flying fox (Pteropus scapulatus).

    PubMed

    Barbosa, Amanda D; Mackie, John T; Stenner, Robyn; Gillett, Amber; Irwin, Peter; Ryan, Una

    2016-06-15

    Little is known about the genetic diversity and pathogenicity of trypanosomes in Australian bats. Recently a novel trypanosome species was identified in an adult female little red flying fox (Pteropus scapulatus) with clinical and pathological evidence of trypanosomosis. The present study used morphology and molecular methods to demonstrate that this trypanosome is a distinct species and we propose the name Trypanosoma teixeirae sp. n. Morphological comparison showed that its circulating trypomastigotes were significantly different from those of Trypanosoma pteropi and Trypanosoma hipposideri, two species previously described from Australian bats. Genetic information was not available for T. pteropi and T. hipposideri but phylogenetic analyses at the 18S ribosomal RNA (rRNA) and glycosomal glyceraldehyde phosphate dehydrogenase (gGAPDH) loci indicated that T. teixeirae sp. n. was genetically distinct and clustered with other bat-derived trypanosome species within the Trypanosoma cruzi clade. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. Prevalence of antibodies to Trypanosoma cruzi, Toxoplasma gondii, Encephalitozonn cuniculi, Sarcocystis neurona, Besnoitia darlingi, and Neospora caninum in North American opossum, Didelphis virginiana, from Southern Louisian

    USDA-ARS?s Scientific Manuscript database

    We examined the prevalence of antibodies to zoonotic protozoan parasites (Trypanosoma cruzi, Toxoplasma gondii, and Encephalitozoon cuniculi) and protozoan’s of veterinary importance (Neospora caninum, Sarcocystis neurona and Besnoitia darlingi) in a population of North American opossums (Didelphis...

  2. Southern plains woodrats (Neotoma micropus) from southern Texas are important reservoirs of two genotypes of Trypanosoma cruzi and host of a putative novel Trypanosoma species.

    PubMed

    Charles, Roxanne A; Kjos, Sonia; Ellis, Angela E; Barnes, John C; Yabsley, Michael J

    2013-01-01

    Trypanosoma cruzi, the causative agent of Chagas' disease, is an important public health and veterinary pathogen. Although human cases are rare in the United States, infections in wildlife, and in some areas domestic dogs, are common. In 2008 and 2010, we investigated T. cruzi prevalence in possible vertebrate reservoirs in southern Texas, with an emphasis on southern plains woodrats (Neotoma micropus). Infection status was determined using a combination of culture isolation, polymerase chain reaction (PCR), and serologic testing. Based on PCR and/or culture, T. cruzi was detected in 35 of 104 (34%) woodrats, 3 of 4 (75%) striped skunks (Mephitis mephitis), 12 of 20 (60%) raccoons (Procyon lotor), and 5 of 28 (18%) other rodents including a hispid cotton rat (Sigmodon hispidus), rock squirrel (Otospermophilus variegatus), black rat (Rattus rattus), and two house mice (Mus musculus). Additionally, another Trypanosoma species was detected in 41 woodrats, of which 27 were co-infected with T. cruzi. Genetic characterization of T. cruzi revealed that raccoon, rock squirrel, and cotton rat isolates were genotype TcIV, while woodrats and skunks were infected with TcI and TcIV. Based on the Chagas Stat-Pak assay, antibodies were detected in 27 woodrats (26%), 13 raccoons (65%), 4 skunks (100%), and 5 other rodents (18%) (two white-ankled mice [Peromyscus pectoralis laceianus], two house mice, and a rock squirrel). Seroprevalence based on indirect immunofluorescence antibody testing was higher for both woodrats (37%) and raccoons (90%), compared with the Chagas Stat-Pak. This is the first report of T. cruzi in a hispid cotton rat, black rat, rock squirrel, and white-ankled mouse. These data indicate that based on culture and PCR testing, the prevalence of T. cruzi in woodrats is comparable with other common reservoirs (i.e., raccoons and opossums) in the United States. However, unlike raccoons and opossums, which tend to be infected with a particular genotype, southern

  3. Southern Plains Woodrats (Neotoma micropus) from Southern Texas Are Important Reservoirs of Two Genotypes of Trypanosoma cruzi and Host of a Putative Novel Trypanosoma Species

    PubMed Central

    Charles, Roxanne A.; Kjos, Sonia; Ellis, Angela E.; Barnes, John C.

    2013-01-01

    Abstract Trypanosoma cruzi, the causative agent of Chagas' disease, is an important public health and veterinary pathogen. Although human cases are rare in the United States, infections in wildlife, and in some areas domestic dogs, are common. In 2008 and 2010, we investigated T. cruzi prevalence in possible vertebrate reservoirs in southern Texas, with an emphasis on southern plains woodrats (Neotoma micropus). Infection status was determined using a combination of culture isolation, polymerase chain reaction (PCR), and serologic testing. Based on PCR and/or culture, T. cruzi was detected in 35 of 104 (34%) woodrats, 3 of 4 (75%) striped skunks (Mephitis mephitis), 12 of 20 (60%) raccoons (Procyon lotor), and 5 of 28 (18%) other rodents including a hispid cotton rat (Sigmodon hispidus), rock squirrel (Otospermophilus variegatus), black rat (Rattus rattus), and two house mice (Mus musculus). Additionally, another Trypanosoma species was detected in 41 woodrats, of which 27 were co-infected with T. cruzi. Genetic characterization of T. cruzi revealed that raccoon, rock squirrel, and cotton rat isolates were genotype TcIV, while woodrats and skunks were infected with TcI and TcIV. Based on the Chagas Stat-Pak assay, antibodies were detected in 27 woodrats (26%), 13 raccoons (65%), 4 skunks (100%), and 5 other rodents (18%) (two white-ankled mice [Peromyscus pectoralis laceianus], two house mice, and a rock squirrel). Seroprevalence based on indirect immunofluorescence antibody testing was higher for both woodrats (37%) and raccoons (90%), compared with the Chagas Stat-Pak. This is the first report of T. cruzi in a hispid cotton rat, black rat, rock squirrel, and white-ankled mouse. These data indicate that based on culture and PCR testing, the prevalence of T. cruzi in woodrats is comparable with other common reservoirs (i.e., raccoons and opossums) in the United States. However, unlike raccoons and opossums, which tend to be infected with a particular genotype

  4. Unraveling the differences of the hydrolytic activity of Trypanosoma cruzi trans-sialidase and Trypanosoma rangeli sialidase: a quantum mechanics-molecular mechanics modeling study.

    PubMed

    Bueren-Calabuig, Juan A; Pierdominici-Sottile, Gustavo; Roitberg, Adrian E

    2014-06-05

    Chagas' disease, also known as American trypanosomiasis, is a lethal, chronic disease that currently affects more than 10 million people in Central and South America. The trans-sialidase from Trypanosoma cruzi (T. cruzi, TcTS) is a crucial enzyme for the survival of this parasite: sialic acids from the host are transferred to the cell surface glycoproteins of the trypanosome, thereby evading the host's immune system. On the other hand, the sialidase of T. rangeli (TrSA), which shares 70% sequence identity with TcTS, is a strict hydrolase and shows no trans-sialidase activity. Therefore, TcTS and TrSA represent an excellent framework to understand how different catalytic activities can be achieved with extremely similar structures. By means of combined quantum mechanics-molecular mechanics (QM/MM, SCC-DFTB/Amberff99SB) calculations and umbrella sampling simulations, we investigated the hydrolysis mechanisms of TcTS and TrSA and computed the free energy profiles of these reactions. The results, together with our previous computational investigations, are able to explain the catalytic mechanism of sialidases and describe how subtle differences in the active site make TrSA a strict hydrolase and TcTS a more efficient trans-sialidase.

  5. Unraveling the Differences of the Hydrolytic Activity of Trypanosoma cruzitrans-Sialidase and Trypanosoma rangeli Sialidase: A Quantum Mechanics–Molecular Mechanics Modeling Study

    PubMed Central

    2015-01-01

    Chagas’ disease, also known as American trypanosomiasis, is a lethal, chronic disease that currently affects more than 10 million people in Central and South America. The trans-sialidase from Trypanosoma cruzi (T. cruzi, TcTS) is a crucial enzyme for the survival of this parasite: sialic acids from the host are transferred to the cell surface glycoproteins of the trypanosome, thereby evading the host’s immune system. On the other hand, the sialidase of T. rangeli (TrSA), which shares 70% sequence identity with TcTS, is a strict hydrolase and shows no trans-sialidase activity. Therefore, TcTS and TrSA represent an excellent framework to understand how different catalytic activities can be achieved with extremely similar structures. By means of combined quantum mechanics–molecular mechanics (QM/MM, SCC-DFTB/Amberff99SB) calculations and umbrella sampling simulations, we investigated the hydrolysis mechanisms of TcTS and TrSA and computed the free energy profiles of these reactions. The results, together with our previous computational investigations, are able to explain the catalytic mechanism of sialidases and describe how subtle differences in the active site make TrSA a strict hydrolase and TcTS a more efficient trans-sialidase. PMID:24814976

  6. Epidemiology of Chagas disease in Guatemala: infection rate of Triatoma dimidiata, Triatoma nitida and Rhodnius prolixus (Hemiptera, Reduviidae) with Trypanosoma cruzi and Trypanosoma rangeli (Kinetoplastida, Trypanosomatidae).

    PubMed

    Monroy, Carlota; Rodas, Antonieta; Mejía, Mildred; Rosales, Regina; Tabaru, Yuichiro

    2003-04-01

    A five-year domiciliary collection in the 22 departments of Guatemala showed that out of 4,128 triatomines collected, 1,675 were Triatoma dimidiata (Latreille, 1811), 2,344 were Rhodnius prolixus Stal 1859, and only 109 were T. nitida Usinger 1939. The Chagas disease parasite, Trypanosoma cruzi, was found in all three species. Their natural infection rates were similar in the first two species (20.6%; 19.1%) and slightly lower in T. nitida(13.8%). However there was no significant difference in the infection rates in the three species (p = 0.131). T. dimidiata males have higher infection rates than females (p = 0.030), whereas for R. prolixus there is no difference in infection rates between males and females (p = 0.114). The sex ratios for all three species were significantly skewed. More males than females were found inside houses for T. dimidiata (p < 0.0001) and T. nitida (p = 0.011); a different pattern was seen for R. prolixus (p = 0.037) where more females were found. Sex ratio is proposed as an index to show the mobility of T. dimidiata in different populations. T. dimidiata is widely distributed in the country, and is also the main vector in at least ten departments, but R. prolixus with higher vectorial capacity is an important vector in at least two departments.

  7. Polymerase chain reaction identification of Trypanosoma brucei rhodesiense in wild tsetse flies from Nkhotakota Wildlife Reserve, Malawi.

    PubMed

    Musaya, Janelisa; Chisi, John; Senga, Edward; Nambala, Peter; Maganga, Emmanuel; Matovu, Enock; Enyaru, John

    2017-03-01

    Trypanosoma brucei rhodesiense is the causative agent of acute human African trypanosomiasis. Identification of T. b. rhodesiense in tsetse populations is essential for understanding transmission dynamics, assessng human disease risk, and monitoring spatiotemporal trends and impact of control interventions. Accurate detection and characterisation of trypanosomes in vectors relies on molecular techniques. For the first time in Malawi, a molecular technique has been used to detect trypanosomes in tsetse flies in Nkhotakota Wildlife Reserve. A polymerase chain reaction (PCR) technique was used to identify the serum resistance associated (SRA) gene of T. b. rhodesiense in tsetse flies. Of 257 tsetse flies that were randomly caught, 42 flies were dissected for microscopic examination. The midguts of 206 flies were positive and were individually put in eppendorf tubes containing phosphate-buffered saline (PBS buffer) for DNA extraction. Internal transcribed spacer (ITS)-PCR was first used to isolate all trypanosome species from the flies. TBR PCR was then used to isolate the Trypanozoon group. T. brucei-positive samples were further evaluated by SRA PCR for the presence of the SRA gene. Of 257 flies caught, 185 (72%) were Glossina morsitans morsitans and 72 (28%) were Glossina pallidipes. Three were tenerals and 242 were mature live flies. Of the 242 flies dissected, 206 were positive, representing an 85.1% infection rate. From 206 infected flies, 106 (51.5%) were positive using ITS-PCR, 68 (33.0%) being mixed infections, 18 (8.7%) T. brucei, 9 (4.4%) Trypanosoma vivax, 4 (1.9%) Trypanosoma godfrey, 3 (1.5%) Trypanosoma congolense savanna, 3 (1.5%) Trypanosoma simae, and 1 (0.4%) Trypanosoma simaetsavo. When subjected to TBR PCR, 107(51.9%) were positive for T. brucei. Of the 107 T. brucei-positive samples, 5 (4.7%) were found to have the SRA gene. These results suggest that wild tsetse flies in Malawi are infected with human-infective trypanosomes that put communities

  8. Acute Chagas outbreaks: molecular and biological features of Trypanosoma cruzi isolates, and clinical aspects of acute cases in Santander, Colombia.

    PubMed

    Díaz, Martha Lucía; Leal, Sandra; Mantilla, Julio César; Molina-Berríos, Alfredo; López-Muñoz, Rodrigo; Solari, Aldo; Escobar, Patricia; González Rugeles, Clara Isabel

    2015-11-26

    Outbreaks of acute Chagas disease associated with oral transmission are easily detected nowadays with trained health personnel in areas of low endemicity, or in which the vector transmission has been interrupted. Given the biological and genetic diversity of Trypanosoma cruzi, the high morbidity, mortality, and the observed therapeutic failure, new characteristics of these outbreaks need to be addressed at different levels, both in Trypanosoma cruzi as in patient response. The aim of this work was to evaluate the patient's features involved in six outbreaks of acute Chagas disease which occurred in Santander, Colombia, and the characteristics of Trypanosoma cruzi clones isolated from these patients, to establish the potential relationship between the etiologic agent features with host behavior. The clinical, pathological and epidemiological aspects of outbreaks were analyzed. In addition, Trypanosoma cruzi clones were biologically characterized both in vitro and in vivo, and the susceptibility to the classical trypanocidal drugs nifurtimox and benznidazole was evaluated. Trypanosoma cruzi clones were genotyped by means of mini-exon intergenic spacer and cytochrome b genes sequencing. All clones were DTU I, and based on the mini-exon intergenic spacer, belong to two genotypes: G2 related with sub-urban, and G11 with rural outbreaks. Girón outbreak clones with higher susceptibility to drugs presented G2 genotype and C/T transition in Cyt b. The outbreaks affected mainly young population (±25.9 years), and the mortality rate was 10 %. The cardiac tissue showed intense inflammatory infiltrate, myocardial necrosis and abundant amastigote nests. However, although the gastrointestinal tissue was congestive, no inflammation or parasites were observed. Although all clones belong to DTU I, two intra-DTU genotypes were found with the sequencing of the mini-exon intergenic spacer, however there is no strict correlation between genetic groups, the cycles of the parasite or

  9. Antiheart antibody-dependent cytotoxicity in the sera of mice chronically infected with Trypanosoma cruzi.

    PubMed Central

    Laguens, R P; Meckert, P C; Chambó, J G

    1988-01-01

    Sera of mice chronically infected with Trypanosoma cruzi contain antibodies that bind to the surface of living adult syngeneic heart muscle cells. In a syngeneic system, with nonadherent spleen mononuclear cells as effector cells and cardiocytes as targets, antibody-dependent cytotoxicity (ADCC), revealed by the liberation of creatine phosphokinase from damaged cardiocytes, was observed after incorporation of serum samples from infected mice. Target damage was decreased after absorption with syngeneic myocardium, but absorption with T. cruzi epimastigotes or trypomastigotes or with syngeneic skeletal muscle had no effect on ADCC. No complement-dependent lysis against heart muscle cells was detected in the same serum samples. These observations indicate that serum from chronically chagasic mice contain antibodies that bind to the surface of living adult syngeneic cardiocytes and are able to exert ADCC, suggesting that they could play a role in the pathogenesis of the heart damage that occurs in Chagas' disease. Images PMID:3126153

  10. Isradipine and lacidipine: effects in vivo and in vitro on Trypanosoma cruzi epimastigotes.

    PubMed

    Núñez-Vergara, L J; Squella, J A; Bollo-Dragnic, S; Marín-Catalán, R; Pino, L; Díaz-Araya, G; Letelier, M E

    1998-01-01

    1. Isradipine and lacidipine, two new drugs that are members of the nitro-aryl-1,4-dihydropyridine family, produced inhibition of both growth cultures and oxygen consumption on epimastigotes of Trypanosoma cruzi Tulahuen strain, at micromolar concentrations. 2. Isradipine was found to be the most potent derivative in both, in growth cultures (I50 = 20.8 microM) and in vivo oxygen uptake (I50 = 31.1 microM). 3. Diltiazem and verapamil, two well-known calcium channel antagonists, lacked inhibitory activity, even at a 100 microM concentration. 4. The present findings indicate that the trypanocide effects exerted by isradipine and lacidipine are not related with a disruption of the calcium homeostasis of the parasite.

  11. Nitro aryl 1,4-dihydropyridine derivatives: effects on Trypanosoma cruzi.

    PubMed

    Núñez-Vergara, L J; Squella, J A; Bollo-Dragnic, S; Morello, A; Repetto, Y; Aldunate, J; Letelier, M E

    1997-09-01

    A series of nitro aryl 1,4-dihydropyridine derivatives produced inhibition of both cell growth and oxygen consumption on Tulahuen and LQ strains, and clone Dm 28c of epimastigotes of Trypanosoma cruzi. Nicardipine was found to be the most potent derivative in both growth cell (I50 = 70 microM) and oxygen uptake (I50 = 26 microM in intact parasites, I50 = 325 microM in situ mitochondria). A correlation between the inhibitory effects on the growth cell and the apparent first order kinetic for the uptake of the 1,4-dihypyridine derivatives by T. cruzi epimastigotes was found. Thus, nicardipine, the most potent derivative, exhibited the highest apparent rate constant, ku, (0.043 min-1). On the other hand, no susceptibility differences by strains and clone Dm 28c to the action of these drugs were found.

  12. Trypanosoma Cruzi Cyp51 Inhibitor Derived from a Mycobacterium Tuberculosis Screen Hit

    SciTech Connect

    Chen, Chiung-Kuang; Doyle, Patricia S.; Yermalitskaya, Liudmila V.; Mackey, Zachary B.; Ang, Kenny K.H.; McKerrow, James H.; Podust, Larissa M.

    2009-02-18

    The two front-line drugs for chronic Trypanosoma cruzi infections are limited by adverse side-effects and declining efficacy. One potential new target for Chagas disease chemotherapy is sterol 14{alpha}-demethylase (CYP51), a cytochrome P450 enzyme involved in biosynthesis of membrane sterols. In a screening effort targeting Mycobacterium tuberculosis CYP51 (CYP51{sub Mt}), we previously identified the N-[4-pyridyl]-formamide moiety as a building block capable of delivering a variety of chemotypes into the CYP51 active site. In that work, the binding modes of several second generation compounds carrying this scaffold were determined by high-resolution co-crystal structures with CYP51{sub Mt}. Subsequent assays against the CYP51 orthologue in T. cruzi, CYP51{sub Tc}, demonstrated that two of the compounds tested in the earlier effort bound tightly to this enzyme. Both were tested in vitro for inhibitory effects against T. cruzi and the related protozoan parasite Trypanosoma brucei, the causative agent of African sleeping sickness. One of the compounds had potent, selective anti-T. cruzi activity in infected mouse macrophages. Cure of treated host cells was confirmed by prolonged incubation in the absence of the inhibiting compound. Discrimination between T. cruzi and T. brucei CYP51 by the inhibitor was largely based on the variability (phenylalanine versus isoleucine) of a single residue at a critical position in the active site. CYP51{sub Mt}-based crystal structure analysis revealed that the functional groups of the two tightly bound compounds are likely to occupy different spaces in the CYP51 active site, suggesting the possibility of combining the beneficial features of both inhibitors in a third generation of compounds to achieve more potent and selective inhibition of CYP51{sub Tc}. Enzyme sterol 14{alpha}-demethylase (CYP51) is a well-established target for anti-fungal therapy and is a prospective target for Chagas disease therapy. We previously identified a

  13. Phylogenetic character mapping of proteomic diversity shows high correlation with subspecific phylogenetic diversity in Trypanosoma cruzi

    PubMed Central

    Telleria, Jenny; Biron, David G.; Brizard, Jean-Paul; Demettre, Edith; Séveno, Martial; Barnabé, Christian; Ayala, Francisco J.; Tibayrenc, Michel

    2010-01-01

    We performed a phylogenetic character mapping on 26 stocks of Trypanosoma cruzi, the parasite responsible for Chagas disease, and 2 stocks of the sister taxon T. cruzi marinkellei to test for possible associations between T. cruzi–subspecific phylogenetic diversity and levels of protein expression, as examined by proteomic analysis and mass spectrometry. We observed a high level of correlation (P < 10−4) between genetic distance, as established by multilocus enzyme electrophoresis, and proteomic dissimilarities estimated by proteomic Euclidian distances. Several proteins were found to be specifically associated to T. cruzi phylogenetic subdivisions (discrete typing units). This study explores the previously uncharacterized links between infraspecific phylogenetic diversity and gene expression in a human pathogen. It opens the way to searching for new vaccine and drug targets and for identification of specific biomarkers at the subspecific level of pathogens. PMID:21059959

  14. The structure of the 80S ribosome from Trypanosoma cruzi reveals unique rRNA components

    PubMed Central

    Gao, Haixiao; Ayub, Maximiliano Juri; Levin, Mariano J.; Frank, Joachim

    2005-01-01

    We present analysis, by cryo-electron microscopy and single-particle reconstruction, of the structure of the 80S ribosome from Trypanosoma cruzi, the kinetoplastid protozoan pathogen that causes Chagas disease. The density map of the T. cruzi 80S ribosome shows the phylogenetically conserved eukaryotic rRNA core structure, together with distinctive structural features in both the small and large subunits. Remarkably, a previously undescribed helical structure appears in the small subunit in the vicinity of the mRNA exit channel. We propose that this rRNA structure likely participates in the recruitment of ribosome onto the 5′ end of mRNA, in facilitating and modulating the initiation of translation that is unique to the trypanosomes. PMID:16014419

  15. Expression of a glycosylphosphatidylinositol-anchored Trypanosoma brucei transferrin-binding protein complex in insect cells.

    PubMed Central

    Chaudhri, M; Steverding, D; Kittelberger, D; Tjia, S; Overath, P

    1994-01-01

    The expression site-associated gene ESAG 6 was previously implicated in transferrin binding in the protozoan parasite Trypanosoma brucei. ESAG 6 and the closely related ESAG 7 of T. brucei strain AnTat1.3 have now been expressed in insect cells using the baculovirus expression system. Expression of ESAG 6 alone in insect cells gives rise to a glycosylated protein of approximately 52 kDa, which is cell surface-associated through a glycosylphosphatidylinositol anchor at its C terminus. The ESAG 7 product of about 42 kDa is also glycosylated, but lacks the glycosylphosphatidylinositol modification, and is located intracellularly. No transferrin-binding activity is observed when either ESAG is expressed independently. However, their expression results in a cell surface complex of ESAG 6 and 7 products that specifically binds transferrin. This shows that both ESAG 6 and 7 products are necessary and sufficient for binding to transferrin. Images PMID:8022802

  16. Aromatic glycosyl disulfide derivatives: evaluation of their inhibitory activities against Trypanosoma cruzi.

    PubMed

    Gutiérrez, Bessy; Muñoz, Christian; Osorio, Luis; Fehér, Krisztina; Illyés, Tünde-Zita; Papp, Zsuzsa; Kumar, Ambati Ashok; Kövér, Katalin E; Sagua, Hernán; Araya, Jorge E; Morales, Patricio; Szilágyi, László; González, Jorge

    2013-06-15

    Aromatic oligovalent glycosyl disulfides and some diglycosyl disulfides were tested against three different Trypanosoma cruzi strains. Di-(β-D-galactopyranosyl-dithiomethylene) benzenes 2b and 4b proved to be the most active derivatives against all three strains of cell culture-derived trypomastigotes with IC50 values ranging from 4 to 11 μM at 37 °C. The inhibitory activities were maintained, although somewhat lowered, at a temperature of 4 °C as well. Three further derivatives displayed similar activities against at least one of the three strains. Low cytotoxicities of the active compounds, tested on confluent HeLa, Vero and peritoneal macrophage cell cultures, resulted in significantly higher selectivity indices (SI) than that of the reference drug benznidazole. Remarkably, several molecules of the tested panel strongly inhibited the parasite release from T. cruzi infected HeLa cell cultures suggesting an effect against the intracellular development of T. cruzi amastigotes as well.

  17. Natively Inhibited Trypanosoma brucei Cathepsin B Structure Determined by Using an X-ray Laser

    PubMed Central

    DePonte, Daniel P.; White, Thomas A.; Rehders, Dirk; Barty, Anton; Stellato, Francesco; Liang, Mengning; Barends, Thomas R.M.; Boutet, Sébastien; Williams, Garth J.; Messerschmidt, Marc; Seibert, M. Marvin; Aquila, Andrew; Arnlund, David; Bajt, Sasa; Barth, Torsten; Bogan, Michael J.; Caleman, Carl; Chao, Tzu-Chiao; Doak, R. Bruce; Fleckenstein, Holger; Frank, Matthias; Fromme, Raimund; Galli, Lorenzo; Grotjohann, Ingo; Hunter, Mark S.; Johansson, Linda C.; Kassemeyer, Stephan; Katona, Gergely; Kirian, Richard A.; Koopmann, Rudolf; Kupitz, Chris; Lomb, Lukas; Martin, Andrew V.; Mogk, Stefan; Neutze, Richard; Shoeman, Robert L.; Steinbrener, Jan; Timneanu, Nicusor; Wang, Dingjie; Weierstall, Uwe; Zatsepin, Nadia A.; Spence, John C. H.; Fromme, Petra; Schlichting, Ilme; Duszenko, Michael; Betzel, Christian; Chapman, Henry N.

    2013-01-01

    The Trypanosoma brucei cysteine protease cathepsin B (TbCatB), which is involved in host protein degradation, is a promising target to develop new treatments against sleeping sickness, a fatal disease caused by this protozoan parasite. The structure of the mature, active form of TbCatB has so far not provided sufficient information for the design of a safe and specific drug against T. brucei. By combining two recent innovations, in vivo crystallization and serial femtosecond crystallography, we obtained the room-temperature 2.1 angstrom resolution structure of the fully glycosylated precursor complex of TbCatB. The structure reveals the mechanism of native TbCatB inhibition and demonstrates that new biomolecular information can be obtained by the “diffraction-before-destruction” approach of x-ray free-electron lasers from hundreds of thousands of individual microcrystals. PMID:23196907

  18. Protein preparation, crystallization and preliminary X-ray analysis of Trypanosoma cruzi nucleoside diphosphate kinase 1.

    PubMed

    Gómez Barroso, J A; Pereira, H; Miranda, M; Pereira, C; Garratt, R C; Aguilar, C F

    2010-07-01

    The flagellated protozoan parasite Trypanosoma cruzi is the aetiological agent of Chagas disease. Nucleoside diphosphate kinases (NDPKs) are enzymes that are involved in energy management and nucleoside balance in the cell. T. cruzi TcNDPK1, a canonical isoform, was overexpressed in Escherichia coli as an N-terminally poly-His-tagged fusion protein and crystallized. Crystals grew after 72 h in 0.2 M MgCl(2), 20% PEG 3350. Data were collected to 3.5 A resolution using synchrotron X-ray radiation at the National Synchrotron Light Laboratory (Campinas, Brazil). The crystals belonged to the trigonal space group P3, with unit-cell parameters a = b = 127.84, c = 275.49 A. Structure determination is under way and will provide relevant information that may lead to the first step in rational drug design for the treatment of Chagas disease.

  19. Trypanosoma cruzi Epimastigotes Are Able to Manage Internal Cholesterol Levels under Nutritional Lipid Stress Conditions

    PubMed Central

    Pereira, Miria Gomes; Visbal, Gonzalo; Salgado, Leonardo T.; Vidal, Juliana Cunha; Godinho, Joseane L. P.; De Cicco, Nuccia N. T.; Atella, Geórgia C.; de Souza, Wanderley; Cunha-e-Silva, Narcisa

    2015-01-01

    Trypanosoma cruzi epimastigotes store high amounts of cholesterol and cholesteryl esters in reservosomes. These unique organelles are responsible for cellular digestion by providing substrates for homeostasis and parasite differentiation. Here we demonstrate that under nutritional lipid stress, epimastigotes preferentially mobilized reservosome lipid stocks, instead of lipid bodies, leading to the consumption of parasite cholesterol reservoirs and production of ergosterol. Starved epimastigotes acquired more LDL-NBD-cholesterol by endocytosis and distributed the exogenous cholesterol to their membranes faster than control parasites. Moreover, the parasites were able to manage internal cholesterol levels, alternating between consumption and accumulation. With normal lipid availability, parasites esterified cholesterol exhibiting an ACAT-like activity that was sensitive to Avasimibe in a dose-dependent manner. This result also implies that exogenous cholesterol has a role in lipid reservoirs in epimastigotes. PMID:26068009

  20. Role of Trypanosoma cruzi Trans-sialidase on the Escape from Host Immune Surveillance

    PubMed Central

    Nardy, Ana F. F. R.; Freire-de-Lima, Celio G.; Pérez, Ana R.; Morrot, Alexandre

    2016-01-01

    Chagas disease is caused by the flagellate protozoan Trypanosoma cruzi, affecting millions of people throughout Latin America. The parasite dampens host immune response causing modifications in diverse lymphoid compartments, including the thymus. T. cruzi trans-sialidase (TS) seems to play a fundamental role in such immunopathological events. This unusual enzyme catalyses the transference of sialic acid molecules from host glycoconjugates to acceptor molecules placed on the parasite surface. TS activity mediates several biological effects leading to the subversion of host immune system, hence favoring both parasite survival and the establishment of chronic infection. This review summarizes current findings on the roles of TS in the immune response during T. cruzi infection. PMID:27047464

  1. Trypanosoma cruzi, the Causal Agent of Chagas Disease: Boundaries between Wild and Domestic Cycles in Venezuela

    PubMed Central

    Herrera, Leidi

    2014-01-01

    Trypanosoma cruzi the etiological agent of American Trypanosomiasis or Chagas disease (ChD) is transmitted by triatomines vectors between mammals including man. T. cruzi has existed for circa 150 Ma in the Americas and nearly 10 million people are currently infected. The overlap between wild and domestic ecotopes where T. cruzi circulates is increasing. Host–parasite interactions have been determined by infection patterns in these cycles, all under natural or laboratorial conditions. This mini-review describes specific parasite niches, such as plant communities or biological corridors between domestic and wild landscapes, in order to help identify risk factors for ChD and define the boundaries between wild and domestic transmission cycles, with an emphasis on research undertaken in Venezuela. PMID:25506587

  2. Trypanosoma brucei parasites occupy and functionally adapt to the adipose tissue in mice

    DOE PAGES

    Trindade, Sandra; Rijo-Ferreira, Filipa; Carvalho, Tania; ...

    2016-05-26

    Trypanosoma brucei is an extracellular parasite that causes sleeping sickness. In mammalian hosts, trypanosomes are thought to exist in two major niches: early in infection, they populate the blood; later, they breach the blood-brain barrier. Working with a well-established mouse model, we discovered that adipose tissue constitutes a third major reservoir for T. brucei. Parasites from adipose tissue, here termed adipose tissue forms (ATFs), can replicate and were capable of infecting a naive animal. ATFs were transcriptionally distinct from bloodstream forms, and the genes upregulated included putative fatty acid β-oxidation enzymes. Consistent with this, ATFs were able to utilize exogenousmore » myristate and form β-oxidation intermediates, suggesting that ATF parasites can use fatty acids as an external carbon source. Lastly, these findings identify the adipose tissue as a niche for T. brucei during its mammalian life cycle and could potentially explain the weight loss associated with sleeping sickness.« less

  3. Trypanosoma brucei Infection in Asymptomatic Greater Kudus (Tragelaphus strepsiceros) on a Game Ranch in Zambia

    PubMed Central

    Siamudaala, Victor; Munyeme, Musso; Nambota, Andrew; Mutoloki, Stephen; Matandiko, Wigganson

    2010-01-01

    Trypomastogotes of Trypanosoma brucei were detected from 4 asymptomatic kudus (Tragelaphus strepsiceros) on a game ranch located approximately 45 km north east of Lusaka, Zambia. Blood smears examined from 14 wildlife species comprising of the impala (Aepyceros melampus), Kafue lechwe (kobus leche kafuensis), sable antelope (Hippotragus niger), tsessebe (Damaliscus lunatus), warthog (Phacochoerus aethiopicus), puku (Kobus vardoni), zebra (Equus burchelli), waterbuck (Kobus ellipsiprymnus), bushbuck (Tragelaphus scriptus), reedbuck (Redunca arundinum), wilderbeest (Connochaetes taurinus), hartebeest (Alcephelus lichtensteini), African buffalo (Syncerus caffer), and kudu (Tragelaphus strepsiceros) showed that only the kudu had T. brucei. Although game ranching has emerged to be a successful ex-situ conservation strategy aimed at saving the declining wildlife population in the National Parks, our findings suggest that it has the potential of aiding the re-distribution of animal diseases. Hence, there is a need for augmenting wildlife conservation with disease control strategies aimed at reducing the risk of disease transmission between wildlife and domestic animals. PMID:20333288

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

    PubMed

    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. Copyright © 2014 Elsevier B.V. All rights reserved.

  5. Structure-guided design of novel Trypanosoma brucei Methionyl-tRNA synthetase inhibitors.

    PubMed

    Huang, Wenlin; Zhang, Zhongsheng; Barros-Álvarez, Ximena; Koh, Cho Yeow; Ranade, Ranae M; Gillespie, J Robert; Creason, Sharon A; Shibata, Sayaka; Verlinde, Christophe L M J; Hol, Wim G J; Buckner, Frederick S; Fan, Erkang

    2016-11-29

    A screening hit 1 against Trypanosoma brucei methionyl-tRNA synthetase was optimized using a structure-guided approach. The optimization led to the identification of two novel series of potent inhibitors, the cyclic linker and linear linker series. Compounds of both series were potent in a T. brucei growth inhibition assay while showing low toxicity to mammalian cells. The best compound of each series, 16 and 31, exhibited EC50s of 39 and 22 nM, respectively. Compounds 16 and 31 also exhibited promising PK properties after oral dosing in mice. Moreover, compound 31 had moderately good brain permeability, with a brain/plasma ratio of 0.27 at 60 min after IP injection. This study provides new lead compounds for arriving at new treatments of human African trypanosomiasis (HAT). Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  6. Phloretin and citrate promote the differentiation rate from epimastigote to metacyclic forms of Trypanosoma cruzi.

    PubMed

    Adroher, F J; Osuna, A; Lupiáñez, J A

    1991-11-01

    We have investigated the effects of the metabolic inhibitors, phloretin, 2-deoxy-D-glucose, maleate and trans-aconitate, as well as two intermediates of the tricarboxylic-acid cycle, acetate and citrate, on the growth and metacyclogenesis of Trypanosoma cruzi. 0.1 mM phloretin increased the percentage of metacyclic forms about 2.7-fold without affecting growth rate, whereas the other inhibitors had no apparent effect on either growth or differentiation rates. The addition of 5 mM citrate stimulated differentiation by about 2.6-fold. When either 10 mM citrate or 10 mM acetate were added, on the other hand, both the growth and differentiation rates were severely inhibited.

  7. Lysosome-like compartments of Trypanosoma cruzi trypomastigotes may originate directly from epimastigote reservosomes.

    PubMed

    Vidal, Juliana C; Alcantara, Carolina DE L; DE Souza, Wanderley; Cunha-E-Silva, Narcisa L

    2017-01-12

    Trypanosoma cruzi epimastigote reservosomes store nutrients taken up during the intense endocytic activity exhibited by this developmental form. Reservosomes were classified as pre-lysosomal compartments. In contrast, trypomastigote forms are not able to take up nutrients from the medium. Interestingly, trypomastigotes also have acidic organelles with the same proteases contained in epimastigote reservosomes. Nevertheless, the origin and function of these organelles have not been disclosed so far. Given the similarities between the compartments of epimastigotes and trypomastigotes, the present study aimed to investigate the origin of metacyclic trypomastigote protease-containing organelles by tracking fluorospheres or colloidal gold particles previously stored in epimastigotes' reservosomes throughout metacyclogenesis. Using three-dimensional reconstruction of serial electron microscopy images, it was possible to find trypomastigote compartments containing the tracer. Our observations demonstrate that the protease-containing compartments from metacyclic trypomastigotes may originate directly from the reservosomes of epimastigotes.

  8. Evaluation of dipeptide nitriles as inhibitors of rhodesain, a major cysteine protease of Trypanosoma brucei.

    PubMed

    Schirmeister, Tanja; Schmitz, Janina; Jung, Sascha; Schmenger, Torsten; Krauth-Siegel, R Luise; Gütschow, Michael

    2017-01-01

    A series of dipeptide nitriles known as inhibitors of mammalian cathepsins were evaluated for inhibition of rhodesain, the cathepsin L-like protease of Trypanosoma brucei. Compound 35 consisting of a Leu residue fitting into the S2 pocket and a triarylic moiety consisting of thiophene, a 1,2,4-oxadiazole and a phenyl ring fitting into the S3 pocket, and compound 33 with a 3-bromo-Phe residue (S2) and a biphenyl fragment (S3) were found to inhibit rhodesain in the single-digit nanomolar range. The observed steep structure-activity relationship could be explained by covalent docking simulations. With their high selectivity indices (ca. 200) and the good antitrypanosomal activity (8μM) the compounds represent promising starting points for new rhodesain inhibitors.

  9. Changes in immunoglobulin levels in zinc-deficient mice infected with Trypanosoma musculi.

    PubMed Central

    Humphrey, P. A.; Lee, C. M.; Ashraf, M.

    1994-01-01

    A metabolic imbalance technique was used to study the effects of zinc deficiency on immunoglobulin levels in mice infected with Trypanosoma musculi or immunized with parasite products. Zinc-deficient mice developed higher numbers of parasitemia earlier and exhibited prolonged infection. Irrespective of the diet, higher IgG1, IgG2b, and IgM levels, lower IgG2a and IgA levels, and uniform IgG3 levels were exhibited primarily by mice infected with T musculi and to a lesser extent by mice immunized with parasite products. Zinc-deficient mice showed smaller increases in IgG1 and IgM, but larger gains in IgG2b compared with mice on full-complement and pair-fed diets. However, IgG2a decreased significantly in zinc-deficient mice. PMID:7932840

  10. Trypanosoma avium incidence, pathogenicity and response to melarsomine in falcons from Kuwait.

    PubMed

    Tarello, W

    2005-03-01

    Epidemiological and clinical studies on Trypanosoma avium are lacking in the Middle East. The aims of this study were to determine the T. avium incidence in falcons from Kuwait, report clinical signs and find an effective therapy. Blood smears from 921 diseased and 56 healthy falcons were examined between May 2003 and April 2004. 12 birds 11.3%) were found infected by T. avium and ten of these were treated with melarsomine (Cymelarsan) at a dosage of 0.25 mg/kg intramuscularly for four days. All affected birds presented clinical signs, including incapacity of flying high, poor appetite, lethargy, loosing weight, weakness, dyspnoea and death. Signs disappeared within 1-7 days after administration of melarsomine. Trypomastigotes were not detected in blood smears made 1-7 days after the end of therapy. This study suggests that T. avium induces disease in falcons and that melarsomine can be an effective therapy eliminating both clinical signs and circulating trypomastigotes.

  11. Trypanosoma cruzi infection in the Mexican state of Guerrero: a seroepidemiological (ELISA) survey of 20 communities.

    PubMed

    Andersson, N; Morales, A; Nava, E; Martinez, E; Rodriguez, I; Young, P; Howard, M K; Miles, M A

    1990-10-01

    The enzyme linked immunosorbent assay (ELISA) was used to analyse 4372 blood samples from residents of 978 households in 20 representative communities in the Mexican state of Guerrero. Seventy-five individuals had very high titres of antibodies against Trypanosoma cruzi. Samples with intermediate optical density values, despite overlapping values with several control positives on a single-well test, did not sustain their positivity at high dilutions. 'Intermediate positives' had a different distribution among the 20 communities to samples sustaining reactivity at high dilutions, indicating possible cross-reactivity with another infectious agent. The finding of seropositive children under the age of 10 years in the Costa Chica, Acapulco and the Tierra Caliente regions, with family clustering of putative cases, indicates that recent transmission must be considered. Very few people interviewed in the 20 communities knew the triatomine bug could transmit a disease.

  12. Mitochondrial and Nucleolar Localization of Cysteine Desulfurase Nfs and the Scaffold Protein Isu in Trypanosoma brucei

    PubMed Central

    Kovářová, Julie; Horáková, Eva; Changmai, Piya; Vancová, Marie

    2014-01-01

    Trypanosoma brucei has a complex life cycle during which its single mitochondrion is subjected to major metabolic and morphological changes. While the procyclic stage (PS) of the insect vector contains a large and reticulated mitochondrion, its counterpart in the bloodstream stage (BS) parasitizing mammals is highly reduced and seems to be devoid of most functions. We show here that key Fe-S cluster assembly proteins are still present and active in this organelle and that produced clusters are incorporated into overexpressed enzymes. Importantly, the cysteine desulfurase Nfs, equipped with the nuclear localization signal, was detected in the nucleolus of both T. brucei life stages. The scaffold protein Isu, an interacting partner of Nfs, was also found to have a dual localization in the mitochondrion and the nucleolus, while frataxin and both ferredoxins are confined to the mitochondrion. Moreover, upon depletion of Isu, cytosolic tRNA thiolation dropped in the PS but not BS parasites. PMID:24243795

  13. Surface charge of trypanosoma cruzi. Binding of cationized ferritin and measurement of cellular electrophoretic mobility.

    PubMed

    De Souza, W; Arguello, C; Martinez-Paloma, A; Trissl, D; Gonzáles-Robles, A; Chiari, E

    1977-08-01

    The surface charge of epimastigote and trypomastigote forms of Trypanosoma cruzi was evaluated by means of binding of cationized ferritin to the cell surface as visualized by electron microscopy, and by direct measurements of the cellular microelectrophoretic mobility (EPM). Epimastigote forms had a mean EPM of -0.52 micrometer-s-1-V-1-cm and were lightly labeled with cationized ferritin. In contrast, bloodstream trypomastigotes had a much higher EPM (-1.14), and the surface was heavily labeled with cationized ferritin. When trypomastigotes from staionary phase cultures were isolated on DEAE cellulose columns, the mean EPM was found to be significantly lower (-0.63), and labeling with cationized ferritin decreased. With a mixed population containing epimastigote, trypomastigote, and intermediate forms, EPM values ranging between -0.70 to -1.14 were found. From these observations we conclude that there is a definite increase in negative surface charge during development from epi- to trypomastigote forms of T. cruzi.

  14. Expression in Escherichia coli of a dominant immunogen of Trypanosoma cruzi recognized by human chagasic sera.

    PubMed Central

    Cotrim, P C; Paranhos, G S; Mortara, R A; Wanderley, J; Rassi, A; Camargo, M E; da Silveira, J F

    1990-01-01

    A genomic clone expressing a Trypanosoma cruzi antigen in Escherichia coli was identified using human chagasic sera. Chagasic antibodies affinity purified on extracts of this clone recognized a high-molecular-weight protein expressed in all developmental stages of the parasite life cycle, as well as in various T. cruzi strains. The antigen is associated with the cytoskeleton of the parasite and localizes along the attachment region between the flagellum and the cell body. Antibodies to the recombinant antigen were detected in the sera of 115 chagasic patients from different endemic regions, but not in sera of patients with leishmaniasis, T. rangeli infection, or other parasitic diseases. Our data suggest that the presence of antibodies to this antigen may be specifically associated with Chagas' disease. Images PMID:1691209

  15. An automatic algorithm for the detection of Trypanosoma cruzi parasites in blood sample images.

    PubMed

    Soberanis-Mukul, Roger; Uc-Cetina, Víctor; Brito-Loeza, Carlos; Ruiz-Piña, Hugo

    2013-12-01

    Chagas disease is a tropical parasitic disease caused by the flagellate protozoan Trypanosoma cruzi (T. cruzi) and currently affecting large portions of the Americas. One of the standard laboratory methods to determine the presence of the parasite is by direct visualization in blood smears stained with some colorant. This method is time-consuming, requires trained microscopists and is prone to human mistakes. In this article we propose a novel algorithm for the automatic detection of T. cruzi parasites, in microscope digital images obtained from peripheral blood smears treated with Wright's stain. Our algorithm achieved a sensitivity of 0.98 and specificity of 0.85 when evaluated against a dataset of 120 test images. Experimental results show the versatility of the method for parasitemia determination.

  16. Gene expression and molecular modeling of the HSP104 chaperone of Trypanosoma cruzi.

    PubMed

    Campos, R A; da Silva, M L; da Costa, G V; Bisch, P M; Peralta, J M; Silva, R; Rondinelli, E; Urményi, T P

    2012-08-06

    Heat shock protein (HSP) 104 is a highly conserved molecular chaperone that catalyzes protein unfolding, disaggregation and degradation under stress conditions. We characterized HSP104 gene structure and expression in Trypanosoma cruzi, a protozoan parasite that causes Chagas' disease. The T. cruzi HSP104 is an 869 amino-acid protein encoded by a single-copy gene that has the highest sequence similarity (76%) with that of T. brucei and the lowest (23%) with that of the human protein. HSP104 transcripts were detected at room temperature, and levels increased after incubation at 37° or 40°C. The HSP104 protein was found at low levels in non-heat-shocked cells, and accumulated continuously up to 24 h at elevated temperatures. We developed a predicted structural model of hexameric T. cruzi HSP104, which showed some conserved features.

  17. New 1,3-thiazole derivatives and their biological and ultrastructural effects on Trypanosoma cruzi.

    PubMed

    de Moraes Gomes, Paulo André Teixeira; de Oliveira Barbosa, Miria; Farias Santiago, Edna; de Oliveira Cardoso, Marcos Veríssimo; Capistrano Costa, Natáli Tereza; Hernandes, Marcelo Zaldini; Moreira, Diogo Rodrigo Magalhães; da Silva, Aline Caroline; Dos Santos, Thiago André Ramos; Pereira, Valéria Rêgo Alves; Brayner Dos Santosd, Fábio André; do Nascimento Pereira, Glaécia Aparecida; Ferreira, Rafaela Salgado; Leite, Ana Cristina Lima

    2016-10-04

    In previous studies, the compound 3-(bromopropiophenone) thiosemicarbazone was described as a potent anti-Trypanosoma cruzi and cruzain inhibitor. In view to optimize this activity, 1,3-thiazole core was used as building-block strategy to access new lead generation of anti T. cruzi agents. In this way a series of thiazole derivatives were synthesized and most of these derivatives exhibited antiparasitic activity similar to benznidazole (Bzd). Among them, compounds (1c) and (1g) presented better selective index (SI) than Bzd. In addition, compounds showed inhibitory activity against the cruzain protease. As observed by electron microscopy, compound (1c) treatment caused irreversible and specific morphological changes on ultrastructure organization of T. cruzi, demonstrating that this class of compounds is killing parasites. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  18. A Brief View of the Surface Membrane Proteins from Trypanosoma cruzi

    PubMed Central

    Pech-Canul, Ángel de la Cruz

    2017-01-01

    Trypanosoma cruzi is the causal agent of Chagas' disease which affects millions of people around the world mostly in Central and South America. T. cruzi expresses a wide variety of proteins on its surface membrane which has an important role in the biology of these parasites. Surface molecules of the parasites are the result of the environment to which the parasites are exposed during their life cycle. Hence, T. cruzi displays several modifications when they move from one host to another. Due to the complexity of this parasite's cell surface, this review presents some membrane proteins organized as large families, as they are the most abundant and/or relevant throughout the T. cruzi membrane. PMID:28656101

  19. Identification of novel Trypanosoma cruzi prolyl oligopeptidase inhibitors by structure-based virtual screening

    NASA Astrophysics Data System (ADS)

    de Almeida, Hugo; Leroux, Vincent; Motta, Flávia Nader; Grellier, Philippe; Maigret, Bernard; Santana, Jaime M.; Bastos, Izabela Marques Dourado

    2016-12-01

    We have previously demonstrated that the secreted prolyl oligopeptidase of Trypanosoma cruzi (POPTc80) is involved in the infection process by facilitating parasite migration through the extracellular matrix. We have built a 3D structural model where POPTc80 is formed by a catalytic α/β-hydrolase domain and a β-propeller domain, and in which the substrate docks at the inter-domain interface, suggesting a "jaw opening" gating access mechanism. This preliminary model was refined by molecular dynamics simulations and next used for a virtual screening campaign, whose predictions were tested by standard binding assays. This strategy was successful as all 13 tested molecules suggested from the in silico calculations were found out to be active POPTc80 inhibitors in the micromolar range (lowest K i at 667 nM). This work paves the way for future development of innovative drugs against Chagas disease.

  20. Trypanosoma irwini n. sp (Sarcomastigophora: Trypanosomatidae) from the koala ( Phascolarctos cinereus).

    PubMed

    McInnes, L M; Gillett, A; Ryan, U M; Austen, J; Campbell, R S F; Hanger, J; Reid, S A

    2009-07-01

    The morphology and genetic characterization of a new species of trypanosome infecting koalas (Phascolarctos cinereus) are described. Morphological analysis of bloodstream forms and phylogenetic analysis at the 18S rDNA and gGAPDH loci demonstrated this trypanosome species to be genetically distinct and most similar to Trypanosoma bennetti, an avian trypanosome with a genetic distance of 0.9% at the 18S rDNA and 10.7% at the gGAPDH locus. The trypanosome was detected by 18S rDNA PCR in the blood samples of 26 out of 68 (38.2%) koalas studied. The aetiological role of trypanosomes in koala disease is currently poorly defined, although infection with these parasites has been associated with severe clinical signs in a number of koalas. Based on biological and genetic characterization data, this trypanosome species infecting koalas is proposed to be a new species Trypanosome irwini n. sp.

  1. Extracellular vesicles from Trypanosoma brucei mediate virulence factor transfer and cause host anemia

    PubMed Central

    Szempruch, Anthony J.; Sykes, Steven E.; Kieft, Rudo; Denison, Lauren; Becker, Allison C.; Gartrell, Anzio; Martin, William J.; Nakayasu, Ernesto S.; Almeida, Igor C.; Hajduk, Stephen L.; Harrington, John M.

    2015-01-01

    Intercellular communication between parasites and with host cells provides mechanisms for parasite development, immune evasion and disease pathology. Bloodstream African trypanosomes produce membranous nanotubes that originate from the flagellar membrane and disassociate into free extracellular vesicles (EVs). Trypanosome EVs contain several flagellar proteins that contribute to virulence and Trypanosoma brucei rhodesiense EVs contain the serum resistance-associated protein (SRA) necessary for human infectivity. T. b. rhodesiense EVs transfer SRA to non-human infectious trypanosomes allowing evasion of human innate immunity. Trypanosome EVs can also fuse with mammalian erythrocytes resulting in rapid erythrocyte clearance and anemia. These data indicate that trypanosome EVs are organelles mediating non-hereditary virulence factor transfer and causing host erythrocyte remodeling inducing anemia. PMID:26771494

  2. Molecular phylogeny of Trypanosoma cruzi from Central America (Guatemala) and a comparison with South American strains.

    PubMed

    Iwagami, M; Higo, H; Miura, S; Yanagi, T; Tada, I; Kano, S; Agatsuma, T

    2007-12-01

    Molecular phylogenetic analysis was carried out for 21 strains of Trypanosoma cruzi, nine of which were obtained from Guatemala and 12 from South America. Phylogenetic trees were constructed using the nucleotide sequences of two nuclear gene regions, dihydrofolate reductase-thymidylate synthase (DHFR-TS) and trypanothione reductase (TR), and contiguous portions of two mitochondrial genes, cytochrome oxidase subunit II (COII) and reduced nicotinamide adenine dinucleotide dehydrogenase subunit 1 (ND1). Possible genetic exchange between the rather divergent lineages of T. cruzi II from South America was suggested in the trees of the two nuclear genes. T. cruzi I strains obtained from Guatemala and Colombia were identical in all the genes examined, but other T. cruzi I isolates from South America were rather polymorphic in the DHFR-TS and mitochondrial genes. No genetic exchange was identified between T. cruzi I populations from Central and South America in the present study.

  3. Familial Analysis of Seropositivity to Trypanosoma cruzi and of Clinical Forms of Chagas Disease

    PubMed Central

    Silva-Grecco, Roseane L.; Balarin, Marly A. S.; Correia, Dalmo; Prata, Aluízio; Rodrigues, Virmondes

    2010-01-01

    A cross-sectional study was carried out in Água Comprida, MG, Brazil, a region previously endemic to Chagas disease whose vectorial transmission was interrupted around 20 year ago. A total of 998 individuals were examined for anti-Trypanosoma cruzi antibodies. Seropositivity was observed in 255 subjects (25.5%), and 743 subjects were negative. Forty-one families with 5–80 individuals with similar environmental conditions were selected for familial analysis. In 15 families, seropositivity to T. cruzi was observed in > 50% of individuals. The segregation analysis confirmed family aggregation for the seropositivity to the T. cruzi. Heart commitment was the major clinical form observed, and in six families, > 50% of the individuals display cardiopathy that may be attributed to T. cruzi infection. Our results support the hypothesis that there is a family aggregation for the seropositivity but without the effect of one major gene. PMID:20064994

  4. Trypanosoma brucei Infection in asymptomatic greater Kudus (Tragelaphus strepsiceros) on a game ranch in Zambia.

    PubMed

    Munang'andu, Hetron Mweemba; Siamudaala, Victor; Munyeme, Musso; Nambota, Andrew; Mutoloki, Stephen; Matandiko, Wigganson

    2010-03-01

    Trypomastogotes of Trypanosoma brucei were detected from 4 asymptomatic kudus (Tragelaphus strepsiceros) on a game ranch located approximately 45 km north east of Lusaka, Zambia. Blood smears examined from 14 wildlife species comprising of the impala (Aepyceros melampus), Kafue lechwe (kobus leche kafuensis), sable antelope (Hippotragus niger), tsessebe (Damaliscus lunatus), warthog (Phacochoerus aethiopicus), puku (Kobus vardoni), zebra (Equus burchelli), waterbuck (Kobus ellipsiprymnus), bushbuck (Tragelaphus scriptus), reedbuck (Redunca arundinum), wilderbeest (Connochaetes taurinus), hartebeest (Alcephelus lichtensteini), African buffalo (Syncerus caffer), and kudu (Tragelaphus strepsiceros) showed that only the kudu had T. brucei. Although game ranching has emerged to be a successful ex-situ conservation strategy aimed at saving the declining wildlife population in the National Parks, our findings suggest that it has the potential of aiding the re-distribution of animal diseases. Hence, there is a need for augmenting wildlife conservation with disease control strategies aimed at reducing the risk of disease transmission between wildlife and domestic animals.

  5. Plasmid vectors and molecular building blocks for the development of genetic manipulation tools for Trypanosoma cruzi.

    PubMed

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

    2013-01-01

    The post genomic era revealed the need for developing better performing, easier to use and more sophisticated genetic manipulation tools for the study of Trypanosoma cruzi, the etiological agent of Chagas disease. In this work a series of plasmids that allow genetic manipulation of this protozoan parasite were developed. First of all we focused on useful tools to establish selection strategies for different strains and which can be employed as expression vectors. On the other hand molecular building blocks in the form of diverse selectable markers, modifiable fluorescent protein and epitope-tag coding sequences were produced. Both types of modules were harboured in backbone molecules conceived to offer multiple construction and sub-cloning strategies. These can be used to confer new properties to already available genetic manipulation tools or as starting points for whole novel designs. The performance of each plasmid and building block was determined independently. For illustration purposes, some simple direct practical applications were conducted.

  6. Loop-mediated isothermal amplification test for Trypanosoma vivax based on satellite repeat DNA.

    PubMed

    Njiru, Z K; Ouma, J O; Bateta, R; Njeru, S E; Ndungu, K; Gitonga, P K; Guya, S; Traub, R

    2011-08-25

    Trypanosoma vivax is major cause of animal trypanosomiasis and responsible for enormous economic burden in Africa and South America animal industry. T. vivax infections mostly run low parasitaemia with no apparent clinical symptoms, making diagnosis a challenge. This work reports the design and evaluation of a loop-mediated isothermal amplification (LAMP) test for detecting T. vivax DNA based on the nuclear satellite repeat sequence. The assay is rapid with results obtained within 35 min. The analytical sensitivity is ∼ 1 trypanosome/ml while that of the classical PCR tests ranged from 10 to 10(3)trypanosomes/ml. The T. vivax LAMP test reported here is simple, robust and has future potential in diagnosis of animal trypanosomiasis in the field.

  7. Urban Trypanosoma (Schizotrypanum) cruzi: pathology in white mice of isolates from Panstrongylus geniculatus.

    PubMed

    De Scorza, C; Urdaneta-Morales, S; Sampson-Ward, L

    1989-12-01

    The histopathological alterations produced in white mice (NMRI strain) by isolates of Trypanosoma (Schizotrypanum) cruzi from Panstrongylus geniculatus captured in dwellings of the city of Caracas, Venezuela are reported. The heart, skeletal muscle, liver, spleen, duodenum, colon, lung, and brain were parasitized by all the isolates. All showed strong myotropism, with elevated virulence and severe histopathological alterations in the cardiac and skeletal muscle, and in the smooth muscle of the duodenum, colon, and lung; parasitization of the mononuclear phagocytic system was discrete. These results, in addition to the morphobiological characters reported in a previous paper, suggest that the isolates in question belong to the same type of parasite. The possible causes of this observation are discussed in the light of the heterogeneity of T. cruzi. The epidemiological significance of the existence of these parasite forms in the urban areas of Caracas is emphasized.

  8. Assembly of the flagellum and its role in cell morphogenesis in Trypanosoma brucei.

    PubMed

    Vaughan, Sue

    2010-08-01

    Eukaryotic flagella are microtubule-based structures required for a variety of functions including cell motility and sensory perception. Most eukaryotic flagella grow out from a cell into the surrounding medium, but when the flagellum of the protozoan parasite Trypanosoma brucei exits the cell via the flagellar pocket, it is attached along the length of the cell body by a cytoskeletal structure called the flagellum attachment zone (FAZ). The exact reasons for flagellum attachment have remained elusive, but evidence is emerging that the attached flagellum plays a major role in cell morphogenesis in this organism. In this review we discuss evidence published in the past four years that is unravelling the role of the flagellum in organelle segregation, inheritance of cell shape and cytokinesis.

  9. Tc45, a dimorphic Trypanosoma cruzi immunogen with variable chromosomal localization, is calreticulin.

    PubMed

    Aguillón, J C; Ferreira, L; Pérez, C; Colombo, A; Molina, M C; Wallace, A; Solari, A; Carvallo, P; Galindo, M; Galanti, N; Orn, A; Billetta, R; Ferreira, A

    2000-01-01

    We demonstrate that Tc45, a polypeptide described as an immunogenetically restricted Trypanosoma cruzi antigen in mice, is calreticulin, a dimorphic molecule encoded by genes with variable chromosomal distribution. Previously we showed that IgG from A.SW (H2s) mice immunized with T. cruzi trypomastigotes or epimastigotes and sera from infected humans recognize Tc45, a 45 kD parasite polypeptide. Herein we describe the cloning, sequencing, and expression of the Tc45 gene. A 98% homology in the deduced amino acid sequence was found with a T. cruzi calreticulin-like molecule and 41% with Leishmania donovani and human calreticulin. In the T. cruzi CL Brener clone and in the Tulahuén strain, the gene is located in two and four chromosomes, respectively. Calreticulin was detected in several T. cruzi clones, in the Tulahuén strain, and in T. rangeli, displaying alternative 43 and 46 kD forms.

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

    PubMed Central

    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

  11. Serologic survey of antibodies to Trypanosoma cruzi in coyotes and red foxes from Pennsylvania and Tennessee.

    PubMed

    Rosypal, Alexa C; Smith, Trynecia; Alexander, Andrew; Weaver, Melanie; Stewart, Richard; Houston, Allan; Gerhold, Richard; Van Why, Kyle; Dubey, Jitender P

    2014-12-01

    Trypanosoma cruzi is a zoonotic parasite of humans and other mammalian hosts with distribution throughout the Americas. Domestic and wild canine species are reservoirs for human T. cruzi infections. The present study examined the prevalence of antibodies to T. cruzi in wild canids from the United States. Sera from 13 red foxes (Vulpes vulpes) and 263 coyotes (Canis latrans), originating in Pennsylvania and Tennessee, were assayed for antibodies to T. cruzi with immunochromatographic tests. Antibodies to T. cruzi were found in 2 of 276 (0.72%) of all wild canids tested. Both T. cruzi-positive wild canids were coyotes and represented 2 of 21 (9.52%) wild canids assayed from Tennessee. Antibodies to T. cruzi were not detected in red fox. Anti-T. cruzi antibodies were not found in any wild canids from Pennsylvania. These results suggest that coyotes are exposed to T. cruzi in Tennessee but not in Pennsylvania.

  12. Prevalence of antibody to Trypanosoma cruzi in pregnant Hispanic women in Houston.

    PubMed

    Di Pentima, M C; Hwang, L Y; Skeeter, C M; Edwards, M S

    1999-06-01

    We assessed the seroprevalence of antibodies to Trypanosoma cruzi among pregnant Hispanic women in Houston. Sera from 2,107 Hispanic and 1,658 non-Hispanic subjects were tested by ELISA for the presence of antibodies to T. cruzi. Twenty-two (0.6%) of 3,765 subjects had sera that were reactive. Seroreactivity was confirmed by hemagglutination assay. Eleven subjects had reactive sera, giving a confirmed seroprevalence of 0.3% (95% CI, 0-1%). Nine sera from Hispanic and two from non-Hispanic women were positive by hemagglutination assay, for a prevalence of 0.4% and 0.1%, respectively, during pregnancy. On the basis of these seroreactivity data, transplacental transmission of T. cruzi could occur in the continental United States. Screening for antibodies to T. cruzi during pregnancy would provide the potential for early intervention in congenital Chagas' disease.

  13. Therapeutic activity of crude ethanolic extract of Artemisia herba alba against Trypanosoma evansi in rabbits

    NASA Astrophysics Data System (ADS)

    Awad, Fathy M.; Hasan, Zainal Abidin Abu; Osman, Abdinasir Yusuf; Ibrahim, Nazlina

    2013-11-01

    The present work was conducted to evaluate the antitrypanosomal efficacy of crude ethanolic extract (CEE) of the aerial parts of Artemisia herba alba against Trypanosoma evansi infection in an animal model. The results indicated low levels of parasitaemia in rabbits administered with crude ethanolic extract (CEE) compared to those from the negative control group. Similarly, there was also haematologically significant difference (p<0.05) where low mean levels of packed cell volume (PCV) was observed in Groups 1-4 respectively. In contrast, there was no statistically significant difference in almost all investigated parameters between positive control and treatment groups of animals. In conclusion, both CEE of A. herba-alba and Berenil® showed relatively a parasitaemia and normal haematological values in infected rabbits, thereby confirming their antiparasitic properties.

  14. Quercetin induces apoptosis of Trypanosoma brucei gambiense and decreases the proinflammatory response of human macrophages.

    PubMed

    Mamani-Matsuda, Maria; Rambert, Jérôme; Malvy, Denis; Lejoly-Boisseau, Hélène; Daulouède, Sylvie; Thiolat, Denis; Coves, Sara; Courtois, Pierrette; Vincendeau, Philippe; Mossalayi, M Djavad

    2004-03-01

    In addition to parasite spread, the severity of disease observed in cases of human African trypanosomiasis (HAT), or sleeping sickness, is associated with increased levels of inflammatory mediators, including tumor necrosis factor (TNF)-alpha and nitric oxide derivatives. In the present study, quercetin (3,3',4',5,7-pentahydroxyflavone), a potent immunomodulating flavonoid, was shown to directly induce the death of Trypanosoma brucei gambiense, the causative agent of HAT, without affecting normal human cell viability. Quercetin directly promoted T. b. gambiense death by apoptosis as shown by Annexin V binding. In addition to microbicidal activity, quercetin induced dose-dependent decreases in the levels of TNF-alpha and nitric oxide produced by activated human macrophages. These results highlight the potential use of quercetin as an antimicrobial and anti-inflammatory agent for the treatment of African trypanomiasis.

  15. Characterization of the genes specifying two metacyclic variable antigen types in Trypanosoma brucei rhodesiense.

    PubMed Central

    Lenardo, M J; Rice-Ficht, A C; Kelly, G; Esser, K M; Donelson, J E

    1984-01-01

    Bloodstream trypanosomes evade the immune system of their mammalian host by sequentially expressing a large number of different variable surface glycoproteins (VSGs). In contrast, metacyclic trypanosomes, the final developmental stage in the tsetse fly, express a much more restricted set of VSGs. These metacyclic VSGs are the first to be exposed to the immune system of the mammalian host after infection and may offer the potential for the eventual development of a vaccine. We have identified cDNAs for two VSGs in cDNA libraries prepared from amplified metacyclic populations of Trypanosoma brucei rhodesiense and show that they correspond to two different metacyclic serotypes. Determination of the cDNA sequences shows that metacyclic VSG mRNAs are similar to VSG mRNAs expressed during the bloodstream stage. Southern blots demonstrate that the metacyclic VSG genes are located near chromosomal telomeres. No evidence of gene rearrangement associated with expression of these VSGs was found. Images PMID:6593722

  16. Attenuated Salmonella sp. as a DNA Delivery System for Trypanosoma cruzi Antigens.

    PubMed

    Bivona, Augusto E; Cerny, Natacha; Alberti, Andrés Sánchez; Cazorla, Silvia I; Malchiodi, Emilio L

    2016-01-01

    Chagas disease is an important neglected disease affecting thousands of people in the Americas. Novel strategies for prophylactic and therapeutic vaccines against the etiological agent, the intracellular protozoan Trypanosoma cruzi, are urgently needed. Vaccines based on attenuated virus and bacteria as a foreign DNA delivery system represent a strong advantage over naked DNA-based vaccines. Here we describe the use of attenuated Salmonella carrying a eukaryotic expression plasmid encoding a T. cruzi antigen. The main advantages of the methodology are the oral administration of the Salmonella-based vaccine and the induction of a strong humoral and cell-mediated immune response at both mucosal and systemic level, favored by the adjuvant effect elicited by the bacteria pathogen-associated molecular patterns.

  17. Standard culture medium allows clonal dilution of Trypanosoma brucei procyclic cells after auto-conditioning.

    PubMed

    Archer, Stuart K

    2009-03-01

    Trypanosoma brucei can be cultured in vitro in the mammalian bloodstream form or in the procyclic (PC) form found in the insect vector. Bloodstream trypanosomes can be cloned by limiting dilution, but PCs can only be diluted in conditioned medium, i.e., medium in which PC cells have previously been grown. It is shown here that this limitation does not apply to the most commonly used PC cell strain, Lister 427, if free radicals are removed from the medium. The reported benefit of conditioning media may arise in part from a process of hemin-catalysed depletion of peroxide ("auto-conditioning") which occurs during extended incubation at growth temperature. Scavenging free radicals by addition of pyruvate also improves PC cell viability. However, other PC cell strains such as Treu 927 require cell-conditioned media unless grown in a 5% CO2 atmosphere. Several other culture parameters that affect growth rates and dilution capability were identified.

  18. Evidence for a degradosome-like complex in the mitochondria of Trypanosoma brucei

    PubMed Central

    Mattiacio, Jonelle L.; Read, Laurie K.

    2009-01-01

    Mitochondrial RNA turnover in yeast involves the degradosome, composed of DSS-1 exoribonuclease and SUV3 RNA helicase. Here, we describe a degradosome-like complex, containing SUV3 and DSS-1 homologues, in the early branching protozoan, Trypanosoma brucei. TbSUV3 is mitochondrially localized and co-sediments with TbDSS-1 on glycerol gradients. Co-immunoprecipitation demonstrates that TbSUV3 and TbDSS-1 associate in a stable complex, which differs from the yeast degradosome in that it is not stably associated with mitochondrial ribosomes. This is the first report of a mitochondrial degradosome-like complex outside of yeast. Our data indicate an early evolutionary origin for the mitochondrial SUV3/DSS-1 containing complex. PMID:19540236

  19. Mapping of B-Cell Epitopes in a Trypanosoma cruzi Immunodominant Antigen Expressed in Natural Infections

    PubMed Central

    Lesénéchal, Mylène; Becquart, Laurence; Lacoux, Xavier; Ladavière, Laurent; Baida, Renata C. P.; Paranhos-Baccalà, Glaucia; da Silveira, José Franco

    2005-01-01

    Tc40 is an immunodominant antigen present in natural Trypanosoma cruzi infections. This immunogen was thoroughly mapped by using overlapping amino acid sequences identified by gene cloning and chemical peptide synthesis. To map continuous epitopes of the Tc40 antigen, an epitope expression library was constructed and screened with sera from human chagasic patients. A major, linear B-cell epitope spanning residues 403 to 426 (PAKAAAPPAA) was identified in the central domain of Tc40. A synthetic peptide spanning this region reacted strongly with 89.8% of the serum samples from T. cruzi-infected individuals. This indicates that the main antigenic site is defined by the linear sequence of the peptide rather than a conformation-dependent structure. The major B-cell epitope of Tc40 shares a high degree of sequence identity with T. cruzi ribosomal and RNA binding proteins, suggesting the existence of cross-reactivity among these molecules. PMID:15699429

  20. Indomethacin Amides as a Novel Molecular Scaffold for Targeting Trypanosoma cruzi Sterol 14α-Demethylase

    PubMed Central

    Konkle, Mary E.; Hargrove, Tatiana Y.; Kleshchenko, Yuliya Y.; von Kries, Jens P.; Ridenour, Whitney; Uddin, Md. Jashim; Caprioli, Richard M.; Marnett, Lawrence J.; Nes, W. David; Villalta, Fernando; Waterman, Michael R.; Lepesheva, Galina I.

    2009-01-01

    Trypanosoma cruzi (TC) causes Chagas disease, which in its chronic stage remains incurable. We have shown recently that specific inhibition of TC sterol 14α-demethylase (TCCYP51) with imidazole derivatives is effective in killing both extracellular and intracellular human stages of TC. An alternative set of TCCYP51 inhibitors has been identified using optical high throughput screening followed by web-database search for similar structures. The best TCCYP51 inhibitor from this search was found to have structural similarity to a class of cyclooxygenase-2-selective inhibitors, the indomethacin-amides. A number of indomethacin-amides were found to bind to TCCYP51, inhibit its activity in vitro and produce strong antiparasitic effects in the cultured TC cells. Analysis of TC sterol composition indicated that the mode of action of the compounds is by inhibition of sterol biosynthesis in the parasite. PMID:19354253

  1. Trypanosoma cruzi Epimastigotes Are Able to Manage Internal Cholesterol Levels under Nutritional Lipid Stress Conditions.

    PubMed

    Pereira, Miria Gomes; Visbal, Gonzalo; Salgado, Leonardo T; Vidal, Juliana Cunha; Godinho, Joseane L P; De Cicco, Nuccia N T; Atella, Geórgia C; de Souza, Wanderley; Cunha-e-Silva, Narcisa

    2015-01-01

    Trypanosoma cruzi epimastigotes store high amounts of cholesterol and cholesteryl esters in reservosomes. These unique organelles are responsible for cellular digestion by providing substrates for homeostasis and parasite differentiation. Here we demonstrate that under nutritional lipid stress, epimastigotes preferentially mobilized reservosome lipid stocks, instead of lipid bodies, leading to the consumption of parasite cholesterol reservoirs and production of ergosterol. Starved epimastigotes acquired more LDL-NBD-cholesterol by endocytosis and distributed the exogenous cholesterol to their membranes faster than control parasites. Moreover, the parasites were able to manage internal cholesterol levels, alternating between consumption and accumulation. With normal lipid availability, parasites esterified cholesterol exhibiting an ACAT-like activity that was sensitive to Avasimibe in a dose-dependent manner. This result also implies that exogenous cholesterol has a role in lipid reservoirs in epimastigotes.

  2. The sesquiterpene lactone dehydroleucodine (DhL) affects the growth of cultured epimastigotes of Trypanosoma cruzi.

    PubMed

    Brengio, S D; Belmonte, S A; Guerreiro, E; Giordano, O S; Pietrobon, E O; Sosa, M A

    2000-04-01

    Here, we report an inhibitory effect of a sesquiterpene lactone dehydroleucodine (DhL) on the growth of Trypanosoma cruzi in culture. At concentrations of the drug between 5 and 10 microg/ml in the medium,the parasites remained alive for at least 4 days. Higher concentrations of DhL were lethal for the parasites within a few hours. The effect of DhL is irreversible. Morphological changes induced by DhL were also observed in the parasites. The effect of DhL was blocked by the presence of reducing substrates such as glutathione or dithiothreitol, but these agents were not able to reverse the effect of DhL if added 2 days after the start of drug exposure.

  3. High throughput sequencing analysis of Trypanosoma brucei DRBD3/PTB1-bound mRNAs.

    PubMed

    Das, Anish; Bellofatto, Vivian; Rosenfeld, Jeffrey; Carrington, Mark; Romero-Zaliz, Rocío; del Val, Coral; Estévez, Antonio M

    2015-01-01

    Trypanosomes are early-branched eukaryotes that show an unusual dependence on post-transcriptional mechanisms to regulate gene expression. RNA-binding proteins are crucial in controlling mRNA fate in these organisms, but their RNA substrates remain largely unknown. Here we have analyzed on a global scale the mRNAs associated with the Trypanosoma brucei RNA-binding protein DRBD3/PTB1, by capturing ribonucleoprotein complexes using UV cross-linking and subsequent immunoprecipitation. DRBD3/PTB1 associates with many transcripts encoding ribosomal proteins and translation factors. Consequently, silencing of DRBD3/PTB1 expression altered the protein synthesis rate. DRBD3/PTB1 also binds to mRNAs encoding the enzymes required to obtain energy through the oxidation of proline to succinate. We hypothesize that DRBD3/PTB1 is a key player in RNA regulon-based gene control influencing protein synthesis in trypanosomes.

  4. Trypanosoma cruzi: Molecular characterization of an RNA binding protein differentially expressed in the parasite life cycle

    PubMed Central

    Pérez-Díaz, Leticia; Duhagon, María Ana; Smircich, Pablo; Sotelo-Silveira, José; Robello, Carlos; Krieger, Marco Aurelio; Goldenberg, Samuel; Williams, Noreen; Dallagiovanna, Bruno; Garat, Beatriz

    2007-01-01

    Molecular studies have shown several peculiarities in the regulatory mechanisms of gene expression in trypanosomatids. Protein coding genes are organized in long polycistronic units that seem to be constitutively transcribed. Therefore, post-transcriptional regulation of gene expression is considered to be the main point for control of transcript abundance and functionality. Here we describe the characterization of a 17 kDa RNA-binding protein from Trypanosoma cruzi (TcRBP19) containing an RNA recognition motive (RRM). This protein is coded by a single copy gene located in a high molecular weight chromosome of T. cruzi. Orthologous genes are present in the TriTryp genomes. TcRBP19 shows target selectivity since among the different homoribopolymers it preferentially binds polyC. TcRBP19 is a low expression protein only barely detected at the amastigote stage localizing in a diffuse pattern in the cytoplasm. PMID:17475252

  5. Induction and regulation of Trypanosoma brucei VSG-specific antibody responses.

    PubMed

    Black, S J; Guirnalda, P; Frenkel, D; Haynes, C; Bockstal, V

    2010-12-01

    The review addresses how infection with Trypanosoma brucei affects the development, survival and functions of B lymphocytes in mice. It discusses (1) the contributions of antibodies to trypanosome clearance from the bloodstream, (2) how B lymphocytes, the precursors of antibody producing plasma cells, interact with membrane form variable surface glycoprotein (VSG), i.e. with monovalent antigen that is free to diffuse within the lipid bilayer of the trypanosome plasma membrane and consequently can cross-link B cell antigen specific receptors by indirect processes only and (3) the extent and underlying causes of dysregulation of humoral immune responses in infected mice, focusing on the impact of wild type and GPI-PLC⁻/⁻ trypanosomes on bone marrow and extramedullary B lymphopoiesis, B cell maturation and survival.

  6. Consequences of telomere shortening at an active VSG expression site in telomerase-deficient Trypanosoma brucei.

    PubMed

    Dreesen, Oliver; Cross, George A M

    2006-12-01

    Trypanosoma brucei evades the host immune response by sequential expression of a large family of variant surface glycoproteins (VSG) from one of approximately 20 subtelomeric expression sites (ES). VSG transcription is monoallelic, and little is known about the regulation of antigenic switching. To explore whether telomere length could affect antigenic switching, we created a telomerase-deficient cell line, in which telomeres shortened at a rate of 3 to 6 bp at each cell division. Upon reaching a critical length, short silent ES telomeres were stabilized by a telomerase-independent mechanism. The active ES telomere progressively shortened and frequently broke. Upon reaching a critical length, the short active ES telomere stabilized, but the transcribed VSG was gradually lost from the population and replaced by a new VSG through duplicative gene conversion. We propose a model in which subtelomeric-break-induced replication-mediated repair at a short ES telomere leads to duplicative gene conversion and expression of a new VSG.

  7. Trypanosoma brucei: Enrichment by UV of intergenic transcripts from the variable surface glycoprotein gene expression site

    SciTech Connect

    Coquelet, H.; Tebabi, P.; Pays, A.; Steinert, M.; Pays, E. )

    1989-09-01

    The expression site for the variable surface glycoprotein (VSG) gene AnTat 1.3A of Trypanosoma brucei is 45 kilobases long and encompasses seven expression site-associated genes (ESAGs). After UV irradiation, several large transcripts from the putative promoter region were strongly enriched. We report that one such major transcript starts near the poly(A) addition site of the first gene (ESAG 7), spans the intergenic region, and extends to the poly(A) addition site of the second gene (ESAG 6), thus bypassing the normal 3' splice site of the ESAG 6 mRNA. Since this transcript is spliced, we conclude that UV irradiation does not inhibit splicing but stabilizes unstable processing products. This demonstrates that at least some intergenic regions of the VSG gene expression site are continuously transcribed in accordance with a polycistronic transcription model.

  8. Solution structure of the glycosylphosphatidylinositol membrane anchor glycan of Trypanosoma brucei variant surface glycoprotein

    SciTech Connect

    Homans, S.W.; Edge, C.J.; Ferguson, M.A.J.; Dwek, R.A.; Rademacher, T.W. )

    1989-04-04

    The average solution conformation of the glycosylphosphatidylinositol (GPI) membrane anchor of Trypanosoma brucei variant surface glycoprotein (VSG) has been determined by using a combination of two-dimensional {sup 1}H-{sup 1}H NMR methods together with molecular orbital calculations and restrained molecular dynamics simulations. This allows the generation of a model to describe the orientation of the glycan with respect to the membrane. This shows that the glycan exists in an extended configuration along the plane of the membrane and spans an area of 600 {angstrom}{sup 2}, which is similar to the cross-sectional area of a monomeric N-terminal VSG domain. Taken together, these observations suggest a possible space-filling role for the GPI anchor that may maintain the integrity of the VSG coat. The potential importance of the GPI glycan as a chemotherapeutic target is discussed in light of these observations.

  9. Mapping replication dynamics in Trypanosoma brucei reveals a link with telomere transcription and antigenic variation.

    PubMed

    Devlin, Rebecca; Marques, Catarina A; Paape, Daniel; Prorocic, Marko; Zurita-Leal, Andrea C; Campbell, Samantha J; Lapsley, Craig; Dickens, Nicholas; McCulloch, Richard

    2016-05-26

    Survival of Trypanosoma brucei depends upon switches in its protective Variant Surface Glycoprotein (VSG) coat by antigenic variation. VSG switching occurs by frequent homologous recombination, which is thought to require locus-specific initiation. Here, we show that a RecQ helicase, RECQ2, acts to repair DNA breaks, including in the telomeric site of VSG expression. Despite this, RECQ2 loss does not impair antigenic variation, but causes increased VSG switching by recombination, arguing against models for VSG switch initiation through direct generation of a DNA double strand break (DSB). Indeed, we show DSBs inefficiently direct recombination in the VSG expression site. By mapping genome replication dynamics, we reveal that the transcribed VSG expression site is the only telomeric site that is early replicating - a differential timing only seen in mammal-infective parasites. Specific association between VSG transcription and replication timing reveals a model for antigenic variation based on replication-derived DNA fragility.

  10. Gene-deleted live-attenuated Trypanosoma cruzi parasites as vaccines to protect against Chagas disease.

    PubMed

    Sánchez-Valdéz, Fernando J; Pérez Brandán, Cecilia; Ferreira, Arturo; Basombrío, Miguel Ángel

    2015-05-01

    Chagas disease is a neglected tropical disease caused by the protozoan parasite Trypanosoma cruzi. This illness is now becoming global, mainly due to congenital transmission, and so far, there are no prophylactic or therapeutic vaccines available to either prevent or treat Chagas disease. Therefore, different approaches aimed at identifying new protective immunogens are urgently needed. Live vaccines are likely to be more efficient in inducing protection, but safety issues linked with their use have been raised. The development of improved protozoan genetic manipulation tools and genomic and biological information has helped to increase the safety of live vaccines. These advances have generated a renewed interest in the use of genetically attenuated parasites as vaccines against Chagas disease. This review discusses the protective capacity of genetically attenuated parasite vaccines and the challenges and perspectives for the development of an effective whole-parasite Chagas disease vaccine.

  11. Antibody profiles induced by Trypanosoma cruzi in chagasic patients with previous or current exposure to mycobacteria.

    PubMed

    Peverengo, Luz; Prochetto, Estefanía; Rodeles, Luz; Valenzuela, Ignacio; Marcipar, Iván Sergio; Bottasso, Oscar; Vicco, Miguel Hernán

    2016-12-01

    Since the immune response mounted by the host to a particular microorganism might be influenced by the acquired immunological experience due to previous contact with other microorganisms, we performed a cross-sectional study to explore the pattern of Trypanosoma cruzi infection-related antibodies in T. cruzi-seropositive individuals presenting concomitant tuberculosis, or the antecedent of BCG vaccination. Sampled individuals were grouped as follows: patients with Chagas disease, not vaccinated with BCG, who further developed pulmonary tuberculosis; individuals with Chagas disease, BCG-vaccinated; and subjects with Chagas disease, presenting neither BCG scar nor tuberculosis disease. Non-vaccinated individuals or without tuberculosis, presented the highest values of anti-PH (P < 0.001), anti-FRA (P < 0.001), anti-p2β (P = 0.0023) and anti-B13 (P < 0.001) antibodies. The present findings constitute the first demonstration of the potential influence of concomitant tuberculosis on Chagas disease.

  12. The Role of Heme and Reactive Oxygen Species in Proliferation and Survival of Trypanosoma cruzi

    PubMed Central

    Paes, Marcia Cristina; Cosentino-Gomes, Daniela; de Souza, Cíntia Fernandes; Nogueira, Natália Pereira de Almeida; Meyer-Fernandes, José Roberto

    2011-01-01

    Trypanosoma cruzi, the protozoan responsible for Chagas disease, has a complex life cycle comprehending two distinct hosts and a series of morphological and functional transformations. Hemoglobin degradation inside the insect vector releases high amounts of heme, and this molecule is known to exert a number of physiological functions. Moreover, the absence of its complete biosynthetic pathway in T. cruzi indicates heme as an essential molecule for this trypanosomatid survival. Within the hosts, T. cruzi has to cope with sudden environmental changes especially in the redox status and heme is able to increase the basal production of reactive oxygen species (ROS) which can be also produced as byproducts of the parasite aerobic metabolism. In this regard, ROS sensing is likely to be an important mechanism for the adaptation and interaction of these organisms with their hosts. In this paper we discuss the main features of heme and ROS susceptibility in T. cruzi biology. PMID:22007287

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

    PubMed

    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.

  14. [Towards the elimination of the transmission of Trypanosoma cruzi in Honduras and Central American countries].

    PubMed

    Ponce, C

    1999-01-01

    Central America is composed of seven countries: Belice, Costa Rica, El Salvador, Guatemala, Honduras, Nicaragua and Panama. Chagas disease exists in all seven countries, but with major prevalence in El Salvador, Guatemala, Honduras and Nicaragua. The main species of triatomine vectors are: Rhodnius prolixus, Triatoma dimidiata y Rhodnius pallescens. In 1997 the Central American countries launched an Initiative for the Vectorial and Transfusion Transmission Control of Chagas disease. The objectives of the Initiative are: 1. elimination of Rhodnius prolixus, 2. control of Triatoma dimidiata and 3. serological screening for Trypanosoma cruzi of 100% of the blood donors. This Initiative is supported by the Resolution for "The Elimination of Transmission of Chagas Disease", of the World Health Assembly in 1998.

  15. Autonomic Dysfunction and Risk Factors Associated with Trypanosoma cruzi Infection among Children in Arequipa, Peru

    PubMed Central

    Bowman, Natalie M.; Kawai, Vivian; Gilman, Robert H.; Bocangel, Cesar; Galdos-Cardenas, Gerson; Cabrera, Lilia; Levy, Michael Z.; Cornejo del Carpio, Juan Geny; Delgado, Freddy; Rosenthal, Lauren; Pinedo-Cancino, Vivian V.; Steurer, Francis; Seitz, Amy E.; Maguire, James H.; Bern, Caryn

    2011-01-01

    Chagas disease affects an estimated 8 million people in Latin America. Infected individuals have 20–30% lifetime risk of developing cardiomyopathy, but more subtle changes in autonomic responses may be more frequent. We conducted a matched case-control study of children in Arequipa, Peru, where triatomine infestation and Trypanosoma cruzi infection are emerging problems. We collected data on home environment, history, physical examination, electrocardiogram, and autonomic testing. Signs of triatomine infestation and/or animals sleeping in the child's room and household members with Chagas disease were associated with increased infection risk. Electrocardiogram findings did not differ between cases and controls. However, compared with control children, infected children had blunted autonomic responses by three different measures, the Valsalva maneuver, the cold pressor test, and the orthostatic test. T. cruzi-infected children show autonomic dysfunction, although the prognostic value of this finding is not clear. Sustained vector control programs are essential to decreasing future T. cruzi infections. PMID:21212207

  16. Lymphocyte muscarinic cholinergic activity and PGE2 involvement in experimental Trypanosoma cruzi infection.

    PubMed

    Sterin-Borda, L; Gorelik, G; Goren, N; Cappa, S G; Celentano, A M; Borda, E

    1996-11-01

    In this paper, we demonstrated that the production of PGE2 by CD8+ T lymphocytes through muscarinic cholinergic receptor (mAChR) activation of lymphocytes from mice acutely infected with nonlethal Trypanosoma cruzi CA-1 strain could enhance resistance to infection. Treatment in vivo with either atropine or cyclooxygenase inhibitors enhanced mortality rates and parasitemia of mice infected with T. cruzi CA-1 strain. The mechanism by which CD8+ T lymphocytes released PGE2 appears to involve the activation of the cells by circulating IgG present in mice infected with T. cruzi CA-1 strain. Binding of these antibodies to mAChR on CD8+ T lymphocytes triggered the release of large amounts of PGE2. The results point to a role of serum antibodies against mAChR in the protection of T. cruzi infection. The prostanoid acting as an immunomodulator contributed to the maintenance of the chronic course of experimental Chagas disease.

  17. Time-dose response of Trypanosoma congolense bloodstream forms to diminazene and isometamidium.

    PubMed

    Kaminsky, R; Chuma, F; Wasike, R P

    1994-04-01

    Trypanosoma congolense bloodstream forms were propagated in vitro axenically in a simplified cultivation medium at 34 degrees C. Viability of a drug-sensitive and a drug-resistant clone were examined for 10 days following exposure to 0.1, 1.0 and 10.0 micrograms ml-1 of diminazene aceturate and 0.1, 1.0 and 10.0 ng ml-1 of isometamidium chloride for various time intervals. Drug-sensitive T. congolense were irreversibly damaged after incubation with 10 micrograms ml-1 or 1 microgram ml-1 diminazene aceturate for 30 min or 2 h, respectively, while drug-resistant trypanosomes were not affected. Exposure to 10 ng ml-1 isometamidium chloride eliminated drug-sensitive trypanosomes after 24 h and drug-resistant trypanosomes after 96 h. The data obtained on in vitro time-dose responses of T. congolense were related to pharmacokinetic data of diminazene and isometamidium in cattle plasma.

  18. Absence of correlation between karyotype profiles of Trypanosoma congolense and resistance to isometamidium chloride.

    PubMed

    Shahada, Francis; Clausen, Peter-Henning; Tietjen, Uwe; Chuma, Takehisa; Okamoto, Karoku

    2007-07-20

    Chromosome profiles of 10 Trypanosoma (T.) congolense populations with different isometamidium sensitivities were compared using the pulsed field gel electrophoresis technique. The aim was to elucidate whether there was a karyotype pattern specific to eight isometamidium resistant phenotypes. Analysis of the profiles indicated that all populations displayed several discrete bands at the region of small, intermediate and large chromosomes. The highest similarity was observed between two isolates originating from Burkina Faso, indicating that they had the same genetic origin. Other eight strains exhibited different patterns in terms of chromosome size and numbers such that there was no characteristic karyotype pattern that was established specifically to identify resistant populations and discriminate them from the sensitive ones. This study has revealed that isometamidium resistance is not correlated to karyotype profile in T. congolense.

  19. Mechanism of Trypanosoma cruzi Placenta Invasion and Infection: The Use of Human Chorionic Villi Explants

    PubMed Central

    Fretes, Ricardo E.; Kemmerling, Ulrike

    2012-01-01

    Congenital Chagas disease, a neglected tropical disease, endemic in Latin America, is associated with premature labor and miscarriage. During vertical transmission the parasite Trypanosoma cruzi (T. cruzi) crosses the placental barrier. However, the exact mechanism of the placental infection remains unclear. We review the congenital transmission of T. cruzi, particularly the role of possible local placental factors that contribute to the vertical transmission of the parasite. Additionally, we analyze the different methods available for studying the congenital transmission of the parasite. In that context, the ex vivo infection with T. cruzi trypomastigotes of human placental chorionic villi constitutes an excellent tool for studying parasite infection strategies as well as possible local antiparasitic mechanisms. PMID:22701129

  20. The Cyclical Development of Trypanosoma vivax in the Tsetse Fly Involves an Asymmetric Division

    PubMed Central

    Ooi, Cher-Pheng; Schuster, Sarah; Cren-Travaillé, Christelle; Bertiaux, Eloise; Cosson, Alain; Goyard, Sophie; Perrot, Sylvie; Rotureau, Brice

    2016-01-01

    Trypanosoma vivax is the most prevalent trypanosome species in African cattle. It is thought to be transmitted by tsetse flies after cyclical development restricted to the vector mouthparts. Here, we investigated the kinetics of T. vivax development in Glossina morsitans morsitans by serial dissections over 1 week to reveal differentiation and proliferation stages. After 3 days, stable numbers of attached epimastigotes were seen proliferating by symmetric division in the cibarium and proboscis, consistent with colonization and maintenance of a parasite population for the remaining lifespan of the tsetse fly. Strikingly, some asymmetrically dividing cells were also observed in proportions compatible with a continuous production of pre- metacyclic trypomastigotes. The involvement of this asymmetric division in T. vivax metacyclogenesis is discussed and compared to other trypanosomatids. PMID:27734008

  1. Familial analysis of seropositivity to Trypanosoma cruzi and of clinical forms of Chagas disease.

    PubMed

    Silva-Grecco, Roseane L; Balarin, Marly A S; Correia, Dalmo; Prata, Aluízio; Rodrigues, Virmondes

    2010-01-01

    A cross-sectional study was carried out in Agua Comprida, MG, Brazil, a region previously endemic to Chagas disease whose vectorial transmission was interrupted around 20 year ago. A total of 998 individuals were examined for anti-Trypanosoma cruzi antibodies. Seropositivity was observed in 255 subjects (25.5%), and 743 subjects were negative. Forty-one families with 5-80 individuals with similar environmental conditions were selected for familial analysis. In 15 families, seropositivity to T. cruzi was observed in > 50% of individuals. The segregation analysis confirmed family aggregation for the seropositivity to the T. cruzi. Heart commitment was the major clinical form observed, and in six families, > 50% of the individuals display cardiopathy that may be attributed to T. cruzi infection. Our results support the hypothesis that there is a family aggregation for the seropositivity but without the effect of one major gene.

  2. Characterization of Trypanosoma brucei gambiense variant surface glycoprotein LiTat 1.5.

    PubMed

    Van Nieuwenhove, L; Rogé, S; Lejon, V; Guisez, Y; Büscher, P

    2012-05-09

    At present, all available diagnostic antibody detection tests for Trypanosoma brucei gambiense human African trypanosomiasis are based on predominant variant surface glycoproteins (VSGs), such as VSG LiTat 1.5. During investigations aiming at replacement of the native VSGs by recombinant proteins or synthetic peptides, the sequence of VSG LiTat 1.5 was derived from cDNA and direct N-terminal amino acid sequencing. Characterization of the VSG based on cysteine distribution in the amino acid sequence revealed an unusual cysteine pattern identical to that of VSG Kinu 1 of T. b. brucei. Even though both VSGs lack the third of four conserved cysteines typical for type A N-terminal domains, they can be classified as type A.

  3. RAP1 is essential for silencing telomeric variant surface glycoprotein genes in Trypanosoma brucei.

    PubMed

    Yang, Xiaofeng; Figueiredo, Luisa M; Espinal, Amin; Okubo, Eiji; Li, Bibo

    2009-04-03

    Trypanosoma brucei expresses variant surface glycoprotein (VSG) genes in a strictly monoallelic fashion in its mammalian hosts, but it is unclear how this important virulence mechanism is enforced. Telomere position effect, an epigenetic phenomenon, has been proposed to play a critical role in VSG regulation, yet no telomeric protein has been identified whose disruption led to VSG derepression. We now identify tbRAP1 as an intrinsic component of the T. brucei telomere complex and a major regulator for silencing VSG expression sites (ESs). Knockdown of tbRAP1 led to derepression of all VSGs in silent ESs, but not VSGs located elsewhere, and resulted in stronger derepression of genes located within 10 kb from telomeres than genes located further upstream. This graduated silencing pattern suggests that telomere integrity plays a key role in tbRAP1-dependent silencing and VSG regulation.

  4. Delineation of the regulated Variant Surface Glycoprotein gene expression site domain of Trypanosoma brucei.

    PubMed

    Sheader, Karen; Berberof, Magali; Isobe, Tomoko; Borst, Piet; Rudenko, Gloria

    2003-05-01

    The African trypanosome Trypanosoma brucei is protected in the bloodstream of the mammalian host by a dense Variant Surface Glycoprotein (VSG) coat. Although an individual cell has hundreds of VSG genes, the active VSG is transcribed in a mutually exclusive fashion from one of about twenty telomeric VSG expression sites. Expression sites are regulated domains flanked by 50 bp repeat arrays and extensive tracts of repetitive elements. We have integrated exogenous rDNA and expression site promoters upstream of the 50 bp repeats of the VO2 VSG expression site. Transcription from both types of exogenous promoter is downregulated and comparable to promoters targeted into the VSG Basic Copy arrays. We show that the upstream exogenous rDNA promoter escapes VSG expression site control, as switching the downstream VO2 VSG expression site on and off does not affect its activity. Therefore, the 50 bp repeat arrays appear to be the boundary of the regulated expression site domain.

  5. A VSG expression site-associated gene confers resistance to human serum in Trypanosoma rhodesiense.

    PubMed

    Xong, H V; Vanhamme, L; Chamekh, M; Chimfwembe, C E; Van Den Abbeele, J; Pays, A; Van Meirvenne, N; Hamers, R; De Baetselier, P; Pays, E

    1998-12-11

    Infectivity of Trypanosoma brucei rhodesiense to humans is due to its resistance to a lytic factor present in human serum. In the ETat 1 strain this character was associated with antigenic variation, since expression of the ETat 1.10 variant surface glycoprotein was required to generate resistant (R) clones. In addition, in this strain transcription of a gene termed SRA was detected in R clones only. We show that the ETat 1.10 expression site is the one selectively transcribed in R variants. This expression site contains SRA as an expression site-associated gene (ESAG) and is characterized by the deletion of several ESAGs. Transfection of SRA into T.b. brucei was sufficient to confer resistance to human serum, identifying this gene as one of those responsible for T.b. rhodesiense adaptation to humans.

  6. Trypanosoma rangeli: a new perspective for studying the modulation of immune reactions of Rhodnius prolixus

    PubMed Central

    Garcia, Eloi S; Castro, Daniele P; Figueiredo, Marcela B; Genta, Fernando A; Azambuja, Patrícia

    2009-01-01

    Insects are exposed to a wide range of microorganisms (bacteria, fungi, parasites and viruses) and have interconnected powerful immune reactions. Although insects lack an acquired immune system they have well-developed innate immune defences that allow a general and rapid response to infectious agents. Over the last few decades we have observed a dramatic increase in the knowledge of insect innate immunity, which relies on both humoral and cellular responses. However, innate reactions to natural insect pathogens and insect-transmitted pathogens, such as parasites, still remain poorly understood. In this review, we briefly introduce the general immune system of insects and highlight our current knowledge of these reactions focusing on the interactions of Trypanosoma rangeli with Rhodnius prolixus, an important model for innate immunity investigation. PMID:19615044

  7. Assembly mechanism of Trypanosoma brucei BILBO1 at the flagellar pocket collar.

    PubMed

    Vidilaseris, Keni; Lesigang, Johannes; Morriswood, Brooke; Dong, Gang

    2015-01-01

    The flagellar pocket is a bulb-like invagination of the plasma membrane that encloses the base of the single flagellum in trypanosomes. It is the site of all endo- and exocytic activity in the parasite and has thus been proposed to be a therapeutic target. At the neck of the flagellar pocket is an electron-dense cytoskeletal structure named the flagellar pocket collar. The protein BILBO1 was the first characterized and remains the only known component of the flagellar pocket collar, with essential functions in the biogenesis of both the flagellar pocket and flagellar pocket collar. We recently reported that the filamentous assembly of Trypanosoma brucei BILBO1 (TbBILBO1) is mediated by its central coiled coil domain and C-terminal leucine zipper. Here, we discuss how TbBILBO1 might assemble at the flagellar pocket collar in T. brucei.

  8. In vitro activity of Etanidazole against the protozoan parasite Trypanosoma cruzi.

    PubMed

    Petray, Patricia B; Morilla, María J; Corral, Ricardo S; Romero, Eder L

    2004-03-01

    We investigated the in vitro action of an hydrosoluble 2-nitroimidazole, Etanidazole (EZL), against Trypanosoma cruzi, the etiologic agent of Chagas disease. EZL displayed lethal activity against isolated trypomastigotes as well as amastigotes of T. cruzi (RA strain) growing in Vero cells or J774 macrophages, without affecting host cell viability. Although not completely equivalent to Benznidazole (BZL), the reference drug for Chagas chemotherapy, EZL takes advantage in exerting its anti-T. cruzi activity for longer periods without serious toxic side effects, as those recorded in BZL-treated patients. Our present results encourage further experiments to study in depth the trypanocidal properties of this drug already licensed for use in human cancers.

  9. Solution structure of Q388A3 PDZ domain from Trypanosoma brucei.

    PubMed

    Mei, Song; Dong, Yuanqiu; Zhang, Jiahai; Zhang, Xuecheng; Tu, Xiaoming

    2016-05-01

    PDZ domains are abundant protein interaction modules that often recognize short amino acid motifs at the C-termini of target proteins and regulate multiple biological processes. So far, no PDZ domain in Trypanosoma brucei, an eukaryotic parasite causing sleeping sickness, has been studied. Q388A3, conserved in the related kinetoplastid parasites, is a 1634-residue protein containing a PDZ domain at its C-terminus. In this work, the solution structure of Q388A3 PDZ domain was solved by NMR spectroscopy. Q388A3 PDZ domain adopts a PDZ-like fold composed by a five-stranded β-sheet capped by two α-helices, which is similar to the PDZ domains from HtrA family proteins. Meanwhile, Q388A3 PDZ domain shows some structural features quite different from HtrA PDZ domain.

  10. Trypanosoma cruzi parasites fight for control of the JAK-STAT pathway by disarming their host

    PubMed Central

    Stahl, Philipp; Schwarz, Ralph T; Debierre-Grockiego, Françoise; Meyer, Thomas

    2014-01-01

    The zoonotic Chagas’ disease is caused by infections with the hemoflagellate Trypanosoma cruzi (T. cruzi) which is endemic in Latin America. Despite recent advances in our understanding of the pathogenesis of the disease, the underlying molecular processes involved in host-parasite interactions are only poorly understood. In particular, the mechanisms for parasite persistence in host cells remain largely unknown. Cytokine-driven transcription factors from the family of STAT (signal transducer and activator of transcription) proteins appear to play a central role in the fight against T. cruzi infection. However, amastigotes proliferating in the cytoplasm of infected host cells develop effective strategies to circumvent the attack executed by STAT proteins. This review highlights the interactions between T. cruzi parasites and human host cells in terms of cytokine signaling and, in particular, discusses the impact of STATs on the balance between parasite invasion and clearance. PMID:26413423

  11. Trypanosoma cruzi: dehydroepiandrosterone (DHEA) and immune response during the chronic phase of the experimental Chagas' disease.

    PubMed

    Caetano, Leony Cristina; Santello, Fabricia Helena; Del Vecchio Filipin, Marina; Brazão, Vânia; Caetano, Luana Naiara; Toldo, Miriam Paula Alonso; Caldeira, Jerri C; do Prado Júnior, José Clóvis

    2009-07-07

    Dehydroepiandrosterone (DHEA) has long been considered as a precursor for many steroid hormones. It also enhances the immune responses against a wide range of viral, bacterial, and parasitic pathogens. The aims of this work were to evaluate the influences of exogenous DHEA treatment on Wistar rats infected with the Y strain of Trypanosoma cruzi during the acute and its influence on the chronic phase of infection. Animals were subcutaneous treated with 40 mg/kg body weight/day of DHEA. DHEA treatment promoted increased lymphoproliferative responses as well as enhanced concentrations of NO and IL-12. So, we point in the direction that our results validate the utility of the use of DHEA as an alternative therapy candidate against T. cruzi.

  12. Trypanosoma cruzi- specific immune responses in subjects from endemic areas of Chagas disease of Argentina

    PubMed Central

    Olivera, Gabriela C.; Albareda, Maria C.; Alvarez, Maria G.; De Rissio, Ana M.; Fichera, Laura E.; Cooley, Gretchen; Yachelini, Pedro; Hrellac, Hugo A.; Riboldi, Hilda; Laucella, Susana A.; Tarleton, Rick L.; Postan, Miriam

    2010-01-01

    Trypanosoma cruzi-specific immune responses were evaluated in a total of 88 subjects living in areas endemic of Chagas disease of Argentina by IFN-γ ELISPOT assays and immunoblotting. Positive T. cruzi antigen-induced IFN-γ responses were detected in 42% of subjects evaluated (15/26 positive by conventional serology and 22/62 seronegative subjects). Using immunoblotting, T. cruzi-specific IgG reactivity was detected in all seropositive subjects and in 11% (7/61) of subjects negative by conventional serology. Measurements of T cell responses and antibodies by immunoblotting, in conjunction with conventional serology, might enhance the capability of detection of exposure to T. cruzi in endemic areas. PMID:20123034

  13. Deciphering RNA Regulatory Elements Involved in the Developmental and Environmental Gene Regulation of Trypanosoma brucei.

    PubMed

    Gazestani, Vahid H; Salavati, Reza

    2015-01-01

    Trypanosoma brucei is a vector-borne parasite with intricate life cycle that can cause serious diseases in humans and animals. This pathogen relies on fine regulation of gene expression to respond and adapt to variable environments, with implications in transmission and infectivity. However, the involved regulatory elements and their mechanisms of actions are largely unknown. Here, benefiting from a new graph-based approach for finding functional regulatory elements in RNA (GRAFFER), we have predicted 88 new RNA regulatory elements that are potentially involved in the gene regulatory network of T. brucei. We show that many of these newly predicted elements are responsive to both transcriptomic and proteomic changes during the life cycle of the parasite. Moreover, we found that 11 of predicted elements strikingly resemble previously identified regulatory elements for the parasite. Additionally, comparison with previously predicted motifs on T. brucei suggested the superior performance of our approach based on the current limited knowledge of regulatory elements in T. brucei.

  14. Sec16 determines the size and functioning of the Golgi in the protist parasite, Trypanosoma brucei.

    PubMed

    Sealey-Cardona, Marco; Schmidt, Katy; Demmel, Lars; Hirschmugl, Tatjana; Gesell, Tanja; Dong, Gang; Warren, Graham

    2014-06-01

    The Sec16 homologue in Trypanosoma brucei has been identified and characterized. TbSec16 colocalizes with COPII components at the single endoplasmic reticulum exit site (ERES), which is next to the single Golgi stack in the insect (procyclic) form of this organism. Depletion of TbSec16 reduces the size of the ERES and the Golgi, and slows growth and transport of a secretory marker to the cell surface; conversely, overexpression of TbSec16 increases the size of the ERES and Golgi but has no effect on growth or secretion. Together these data suggest that TbSec16 regulates the size of the ERES and Golgi and this size is set for optimal growth of the organism.

  15. Trypanosoma cruzi-associated cerebrovascular disease: a case-control study in Eastern Colombia.

    PubMed

    Leon-Sarmiento, Fidias E; Mendoza, Eder; Torres-Hillera, Martin; Pinto, Neyla; Prada, Janette; Silva, Clara A; Vera, Silvia J; Castillo, Erwin; Valderrama, Vladimir; Prada, Didier G; Bayona-Prieto, Jaime; Garcia, Ingrid

    2004-01-15

    Trypanosoma cruzi infection is a common cause of cardiopathy in South America leading it eventually to an established stroke; however, the association between T. cruzi infection itself and cerebrovascular disease is still unknown. We did a case-control study at Eastern Colombia and found that T. cruzi infection was more frequent and statistically significant in stroke cases (24.4%) than controls (1.9%), (Chi square: 21.72; OR: 16.13; 95% confidence interval (CI): 3.64-71.4; p<0.00001). After removing the seropositive patients with cardiological abnormalities, the significance still remained by multivariate analysis (p<0.05). This is the first case-control study that demonstrated a significant link between this infection and symptomatic cerebrovascular disease, mainly ischemic, regardless of cardiac abnormalities. Therefore, we recommend that patients with stroke must be screened for T. cruzi infection if they currently live or have lived in places where this parasite is considered endemic.

  16. Heterologous expression of a plant arginine decarboxylase gene in Trypanosoma cruzi.

    PubMed

    Carrillo, Carolina; Serra, María P; Pereira, Claudio A; Huber, Alejandra; González, Nélida S; Algranati, Israel D

    2004-11-01

    Wild-type Trypanosoma cruzi epimastigotes lack arginine decarboxylase (ADC) enzymatic activity. However, the transformation of these parasites with a recombinant plasmid containing the oat ADC cDNA coding region gave rise to the transient heterologous expression of the enzyme, suggesting the absence of endogenous mechanisms that could inhibit the expression of a hypothetical own ADC gene or the assay used to measure its enzymatic activity. The foreign ADC enzyme expressed in the transgenic T. cruzi was characterized by identification of the products, the stoichiometry of the catalysed reaction, the specific inhibition by alpha-difluoromethylarginine (DFMA) and the study of its metabolic turnover. The half-life of the heterologous ADC activity in T. cruzi was about 150 min. Bioinformatics studies and polymerase chain reaction (PCR) analyses seem to indicate the absence of ADC-like DNA sequences in the wild-type T. cruzi genome.

  17. Trypanosoma cruzi P21: a potential novel target for chagasic cardiomyopathy therapy

    PubMed Central

    Teixeira, Thaise Lara; Machado, Fabrício Castro; Alves da Silva, Aline; Teixeira, Samuel Cota; Borges, Bruna Cristina; dos Santos, Marlus Alves; Martins, Flávia Alves; Brígido, Paula Cristina; Rodrigues, Adele Aud; Notário, Ana Flávia Oliveira; Ferreira, Bruno Antônio; Servato, João Paulo Silva; Deconte, Simone Ramos; Lopes, Daiana Silva; Ávila, Veridiana Melo Rodrigues; Araújo, Fernanda de Assis; Tomiosso, Tatiana Carla; Silva, Marcelo José Barbosa; da Silva, Claudio Vieira

    2015-01-01

    Chagas disease, which is caused by the parasite Trypanosoma cruzi, is an important cause of cardiomyopathy in Latin America. It is estimated that 10%–30% of all infected individuals will acquire chronic chagasic cardiomyopathy (CCC). The etiology of CCC is multifactorial and involves parasite genotype, host genetic polymorphisms, immune response, signaling pathways and autoimmune progression. Herein we verified the impact of the recombinant form of P21 (rP21), a secreted T. cruzi protein involved in host cell invasion, on progression of inflammatory process in a polyester sponge-induced inflammation model. Results indicated that rP21 can recruit immune cells induce myeloperoxidase and IL-4 production and decrease blood vessels formation compared to controls in vitro and in vivo. In conclusion, T. cruzi P21 may be a potential target for the development of P21 antagonist compounds to treat chagasic cardiomyopathy. PMID:26574156

  18. Iron superoxide dismutases in eukaryotic pathogens: new insights from Apicomplexa and Trypanosoma structures.

    PubMed

    Phan, Isabelle Q H; Davies, Douglas R; Moretti, Nilmar Silvio; Shanmugam, Dhanasekaran; Cestari, Igor; Anupama, Atashi; Fairman, James W; Edwards, Thomas E; Stuart, Kenneth; Schenkman, Sergio; Myler, Peter J

    2015-05-01

    Prior studies have highlighted the potential of superoxide dismutases as drug targets in eukaryotic pathogens. This report presents the structures of three iron-dependent superoxide dismutases (FeSODs) from Trypanosoma cruzi, Leishmania major and Babesia bovis. Comparison with existing structures from Plasmodium and other trypanosome isoforms shows a very conserved overall fold with subtle differences. In particular, structural data suggest that B. bovis FeSOD may display similar resistance to peroxynitrite-mediated inactivation via an intramolecular electron-transfer pathway as previously described in T. cruzi FeSOD isoform B, thus providing valuable information for structure-based drug design. Furthermore, lysine-acetylation results in T. cruzi indicate that acetylation occurs at a position close to that responsible for the regulation of acetylation-mediated activity in the human enzyme.

  19. The natural compounds piperovatine and piperlonguminine induce autophagic cell death on Trypanosoma cruzi.

    PubMed

    Veiga-Santos, Phercyles; Desoti, Vânia Cristina; Miranda, Nathielle; Ueda-Nakamura, Tânia; Dias-Filho, Benedito Prado; Silva, Sueli Oliveira; Cortez, Diogenes Aparício Garcia; de Mello, João Carlos Palazzo; Nakamura, Celso Vataru

    2013-03-01

    The currently available treatments for Chagas disease show limited therapeutic potential and are associated with serious side effects. Our group has been attempting to find alternative drugs isolated from natural products as a potential source of pharmacological agents against Trypanosoma cruzi. Here, we demonstrate the antitrypanosomal activity of the amides piperovatine and piperlonguminine isolated from Piper ovatum against epimastigotes and intracellular amastigotes. We also investigated the mechanisms of action of these compounds on extracellular amastigote and epimastigote forms of T. cruzi. These amides showed low toxicity to LLCMK(2) mammalian cells. By using transmission and scanning electron microscopy, we observed that the compounds caused severe alterations in T. cruzi. These alterations were mainly located in plasma membrane and mitochondria. Furthermore, the study of treated parasites labeled with Rh123, PI and MDC corroborate with our TEM data. These mitochondrial dysfunctions induced by the amides might trigger biochemical alterations that lead to cell death. Altogether, our data evidence a possible autophagic process.

  20. Oral exposure to Phytomonas serpens attenuates thrombocytopenia and leukopenia during acute infection with Trypanosoma cruzi.

    PubMed

    da Silva, Rosiane V; Malvezi, Aparecida D; Augusto, Leonardo da Silva; Kian, Danielle; Tatakihara, Vera Lúcia H; Yamauchi, Lucy M; Yamada-Ogatta, Sueli F; Rizzo, Luiz V; Schenkman, Sergio; Pinge-Filho, Phileno

    2013-01-01

    Mice infected with Trypanosoma cruzi, the agent of Chagas disease, rapidly develop anemia and thrombocytopenia. These effects are partially promoted by the parasite trans-sialidase (TS), which is shed in the blood and depletes sialic acid from the platelets, inducing accelerated platelet clearance and causing thrombocytopenia during the acute phase of disease. Here, we demonstrate that oral immunization of C57BL/6 mice with Phytomonas serpens, a phytoflagellate parasite that shares common antigens with T. cruzi but has no TS activity, reduces parasite burden and prevents thrombocytopenia and leukopenia. Immunization also reduces platelet loss after intraperitoneal injection of TS. In addition, passive transfer of immune sera raised in mice against P. serpens prevented platelet clearance. Thus, oral exposure to P. serpens attenuates the progression of thrombocytopenia induced by TS from T. cruzi. These findings are not only important for the understanding of the pathogenesis of T. cruzi infection but also for developing novel approaches of intervention in Chagas disease.