Chagas Disease and Leishmaniasis, Sympatric Zoonoses Present in Human from Rural Communities of Venezuela.

PURPOSE: Trypanosoma cruzi and Leishmania spp. coexist in several endemic areas, and there are few studies of Chagas disease and leishmaniasis coinfection worldwide; for this reason, the objective of this work was to determine the Chagas disease and leishmaniasis coinfection in several rural communities co-endemic for these diseases. METHODS: A total of 1107 human samples from six co-endemic rural communities of Cojedes state, Venezuela, were analyzed. Serum samples were evaluated by ELISA, indirect hemagglutination, and indirect immunofluorescence for Chagas disease diagnosis, and individuals were evaluated for leishmaniasis by leishmanin skin test (LST). Approximately, 30% of the individuals were also analyzed by PCR (blood clot samples) for T. cruzi and for Leishmania spp. RESULTS: The 14.7% of the individuals were positive to Trypanosoma cruzi infection by serology, and 25.8% were positive to Leishmania spp. current or past infection by LST. Among the group with PCR results, 7.8% were positive for T. cruzi, and 9.4% for Leishmania spp. The coinfection T. cruzi/Leishmania spp. was 6.5%. The T. cruzi DTUs of the positive blood clot samples were TcI, revealed using the molecular markers: (i) intergenic region of the miniexon, (ii) D7 divergent domain of the 24Sα rDNA, (iii) size-variable domain of the 18S rDNA, and (iv) hsp60-PCR-RFLP (EcoRV). The Leishmania species identified were L. (Leishmania) mexicana and L. (Viannia) braziliensis. CONCLUSION: A high prevalence was found for T. cruzi and Leishmania spp. single and coinfections in almost all communities studied, being these results of relevance for the implementation of control programs in co-endemic areas.

Parasitological and molecular characterization of Trypanosoma cruzi of triatomines and mammals from endemic areas for Chagas disease in Venezuela.

It is estimated that 6-7 million people worldwide are infected with Trypanosoma cruzi, the parasite that causes Chagas disease. In Venezuela, Chagas disease remains a public health problem. In this work, T. cruzi isolates from six species of triatomines and mammals of the orders Didelphimorphia and Xenarthra, captured in rural communities of Monagas, underwent parasitological and molecular characterization. A total of 471 triatomines and 17 mammals were captured, with a natural infection rate of 41.4% and 70.6%, respectively. In the male NMRI mouse model used for parasitological characterization (prepatent period, parasitemia curve, mouse mortality, and tissular parasitism), T. cruzi isolates exhibited high lethality due to their pronounced virulence, irrespective of the parasite load in each mouse, resulting in a mortality rate of 75%. Among the vector isolates, in the mouse model, only 2 out of 6 remained alive, while the rest perished during the evaluation. Conversely, the isolates from mammals proved fatal for all the inoculated mice. All isolates were identified as belonging to DTU TcI, based on the molecular markers such as the intergenic region of the miniexon, D7 divergent domain of the 24Sα rDNA, size-variable domain of the 18S rDNA, and hsp60-PCR-RFLP-EcoRV. This study demonstrates the presence of vectors and mammalian reservoirs naturally infected with T. cruzi in communities of Monagas, the 9th largest and 9th most populous state in Venezuela. This situation represents a neglected epidemiological problem demanding urgent attention and imperative health care intervention.

Molecular detection of the natural infection by trypanosomatid parasites in Didelphis marsupialis from a rural area in northern Colombia. / Detección molecular de infección natural por parásitos tripanosomátidos en Didelphis marsupialis de una zona rural del norte de Colombia.

We studied the prevalence of infection by trypanosomatid parasites in Didelphis marsupialis and its relationship with morphological/age aspects in a rural area of El Carmen de Bolivar, Colombia. Five visits were made to the Vereda El Alférez; each of which lasted three consecutive nights. During these visits, Tomahawk® traps were installed in the peridomestic and wild ecotopes of the Vereda El Alférez. Body measurements, sex and age were determined from the collected animals. Blood was extracted by cardiopuncture, after sedation, in order to obtain total deoxyribonucleic acid (DNA) and amplify the conserved region of the kinetoplast minicircle DNA (kDNA) of parasitic trypanosomatids. The association between morphological parameters of didelphids and their frequency of infection by parasitic trypanosomatids was determined by binomial regression. Thirty D. marsupialis specimens (60.0% females and 40.0% males/66.7% adults and 33.3% juveniles) were collected. Molecular diagnosis revealed a frequency of trypanosomatid parasite infection of 46.7%. Stage (p=0.024) was a determinant for infection. We discuss the role of D. marsupialis as a potential reservoir of parasitic trypanosomatids in the Vereda El Alférez.

Se evaluó la prevalencia de infección por parásitos tripanosomátidos en Didelphis marsupialis y su relación con los aspectos morfológicos/etarios en una zona rural de El Carmen de Bolívar, Colombia. En cinco visitas (2018-2019) se instalaron trampas Tomahawk® en los ecótopos peridoméstico y silvestre en la Vereda El Alférez, durante tres noches consecutivas/visita. A los animales recolectados, se les determinaron medidas corporales, sexo y edad; y se les extrajo sangre por cardiopuntura, previa sedación, para extracción del ácido desoxirribonucleico (ADN) total y amplificación de la región conservada del ADN de minicírculos de kinetoplasto (ADNk) de parásitos tripanosomátidos. La asociación entre parámetros morfológicos de los didélfidos y su frecuencia de infección por parásitos tripanosomátidos fue determinada mediante una regresión binomial. Se recolectaron 30 individuos de D. marsupialis (60,0% hembras y 40,0% machos/66,7% adultos y 33,3% juveniles). El diagnóstico molecular reveló una frecuencia de infección por parásitos tripanosomátidos del 46,7%. El estadio (p=0,024) fue determinante para la infección. Se discute el papel de D. marsupialis como potencial reservorio de parásitos tripanosomátidos en la zona evaluada.

Geographic Distribution of the Genus Panstrongylus Berg, 1879 in the Neotropic with Emphasis on Trypanosoma cruzi Vectors.

Panstrongylus is a Neotropical taxa of 16 species, some more widespread than others, that act as vectors of Trypanosoma cruzi, the etiologic agent of Chagas disease (CD). This group is associated with mammalian reservoir niches. There are few studies of the biogeography and niche suitability of these triatomines. Using zoo-epidemiological occurrence databases, the distribution of Panstrongylus was determined based on bioclimatic modelling (DIVA GIS), parsimonious niche distribution (MAXENT), and parsimony analysis of endemic species (PAE). Through 517 records, a wide presence of P. geniculatus, P. rufotuberculatus, P. lignarius, and P. megistus was determined and recorded as frequent vectors of T. cruzi in rainforest habitats of 24-30 °C. These distributions were modeled with AUC >0.80 and <0.90, as well as with the seasonality of temperature, isothermality, and precipitation as relevant bioclimatic variables. Individual traces for each taxon in Panstrongylus-1036 records-showed widely dispersed lines for frequent vectors P. geniculatus, P. lignarius, P. rufotuberculatus, and P. megistus. Other occasional vectors showed more restricted dispersal, such as P. howardi, P. humeralis, P. lenti, P. lutzi, P. tupynambai, P. noireaiui, and P. chinai. Areas of defined environmental variation, geological change, and trans domain fluid fauna, such as the American Transition Zone and the Pacific Domain of Morrone, had the highest Panstrongylus diversity. Pan-biogeographic nodes appear to be areas of the greatest species diversity that act as corridors connecting biotopes and allowing fauna migration. Vicariance events in the geologic history of the continent need to be investigated. The geographical distribution of Panstrongylus overlapped with CD cases and Didelphis marsupialis/Dasypus novemcinctus presence, two important reservoirs in Central and South America. The information derived from the distribution of Panstrongylus provides knowledge for surveillance and vector control programs. It would increase information on the most and less relevant vector species of this zoonotic agent, for monitoring their population behavior.

Detección molecular de la infección natural por parásitos tripanosomátidos en Didelphis marsupialis de una zona rural del norte de Colombia / Molecular detection of the natural infection by trypanosomatid parasites in Didelphis marsupialis from a rural area in northern Colombia

Se evaluó la prevalencia de infección por parásitos tripanosomátidos en Didelphis marsupialis y su relación con los aspectos morfológicos/etarios en una zona rural de El Carmen de Bolívar, Colombia. En cinco visitas (2018-2019) se instalaron trampas Tomahawk® en los ecótopos peridoméstico y silvestre en la Vereda El Alférez, durante tres noches consecutivas/visita. A los animales recolectados, se les determinaron medidas corporales, sexo y edad; y se les extrajo sangre por cardiopuntura, previa sedación, para extracción del ácido desoxirribonucleico (ADN) total y amplificación de la región conservada del ADN de minicírculos de kinetoplasto (ADNk) de parásitos tripanosomátidos. La asociación entre parámetros morfológicos de los didélfidos y su frecuencia de infección por parásitos tripanosomátidos fue determinada mediante una regresión binomial. Se recolectaron 30 individuos de D. marsupialis (60,0% hembras y 40,0% machos/66,7% adultos y 33,3% juveniles). El diagnóstico molecular reveló una frecuencia de infección por parásitos tripanosomátidos del 46,7%. El estadio (p=0,024) fue determinante para la infección. Se discute el papel de D. marsupialis como potencial reservorio de parásitos tripanosomátidos en la zona evaluada.

We studied the prevalence of infection by trypanosomatid parasites in Didelphis marsupialis and its relationship with morphological/age aspects in a rural area of El Carmen de Bolivar, Colombia. Five visits were made to the Vereda El Alférez; each of which lasted three consecutive nights. During these visits, Tomahawk® traps were installed in the peridomestic and wild ecotopes of the Vereda El Alférez. Body measurements, sex and age were determined from the collected animals. Blood was extracted by cardiopuncture, after sedation, in order to obtain total deoxyribonucleic acid (DNA) and amplify the conserved region of the kinetoplast minicircle DNA (kDNA) of parasitic trypanosomatids. The association between morphological parameters of didelphids and their frequency of infection by parasitic trypanosomatids was determined by binomial regression. Thirty D. marsupialis specimens (60.0% females and 40.0% males/66.7% adults and 33.3% juveniles) were collected. Molecular diagnosis revealed a frequency of trypanosomatid parasite infection of 46.7%. Stage (p=0.024) was a determinant for infection. We discuss the role of D. marsupialis as a potential reservoir of parasitic trypanosomatids in the Vereda El Alférez.

Genetic diversity of Aedes aegypti in the central-upper Paraná Cross-Border axis in Paraguay. / Diversidad genética de aedes aegypti en el eje transfronterizo Central-Alto Paraná en Paraguay.

OBJECTIVE: To determine the genetic diversity of Aedes aegypti in the Central-Alto Paraná cross-border road corridor of Paraguay, an area that has reports of dengue cases. MATERIALS AND METHODS: Twenty adult females were selected from hatching Ae. aegypti eggs from households geolocated in the departments of Alto Paraná, Caaguazú, Cordillera and Central, between 2018 and 2019. DNA was extracted from the tissue of females for amplifying their polymorphic patterns by random amplification of polymorphic DNA by PCR (RAPD-PCR), using primers H3 and B03 in order to identify genetic parameters of population diversity. The relationships between mosquito populations according to locality were observed by unpaired arithmetic mean analysis. We used DIVA-GIS 7.3.0 and MAXENT to analyze the suitable areas of actual and potential geographic distribution of these Ae. aegypti populations. RESULTS: Forty loci were identified by RAPD-PCR profiling, with moderate gene differentiation (Gst = 0.12). The cross-border corridor presented bioclimatic conditions for the presence of variant populations of Ae. aegypti, with precipitation in the warmest quarter and mean temperature in the driest quarter being determinant in the distribution. CONCLUSIONS: There is evidence of moderate genetic diversity in Ae. aegypti populations from areas that have reported dengue cases in the cross-border road corridor linking the Central and Alto Paraná departments of Paraguay. The study of genetic variability of Ae. aegypti is very useful for entomo-epidemiological surveillance and evaluation of possible resistance to chemical control.

OBJETIVOS: Conocer la diversidad genética de Aedes aegypti en el corredor vial transfronterizo Central-Alto Paraná de Paraguay, con registros de casos de dengue. MATERIALES Y MÉTODOS: Se seleccionaron veinte hembras adultas de la eclosión de huevos de Ae. aegypti procedentes de casas geolocalizadas en los departamentos de Alto Paraná, Caaguazú, Cordillera y Central, entre el 2018 y 2019. Se extrajo ADN del tejido de las hembras para amplificación aleatoria de sus patrones polimórficos mediante amplificación aleatoria del ADN polimórfico por PCR (RAPD-PCR), usando cebadores H3 y B03 a fin de conocer parámetros genéticos de diversidad poblacional. Las relaciones entre las poblaciones de mosquitos según la localidad fueron visualizadas mediante análisis no apareado de la media aritmética. Las áreas idóneas de distribución geográfica real y potencial de estas poblaciones de Ae. aegypti fueron analizadas mediante DIVA-GIS 7.3.0 y MAXENT. RESULTADOS: Se identificaron 40 loci mediante perfiles RAPD-PCR, con diferenciación génica moderada (Gst = 0,12). El corredor transfronterizo presentó condiciones bioclimáticas para la presencia de poblaciones variantes de Ae. aegypti, siendo determinantes en la distribución la precipitación del trimestre más cálido y la temperatura media del trimestre más seco. CONCLUSIONES: Se evidencia que existe diversidad genética moderada en las poblaciones de Ae. aegypti procedentes de zonas con registros de casos de dengue ubicadas en el corredor vial transfronterizo que une los departamentos Central y Alto Paraná de Paraguay. El estudio de variabilidad genética de Ae. aegypti es de gran utilidad para la vigilancia entomoepidemiológica y evaluación de posibles eventos de resistencia al control químico.

Molecular diagnosis of trypanosomatids in Didelphis marsupialis from Los Montes de María: a first report of Trypanosoma rangeli from Colombian Caribbean region.

Didelphis marsupialis is a primary reservoir of Trypanosoma cruzi, etiologic agent of American Trypanosomiasis-AT or Chagas Disease-CD, in America. Some findings of Trypanosoma rangeli have been recorded in this mammal, in sympatry with T. cruzi. In Los Montes de María, Bolívar, Colombian Caribbean, triatomine insects and potential parasite host has been registered, but little is known about the relationship between these parasites and D. marsupialis. We investigated the natural trypanosomatids infection rate in D. marsupialis, applying a parasitological and molecular diagnosis. Twenty D. marsupialis was investigated between 2018 and 2019 using 21 Tomahawk® traps placed on the sylvatic/domestic corridors. Blood was drawn by cardiopuncture after sedation. An aliquot of blood samples was cultured in Novy, Nicolle, McNeal/Roswell Park Memorial Institute medium at 24 °C/60 days for the detection of motile trypomastigotes. Parasite DNA was obtained by salting out methods from positive blood cultures. Trypanosomatids diagnosis was done by Polymerase Chain Reaction-sequencing of V7V8 region of 18S ribosomal RNA (18S-rRNA) gene. Amplicons were sequenced, and consensus sequences were aligned with reference sequences from GenBank. Four isolates corresponded to T. rangeli (20%) and one to T. cruzi (5%). The natural infection of D. marsupialis by T. rangeli and T. cruzi constitutes the first record of these parasites in didelphids in Los Montes de María and the first record of T. rangeli in this marsupial, in the Colombian Caribbean.

Diversidad genética de aedes aegypti en el eje transfronterizo Central-Alto Paraná en Paraguay / Genetic diversity of Aedes aegypti in the central-upper Paraná Cross-Border axis in Paraguay

RESUMEN Objetivos: Conocer la diversidad genética de Aedes aegypti en el corredor vial transfronterizo Central-Alto Paraná de Paraguay, con registros de casos de dengue. Materiales y métodos: Se seleccionaron veinte hembras adultas de la eclosión de huevos de Ae. aegypti procedentes de casas geolocalizadas en los departamentos de Alto Paraná, Caaguazú, Cordillera y Central, entre el 2018 y 2019. Se extrajo ADN del tejido de las hembras para amplificación aleatoria de sus patrones polimórficos mediante amplificación aleatoria del ADN polimórfico por PCR (RAPD-PCR), usando cebadores H3 y B03 a fin de conocer parámetros genéticos de diversidad poblacional. Las relaciones entre las poblaciones de mosquitos según la localidad fueron visualizadas mediante análisis no apareado de la media aritmética. Las áreas idóneas de distribución geográfica real y potencial de estas poblaciones de Ae. aegypti fueron analizadas mediante DIVA-GIS 7.3.0 y MAXENT. Resultados: Se identificaron 40 loci mediante perfiles RAPD-PCR, con diferenciación génica moderada (Gst = 0,12). El corredor transfronterizo presentó condiciones bioclimáticas para la presencia de poblaciones variantes de Ae. aegypti, siendo determinantes en la distribución la precipitación del trimestre más cálido y la temperatura media del trimestre más seco. Conclusiones: Se evidencia que existe diversidad genética moderada en las poblaciones de Ae. aegypti procedentes de zonas con registros de casos de dengue ubicadas en el corredor vial transfronterizo que une los departamentos Central y Alto Paraná de Paraguay. El estudio de variabilidad genética de Ae. aegypti es de gran utilidad para la vigilancia entomoepidemiológica y evaluación de posibles eventos de resistencia al control químico.

ABSTRACT Objective: To determine the genetic diversity of Aedes aegypti in the Central-Alto Paraná cross-border road corridor of Paraguay, an area that has reports of dengue cases. Materials and methods: Twenty adult females were selected from hatching Ae. aegypti eggs from households geolocated in the departments of Alto Paraná, Caaguazú, Cordillera and Central, between 2018 and 2019. DNA was extracted from the tissue of females for amplifying their polymorphic patterns by random amplification of polymorphic DNA by PCR (RAPD-PCR), using primers H3 and B03 in order to identify genetic parameters of population diversity. The relationships between mosquito populations according to locality were observed by unpaired arithmetic mean analysis. We used DIVA-GIS 7.3.0 and MAXENT to analyze the suitable areas of actual and potential geographic distribution of these Ae. aegypti populations. Results: Forty loci were identified by RAPD-PCR profiling, with moderate gene differentiation (Gst = 0.12). The cross-border corridor presented bioclimatic conditions for the presence of variant populations of Ae. aegypti, with precipitation in the warmest quarter and mean temperature in the driest quarter being determinant in the distribution. Conclusions: There is evidence of moderate genetic diversity in Ae. aegypti populations from areas that have reported dengue cases in the cross-border road corridor linking the Central and Alto Paraná departments of Paraguay. The study of genetic variability of Ae. aegypti is very useful for entomo-epidemiological surveillance and evaluation of possible resistance to chemical control.

Susceptibilidad a deltametrina de Aedes (Stegomyia) aegypti (Linnaeus 1676) (Diptera: Culicidae) y determinación de áreas de influencia para su presencia en un municipio del Departamento Central, Paraguay / Susceptibility to deltametrin of Aedes (Stegomyia) aegypti(Linnaeus 1676) (Diptera: Culicidae) and determination from areas of influence for its presence in a municipality of the Centraldepartment, Paraguay

RESUMEN Introducción. Paraguay es un país endémico para el dengue y la intervención primaria para esta arbovirosis es el control químico de su vector Aedes aegypti, siendo necesario estudiar su resistencia frente a insecticidas utilizados para el control y conocer su sostenibilidad en espacio y tiempo. Objetivo. Determinar la susceptibilidad de poblaciones silvestres de Ae. aegypti procedentes de Villa Elisa-Central, frente al adulticida deltametrina y conocer el área potencial de distribución de estas poblacionesresistentes/susceptibles. Metodología. Se utilizaron mosquitos Ae. aegypti adultos de la Filial 1 obtenidos de ovitrampas colocadas en viviendas geo-localizadas entre diciembre 2017-marzo 2018. Las hembras resultantes de la cría en condiciones ad hoc en laboratorio fueron expuestas a concentraciones crecientes del insecticida. La distribución real de las poblaciones de mosquitos y su área de influencia, fueron modeladas mediante DIVA-GIS. Resultados. La mortalidad promedio observada para cada dosis de deltametrina fue: 5,3% para (0,03%); 47,4% para (0,15%) y 61,2% para (0,3). Se observó áreas de influencia para la presencia de Ae. aegypti. Conclusión. los valores correspondientes a la mortalidad resultaron inferiores al 98%, lo cual fue indicativo de resistencia a dosis diagnóstica. El modelo de distribución geográfica de la población evaluada demostró la existencia de áreas idóneas con condiciones bioclimáticas óptimas (niveles de precipitación, temperatura y humedad), propicias para la aparición de reservorios de agua y presencia del vector, tanto en Villa Elisa, como en los distritos localizados en la periferia tales como San Lorenzo, Lambaré, Ñemby, San Antonio y Fernando de la Mora.

ABSTRACT Introduction. Paraguay is an endemic country for dengue and the primary intervention for this arbovirus is the chemical control of its vector Aedes aegypti, being necessary to study its resistance against insecticides used for control and to know its sustainability in space and time. Objective. To determine the susceptibility of wild populations of Ae. aegypti from Villa Elisa-Central, against the adulticide deltamethrin and to know the potential area of ​​distribution of these resistant/susceptible populations. Methodology. Mosquitoes Ae. aegypti adults from Filial 1 obtained from ovitraps placed in geo-located dwellings between December 2017 and March 2018. The females resulting from rearing under ad hoc conditions in the laboratory were exposed to increasing concentrations of the insecticide. The actual distribution of mosquito populations and their area of ​​influence were modeled using DIVA-GIS. Results. The average mortality observed for each dose of deltamethrin was: 5.3% for (0.03%); 47.4% for (0.15%) and 61.2% for (0.3). Areas of influence were observed for the presence of Ae. aegypti. Conclusion. The values ​​corresponding to mortality were lower than 98%, which was indicative of resistance to diagnostic dose. The geographic distribution model of the evaluated population demonstrated the existence of suitable areas with optimal bioclimatic conditions (levels of precipitation, temperature and humidity), conducive to the appearance of water reservoirs and the presence of the vector, both in Villa Elisa and in the districts located on the outskirts such as San Lorenzo, Lambaré, Ñemby, San Antonio and Fernando de la Mora.

Evaluación de estuches comerciales para el diagnóstico inmunológico y molecular de la enfermedad de Chagas en zonas endémicas de Venezuela / Evaluation of commercial kits for the immunological and molecular diagnosis of Chagas disease in endemic areas of Venezuela

IntroducciónLa sensibilidad y especificidad de las técnicas de diagnóstico para la enfermedad de Chagas dependen en gran parte de los antígenos y las dianas utilizadas y de la respuesta inmunológica y las características de la infección de la población donde se aplica, de allí la necesidad de la evaluación de las técnicas de diagnóstico disponibles en un área determinada, por lo que el objetivo de este trabajo fue evaluar dos estuches comerciales para el diagnóstico inmunológico y molecular de la enfermedad de Chagas en zonas endémicas de Venezuela.MétodosSe evaluaron los estuches: Chagas ELISA IgG+IgM® y Speed Oligo Chagas® (Vircell®, Granada, España). Se valoraron con 129 muestras (35 de pacientes en fase aguda, 33 en fase crónica, 31 de pacientes con otras enfermedades y 30 de individuos sanos). Se compararon los resultados con los obtenidos en las pruebas convencionales ELISA y PCR-ADN satélite de Trypanosoma cruzi.ResultadosCon Chagas ELISA IgG+IgM® se obtuvo una sensibilidad de 94,1% y especificidad de 93,4%, con Speed Oligo Chagas® se obtuvo una sensibilidad de 92,6% y especificidad de 100%, valores similares a los obtenidos con ELISA y PCR-ADNsat convencionales.ConclusiónLa sensibilidad y especificidad de los estuches comerciales evaluados los hacen adecuados para el diagnóstico de la enfermedad de Chagas en zonas endémicas de Venezuela.

IntroductionThe sensitivity and specificity of diagnostic techniques for Chagas disease depend largely on the antigens and targets used and on the immune response and characteristics of the infection of the population where it is applied, hence the need for evaluation of the diagnostic techniques available in a given area. So, the objective of this work was to evaluate two commercial kits for the immunological and molecular diagnosis of Chagas disease in endemic areas of Venezuela.MethodsThe evaluated kits were: Chagas ELISA IgG+IgM® and Speed Oligo Chagas® (Vircell®, Granada, Spain). They were evaluated with 129 samples (35 from patients in the acute phase, 33 in the chronic phase, 31 from patients with other diseases, and 30 from healthy individuals). The results were compared with those obtained in the conventional ELISA and PCR-satellite DNA tests for Trypanosoma cruzi.ResultsWith Chagas ELISA IgG+IgM® a sensitivity of 94.1% and specificity of 93.4% were obtained, with Speed Oligo Chagas® a sensitivity of 92.6% and specificity of 100% were achieved, values similar to those showed by conventional ELISA and satDNA-PCR.ConclusionThe sensitivity and specificity of the commercial kits evaluated make them suitable for the diagnosis of Chagas disease in endemic areas of Venezuela.

Infections and Coinfections by Trypanosomatid Parasites in a Rural Community of Venezuela.

INTRODUCTION: Trypanosoma cruzi, Trypanosoma rangeli and Leishmania spp. are parasites that coexist in several endemic areas. The identification of these parasites in hosts is important for the control programs. METHODS: 216 samples from human blood (101), blood of other mammals (45) and triatomine intestinal content and hemolymph (70), from an endemic area of Venezuela, were analysed. The samples were evaluated by; serology (only humans) and PCR for T. cruzi in human, other mammals and triatomines, PCR for T. rangeli in mammals-including human and triatomines and PCR for Leishmania in mammals-including human. RESULTS: The 9.9% of the human samples were positive for T. cruzi by serology, 11.9% by PCR, 4% for T. rangeli PCR and none for Leishmania spp. PCR. 60% of the samples of other mammals showed DNA amplification for T. cruzi, 42.2% for T. rangeli and 4.4% for Leishmania spp. 61.4% of the triatomine samples showed DNA amplification for T. cruzi and 10% for T. rangeli. CONCLUSIONS: High T. cruzi infection was detected in mammals and triatomines compared with T. rangeli. Low leishmanial infection was detected in other mammals. It is the first time that T. cruzi/T. rangeli coinfection, in humans, Canis familiaris (dog), and Bos Taurus (cow), were reported world-wide, and that this coinfection was described in Tamandua tetradactyla (anteater) from Venezuela. The coinfection T. cruzi/T. rangeli in mammals-including humans and triatomines, and coinfection T. cruzi/Leishmania spp. in non-human mammals, show the risk for trypanosomic zoonoses in this endemic area.

Evaluation of commercial kits for the immunological and molecular diagnosis of Chagas disease in endemic areas of Venezuela.

INTRODUCTION: The sensitivity and specificity of diagnostic techniques for Chagas disease depend largely on the antigens and targets used and on the immune response and characteristics of the infection of the population where it is applied, hence the need for evaluation of the diagnostic techniques available in a given area. So, the objective of this work was to evaluate two commercial kits for the immunological and molecular diagnosis of Chagas disease in endemic areas of Venezuela. METHODS: The evaluated kits were: Chagas ELISA IgG+IgM® and Speed Oligo Chagas® (Vircell®, Granada, Spain). They were evaluated with 129 samples (35 from patients in the acute phase, 33 in the chronic phase, 31 from patients with other diseases, and 30 from healthy individuals). The results were compared with those obtained in the conventional ELISA and PCR-satellite DNA tests for Trypanosoma cruzi. RESULTS: With Chagas ELISA IgG+IgM® a sensitivity of 94.1% and specificity of 93.4% were obtained, with Speed Oligo Chagas® a sensitivity of 92.6% and specificity of 100% were achieved, values similar to those showed by conventional ELISA and satDNA-PCR. CONCLUSION: The sensitivity and specificity of the commercial kits evaluated make them suitable for the diagnosis of Chagas disease in endemic areas of Venezuela.

Manifestaciones oculares de la toxocariasis en escolares del estado Anzoátegui en Venezuela / Ocular manifestations of toxocariasis in schoolchildren from the state from the Anzoátegui state in Venezuela

RESUMEN El objetivo de este estudio fue determinar manifestaciones oculares de la toxocariasis en escolares. Se realizó un estudio en dos escuelas del estado Anzoátegui en Venezuela en el 2019. Se empleó la prueba de ELISA para determinar los anticuerpos IgG contra Toxocara spp. Las familias completaron un cuestionario y los niños fueron evaluados clínicamente por pediatras y oftalmólogos. Participaron 118 niños, el 18,6% presentó anticuerpos anti-Toxocara spp. Las manifestaciones clínicas con asociación estadísticamente significativa fueron las reacciones alérgicas, epífora y disminución de la agudeza visual. En la evaluación oftalmológica se encontró queratitis, uveítis, iritis, granuloma retiniano, endoftalmitis, amaurosis, leucocoria, desprendimiento de retina y endotropía. Los hallazgos muestran una alta frecuencia de enfermedad ocular en niños con toxocariasis de un estado de Venezuela.

ABSTRACT The objective of this study was to determine ocular manifestations of toxocariasis in schoolchildren. A study was conducted in two schools in the Anzoátegui state in Venezuela in 2019. The ELISA test was used to determine IgG antibodies against Toxocara spp. The families completed a questionnaire, and the children were clinically evaluated by pediatricians and ophthalmologists. 118 children participated, 18.6% presented anti-Toxocara spp. The clinical manifestations with a statistically significant association were allergic reactions, epiphora, and decreased visual acuity. The ophthalmological evaluation found keratitis, uveitis, iritis, retinal granuloma, endophthalmitis, amaurosis, leukocoria, retinal detachment and endotropia. The findings show a high frequency of eye disease in children with toxocariasis from a state of Venezuela.

Ecopathogenic complexes of American trypanosomiasis in endemic areas of Venezuela: Diagnosis and variability of Trypanosoma cruzi.

BACKGROUND & OBJECTIVES: Trypanosoma cruzi, the causative agent of American trypanosomiasis, has been reported in 180 mammalian species and 154 triatomines species of Neotropic. This is a clonal parasite with variable biological behaviour, associated with the genetics of the parasite and its hosts. To know the eco-pathogenic complex of this zoonosis, it was proposed to characterize T. cruzi isolates obtained from triatomines and domestic, peridomestic and wild mammals of the eastern and central-western regions of Venezuela. METHODS: The positivity to T. cruzi was established and the isolates were genetically characterized by PCR amplification of the mini-exon gene, the DNA coding for 24Sa and 18S rRNA, and polymorphic sequences-RFLPs. The sampling sites were georeferenced using the MapSource Software and ArcGis 9.3 programs to generate distribution maps of the isolates. RESULTS: Of the 460 hosts (205 triatomines and 255 mammals), 49% were positive for the parasite. On the other hand, 38 isolates obtained from the triatomines and 23 isolates obtained from mammals were evaluated. The TcI genotype predominated in most of the isolates; however, in those obtained from triatomines the presence of the TcIII genotype in single infections and TcI + TcIII or TcI + TcIV in mixed infections was also evidenced. INTERPRETATION & CONCLUSION: There is a possibility that the triatomines act as biological syringes for these genotypes associated exclusively to them. The heterogeneity in T. cruzi isolates demonstrated the complexity of parasitosis in these regions, presenting its control and prevention as a challenge.

First molecular detection of Trypanosoma cruzi, T. rangeli and Leishmania spp. in capybaras.

Hydrochoerus hydrochaeris (capybara), is a widely distributed rodent in Latin America, with exploitation for food purposes and also used in leather industry products. The infection of this rodent by trypanosomatids may not be detected by parasitological methods, due to low parasitemias. The Capybaras blood samples from the Apure State were collected on filter paper, DNA was extracted and PCR was performed. The PCR technique was used for the detection of Trypanosoma cruzi satellite and kinetoplast DNA, T. rangeli miniexon, T. evansi RIME sequence, and DNA encoding ribosomal RNA and internal transcribed spacer 1 from Leishmania spp. Of the 16 evaluated samples, 12 (75%) were positive for T. cruzi, two for T. rangeli (12.5%), one for Leishmania spp. (6.3%) and none for T. evansi. Regarding coinfection, the two specimens infected with T. rangeli were also infected with T. cruzi (12.5%) and the positive sample for Leishmania spp. was also infected with T. cruzi (6.3%). The results shown in this study represent the first finding of T. cruzi infection, detected by molecular methods, world-wide and the first time that T. rangeli and Leishmania spp. have been found in capybaras. In addition, we report coinfections by T. cruzi/T. rangeli and T. cruzi/Leishmania spp. in H. hydrochaeris for the first time world-wide. Capybaras are widely managed as a source of animal protein, the results obtained require evaluating their possible role as a reservoir in trypanosomiasis and leishmaniasis. A 'One Health' approach through combination of ecological, veterinary and human health including the diagnosis and treatment of diseases of both humans and animals is essential for the development of more successful health programs.

[Ocular manifestations of toxocariasis in schoolchildren from the state from the Anzoátegui state in Venezuela]. / Manifestaciones oculares de la toxocariasis en escolares del estado Anzoátegui en Venezuela.

The objective of this study was to determine ocular manifestations of toxocariasis in schoolchildren. A study was conducted in two schools in the Anzoátegui state in Venezuela in 2019. The ELISA test was used to determine IgG antibodies against Toxocara spp. The families completed a questionnaire, and the children were clinically evaluated by pediatricians and ophthalmologists. 118 children participated, 18.6% presented anti-Toxocara spp. The clinical manifestations with a statistically significant association were allergic reactions, epiphora, and decreased visual acuity. The ophthalmological evaluation found keratitis, uveitis, iritis, retinal granuloma, endophthalmitis, amaurosis, leukocoria, retinal detachment and endotropia. The findings show a high frequency of eye disease in children with toxocariasis from a state of Venezuela.

El objetivo de este estudio fue determinar manifestaciones oculares de la toxocariasis en escolares. Se realizó un estudio en dos escuelas del estado Anzoátegui en Venezuela en el 2019. Se empleó la prueba de ELISA para determinar los anticuerpos IgG contra Toxocara spp. Las familias completaron un cuestionario y los niños fueron evaluados clínicamente por pediatras y oftalmólogos. Participaron 118 niños, el 18,6% presentó anticuerpos anti-Toxocara spp. Las manifestaciones clínicas con asociación estadísticamente significativa fueron las reacciones alérgicas, epífora y disminución de la agudeza visual. En la evaluación oftalmológica se encontró queratitis, uveítis, iritis, granuloma retiniano, endoftalmitis, amaurosis, leucocoria, desprendimiento de retina y endotropía. Los hallazgos muestran una alta frecuencia de enfermedad ocular en niños con toxocariasis de un estado de Venezuela.

Utility of a Fluid Library with Samples of Humans, Reservoirs and Vectors Collected in Filter Paper, for Retrospective Diagnosis of American Trypanosomiasis in Endemic Areas of Venezuela.

INTRODUCTION: We define a fluid library as a library of samples of different biological fluids (from humans, animals or vectors) collected and properly stored on filter paper, which allows retrospective studies, especially of diagnosis or detection of infectious agents in these samples, using different techniques. The objective of this work was the retrospective diagnosis of American trypanosomiasis by PCR in a Venezuelan endemic area using a fluid library. METHODS: A fluid library with samples that had been collected on filter paper, 5 years ago, was used for the detection of Trypanosoma cruzi DNA. 165 blood samples of humans, 30 samples of 25 animals (Didelphis marsupialis, Canis familiaris, Equus asinus and Felis catus) and 8 samples of vectors from endemic areas of Anzoátegui state, were analysed by PCR. RESULTS: The results revealed that 16.4% of the humans samples were positive, 11.1% of those detected positive were children younger than 10 years old, and 26.72% young people aged 11-20 years, suggesting that T. cruzi infection has been active for the past two decades. 56% of the animal samples showed amplification; Didelphis marsupialis 66%, Canis familiaris 54.5%, Equus asinus 50%, and Felis catus 33.3%. On the other hand, positivity (50%) was detected in the studied vectors, of which the 3 most important species in Venezuela (Rhodnius prolixus, Triatoma maculata and Panstrongylus geniculatus) were involved. CONCLUSIONS: The PCR using a fluid library allowed the detection of T. cruzi DNA in old samples from the three host of the epidemiological chain, suggesting that retrospective diagnosis can be made through this strategy and demonstrate that there has been active transmission, which helps to clarify the epidemiological situation in areas where there are no previous reports.

Detection of Trypanosoma cruzi DNA in false negative samples of collected triatomines, xenodiagnosis material, and biopsies of experimentally infected animals.

Direct test over the gut material from triatomine vectors and xenodiagnosis over mammalian hosts are classical techniques for Trypanosoma cruzi parasitological diagnosis. Nevertheless, negative results can be a source of uncertainty. Experimental models have allowed evaluating the tissue invasion of different strains of T. cruzi, but conventional techniques for tissue biopsies involve time-consuming and elaborated procedures and have low sensitivity. Gut material of collected triatomines (microscopically negative) (n = 114), material of mammal xenodiagnoses (microscopically negative) (n = 138), and biopsy material (microscopically negative) from experimentally infected animals (n = 34) with isolates from endemic areas of Chagas' disease from Venezuela were used for DNA extraction and PCR for the amplification of kinetoplast DNA (kDNA) and satellite DNA (sDNA) of T. cruzi. Positive PCR was observed in 53.6% of collected triatomine material, 15.8% of parasitological negative xenodiagnosis material, and 70.6% in biopsies, revealing underestimation by the parasitological tests and the valour of this analysis with preserved material. Anzoátegui was the state with the highest percentage of infection, and the triatomine species Rhodnius prolixus and Panstrongylus geniculatus had the highest percentages of infection. Didelphis marsupialis and Canis familiaris were the most infected by T. cruzi revealed by PCR of xenodiagnosis material. In addition, the PCR technique allowed demonstrating the invasion of T. cruzi in all tissues analyzed, constituting a molecular marker of tissue invasion.

Infección natural por Trypanosoma cruzi en triatominos que habitan en la palma corozo ( Acrocomia aculeta ) en regiones del oriente de Venezuela / Triatominae in Palm Trees and Their Natural Infection by Trypanosoma cruzi in Regions of Eastern Venezuela

RESUMEN Objetivos Conocer la infestación natural por triatominos y su infección por Trypanosoma cruzi (T. cruzi) en Acrocomia aculeata (A. aculeata) o palma corozo en el estado Anzoátegui, Venezuela. Materiales y métodos Se estudió la infestación triatomínica y su infección por T. cruzi en A. aculeata desafectadas en campañas fitosanitarias. La presencia del parásito se determinó por microscopia y PCR-kDNA, y se realizó su caracterización mediante marcadores moleculares. Resultados Se encontraron 14 palmeras con infestación triatomínica, el 48,8 % de los ejemplares correspondieron a Rhodnius prolixus y el 48,2 % a Triatoma maculata, con desarrollo ontogénico hacia el adulto. Las pruebas parasitológicas y moleculares, su morfología típica y la infección en el modelo murino revelaron la presencia de T. cruzi en 54,8 % en promedio, para ambas especies de triatominos, con circulación del genotipo TcI de T. cruzi. Conclusiónes Se reportó para el estado Anzoátegui en Venezuela, la infestación de palma corozo con Rhodnius prolixus y Triatoma maculata y la presencia de subpoblaciones TcI de T. cruzi, siendo esta palma el hábitat peridomiciliar del binomio triatominos-T. cruzi y posible bioindicador de riesgo de infección para poblaciones humanas circunvecinas.

ABSTRACT Introduction To know the natural infestation by triatominae and their infection by Trypanosoma cruzi (T. cruzi) in Acrocomia Aculeata (A. aculeata) or coyol palm in the state of Anzoátegui, Venezuela. Materials and Methods Triatominic infestation and its infection by T. cruzi was studied in non-affected A. aculeata in phytosanitary campaigns. The presence of the parasite was determined by microscopy and PCR-kDNA, and its characterization was made by means of molecular markers. Results Fourteen palm trees with triatominic infestation were found; 48.8% of the individuals corresponded to Rhodnius prolixus and 48.2% to Maculata Triatoma, with ontogenetic development towards adult. The parasitology and molecular tests, their typical morphology and the infection in the murine model revealed the presence of T. cruzi in an average of 54,8%, for both species of triatominae, with circulation of the TcI genotype of T. cruzi. Conclusions The infestation of coyol palm trees with Rhodnius prolixus and Maculata Triatoma was reported for the state of Anzoátegui in Venezuela, as well as the presence of TcI sub-populations of T. cruzi, being this palm tree the peridomicilar habitat of the triatominae-T. cruzi binomial and possible bioindicador of risk of infection for surrounding human populations.

[Triatominae in Palm Trees and Their Natural Infection by Trypanosoma cruzi in Regions of Eastern Venezuela]. / Infección natural por Trypanosoma cruzi en triatominos que habitan en la palma corozo ( Acrocomia aculeta ) en regiones del oriente de Venezuela.

INTRODUCTION: To know the natural infestation by triatominae and their infection by Trypanosoma cruzi (T. cruzi) in Acrocomia Aculeata (A. aculeata) or coyol palm in the state of Anzoátegui, Venezuela. MATERIALS AND METHODS: Triatominic infestation and its infection by T. cruzi was studied in non-affected A. aculeata in phytosanitary campaigns. The presence of the parasite was determined by microscopy and PCR-kDNA, and its characterization was made by means of molecular markers. RESULTS: Fourteen palm trees with triatominic infestation were found; 48.8% of the individuals corresponded to Rhodnius prolixus and 48.2% to Maculata Triatoma, with ontogenetic development towards adult. The parasitology and molecular tests, their typical morphology and the infection in the murine model revealed the presence of T. cruzi in an average of 54,8%, for both species of triatominae, with circulation of the TcI genotype of T. cruzi. CONCLUSIONS: The infestation of coyol palm trees with Rhodnius prolixus and Maculata Triatoma was reported for the state of Anzoátegui in Venezuela, as well as the presence of TcI sub-populations of T. cruzi, being this palm tree the peridomicilar habitat of the triatominae-T. cruzi binomial and possible bioindicador of risk of infection for surrounding human populations.

OBJETIVOS: Conocer la infestación natural por triatominos y su infección por Trypanosoma cruzi (T. cruzi) en Acrocomia aculeata (A. aculeata) o palma corozo en el estado Anzoátegui, Venezuela. MATERIALES Y MÉTODOS: Se estudió la infestación triatomínica y su infección por T. cruzi en A. aculeata desafectadas en campañas fitosanitarias. La presencia del parásito se determinó por microscopia y PCR-kDNA, y se realizó su caracterización mediante marcadores moleculares. RESULTADOS: Se encontraron 14 palmeras con infestación triatomínica, el 48,8 % de los ejemplares correspondieron a Rhodnius prolixus y el 48,2 % a Triatoma maculata, con desarrollo ontogénico hacia el adulto. Las pruebas parasitológicas y moleculares, su morfología típica y la infección en el modelo murino revelaron la presencia de T. cruzi en 54,8 % en promedio, para ambas especies de triatominos, con circulación del genotipo TcI de T. cruzi. CONCLUSIÓNES: Se reportó para el estado Anzoátegui en Venezuela, la infestación de palma corozo con Rhodnius prolixus y Triatoma maculata y la presencia de subpoblaciones TcI de T. cruzi, siendo esta palma el hábitat peridomiciliar del binomio triatominos-T. cruzi y posible bioindicador de riesgo de infección para poblaciones humanas circunvecinas.