RESUMO
Of the currently known 140 species in the family Reduviidae, subfamily Triatominae, those which are most important as vectors of the aetiologic agent of Chagas disease, Trypanosoma cruzi, belong to the tribes Triatomini and Rhodniini. The latter not only transmit T. cruzi but also Trypanosoma rangeli, which is considered apathogenic for the mammalian host but can be pathogenic for the vectors. Using different molecular methods, two main lineages of T. cruzi have been classified, T. cruzi I and T. cruzi II. Within T. cruzi II, five subdivisions are recognized, T. cruzi IIa-IIe, according to the variability of the ribosomal subunits 24Salpha rRNA and 18S rRNA. In T. rangeli, differences in the organization of the kinetoplast DNA separate two forms denoted T. rangeli KP1+ and KP1-, although differences in the intergenic mini-exon gene and of the small subunit rRNA (SSU rRNA) suggest four subpopulations denoted T. rangeli A, B, C and D. The interactions of these subpopulations of the trypanosomes with different species and populations of Triatominae determine the epidemiology of the human-infecting trypanosomes in Latin America. Often, specific subpopulations of the trypanosomes are transmitted by specific vectors in a particular geographic area. Studies centered on trypanosome-triatomine interaction may allow identification of co-evolutionary processes, which, in turn, could consolidate hypotheses of the evolution and the distribution of T. cruzi/T. rangeli-vectors in America, and they may help to identify the mechanisms that either facilitate or impede the transmission of the parasites in different vector species. Such mechanisms seem to involve intestinal bacteria, especially the symbionts which are needed by the triatomines to complete nymphal development and to produce eggs. Development of the symbionts is regulated by the vector. T. cruzi and T. rangeli interfere with this system and induce the production of antibacterial substances. Whereas T. cruzi is only subpathogenic for the insect host, T. rangeli strongly affects species of the genus Rhodnius and this pathogenicity seems based on a reduction of the number of symbionts.
Assuntos
Vetores de Doenças , Interações Hospedeiro-Parasita , Triatominae/fisiologia , Triatominae/parasitologia , Trypanosoma cruzi/fisiologia , Trypanosoma cruzi/patogenicidade , Animais , DNA de Cinetoplasto/genética , DNA de Protozoário/genética , DNA Ribossômico/genética , América Latina , Filogenia , Polimorfismo Genético , Trypanosoma cruzi/classificação , Trypanosoma cruzi/genéticaRESUMO
We present data on the molecular characterisation of strains of Trypanosoma rangeli isolated from naturally infected Rhodnius ecuadoriensis in Peru, from Rhodnius colombiensis, Rhodnius pallescens and Rhodnius prolixus in Colombia, and from Rhodnius pallescens in Panama. Strain characterisation involved a duplex PCR with S35/S36/KP1L primers. Mini-exon gene analysis was also carried out using TrINT-1/TrINT-2 oligonucleotides. kDNA and mini-exon amplification indicated dimorphism within both DNA sequences: (i) KP1, KP2 and KP3 or (ii) KP2 and KP3 products for kDNA, and 380 bp or 340 bp products for the mini-exon. All T. rangeli strains isolated from R. prolixus presented KP1, KP2 and KP3 products with the 340 bp mini-exon product. By contrast, all T. rangeli strains isolated from R. ecuadoriensis, R. pallescens and R. colombiensis, presented profiles with KP2 and KP3 kDNA products and the 380 bp mini-exon product. Combined with other studies, these results provide evidence of co-evolution of T. rangeli strains associated with different Rhodnius species groups east and west of the Andean mountains.
Assuntos
Evolução Molecular , Rhodnius/parasitologia , Trypanosoma/genética , Animais , Colômbia , DNA Intergênico/genética , DNA de Cinetoplasto/genética , DNA de Protozoário/genética , Éxons/genética , Interações Hospedeiro-Parasita/genética , Panamá , Peru , Filogenia , Trypanosoma/classificaçãoRESUMO
Domiciliated Rhodnius prolixus and sylvatic R. colombiensis were analyzed in order to confirm their genetic divergence and verify the risk that the latter represents in the domiciliation process, and to provide tools for identifying the sources of possible reinfestation by triatomines in human dwellings allowing control programs to be undertaken. Comparison of random amplified polymorphic DNA amplification patterns and cluster analysis suggests reproductive discontinuity between the two species. The calculated statistical F value of 0.24 and effective migration rate of 0.6 individuals per generation are insufficient to maintain genetic homogeneity between them and confirm the absence of present genetic flow. R. colombiensis presents higher intrapopulation variability. Polymerase chain reaction of ribosomal DNA supports these findings. The low genetic flow between the two species implies that R. colombiensis do not represent an epidemiological risk for the domiciliary transmission of Trypanosoma cruzi in the Tolima Department. The lower variability of the domiciliated R. prolixus could result in greater susceptibility to the use of pesticides in control programs.
Assuntos
DNA Ribossômico/análise , Insetos Vetores/genética , Técnica de Amplificação ao Acaso de DNA Polimórfico , Rhodnius/genética , Animais , Primers do DNA/análise , Primers do DNA/genética , Humanos , Polimorfismo Genético , Especificidade da EspécieRESUMO
Human Chagas disease is a purely accidental occurrence. As humans came into contact with the natural foci of infection might then have become infected as a single addition to the already extensive host range of Trypanosoma cruzi that includes other primates. Thus began a process of adaptation and domiciliation to human habitations through which the vectors had direct access to abundant food as well as protection from climatic changes and predators. Our work deals with the extraction and specific amplification by polymerase chain reaction of T. cruzi DNA obtained from mummified human tissues and the positive diagnosis of Chagas disease in a series of 4, 000-year-old Pre-Hispanic human mummies from the northern coast of Chile. The area has been inhabited at least for 7,000 years, first by hunters, fishers and gatherers, and then gradually by more permanent settlements. The studied specimens belonged to the Chinchorro culture, a people inhabiting the area now occupied by the modern city of Arica. These were essentially fishers with a complex religious ideology, which accounts for the preservation of their dead in the way of mummified bodies, further enhanced by the extremely dry conditions of the desert. Chinchorro mummies are, perhaps, the oldest preserved bodies known to date.
Assuntos
Doença de Chagas/transmissão , Emigração e Imigração , Transmissão Vertical de Doenças Infecciosas , Múmias/parasitologia , Trypanosoma cruzi , Animais , Doença de Chagas/história , Doença de Chagas/parasitologia , Chile , DNA de Protozoário/análise , História Antiga , Humanos , Transmissão Vertical de Doenças Infecciosas/história , Trypanosoma cruzi/isolamento & purificaçãoRESUMO
Trypanosoma rangeli can infect humans as well as the same domestic and wild animals and triatomine vectors infected by Trypanosoma cruzi in Central and South America. This overlapping distribution complicates the epidemiology of American trypanosomiasis due to the cross-reactivity between T. rangeli and T. cruzi antigens and the presence of conserved DNA sequences in these parasites. We have isolated a T. rangeli-specific DNA repetitive element which is represented in approximately 103 copies per parasite genome and is distributed in several chromosomal bands. The 542-bp nucleotide sequence of this element, named P542, was determined and a PCR assay was standardized for its amplification. The sensitivity of the assay is high, allowing the detection of one tenth of the DNA content of a single parasite. The presence of the P542 element was confirmed in 11 T. rangeli isolates from mammalian hosts and insect vectors originating from several countries in Latin America. Negative amplification was observed with different T. cruzi strains and other trypanosomatids. The potential field application of the P542 PCR assay was investigated in simulated samples containing T. rangeli and/or T. cruzi and intestinal tract and feces of Rhodnius prolixus. Epidemiological studies were conducted in DNA preparations obtained from the digestive tracts of 12 Rhodnius colombiensis insects collected in a sylvatic area in Colombia. Positive amplification of the P542 element was obtained in 9/12 insects. We have also compared in the same samples the diagnostic performance of two PCR assays for the amplification of the variable domain of minicircle kinetoplast DNA (kDNA) and of the large subunit (LSU) of the ribosomal RNA gene of T. cruzi and T. rangeli. Data indicate that the kDNA PCR assay does not allow diagnosis of mixed infections in most insects. On the other hand, the PCR assay of the LSU RNA gene showed lower sensitivity in the detection of T. rangeli than the PCR assay of the P542 element. It is predicted that the use of sensitive detection techniques will indicate that the actual distribution of T. rangeli in America is wider than presumed.
Assuntos
Doença de Chagas/epidemiologia , DNA de Protozoário/química , Sequências Repetitivas de Ácido Nucleico , Trypanosoma/isolamento & purificação , Animais , Sequência de Bases , Southern Blotting , Doença de Chagas/diagnóstico , Doença de Chagas/parasitologia , Colômbia/epidemiologia , Enzimas de Restrição do DNA , DNA de Cinetoplasto/química , DNA de Cinetoplasto/isolamento & purificação , DNA de Protozoário/isolamento & purificação , Diagnóstico Diferencial , Eletroforese em Gel de Ágar , Eletroforese em Gel de Campo Pulsado , Humanos , Insetos Vetores/parasitologia , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , RNA Ribossômico/genética , Rhodnius/parasitologia , Sensibilidade e Especificidade , Análise de Sequência de DNA , Especificidade da Espécie , Trypanosoma/genéticaRESUMO
A segment of DNA unique to the kinetoplast of Trypanosoma cruzi was isolated from spontaneously mummified human remains from the coastal area of northern Chile at sites dated from 2000 BC to about AD 1400. Following rehydration of the desiccated human tissue samples of heart, esophagus, or colon, the samples were extracted and primers employed to bind to a 330 bp kinetoplast minicircle DNA sequence present in T. cruzi. This segment was then amplified using the polymerase chain reaction (PCR), and the target segment was visualized by gel electrophoresis. This method enables the identification of Chagas' disease in an ancient body in the absence of recognizable anatomic pathological changes.
Assuntos
DNA de Protozoário/isolamento & purificação , Múmias , Trypanosoma cruzi/genética , Adolescente , Adulto , Animais , Doença de Chagas/epidemiologia , Doença de Chagas/transmissão , Criança , Chile/epidemiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da PolimeraseRESUMO
Several groups have recently developed molecular tests for the detection of Trypanosoma cruzi, the causative agent of Chagas' disease. Polymerase chain reaction (PCR) amplification of kinetoplast minicircle DNA sequences appears to be the most sensitive method. However, the specificity of PCR-based diagnostic methods was challenged when the complete sequence of Trypanosoma rangeli DNA minicircles was discovered. In the present study, we conducted. PCR experiments using the S35/S36 primers in Rhodnius prolixus and Balb/c mice with single and mixed infections of T. cruzi and/or T. rangeli. In single infections, the profile of each trypanosome was easily distinguishable in haemolymph, salivary gland and intestinal tissues and faeces of insect vectors. In mixed infections of anterior intestine (where T. rangeli is more predominant than T. cruzi), the DNA amplification profile of both parasites was observed simultaneously. Conversely, only the T. cruzi profile was observed in rectal ampulla (where T. cruzi is more abundant than T. rangeli). In mice with single infections of T. cruzi or T. rangeli, the profiles of amplified DNA were easily distinguishable in each case. The T. cruzi profile was dominant in most mixed infections, probably due to the fact that T. cruzi minicircles are more abundant and consequently compete more eagerly for annealing with the S35/S36 primers. In cases of mixed infections where T. rangeli was initially more abundant than T. cruzi, the specific T. rangeli 760 bp band was present for 7 days after infection and then this band and others ranging from 300 to 450 bp disappeared and only the typical T. cruzi 330 bp band remained. The S35/S36 primers used in polyacrylamide gel electrophoresis (PAGE) detected T. cruzi specifically, and prevented misdiagnosis due to the presence of T. rangeli. This technique can also be used to identify parasites in different stages of the infection (acute or chronic) in vertebrate hosts and to localize the parasites in the insect vector.
Assuntos
Doença de Chagas/diagnóstico , DNA de Cinetoplasto/análise , Insetos Vetores/parasitologia , Rhodnius/parasitologia , Trypanosoma/isolamento & purificação , Tripanossomíase/diagnóstico , Animais , Sequência de Bases , Doença de Chagas/parasitologia , Sequência Consenso , DNA de Cinetoplasto/isolamento & purificação , Camundongos , Dados de Sequência Molecular , Reação em Cadeia da Polimerase/métodos , Especificidade da Espécie , Trypanosoma/genética , Trypanosoma cruzi/genética , Trypanosoma cruzi/isolamento & purificação , Tripanossomíase/parasitologiaRESUMO
Extracts of 176 species of Colombian plant seeds, corresponding to 49 families and 147 genera, were tested for detecting agglutinins against human red blood cells from A+, B+ and O+ groups, dog, horse and rabbit. Extracts with haemagglutination activity were used for agglutination of Trypanosoma cruzi and T. rangeli. In addition the hemolymph of 16 native species of invertebrates were tested in the same conditions. Serial dilution of extracts were used for agglutination reactions. Both T. cruzi and T. rangeli epimastigotes showed agglutination with the extract of seven different species of plant seeds and with two types of haemolymph of invertebrates. The seeds of five plants exclusively agglutinated the epimastigotes of T. cruzi and thus can be used for the differentiation between culture forms of the trypanosomes. The secretion of the lung of a snail (Bulimus sp.) lysed entirely the epimastigotes of T. cruzi but did not affect T. rangeli forms. No extracts were found which agglutinated or lysed exclusively the epimastigotes of T. rangeli.