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1.
Mem Inst Oswaldo Cruz ; 116: e200634, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33787768

RESUMO

The availability of Trypanosomatid genomic data in public databases has opened myriad experimental possibilities that have contributed to a more comprehensive understanding of the biology of these parasites and their interactions with hosts. In this review, after brief remarks on the history of the Trypanosoma cruzi and Leishmania genome initiatives, we present an overview of the relevant contributions of genomics, transcriptomics and functional genomics, discussing the primary obstacles, challenges, relevant achievements and future perspectives of these technologies.


Assuntos
Genoma de Protozoário/genética , Leishmania/genética , Trypanosoma cruzi/genética , Biologia Computacional , Genômica
2.
Rev Soc Bras Med Trop ; 54: e0873-2020, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33759934

RESUMO

Chagas disease is caused by the protozoan Trypanosoma cruzi. Seven lineages have been identified based on different molecular markers, namely TcI, TcII, TcIII, TcIV, TcV, TcVI, and TcBat. Dogs play the role of epidemiological sentinels being domestic reservoirs of T. cruzi. The aim of the current study was to report the first case of CD in a domestic dog in Manaus, Amazonas, Brazil, infected with T. cruzi DTU TcIV. We hope our report encourages veterinarians and surveillance professionals to a take a deeper look at T. cruzi infection in domestic animals.


Assuntos
Doença de Chagas , Trypanosoma cruzi , Animais , Brasil , Doença de Chagas/diagnóstico , Doença de Chagas/veterinária , Cães , Genótipo , Trypanosoma cruzi/genética
3.
PLoS Genet ; 16(12): e1009170, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33326438

RESUMO

Analysis of genetic polymorphism is a powerful tool for epidemiological surveillance and research. Powerful inference from pathogen genetic variation, however, is often restrained by limited access to representative target DNA, especially in the study of obligate parasitic species for which ex vivo culture is resource-intensive or bias-prone. Modern sequence capture methods enable pathogen genetic variation to be analyzed directly from host/vector material but are often too complex and expensive for resource-poor settings where infectious diseases prevail. This study proposes a simple, cost-effective 'genome-wide locus sequence typing' (GLST) tool based on massive parallel amplification of information hotspots throughout the target pathogen genome. The multiplexed polymerase chain reaction amplifies hundreds of different, user-defined genetic targets in a single reaction tube, and subsequent agarose gel-based clean-up and barcoding completes library preparation at under 4 USD per sample. Our study generates a flexible GLST primer panel design workflow for Trypanosoma cruzi, the parasitic agent of Chagas disease. We successfully apply our 203-target GLST panel to direct, culture-free metagenomic extracts from triatomine vectors containing a minimum of 3.69 pg/µl T. cruzi DNA and further elaborate on method performance by sequencing GLST libraries from T. cruzi reference clones representing discrete typing units (DTUs) TcI, TcIII, TcIV, TcV and TcVI. The 780 SNP sites we identify in the sample set repeatably distinguish parasites infecting sympatric vectors and detect correlations between genetic and geographic distances at regional (< 150 km) as well as continental scales. The markers also clearly separate TcI, TcIII, TcIV and TcV + TcVI and appear to distinguish multiclonal infections within TcI. We discuss the advantages, limitations and prospects of our method across a spectrum of epidemiological research.


Assuntos
Código de Barras de DNA Taxonômico/métodos , Genoma de Protozoário , Metagenoma , Metagenômica/métodos , Trypanosoma cruzi/genética , Sequenciamento Completo do Genoma/métodos , Animais , Custos e Análise de Custo , Código de Barras de DNA Taxonômico/economia , Código de Barras de DNA Taxonômico/normas , Vetores de Doenças , Hemípteros/parasitologia , Metagenômica/economia , Metagenômica/normas , Polimorfismo Genético , Trypanosoma cruzi/patogenicidade , Virulência/genética , Sequenciamento Completo do Genoma/economia , Sequenciamento Completo do Genoma/normas
4.
PLoS Negl Trop Dis ; 14(12): e0008969, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33347472

RESUMO

CD8 T cells are regarded as pivotal players in both immunoprotection and immunopathology following Trypanosoma cruzi infection. Previously, we demonstrated the expansion of CD8+ T lymphocytes in the spleen of T. cruzi-infected mice under treatment with benznidazole (N-benzyl-2-nitroimidazole acetamide; Bz), a drug available for clinical therapy. This finding underlies the concept that the beneficial effects of Bz on controlling acute T. cruzi infection are related to a synergistic process between intrinsic trypanocidal effect and indirect triggering of the active immune response. In the present study, we particularly investigated the effect of Bz treatment on the CD8+ T cell subset following T. cruzi infection. Herein we demonstrated that, during acute T. cruzi infection, Bz treatment reduces and abbreviates the parasitemia, but maintains elevated expansion of CD8+ T cells. Within this subset, a remarkable group of CD8low cells was found in both Bz-treated and non-treated infected mice. In Bz-treated mice, early pathogen control paralleled the lower frequency of recently activated CD8low cells, as ascertained by CD69 expression. However, the CD8low subset sustains significant levels of CD44highCD62Llow and CD62LlowT-bethigh effector memory T cells, in both Bz-treated and non-treated infected mice. These CD8low cells also comprise the main group of spontaneous interferon (IFN)-γ-producing CD8+ T cells. Interestingly, following in vitro anti-CD3/CD28 stimulation, CD8+ T cells from Bz-treated T. cruzi-infected mice exhibited higher frequency of IFN-γ+ cells, which bear mostly a CD8low phenotype. Altogether, our results point to the marked presence of CD8low T cells that arise during acute T. cruzi infection, with Bz treatment promoting their significant expansion along with a potential effector program for high IFN-γ production.


Assuntos
Doença de Chagas/tratamento farmacológico , Nitroimidazóis/uso terapêutico , Tripanossomicidas/uso terapêutico , Trypanosoma cruzi/efeitos dos fármacos , Doença Aguda , Animais , Linfócitos T CD8-Positivos/imunologia , Doença de Chagas/parasitologia , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Baço/imunologia , Trypanosoma cruzi/genética
5.
PLoS Negl Trop Dis ; 14(12): e0008932, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33332357

RESUMO

BACKGROUND: Chagas disease is a neglected zoonosis of growing concern in the southern US, caused by the parasite Trypanosoma cruzi. We genotyped parasites in a large cohort of PCR positive dogs to shed light on parasite transmission cycles and assess potential relationships between parasite diversity and serological test performance. METHODOLOGY/PRINCIPAL FINDINGS: We used a metabarcoding approach based on deep sequencing of T. cruzi mini-exon marker to assess parasite diversity. Phylogenetic analysis of 178 sequences from 40 dogs confirmed the presence of T. cruzi discrete typing unit (DTU) TcI and TcIV, as well as TcII, TcV and TcVI for the first time in US dogs. Infections with multiple DTUs occurred in 38% of the dogs. These data indicate a greater genetic diversity of T. cruzi than previously detected in the US. Comparison of T. cruzi sequence diversity indicated that highly similar T. cruzi strains from these DTUs circulate in hosts and vectors in Louisiana, indicating that they are involved in a shared T. cruzi parasite transmission cycle. However, TcIV and TcV were sampled more frequently in vectors, while TcII and TcVI were sampled more frequently in dogs. CONCLUSIONS/SIGNIFICANCE: These observations point to ecological host-fitting being a dominant mechanism involved in the diversification of T. cruzi-host associations. Dogs with negative, discordant or confirmed positive T. cruzi serology harbored TcI parasites with different mini-exon sequences, which strongly supports the hypothesis that parasite genetic diversity is a key factor affecting serological test performance. Thus, the identification of conserved parasite antigens should be a high priority for the improvement of current serological tests.


Assuntos
Doença de Chagas/veterinária , Éxons/genética , Variação Genética , Trypanosoma cruzi/genética , Animais , Doença de Chagas/epidemiologia , Doença de Chagas/parasitologia , Doença de Chagas/transmissão , Estudos de Coortes , Cães , Genótipo , Humanos , Louisiana/epidemiologia , Filogenia , Testes Sorológicos/veterinária , Trypanosoma cruzi/imunologia , Trypanosoma cruzi/fisiologia , Zoonoses
6.
Mem Inst Oswaldo Cruz ; 115: e200142, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33053076

RESUMO

BACKGROUND: Calpains are present in almost all organisms and comprise a family of calcium-dependent cysteine peptidases implicated in crucial cellular functions. Trypanosoma cruzi, the causative agent of Chagas disease, presents an expansion on this gene family with unexplored biological properties. OBJECTIVES: Here, we searched for calpains in the T. cruzi genome, evaluated the mRNA levels, calpain activity and the protein expression and determined the cellular localisation in all three parasite life cycle forms. METHODS/FINDINGS: Sixty-three calpain sequences were identified in T. cruzi CL Brener genome, with fourteen domain arrangements. The comparison of calpain mRNA abundance by quantitative polymerase chain reaction (qPCR) revealed seven up-regulated sequences in amastigotes and/or bloodstream trypomastigotes and five in epimastigotes. Western Blotting analysis revealed seven different molecules in the three parasite forms, and one amastigote-specific, while no proteolytic activity could be detected. Flow cytometry assays revealed a higher amount of intracellular calpains in amastigotes and/or trypomastigotes in comparison to epimastigotes. Finally, ultrastructural analysis revealed the presence of calpains in the cytoplasm, vesicular and plasma membranes of the three parasite forms, and in the paraflagellar rod in trypomastigotes. CONCLUSION: Calpains are differentially expressed and localised in the T. cruzi life cycle forms. This study adds data on the calpain occurrence and expression pattern in T. cruzi.


Assuntos
Calpaína/genética , Trypanosoma cruzi/genética , Animais , Western Blotting , Calpaína/metabolismo , Doença de Chagas , Estágios do Ciclo de Vida , RNA Mensageiro/genética
7.
PLoS Pathog ; 16(8): e1008781, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32810179

RESUMO

Chagas disease is caused by Trypanosoma cruzi, a protozoan parasite that has a heterogeneous population composed of a pool of strains with distinct characteristics, including variable levels of virulence. In previous work, transcriptome analyses of parasite genes after infection of human foreskin fibroblasts (HFF) with virulent (CL Brener) and non-virulent (CL-14) clones derived from the CL strain, revealed a reduced expression of genes encoding parasite surface proteins in CL-14 compared to CL Brener during the final steps of the intracellular differentiation from amastigotes to trypomastigotes. Here we analyzed changes in the expression of host genes during in vitro infection of HFF cells with the CL Brener and CL-14 strains by analyzing total RNA extracted from cells at 60 and 96 hours post-infection (hpi) with each strain, as well as from uninfected cells. Similar transcriptome profiles were observed at 60 hpi with both strains compared to uninfected samples. However, at 96 hpi, significant differences in the number and expression levels of several genes, particularly those involved with immune response and cytoskeleton organization, were observed. Further analyses confirmed the difference in the chemokine/cytokine signaling involved with the recruitment and activation of immune cells such as neutrophils upon T. cruzi infection. These findings suggest that infection with the virulent CL Brener strain induces a more robust inflammatory response when compared with the non-virulent CL-14 strain. Importantly, the RNA-Seq data also exposed an unexplored role of fibroblasts as sentinel cells that may act by recruiting neutrophils to the initial site of infection. This role for fibroblasts in the regulation of the inflammatory response during infection by T. cruzi was corroborated by measurements of levels of different chemokines/cytokines during in vitro infection and in plasma from Chagas disease patients as well as by neutrophil activation and migration assays.


Assuntos
Doença de Chagas/metabolismo , Fibroblastos , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Ativação de Neutrófilo , Neutrófilos , Trypanosoma cruzi/metabolismo , Doença de Chagas/genética , Doença de Chagas/patologia , Fibroblastos/metabolismo , Fibroblastos/parasitologia , Fibroblastos/patologia , Humanos , Neutrófilos/metabolismo , Neutrófilos/parasitologia , Neutrófilos/patologia , Trypanosoma cruzi/genética , Trypanosoma cruzi/patogenicidade , Fatores de Virulência/genética , Fatores de Virulência/metabolismo
8.
PLoS One ; 15(8): e0237180, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32750094

RESUMO

BACKGROUND: Chagas disease, caused by the intracellular parasite Trypanosoma cruzi, is one of the most important parasitological infections in the Americas. It is estimated to infect approximately 6 million people from mostly low income countries in Latin America, although recent infections have been reported in southern US states. Several studies have described an extensive genetic diversity among T. cruzi isolates throughout its geographic distribution in the American continent. This diversity has been correlated with the pathology developed during an infection. However, due to a lack of a single reliable test, current diagnosis practices of the disease are not straightforward since several different tests are applied. The use of current genomic sequence data allows for the selection of molecular markers (MM) that have the ability to identify the Discrete Typing Unit (DTU) of T. cruzi in a given infection, without the need of any sequencing reaction. METHODOLOGY/PRINCIPAL FINDINGS: Applying three criteria on the genomic sequencing data of four different phylogenetic lineages of T. cruzi, we designed several molecular tests that can be used for the molecular typing of the parasite. The criteria used were: (1) single-copy orthologs of T. cruzi, (2) T. cruzi unique loci, and (3) T. cruzi polymorphic loci. All criteria combined allowed for the selection of 15 MM, 12 of which were confirmed to be functional and replicable in the laboratory with sylvatic samples. Furthermore, one MM produced distinct polymerase chain reaction (PCR) amplicon sizes among distinct T. cruzi DTUs, allowing the use of a AFLP-PCR test to distinguish DTUs I, II/IV, V and VI. Whereas two MM can differentiate DTUs I, II, IV and V/VI out of the six current DTUs with a PCR-RFLP test. CONCLUSIONS/SIGNIFICANCE: The designed molecular tests provide a practical and inexpensive molecular typing test for the majority of DTUs of T. cruzi, excluding the need to perform any sequencing reaction. This provides the scientific community with an additional specific, quick and inexpensive test that can enhance the understanding of the correlation between the DTU of T. cruzi and the pathology developed during the infection.


Assuntos
Análise do Polimorfismo de Comprimento de Fragmentos Amplificados/métodos , Doença de Chagas/diagnóstico , Polimorfismo de Fragmento de Restrição/genética , Trypanosoma cruzi/genética , Doença de Chagas/parasitologia , DNA de Protozoário/genética , Loci Gênicos , Variação Genética , Genoma de Protozoário/genética , Humanos , Tipagem Molecular/métodos , Filogenia , Polimorfismo de Nucleotídeo Único
9.
Mem Inst Oswaldo Cruz ; 115: e200019, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32696913

RESUMO

BACKGROUND: NME23/NDPKs are well conserved proteins found in all living organisms. In addition to being nucleoside diphosphate kinases (NDPK), they are multifunctional enzymes involved in different processes such as DNA stability, gene regulation and DNA repair among others. TcNDPK1 is the canonical NDPK isoform present in Trypanosoma cruzi, which has nuclease activity and DNA-binding properties in vitro. OBJECTIVES: In the present study we explored the role of TcNDPK1 in DNA damage responses. METHODS: TcNDPK1 was expressed in mutant bacteria and yeasts and over-expressed in epimastigotes. Mutation frequencies, tolerance to genotoxic agents and activity of DNA repair enzymes were evaluated. FINDINGS: Bacteria decreased about 15-folds the spontaneous mutation rate and yeasts were more resistant to hydrogen peroxide and to UV radiation than controls. Parasites overexpressing TcNDPK1 were able to withstand genotoxic stresses caused by hydrogen peroxide, phleomycin and hidroxyurea. They also presented less genomic damage and augmented levels of poly(ADP)ribose and poly(ADP)ribose polymerase, an enzyme involved in DNA repair. MAIN CONCLUSION: These results strongly suggest a novel function for TcNDPK1; its involvement in the maintenance of parasite's genome integrity.


Assuntos
Dano ao DNA , Núcleosídeo-Difosfato Quinase/metabolismo , Trypanosoma cruzi/enzimologia , Reparo do DNA , Núcleosídeo-Difosfato Quinase/genética , Poli(ADP-Ribose) Polimerases , Trypanosoma cruzi/genética
10.
Exp Parasitol ; 215: 107931, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32464222

RESUMO

Chagas disease is a public health problem in America. Its parasite, Trypanosoma cruzi, presents different discrete typing units (DTUs), colonizes organs of mammalian hosts in chronic infections, and presents tropism for particular organs in experimental infections. We evaluated T. cruzi tropism towards organs on the naturally infected rodent Octodon degus, identifying the parasites' DTUs, by means of conventional PCR and hybridization. Almost all the analyzed organs presented T. cruzi. More than 42% of the tested oesophagus, skin, skeletal muscle, brain and intestine showed T. cruzi DNA. Other nine types of organs were infected in over 15%. These results suggest that there is some tropism by T. cruzi in chronically infected O. degus. DTU TcV was present in 92.5% of infected organs with identified DTUs; this DTU is frequently reported in human infections in the Southern Cone of South America. Few organs showed mixed DTU infections. This is one of the few reports on the outcome of chronic natural T. cruzi-infection in wild mammal hosts exposed to naturally infected vectors.


Assuntos
Doença de Chagas/veterinária , Octodon/parasitologia , Doenças dos Roedores/patologia , Doenças dos Roedores/parasitologia , Animais , Animais Selvagens , Doença de Chagas/parasitologia , Doença de Chagas/patologia , DNA de Protozoário/isolamento & purificação , Feminino , Masculino , Trypanosoma cruzi/classificação , Trypanosoma cruzi/genética
11.
Mem Inst Oswaldo Cruz ; 115: e190457, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32428081

RESUMO

BACKGROUND Imitation SWItch (ISWI) ATPase is the catalytic subunit in diverse chromatin remodeling complexes. These complexes modify histone-DNA interactions and therefore play a pivotal role in different DNA-dependent processes. In Trypanosoma cruzi, a protozoan that controls gene expression principally post-transcriptionally, the transcriptional regulation mechanisms mediated by chromatin remodeling are poorly understood. OBJECTIVE To characterise the ISWI remodeler in T. cruzi (TcISWI). METHODS A new version of pTcGW vectors was constructed to express green fluorescent protein (GFP)-tagged TcISWI. CRISPR-Cas9 system was used to obtain parasites with inactivated TcISWI gene and we determined TcISWI partners by cryomilling-affinity purification-mass spectrometry (MS) assay as an approximation to start to unravel the function of this protein. FINDINGS Our approach identified known ISWI partners [nucleoplasmin-like protein (NLP), regulator of chromosome condensation 1-like protein (RCCP) and phenylalanine/tyrosine-rich protein (FYRP)], previously characterised in T. brucei, and new components in TcISWI complex [DRBD2, DHH1 and proteins containing a domain characteristic of structural maintenance of chromosomes (SMC) proteins]. Data are available via ProteomeXchange with identifier PXD017869. MAIN CONCLUSIONS In addition to its participation in transcriptional silencing, as it was reported in T. brucei, the data generated here provide a framework that suggests a role for TcISWI chromatin remodeler in different nuclear processes in T. cruzi, including mRNA nuclear export control and chromatin compaction. Further work is necessary to clarify the TcISWI functional diversity that arises from this protein interaction study.


Assuntos
Adenosina Trifosfatases/genética , Montagem e Desmontagem da Cromatina/genética , Fatores de Transcrição/genética , Trypanosoma cruzi/genética , Animais , Western Blotting , Citometria de Fluxo , Regulação da Expressão Gênica
12.
Dis Markers ; 2020: 8074314, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32184904

RESUMO

Chagas' disease is a neglected tropical disease caused by Trypanosoma cruzi which is endemic throughout Latin America and is spread by worldwide migration. Diagnosis is currently limited to serological and molecular techniques having variations regarding their sensitivity and specificity. This work was aimed at developing a new sensitive, applicable, and cost-effective molecular diagnosis technique for loop-mediated isothermal amplification-based detection of T. cruzi (Tc-LAMP). The results led to determining a highly homologous satellite repeat region (231 bp) among parasite strains as a molecular marker for diagnosing the disease. Tc-LAMP was performed correctly for detecting parasite DNA (5 fg for the CL Brener strain and 50 fg for the DM28, TcVI, and TcI strains). Assay results proved negative for DNA from 16 helminth species and 7 protozoa, including Leishmania spp. Tc-LAMP based on the highly repeated T. cruzi satellite region is thus proposed as an important alternative for diagnosing T. cruzi infection, overcoming other methods' limitations such as their analytic capability, speed, and requiring specialized equipment or highly trained personnel. Tc-LAMP could be easily adapted for point-of-care testing in areas having limited resources.


Assuntos
Doença de Chagas/diagnóstico , DNA Satélite/genética , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Amplificação de Ácido Nucleico/métodos , Trypanosoma cruzi/isolamento & purificação , DNA de Protozoário/genética , Marcadores Genéticos , Humanos , Técnicas de Diagnóstico Molecular/economia , Técnicas de Amplificação de Ácido Nucleico/economia , Testes Imediatos , Sensibilidade e Especificidade , Trypanosoma cruzi/genética
13.
PLoS Negl Trop Dis ; 14(3): e0008007, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32196491

RESUMO

Investigations into intracellular replication and differentiation of Trypanosoma cruzi within the mammalian host have been restricted by limitations in our ability to detect parasitized cells throughout the course of infection. We have overcome this problem by generating genetically modified parasites that express a bioluminescent/fluorescent fusion protein. By combining in vivo imaging and confocal microscopy, this has enabled us to routinely visualise murine infections at the level of individual host cells. These studies reveal that intracellular parasite replication is an asynchronous process, irrespective of tissue location or disease stage. Furthermore, using TUNEL assays and EdU labelling, we demonstrate that within individual infected cells, replication of both mitochondrial (kDNA) and nuclear genomes is not co-ordinated within the parasite population, and that replicating amastigotes and non-replicating trypomastigotes can co-exist in the same cell. Finally, we report the presence of distinct non-canonical morphological forms of T. cruzi in the mammalian host. These appear to represent transitional forms in the amastigote to trypomastigote differentiation process. Therefore, the intracellular life-cycle of T. cruzi in vivo is more complex than previously realised, with potential implications for our understanding of disease pathogenesis, immune evasion and drug development. Dissecting the mechanisms involved will be an important experimental challenge.


Assuntos
Doença de Chagas/parasitologia , Replicação do DNA , Estágios do Ciclo de Vida , Trypanosoma cruzi/crescimento & desenvolvimento , Animais , Modelos Animais de Doenças , Feminino , Genes Reporter , Microscopia Intravital/métodos , Camundongos SCID , Microscopia Confocal/métodos , Coloração e Rotulagem/métodos , Trypanosoma cruzi/genética
14.
PLoS Negl Trop Dis ; 14(3): e0007910, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32150562

RESUMO

Anthropogenic environmental alterations such as urbanization can threaten native populations as well as create novel environments that allow human pests and pathogens to thrive. As the number and size of urban environments increase globally, it is more important than ever to understand the dispersal dynamics of hosts, vectors and pathogens of zoonotic disease systems. For example, a protozoan parasite and the causative agent of Chagas disease in humans, Trypanosoma cruzi, recently colonized and spread through the city of Arequipa, Peru. We used population genomic and phylogenomic tools to analyze whole genomes of 123 T. cruzi isolates derived from vectors and non-human mammals throughout Arequipa to determine patterns of T. cruzi dispersal. The data show significant population genetic structure within city blocks-parasites in the same block tend to be very closely related-but no population structure among blocks within districts-parasites in neighboring blocks are no more closely related to one another than to parasites in distant districts. These data suggest that T. cruzi dispersal within a block occurs regularly and that occasional long-range dispersal events allow the establishment of new T. cruzi populations in distant blocks. Movement of domestic animals may be the primary mechanism of inter-block and inter-district T. cruzi dispersal.


Assuntos
Animais Domésticos/parasitologia , Doença de Chagas/epidemiologia , Doença de Chagas/parasitologia , Transmissão de Doença Infecciosa , Genótipo , Filogenia , Trypanosoma cruzi/isolamento & purificação , Animais , Doença de Chagas/transmissão , Vetores de Doenças , Humanos , Epidemiologia Molecular , Peru/epidemiologia , Trypanosoma cruzi/classificação , Trypanosoma cruzi/genética
15.
Biochim Biophys Acta Mol Cell Res ; 1867(7): 118694, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32151656

RESUMO

Dot1 enzymes are histone methyltransferases that mono-, di- and trimethylate lysine 79 of histone H3 to affect several nuclear processes. The functions of these different methylation states are still largely unknown. Trypanosomes, which are flagellated protozoa that cause several parasitic diseases, have two Dot1 homologues. Dot1A catalyzes the mono- and dimethylation of lysine 76 during late G2 and mitosis, and Dot1B catalyzes trimethylation, which is a modification found in all stages of the cell cycle. Here, we generated Trypanosoma cruzi lines lacking Dot1B. Deletion of one allele resulted in parasites with increased levels of mono- and dimethylation and a reduction in H3K76me3. In the full knockout (DKO), no trimethylation was observed. Both the DKO and the single knockout (SKO) showed aberrant morphology and decreased growth due to cell cycle arrest after G2. This phenotype could be rescued by caffeine in the DKO, as caffeine is a checkpoint inhibitor of the cell cycle. The knockouts also phosphorylated γH2A without producing extensive DNA breaks, and Dot1B-depleted cells were more susceptible to general checkpoint kinase inhibitors, suggesting that a lack of H3K76 trimethylation prevents the initiation and/or completion of cytokinesis.


Assuntos
Doença de Chagas/genética , Histona-Lisina N-Metiltransferase/genética , Mitose/genética , Trypanosoma cruzi/genética , Ciclo Celular/genética , Doença de Chagas/parasitologia , Histonas/genética , Lisina/genética , Metilação/efeitos dos fármacos , Proteínas Nucleares/genética , Fosforilação/genética , Trypanosoma cruzi/patogenicidade
16.
Acta Trop ; 205: 105392, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32061627

RESUMO

There are 8 million people with Chagas disease worldwide and in El Salvador approximately 39% of the population is at risk of contracting the disease. One of the principal challenges in mitigating Chagas is evaluating the role of the vector ecology of triatomine species in the transmission of the Trypanosoma cruzi parasite in anthropogenically modified habitats, where new patterns of transmission frequently arise. Field studies of triatomine vector ecology in El Salvador have largely focused on describing parameters that contribute to infestation patterns, which may themselves be rooted in the morphological variability that exists in triatomine populations. The objective of this study was to evaluate the morphology of the vector species Triatoma dimidiata with respect to the characteristics of the ecological landscape the vector inhabits throughout El Salvador. We used image analyses to evaluate T. dimidiata morphological variability and then used Geographic Information Systems to intersect the morphological point-data with map layers containing different environmental characteristics. Our study found that the variation in the size, shape, and coloration of T. dimidiata varied in relation to elevation, Holdridge life zone, soil type and land use. We further characterize the local morphological adaptations of T. dimidiata with respect to the local ecological, biological, and geographical conditions in El Salvador. We suggest that future studies consider a molecular exploration of local T. dimidiata species complex in El Salvador, especially since morphological studies of triatomine species complex have found that variability correlate with the genetic variability of the population.


Assuntos
Distribuição Animal , Doença de Chagas/transmissão , Ecossistema , Insetos Vetores/fisiologia , Triatoma/fisiologia , Trypanosoma cruzi/fisiologia , Animais , Doença de Chagas/epidemiologia , El Salvador/epidemiologia , Insetos Vetores/parasitologia , Triatoma/parasitologia , Trypanosoma cruzi/genética
17.
BMC Infect Dis ; 20(1): 143, 2020 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-32059706

RESUMO

BACKGROUND: Chagas disease is caused by the haemoflagellate protozoan Trypanosoma cruzi. Currently, T. cruzi recognizes seven discrete typing units (DTUs): TcI to TcVI and Tcbat. The genetic diversity of T. cruzi is suspected to influence the clinical outcome. Acute clinical manifestations, which include myocarditis and meningoencephalitis, are sometimes fatal; occur most frequently in children and in immunocompromised individuals. Acute disease is often overlooked, leading to a poor prognosis. CASE PRESENTATION: A 38-year-old man from a subtropical area of the Andes mountains of Ecuador was hospitalized after 3 weeks of evolution with high fever, chills, an enlarged liver, spleen, and lymph nodes, as well as facial edema. ECG changes were also observed. T. cruzi was identified in blood smears, culture and amplification of DNA by PCR. Tests for anti-T. cruzi IgG and IgM and HIV were negative. Molecular typing by restriction fragment length polymorphism (PCR-RFLP) determined the parasite to DTU TcI. In the absence of a timely anti-T. cruzi medication, the patient died. CONCLUSIONS: This is a case of severe pathogenicity and the virulence of a DTU TcI strain in an adult patient. The severe acute Chagas disease was probably overlooked due to limited awareness and its low incidence. Our findings suggest that T. cruzi DTU TcI strains circulating in Ecuador are capable of causing fatal acute disease. Early diagnosis and prompt treatment is of paramount importance to avoid fatalities in acute infections.


Assuntos
Doença de Chagas/etiologia , Trypanosoma cruzi/genética , Trypanosoma cruzi/patogenicidade , Adulto , Doença de Chagas/parasitologia , Equador , Variação Genética , Humanos , Masculino , Tipagem Molecular , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Trypanosoma cruzi/classificação
18.
PLoS Negl Trop Dis ; 14(1): e0007770, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-32004318

RESUMO

BACKGROUND: Genetic exchange in Trypanosoma cruzi is controversial not only in relation to its frequency, but also to its mechanism. Parasexual genetic exchange has been proposed based on laboratory hybrids, but population genomics strongly suggests meiosis in T. cruzi. In addition, mitochondrial introgression has been reported several times in natural isolates although its mechanism is not fully understood yet. Moreover, hybrid T. cruzi DTUs (TcV and TcVI) have inherited at least part of the kinetoplastic DNA (kDNA = mitochondrial DNA) from both parents. METHODOLOGY/PRINCIPAL FINDINGS: In order to address such topics, we sequenced and analyzed fourteen nuclear DNA fragments and three kDNA maxicircle genes in three TcI stocks which are natural clones potentially involved in events of genetic exchange. We also deep-sequenced (a total of 6,146,686 paired-end reads) the minicircle hypervariable region (mHVR) of the kDNA in such three strains. In addition, we analyzed the DNA content by flow cytometry to address cell ploidy. We observed that most polymorphic sites in nuclear loci showed a hybrid pattern in one cloned strain and the other two cloned strains were compatible as parental strains (or nearly related to the true parents). The three clones had almost the same ploidy and the DNA content was similar to the reference strain Sylvio (a nearly diploid strain). Despite maxicircle genes evolve faster than nuclear housekeeping ones, we detected no polymorphisms in the sequence of three maxicircle genes showing mito-nuclear discordance. Lastly, the hybrid stock shared 66% of its mHVR clusters with one putative parent and 47% with the other one; in contrast, the putative parental stocks shared less than 30% of the mHVR clusters between them. CONCLUSIONS/SIGNIFICANCE: The results suggest a reductive division, a natural hybridization, biparental inheritance of the minicircles in the hybrid and maxicircle introgression. The models including such phenomena and explaining the relationships between these three clones are discussed.


Assuntos
DNA de Protozoário/genética , Hibridização Genética , Trypanosoma cruzi/classificação , Trypanosoma cruzi/genética , DNA de Cinetoplasto/genética , Genes de Protozoários , Sequenciamento de Nucleotídeos em Larga Escala , Ploidias , Análise de Sequência de DNA
19.
Biochim Biophys Acta Mol Basis Dis ; 1866(5): 165691, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32006572

RESUMO

More than 110 years has passed since the first publications on Chagas disease, and it still urges the necessity of understanding it as a complex socioenvironmental issue in which components of diverse nature converge and interact beyond the biomedical and epidemiological aspects. The current scenarios of the issue, both rural and Latin American as urban and global, demand that the education on Chagas disease include all possible contexts: where there are insect vectors and where there are not; inside and outside Latin America; in rural, periurban, and urban areas; in formal and non-formal educational environments. We consider essential the requirement of both an integral approach that overcomes the biomedical aspect to include the multidimensionality of the issue and a dialogical educational perspective that allows individuals and communities to analyze, decide, and lead contextualized prevention and promotion actions regarding their health. In this study, we surveyed, described, and critically analyzed studies approaching the link education-Chagas disease in scientific publications from the last 15 years. We aimed at contributing methodological-theoretical elements to (re)think the development of educational research and experiences that truly help facing this issue. From the electronic search of scientific literature in 6 databases, we found 426 articles, out of which we selected 25. We incorporated 10 articles from other sources to this initial corpus and performed both qualitative and quantitative analyses over the total number [35] to characterize the studied works in general, focusing on the conceptions on the Chagas disease issue and the underlying health education approaches.


Assuntos
Doença de Chagas/prevenção & controle , Doenças Negligenciadas/prevenção & controle , Educação de Pacientes como Assunto , Triatoma/parasitologia , Trypanosoma cruzi/patogenicidade , Animais , Pesquisa Biomédica/estatística & dados numéricos , Doença de Chagas/epidemiologia , Doença de Chagas/parasitologia , Doença de Chagas/transmissão , Controle de Doenças Transmissíveis/economia , Controle de Doenças Transmissíveis/organização & administração , Erradicação de Doenças/economia , Erradicação de Doenças/organização & administração , Vetores de Doenças , Carga Global da Doença , Humanos , Doenças Negligenciadas/epidemiologia , Doenças Negligenciadas/parasitologia , Política , Publicações/estatística & dados numéricos , Trypanosoma cruzi/genética , Trypanosoma cruzi/isolamento & purificação
20.
Biochim Biophys Acta Mol Basis Dis ; 1866(5): 165689, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32001300

RESUMO

This review is a perspective on the history of Chagas disease, and it adopts a novel approach from literary studies, historical documents and the science and epidemiology of the nature of the disease. From this analysis, comes the review's working definition of the Contact Zone (CZ): "the space in which geographically and historically separated people come into contact with each other and establish long-lasting relationships, which usually involve coercive conditions, radical inequality and intolerable conflict." In the Patient-Physician CZ, we verified the triple transition phenomena: the American trypanosomiasis shifted from a rural, acute, and vectorial transmitted disease to an urban, chronic and non-vectorial disease. In the Academic CZ, we describe the original disagreements which denied the existence of the disease and the current controversies about pathogenic mechanisms and etiological treatment. From the News from Latin America, and in the Original CZ, we will review the evolution of different forms of transmission. As in any good story, research across broad disciplines is necessary to reveal historical perspectives, scientific approaches, and the epidemiology of the disease, which has a prequel of 9000 years and an open ending: thus, we explore across the Global CZ, with its multiple and unexpected actors.


Assuntos
Doença de Chagas/história , Erradicação de Doenças/organização & administração , Doenças Endêmicas/história , Doenças Negligenciadas/história , Trypanosoma cruzi/patogenicidade , Animais , Restos Mortais/parasitologia , Doença de Chagas/epidemiologia , Doença de Chagas/prevenção & controle , Doença de Chagas/transmissão , DNA de Protozoário/isolamento & purificação , Erradicação de Doenças/história , Erradicação de Doenças/tendências , Vetores de Doenças , Doenças Endêmicas/prevenção & controle , Antropologia Forense/história , Carga Global da Doença , História do Século XIX , História do Século XX , História do Século XXI , História Antiga , Humanos , Doenças Negligenciadas/epidemiologia , Doenças Negligenciadas/parasitologia , Doenças Negligenciadas/prevenção & controle , Triatoma/parasitologia , Trypanosoma cruzi/genética , Trypanosoma cruzi/isolamento & purificação
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