[Circulación de Leishmania infantum y Trypanosoma cruzi en perros domésticos de áreas urbanas de Sincelejo, región Caribe de Colombia].

Introducción. La enfermedad de Chagas y la leishmaniasis tradicionalmente se han considerado zoonosis endémicas de áreas rurales del país. Sin embargo, la aparición de casos de estas enfermedades en áreas urbanas sugiere nuevos ciclos de circulación de estos parásitos. Por esta razón, se ha propuesto a los perros como centinelas de estos agentes zoonóticos, dado su rol como huéspedes accidentales o reservorios. Objetivo. Evaluar la circulación silenciosa de Leishmania spp. y Trypanosoma cruzi en perros de zonas urbanas de la ciudad de Sincelejo, Sucre. Materiales y métodos. Se analizaron 100 muestras de sangre de perros para amplificar la región ITS1 de Leishmania spp. Las muestras positivas se utilizaron para amplificar la región conservada del minicírculo del ADN del cinetoplasto de Leishmania infantum y para el análisis de polimorfismos de longitud de fragmentos de restricción con la endonucleasa HaeIII. Por otra parte, se amplificó un fragmento del ADN satelital de T. cruzi. Además, se evaluó la presencia de infecciones por Ehrlichia canis y Anaplasma platys, como potencialmente modificadoras de las manifestaciones clínicas. Resultados. De los 100 perros estudiados, se detectó: Leishmania spp. en 32, T. cruzi en 12, ambos parásitos en 7 y L. infantum en 18. Se encontraron infecciones por anaplasmatáceos en 18, y coinfecciones por bacterias y parásitos en 8 de los perros. En general, 47 de los animales estaban infectados por, al menos, un agente etiológico. Conclusión. Se demuestra la circulación de L. infantum y T. cruzi en zonas urbanas de Sincelejo, así como coinfecciones de estos parásitos junto con parásitos de la familia Anaplasmataceae. El presente estudio demuestra la conveniencia del uso de perros en la vigilancia epidemiológica de estos agentes zoonóticos.

Circulación de Leishmania infantum y Trypanosoma cruzi en perros domésticos de áreas urbanas de Sincelejo, región Caribe de Colombia / Silent circulation of Leishmania infantum and Trypanosoma cruzi among urban dogs from Sincelejo city, Caribbean region of Colombia

Introducción. La enfermedad de Chagas y la leishmaniasis tradicionalmente se han considerado zoonosis endémicas de áreas rurales del país. Sin embargo, la aparición de casos de estas enfermedades en áreas urbanas sugiere nuevos ciclos de circulación de estos parásitos. Por esta razón, se ha propuesto a los perros como centinelas de estos agentes zoonóticos, dado su rol como huéspedes accidentales o reservorios. Objetivo. Evaluar la circulación silenciosa de Leishmania spp. y Trypanosoma cruzi en perros de zonas urbanas de la ciudad de Sincelejo, Sucre. Materiales y métodos. Se analizaron 100 muestras de sangre de perros para amplificar la región ITS1 de Leishmania spp. Las muestras positivas se utilizaron para amplificar la región conservada del minicírculo del ADN del cinetoplasto de Leishmania infantum y para el análisis de polimorfismos de longitud de fragmentos de restricción con la endonucleasa HaelII. Por otra parte, se amplificó un fragmento del ADN satelital de T. cruzi. Además, se evaluó la presencia de infecciones por Ehrlichia canis y Anaplasma platys, como potencialmente modificadoras de las manifestaciones clínicas. Resultados. De los 100 perros estudiados, se detectó: Leishmania spp. en 32, T. cruzi en 12, ambos parásitos en 7 y L. infantum en 18. Se encontraron infecciones por anaplasmatáceos en 18, y coinfecciones por bacterias y parásitos en 8 de los perros. En general, 47 de los animales estaban infectados por, al menos, un agente etiológico. Conclusión. Se demuestra la circulación de L. infantum y T. cruzi en zonas urbanas de Sincelejo, así como coinfecciones de estos parásitos junto con parásitos de la familia Anaplasmataceae. El presente estudio demuestra la conveniencia del uso de perros en la vigilancia epidemiológica de estos agentes zoonóticos.

Introduction: Leishmania infantum and Trypanosoma cruzi are considered endemic zoonotic agents in rural areas of the country; however, there is a high risk of urbanization due to anthropogenic processes. For this reason, dogs have been proposed as sentinels of these zoonoses given their role as patients, hosts and/or reservoirs. Objective: To assess the silent circulation of Leishmania spp. and T. cruzi parasites in canines from urban areas of Sincelejo, Sucre. Materials and methods: One hundred canine blood samples were used to amplify the ITS1 region of Leishmania spp. Positive samples were used to amplify the conserved region of the kinetoplast DNA minicircle of L. infantum and for restriction fragment length polymorphism analysis with HaelII endonuclease. In addition, a satellite DNA of T. cruzi was amplified. Also, the presence of Ehrlichia canis and Anaplasma platys was evaluated as infections that can influence clinical symptoms and health of animals. Results: Leishmania spp. was detected in 32% (32/100) and T. cruzi in 12% (12/100) of the animals, and 7% (7/100) of the samples were positive for both parasites. Also, L. infantum and infections with Anaplasmataceae family parasites were both detected in 18 % (18/100) of the samples. In the same way, co-infections with bacteria and parasites were found in 8 % (8/100) of the animals. Overall, 47 % (47/100) of the animals were infected with at least one agent. Conclusion: The circulation of L. infantum and T. cruzi, as well as co-infections of pathogens of the Anaplasmataceae family, is demonstrated in urban areas of Sincelejo. The present study demonstrates the convenience of canines as epidemiological surveillance sentinels of these zoonotic agents.

Pintomyia evansi (Diptera: Psychodidae) larvae susceptibility to hydrated lime under laboratory conditions.

This study evaluated the effect of Calcium hydroxide (Ca(OH)2 Mg(OH)2) on third stages Pi. evansi larvae mortality under experimental laboratory conditions. Three treatments containing a mixture of phlebotomine natural breeding soil (substrate) and Calcium hydroxide at different concentrations were used: Treatment 1 (T1), 1 kg of substrate mixed with 56.2 g of lime; Treatment 2 (T2), 1 kg of substrate mixed with 62.5 g of lime; and Treatment 3 (T3), 1 kg of substrate mixed with 70 g of lime. in addition, a sample of substrate without lime was used as a control for each treatment. The mortality in T1 was 1% at 24 h and 12% at 48 h, reaching a maximum of 56% at 72 h of exposure. For T2, mortality was progressive, starting with 12% at 12 h, 36% at 24 h, 52% at 48 h, and 100% at 72 h; while T3 showed mortality percentages of 94% and 100% between 12 and 24 h of exposure. Therefore, T3 was the most effective to according to the Kaplan-Meier survival analysis. This study showed that treatments over 62 g of Calcium hydroxide per 1 kg of substrate offer a starting point for immature stage control under laboratory conditions. With these results, we propose to evaluate the cost-effectiveness and feasibility of the application, of the latter concentration, under field conditions in urban environments for its application in vector control programs.

A NEW RECORD OF Leishmania (Viannia) guyanensis (Trypanosomatidae) FROM THE PACIFIC COAST OF COLOMBIA / Un nuevo registro de Leishmania (Viannia) guyanensis (Trypanosomatidae) de la costa pacífica de Colombia

ABSTRACT The aim of this study was the identification of Leishmania species that causes cutaneous leishmaniasis in a patient from Buenaventura, Valle del Cauca, on the Pacific coast of Colombia. Clinical samples were obtained from a 29 years-old male who presented a distinct ulcer with raised borders on his neck. Samples were taken for direct microscopic examination, parasite culture, and molecular identification of the infecting Leishmania species by sequencing of the cytochrome b gene. Direct examination was positive for amastigotes of Leishmania but the culture was negative. The infecting parasite species was identified as L. (V.) guyanensis by means of the nucleotide sequence of a 509 bp fragment of the cytochrome b gene. We report the presence of L. (V.) guyanensis in rural areas of Buenaventura in Valle del Cauca, and the expansion of the geographical distribution of this species in the Pacific region of Colombia.

RESUMEN El objetivo de este estudio fue identificar la especie de Leishmania causante de la leishmaniasis cutánea en un paciente de Buenaventura, Valle del Cauca, en la costa Pacífica de Colombia. Se obtuvieron muestras clínicas de un varón de 29 años de edad que presentó una úlcera distintiva con bordes levantados en el cuello. Se tomaron muestras para examen microscópico directo, cultivo de parásitos e identificación molecular de la especie infectante de Leishmania mediante secuenciación del gen del citocromo b. El examen directo fue positivo para amastigotes de Leishmania pero el cultivo fue negativo. La especie parasitaria infectante se identificó como L. (V.) guyanensis por medio de la secuencia de nucleótidos de un fragmento de 509 pb del gen citocromo b. Con este reporte notificamos la presencia de L. (V.) guyanensis en zona rural del municipio de Buenaventura en el Valle del Cauca y la expansión de la distribución geográfica de esta especie en la región Pacífica de Colombia.

Development of Lutzomyia evansi immature stages in peridomiciliary environment in a leishmaniasis urban focus in the Colombian Caribbean.

In the Caribbean region of Colombia, Lutzomyia evansi is recognized as the vector for Leishmania infantum and Leishmania braziliensis. Identifying breeding sites and surveying abundance of immature phlebotomine sand flies in urban foci of leishmaniasis are useful tool to design new vector control strategies. The objective of this study was to describe the natural breeding sites of Lu. evansi in peridomiciliary vegetation in a peri-urban area of the Colombian Caribbean region. Between 2013 and 2015, 466 microhabitats were sampled, collecting 621 kg of soil samples. The explored microhabitats were bases and tree holes, fallen trees, animal caves, leaf litter, domestic animal shelters, and the inside of dwellings. The immature phlebotomines were recovered by direct search under the stereoscope and incubation of soil samples. In total, 103 microhabitats, associated with 17 arboreal species, were identified as natural breeding sites. Of 422 immature sandflies detected, 98.6% were found in soils at the base of the trees. Eight species of the genus Lutzomyia were identified, of which Lu. evansi (52.6%) was the most abundant, followed by Lu. rangeliana, Lu. cayennensis cayennensis, Lu. atroclavata, Lu. micropyga, Lu. trinidadensis, Lu. dubitans and Lu. gomezi. The arboreal species Cordia alba was the most used by phlebotomines for the development of their immature stages. From 63 natural breeding sites identified 268 immatures were recovered including 176 Lu. evansi. The accumulated precipitation showed correlation (R2 = 0.643, p = 0.013) with the abundance of developmental stages, which increased in September and October. The natural breeding sites of Lu. evansi exhibited a local pattern of occurrence dependent on rainfall. The physicochemical analysis of the soil samples showed that the natural sites for C. alba were categorized as fertile loam soils. This is the first systematic study that estimates the temporal variation of immature sand flies in peridomiciliary vegetation in a peri-urban focus of leishmaniasis in Colombia.

DNA barcoding and fauna of phlebotomine sand flies (Diptera: Psychodidae: Phlebotominae) from Los Tuxtlas, Veracruz, Mexico.

Mexico has great diversity of phlebotomine sand flies related to cases of leishmaniasis, yet few studies have dressed the molecular taxonomy of these sand fly species. The use of the cytochrome oxidase subunit 1 (COI) gene, as a DNA Barcode has facilitated the molecular identification of sand flies species worldwide. We use the DNA barcode as a useful tool for the identification of phlebotomine sand flies of the natural reserve Los Tuxtlas from Veracruz, México. A fragment of 536 bp of the COI gene was obtained from 36 individuals belonging to eight species of five genera (Dampfomyia, Lutzomyia, Psathyromyia, Psychodopygus and Brumptomyia) with coverage between 92-100%, and found similarities ranging from 93-98% with other New World phlebotomine sand flies. The NJ dendogram grouped sand flies into eight clusters according to identified species, supported by bootstrap of 97%-100%. In conclusion, all phlebotomine sand flies were correctly identified and agree with the morphological identification, also could separate genetics the isomorphic females of the genus Brumptomyia.

First record of the sand fly Warileya (Hertigia) hertigi from Antioquia, Colombia

Introduction: The genus Warileya is one of the least-known taxa of sandflies, comprising only nine species, i.e., W. (Warileya) phlebotomanica, W. (Hertigia) hertigi, W. (W.) rotundipennis, W. (W.) nigrosacculus, W. (W.) yungasi, W. (W.) fourgassiensis, W. (W.) lumbrerasi, W. (W.) euniceae and W. (W.) leponti. Objective: To document the presence of a species of the genus Warileya in Antioquia, Colombia. Materials and methods: Sandflies were collected in a cavern of the Cañón del Río Claro Natural Reserve, of the municipality of San Francisco, Antioquia department, Colombia. Phlebotomine sampling was carried out using a CDC light trap during three consecutive nights in May of 2008. Taxonomical determination was based on a revision of the type material of the species and through the use of standard keys for American sandflies. Results: Five male and two female sandflies were taxonomically identified as W. (H.) hertigi. In both sexes, the absence of setal scars in the anepisternum, proepimeron and clypeus; the presence of two transverse rows of setal scars in the tergites; and the short length of the vena gamma were notable. Conclusion: The finding of W. (H.) hertigi increases the number of sandfly species found in Antioquia department to 64. In total, 164 sandfly species have been recorded in Colombia.

Genetic variability of Aedes aegypti in the department of Sucre, Colombia, by analysis of the nucleotide sequence of the mitochondrial ND4 gene

Introduction. Aedes aegypti is the most important mosquito species in America for the transmission of viruses of dengue, Zika, Chikungunya and yellow fever. Ecological factors as well as chemical controls can affect the genetic composition of Ae. aegypti populations, which is why its genetic characterization is necessary. Objective. To determine the genetic variability of Ae. aegypti populations in four municipalities of Sucre department, Colombia. Materials and methods. Larvae of Ae. aegypti, collected in the municipalities of Sincelejo, Sampués, Corozal and Guaranda, Sucre department, were reared under laboratory conditions to adult stage. A segment of the mitochondrial ND4 gene which codes for the subunit 4 of the enzyme NADH-dehydrogenase was used as genetic marker. The genetic analysis included the estimation of parameters of nucleotide and haplotype diversity, genetic structure and gene flow. Results. One hundred and eight partial sequences of 357 nucleotides and four nucleotide haplotypes of the ND4 gene of Ae. aegypti were obtained. A significantly high genetic differentiation was found between the Sampués and Guaranda populations (FST=0.59467), Sincelejo and Sampués (FST=0.25637), and Corozal and Guaranda (FST=0.22237). A high gene flow (Nm=infinite) was observed among the populations of Sincelejo and Corozal. Conclusion. There are genetic differences between the Ae. aegypti populations from the municipalities of Sucre department. The presence of a new haplotype of the mitochondrial ND4 gene of Ae. aegypti in Colombia was recorded, detected in the municipality of Sincelejo.

Primer registro del flebotomíneo Warileya (Hertigia) hertigi en Antioquia, Colombia / First record of the sand fly Warileya (Hertigia) hertigi from Antioquia, Colombia

Resumen Introducción. El género Warileyaes uno de los taxones menos conocidos de los flebotomíneos, el cual está integrado por solo nueve especies: Warileya (Warileya) phlebotomanica, W. (Hertigia) hertigi, W. (W.) rotundipennis, W. (W.) nigrosacculus, W. (W.) yungasi, W. (W.) fourgassiensis, W. (W.) lumbrerasi, W. (W.) euniceae y W. (W.) leponti. Objetivo. Registrar el hallazgo de una especie del género Warileya en Antioquia, Colombia. Materiales y métodos. Los flebotomíneos se recolectaron en una caverna de la Reserva Natural Cañón del Río Claro del municipio de San Francisco, departamento de Antioquia, Colombia. El muestreo entomológico sehizo con una trampa de luz de tipo CDC, durante tres noches consecutivas, en mayo de 2008. La determinación taxonómica se basó en la revisión del material tipo de la especie y de las claves estándar para flebotomíneos americanos. Resultados. Se identificaron taxonómicamente cinco machos y dos hembras de flebotomíneos comoW. (H.) hertigi. En ambos sexos fue notoria la ausencia de cicatrices de setas en el anepisternón, el proepimerón y el clípeo, la presencia de dos hileras transversales de cicatrices de setas en los tergitos, y la longitud corta de la vena gamma. Conclusión. El hallazgo de W. (H.) hertigi incrementa a 64 el número de especies de flebotomíneos encontradas en el departamento de Antioquia. En total, 164 especies de flebotomíneos han sido registradas en Colombia.

Abstract Introduction: The genus Warileyais one of the least-known taxa of sandflies, comprising only nine species, i.e., W. (Warileya) phlebotomanica, W. (Hertigia) hertigi, W. (W.) rotundipennis, W. (W.) nigrosacculus, W. (W.) yungasi, W. (W.) fourgassiensis, W. (W.) lumbrerasi, W. (W.) euniceae and W. (W.) leponti. Objective: To document the presence of a species of the genus Warileya in Antioquia, Colombia. Materials and methods: Sandflies were collected in a cavern of the Cañóndel Río Claro Natural Reserve, of the municipality of San Francisco, Antioquia department, Colombia. Phlebotomine sampling was carried out using a CDC light trap during three consecutive nights in May of 2008. Taxonomical determination was based on a revision of the type material of the species and through the use of standard keys for American sandflies. Results: Five male and two female sandflies were taxonomically identified as W. (H.) hertigi. In both sexes, the absence of setal scars in the anepisternum, proepimeron and clypeus; the presence of two transverse rows of setal scars in the tergites; and the short length of the vena gamma were notable. Conclusion: The finding of W. (H.) hertigi increases the number of sandfly species found in Antioquia department to 64. In total, 164 sandfly species have been recorded in Colombia.

Variabilidad genética de Aedes aegypti en el departamento de Sucre, Colombia, mediante el análisis de la secuencia de nucleótidos del gen mitocondrial ND4 / Genetic variability of Aedes aegypti in the department of Sucre, Colombia, by analysis of the nucleotide sequence of the mitochondrial ND4 gene

Resumen Introducción. Aedes aegypti es la especie de mosquito de mayor relevancia en América por transmitir los virus del dengue, del Zika, del chikungunya y de la fiebre amarilla. Tanto factores ecológicos como el control químico, pueden influir en la composición genética de las poblaciones de Ae. aegypti, por lo cual es necesaria su caracterización. Objetivo. Determinar la variabilidad genética de las poblaciones de Ae. aegypti en cuatro municipios del departamento de Sucre, Colombia. Materiales y métodos. Larvas de Ae. aegypti, recolectadas en los municipios de Sincelejo, Sampués, Corozal y Guaranda del departamento de Sucre, fueron criadas en laboratorio hasta el estado adulto. Como marcador genético, se utilizó un segmento del gen mitocondrial ND4, que codifica para la subunidad 4 de la enzima NADH-deshidrogenasa. El análisis genético incluyó la estimación de parámetros de diversidad de nucleótidos, haplotipos, de estructura genética y de flujo de genes. Resultados. Se obtuvieron 108 secuencias parciales de 357 nucleótidos y cuatro haplotipos de nucleótidos del gen ND4 de Ae. aegypti. Se encontró una diferenciación genética significativamente alta entre las poblaciones de Sampués y Guaranda mediante el índice de fijación (F ST =0,59467), las de Sincelejo y Sampués (F ST = 0,25637), y las de Corozal y Guaranda (F ST = 0,22237). Se evidenció un gran flujo de genes (Nm=infinito) entre las poblaciones de Sincelejo y Corozal. Conclusión. Existen diferencias genéticas entre las poblaciones del mosquito Ae. aegypti de los municipios del departamento de Sucre. Se registra la presencia de un nuevo haplotipo del gen mitocondrial ND4 de Ae. aegypti en Colombia, el cual fue detectado en el municipio de Sincelejo.

Abstract Introduction. Aedes aegypti is the most important mosquito species in America for the transmission of viruses of dengue, Zika, Chikungunya and yellow fever. Ecological factors as well as chemical controls can affect the genetic composition of Ae. aegypti populations, which is why its genetic characterization is necessary. Objective. To determine the genetic variability of Ae. aegypti populations in four municipalities of Sucre department, Colombia. Materials and methods. Larvae of Ae. aegypti, collected in the municipalities of Sincelejo, Sampués, Corozal and Guaranda, Sucre department, were reared under laboratory conditions to adult stage. A segment of the mitochondrial ND4 gene which codes for the subunit 4 of the enzyme NADH-dehydrogenase was used as genetic marker. The genetic analysis included the estimation of parameters of nucleotide and haplotype diversity, genetic structure and gene flow. Results. One hundred and eight partial sequences of 357 nucleotides and four nucleotide haplotypes of the ND4 gene of Ae. aegypti were obtained. A significantly high genetic differentiation was found between the Sampués and Guaranda populations (F ST =0.59467),Sincelejo and Sampués (F ST =0.25637), and Corozal and Guaranda (F ST =0.22237). A high gene flow (Nm=infinite) was observed among the populations of Sincelejo and Corozal. Conclusion. There are genetic differences between the Ae. aegypti populations from the municipalities of Sucre department. The presence of a new haplotype of the mitochondrial ND4 gene of Ae. aegypti in Colombia was recorded, detected in the municipality of Sincelejo.

[Estimation of time detection limit for human cytochrome b in females of Lutzomyia evansi].

INTRODUCTION: Molecular biology techniques have allowed a better knowledge of sources of blood meals in vector insects. However, the usefulness of these techniques depends on both the quantity of ingested blood and the digestion process in the insect. OBJECTIVE: To identify the time limit for detection of the human cytochrome b (Cyt b) gene in experimentally fed females of Lutzomyia evansi. MATERIALS AND METHODS: Eight groups of L. evansi females were fed on human blood and sacrificed at intervals of 24 hours post-ingestion. Total DNA was extracted from each female and a segment of 358 bp of Cyt b was amplified. In order to eliminate false positives, amplification products were subjected to a restriction fragment length polymorphism (RFLP) analysis. RESULTS: The human Cyt b gene segment was detected in 86% (49/57) of the females of L. evansi, from 0 to 168 hours after blood ingestion. In 7% (4/57) of the individuals we amplified insect DNA, while in the remaining 7%, the band of interest was not amplified. We did not find any statistical differences between groups of females sacrificed at different times post-blood meal regarding the amplification of the human Cyt b gene segment or the number of samples amplified. CONCLUSION: The human Cyt b gene segment was detectable in L. evansi females up to 168 hours after blood ingestion.

Estimación del tiempo límite de detección del gen citocromo b de humanos en hembras de Lutzomyia evansi / Estimation of time detection limit for human cytochrome b in females of Lutzomyia evansi

Resumen Introducción. Las técnicas de biología molecular han permitido ampliar el conocimiento sobre las fuentes de ingestión de sangre de los insectos vectores. Sin embargo, la utilidad de estas técnicas depende de la cantidad de sangre ingerida y del proceso de digestión en el insecto. Objetivo. Determinar el tiempo límite de detección del gen citocromo b (Cyt b) de humanos en hembras de Lutzomyia evansi alimentadas experimentalmente. Materiales y métodos. Se evaluaron ocho grupos de hembras de L. evansi alimentadas con sangre humana, las cuales fueron sacrificadas en intervalos de 24 horas desde el momento de la ingestión sanguínea. Se extrajo el ADN total de cada hembra y se amplificó un segmento de 358 pb del gen Cyt b. Los productos amplificados fueron sometidos a un análisis de polimorfismos en la longitud de los fragmentos de restricción (Restriction Fragment Length Polymorphism, RFLP), con el fin de descartar falsos positivos. Resultados. El segmento del gen Cyt b de humanos fue detectado en 86 % (49/57) de las hembras de L. evansi a partir de las 0 horas y hasta 168 horas después de la ingestión de sangre. En 7 % (4/57) de los individuos se amplificó el ADN del insecto y en el 7 % restante no se amplificó la banda de interés. No se encontraron diferencias estadísticas en cuanto a la amplificación del segmento del gen Cyt b de humanos ni al número de muestras amplificadas entre los grupos de hembras sacrificadas a distintas horas después de la ingestión. Conclusión. El segmento del gen Cyt b de humanos fue detectable en hembras de L. evansi hasta 168 horas después de la ingestión de sangre.

Abstract Introduction: Molecular biology techniques have allowed a better knowledge of sources of blood meals in vector insects. However, the usefulness of these techniques depends on both the quantity of ingested blood and the digestion process in the insect. Objective: To identify the time limit for detection of the human cytochrome b (Cyt b) gene in experimentally fed females of Lutzomyia evansi. Materials and methods: Eight groups of L. evansi females were fed on human blood and sacrificed at intervals of 24 hours post-ingestion. Total DNA was extracted from each female and a segment of 358 bp of Cyt b was amplified. In order to eliminate false positives, amplification products were subjected to a restriction fragment length polymorphism (RFLP) analysis. Results: The human Cyt b gene segment was detected in 86% (49/57) of the females of L. evansi, from 0 to 168 hours after blood ingestion. In 7% (4/57) of the individuals we amplified insect DNA, while in the remaining 7%, the band of interest was not amplified. We did not find any statistical differences between groups of females sacrificed at different times post-blood meal regarding the amplification of the human Cyt b gene segment or the number of samples amplified. Conclusion: The human Cyt b gene segment was detectable in L. evansi females up to 168 hours after blood ingestion.

DNA barcode for identification of immature stages of sand flies (Diptera: Psychodidae) collected from natural breeding sites.

Although phlebotomine sand flies breeding sites have been identified and recorded by several studies, the microhabitats exploited by these insects remain little-known and hard to find. In this context, the difficulty of finding immature stages, and the limited number of taxonomic studies to identify immature stages of phlebotomine sand flies, are considered the major obstacles when attempting a complete inventory of Lutzomyia species. The objective of this study is to validate Cytochrome Oxidase I (Barcode region) as a marker for the identification of immature stages of Lutzomyia species recovered from natural breeding sites in Colombia. Among 142 collected sand flies, 18 immature individuals that did not complete their life cycle were identified to species level through sequencing of the COI gene. Values of K2P genetic distance between 0.002-0.031 allowed the identification of larvae at species level. The bootstrap support values (96%) in the Neighbor-Joining dendrogram were consistent for the majority of the established MOTUS of Lutzomyia atroclavata, Lutzomyia micropyga, Lutzomyia serrana, Lutzomyia cayennensis, Lutzomyia rangeliana, Lutzomyia shannoni and some species of the genus Brumptomyia. The COI gene is validated as a marker for the identification of immature stages of the genus Lutzomyia.

[First report of the F1534C mutation associated with cross-resistance to DDT and pyrethroids in Aedes aegypti from Colombia].

INTRODUCTION: The main strategy for the control of Aedes aegypti, vector of dengue, chikungunya and Zika viruses, is based on the use of insecticides to reduce its populations. However, their use has led to insect resistance to these chemicals. Objective: To determine the presence of the F1534C mutation associated with cross-resistance to DDT and pyrethroids in A. aegypti in Sincelejo, Colombia. Materials and methods: We studied nine specimens of A. aegypti that showed resistance to lambdacyhalothrin in bioassays developed by the Secretaría de Salud de Sucre. We used a semi-nested PCR as previously described by Harris, et al., to amplify exon 31 of the para gene of the voltage-dependent sodium channel of A. aegypti. We sequenced, edited, and analyzed PCR products with the MEGA 5 software. Results: We detected the wild and mutant alleles of exon 31 in all of the nine mosquitoes tested, and observed the substitution of thymine for guanine in the nucleotide sequence of the mutant allele, producing a change to UGC in the UUC codon, which led to the replacement of phenylalanine by cysteine in residue 1534 of the protein. Conclusion: The nine mosquitoes analyzed presented a heterozygote genotype for the F1534C mutation, whose phenotypic effect is knockdown resistance (kdr) to DDT and pyrethroids.

Comparison of 16S and COX1 genes mitochondrial regions and their usefulness for genetic analysis of ticks (Acari: Ixodidae).

INTRODUCTION: In recent decades the analysis of mitochondrial genes has been used for population and phylogenetic studies of ticks allowing many advances in their systematics. Mitochondrial ribosomal 16S (16S) subunit is one of the most frequently used among those genes available for tick analysis, whereas cytochrome oxidase gene 1 (COX1) has recently been used and proposed as an alternative to the traditional 16S gene marker.  OBJECTIVE: To evaluate the usefulness of 16S and COX1 in genetic studies of ticks by analyzing sequences of three species commonly found in the Caribbean region of Colombia.  RESULTS: The analysis of both genes sequences allowed us to identify the three species with high levels of confidence and interspecific genetic divergence (19-22%), although only COX1 allowed us to detect intraspecific genetic variability (up to ~0.8%). A substitution saturation analysis indicated that the 16S gene was not saturated with transitions while the COX1 gene showed saturation distances starting at ~17%.  CONCLUSION: Our results indicated that the 16S gene seems to have better features for interspecific phylogenetic analyses because of its high level of genetic divergence and low saturation pattern, while the COX1 gene appears to be more useful for intraspecific genetic variability studies. However, as our study was conducted at a local scale, future studies at different biogeographical scales would help to establish its usefulness in wider and more complex scenarios.

FAMILY PSYCHODIDAE.

A catalogue is presented of the species of haematophagous and non-haematophagous psychodids recorded in Colombia. The list comprises 199 species distributed among five subfamilies and 16 genera, as follows: Subfamily Bruchomyiinae, genus Nemopalpus Macquart, 1838 (4 species); subfamily Phlebotominae, genera Brumptomyia França & Parrot, 1921 (8 species), Lutzomyia França, 1924 (153 species) and Warileya Hertig, 1948 (2 species); subfamily Psychodinae, genera Arisemus Satchell, 1955 (3 species), Australopericoma Vaillant, 1975 (1 species), Balbagathis Quate, 1996 (1 species), Clogmia Enderlein, 1937 (1 species), Didicrum Enderlein, 1937 (1 species), Feuerborniella Vaillant, 1971 (1 species), Lepidiella Enderlein, 1937 (1 species), Maruina Müller, 1895 (4 species), Paramormia Enderlein, 1935 (1 species), Parasetomima Duckhouse, 1968 (1 species) and Psychoda Latreille, 1796 (7 species); subfamily Sycoracinae, genus Sycorax Haliday, 1839 (5 species); and subfamily Trichomyiinae, genus Trichomyia Haliday, 1839 (5 species).

Comparación y utilidad de las regiones mitocondriales de los genes 16S y COX1 para los análisis genéticos en garrapatas (Acari: Ixodidae) / Comparison of 16S and COX1 genes mitochondrial regions and their usefulness for genetic analysis of ticks (Acari: Ixodidae)

Introducción. En las últimas décadas, el análisis de los genes mitocondriales se ha utilizado en los estudios poblacionales y filogenéticos de garrapatas, lo cual ha permitido numerosos avances en la sistemática de estos ácaros. El gen mitocondrial de la subunidad 16S del ARN ribosómico ( 16S ) es uno de los más usados, mientras que el gen mitocondrial de la citocromo oxidasa 1 ( COX1 ) se ha empleado recientemente y se propone como un marcador genético alternativo frente al 16S . Objetivo. Evaluar la utilidad de los genes 16S y COX1 en los estudios genéticos de las garrapatas mediante el análisis de secuencias en tres especies de la región Caribe de Colombia. Resultados. El análisis de secuencias mostró que los dos genes permitieron identificar las tres especies con mucha confiabilidad y con niveles de divergencia genética interespecífica relativamente similares (19 a 22 %), aunque solo el gen COX1 permitió detectar la variabilidad genética intraespecífica (hasta de ˜0,8 %). El análisis de saturación de sustituciones indicó que el gen 16S no se saturó con transiciones, mientras que el COX1 mostró saturación a partir de distancias de ˜17 %. Conclusión. Los resultados indicaron que el gen 16S parece tener mejores características para los análisis filogenéticos interespecíficos dada su alta divergencia genética y baja saturación de transiciones, mientras que el gen COX1 parece ser más útil para estudios de variabilidad genética intraespecífica. Sin embargo, dado que el estudio se hizo a escala local, se requieren más investigaciones en diferentes escalas biogeográficas para establecer su utilidad en circunstancias más amplias y complejas.

Introduction: In recent decades the analysis of mitochondrial genes has been used for population and phylogenetic studies of ticks allowing many advances in their systematics. Mitochondrial ribosomal 16S ( 16S ) subunit is one of the most frequently used among those genes available for tick analysis, whereas cytochrome oxidase gene 1 ( COX1 ) has recently been used and proposed as an alternative to the traditional 16S gene marker. Objective: To evaluate the usefulness of 16S and COX1 in genetic studies of ticks by analyzing sequences of three species commonly found in the Caribbean region of Colombia. Results: The analysis of both genes sequences allowed us to identify the three species with high levels of confidence and interspecific genetic divergence (19-22%), although only COX1 allowed us to detect intraspecific genetic variability (up to ˜0.8%). A substitution saturation analysis indicated that the 16S gene was not saturated with transitions while the COX1 gene showed saturation distances starting at ˜17%. Conclusion: Our results indicated that the 16S gene seems to have better features for interspecific phylogenetic analyses because of its high level of genetic divergence and low saturation pattern, while the COX1 gene appears to be more useful for intraspecific genetic variability studies. However, as our study was conducted at a local scale, future studies at different biogeographical scales would help to establish its usefulness in wider and more complex scenarios.

DETECCIÓN DE ANTICUERPOS ANTI-Leishmania (Trypanosomatidae) EN POBLACIONES CANINAS DEL DEPARTAMENTO DE SUCRE, COLOMBIA / Detection of Anti-Leishmania Antibodies (Trypanosomatidae) in Canine Populations in the Department of Sucre, Colombia

El objetivo de la investigación fue determinar la prevalencia de anticuerpos anti-Leishmania en perros del departamento de Sucre, Colombia. Se analizaron 122 perros del área rural de los municipios de Ovejas, Sampués y Sincelejo, mediante la técnica de inmunofluorescencia indirecta (IFI). Anticuerpos anti-Leishmania fueron detectados en el 69,6 % (IC95 % = 61,1 - 78,2) de la población canina estudiada. En el municipio de Sincelejo la seropositividad fue del 72 % (IC95 % = 61,1 - 82,8), en el municipio de Sampués del 68,4 % (IC95 % = 52,3 - 84,5) y en el municipio de Ovejas del 55,5 % (IC95 % = 21,2 - 86,3). Los animales seropositivos fueron principalmente caninos jóvenes y adultos. El 17,6 % de los perros seropositivos mostró signos clínicos compatibles con leishmaniasis canina, entre los cuales la onicogrifosis, alopecia y caquexia fueron los más comunes. La alta frecuencia de caninos con anticuerpos a Leishmania detectada en el presente estudio, pone de manifiesto la hiperendemicidad de la leishmaniasis canina en esta zona del país, así como el alto riesgo de brotes epidémicos de la enfermedad.

The aim of this survey wasto determine prevalence of antibodies against Leishmania in dogs from Sucre, Colombia. We analyzed 122 dogs in rural areas from municipalities of Sampués, Ovejas and Sincelejo, by indirect immunofluorescence (IFA). Anti-Leishmania antibodies were detected in 69.6 % (95 % CI = 61.1 to 78.2) of the selected canine population. In the municipality of Sincelejo, Sampues and Ovejas, the seropositivities were 72 % (95 % CI = 61.1 to 82.8), 68.4 % (95 % CI = 52.3 to 84.5) and 55.5 % (95 % CI = 21.2 to 86.3) respectively. Seropositive animals were mainly young and adult dogs. The 17.6 % of seropositive dogs showed clinical signs compatible with canine leishmaniasis; onychogryphosis, alopecia and cachexia were the most common ones. The detected high frequency of canines with antibodies against Leishmania in the present study, highlights the hyperendemicity status of canine leishmaniasis in this area of ​​the country and the high risk of disease outbreaks.

REGISTROS NUEVOS DE ESPECIES DE Lutzomyia (DIPTERA: PSYCHODIDAE) EN EL DEPARTAMENTO DE CESAR, COLOMBIA / New Records of Species of Lutzomyia (Diptera: Psychodidae) From the Department of Cesar, Colombia

El estudio de los insectos asociados a la epidemiología de la leishmaniasis, en la costa Caribe colombiana, se ha concentrado en los departamentos de Sucre y Córdoba, por consiguiente existe un escaso conocimiento de la fauna de flebotomíneos del resto de la región. En la presente nota se presentan cuatro nuevos registros de Lutzomyia spp., para el departamento de Cesar. Los insectos fueron colectados con dos trampas de luz tipo CDC, equipadas con LED de color azul, en el Balneario Hurtado de la ciudad de Valledupar, Cesar. Se recolectaron en total 50 flebotomíneos, los cuales estuvieron representados por las especies Lutzomyia cayennensis cayennensis (38 %), L. evansi (28 %), L. trinidadensis (14 %), L. venezuelensis (10 %), L. micropyga (6 %) y L. rangeliana (2 %). Como primeros registros para el departamento sobresalen L. evansi, L. venezuelensis, L. micropyga y L. rangeliana, con lo que se eleva a ocho el número de especies reportadas hasta la fecha en el Cesar. Entre los nuevos registros se destaca el hallazgo de L. evansi, reconocido vector de Leishmania spp. en la costa Caribe de Colombia.

Studies of the insects associated with the epidemiological cycle of leishmaniasis on the Caribbean coast of Colombia have been focused on the departments of Sucre and Córdoba. The phlebotomine sand fly fauna of the rest of the Caribbean region remains little known. In the present study four species of the genus Lutzomyia are reported from the department of Cesar for the first time. Sand flies were sampled using two CDC light traps with blue color LEDs at Balneario Hurtado, a tourist area in the city of Valledupar. A total of 50 phlebotomine sand flies were collected, of which the species Lutzomyia cayennensis cayennensis (38 % of the sample) and L. trinidadensis (14 %) were already known from the departament. Specimens of L. evansi (28 %), L. venezuelensis (10 %), L. micropyga (6 %) and L. rangeliana (2 %) represent the first records of these species for Cesar, raising the number of known occurrences from the department to eight species. Noteworthy among the new records is the sandfly L. evansi, recognized vector de Leishmania spp. in the Colombian Caribbean coast.

Nuevos hallazgo de flebotomíneos (diptera: psyhodidae) en la sierra nevada dee santa marta, Colombia / New Records of Phlebotomine Sand Flies (Diptera: Psychodidae) at Sierra Nevada de Santa Marta, Colombia

Los insectos relacionados con la transmisión de los patógenos causantes de las leishmaniasis han sido poco estudiados en la Sierra Nevada de Santa Marta, Colombia, incluido el departamento de Magdalena, donde a la fecha están registradas trece especies del género Lutzomyia. En la presente nota se informa el hallazgo de tres especies y un subgénero adicionales en la región. Se recolectaron 885 flebotomíneos en Seywiaka y las veredas Las Tinajas y Calabazo, estribaciones de la Sierra Nevada de Santa Marta (117-130 m s.n.m.). El 84 % de los ejemplares se obtuvieron con trampa CDC, el 11 % con trampa Shannon y el 5 % fueron capturados, en reposo, con un dispositivo eléctrico de succión. Se identificaron nueve especies, Lu. gomezi, Lu. panamensis, Lu. trinidadensis, Lu. carpenteri, Lu. evansi, Lu. dysponeta, Lu. dubitans, Lu. shannoni, y Lu. micropyga, la más abundante fue Lu. gomezi (69 %), seguida por Lu. panamensis (14 %). También se recolectaron ejemplares de la serie Lu. osornoi del subgénero Helcocyrtomyia. Entre el material hallado sobresalen Lu. carpenteri, Lu. dubitans y Lu. dysponeta como primeros registros para el departamento del Magdalena, además de Lu. (Helcocyrtomyia) sp., que representa el primer informe del subgénero en el Caribe colombiano.

Phlebotomine sand flies, vectors of leishmaniasis, have not been well studied in the Sierra Nevada de Santa Marta, and likewise, are not well known in other regions of the Department of Magdalena, Colombia. To date only thirteen species of Lutzomyia have been recorded as occurring in the Department. The present note adds three species and includes an additional subgenus. Collections were made in the lower foothills of the Sierra Nevada de Santa Marta at elevations ranging from 117-130 m in the communities of Seywiaka, Las Tinajas and Calabazo. Eighty-four percent of the 885 phlebotomines sand flies collected were obtained from CDC light traps, 11 % from Shannon trap and 5 % from typical resting sites using an electric aspirator. The following nine species were identified from the collections: Lutzomyia gomezi, Lu. panamensis, Lu. trinidadensis, Lu. carpenteri, Lu. evansi, Lu. dysponeta, Lu. dubitans, Lu. shannoni, and Lu. micropyga. The most abundant species were Lu. gomezi and Lu. panamensis, which, respectively, accounted for 69 % and 14 % of the specimens. Of the nine species, Lu. carpenteri, Lu. dubitans and Lu. dysponeta represent new records for the Department of Magdalena. Also, a few female specimens were encountered of a species belonging to the Lu. osornoi series of the subgenus Helcocyrtomyia, which represents the first record of this subgenus in the Caribbean region of Colombia.