BackgroundMosquitoes host and pass on to humans a variety of disease-causing pathogens such as infectious viruses and other parasitic microorganisms. The emergence and spread of insecticide resistance is threatening the effectiveness of current control measures for common mosquito vector borne diseases, such as malaria, dengue and Zika. Therefore, the emerging resistance to the widely used pyrethroid insecticides is an alarming problem for public health. Herein we demonstrated the use of RNA interference (RNAi) to increase susceptibility of adult mosquitoes to a widely used pyrethroid insecticide.MethodsExperiments were performed on a field-collected pyrethroid resistant strain of Ae. aegypti (Rio de Janeiro; RJ). Larvae from the resistant Ae. aegypti population were soaked with double-stranded RNAs (dsRNAs) that correspond either to voltage-gate sodium channel (VGSC), P-glycoprotein, or P450 detoxification genes and reared to adulthood. Adult mortality rates in the presence of various Deltamethrin pyrethroid concentrations were used to assess mosquito insecticide susceptibility.ResultsWe characterized the RJ Ae. aegypti strain with regard to its level of resistance to a pyrethroid insecticide and found that it was approximately 6 times more resistant to Deltamethrin compared to the laboratory Rockefeller strain. The RJ strain displayed a higher frequency of Val1016Ile and Phe1534Cys substitutions of the VGSC gene. The resistant strain also displayed a higher basal expression level of VGSC compared to the Rockefeller strain. When dsRNA-treated mosquitoes were subjected to a standard pyrethroid contact bioassay, only dsRNA targeting VGSC increased the adult mortality of the pyrethroid resistant strain. The dsRNA treatment proved effective in increasing adult mosquito susceptibility over a range of pyrethroid concentrations and these results were associated with dsRNA-specific small interfering RNAs in treated adults, and the corresponding specific down regulation of VGSC gene expression level. Finally, we demonstrated that the efficiency of our approach was further improved by ‘tiling’ along the VGSC gene in order to identify the most potent dsRNA sequences.ConclusionsThese results demonstrate that dsRNA applied to mosquito larvae retains its biological activity into adulthood. Thus, the RNAi system reported here could be a useful approach to control the widespread insecticide resistance in mosquitoes and other insect vectors of human diseases.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-016-1634-y) contains supplementary material, which is available to authorized users.
O objetivo da investigação foi analisar a variação da diversidade e abundância das espécies de Culicidae e sua relação com algumas variáveis ambientais, bem como estimar a taxa de paridade de Anopheles cruzii Dyar & Knab, 1908. O estudo foi desenvolvido em Floresta Ombrófila Densa da Mata Atlântica, localizada no Estado do Paraná, denominada de Floresta Estadual do Palmito. As capturas foram executadas quinzenalmente, de dezembro 2006 a março 2007, com a técnica de "aspiração menor", nos crepúsculos vespertino e matutino, iniciando antes dos crepúsculos e finalizando após os crepúsculos. Foram detectadas 25 espécies, sendo as mais abundantes, Anopheles cruzii (65,2%), Culex sachettae Sirivanakarn & Jacob, 1982 (11,2%) e Anopheles bellator Dyar & Knab, 1906 (8,5%). De acordo com análise de variância, ocorreu diferença significativa na freqüência entre os períodos crepusculares para seguintes espécies: Aedes scapularis (Rondone, 1848) (p = 0,03651), Coquillettidia chrysonotum (Peryassu, 1922) (p = 0,00795), Mansonia fonsecai (Pinto, 1932) (p = 0,00804), e Runchomyia theobaldi Lane & Cerqueira, 1934 (p = 0,01996). Não houve correlação significativa com a abundância das principais espécies capturadas e as médias dos fatores abióticos avaliados. A taxa de paridade de Anopheles cruzii atingiu média percentual de 48%, não existindo correlação entre a abundância e a taxa de paridade. O crepúsculo exerce influência no comportamento apetente das espécies de Culicidae. A comparação de similaridade entre os crepúsculos apontou para elevada semelhança da composição específica. Anopheles cruzii atingiu dominância nos períodos vespertino e matutino, sendo a diversidade de forma geral reduzida nestes períodos.
INTRODUCTION: The precise identification of the genetic variants of the dengue virus is important to understand its dispersion and virulence patterns and to identify the strains responsible for epidemic outbreaks. This study investigated the genetic variants of the capsid-premembrane junction region fragment in the dengue virus serotypes 1 and 2 (DENV1-2). METHODS: Samples from 11 municipalities in the State of Paraná, Brazil, were provided by the Central Laboratory of Paraná. They were isolated from the cell culture line C6/36 (Aedes albopictus) and were positive for indirect immunofluorescence. Ribonucleic acid (RNA) extracted from these samples was submitted to the reverse transcription polymerase chain reaction (RT-PCR) and nested PCR. RESULTS: RT-PCR revealed that 4 of the samples were co-infected with both serotypes. The isolated DENV-1 sequences were 95-100% similar to the sequences of other serotype 1 strains deposited in GenBank. Similarly, the isolated DENV-2 sequences were 98-100% similar to other serotype 2 sequences in GenBank. According to our neighbor-joining tree, all strains obtained in this study belonged to genotype V of DENV-1. The DENV-2 strains, by contrast, belonged to the American/Asian genotypes. CONCLUSIONS: The monitoring of circulating strains is an important tool to detect the migration of virus subtypes involved in dengue epidemics.
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