RESUMEN
Trypanosoma cruzi, the protozoan causative of Chagas disease (ChD), exhibits striking genetic and phenotypic intraspecific diversity, along with ecoepidemiological complexity. Human-pathogen interactions lead to distinct clinical presentations of ChD. In 2009, an international consensus classified T. cruzi strains into six discrete typing units (DTUs), TcI to TcVI, later including TcBat, and proposed reproducible genotyping schemes for DTU identification. This article aims to review the impact of classifying T. cruzi strains into DTUs on our understanding of biological, ecoepidemiological, and pathogenic aspects of T. cruzi. We will explore the likely origin of DTUs and the intrinsic characteristics of each group of strains concerning genome organization, genomics, and susceptibility to drugs used in ChD treatment. We will also provide an overview of the association of DTUs with mammalian reservoirs, and summarize the geographic distribution, and the clinical implications, of prevalent specific DTUs in ChD patients. Throughout this review, we will emphasize the crucial roles of both parasite and human genetics in defining ChD pathogenesis and chemotherapy outcome.
RESUMEN
In recent years, proteasome involvement in the damage response induced by ionizing radiation (IR) became evident. However, whether proteasome plays a direct or indirect role in IR-induced damage response still unclear. Trypanosoma cruzi is a human parasite capable of remarkable high tolerance to IR, suggesting a highly efficient damage response system. Here, we investigate the role of T. cruzi proteasome in the damage response induced by IR. We exposed epimastigotes to high doses of gamma ray and we analyzed the expression and subcellular localization of several components of the ubiquitin-proteasome system. We show that proteasome inhibition increases IR-induced cell growth arrest and proteasome-mediated proteolysis is altered after parasite exposure. We observed nuclear accumulation of 19S and 20S proteasome subunits in response to IR treatments. Intriguingly, the dynamic of 19S particle nuclear accumulation was more similar to the dynamic observed for Rad51 nuclear translocation than the observed for 20S. In the other hand, 20S increase and nuclear translocation could be related with an increase of its regulator PA26 and high levels of proteasome-mediated proteolysis in vitro. The intersection between the opposed peaks of 19S and 20S protein levels was marked by nuclear accumulation of both 20S and 19S together with Ubiquitin, suggesting a role of ubiquitin-proteasome system in the nuclear protein turnover at the time. Our results revealed the importance of proteasome-mediated proteolysis in T. cruzi IR-induced damage response suggesting that proteasome is also involved in T. cruzi IR tolerance. Moreover, our data support the possible direct/signaling role of 19S in DNA damage repair. Based on these results, we speculate that spatial and temporal differences between the 19S particle and 20S proteasome controls proteasome multiple roles in IR damage response.
Asunto(s)
Complejo de la Endopetidasa Proteasomal/metabolismo , Radiación Ionizante , Trypanosoma cruzi/metabolismo , Trypanosoma cruzi/efectos de la radiación , Ubiquitina/metabolismo , Reparación del ADN , Proteolisis , Respuesta de Proteína DesplegadaRESUMEN
BACKGROUND: Trypanosoma cruzi, the etiologic agent of Chagas disease, is currently divided into six discrete typing units (DTUs), named TcI-TcVI. CL Brener, the reference strain of the T. cruzi genome project, is a hybrid with a genome assembled into 41 putative chromosomes. Gene copy number variation (CNV) is well documented as an important mechanism to enhance gene expression and variability in T. cruzi. Chromosomal CNV (CCNV) is another level of gene CNV in which whole blocks of genes are expanded simultaneously. Although the T. cruzi karyotype is not well defined, several studies have demonstrated a significant variation in the size and content of chromosomes between different T. cruzi strains. Despite these studies, the extent of diversity in CCNV among T. cruzi strains based on a read depth coverage analysis has not been determined. RESULTS: We identify the CCNV in T. cruzi strains from the TcI, TcII and TcIII DTUs, by analyzing the depth coverage of short reads from these strains using the 41 CL Brener chromosomes as reference. This study led to the identification of a broader extent of CCNV in T. cruzi than was previously speculated. The TcI DTU strains have very few aneuploidies, while the strains from TcII and TcIII DTUs present a high degree of chromosomal expansions. Chromosome 31, which is the only chromosome that is supernumerary in all six T. cruzi samples evaluated in this study, is enriched with genes related to glycosylation pathways, highlighting the importance of glycosylation to parasite survival. CONCLUSIONS: Increased gene copy number due to chromosome amplification may contribute to alterations in gene expression, which represents a strategy that may be crucial for parasites that mainly depend on post-transcriptional mechanisms to control gene expression.
Asunto(s)
Variaciones en el Número de Copia de ADN/genética , Genoma de Protozoos/genética , Trypanosoma cruzi/genética , ADN Protozoario/genética , Expresión Génica/genética , Variación Genética/genética , Genómica/métodos , GlicosilaciónRESUMEN
Schistosomiasis is a neglected tropical disease, and after malaria, is the second most important tropical disease in public health. A vaccine that reduces parasitemia is desirable to achieve mass treatment with a low cost. Although potential antigens have been identified and tested in clinical trials, no effective vaccine against schistosomiasis is available. Y-box-binding proteins (YBPs) regulate gene expression and participate in a variety of cellular processes, including transcriptional and translational regulation, DNA repair, cellular proliferation, drug resistance, and stress responses. The Schistosoma mansoni ortholog of the human YB-1, SMYB1, is expressed in all stages of the parasite life cycle. Although SMYB1 binds to DNA or RNA oligonucleotides, immunohistochemistry assays demonstrated that it is primarily localized in the cytoplasm of parasite cells. In addition, SMYB1 interacts with a protein involved in mRNA processing, suggesting that SMYB1 functions in the turnover, transport, and/or stabilization of RNA molecules during post-transcriptional gene regulation. Here we report the potential of SMYB1 as a vaccine candidate. We demonstrate that recombinant SMYB1 stimulates the production of high levels of specific IgG1 antibodies in a mouse model. The observed levels of specific IgG1 and IgG2a antibodies indicate an actual protection against cercariae challenge. Animals immunized with rSMYB1 exhibited a 26% reduction in adult worm burden and a 28% reduction in eggs retained in the liver. Although proteins from the worm tegument are considered optimal targets for vaccine development, this study demonstrates that unexposed cytoplasmic proteins can reduce the load of intestinal worms and the number of eggs retained in the liver.
RESUMEN
THE SPLICED LEADER (SL) IS A GENE THAT GENERATES A FUNCTIONAL NCRNA THAT IS COMPOSED OF TWO REGIONS: an intronic region of unknown function (SLi) and an exonic region (SLe), which is transferred to the 5' end of independent transcripts yielding mature mRNAs, in a process known as spliced leader trans-splicing (SLTS). The best described function for SLTS is to solve polycistronic transcripts into monocistronic units, specifically in Trypanosomatids. In other metazoans, it is speculated that the SLe addition could lead to increased mRNA stability, differential recruitment of the translational machinery, modification of the 5' region or a combination of these effects. Although important aspects of this mechanism have been revealed, several features remain to be elucidated. We have analyzed 157 SLe sequences from 148 species from seven phyla and found a high degree of conservation among the sequences of species from the same phylum, although no considerable similarity seems to exist between sequences of species from different phyla. When analyzing case studies, we found evidence that a given SLe will always be related to a given set of transcripts in different species from the same phylum, and therefore, different SLe sequences from the same species would regulate different sets of transcripts. In addition, we have observed distinct transcript categories to be preferential targets for the SLe addition in different phyla. This work sheds light into crucial and controversial aspects of the SLTS mechanism. It represents a comprehensive study concerning various species and different characteristics of this important post-transcriptional regulatory mechanism.
RESUMEN
The main consequence of oxidative stress is the formation of DNA lesions, which can result in genomic instability and lead to cell death. Guanine is the base that is most susceptible to oxidation, due to its low redox potential, and 8-oxoguanine (8-oxoG) is the most common lesion. These characteristics make 8-oxoG a good cellular biomarker to indicate the extent of oxidative stress. If not repaired, 8-oxoG can pair with adenine and cause a G:C to T:A transversion. When 8-oxoG is inserted during DNA replication, it could generate double-strand breaks, which makes this lesion particularly deleterious. Trypanosoma cruzi needs to address various oxidative stress situations, such as the mammalian intracellular environment and the triatomine insect gut where it replicates. We focused on the MutT enzyme, which is responsible for removing 8-oxoG from the nucleotide pool. To investigate the importance of 8-oxoG during parasite infection of mammalian cells, we characterized the MutT gene in T. cruzi (TcMTH) and generated T. cruzi parasites heterologously expressing Escherichia coli MutT or overexpressing the TcMTH enzyme. In the epimastigote form, the recombinant and wild-type parasites displayed similar growth in normal conditions, but the MutT-expressing cells were more resistant to hydrogen peroxide treatment. The recombinant parasite also displayed significantly increased growth after 48 hours of infection in fibroblasts and macrophages when compared to wild-type cells, as well as increased parasitemia in Swiss mice. In addition, we demonstrated, using western blotting experiments, that MutT heterologous expression can influence the parasite antioxidant enzyme protein levels. These results indicate the importance of the 8-oxoG repair system for cell viability.
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Daño del ADN , Guanina/análogos & derivados , Estrés Oxidativo , Trypanosoma cruzi/fisiología , Animales , Supervivencia Celular , Células Cultivadas , Enfermedad de Chagas/parasitología , Enfermedad de Chagas/patología , Modelos Animales de Enfermedad , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Femenino , Fibroblastos/parasitología , Expresión Génica , Guanina/metabolismo , Peróxido de Hidrógeno/toxicidad , Macrófagos/parasitología , Ratones , Datos de Secuencia Molecular , Oxidorreductasas actuantes sobre Donantes de Grupo CH-NH/genética , Oxidorreductasas actuantes sobre Donantes de Grupo CH-NH/metabolismo , Parasitemia/parasitología , Parasitemia/patología , Pirofosfatasas/genética , Pirofosfatasas/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Análisis de Secuencia de ADN , Trypanosoma cruzi/efectos de los fármacos , Trypanosoma cruzi/enzimología , Trypanosoma cruzi/crecimiento & desarrolloRESUMEN
SMYB1 is a Schistosoma mansoni protein highly similar to members of the Y-box binding protein family. Similar to other homologues, SMYB1 is able to bind double- and single-stranded DNA, as well as RNA molecules. The characterization of proteins involved in the regulation of gene expression in S. mansoni is of great importance for the understanding of molecular events that control morphological and physiological changes in this parasite. Here we demonstrate that SMYB1 is located in the cytoplasm of cells from different life-cycle stages of S. mansoni, suggesting that this protein is probably acting in mRNA metabolism in the cytoplasm and corroborating previous findings from our group that showed its ability to bind RNA. Protein-protein interactions are important events in all biological processes, since most proteins execute their functions through large supramolecular structures. Yeast two-hybrid screenings using SMYB1 as bait identified a partner in S. mansoni similar to the SmD3 protein of Drosophila melanogaster (SmRNP), which is important in the assembly of small nuclear ribonucleoprotein complexes. Also, pull-down assays were conducted using immobilized GST-SMYB1 proteins and confirmed the SMYB1-SmRNP interaction. The interaction of SMYB1 with a protein involved in mRNA processing suggests that it may act in processes such as turnover, transport and stabilization of RNA molecules.
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Proteínas del Helminto/metabolismo , ARN de Helminto/metabolismo , ARN Mensajero/metabolismo , Schistosoma mansoni/metabolismo , Animales , Anticuerpos Antihelmínticos/sangre , Anticuerpos Antihelmínticos/inmunología , Transporte Biológico , Citoplasma/metabolismo , Femenino , Biblioteca de Genes , Proteínas del Helminto/genética , Inmunohistoquímica , Masculino , ARN de Helminto/genética , ARN Mensajero/genética , Conejos , Schistosoma mansoni/genética , Técnicas del Sistema de Dos HíbridosRESUMEN
The protozoan Trypanosoma cruzi, its mammalian reservoirs, and vectors have existed in nature for millions of years. The human infection, named Chagas disease, is a major public health problem for Latin America. T. cruzi is genetically highly diverse and the understanding of the population structure of this parasite is critical because of the links to transmission cycles and disease. At present, T. cruzi is partitioned into six discrete typing units (DTUs), TcI-TcVI. Here we focus on the current status of taxonomy-related areas such as population structure, phylogeographical and eco-epidemiological features, and the correlation of DTU with natural and experimental infection. We also summarize methods for DTU genotyping, available for widespread use in endemic areas. For the immediate future multilocus sequence typing is likely to be the gold standard for population studies. We conclude that greater advances in our knowledge on pathogenic and epidemiological features of these parasites are expected in the coming decade through the comparative analysis of the genomes from isolates of various DTUs.
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Enfermedad de Chagas/epidemiología , Enfermedad de Chagas/parasitología , Trypanosoma cruzi/clasificación , Animales , Quirópteros/parasitología , Evolución Molecular , Variación Genética , Genoma de Protozoos , Genómica , Humanos , Tipificación de Secuencias Multilocus , Filogeografía , Terminología como Asunto , Trypanosoma cruzi/genética , Trypanosoma cruzi/patogenicidadRESUMEN
Therapeutic failure of benznidazole (BZ) is widely documented in Chagas disease and has been primarily associated with variations in the drug susceptibility of Trypanosoma cruzi strains. In humans, therapeutic success has been assessed by the negativation of anti-T. cruzi antibodies, a process that may take up to 10 years. A protocol for early screening of the drug resistance of infective strains would be valuable for orienting physicians towards alternative therapies, with a combination of existing drugs or new anti-T. cruzi agents. We developed a procedure that couples the isolation of parasites by haemoculture with quantification of BZ susceptibility in the resultant epimastigote forms. BZ activity was standardized with reference strains, which showed IC50 to BZ between 7.6-32 µM. The assay was then applied to isolates from seven chronic patients prior to administration of BZ therapy. The IC50 of the strains varied from 15.6 ± 3-51.4 ± 1 µM. Comparison of BZ susceptibility of the pre-treatment isolates of patients considered cured by several criteria and of non-cured patients indicates that the assay does not predict therapeutic outcome. A two-fold increase in BZ resistance in the post-treatment isolates of two patients was verified. Based on the profile of nine microsatellite loci, sub-population selection in non-cured patients was ruled out.
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Enfermedad de Chagas/tratamiento farmacológico , Nitroimidazoles/uso terapéutico , Tripanocidas/uso terapéutico , Trypanosoma cruzi/efectos de los fármacos , Adulto , Enfermedad de Chagas/parasitología , Resistencia a Medicamentos , Femenino , Humanos , Dosificación Letal Mediana , Masculino , Repeticiones de Microsatélite , Persona de Mediana Edad , Nitroimidazoles/farmacología , Pruebas de Sensibilidad Parasitaria , Resultado del Tratamiento , Tripanocidas/farmacología , Trypanosoma cruzi/genéticaRESUMEN
Therapeutic failure of benznidazole (BZ) is widely documented in Chagas disease and has been primarily associated with variations in the drug susceptibility of Trypanosoma cruzi strains. In humans, therapeutic success has been assessed by the negativation of anti-T. cruzi antibodies, a process that may take up to 10 years. A protocol for early screening of the drug resistance of infective strains would be valuable for orienting physicians towards alternative therapies, with a combination of existing drugs or new anti-T. cruzi agents. We developed a procedure that couples the isolation of parasites by haemoculture with quantification of BZ susceptibility in the resultant epimastigote forms. BZ activity was standardized with reference strains, which showed IC50 to BZ between 7.6-32 µM. The assay was then applied to isolates from seven chronic patients prior to administration of BZ therapy. The IC50 of the strains varied from 15.6 ± 3-51.4 ± 1 µM. Comparison of BZ susceptibility of the pre-treatment isolates of patients considered cured by several criteria and of non-cured patients indicates that the assay does not predict therapeutic outcome. A two-fold increase in BZ resistance in the post-treatment isolates of two patients was verified. Based on the profile of nine microsatellite loci, sub-population selection in non-cured patients was ruled out.
Asunto(s)
Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Enfermedad de Chagas , Nitroimidazoles , Tripanocidas , Trypanosoma cruzi , Enfermedad de Chagas , Resistencia a Medicamentos , Repeticiones de Microsatélite , Nitroimidazoles , Pruebas de Sensibilidad Parasitaria , Resultado del Tratamiento , Tripanocidas , Trypanosoma cruziRESUMEN
We have previously demonstrated selection favoring the JG strain of Trypanosoma cruzi in hearts of BALB/c mice that were chronically infected with an equal mixture of the monoclonal JG strain and a clone of the Colombian strain, Col1.7G2. To evaluate whether cell invasion efficiency drives this selection, we infected primary cultures of BALB/c cardiomyocytes using these same T. cruzi populations. Contrary to expectation, Col1.7G2 parasites invaded heart cell cultures in higher numbers than JG parasites; however, intracellular multiplication of JG parasites was more efficient than that of Col1.7G2 parasites. This phenomenon was only observed for cardiomyocytes and not for cultured Vero cells. Double infections (Col1.7G2 + JG) showed similar results. Even though invasion might influence tissue selection, our data strongly suggest that intracellular development is important to determine parasite tissue tropism.
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Interacciones Huésped-Parásitos , Miocitos Cardíacos/parasitología , Tropismo/fisiología , Trypanosoma cruzi/crecimiento & desarrollo , Animales , Femenino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos DBA , Factores de Tiempo , Trypanosoma cruzi/clasificación , Trypanosoma cruzi/genéticaRESUMEN
We have previously demonstrated selection favoring the JG strain of Trypanosoma cruziin hearts of BALB/c mice that were chronically infected with an equal mixture of the monoclonal JG strain and a clone of the Colombian strain, Col1.7G2. To evaluate whether cell invasion efficiency drives this selection, we infected primary cultures of BALB/c cardiomyocytes using these same T. cruzi populations. Contrary to expectation, Col1.7G2 parasites invaded heart cell cultures in higher numbers than JG parasites; however, intracellular multiplication of JG parasites was more efficient than that of Col1.7G2 parasites. This phenomenon was only observed for cardiomyocytes and not for cultured Vero cells. Double infections (Col1.7G2 + JG) showed similar results. Even though invasion might influence tissue selection, our data strongly suggest that intracellular development is important to determine parasite tissue tropism.
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Animales , Femenino , Ratones , Interacciones Huésped-Parásitos , Miocitos Cardíacos , Tropismo/fisiología , Trypanosoma cruzi/crecimiento & desarrollo , Ratones Endogámicos BALB C , Ratones Endogámicos DBA , Factores de Tiempo , Trypanosoma cruzi , Trypanosoma cruziRESUMEN
The Trypanosoma cruzi taxon is composed of 2 major lineages, T cruzi I and T cruzi II. The clinical symptoms of Chagas disease are highly variable, and their geographic distribution is correlated with the distribution of the parasite lineages. In Colombia and northern South America, T cruzi I lineage is associated with chagasic cardiomyopathy. Alternatively, in the countries south cone of South America, there is a predominance of T cruzi II, which is associated with cardiomyopathy and digestive diseases. We report for the first time a mixed infection consisting of both T cruzi I and T cruzi II detected in the esophagus and in the heart, respectively, of a cardiomyopathic patient from an endemic area in Santander, Colombia. This finding has epidemiological relevance related to the association of T cruzi II with the clinical manifestations of Chagas disease and its frequency in Colombia and countries in northern South America.
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Cardiomiopatía Chagásica/parasitología , Trypanosoma cruzi/aislamiento & purificación , Resultado Fatal , Humanos , Masculino , Persona de Mediana Edad , Trypanosoma cruzi/clasificaciónRESUMEN
BACKGROUND: During its development, the parasite Schistosoma mansoni is exposed to different environments and undergoes many morphological and physiological transformations as a result of profound changes in gene expression. Characterization of proteins involved in the regulation of these processes is of importance for the understanding of schistosome biology. Proteins containing zinc finger motifs usually participate in regulatory processes and are considered the major class of transcription factors in eukaryotes. It has already been shown, by EMSA (Eletrophoretic Mobility Shift Assay), that SmZF1, a S. mansoni zinc finger (ZF) protein, specifically binds both DNA and RNA oligonucleotides. This suggests that this protein might act as a transcription factor in the parasite. METHODOLOGY/PRINCIPAL FINDINGS: In this study we extended the characterization of SmZF1 by determining its subcellular localization and by verifying its ability to regulate gene transcription. We performed immunohistochemistry assays using adult male and female worms, cercariae and schistosomula to analyze the distribution pattern of SmZF1 and verified that the protein is mainly detected in the cells nuclei of all tested life cycle stages except for adult female worms. Also, SmZF1 was heterologously expressed in mammalian COS-7 cells to produce the recombinant protein YFP-SmZF1, which was mainly detected in the nucleus of the cells by confocal microscopy and Western blot assays. To evaluate the ability of this protein to regulate gene transcription, cells expressing YFP-SmZF1 were tested in a luciferase reporter system. In this system, the luciferase gene is downstream of a minimal promoter, upstream of which a DNA region containing four copies of the SmZF1 putative best binding site (D1-3DNA) was inserted. SmZF1 increased the reporter gene transcription by two fold (p
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Proteínas del Helminto/metabolismo , Schistosoma mansoni/metabolismo , Esquistosomiasis mansoni/parasitología , Factores de Transcripción/metabolismo , Animales , Células COS , Núcleo Celular/metabolismo , Chlorocebus aethiops , Femenino , Proteínas del Helminto/genética , Humanos , Masculino , Unión Proteica , Transporte de Proteínas , Schistosoma mansoni/genética , Schistosoma mansoni/crecimiento & desarrollo , Esquistosomiasis mansoni/metabolismo , Factores de Transcripción/genéticaRESUMEN
Virus-host biological interaction is a continuous coevolutionary process involving both host immune system and viral escape mechanisms. Flaviviridae family is composed of fast evolving RNA viruses that infects vertebrate (mammals and birds) and/or invertebrate (ticks and mosquitoes) organisms. These host groups are very distinct life forms separated by a long evolutionary time, so lineage-specific anti-viral mechanisms are likely to have evolved. Flaviviridae viruses which infect a single host lineage would be subjected to specific host-induced pressures and, therefore, selected by them. In this work we compare the genomic evolutionary patterns of Flaviviridae viruses and their hosts in an attempt to uncover coevolutionary processes inducing common features in such disparate groups. Especially, we have analyzed dinucleotide and codon usage patterns in the coding regions of vertebrate and invertebrate organisms as well as in Flaviviridae viruses which specifically infect one or both host types. The two host groups posses very distinctive dinucleotide and codon usage patterns. A pronounced CpG under-representation was found in the vertebrate group, possibly induced by the methylation-deamination process, as well as a prominent TpA decrease. The invertebrate group displayed only a TpA frequency reduction bias. Flaviviridae viruses mimicked host nucleotide motif usage in a host-specific manner. Vertebrate-infecting viruses possessed under-representation of CpG and TpA, and insect-only viruses displayed only a TpA under-representation bias. Single-host Flaviviridae members which persistently infect mammals or insect hosts (Hepacivirus and insect-only Flavivirus, respectively) were found to posses a codon usage profile more similar to that of their hosts than to related Flaviviridae. We demonstrated that vertebrates and mosquitoes genomes are under very distinct lineage-specific constraints, and Flaviviridae viruses which specifically infect these lineages appear to be subject to the same evolutionary pressures that shaped their host coding regions, evidencing the lineage-specific coevolutionary processes between the viral and host groups.
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Evolución Biológica , Flaviviridae/genética , Interacciones Huésped-Patógeno , Animales , Secuencia de Bases , Codón , Vectores de Enfermedades , Flaviviridae/clasificación , FilogeniaRESUMEN
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.
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Cardiomiopatía Chagásica/genética , Enfermedad de Chagas/genética , Proteínas de la Membrana/genética , NADH Deshidrogenasa/genética , Proteínas Protozoarias/genética , Trypanosoma cruzi/genética , Animales , Brasil/epidemiología , Cardiomiopatía Chagásica/epidemiología , Cardiomiopatía Chagásica/parasitología , Enfermedad de Chagas/epidemiología , Enfermedad de Chagas/parasitología , Perfilación de la Expresión Génica , Humanos , Datos de Secuencia Molecular , Trypanosoma cruzi/aislamiento & purificaciónRESUMEN
We have previously demonstrated that both parasite genetic variability and host genetic background were important in determining the differential tissue distribution of the Col1.7G2 and JG T. cruzi monoclonal strains after artificial infections in mice. We observed that the JG strain was most prevalent in hearts of mouse lineages with the MHC haplotype H-2(d) (BALB/c and DBA2), while Col1.7G2 was predominant in hearts from C57BL/6 mice, which have the H-2(b) haplotype. To assess whether the MHC gene region indeed influenced tissue tropism of T. cruzi, we used the same two parasite strains to infect C57BL/6 (H-2(b)) and C57BLKS/J (H-2(d)) mice; the latter strain results from the introgression of DBA2 MHC region into the C57BL/6 background. We also performed ex vivo infections of cardiac explants from four congenic mice lineages with the H-2(b) and H-2(d) haplotypes arranged in two different genetic backgrounds: C57BLKS/J (H-2(d)) versus C57BL/6 (H-2(b)) and BALB/c (H-2(d)) versus BALB/B10-H2(b) (H-2(b)). In agreement with our former observations, Col1.7G2 was predominant in hearts from C57BL/6 mice (H-2(b)), but we observed a clear predominance of the JG strain in hearts from C57BLKS/J animals (H-2(d)). In the ex vivo experiments Col1.7G2 also prevailed in explants from H-2(b) animals while no predominance of any of the strains was observed in H-2(d) mice explants, regardless of the genetic background. These observations clearly demonstrate that the MHC region influences the differential tissue distribution pattern of infecting T. cruzi strains, which by its turn may be in a human infection the determinant for the clinical forms of the Chagas disease.
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Interacciones Huésped-Parásitos/genética , Complejo Mayor de Histocompatibilidad/genética , Tropismo , Trypanosoma cruzi/patogenicidad , Animales , Secuencia de Bases , Cartilla de ADN , Haplotipos , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Especificidad de Órganos , Reacción en Cadena de la PolimerasaRESUMEN
Trypanosomes are unicellular eukaryotes that cause disease in humans and other mammals. Trypanosoma cruzi and Trypanosoma brucei are the causative agents, respectively, of Chagas disease in the Americas and sleeping sickness in sub-Saharan Africa. To better comprehend the interaction of these parasites with their hosts, understanding the mechanisms involved in the generation of genetic variability is critical. One such mechanism is mismatch repair (MMR), which has a crucial, evolutionarily conserved role in maintaining the fidelity of DNA replication, as well as acting in other cellular processes, such as DNA recombination. Here we have attempted to complement T. brucei MMR through the expression of MSH2 from T. cruzi. Our results show that T. brucei MSH2-null mutants are more sensitive to hydrogen peroxide (H2O2) than wild type cells, suggesting the involvement of MSH2 in the response to oxidative stress in this parasite. This phenotype is reverted by the expression of either the T. cruzi or the T. brucei MSH2 protein in the MSH2-null mutants. In contrast, MMR complementation, as assessed by resistance to N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and microsatellite instability, was not achieved by the heterologous expression of T. cruzi MSH2. This finding, associated to the demonstration that mutation of MLH1, another component of the MMR system, did not affect sensitivity of T. brucei cells to H2O2, suggests an additional role of MSH2 in dealing with oxidative damage in these parasites, which may occur independently of MMR.