RESUMO
The study focuses on identifying and understanding the ecological dynamics of Triatoma breyeri in Bolivia. Morphological identification and molecular analysis using gene fragments (COI, CytB and 16S) confirms T. breyeri's presence and its relation to other species. The species has been consistently found in the Estancia-Mataral-La Palma region since 2010 but has not spread to other regions in Bolivia. The region of occurrence is a small characteristic dry inter-Andean valley. A MaxEnt model suggests part of the Bolivian Montane Dry Forest ecoregion serves as a unique habitat within its range. The infrequent presence in Bolivia and the distance from its main range in Argentina suggest recent accidental introduction, possibly through human transport. Further research is needed to comprehend its persistence in this small area of Bolivia.
Assuntos
Triatoma , Animais , Bolívia , Triatoma/genética , Triatoma/fisiologia , Ecossistema , FilogeniaRESUMO
BACKGROUND: Chagas disease is caused by Trypanosoma cruzi, whose genetic structure is divided into six discrete typing units (DTUs) known as TcI-TcVI. In the Yucatan Peninsula, Mexico, information regarding the DTUs circulating in wild mammals is scarce, while this is important knowledge for our understanding of T. cruzi transmission dynamics. METHODS: In the current study, we sampled wild mammals in a sylvatic site of the Yucatan Peninsula and assessed their infection with T. cruzi by PCR. Then, for infected mammals, we amplified and sequenced nuclear and mitochondrial T. cruzi genetic markers for DTU identification. RESULTS: In total, we captured 99 mammals belonging to the orders Chiroptera, Rodentia and Didelphimorphia. The prevalence of infection with T. cruzi was 9% (9/99; 95% CI [5, 16]), and we identified TcI in a Jamaican fruit bat, Artibeus jamaicensis. Moreover, we fortuitously identified Trypanosoma dionisii in another Jamaican fruit bat and detected an unidentified Trypanosoma species in a third specimen. While the latter discoveries were not expected because we used primers designed for T. cruzi, this study is the first to report the identification of T. dionisii in a bat from Yucatan, Mexico, adding to a recent first report of T. dionisii in bats from Veracruz, and first report of this Trypanosoma species in Mexico. CONCLUSION: Further research is needed to enhance our knowledge of T. cruzi DTUs and Trypanosoma diversity circulating in wildlife in Southeastern Mexico.
Assuntos
Doença de Chagas , Quirópteros , Trypanosoma cruzi , Animais , México/epidemiologia , Quirópteros/parasitologia , Trypanosoma cruzi/genética , Trypanosoma cruzi/isolamento & purificação , Doença de Chagas/veterinária , Doença de Chagas/epidemiologia , Doença de Chagas/transmissão , Reação em Cadeia da Polimerase , DNA de Protozoário , Prevalência , Trypanosoma/isolamento & purificação , Trypanosoma/genética , Trypanosoma/classificação , Roedores/parasitologiaRESUMO
BACKGROUND: Trypanosoma cruzi, the etiological agent of Chagas Disease, exhibits remarkable genetic diversity and is classified into different Discrete Typing Units (DTUs). Strain typing techniques are crucial for studying T. cruzi, because their DTUs have significant biological differences from one another. However, there is currently no methodological strategy for the direct typing of biological materials that has sufficient sensitivity, specificity, and reproducibility. The high diversity and copy number of the minicircle hypervariable regions (mHVRs) makes it a viable target for typing. METHODOLOGY/PRINCIPAL FINDINGS: Approximately 24 million reads obtained by amplicon sequencing of the mHVR were analyzed for 62 strains belonging to the six main T. cruzi DTUs. To build reference databases of mHVR diversity for each DTU and to evaluate this target as a typing tool. Strains of the same DTU shared more mHVR clusters than strains of different DTUs, and clustered together. Different identity thresholds were used to build the reference sets of the mHVR sequences (85% and 95%, respectively). The 95% set had a higher specificity and was more suited for detecting co-infections, whereas the 85% set was excellent for identifying the primary DTU of a sample. The workflow's capacity for typing samples obtained from cultures, a set of whole-genome data, under various simulated PCR settings, in the presence of co-infecting lineages and for blood samples was also assessed. CONCLUSIONS/SIGNIFICANCE: We present reference databases of mHVR sequences and an optimized typing workflow for T. cruzi including a simple online tool for deep amplicon sequencing analysis (https://ntomasini.github.io/cruzityping/). The results show that the workflow displays an equivalent resolution to that of the other typing methods. Owing to its specificity, sensitivity, relatively low cost, and simplicity, the proposed workflow could be an alternative for screening different types of samples.
Assuntos
Doença de Chagas , Trypanosoma cruzi , Humanos , Trypanosoma cruzi/genética , Reprodutibilidade dos Testes , Doença de Chagas/diagnóstico , Reação em Cadeia da Polimerase , Análise de Sequência de DNA , Genótipo , Variação GenéticaRESUMO
PURPOSE: The objective of this study was to molecularly characterize Mexican isolates of T. cruzi obtained from infected triatomine bugs (the vectors of T. cruzi) and to evaluate their susceptibility to Nifurtimox (NFX). METHODS: Three isolates obtained from Triatoma dimidiata (collected in the State of Veracruz) and one isolate obtained from Triatoma bassolsae (collected in the State of Puebla) were molecularly characterized and the expression of genes associated with natural resistance to NFX was analyzed by qPCR. RESULTS: Molecular characterization by PCR showed that isolates Zn3, Zn5, and SRB1 belong to the DTU TcI, while isolate Sum3 belongs to TcIV. The latter was also confirmed by sequencing of mitochondrial genes. Isolate Zn5 was the most sensitive to treatment with NFX (IC50, 6.8 µM), isolates SRB1 and Zn3 were partially resistant (IC50, 12.8 µM and 12.7 µM) and isolate Sum3 showed a high degree of resistance to NFX (IC50, 21.4 µM). We also found an association between decreased NTR1 or OYE gene expression with NFX resistance. CONCLUSION: Our results also evidenced a high variability in the susceptibility to NFX of these T. cruzi isolates Central and Southeastern Mexico, suggesting the presence of naturally resistant isolates circulating in the country. These results have important implications for defining treatment policies for patients with Chagas disease.
Assuntos
Doença de Chagas , Triatoma , Trypanosoma cruzi , Animais , Humanos , Trypanosoma cruzi/genética , Nifurtimox/farmacologia , México , Insetos Vetores , GenótipoRESUMO
The genus Triatoma contains numerous species, principal or secondary vectors of Chagas disease, which have been included in the three main lineages of Triatomini tribe based on morphological and biogeographical characteristics: North American, South American, and T. dispar complex. The three members of the T. dispar complex are distributed in Ecuador. This complex has been scarcely studied through molecular approaches, and the taxonomic position of this complex is not confirmed. In this study, we explored the phylogenetic relationships within the genus Triatoma, including five species from North and Central America, six from South America, and the three species belonging to the T. dispar complex. Partial sequences of four mitochondrial genes (Cyt b, COII, 16S-rRNA, 12S-rRNA) and two nuclear genes (18S-rRNA, ITS2) were obtained from 74 specimens. Phylogenetic trees were built with concatenated and single sequences through maximum likelihood (ML), maximum parsimony (MP), and Bayesian methods. The trees built using concatenated sequences showed three main branches (clusters) highly supported by significant bootstrap values; the T. dispar complex appeared as a monophyletic group separate from species of North and Central American origin and South American origin. On the contrary, for each gene tree, the three main clusters were not always significantly supported, mostly because genetic information is dramatically reduced when a single gene is considered. Consequently, concatenation of genes gives relevant results and is highly recommended for further in-depth examination of the relationships of several species and complexes of triatomines that remain unresolved. Moreover, our current molecular data fully revealed the division of genus Triatoma into at least three main genetic groups.
Assuntos
Genes de Insetos , Genes Mitocondriais , Insetos Vetores/genética , Filogenia , Triatoma/classificação , Triatoma/genética , Animais , Teorema de Bayes , Biodiversidade , América Central , Doença de Chagas/transmissão , Citocromos b/genética , DNA Mitocondrial , DNA Espaçador Ribossômico , Complexo IV da Cadeia de Transporte de Elétrons/genética , Variação Genética , Haplótipos , Proteínas de Insetos/genética , América do Norte , RNA Ribossômico/análise , RNA Ribossômico 16S , RNA Ribossômico 18S , Análise de Sequência de DNA , América do SulRESUMO
Rhodnius ecuadoriensis is one of the most important vector species of Chagas disease in Ecuador. This species is distributed in the Central coast region and in the south Andean region, and an incipient speciation process between these geographical populations was previously proposed. The current population genetics study only focused on the Central coast region and analyzed 96 sylvatic specimens of R. ecuadoriensis associated with Phytelephas aequatorialis palm trees. We used Cytb and 16S-rRNA sequences and a Cytb-16S-rRNA concatenated set to explore (i) the genetic variability, spatial structuring, and demographic history of R. ecuadoriensis, and to determine (ii) the relationship between the genetic and climatic variabilities. A particularly high genetic variability was observed without detectable general genetic structure; only some terminal genetic clusters were observed. We did not observe isolation by geographical distance (IBD), and it is likely that ancient expansion occurred, according to Fs index and mismatch distribution for Cytb-16S-rRNA concatenated sequences. Hierarchical clustering showed that the current locality origins of the bugs were grouped into four bioclimatic clusters. Genetic and bioclimatic distances were not correlated, but some genetic clusters were associated with bioclimatic ones. The results showed an ancient evolution of the species in the region with a possible old expansion. The absence of spatial genetic structure could be due to climatic conditions (possible selection of singular genotypes) and to passive transportation of palms tree materials where R. ecuadoriensis are living.
Assuntos
Doença de Chagas/transmissão , Insetos Vetores/genética , Rhodnius/genética , Clima Tropical , Animais , Variação GenéticaRESUMO
Limited genetic data are currently available for three vectors of Chagas disease in Ecuador, Panstrongylus howardi, P. chinai, and P. rufotuberculatus. Previously regarded as mainly sylvatic, these species have been poorly studied. Recently, they have been more frequently reported in domiciles and peridomiciles and are now considered true secondary vectors of Chagas disease in a country where an estimated 200,000 people are infected by Trypanosoma cruzi, a causative agent of this disease. In order to fill this gap, we obtained DNA for sequencing from 53 insects belonging to these three species and mainly sampled from the two Ecuadorian provinces of Loja and Manabí. We used six mitochondrial loci (COI, COII, ND4, CytB, 16S, and 12S) and two nuclear ones (ITS2 and 18S). We interpreted the phylogenetic trees built with single and concatenated data through maximum likelihood, Bayesian Markov chain Monte Carlo, and maximum parsimony methods. We provide evidence that P. chinai and P. howardi are indeed two supported species closely related and derived from a common ancestor. Additionally, the phylogenetic position of P. rufotuberculatus was confirmed as being distant from P. chinai and P. howardi and clustered with Triatoma dimidiata, a species belonging to the Northern American Triatoma clade.
Assuntos
Proteínas Mitocondriais/genética , Panstrongylus/classificação , RNA Ribossômico 18S/genética , Análise de Sequência de DNA/métodos , Animais , Doença de Chagas/transmissão , Equador , Especiação Genética , Humanos , Análise Multinível , Tipagem de Sequências Multilocus , Panstrongylus/genética , Panstrongylus/parasitologia , FilogeniaRESUMO
Insects of the genus Rhodnius are broadly involved in Chagas disease transmission. In French Guiana, where the disease remains a public health problem, R. robustus and R. pictipes are vectors, but so far few genetic analyses of these local species have been reported. Here, we explored three mitochondrial genes (Cytb, Lsu-rRNA, and ND1) and one nuclear gene (D2) in 49 adult specimens morphologically characterized as R. robustus. We analyzed genetic polymorphisms and haplotype distributions, and we built phylogenetic trees using the available GenBank sequences from R. robustus and related species. The molecular taxonomy analysis confirmed that 35 insects, closely related to Brazilian species and separated by a few mutations, truly belong to R. robustus; two others were attributed to the R. prolixus complex and for 12 no sequence was obtained. The geographical haplotype distribution indicates a likely geographical structuring and evidenced true differentiation between the two main urban centers, Cayenne and Saint-Laurent-du-Maroni.
Assuntos
Doença de Chagas/transmissão , Insetos Vetores/genética , Rhodnius/genética , Animais , Guiana Francesa , Genes de Insetos/genética , HaplótiposRESUMO
BACKGROUND: For several years, the wild populations of Triatoma infestans, main vector of Trypanosoma cruzi causing Chagas disease, have been considered or suspected of being a source of reinfestation of villages. The number of sites reported for the presence of wild T. infestans, often close to human habitats, has greatly increased, but these data are scattered in several publications, and others obtained by our team in Bolivia have not been published yet. METHODOLOGY/PRINCIPAL FINDINGS: Herein is compiled the largest number of wild sites explored for the presence of T. infestans collected with two methods The standardized methods aimed to determine the relationship between wild T. infestans and the ecoregion, and the directed method help to confirm the presence/absence of triatomines in the ecoregions. Entomological indices were compared between ecoregions and an environmental niche modelling approach, based on bioclimatic variables, was applied. The active search for wild T. infestans in Bolivia suggests a discontinuous distribution from the Andean valleys to the lowlands (Chaco), while the models used suggest a continuous distribution between the two regions and very large areas where wild populations remain to be discovered. The results compile the description of different habitats where these populations were found, and we demonstrate that the environmental niches of wild and domestic populations, defined by climatic variables, are similar but not equivalent, showing that during domestication, T. infestans has conquered new spaces with wider ranges of temperature and precipitation. CONCLUSIONS/SIGNIFICANCE: The great diversity of wild T. infestans habitats and the comparison of their ecological niches with that of domestic populations confirm the behavioural plasticity of the species that increase the possibility of contact with humans. The result of the geographical distribution model of the wild populations calls for more entomological vigilance in the corresponding areas in the Southern Cone countries and in Bolivia. The current presentation is the most comprehensive inventory of wild T. infestans-positive sites that can be used as a reference for further entomological vigilance in inhabited areas.
Assuntos
Doença de Chagas/transmissão , Ecossistema , Insetos Vetores/crescimento & desenvolvimento , Triatoma/crescimento & desenvolvimento , Animais , Bolívia/epidemiologia , Entomologia , Meio Ambiente , Humanos , Trypanosoma cruziRESUMO
The Amazon region has recently been considered as endemic in Latin America. In Bolivia, the vast Amazon region is undergoing considerable human migrations and substantial anthropization of the environment, potentially renewing the danger of establishing the transmission of Chagas disease. The cases of human oral contamination occurring in 2010 in the town of Guayaramerín provided reasons to intensify research. As a result, the goal of this study was to characterize the species of sylvatic triatomines circulating in the surroundings of Yucumo (Beni, Bolivia), a small Amazonian city at the foot of the Andes between the capital (La Paz) and Trinidad the largest city of Beni. The triatomine captures were performed with mice-baited adhesive traps mostly settled in palm trees in forest fragments and pastures. Species were identified by morphological observation, dissection of genitalia, and sequencing of three mitochondrial gene fragments and one nuclear fragment. Molecular analysis was based on (i) the identity score of the haplotypes with GenBank sequences through the BLAST algorithm and (ii) construction of phylogenetic trees. Thirty-four triatomines, all belonging to the Rhodnius genus, of which two were adult males, were captured in palm trees in forest fragments and pastures (overall infestation rate, 12.3%). The morphology of the phallic structures in the two males confirmed the R. stali species. For the other specimens, after molecular sequencing, only one specimen was identified with confidence as belonging to Rhodnius robustus, the others belonged to one of the species of the Rhodnius pictipes complex, probably Rhodnius stali. The two species, R. robustus and R. stali, had previously been reported in the Alto Beni region (edge of the Amazon region), but not yet in the Beni department situated in the Amazon region. Furthermore, the difficulties of molecular characterization of closely related species within the three complexes of the genus Rhodnius are highlighted and discussed.
Assuntos
Doença de Chagas/transmissão , DNA Mitocondrial/genética , Proteínas de Insetos/genética , Insetos Vetores/classificação , Filogenia , Rhodnius/classificação , Trypanosoma cruzi/genética , Animais , Arecaceae , Bolívia/epidemiologia , Núcleo Celular/genética , Doença de Chagas/epidemiologia , Feminino , Haplótipos , Humanos , Insetos Vetores/genética , Insetos Vetores/parasitologia , Masculino , Mitocôndrias/genética , Rhodnius/genética , Rhodnius/parasitologia , Trypanosoma cruzi/isolamento & purificaçãoRESUMO
Trypanosoma cruzi, the causative agent of Chagas disease, presents wide genetic diversity. Currently, six discrete typing units (DTUs), named TcI to TcVI, and a seventh one called TcBat are used for strain typing. Beyond the debate concerning this classification, this systematic review has attempted to provide an inventory by compiling the results of 137 articles that have used it. A total of 6,343 DTU identifications were analyzed according to the geographical and host origins. Ninety-one percent of the data available is linked to South America. This sample, although not free of potential bias, nevertheless provides today's picture of T. cruzi genetic diversity that is closest to reality. DTUs were genotyped from 158 species, including 42 vector species. Remarkably, TcI predominated in the overall sample (around 60%), in both sylvatic and domestic cycles. This DTU known to present a high genetic diversity, is very widely distributed geographically, compatible with a long-term evolution. The marsupial is thought to be its most ancestral host and the Gran Chaco region the place of its putative origin. TcII was rarely sampled (9.6%), absent, or extremely rare in North and Central America, and more frequently identified in domestic cycles than in sylvatic cycles. It has a low genetic diversity and has probably found refuge in some mammal species. It is thought to originate in the south-Amazon area. TcIII and TcIV were also rarely sampled. They showed substantial genetic diversity and are thought to be composed of possible polyphyletic subgroups. Even if they are mostly associated with sylvatic transmission cycles, a total of 150 human infections with these DTUs have been reported. TcV and TcVI are clearly associated with domestic transmission cycles. Less than 10% of these DTUs were identified together in sylvatic hosts. They are thought to originate in the Gran Chaco region, where they are predominant and where putative parents exist (TcII and TcIII). Trends in host-DTU specificities exist, but generally it seems that the complexity of the cycles and the participation of numerous vectors and mammal hosts in a shared area, maintains DTU diversity.
Assuntos
Doença de Chagas/parasitologia , Trypanosoma cruzi/classificação , Animais , Evolução Biológica , América Central , DNA de Protozoário/genética , Variação Genética , Genótipo , Geografia , Humanos , América do Norte , América do Sul , Trypanosoma cruzi/isolamento & purificaçãoRESUMO
BACKGROUND: Chagas disease is a major public health problem in Latin America. Its etiologic agent, Trypanosoma cruzi, is mainly transmitted through the contaminated faeces of blood-sucking insects called triatomines. Triatoma infestans is the main vector in various countries in South America and recently, several foci of wild populations of this species have been described in Bolivia and other countries. These wild populations are suspected of affecting the success of insecticide control campaigns being carried out in South America. To assess the risk that these T. infestans populations pose to human health, it is helpful to determine blood meal sources. METHODS: In the present work, blood meals were identified in various Bolivian wild T. infestans populations and in three specific areas, in both wild and intra-peridomestic populations to assess the links between wild and domestic cycles of T. cruzi transmission. PCR-HDA and sequencing of Cytb gene were used to identify these blood meal sources. RESULTS AND DISCUSSION: Fourteen vertebrate species were identified as wild blood meal sources. Of those, the most prevalent species were two Andean endemic rodents, Octodontomys gliroides (36%) and Galea musteloides (30%), while humans were the third most prevalent source (18.7%). Of 163 blood meals from peridomestic areas, more than half were chickens, and the others were generally domestic animals or humans. Interestingly, blood from wild animals was identified in triatomines captured in the peridomestic and domestic environment, and blood from domestic animals was found in triatomines captured in the wild, revealing links between wild and domestic cycles of T. cruzi transmission. CONCLUSION: The current study suggests that wild T. infestans attack humans in the wild, but is also able to bite humans in domestic settings before going back to its natural environment. These results support the risk to human health posed by wild populations of T. infestans.
Assuntos
Animais Domésticos/parasitologia , Animais Selvagens/parasitologia , Doença de Chagas/veterinária , Insetos Vetores/parasitologia , Triatoma/parasitologia , Trypanosoma cruzi/fisiologia , Animais , Animais Domésticos/sangue , Animais Domésticos/classificação , Animais Selvagens/sangue , Animais Selvagens/classificação , Sangue/parasitologia , Bolívia/epidemiologia , Doença de Chagas/epidemiologia , Doença de Chagas/parasitologia , Doença de Chagas/transmissão , Humanos , Insetos Vetores/fisiologia , Triatoma/fisiologia , Trypanosoma cruzi/genética , Trypanosoma cruzi/isolamento & purificaçãoRESUMO
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.
Assuntos
Tipagem de Sequências Multilocus/métodos , Parasitologia/métodos , Trypanosoma cruzi/genética , GenótipoRESUMO
Trypanosoma cruzi, the causative agent of Chagas disease, is subdivided into six discrete typing units (DTUs; TcI-TcVI) of which TcI is ubiquitous and genetically highly variable. While clonality is the dominant mode of propagation, recombinant events play a significant evolutive role. Recently, foci of wild Triatoma infestans have been described in Bolivia, mainly infected by TcI. Hence, for the first time, we evaluated the level of genetic exchange within TcI natural potentially panmictic populations (single DTU, host, area and sampling time). Seventy-nine TcI stocks from wild T. infestans, belonging to six populations were characterized at eight microsatellite loci. For each population, Hardy-Weinberg equilibrium (HWE), linkage disequilibrium (LD), and presence of repeated multilocus genotypes (MLG) were analyzed by using a total of seven statistics, to test the null hypothesis of panmixia (H0). For three populations, none of the seven statistics allowed to rejecting H0; for another one the low size did not allow us to conclude, and for the two others the tests have given contradictory results. Interestingly, apparent panmixia was only observed in very restricted areas, and was not observed when grouping populations distant of only two kilometers or more. Nevertheless it is worth stressing that for the statistic tests of "HWE", in order to minimize the type I error (i. e. incorrect rejection of a true H0), we used the Bonferroni correction (BC) known to considerably increase the type II error ( i. e. failure to reject a false H0). For the other tests (LD and MLG), we did not use BC and the risk of type II error in these cases was acceptable. Thus, these results should be considered as a good indicator of the existence of panmixia in wild environment but this must be confirmed on larger samples to reduce the risk of type II error.
Assuntos
Triatoma/parasitologia , Trypanosoma cruzi/isolamento & purificação , Animais , Bolívia , Variação Genética , Interações Hospedeiro-Parasita , Desequilíbrio de Ligação , Repetições de Microssatélites/genética , Triatoma/genéticaRESUMO
Triatoma infestans, the major vector of Chagas disease south of the Amazon in South America, has a large distribution of wild populations, contrary to what has previously been stated. These populations have been suspected of being the source of reinfestation of human habitats and could impede the full success of vector control campaigns. This study examined gene flow between intra-peridomestic populations and wild populations collected in the surround areas in three Andean localities in Bolivia. The populations were defined according to temporal, ecological, and spatial criteria. After DNA extraction from the legs of each insect, the samples were analyzed using seven microsatellite markers. First, the analysis of molecular variance (AMOVA) detected an absence of differentiation between wild and intra-peridomestic populations, although strong structuring was observed between the populations within each environment. Then for some populations, the Bayesian method of assignment to inferred populations showed very similar assignment patterns of the members of wild or intra-peridomestic populations in each locality. Finally, the detection of the first-generation migrants within the different populations provided evidence of insect displacement from the wild to the intra-peridomestic environment. This result indicates that, after control campaigns in the Andes, controlling this new paradigm of vector transmission risk stemming from the invasion of human habitats by wild populations of T. infestans requires long-term maintenance of public monitoring to keep the risk at a minimal level. Since wild populations of T. infestans have also been detected elsewhere in Argentina, Paraguay, and Chile, there is an urgent need to take these populations into account in future monitoring of Chagas disease transmission.
Assuntos
Ecossistema , Triatoma/genética , Animais , Bolívia , Feminino , Loci Gênicos/genética , Variação Genética , Genótipo , Geografia , Humanos , Masculino , Repetições de Microssatélites/genética , Filogenia , Dinâmica PopulacionalRESUMO
Trypanosoma cruzi the agent of Chagas disease is a monophyletic but heterogeneous group conformed by several Discrete Typing Units (DTUs) named TcI to TcVI characterized by genetic markers. The trans-sialidase (TS) is a virulence factor involved in cell invasion and pathogenesis that is differentially expressed in aggressive and less virulent parasite stocks. Genes encoding TS-related proteins are included in a large family divided in several groups but only one of them contains TS genes. Two closely related genes differing in a T/C transition encode the enzymatically active TS (aTS) and a lectin-like TS (iTS). We quantified the aTS/iTS genes from TcII and TcVI aggressive and TcI low virulent strains and found variable aTS number (1-32) per haploid genome. In spite of being low TS enzyme-expressers, TcI strains carry 28-32 aTS gene copies. The intriguing absence of iTS genes in TcI strains together with the presence of aTS/iTS in TcII and TcVI strains (virulent) were observed. Moreover, after sequencing aTS/iTS from 38 isolates collected along the Americas encompassing all DTUs, the persistent absence of the iTS gene in TcI, TcIII and TcIV was found. In addition, the sequence clustering together with T/C transition analysis correlated to DTUs of T. cruzi. The consistence of TS results with both evolutionary genome models proposed for T. cruzi, namely the "Two Hybridization" and the "Three Ancestor" was discussed and reviewed to fit present findings. Parasite stocks to attempt genetic KO or to assay the involvement of iTS in parasite biology and virulence are finally available.
Assuntos
Genes de Protozoários , Glicoproteínas/genética , Neuraminidase/genética , Trypanosoma cruzi/classificação , Trypanosoma cruzi/genética , Fatores de Virulência/genética , Doença de Chagas/parasitologia , Códon , Genoma de Protozoário , Dados de Sequência Molecular , Filogenia , Polimorfismo de Nucleotídeo ÚnicoRESUMO
The TcI discrete typing unit (DTU) of Trypanosoma cruzi is the most abundant and widely spread in the Americas. It is found in a wide range of triatomine and mammal species, which are distributed throughout the Americas in sylvatic and domestic environments. Previous studies based on intergenic sequences of the mini-exon gene (SL-IR) have identified five genotype groups within TcI. Based in the large number of sequences available in GenBank, the present study conducted an exhaustive revision of the sequence variability of the SL-IR within TcI using 244 sequences from isolates, cellular or molecular clones, from 11 Latin American countries. First, the evolutionary branching between strains was examined by analyzing only the single nucleotide polymorphism (SNP) deleting the microsatellite region and the gaps from the total alignment. Then the variability of the microsatellite region was re-analyzed alone using principal component analysis (PCA). After haplotype reconstruction using the PHASE algorithm, because of the presence of several ambiguous nucleotides in the SNP region, a total of 131 different haplotypes were obtained. The topology reveals how difficult it is to identify an obvious structure in TcI for most of the parameters examined. Somewhat genetic and geographical structures exist, but no structure was depicted with cycle and host origins. Indeed, the long-lasting evolution with possible recombination events, the occurrence of several waves of geographical dispersions (old and recent), and the high flow of strains between sylvatic and domestic cycles partially hide the major evolutionary trends within TcI. Moreover, we identified several problems in previous analyses, and concluded that in absence of supplementary studies of TcI phylogeny with other genetic markers, it is hazardous to use only the mini-exon intergenic region as a relevant marker of the substructure within TcI.
Assuntos
DNA Intergênico/genética , Éxons/genética , Trypanosoma cruzi/genética , Animais , Sequência de Bases , Doença de Chagas/parasitologia , DNA de Protozoário/genética , Evolução Molecular , Haplótipos , Humanos , Dados de Sequência Molecular , Polimorfismo de Nucleotídeo Único/genética , Análise de Componente Principal , Alinhamento de Sequência , América do SulRESUMO
In the Gran Chaco region the reinfestation by Triatoma infestans remains a major problem for control of Chagas disease. Trypanosoma cruzi the agent of the illness presents a broad genetic intraspecific variability which is poorly documented in the Bolivian Gran Chaco. This work presents the identification of the discrete typing units (DTUs) currently recognized for T. cruzi in T. infestans populations collected before and after residual insecticide spraying in four villages in this region. Before spraying, of 84 samples, the frequencies of the DTUs identified by using the multiplex PCR based on the non transcribed spacer of the mini-exon gene (MMPCR) were 0.21 for TcI, 0.70 for TcII/TcV/TcVI, and 0.17 for TcIII/TcIV and no significant difference was observed after spraying (76 samples). Moreover 13% of the total sample corresponds to T. infestans specimens with mixed infection of DTUs of which three were TcII/TcV/TcVI with TcIII/TcIV. The partial sequences of T. cruzi Gpi gene obtained from 14 PCR products agree the MMPCR DTU identification and allowed to precise the occurrence of TcIII, TcII and hybrid TcV/TcVI stocks which were not discriminated by the MMPCR. Given the high prevalence of hybrid stocks, the authors ask whether the recombination event at the origin of hybrids would have taken place in the Gran Chaco where the putative parents are also present.
Assuntos
Triatoma/parasitologia , Trypanosoma cruzi/genética , Animais , Bolívia/epidemiologia , Doença de Chagas/epidemiologia , Doença de Chagas/parasitologia , DNA de Protozoário , Feminino , Genótipo , Glucose-6-Fosfato Isomerase/genética , Humanos , Controle de Insetos , Masculino , Filogenia , Proteínas de Protozoários/genéticaRESUMO
Trypanosoma cruzi, the agent of Chagas disease, presents a predominantly clonal structure that has been shaped by recombination events leading to six genetic groups (DTUs, discrete typing units, TcI-TcVI). Several conventional and unconventional genetic exchange events have been described, including hybridization and mitochondrial introgression, which is explored here among Bolivian and Peruvian strains belonging to TcI because recombination events have been previously suspected by means of the MLMT method (multilocus microsatellite typing). We analyzed the variation of one nuclear (Gpi) and one mitochondrial (Nd1) gene among 60 TcI strains and 15 reference strains belonging to the six DTUs. The results clearly showed that one strain isolated from Triatoma infestans in the Cochabamba department (Bolivia) presented a genotype TcI for Gpi and a mitochondrial Nd1 genotype common to the DTUs TcIII, IV, V, and VI; this can be interpreted as a mitochondrial introgression event between distant DTUs. These kinds of events, although probably scarce, may have played an important role in the adaptive evolution of the species.
Assuntos
Doença de Chagas/parasitologia , Genes Mitocondriais , Genes de Protozoários , Hibridização Genética , Trypanosoma cruzi/genética , Animais , Sequência de Bases , Doença de Chagas/veterinária , Variação Genética , Haplótipos , Humanos , Dados de Sequência Molecular , Filogenia , Recombinação Genética , América do Sul , Triatoma/parasitologiaRESUMO
BACKGROUND: The current persistence of Triatoma infestans (one of the main vectors of Chagas disease) in some domestic areas could be related to re-colonization by wild populations which are increasingly reported. However, the infection rate and the genetic characterization of the Trypanosoma cruzi strains infecting these populations are very limited. METHODOLOGY/PRINCIPAL FINDINGS: Of 333 wild Triatoma infestans specimens collected from north to south of a Chagas disease endemic area in Bolivia, we characterized 234 stocks of Trypanosoma cruzi using mini-exon multiplex PCR (MMPCR) and sequencing the glucose phosphate isomerase (Gpi) gene. Of the six genetic lineages ("discrete typing units"; DTU) (TcI-VI) presently recognized in T. cruzi, TcI (99.1%) was overdominant on TcIII (0.9%) in wild Andean T. infestans, which presented a 71.7% infection rate as evaluated by microscopy. In the lowlands (Bolivian Chaco), 17 "dark morph" T. infestans were analyzed. None of them were positive for parasites after microscopic examination, although one TcI stock and one TcII stock were identified using MMPCR and sequencing. CONCLUSIONS/SIGNIFICANCE: By exploring large-scale DTUs that infect the wild populations of T. infestans, this study opens the discussion on the origin of TcI and TcV DTUs that are predominant in domestic Bolivian cycles.