Identification and classification of differentially expressed genes in pyrethroid-resistant Culex pipiens pallens
Culex pipiens pallens is an important vector that transmits Bancroftian filariasis, Japanese encephalitis and other diseases that pose a serious threat to human health. Extensive and improper use of insecticides has caused insecticide resistance in mosquitoes, which has become an important obstacle to the control of mosquito-borne diseases. It is crucial to investigate the underlying mechanism of insecticide resistance. The aims of this study were to identify genes involved in insecticide resistance based on the resistance phenotype, gene expression profile and single-nucleotide polymorphisms (SNPs) and to screen for major genes controlling insecticide resistance. Using a combination of SNP and transcriptome data, gene expression quantitative trait loci (eQTLs) were studied in deltamethrin-resistant mosquitoes. The most differentially expressed pathway in the resistant group was identified, and a regulatory network was built using these SNPs and the differentially expressed genes (DEGs) in this pathway. The major candidate genes involved in the control of insecticide resistance were analyzed by qPCR, siRNA microinjection and CDC bottle bioassays. A total of 85 DEGs that encoded putative detoxification enzymes (including 61 P450s) were identified in this pathway. The resistance regulatory network was built using SNPs, and these metabolic genes, and a major gene, CYP9AL1, were identified. The functional role of CYP9AL1 in insecticide resistance was confirmed by siRNA microinjection and CDC bottle bioassays. Using the eQTL approach, we identified important genes in pyrethroid resistance that may aid in understanding the mechanism underlying insecticide resistance and in targeting new measures for resistance monitoring and management.
KeywordsCulex pipiens pallens Deltamethrin resistance eQTL P450s Regulatory network
We thank Meng-Xue Hu for her assistance with mosquito collection.
BS, C-LZ and NX conceived the study. NX collected the data and performed the experiment. X-HS performed the data analysis. Z-HL performed the supplementary experiment duiring revision. YX, DZ and YS supported the development of the statistical methodology. NX and BS drafted the manuscript. All authors critically reviewed, read and approved the final manuscript.
This study was funded by the National Natural Science Foundation of China (Grant numbers 81471984, 81672056, 81672058 and 81772227), the National Institutes of Health (NIH) of the USA (Grant number 2R01AI075746) and the Priority Academic Program Development of Jiangsu Higher Education Institutions.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflicts of interest.
Research involving human participants and/or animals
This article does not contain any studies with human participants or animals performed by any of the authors.
- Aizoun N, Aikpon R, Padonou GG, Oussou O, Oke-Agbo F, Gnanguenon V, Osse R, Akogbeto M (2013) Mixed-function oxidases and esterases associated with permethrin, deltamethrin and bendiocarb resistance in Anopheles gambiae s.l. in the southnorth transect Benin, West Africa. Parasit Vectors 6:223CrossRefGoogle Scholar
- Bass C, Puinean AM, Andrews M, Cutler P, Daniels M, Elias J, Paul VL, Crossthwaite AJ, Denholm I, Field LM, Foster SP, Lind R, Williamson MS, Slater R (2011) Mutation of a nicotinic acetylcholine receptor b subunit is associated with resistance to neonicotinoid insecticides in the aphid Myzus persicae. BMC Neurosci 12(1):51CrossRefGoogle Scholar
- Canales M, Naranjo V, Almazan C, Molina R, Tsuruta SA, Szabo MP, Manzano-Roman R, Pérez de la Lastra JM, Kocan KM, Jiménez MI, Lucientes J, Villar M, de la Fuente J (2009) Conservation and immunogenicity of the mosquito ortholog of the tick-protective antigen, subolesin. Parasitol Res 105:97–111CrossRefGoogle Scholar
- Chesler EJ, Lu L, Shou S, Qu Y, Gu J, Wang J, Hsu HC, Mountz JD, Baldwin NE, Langston MA, Threadgill DW, Manly KF, Williams RW (2005) Complex trait analysis of gene expression uncovers polygenic and pleiotropic networks that modulate nervous system function. Nat Genet 37(3):233–242CrossRefGoogle Scholar
- Edi CV, Djogbenou L, Jenkins AM, Regna K, Muskavitch MA, Poupardin R, Jones CM, Essandoh J, Kétoh GK, Paine MJ, Koudou BG, Donnelly MJ, Ranson H, Weetman D (2014) CYP6 P450 enzymes and ACE-1 duplication produce extreme and multiple insecticide resistance in the malaria mosquito Anopheles gambiae. PLoS Genet 10:e1004236CrossRefGoogle Scholar
- Faucon F, Gaude T, Dusfour I, Navratil V, Corbel V, Juntarajumnong W, Girod R, Poupardin R, Boyer F, Reynaud S, David JP (2017) In the hunt for genomic markers of metabolic resistance to pyrethroids in the mosquito Aedes aegypti: an integrated next-generation sequencing approach. PLoS Negl Trop Dis 11(4)Google Scholar
- Gething PW, Elyazar IR, Moyes CL, Smith DL, Battle KE, Guerra CA, Patil AP, Tatem AJ, Howes RE, Myers MF, George DB, Horby P, Wertheim HF, Price RN, Müeller I, Baird JK, Hay SI (2012) A long neglected world malaria map: Plasmodium vivax endemicity in 2010. PLoS Negl Trop Dis 6(9):e1814CrossRefGoogle Scholar
- WHO (2013) Test procedures for insecticide resistance monitoring in malaria vector mosquitoes. World Health Organization, GenevaGoogle Scholar
- Zhao L, Zhang N, Ma PF, Liu Q, Li DZ, Guo ZH (2013) Statistical summary of contigs and final transcripts assembled by Trinity and TGICL. Plos One 10:23Google Scholar