Screening of insecticide resistance in Aedes aegypti populations collected from parishes in Eastern Jamaica.

Owing to the increased reports in Aedes-borne diseases in the Caribbean and Latin America, the United States Agency for International Development assisted the Jamaican Ministry of Health and Wellness in conducting insecticide susceptibility tests on Aedes aegypti populations. Sentinel sites were established in seven parishes of Jamaica (St. Catherine, Kingston and St. Andrew, St. Thomas, Portland, St. Mary and St. Ann) and Aedes aegypti eggs were collected, reared to adults per collected population and their susceptibility to varying pyrethroids and organophosphates were tested using the World Health Organization paper bioassays for these insecticides. The Centers for Disease Control and Prevention bottle bioassay was used to assess susceptibility to the carbamate, bendiocarb. The voltage gated sodium channel gene mutations V1016I and I1011V, normally associated with pyrethroid resistance, were also analysed. The results showed that Aedes aegypti collected from all parishes exhibited resistance to pyrethroids at the following concentrations, permethrin 0.25-2.5%; deltamethrin 0.03-0.15%; lambda-cyhalothrin 0.03-0.3%; and etofenprox 0.5-2.5%. The insecticide deltamethrin at concentration 0.3% was the only pyrethroid tested that resulted in high mortality, 94.9 ± 0.34% knockdown within 1 hour of exposure and 98.95 ± 0.01% mortality (p <0.01) at 24 hours post exposure. The frequency of the voltage gated sodium channel gene mutation V1016I was high in the tested population, possibly accounting for the reduced sensitivity to pyrethroids. Organophosphate resistance was also observed in all populations tested. Mortality rates for 0.8% Malathion was 0.8 ± 0.70-60.68 ± 0.38% after 24 hour and 0.00-47.10 ± 3.02%, for pirimiphos-methyl 0.21%. Bendiocarb applied as 12.5 µg/ bottle resulted in mortality rates of 76.25 ± 4.30-100 ± 0.00% after 30 minutes of exposure. The results showed that Ae. aegypti from the seven parishes analysed demonstrated resistance to the insecticides tested. Deltamethrin and bendiocarb at concentrations 0.3% and 12.5µg respectively, were considered most effective, causing high mortality in the local populations. Routine monitoring and evaluations of Ae. aegypti populations from the included parishes are recommended. Additionally, the study results represent the most comprehensive testing to date with local Aedes aegypti populations distributed across different parishes of Jamaica and should be useful to guide national and sub national strategies for vector control and surveillance.

Identification of descriptors for structure-activity relationship in ruthenium (II) mixed compounds with antiparasitic activity.

Herein is presented the synthesis, characterization, electrochemical studies, DFT calculations and in vitro evaluation of amoebicidal activity in trophozoites of Entamoeba histolytica of twenty ruthenium (II) mixed compounds with general formulae: [Ru(pdto)(E-E)]Clx (E-E bidentate, either neutral or negatively charged ligands). For compounds under study, O-O, N-O and N-N auxiliary donor ligands demonstrate to have a crucial impact on the electronic properties and that it is possible to modulate the antiparasitic activity. Among analyzed complexes, only four present a better performance compared to typically used metronidazole drug (IC50 < 6.80 µmol/L) to treat amebiasis disease. For studied compounds, structure-activity relationships are strongly determined by either the redox potential (E1/2) of RuII/RuIII and calculated molar volume (V) of the complexes.

Molecular phylogeny of Culex subgenus Melanoconion (Diptera: Culicidae) based on nuclear and mitochondrial protein-coding genes.

The subgenus Melanoconion of the mosquito genus Culex is taxonomically diverse and is widely distributed in the Neotropical Region, with 10 species occurring in the Nearctic Region. Species of this subgenus pose a taxonomical challenge because morphological identification is based largely on anatomical characters of the male genitalia. We addressed the monophyly of the Spissipes and Melanoconion Sections of the subgenus Melanoconion and some of the informal groups in each section. Our sample taxa included 97 specimens representing 43 species, from which we analysed fragments of two single-copy nuclear genes (CAD, HB) and one mitochondrial gene (COI). Phylogenetic relationships within the subgenus are presented based on results of maximum-likelihood and Bayesian analyses using a multi-locus matrix of DNA sequences. We show a molecular phylogeny of Melanoconion in which both sections were recovered as monophyletic groups. The monophyly of the Atratus and Pilosus groups was confirmed. Within each section, other monophyletic groups were recovered highlighting the potential need for future nomenclature rearrangement. The phylogenetic signal contained in nuclear genes, when analysed together, was more informative than each gene analysed separately, corroborating monophyly of Melanoconion relative to Culex (Culex) species included in the analyses, the Melanoconion and Spissipes Sections and some species groups. Our results provide new information for the classification of the subgenus and additional data that can be used to improve species identification when a more representative taxon sampling is available.

Water-Soluble Ruthenium (II) Chiral Heteroleptic Complexes with Amoebicidal in Vitro and in Vivo Activity.

Three water-soluble Ru(II) chiral heteroleptic coordination compounds [Ru(en)(pdto)]Cl2 (1), [Ru(gly)(pdto)]Cl (2), and [Ru(acac)(pdto)]Cl (3), where pdto = 2,2'-[1,2-ethanediylbis-(sulfanediyl-2,1-ethanediyl)]dipyridine, en = ethylendiamine, gly = glycinate, and acac = acetylacetonate, have been synthezised and fully characterized. The crystal structures of compounds 1-3 are described. The IC50 values for compounds 1-3 are within nanomolar range (14, 12, and 6 nM, respectively). The cytotoxicity for human peripheral blood lymphocytes is extremely low (>100 µM). Selectivity indexes for Ru(II) compounds are in the range 700-1300. Trophozoites exposed to Ru(II) compounds die through an apoptotic pathway triggered by ROS production. The orally administration to infected mice induces a total elimination of the parasite charge in mice faeces 1-2-fold faster than metronidazole. Besides, all compounds inhibit the trophozoite proliferation in amoebic liver abscess induced in hamster. All our results lead us to propose these compounds as promising candidates as antiparasitic agents.

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.

Mitochondrial COI gene as a tool in the taxonomy of mosquitoes Culex subgenus Melanoconion.

The subgenus Melanoconion is the second largest subgenus within the genus Culex, with 160 described species. Several of the species are proven vectors of arboviruses, including West Nile virus, Venezuelan equine encephalitis virus complex and Eastern equine encephalomyelitis virus. Species of Melanoconion are well distributed from southern North America to most countries of South America and display the highest species diversity in tropical regions. Taxonomical identification within this group has been primarily based on morphological characters, with the male genitalia as the source of the most solid diagnostic features. The difficulty in reaching accurate species determinations when studying specimens of Culex (Melanoconion) has been extensively documented as a real limitation to expand knowledge of these insects. We tested the utility of the mitochondrial gene COI as a complementary tool in the taxonomy of Melanoconion. Using a data set of 120 COI sequences from Culex specimen captured in several localities in Brazil, the utility of COI barcodes for species delimitation is discussed through the evaluation of genetic divergences among specimens and the clustering patterns of species in three topologies obtained with Neighbor Joining, Maximum Likelihood and Bayesian phylogenetic inference. For all specimens included in this study a previous morphological examination was performed, and most of the taxonomical determinations were corroborated using the COI barcode. We generated COI sequences that belong to 48 species of Melanoconion, with a mean intraspecific K2P genetic divergence of 3%; and all interspecific divergence values higher than the intraspecific divergence values. This is the first comprehensive study of subgenus Melanoconion, with evidence of COI as a useful and accessible DNA barcode.

Catalog of the subgenus Melanoconion of Culex (Diptera: Culicidae) for South America.

Species of Culex (Melanoconion) Theobald are recognized as vectors of arboviruses. The species of this subgenus pose a real taxonomic challenge. The current classification of the subgenus recognizes a total of 160 species divided in two major sections, Melanoconion and Spissipes; and several non-formal groupings within each section. We gathered bibliographic records of the subgenus in South America, with particular focus on the period of time after the publication of the Catalog by Pecor et al. (1992) until present time. This compilation included 139 species occurring in South American countries with all the relevant bibliographic sources, including the corresponding information for those medically important species.

Study on natural breeding sites of sand flies (Diptera: Phlebotominae) in areas of Leishmania transmission in Colombia.

BACKGROUND: The location of the microhabitats where immature phlebotomine sand flies of the genus Lutzomyia develop is one of the least-known aspects of this group of medically important insects. For this reason strategies of source reduction approach for their control have not been possible in contrast to other insect vectors (such as mosquitoes), because their juvenile stages in terrestrial microhabitats is difficult to detect. METHODS: Direct examination of soil samples, incubation of substrates and the use of emergence traps were the methods used to identify juvenile stages in 160 soil samples from urban and forest habitats within the foci of Leishmania transmission in Colombia. Immatures collected were identified subsequent from the rearing and emergence of adults using taxonomic keys or the analysis of the mitochondrial marker cytochrome oxidase I. Plant species associated with the natural breeding sites were identified and physicochemical properties of the soils were analyzed. RESULTS: A total of 38 (23.7%) sampling sites were identified as breeding sites, 142 phlebotomine sand flies were identified, belonging to 13 species of the genus Lutzomyia and two of Brumptomyia. The greatest numbers of immature were found within the tabular roots (51 immature sand flies from eight positive sites) and bases of trees (35 immature sand flies from 11 sites). The characterization and presence of the tree species (mainly Ceiba pentadra, Anacardium excelsum, Pseudosamanea guachapale) and the physicochemical properties (relative humidity and carbon/nitrogen ratio) of the soils associated with these breeding sites are significant factors in explaining the diversity and abundance of phlebotomine sand flies. CONCLUSIONS: Immature phlebotomine sand flies of the genus Lutzomyia in Colombia can be found in a wide variety of breeding sites rich in organic matter, high relative humidity and are associated with a typical vegetation of each locality. These results provide new perspectives for the study of the ecology of the genus Lutzomyia in Colombia and the development of vector control strategies.

Flebotominos adultos e inmaduros (Diptera: Psychodidae): registros para el Caribe colombiano / Adult and immature phlebotomine sandflies (Diptera: Psychodidae): records for the Caribbean región of Colombia

Elaborar inventarios de lutzomyia spp., sin considerar los estados inmaduros de dichos insectos, provee información parcial. Investigar los estados inmaduros de lutzomyia, es un reto y necesidad actual especialmente en áreas con transmisión de leishmania spp. El objetivo de este estudio fue detectar sitios de cría naturales, para un inventario de flebotominos adultos e inmaduros del municipio de Colosó (Sucre, Colombia). Los flebotominos fueron recolectados entre mayo y diciembre de 2009, en la estación experimental de fauna silvestre de Colosó. La detección de estados inmaduros se desarrolló por revisión directa de muestras de suelo y árboles, incubación de estos sustratos en laboratorio y por trampas de emergencia. La colecta de adultos se realizó por búsqueda activa diurna con aspiradores bucales en sitios de reposo, principalmente en bases, huecos y raíces tabulares de árboles. Se aislaron 44 inmaduros de flebotominos, de los cuales 32 correspondieron a las especies lutzomyia migonei, Lu. dubitans, Lu. serrana, Lu. cayennensis cayennensis, Lu. micropyga, Lu. evansi, Lu. gorbitzi, Lu. ovallesi y Lu. shannoni. Así mismo, se colectaron 1231 ejemplares adultos entre los cuales Lu. evansi, Lu. micropyga y Lu. trinidadensis fueron, en su orden, las especies más abundantes. Lu. migonei y Lu. gorbitzi constituyen, respectivamente, primeros registros para el departamento de Sucre y la Costa Caribe. Es necesario incluir muestreos de insectos inmaduros como información complementaria en estudios sobre flebotominos y así reunir información sólida para elaborar inventarios de especies señalando los potenciales vectores en focos de leishmaniasis.

Elaborating an inventory for Lutzomyia spp., without considering the immature forms of such insects, gives only a partial information about the species. Investigating immature phlebotomines represents a challenge and urgency especially in Leishmania spp. transmission areas. The objective of this study was to detect natural breeding sites, for an inventory of immature and adult phlebotomine community in the municipality of Colosó, Department of Sucre. Phlebotomine sandflies were collected between May and December of 2009, at the wild life experimental station in Colosó. Detection of immature stages was approached by direct visualization of soil and tree-substrate samples, incubation of substrate in laboratory conditions and use of emergence traps. Adult sampling was conducted by active diurnal search in resting places using bucal aspirators to collect the insects. Resting places were predominantly associated with tree trunks, such as holes in trunks and buttress roots. We collected 44 immatures of phlebotomine sandflies of which 32 belong to the following species: Lutzomyia migonei, Lu. dubitans, Lu. serrana, Lu. cayennensis cayennensis, Lu. micropyga, Lu. evansi, Lu. gorbitzi, Lu. ovallesi and Lu. shannoni. Also, up to 1231 adult individuals were collected and the most abundant species in descending order were Lu. evansi, Lu. micropyga and Lu. trinidadensis. The species Lu. migonei and Lu. gorbitzi are worth noticing given the fact that they represent new records for the Department of Sucre as well as the Caribbean Region in the country. It is necessary to include immature sampling as complementary information on phlebotomine surveys and in this way gather solid information to release proper species inventories with the remarks on potential vectors in leishmaniasis foci.