Virome analyses of Amblyomma cajennense and Rhipicephalus microplus ticks collected in Colombia.

Tick-borne viruses (TBV) have gained public health relevance in recent years due to the recognition of human-associated fatal cases and the increase in tick-borne disease and transmission. However, many tick species have not been studied for their potential to transmit pathogenic viruses, especially those found in Latin America. To gain better understanding of the tick virome, we conducted targeted amplification using broadly-reactive consensus-degenerate pan-viral targeting viruses from the genera Flavivirus, Bandavirus, Uukuvirus, and Orthonairovirus genus. Additionally, we conducted unbiased metagenomic analyses to investigate the presence of viral RNA sequences in Amblyomma cajennense, A. patinoi and Rhipicephalus microplus ticks collected from a horse slaughter plant in Medellín, Colombia. While no viral products were detected by PCR, results of the metagenomic analyses revealed the presence of viral genomes belonging to the genera Phlebovirus, Bandavirus, and Uukuvirus, including Lihan Tick Virus (LTV), which was previously reported in Rhipicephalus microplus from Colombia. Overall, the results emphasized the enormous utility of the next-generation sequencing in identifying virus genetic diversity presents in ticks and other species of vectors and reservoirs.

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.

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.

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.

Revisiting the heterogeneous global genomic population structure of Leishmania infantum.

Leishmania infantum is the main causative agent responsible for visceral leishmaniasis (VL), a disease with global distribution. The genomic structure and genetic variation of this species have been widely studied in different parts of the world. However, in some countries, this information is still yet unknown, as is the genomic behaviour of the main antigens used in VL diagnosis (rK39 and rK28), which have demonstrated variable sensitivity and specificity in a manner dependent on the geographic region analysed. The objective of this study was to explore the genomic architecture and diversity of four Colombian L. infantum isolates obtained in this study and to compare these results with the genetic analysis of 183 L. infantum isolates from across the world (obtained from public databases), as well as to analyse the whole rK39 and rK28 antigen sequences in our dataset. The results showed that, at the global level, L. infantum has high genetic homogeneity and extensive aneuploidy. Furthermore, we demonstrated that there are distinct populations of L. infantum circulating in various countries throughout the globe and that populations of distant countries have close genomic relationships. Additionally, this study demonstrated the high genetic variability of the rK28 antigen worldwide. In conclusion, our study allowed us to (i) expand our knowledge of the genomic structure of global L. infantum; (ii) describe the intra-specific genomic variability of this species; and (iii) understand the genomic characteristics of the main antigens used in the diagnosis of VL. Additionally, this is the first study to report whole-genome sequences of Colombian L. infantum isolates.

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.

Molecular detection of Anaplasma spp. in domestics dogs from urban areas of Soledad, Atlantico, Colombia.

Tick-borne pathogens are etiological agents of some zoonotic diseases, causing important consequences in animal and human health. These are emerging around the world, especially in tropical countries including Colombia. Domestic dogs play an essential role in the epidemiology of several zoonotic tick-borne pathogens. We performed the detection of bacteria from Anaplasmataceae family and parasites from the Piroplasmida order, in 85 domestic dogs from Soledad municipality, Atlantico, Colombia. Peripheral blood smears, detection by duplex PCR assay (ss rRNA 16S, from bacteria and the ITS-1, of ribosomal DNA from parasites), and DNA sequencing by Sanger method were done. Taxonomic identification was made by phylogenetics analysis of the DNA sequences. The gene sequences analysis showed that 12.9% of the dogs were infected with Anaplasma spp. Infection was higher in young dogs (OR=4.72, 95%CI 1.267-17.584). Besides that, 3.5% of them showed inclusions (morulae) compatible with bacteria from the order Rickettsiales. A coinfection with Babesia spp. and a Rickettsiales bacterial pathogens was found. The frequency of Anaplasma spp. detected in domestic dogs in Soledad highlights the need to improve diagnosis and control measures, to prevent the risk of transmission of these pathogens among ticks, dogs and humans exposed in the area.

[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 barcoding to identify species of phlebotomine sand fly (Diptera: Psychodidae) in the mixed leishmaniasis focus of the Colombian Caribbean.

Identification of the species of phlebotomine sand flies present in each focus of leishmaniasis is necessary to incriminate vectors and implement vector control strategies. Although the cytochrome oxidase I (COI) gene has been proposed as a barcode for the identification of animal species, less than 20% of New World phlebotomines have been characterized to date. In this study DNA barcoding was used to identify phlebotomine species of the mixed leishmaniasis focus in the Colombian Caribbean by means of three evolutionary models: Kimura's two parameter (K2P) nucleotide substitution model, that of (Tamura and Nei, 1993) (TN93) and proportional sequence divergence (p-distances). A 681bp sequence of the COI gene was obtained from 66 individuals belonging to 19 species of the genus Lutzomyia (Lu. abonnenci, Lu. atroclavata, Lu. bicolor, Lu. carpenteri, Lu. cayennensis cayennensis, Lu. dubitans, Lu. evansi, Lu. gomezi, Lu. gorbitzi, Lu. longipalpis, Lu. micropyga, Lu. migonei, Lu. panamensis, Lu. (Psathyromyia) sp., Lu. rangeliana, Lu. serrana, Lu. shannoni, Lu. trinidadensis and Lu. venezuelensis) and one of Brumptomyia (B. mesai). The genetic divergence values for TN93 among individuals of the same species fluctuated up to 3.2% (vs. 2.9% for K2P and 2.8% for p-distances), while the values between species ranged from 8.8-43.7% (vs. 6.8-19.6% for K2P and 6.6-17.4% for p-distances). A dendrogram constructed by means of the Neighbor-Joining method grouped phlebotomines into 20 clusters according to species, with bootstrap values of up to 100% in those with more than one individual. However, loss of the phylogenetic signal of the gene COI was observed at the supraspecific level as a consequence of substitutional saturation. In conclusion, irrespective of the evolutionary model selected, all phlebotomines were correctly assigned to species, showing 100% concordance with morphological identification.

Evidence for anthropophily in five species of phlebotomine sand flies (Diptera: Psychodidae) from northern Colombia, revealed by molecular identification of bloodmeals.

Identification of the bloodmeal sources of phlebotomine sand flies is fundamental to determining which species are anthropophilic and understanding the transmission of Leishmania parasites in natural epidemiological settings. The objective of this study was to identify sand fly bloodmeals in the mixed leishmaniasis focus of the department of Sucre, northern Colombia. In all 141 engorged female sand flies were analyzed, after being captured in intradomiciliary, peridomiciliary and extradomiciliary habitats with Shannon and CDC traps and by active searching in diurnal resting sites. Bloodmeals were identified by sequencing and analysis of a 358bp fragment of the mitochondrial gene Cytochrome b (CYB) and a 330bp fragment of the nuclear gene prepronociceptin (PNOC). Using both genes 105 vertebrate bloodmeals were identified, with an efficiency of 72% for CYB but only 7% for PNOC. Ten species of vertebrates were identified as providing bloodmeal sources for 8 sand fly species: Homo sapiens (Lutzomyia evansi, Lutzomyia panamensis, Lutzomyia micropyga, Lutzomyia shannoni and Lutzomyia atroclavata), Equus caballus (L. evansi, L. panamensis and Lutzomyia cayennensis cayennensis), Equus asinus (L. evansi and L. panamensis), Bos taurus (L. evansi, L. panamensis and L. c. cayennensis), Tamandua mexicana (L. shannoni and Lutzomyia trinidadensis), Proechimys guyanensis (L. evansi, L. panamensis and L. c. cayennensis), Mabuya sp. (Lutzomyia micropyga), Anolissp. (L. micropyga), Sus scrofa (L. evansi and Lutzomyia gomezi) and Gallus gallus (L. evansi). Cattle, donkeys, humans and pigs were significantly more important than other animals (P=0.0001) as hosts of L. evansi, this being the most abundant sand fly species. The five Lutzomyia species in which blood samples of human origin were detected included L. micropyga and L. atroclavata, constituting the first evidence of anthropophily in both species.

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.

Diferencias genéticas entre poblaciones de Aedes aegypti de municipios del norte de Colombia, con baja y alta incidencia de dengue / Genetic differences between populations of Aedes aegypti from municipalities in northern Colombia, with high and low dengue incidence

Introducción. Aedes aegypti es el principal vector del dengue en zonas urbanas. A pesar de su importancia epidemiológica, se desconoce la variabilidad genética de las poblaciones del vector en Colombia. Objetivo. Determinar la variabilidad genética del gen mitocondrial ND4 , que codifica para la subunidad 4 de la enzima NADH-deshidrogenasa, entre poblaciones de Ae. aegypti de los municipios de Sincelejo y Guaranda, donde se registra alta y baja incidencia de dengue, respectivamente. Materiales y métodos. A partir del material genético extraído de 36 hembras de Ae. aegypti , se determinó la secuencia parcial del gen mitocondrial ND4 y se estimaron los parámetros de diversidad de nucleótidos, diversidad haplotípica, estructura genética y flujo de genes entre las poblaciones de Sincelejo y Guaranda. También, se analizó la varianza molecular y se construyó una red haplotípica. Resultados. Se obtuvieron 36 secuencias de nucleótidos de 282 pb; éstas presentaron doce sitios polimórficos y se agruparon en diez haplotipos, dos presentes en ambas poblaciones, tres exclusivos de la población de Sincelejo y cinco de la población de Guaranda. Los estimadores de estructura genética ( F ST =0,15) y de flujo de genes ( Nm =1,40) evidencian diferenciación genética y un limitado intercambio de genes entre las poblaciones. Conclusión. Las poblaciones de Ae. aegypti de Sincelejo y Guaranda son genéticamente divergentes.

Introduction: Aedes aegypti is the principal vector of dengue in urban areas. Despite its epidemiological importance, the genetic variability of Colombian populations of this species is unknown. Objetive: To determine the genetic variability of mitocondrial gene ND4, which codes for subunit 4 of the enzyme NADH deshydrogenase, between populations of Ae. aegypti from municipalities of Sincelejo and Guaranda. The incidences of dengue reported from these two localities are high and low, respectively. Materials and methods: Genetic material extracted from 36 females of Ae. aegypti was used to determine the partial sequence of the mitocondrial gene ND4 as well as to estimate the parameters of nucleotidic and haplotypic diversities, genetic structure and gene flow between the Sincelejo and Guaranda populations. The molecular variance was also analysed and a haplotypic network constructed. Results: In all 36 nucleotide sequences of 282pb were obtained. These presented 12 polymorphic sites and could grouped into 10 haplotypes, two of them present in both populations, three exclusive to the Sincelejo population and five to that of Guaranda. The estimators of genetic structure ( F ST = 0.15) and gene flow ( Nm = 1.40) are both indicative of genetic differentiation and a limited exchange of genes between the populations. Conclusions: The Sincelejo and Guaranda populations of Ae. aegypti are genetically divergent.

[Genetic differences between populations of Aedes aegypti from municipalities in northern Colombia, with high and low dengue incidence]. / Diferencias genéticas entre poblaciones de Aedes aegypti de municipios del norte de Colombia, con baja y alta incidencia de dengue.

INTRODUCTION: Aedes aegypti is the principal vector of dengue in urban areas. Despite its epidemiological importance, the genetic variability of Colombian populations of this species is unknown. OBJETIVE: To determine the genetic variability of mitocondrial gene ND4, which codes for subunit 4 of the enzyme NADH deshydrogenase, between populations of Ae. aegypti from municipalities of Sincelejo and Guaranda. The incidences of dengue reported from these two localities are high and low, respectively. MATERIALS AND METHODS: Genetic material extracted from 36 females of Ae. aegypti was used to determine the partial sequence of the mitocondrial gene ND4 as well as to estimate the parameters of nucleotidic and haplotypic diversities, genetic structure and gene flow between the Sincelejo and Guaranda populations. The molecular variance was also analysed and a haplotypic network constructed. RESULTS: In all 36 nucleotide sequences of 282 pb were obtained. These presented 12 polymorphic sites and could grouped into 10 haplotypes, two of them present in both populations, three exclusive to the Sincelejo population and five to that of Guaranda. The estimators of genetic structure (FST = 0.15) and gene flow (Nm = 1.40) are both indicative of genetic differentiation and a limited exchange of genes between the populations. CONCLUSIONS: The Sincelejo and Guaranda populations of Ae. aegypti are genetically divergent.

tRNASer (UCN) MITOCONDRIAL DE Lutzomyia hartmanni PREDICCIÓN DE LA ESTRUCTURA SECUNDARIA DEL tRNASer (UCN) MITOCONDRIAL DEL FLEBOTOMÍNEO Lutzomyia hartmanni (DIPTERA: PSYCHODIDAE) / Prediction of the Secondary Structure of the Mitochondrial tRNASer (UCN) of Lutzomyia hartmanni (Diptera: Psychodidae)

Lutzomyia (Helcocyrtomyia) hartmanni es un flebotomíneo implicado en la transmisión de Leishmania (Viannia) colombiensis, uno de los agentes etiológicos de leishmaniasis cutánea en Colombia. El objetivo de este trabajo fue explorar la utilidad potencial del RNA de transferencia mitocondrial para Serina (UCN) (tRNASer), en la discriminación taxonómica de L. hartmanni. El DNA mitocondrial se extrajo, amplificó y secuenció a partir de material entomológico recolectado en Envigado, Antioquia, Colombia. El gen tRNASer de L. hartmanni mostró una longitud de 68 pares de bases, con un contenido AT del 80,9%. Éste se diferencia de los demás tRNASer de Lutzomyia conocidos a la fecha tanto por sustituciones en la secuencia primaria de nucleótidos como por los cambios que éstas generan en la estructura secundaria. El número de apareamientos intracatenarios fue siete en el brazo aceptor del aminoácido, tres en el brazo dihidrouridina (DHU), cinco en el brazo del anticodón y cinco en el brazo ribotimidina-pseudouridina-citosina (T C). El tamaño de las lupas DHU, anticodón, variable y T C correspondió a cinco, siete, cuatro y ocho nucleótidos, respectivamente. La ausencia notoria de pares de bases no-Watson-Crick en los cuatro brazos del tRNASer de L. hartmanni, la distingue de otras especies de Lutzomyia.

Lutzomyia (Helcocyrtomyia) hartmanni is a sand fly that has been implicated in the transmission of Leishmania (Viannia) colombiensis, an etiologic agent of cutaneous leishmaniasis in Colombia. The objective of this work was to explore the potential usefulness of the mitochondrial serine transfer RNA (UCN) (tRNASer) in the taxonomic determination of L. hartmanni. Mitochondrial DNA was extracted, amplified and sequenced from entomological material collected in Envigado, Antioquia, Colombia. The tRNASer gene length was 68 nucleotide pairs, with an average adenine-thymine content of 80,9%. The studied tRNASer differs from other sand fly tRNASer known to date, on the basis of its primary and secondary structure. The observed number of intrachain base pairing was 7 in the acceptor arm, 3 in the dihydrouridine (DHU) arm, 5 in the anticodon arm, and 5 in the ribothymidine-pseudouridine-cytosine (T C) arm. The size of the DHU, anticodon, variable and T C loops was estimated to be 5, 7, 4, and 8 nucleotides, respectively. The notorious absence of non-Watson-Crick base pairs in the four arms of the tRNASer distinguishes that of L. hartmanni from others Lutzomyia spp.

Trichomyia andina sp. nov., un nuevo psicódido no hematófago (Diptera: Psychodidae: Trichomyiinae) de Colombia / Trichomyia andina sp. nov., a new non-hematophagous psychodid fly (Diptera: Psychodidae: Trichomyiinae) of Colombia

Se describe e ilustra una nueva especie de psicódido no hematófago, Trichomyia andina, a partir de machos recolectados en el Departamento de Risaralda, Colombia. Este psicódido pertenece al subgénero Opisthotrichomyia Bravo, 2001, y se distingue de las especies co-génericas por presentar un gonostilo ancho, multipartito, parcialmente serrado e infuscado, que termina en una larga punta translucida dirigida hacia adentro y atrás. El subgénero Opisthotrichomyia queda integrado ahora por siete especies, incluidas T. brevitarsa (Rapp, 1945), T. vargasi Barretto, 1954, T. nocturna Bravo, 2001, T. festiva Bravo, 2001, T. fluminensis Bravo, 2001, T. riodocensis Alexander, Freitas & Quate, 2001, y T. andina, sp. nov.

Trichomyia andina, a new species of non-hematophagous psychodid fly is described and illustrated from males collected in the Risaralda Department, Colombia. This species belongs to the subgenus Opisthotrichomyia Bravo, 2001 and is distinguished from congeners in having a gonostyle broad, multipartite, partially serrated and infuscated, with a long hyaline apex directed inward and backward. The subgenus Opisthotrichomyia is now comprised of seven species, including T. brevitarsa (Rapp 1945), T. vargasi Barretto, 1954, T. nocturna Bravo, 2001, T. festiva Bravo, 2001, T. fluminensis Bravo, 2001, T. riodocensis Alexander, Freitas & Quate, 2001, and T. andina, sp. nov.

Trece registros nuevos de Lutzomyia (Diptera: Psychodidae) para el departamento de Vichada, Orinoquia Colombiana / Thirteen new records of Lutzomyia (Diptera: Psychodidae) for the department of Vichada, Colombian Orinoquia

Se informa el primer hallazgo de Lutzomyia trinidadensis (Newstead, 1922), L. bettinii Feliciangeli, Ramírez-Pérez & Ramírez, 1988, L. flaviscutellata (Mangabeira, 1942), L. yuilli Young & Porter, 1972, L. saulensis (Floch & Abonnenc, 1944), L. runoides (Fairchild & Hertig, 1953), L. ayrozai (Barretto & Coutinho, 1940), L. claustrei Abonnenc, Leger & Fauran, 1979, L. walkeri (Newstead, 1914), L. preclara Young & Arias, 1984, L. (geniculata) sp., L. (squamiventris) sp., y L. (Evandromyia) sp., en el departamento de Vichada, Colombia. También se confirma la presencia de L. antunesi (Coutinho, 1939) y L. aragaoi (Costa Lima, 1932) en este departamento. Los flebotomíneos fueron recolectados en los municipios de Puerto Carreño, Cumaribo, y La Primavera, con trampas de luz tipo CDC, trampa Shannon y atrayente humano, y mediante búsqueda activa en lugares de reposo. Se discute la importancia epidemiológica de estas especies de flebotomíneos en relación con la leishmaniosis cutánea.

Lutzomyia trinidadensis (Newstead, 1922), L. bettinii Feliciangeli, Ramírez-Pérez & Ramírez, 1988, L. flaviscutellata (Mangabeira, 1942), L. yuilli Young & Porter, 1972, L. saulensis (Floch & Abonnenc, 1944), L. runoides (Fairchild & Hertig, 1953), L. ayrozai (Barretto & Coutinho, 1940), L. claustrei Abonnenc, Leger & Fauran, 1979, L. walkeri (Newstead, 1914), L. preclara Young & Arias, 1984, L. (geniculata) sp., L. (squamiventris) sp., and L. (Evandromyia) sp., are recorded for the first time in the department of Vichada, Colombia. In addition, the presence of L. antunesi (Coutinho, 1939) and L. aragaoi (Costa Lima, 1932) in this department is confirmed. Sand flies were collected in the municipalities of Puerto Carreño, Cumaribo, and La Primavera, using CDC light traps, Shannon trap, and human bait, and by active search in resting places. The epidemiological importance of these sand fly species is discussed in relation with the cutaneous leishmaniasis.

Una nueva especie de Trichomyia Haliday (Diptera, Psychodidae) de Los Montes de María, Colombia / A new species of Trichomyia Haliday (Diptera, Psychodidae) from Los Montes de María, Colombia

Una nueva especie de Trichomyia Haliday (Diptera, Psychodidae) de Los Montes de María, Colombia. Trichomyia colosensis n. sp. es descrita e ilustrada a partir de ejemplares machos recolectados con una trampa modificada CDC en la Reserva Forestal Protectora Serranía de Coraza y Montes de María, sobre la Costa Caribe Colombiana. Hasta la fecha están registradas dos especies de Trichomyia en el Caribe colombiano, T. brevitarsa (Rapp, 1945) y T. colosensis n. sp.

A new species of Trichomyia Haliday (Diptera, Psychodidae) from Los Montes de María, Colombia. Trichomyia colosensis n. sp. is described and illustrated from male specimens collected with a modified CDC light trap in the Reserva Forestal Protectora Serranía de Coraza y Montes de María, on the Caribbean Coast of Colombia. To date, two species of Trichomyia have been reported from the Colombian Caribbean, T. brevitarsa (Rapp, 1945) and T. colosensis n. sp.

Flebotomíneos (Diptera: Psychodidae) antropofílicos de importancia en salud pública en Los Montes de María, Colombia / Anthropophilic sand flies (Diptera: Psychodidae) of public health importance in Los Montes de María, Colombia

INTRODUCCIÓN: la leishmaniasis cutánea es una enfermedad endémica en Los Montes de María, Colombia, pero hasta la fecha se desconocían los flebotomíneos que pican al humano en el área. OBJETIVO: determinar las especies de Lutzomyia que, por sus hábitos antropofílicos y antecedentes vectoriales, podrían mantener el ciclo epidemiológico de la leishmaniasis cutánea en la región. MÉTODOS: se recolectaron los insectos con cebo humano protegido durante enero y agosto de 2005, que comprende parte de la época seca, así como en abril y noviembre del mismo año, pertenecientes a la temporada de lluvias. Tres personas equipadas con aspiradores bucales y linternas se ubicaron entre las 18:00 y 24:00 h dentro de una vivienda, donde se había registrado un caso de leishmaniasis cutánea. RESULTADOS: se obtuvieron 567 flebotomíneos del género Lutzomyia, distribuidos en 504 hembras y 63 machos. La constitución de la fauna de flebotomíneos antropofílicos correspondió a 97,5 % Lutzomyia evansi, 1,23 % Lutzomyia cayennensis cayennensis, 0,5 % Lutzomyia panamensis, 0,5 % Lutzomyia dubitans y 0,2 % Lutzomyia gomezi. La tasa de picadura de las hembras de Lutzomyia evansi fue de 1,5 en enero, 1,4 en abril, 0,85 en agosto y 0,6 en noviembre. En las demás especies la tasa de picadura mostró valores de 0 en enero y abril, e iguales o inferiores a 0,024 en agosto y noviembre. CONCLUSIONES: algunas de las especies de flebotomíneos encontradas podrían participar en el ciclo epidemiológico de la leishmaniasis cutánea en la región.

INTRODUCTION: cutaneous leishmaniasis is an endemic disease in Los Montes de María, Colombia, but the identity of the human-biting sand fly species in the region remained unknown until now. OBJECTIVE: to determine the Lutzomyia species that could be involved in the epidemiological cycle of cutaneous leishmaniasis in this area, considering their anthropophilic habits and epidemiological background. METHODS: sand flies were sampled using protected human baits, during the dry season in January and August, 2005, and the rainy period in April and November, 2005. Human bait collections were made from 18:00 to 24:00 hours by three men equipped with mouth aspirators and torches, inside a house where a cutaneous leishmaniasis case had occurred the last year. RESULTS: a total of 567 sand flies of the genus Lutzomyia were collected, comprising 504 females and 63 males. The composition of anthropophilic sand fly fauna was 97.5 % of Lu. evansi, 1.23 % of Lu. cayennensis cayennensis, 0.5 % of Lu. panamensis, 0.5 % of Lu. dubitans, and 0.2 % of Lu. gomezi. The L. evansi female's biting rate on humans was 1.5 in January, 1.4 in April, 0.85 in August, and 0.6 in November. Other Lutzomyia species exhibited human-biting rates equal to 0 in January and April; and equal to or lower than 0,024 in August and November. CONCLUSIONS: some of these sand fly species might play a role in the epidemiological cycle of cutaneous leishmaniasis in the region.

Brumptomyia hamata (PSYCHODIDAE), UN NUEVO APORTE A LA FAUNA FLEBOTOMÍNEA DEL CARIBE COLOMBIANO / Brumptomyia hamata (Psychodidae), A New Addition To The Phlebotomine Fauna Of The Colombian Caribbean

La fauna flebotomínea del Caribe colombiano es aún poco conocida, por lo que se hace necesario llevar a cabo estudios para determinar la riqueza de especies de la región. Con este propósito se desarrollaron muestreos entomológicos en enero y marzo de 2007, en el municipio de Colosó, departamento de Sucre, con dos trampas CDC modificadas. En el material recolectado se identificaron dos machos de Brumptomyia hamata, que constituyen el primer registro de este flebotomíneo para el Caribe colombiano. Las características morfológicas observadas en los ejemplares de B. hamata son consistentes con la descripción original de la especie, excepto por las espinas del estilo que se distribuyen en tres grupos. Se plantea que tales diferencias pueden corresponder a variaciones intraespecíficas entre poblaciones geográficas.

The sand fly fauna of the Colombian Caribbean is still not completely known and further surveys are needed in order to determine the richness of species over the region. Entomological samplings were performed in the municipality of Colosó , department of Sucre , using two modified CDC traps, in January and March of 2007. Two males belonging to Brumptomyia hamata were identified. It is the first record of this sand fly species from the Colombian Caribbean. With the exception of the distribution of spines on the style in three distinct groups, all other morphological characters of the individuals of B. hamata agree with the original description of the species. The observed difference could be attributed to intraspecific variation among geographic populations.