Interrogating the transmission dynamics of Trypanosoma cruzi (Trypanosomatida, Trypanosomatidae) by Triatoma venosa (Hemiptera: Reduviidae) after the elimination of vector transmission by Rhodnius prolixus in Boyacá eastern Colombia.

Chagas disease (CD) is a parasitic zoonosis (Trypanosoma cruzi) that is endemic in Colombia. Vector control of Rhodnius prolixus, the main domestic T. cruzi vector, has been achieved in a large part of the area with historically vector transmission of CD. It is necessary to understand the ecological behavior characteristics of local native vectors to ensure sustainability of the vector control programs. To evaluate the long-term success of a recent vector control campaign in the Boyacá department (Colombia), we used a combined strategy of entomological surveillance with co-existing canine surveillance from ten rural villages within six municipalities of the Tenza valley region (Boyacá, Colombia): Chinavita, Garagoa, Guateque, Somondoco, Sutatenza and Tenza, with historical reports of R. prolixus and secondary vectors. Collected triatomines and canine whole blood were analyzed for T. cruzi infection and genotyping. Triatomine bugs specimens were evaluated for blood meal source. Canine serology was performed using two distinct antibody assays. In total, 101 Triatoma venosa were collected by active search in domestic and peridomestic habitats. A natural infection prevalence of 13.9% (14/101) and four feeding sources were identified: human, dog, rat, and hen. A frequency infection of 46.5% (40/87) was observed from two independent serological tests and T. cruzi DNA was detected in 14 dogs (16.4%). Only TcIsylvatic DTU was detected. The results suggest that T. venosa present eco-epidemiological characteristics to maintain the transmission of T. cruzi in Tenza valley. This species has reinfested the intervened households and it has an active role in domestic and peridomestic transmission of T. cruzi due to their infection rates and feeding behavior. Therefore, this species should be considered as epidemiologically relevant for vector control strategies. Moreover, there is a need for human serological studies to have a close up of risk they are exposed to.

Curvas de fusión de regiones genómicas específicas: una herramienta prometedora para el diagnóstico y tipificación de las especies causantes de la leishmaniasis cutánea en Colombia / High-resolution melting analysis based on specific genomic regions: A promising tool for the diagnosis and typing of species causing cutaneous leishmaniasis in Colombia

Resumen Introducción. La leishmaniasis cutánea es una enfermedad causada por parásitos del género Leishmania que tiene gran incidencia en Colombia. El diagnóstico y la identificación de la especie infecciosa son factores críticos en el momento de escoger e iniciar el tratamiento. Actualmente, los métodos de diagnóstico y tipificación requieren procedimientos complejos, por lo que es necesario validar nuevos marcadores moleculares y métodos que simplifiquen el proceso. Objetivo. Desarrollar una herramienta basada en la reacción en cadena de la polimerasa (PCR) con curvas de fusión (High Resolution Melting; PCR-HRM) para el diagnóstico y tipificación de las tres especies de Leishmania de importancia epidemiológica en casos de leishmaniasis cutánea en Colombia. Materiales y métodos. Los genomas de Leishmania panamensis, L. braziliensis y L. guyanensis se compararon mediante métodos bioinformáticos. Las regiones específicas de especie identificadas se validaron mediante PCR. Para los marcadores seleccionados se diseñó una PCR-HRM y se estimaron algunos parámetros de validez y seguridad usando aislamientos de pacientes colombianos caracterizados previamente mediante PCR y análisis de polimorfismos en la longitud de los fragmentos de restricción (Restriction Fragment Length Polymorphism - RFLP; PCR-RFLP) del gen hsp70. Resultados. El análisis genómico comparativo mostró 24 regiones específicas de especie. Sin embargo, la validación mediante PCR solo identificó un marcador específico para cada especie de Leishmania. Los otros marcadores mostraron amplificación cruzada. El límite de detección para los tres marcadores seleccionados fue de un parásito, mientras que la sensibilidad, la especificidad, el valor predictivo positivo y el negativo fueron de 91,4, 100, 100 y 75 %, respectivamente. Conclusiones. Las tres regiones seleccionadas pueden emplearse como marcadores moleculares en el diagnóstico y tipificación de las especies causantes de la leishmaniasis cutánea en Colombia.

Abstract Introduction: Cutaneous leishmaniasis, caused by parasites of the genus Leishmania, is a disease with high incidence in Colombia. The diagnosis and identification of the infectious species are critical factors when selecting and initiating treatment. Currently, the methods for diagnosing and typing cutaneous leishmaniasis require complicated procedures and there is a need for the validation of new molecular markers and methods to simplify the process. Objective: To develop a tool based in PCR melting curves (PCR-HRM) for the diagnosis and typing of the three Leishmania species of epidemiological importance for cutaneous leishmaniasis in Colombia. Materials and methods: The genomes of Leishmania panamensis, L. braziliensis and L. guyanensis were compared with bioinformatic methods. The species-specific regions were then validated using PCR. For the selected markers, a PCR-HRM was designed, and validity and security parameters were estimated using isolates from Colombian patients previously characterized by PCR-RFLP of the hsp70 gene. Results: The comparative genomic analysis yielded 24 species-specific regions. However, the PCR validation identified only one marker that was specific to each Leishmania species. The other markers showed cross amplification. The detection limit for the three selected markers was one parasite. The sensitivity, specificity, predictive positive and negative values were 91.4%, 100%, 100% and 75%, respectively. Conclusions: The three selected regions can be used as molecular markers in the diagnosis and typing of the causative species of cutaneous leishmaniasis in Colombia.

Aedes albopictus (Skuse, 1894) infected with the American-Asian genotype of dengue type 2 virus in Medellín suggests its possible role as vector of dengue fever in Colombia.

INTRODUCTION: Aedes aegypti and Ae. albopictus are recognized vectors of dengue, yellow fever, chikungunya and Zika arboviruses in several countries worldwide. In Colombia, Ae. albopictus geographical distribution has increased to include highly populated cities such as Cali and Medellín. Although this species has been frequently found in urban and semi-urban zones in the country, its role as vector of the dengue fever is poorly known. OBJECTIVE: To identify the presence of Ae. albopictus specimens naturally infected with dengue virus collected in Medellín. MATERIALS AND METHODS: Insects were collected in the Universidad Nacional de Colombia campus in Medellín. Individuals were classified as Ae. albopictus and confirmed by DNA barcode region analysis. Mosquitoes were processed for dengue virus identification, and a fragment of the NS3 gen was sequenced and compared with DENV-2 genotypes reported in the literature. RESULTS: Sequence analysis of COI indicated Ae. albopictus individuals were similar to those recently reported in Colombia, and genetically close to those from other regions worldwide. Among the pools tested one was positive for DENV-2, and the NS3 analysis indicated it belonged to the Asian-American clade. CONCLUSION: We report the presence Ae. albopictus naturally infected with the Asian-American genotype of DENV-2 in Colombia. The presence of Ae. albopictus specimens carrying the most common genotype infecting humans in a highly populated city such as Medellín indicates its potential role as dengue vector in Colombia and highlights the relevance of including it in current vector surveillance strategies.

Virological surveillance of Aedes (Stegomyia) aegypti and Aedes (Stegomyia) albopictus as support for decision making for dengue control in Medellín.

INTRODUCTION: Dengue represents an important public health problem in Colombia. No treatment is available and the vaccine has not been approved in all countries, hence, actions should be strengthened to mitigate its impact through the control of Aedes aegypti, the vector mosquito. In Colombia, surveillance is done using entomological indexes and case notification, which is usually informed late, leading to untimely interventions. Viral detection in urban mosquitoes using molecular techniques provides more accurate entomological information for decision-making. OBJECTIVE: To report results of virological surveillance in Aedes specimens collected during routine entomological activities of the Secretaría de Salud de Medellín. MATERIALS AND METHODS: Specimens were collected during two periods in each of which we selected 18 dwellings around each one of the 250 larva traps arranged for mosquitoe surveillance, as well as 70 educational institutions and 30 health centers. Specimens were identified morphologically, and divided in pools for viral detection using reverse transcription polymerase chain reaction (RT-PCR). We calculated the minimum infection rate and the adult infestation index for each group. RESULTS: We collected 1,507 adult mosquitoes, 10 of which were identified as A. albopictus. Out of the 407 pools, 132 (one of them Ae. albopictus) were positive, and 14.39% were A. aegypti males. The minimum infection rates for Ae. aegypti were 120.07 and 69,50 for the first and second periods, respectively, and the adult infestation index was higher in educational institutions (23.57%). CONCLUSIONS: Using RT-PCR we identified natural infectivity and vertical transmission of dengue virus in A. aegypti and A. albopictus. We suggest the use of molecular techniques in arbovirosis surveillance and control programs in Colombia.

Infección de Aedes albopictus (Skuse, 1894) con el genotipo asiático-americano del virus del dengue serotipo 2 en Medellín y su posible papel como vector del dengue en Colombia / Aedes albopictus (Skuse, 1894) infected with the American-Asian genotype of dengue type 2 virus in Medellín suggests its possible role as vector of dengue fever in Colombia

Resumen Introducción. Aedes aegypti y Ae. albopictus son reconocidos vectores de arbovirus como los del dengue, la fiebre amarilla, el chikungunya y el Zika, en regiones tropicales y subtropicales del mundo. En Colombia, la distribución geográfica de Ae. albopictus ha sufrido un incremento y hoy incluye ciudades como Cali y Medellín. Hasta ahora, sin embargo, no se ha recabado información concluyente sobre su infección viral y su capacidad de transmisión a los humanos. Objetivo. Determinar la infección natural por dengue en ejemplares de Ae. albopictus recolectados en un área urbana de Medellín. Materiales y métodos. Se recolectaron individuos de Ae. albopictus en el campus de la Universidad Nacional de Colombia, sede Medellín. Se confirmó su clasificación taxonómica mediante el análisis del gen citocromo oxidasa I (COI), y se extrajo el ARN total para la identificación del virus del dengue y de los respectivos serotipos. La presencia del genotipo DENV se infirió mediante el análisis del gen NS3. Resultados. El análisis del COI corroboró el estatus taxonómico de Ae. albopictus. Uno de los mosquitos procesados fue positivo para DENV-2 y el análisis del NS3 mostró una gran similitud con el genotipo asiático-americano. Conclusión. Se reporta la infección con DENV-2 en Ae. albopictus en Medellín, Colombia. La presencia del genotipo asiático-americano en una zona urbana sugiere su posible circulación entre humanos y en Ae. albopictus, lo cual alerta sobre su eventual papel en la transmisión del DENV-2, y sobre la necesidad de incluir esta especie en la vigilancia entomológica en Colombia.

Abstract Introduction: Aedes aegypti and Ae. albopictus are recognized vectors of dengue, yellow fever, chikungunya and Zika arboviruses in several countries worldwide. In Colombia, Ae. albopictus geographical distribution has increased to include highly populated cities such as Cali and Medellín. Although this species has been frequently found in urban and semi-urban zones in the country, its role as vector of the dengue fever is poorly known. Objective: To identify the presence of Ae. albopictus specimens naturally infected with dengue virus collected in Medellín. Materials and methods: Insects were collected in the Universidad Nacional de Colombia campus in Medellín. Individuals were classified as Ae. albopictus and confirmed by DNA barcode region analysis. Mosquitoes were processed for dengue virus identification, and a fragment of the NS3 gen was sequenced and compared with DENV-2 genotypes reported in the literature. Results: Sequence analysis of COI indicated Ae. albopictus individuals were similar to those recently reported in Colombia, and genetically close to those from other regions worldwide. Among the pools tested one was positive for DENV-2, and the NS3 analysis indicated it belonged to the Asian-American clade. Conclusion: We report the presence Ae. albopictus naturally infected with the Asian-American genotype of DENV-2 in Colombia. The presence of Ae. albopictus specimens carrying the most common genotype infecting humans in a highly populated city such as Medellín indicates its potential role as dengue vector in Colombia and highlights the relevance of including it in current vector surveillance strategies.

Vigilancia virológica de Aedes (Stegomyia) aegypti y Aedes (Stegomyia) albopictus como apoyo para la adopción de decisiones en el control del dengue en Medellín / Virological surveillance of Aedes (Stegomyia) aegypti and Aedes (Stegomyia) albopictus as support for decision making for dengue control in Medellín

Resumen Introducción. El dengue en Colombia representa un grave problema de salud y, dado que no existe un tratamiento efectivo para la enfermedad y la vacuna no se ha aprobado en todos los países, se deben fortalecer acciones para mitigar su impacto mediante el control de Aedes aegypti, el mosquito vector. La vigilancia en el país se hace con base en los índices entomológicos y en la notificación de casos, la cual es frecuentemente tardía y por ello conduce a falta de oportunidad en las intervenciones. La detección viral en mosquitos urbanos mediante técnicas moleculares proporciona información entomológica más precisa para la adopción de decisiones. Objetivo. Reportar los resultados de la vigilancia virológica de especímenes de Aedes spp. recolectados durante actividades entomológicas rutinarias de la Secretaría de Salud de Medellín. Materiales y métodos. Los ejemplares se recolectaron durante dos periodos, en cada uno de los cuales se seleccionaron 18 viviendas alrededor de cada una de las 250 trampas para larvas dispuestas para la vigilancia entomológica, así como 70 instituciones educativas y 30 centros de salud. Los ejemplares se identificaron y se conformaron grupos para la detección viral mediante reacción en cadena de la polimerasa con transcripción inversa (Reverse Transcription Polymerase Chain Reaction, RT-PCR). Se calculó la tasa mínima de infección y el índice de infestación en adultos. Resultados. Se recolectaron 1.507 mosquitos, 10 de los cuales eran Ae. albopictus. De los 407 grupos conformados, 132 (uno de ellos de Ae. albopictus) fueron positivos, y 14,39 % correspondió a machos de Ae. aegypti. La tasa mínima de infección para Ae. aegypti fue de 120,07 y 69,50 en el primer y segundo períodos, respectivamente, y el índice de infestación en adultos fue mayor en las instituciones educativas (23,57 %). Conclusión. Mediante la RT-PCR se detectaron la infección natural y la transmisión vertical del virus del dengue en Ae. aegypti y en Ae. albopictus. Se propone considerar la incorporación de estas técnicas moleculares en los programas de vigilancia y control de las arbovirosis en el país.

Abstract Introduction: Dengue represents an important public health problem in Colombia. No treatment is available and the vaccine has not been approved in all countries, hence, actions should be strengthened to mitigate its impact through the control of Aedes aegypti, the vector mosquito. In Colombia, surveillance is done using entomological indexes and case notification, which is usually informed late, leading to untimely interventions. Viral detection in urban mosquitoes using molecular techniques provides more accurate entomological information for decision-making. Objective: To report results of virological surveillance in Aedes specimens collected during routine entomological activities of the Secretaría de Salud de Medellín. Materials and methods: Specimens were collected during two periods in each of which we selected 18 dwellings around each one of the 250 larva traps arranged for mosquitoe surveillance, as well as 70 educational institutions and 30 health centers. Specimens were identified morphologically, and divided in pools for viral detection using reverse transcription polymerase chain reaction (RT-PCR). We calculated the minimum infection rate and the adult infestation index for each group. Results: We collected 1,507 adult mosquitoes, 10 of which were identified as A. albopictus. Out of the 407 pools, 132 (one of them Ae. albopictus) were positive, and 14.39% were A. aegypti males. The minimum infection rates for Ae. aegypti were 120.07 and 69,50 for the first and second periods, respectively, and the adult infestation index was higher in educational institutions (23.57%). Conclusions: Using RT-PCR we identified natural infectivity and vertical transmission of dengue virus in A. aegypti and A. albopictus. We suggest the use of molecular techniques in arbovirosis surveillance and control programs in Colombia.

Curvas de fusión de regiones genómicas específicas: una herramienta prometedora para el diagnóstico y tipificación de las especies causantes de la leishmaniasis cutánea en Colombia.

Introducción. La leishmaniasis cutánea es una enfermedad causada por parásitos del género Leishmania que tiene gran incidencia en Colombia. El diagnóstico y la identificación de la especie infecciosa son factores críticos en el momento de escoger e iniciar el tratamiento. Actualmente, los métodos de diagnóstico y tipificación requieren procedimientos complejos, por lo que es necesario validar nuevos marcadores moleculares y métodos que simplifiquen el proceso.Objetivo. Desarrollar una herramienta basada en la reacción en cadena de la polimerasa (PCR) con curvas de fusión (High Resolution Melting; PCR-HRM) para el diagnóstico y tipificación de las tres especies de Leishmania de importancia epidemiológica en casos de leishmaniasis cutánea en Colombia.Materiales y métodos. Los genomas de Leishmania panamensis, L. braziliensis y L. guyanensis se compararon mediante métodos bioinformáticos. Las regiones específicas de especie identificadas se validaron mediante PCR. Para los marcadores seleccionados se diseñó una PCR-HRM y se estimaron algunos parámetros de validez y seguridad usando aislamientos de pacientes colombianos caracterizados previamente mediante PCR y análisis de polimorfismos en la longitud de los fragmentos de restricción (Restriction Fragment Length Polymorphism - RFLP; PCR-RFLP) del gen hsp70.Resultados. El análisis genómico comparativo mostró 24 regiones específicas de especie. Sin embargo, la validación mediante PCR solo identificó un marcador específico para cada especie de Leishmania. Los otros marcadores mostraron amplificación cruzada. El límite de detección para los tres marcadores seleccionados fue de un parásito, mientras que la sensibilidad, la especificidad, el valor predictivo positivo y el negativo fueron de 91,4, 100, 100 y 75 %, respectivamente.Conclusiones. Las tres regiones seleccionadas pueden emplearse como marcadores moleculares en el diagnóstico y tipificación de las especies causantes de la leishmaniasis cutánea en Colombia.

Specific primers design based on the superoxide dismutase b gene for Trypanosoma cruzi as a screening tool: Validation method using strains from Colombia classified according to their discrete typing unit.

OBJECTIVE: To classify 21 new isolates of Trypanosoma cruzi (T. cruzi) according to the Discrete Typing Unit (DTU) which they belong to, as well as tune up a new pair of primers designed to detect the parasite in biological samples. METHODS: Strains were isolated, DNA extracted, and classified by using three Polymerase Chain Reactions (PCR). Subsequently this DNA was used along with other isolates of various biological samples, for a new PCR using primers designed. Finally, the amplified fragments were sequenced. RESULTS: It was observed the predominance of DTU I in Colombia, as well as the specificity of our primers for detection of T. cruzi, while no band was obtained when other species were used. CONCLUSIONS: This work reveals the genetic variability of 21 new isolates of T. cruzi in Colombia.Our primers confirmed their specificity for detecting the presence of T. cruzi.

[Investigation of vectors and reservoirs in an acute Chagas outbreak due to possible oral transmission in Aguachica, Cesar, Colombia]. / Investigación de vectores y reservorios en brote de Chagas agudo por posible transmisión oral en Aguachica, Cesar, Colombia.

Colombia recorded 11 cases of acute Chagas disease and 80 cases of oral contamination with Trypanosoma cruzi. The current study analyzes the entomological and parasitological characteristics of the outbreak in Aguachica, Cesar Department, in 2010. An interdisciplinary group of health professionals and regional university personnel conducted the laboratory tests in the patients and the investigation of the transmission focus. Eleven cases of acute Chagas diseases were detected in a single family in a dwelling with domiciliated triatomines and Rhodnius pallescens, Pantrongylus geniculatus, Eratyrus cuspidatus, and two Didelphis marsupialis opossums infected with T. cruzi in Attalea butyracea and Elaeis oleifera palm trees in the urban area of Aguachica. The study analyzes the role of R. pallescens and palm trees in the wild cycle of T. cruzi and in oral transmission of Chagas disease. Sporadic incursions by wild R. pallescens, P. geniculatus, and E. cuspidatus from the nearby palm trees into human dwellings may cause increasingly frequent outbreaks of oral Chagas disease.

Investigación de vectores y reservorios en brote de Chagas agudo por posible transmisión oral en Aguachica, Cesar, Colombia / Investigation of vectors and reservoirs in an acute Chagas outbreak due to possible oral transmission in Aguachica, Cesar, Colombia / Investigação dos vetores e reservatórios ao surto da doença de Chagas agudo por possível transmissão oral em Aguachica, Cesar, Colômbia

Colombia tiene un registro de 11 casos de Chagas agudo y 80 casos por contaminación oral con Trypanosoma cruzi. Este trabajo analiza los hallazgos entomológicos y parasitológicos del brote de Aguachica, Cesar, en 2010. Un grupo interdisciplinario de profesionales de la salud y de universidades regionales realizó las pruebas de laboratorio a los pacientes y el estudio del foco de transmisión. Se detectaron 11 casos agudos de enfermedad de Chagas en una sola familia con vivienda sin triatominos domiciliados y, Rhodnius pallescens, Pantrongylus geniculatus, Eratyrus cuspidatus y dos Didelphis marsupialis infectados con T. cruzi en palmas de Attalea butyracea y Elaeis oleifera del área urbana de Aguachica. Se analiza la participación del R. pallescens y el rol de las palmas en el ciclo silvestre de T. cruzi y para la transmisión oral de la enfermedad de Chagas. Incursiones esporádicas de R. pallescens, P. geniculatus y E. cuspidatus silvestres desde palmas cercanas al domicilio humano pueden provocar brotes cada vez más frecuentes de Chagas oral.

Colombia recorded 11 cases of acute Chagas disease and 80 cases of oral contamination with Trypanosoma cruzi. The current study analyzes the entomological and parasitological characteristics of the outbreak in Aguachica, Cesar Department, in 2010. An interdisciplinary group of health professionals and regional university personnel conducted the laboratory tests in the patients and the investigation of the transmission focus. Eleven cases of acute Chagas diseases were detected in a single family in a dwelling with domiciliated triatomines and Rhodnius pallescens, Pantrongylus geniculatus, Eratyrus cuspidatus, and two Didelphis marsupialis opossums infected with T. cruzi in Attalea butyracea and Elaeis oleifera palm trees in the urban area of Aguachica. The study analyzes the role of R. pallescens and palm trees in the wild cycle of T. cruzi and in oral transmission of Chagas disease. Sporadic incursions by wild R. pallescens, P. geniculatus, and E. cuspidatus from the nearby palm trees into human dwellings may cause increasingly frequent outbreaks of oral Chagas disease.

A Colômbia tem registro de 11 surtos da doença de Chagas e 80 casos agudos por contaminação oral com Trypanosoma cruzi. Esta pesquisa analisa os achados entomológicos e parasitológicos do surto de Aguachica, Cesar, em 2010. Um grupo interdisciplinar de profissionais da saúde e de universidades regionais efetuou os testes laboratoriais nos pacientes e o estudo de foco de transmissão. Encontraram 11 casos agudos da doença de Chagas em uma única família com domicilio sem triatomíneos e Rhodnius pallescens, Pantrongylus geniculatus, Eratyrus cuspidatus e dois Didelphis marsupialis infectados com T. cruzi em palmeiras de Attalea butyracea e Elaeis oleifera da área urbana de Aguachica. Analisa-se a participação de R. pallescens e a função das palmeiras em o ciclo silvestre de T. cruzi e para a transmissão oral da doença de Chagas. Incursões esporádicas de R. pallescens, P. geniculatus e E. cuspidatus silvestres das palmeiras próximas ao domicilio humano podem provocar surtos cada vez mais frequentes de Chagas oral.

Molecular evidence of demographic expansion of the chagas disease vector Triatoma dimidiata (Hemiptera, Reduviidae, Triatominae) in Colombia.

BACKGROUND: Triatoma dimidiata is one of the most significant vectors of Chagas disease in Central America and Colombia, and, as in most species, its pattern of genetic variation within and among populations is strongly affected by its phylogeographic history. A putative origin from Central America has been proposed for Colombian populations, and high genetic differentiation among three biographically different population groups has recently been evidenced. Analyses based on putatively neutral markers provide data from which past events, such as population expansions and colonization, can be inferred. We analyzed the genealogies of the nicotinamide adenine dinucleotide dehydrogenase 4 (ND4) and the cytochrome oxidase subunit 1-mitochondrial genes, as well as partial nuclear ITS-2 DNA sequences obtained across most of the eco-geographical range in Colombia, to assess the population structure and demographic factors that may explain the geographical distribution of T. dimidiata in this country. RESULTS: The population structure results support a significant association between genetic divergence and the eco-geographical location of population groups, suggesting that clear signals of demographic expansion can explain the geographical distribution of haplotypes of population groups. Additionally, empirical date estimation of the event suggests that the population's expansion can be placed after the emergence of the Panama Isthmus, and that it was possibly followed by a population fragmentation process, perhaps resulting from local adaptation accomplished by orographic factors such as geographical isolation. CONCLUSION: Inferences about the historical population processes in Colombian T. dimidiata populations are generally in accordance with population expansions that may have been accomplished by two important biotic and orographic events such as the Great American Interchange and the uplift of the eastern range of the Andes mountains in central Colombia.

Genetic, cytogenetic and morphological trends in the evolution of the Rhodnius (Triatominae: Rhodniini) trans-Andean group.

The Rhodnius Pacific group is composed of three species: Rhodnius pallescens, R. colombiensis and R. ecuadoriensis, which are considered important vectors of trypanosomes (Trypanosoma cruzi and T. rangeli) infecting humans. This group is considered as a recent trans-Andean lineage derived from the widespread distributed sister taxa R. pictipes during the later uplift of northern Andes mountain range. The widest spread species R. pallescens may be a complex of two divergent lineages with different chromosomal attributes and a particular biogeographical distribution across Central America and Colombia with several southern populations in Colombia occupying the same sylvatic habitat as its sister species R. colombiensis. Although the taxonomy of Rhodnius Pacific group has been well studied, the unresolved phylogenetic and systematic issues are the target of this paper. Here we explore the molecular phylogeography of this species group analyzing two mitochondrial (ND4 and cyt b) and one nuclear (D2 region of ribosomal 28S gene) gene sequences. The molecular analyses suggest an early divergence of the species R. ecuadoriensis and R. colombiensis, followed by a recent expansion of R. pallescens lineages. The phylogenetic relationship between sympatric R. pallescens Colombian lineage and R. colombiensis was further explored using wing morphometry, DNA genome size measurements, and by analyzing chromosomal behavior of hybrids progeny obtained from experimental crosses. Our results suggest that the diversification of the two R. pallescens lineages was mainly influenced by biogeographical events such as (i) the emergence of the Panama Isthmus, while the origin and divergence of R. colombiensis was associated with (ii) the development of particular genetic and chromosomal features that act as isolation mechanisms from its sister species R. pallescens (Colombian lineage). These findings provide new insights into the evolution of the Rhodnius Pacific group and the underlying biological processes that occurred during its divergence.

Identificación de nuevos escenarios epidemiológicos para la enfermedad de Chagas en la región momposina, norte de Colombia / Identification of new epidemiological scenarios for Chagas disease in the Momposina region, North of Colombia

Introducción. La región momposina es una de las zonas de mayor interés en Colombia para estudiar nuevos escenarios epidemiológicos de la enfermedad de Chagas. Objetivo. Determinar el riesgo epidemiológico de infección con Trypanosoma cruzi en seis poblaciones humanas de la región momposina, según el patrón climático bimodal característico de Colombia. Materiales y métodos. Se hicieron cuatro muestreos, dos en época de lluvia y dos en sequía. Las heces de triatominos, como también muestras de sangre de humanos y otros animales mamíferos reservorios, se evaluaron por ELISA, IFI y PCR. Además, para determinar factores de riesgo se analizaron las condiciones de las viviendas, el estado alimentario de triatominos y la participación de los perros en el ciclo de transmisión. Resultados. Los insectos y mamíferos, incluyendo los humanos, presentaron altas tasas de infección por T. cruzi , con diferencias entre las épocas climáticas y según la vegetación asociada. Las tasas de infección por T. cruzi en perros fue de 15 % y, en humanos, de 16,8 %. La gran densidad de población de insectos, el alto porcentaje de insectos alimentados y la alta tasa de infección, indican que en la época seca hay mayor riesgo de infección. Conclusión. Se confirma la presencia y circulación de un foco peridoméstico de T. cruzi y se describen algunos factores de riesgo eco-epidemiológicos para la zona, que sugieren nuevos escenarios epidemiológicos para la enfermedad de Chagas en Colombia.

Introduction: The Momposina region is one of the most interesting areas to study new epidemiological scenarios for Chagas disease in Colombia. Objectives: To determine the presence of a source of peridomestic transmission of T. cruzi and its epidemiological risk in the Momposina region, based on the bimodal weather pattern characteristic of Colombia. Materials and methods: Four surveys over two years (two in the rainy season and two during the dry one) were conducted. Triatomines feces and blood samples from human and reservoirs were evaluated for presence of antibodies and parasites by ELISA, IFI and PCR. The conditions of housing, feeding triatomine state and involvement of dogs in the transmission were assessed. Results. High rates of infection with T. cruzi in insects and wild animals were found. Infection rates of T. cruzi in dogs (15%) and humans (16.8%) were found. The results obtained in this study indicated that in the dry season there is increased risk of infection with T. cruzi , given the higher population density of insects, the higher percentage of fed insects and the higher rate of infection. Conclusion: These results confirm the presence and movement of a peridomestic outbreak of T. cruzi and describes some risk factors for the eco-epidemiological area, suggesting new epidemiological scenarios for Chagas disease in Colombia.

Eco-geographical differentiation among Colombian populations of the Chagas disease vector Triatoma dimidiata (Hemiptera: Reduviidae).

Triatoma dimidiata is currently the main vector of Chagas disease in Mexico, most Central American countries and several zones of Ecuador and Colombia. Although this species has been the subject of several recent phylogeographic studies, the relationship among different populations within the species remains unclear. To elucidate the population genetic structure of T. dimidiata in Colombia, we analyzed individuals from distinct geographical locations using the cytochrome c oxidase subunit 1 gene and 7 microsatellite loci. A clear genetic differentiation was observed among specimens from three Colombian eco-geographical regions: Inter Andean Valleys, Caribbean Plains and Sierra Nevada de Santa Marta mountain (SNSM). Additionally, evidence of genetic subdivision was found within the Caribbean Plains region as well as moderate gene flow between the populations from the Caribbean Plains and SNSM regions. The genetic differentiation found among Colombian populations correlates, albeit weakly, with an isolation-by-distance model (IBD). The genetic heterogeneity among Colombian populations correlates with the eco-epidemiological and morphological traits observed in this species across regions within the country. Such genetic and epidemiological diversity should be taken into consideration for the development of vector control strategies and entomological surveillance.

[Identification of new epidemiological scenarios for Chagas disease in the Momposina region, North of Colombia]. / Identificación de nuevos escenarios epidemiológicos para la enfermedad de Chagas en la región momposina, norte de Colombia.

INTRODUCTION: The Momposina region is one of the most interesting areas to study new epidemiological scenarios for Chagas disease in Colombia. OBJECTIVES: To determine the presence of a source of peridomestic transmission of T. cruzi and its epidemiological risk in the Momposina region, based on the bimodal weather pattern characteristic of Colombia. MATERIALS AND METHODS: Four surveys over two years (two in the rainy season and two during the dry one) were conducted. Triatomines feces and blood samples from human and reservoirs were evaluated for presence of antibodies and parasites by ELISA, IFI and PCR. The conditions of housing, feeding triatomine state and involvement of dogs in the transmission were assessed. RESULTS: High rates of infection with T. cruzi in insects and wild animals were found. Infection rates of T. cruzi in dogs (15%) and humans (16.8%) were found. The results obtained in this study indicated that in the dry season there is increased risk of infection with T. cruzi , given the higher population density of insects, the higher percentage of fed insects and the higher rate of infection. CONCLUSION: These results confirm the presence and movement of a peridomestic outbreak of T. cruzi and describes some risk factors for the eco-epidemiological area, suggesting new epidemiological scenarios for Chagas disease in Colombia.

Reconstructing Native American population history.

The peopling of the Americas has been the subject of extensive genetic, archaeological and linguistic research; however, central questions remain unresolved. One contentious issue is whether the settlement occurred by means of a single migration or multiple streams of migration from Siberia. The pattern of dispersals within the Americas is also poorly understood. To address these questions at a higher resolution than was previously possible, we assembled data from 52 Native American and 17 Siberian groups genotyped at 364,470 single nucleotide polymorphisms. Here we show that Native Americans descend from at least three streams of Asian gene flow. Most descend entirely from a single ancestral population that we call 'First American'. However, speakers of Eskimo-Aleut languages from the Arctic inherit almost half their ancestry from a second stream of Asian gene flow, and the Na-Dene-speaking Chipewyan from Canada inherit roughly one-tenth of their ancestry from a third stream. We show that the initial peopling followed a southward expansion facilitated by the coast, with sequential population splits and little gene flow after divergence, especially in South America. A major exception is in Chibchan speakers on both sides of the Panama isthmus, who have ancestry from both North and South America.

Morphometric and molecular evidence of intraspecific biogeographical differentiation of Rhodnius pallescens (HEMIPTERA: REDUVIIDAE: RHODNIINI) from Colombia and Panama.

Rhodnius pallescens is considered the main vector of Chagas disease in Panama and a relevant secondary vector in northern Colombia. Previous data reported that this species presents cytogenetically heterogeneous populations, which are probably biogeographically segregated. To provide new information on the diversity of R. pallescens, we compared several populations from Colombia and Panama based on the morphometric analyses of wings, mitochondrial cytochrome b (cyt b) gene sequencing, and genomic DNA measurements. Although no differences in DNA amount were detected, significant differences in cyt b sequences as well as wing size and shape were identified among populations. The results obtained in this work indicate R. pallescens comprises two evolutionary lineages with genetic and morphological differences that could be explained by their geographic isolation in distinct ecological zones. These results provide new insight into R. pallescens population diversity and the underlying biological processes that shape its evolution.

Actividad tripanocida y citotóxica de extractos de plantas colombianas / Trypanocidal and cytotoxic activity of extracts of Colombian plants

Introducción. El tratamiento de la enfermedad de Chagas está basado en sólo dos medicamentos de eficacia limitada y con importantes efectos colaterales. La gran biodiversidad de la flora colombiana hace de la bioprospección una alternativa potencial en la búsqueda de nuevos antiparasitarios. Objetivo. Evaluar in vitro el potencial tripanocida y la citotoxicidad de extractos obtenidos de 23 plantas colombianas. Materiales y métodos. Se obtuvieron extractos de hojas, de tallos o de la planta entera, en solventes de diferente polaridad. La actividad contra epimastigotes y la citotoxicidad se evaluaron por el micrométodo enzimático con MTT. Los extractos activos contra epimastigotes y con baja citotoxicidad se evaluaron también en tripomastigotes y amastigotes intracelulares. Resultados. Se reporta la actividad tripanocida de 13 plantas colombianas y se confirma el efecto biológico de cuatro especies previamente evaluadas. Cuatro extractos activos en epimastigotes también fueron activos en tripomastigotes y, uno de ellos, en amastigotes. Este extracto fue aislado de la planta Hieronyma antioquensis, y presentó CI50 de 3,125, 11,48 y 2,85 µg/ml, e índices de selectividad de 25,7 y 27, para epimastigotes, tripomastigotes y amastigotes, respectivamente. Los resultados sugieren que este extracto es un candidato promisorio para el tratamiento de la enfermedad de Chagas. Conclusión. La flora colombiana es una fuente potencial de nuevas sustancias para la quimioterapia contra la enfermedad de Chagas. El micrométodo enzimático con MTT es una herramienta útil para la tamización de la actividad biológica en epimastigotes y posterior selección para ensayos con otros estadios del parásito.

Introduction. The treatment of Chagas disease is based on only two drugs with limited efficacy and significant side effects. The rich biodiversity of the Colombian flora makes bio-prospecting a potential alternative in the search for new antiparasitic drugs. Objective. Potential trypanocidal activity and cytotoxicity was assessed in extracts from 23 Colombian plants. Materials and methods. Extracts of leaves, stems, or of the whole plants were obtained in solvents of a range of polarities. The activity against Trypanosoma cruzi epimastigotes and the cytotoxicity were evaluated by the MTT enzymatic micro-method. Extracts active against epimastigotes and with lowcytotoxicity were also tested on trypomastigotes and intracellular amastigotes. Results. Among the extracts, biological activity was confirmed in 4 species. The extracts were active on epimastigotes and trypomastigotes; one was active also against amastigotes. The latter extract was isolated from the plant Hieronyma antioquensis and presented IC50 of 3.1 mg/ml for epimastigotes, 11.5mg/ml for trypomastigotes and 2.9 mg/ml for amastigotes. The selectivity indexes were 25, 7, and 27 respectively. Conclusions. The extract from H. antioquensis proved a promising candidate for Chagas disease treatment. Futhermore, the MTT enzymatic micromethod was a useful tool for screening biological activity on epimastigotes and other stages of the parasite for further extract trials.

Mitochondrial dysfunction in Trypanosoma cruzi: the role of Serratia marcescens prodigiosin in the alternative treatment of Chagas disease.

BACKGROUND: Chagas disease is a health threat for many people, mostly those living in Latin America. One of the most important problems in treatment is the limitation of existing drugs. Prodigiosin, produced by Serratia marcescens (Rhodnius prolixus endosymbiont), belongs to the red-pigmented bacterial prodiginine family, which displays numerous biological activities, including antibacterial, antifungal, antiprotozoal, antimalarial, immunosuppressive, and anticancer properties. Here we describe its effects on Trypanosoma cruzi mitochondria belonging to Tc I and Tc II. RESULTS: Parasites exposed to prodigiosin altered the mitochondrial function and oxidative phosphorylation could not have a normal course, probably by inhibition of complex III. Prodigiosin did not produce cytotoxic effects in lymphocytes and Vero cells and has better effects than benznidazole. Our data suggest that the action of prodigiosin on the parasites is mediated by mitochondrial structural and functional disruptions that could lead the parasites to an apoptotic-like cell death process. CONCLUSIONS: Here, we propose a potentially useful trypanocidal agent derived from knowledge of an important aspect of the natural life cycle of the parasite: the vector-parasite interaction. Our results indicate that prodigiosin could be a good candidate for the treatment of Chagas disease.

[Trypanocidal and cytotoxic activity of extracts of Colombian plants]. / Actividad tripanocida y citotóxica de extractos de plantas colombianas.

INTRODUCTION: The treatment of Chagas disease is based on only two drugs with limited efficacy and significant side effects. The rich biodiversity of the Colombian flora makes bio-prospecting a potential alternative in the search for new antiparasitic drugs. OBJECTIVE: Potential trypanocidal activity and cytotoxicity was assessed in extracts from 23 Colombian plants. MATERIALS AND METHODS: Extracts of leaves, stems, or of the whole plants were obtained in solvents of a range of polarities. The activity against Trypanosoma cruzi epimastigotes and the cytotoxicity were evaluated by the MTT enzymatic micro-method. Extracts active against epimastigotes and with low cytotoxicity were also tested on trypomastigotes and intracellular amastigotes. RESULTS: Among the extracts, biological activity was confirmed in 4 species. The extracts were active on epimastigotes and trypomastigotes; one was active also against amastigotes. The latter extract was isolated from the plant Hieronyma antioquensis and presented IC(50) of 3.1 mg/ml for epimastigotes, 11.5 mg/ml for trypomastigotes and 2.9 mg/ml for amastigotes. The selectivity indexes were 25, 7, and 27 respectively. CONCLUSIONS: The extract from H. antioquensis proved a promising candidate for Chagas disease treatment. Futhermore, the MTT enzymatic micromethod was a useful tool for screening biological activity on epimastigotes and other stages of the parasite for further extract trials.