Dataset Information

Functional characterization of an arrestin gene on insecticide resistance of Culex pipiens pallens.

ABSTRACT: Continuous and excessive application of insecticides has resulted in the rapid development of insecticide resistance in several mosquito species, including Culex pipiens pallens. Previous studies in our laboratory found that arrestin gene expression was higher in the deltamethrin-resistant (DR) strain than in the deltamethrin-susceptible (DS) strain of Cx. pipiens pallens. Similarly, other studies reported that arrestin was highly expressed in permethrin-resistant Cx. quinquefasciatus and in dichlorodiphenyltrichloroethane (DDT)-resistant Drosophila melanogaster.Full-length cDNAs of an arrestin gene were cloned from Cx. pipiens pallens via polymerase chain reaction (PCR) and rapid amplification of cDNA end (RACE). The mRNA levels of the arrestin gene in the whole life cycle of DR and DS strains of Cx. pipiens pallens were investigated via quantitative real-time PCR. In addition, the relationship between arrestin and deltamethrin (DM) resistance were identified using genetic overexpression strategies and arrestin RNAi in mosquito cells. Cell viability was analyzed with cholecystokinin octapeptide after DM treatment. Moreover, the mRNA levels of cytochrome P450 6A1 (CYP6A1) and opsin in the transfected cells and controls were analyzed.Complete arrestin gene sequence was cloned and expressed throughout the life cycle of Cx. pipiens pallens. Moreover, arrestin was significantly upregulated in the DR strain, compared with that in the DS strain at the egg, pupae, and adult stages. Arrestin overexpression comparably increased the mosquito cell viability, whereas arrestin knockdown by siRNA decreased mosquito cell viability with deltamethrin (DM) treatment. Meanwhile, the mRNA levels of CYP6A1 and opsin were upregulated in mosquito cells transfected with arrestin and downregulated in mosquito cells with arrestin knockdown.This study presented the first evidence that arrestin might be associated with insecticide resistance in Cx. pipiens pallens.

SUBMITTER: Sun Y

PROVIDER: S-EPMC3425237 | biostudies-literature | 2012 Jul

REPOSITORIES: biostudies-literature

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Functional characterization of an arrestin gene on insecticide resistance of Culex pipiens pallens.

Sun Yan Y Zou Ping P Yu Xin-You XY Chen Chen C Yu Jing J Shi Lin-Na LN Hong Shan-Chao SC Zhou Dan D Chang Xue-Lian XL Wang Wei-Jie WJ Shen Bo B Zhang Dong-Hui DH Ma Lei L Zhu Chang-Liang CL

Parasites & vectors 20120706

<h4>Background</h4>Continuous and excessive application of insecticides has resulted in the rapid development of insecticide resistance in several mosquito species, including Culex pipiens pallens. Previous studies in our laboratory found that arrestin gene expression was higher in the deltamethrin-resistant (DR) strain than in the deltamethrin-susceptible (DS) strain of Cx. pipiens pallens. Similarly, other studies reported that arrestin was highly expressed in permethrin-resistant Cx. quinquef ...[more]

PMID: 22768923

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Project description:Mosquito control based on chemical insecticides is considered as an important element of the current global strategies for the control of mosquito-borne diseases. Unfortunately, the development of insecticide resistance of important vector mosquito species jeopardizes the effectiveness of insecticide-based mosquito control. In contrast to target site resistance, other mechanisms are far from being fully understood. Global protein profiles among cypermethrin-resistant, propoxur-resistant, dimethyl-dichloro-vinyl-phosphate-resistant and susceptible strain of Culex pipiens pallens were obtained and proteomic differences were evaluated by using isobaric tags for relative and absolute quantification labeling coupled with liquid chromatography/tandem mass spectrometric analysis. A susceptible strain of Culex pipiens pallens showed elevated resistance levels after 25 generations of insecticide selection, through iTRAQ data analysis detected 2,502 proteins, of which 1,513 were differentially expressed in insecticide-selected strains compared to the susceptible strain. Finally, midgut differential protein expression profiles were analyzed, and 62 proteins were selected for verification of differential expression using iTRAQ and parallel reaction monitoring strategy, respectively. iTRAQ profiles of adaptation selection to three insecticide strains combined with midgut profiles revealed that multiple insecticide resistance mechanisms operate simultaneously in resistant insects of Culex pipiens pallens. Significant molecular resources were developed for Culex pipiens pallens, potential candidates were involved in metabolic resistance and reducing penetration or sequestering insecticide. Future research that is targeted towards RNA interference of the identified metabolic targets, such as cuticular proteins, cytochrome P450s, glutathione S-transferases and ribosomal proteins proteins and biological pathways (drug metabolism-cytochrome P450, metabolism of xenobiotics by cytochrome P450, oxidative phosphorylation, ribosome) could lay the foundation for a better understanding of the genetic basis of insecticide resistance in Culex pipiens pallens.

Project description:BackgroundCulex pipiens pallens is the most abundant Culex mosquito species in northern China and is an important vector of bancroftian filariasis and, potentially, West Nile virus. Insecticides, particularly pyrethroids, are widely used for adult mosquito control. Insecticide resistance has become common in several mosquito species, and vector control is the main method currently available to prevent disease transmission. The voltage-gated sodium channel (Vgsc) gene is the target site of pyrethroids, and mutations in this gene cause knockdown resistance (kdr).MethodsCulex pipiens pallens larvae were collected from May to November over two decades, from 1992 to 2018, in four cities in Shandong Province, China. The World Health Organization (WHO) standard resistance bioassay was applied to test the resistance levels of Cx. p. pallens larvae to five different insecticides and to test deltamethrin resistance in adults, using the F1 generation. Mutations at Vgsc codon 1014 were also screened in 471 adult samples collected in 2014 to determine the association between kdr mutations and phenotypic resistance.ResultsLarval resistance against deltamethrin showed an increasing trend from the 1990s until 2018, which was statistically significant in all populations; resistance to cypermethrin increased significantly in mosquitoes from the Zaozhuang population. However, larval resistance to other insecticides remained relatively stable. Larval resistance against deltamethrin was consistent with adult bioassays in 2014, in which all tested populations were highly resistant, with mortality rates ranging from 39.4 to 55.23%. The L1014S and L1014F mutations were both observed in five Cx. p. pallens populations, with L1014F significantly associated with deltamethrin resistance.ConclusionsThe long-term dataset from Shandong demonstrates major increases in pyrethroid resistance over a 20-year period. The L1014F kdr mutation may be considered a viable molecular marker for monitoring pyrethroid resistance in Cx. p. pallens.

Project description:BackgroundThe overuse of insecticides to control insect vectors has promoted extensive insecticide resistance in mosquitoes. In this study, the functions of microRNA (miR)-279-3p and its target CYP325BB1 in the regulation of deltamethrin resistance in Culex pipiens pallens was investigated.MethodsQuantitative real-time reverse transcription PCR was used to detect the expression levels of miR-279-3p and CYP325BB1. Then, the dual-luciferase reporter assay system, RNA interference, CDC bottle bioassay and Cell Counting Kit-8 (CCK-8) assay were used to explore the roles of these molecules in deltamethrin resistance both in vivo and in vitro.ResultsThe expression patterns of miR-279-3p and CYP325BB1 were compared between deltamethrin-sensitive (DS-strain) and deltamethrin-resistant (DR-strain) mosquitoes. Luciferase activity was downregulated by miR-279-3p, the effect of which was ablated by a mutation of the putative binding site for CYP325BB1. In DR-strain mosquitoes, the expression of miR-279-3p was increased by microinjection and oral feeding of miR-279-3p agomir (mimic). CYP325BB1 mRNA levels were downregulated, which resulted in a higher mortality of the mosquitoes in miR-279-3p mimic-treated groups. In the DS-strain mosquitoes, microinjection of a miR-279-3p inhibitor decreased miR-279-3p expression, whereas the expression of CYP325BB1 was increased; the mortality of these mosquitoes decreased significantly. In addition, overexpression of pIB/V5-His-CYP325BB1 changed the sensitivity of C6/36 cells to deltamethrin in vitro. Also in DR-strain mosquitoes, downregulation of CYP325BB1 expression by microinjection of si-CYP325BB1 increased mosquito mortality in vivo.ConclusionsThese findings provide empirical evidence of the involvement of miRNAs in the regulation of insecticide resistance and indicate that miR-279-3p suppresses the expression of CYP325BB1, which in turn decreases deltamethrin resistance, resulting in increased mosquito mortality. Taken together, the results provide important information for use in the development of future mosquito control strategies.

Dataset Information

Functional characterization of an arrestin gene on insecticide resistance of Culex pipiens pallens.

Publications

Functional characterization of an arrestin gene on insecticide resistance of Culex pipiens pallens.

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