{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,4]],"date-time":"2026-07-04T03:30:26Z","timestamp":1783135826283,"version":"3.54.6"},"reference-count":43,"publisher":"MDPI AG","issue":"12","license":[{"start":{"date-parts":[[2024,12,15]],"date-time":"2024-12-15T00:00:00Z","timestamp":1734220800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"National Institutes of Health","award":["U19 AI129326"],"award-info":[{"award-number":["U19 AI129326"]}]},{"name":"National Institutes of Health","award":["D43 TW001505"],"award-info":[{"award-number":["D43 TW001505"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Genes"],"abstract":"<jats:p>Background: The swift expansion of the invasive malaria vector Anopheles stephensi throughout Africa presents a major challenge to malaria control initiatives. Unlike the native African vectors, An. stephensi thrives in urban settings and has developed resistance to multiple classes of insecticides, including pyrethroids, organophosphates, and carbamates. Methods: Insecticide susceptibility tests were performed on field-collected An. stephensi mosquitoes from Awash Sebac Kilo, Ethiopia, to assess insecticide resistance levels. Illumina RNA-seq analysis was then employed to compare the transcriptomes of field-resistant populations and susceptible laboratory strains (STE2). Results: An. stephensi populations exhibited high levels of resistance to both deltamethrin (mortality, 39.4 \u00b1 6.0%) and permethrin (mortality, 59.3 \u00b1 26.3%) in WHO tube bioassays. RNA-seq analysis revealed that both field-resistant and field-unexposed populations exhibited increased expressions of genes associated with pyrethroid resistance, including esterases, P450s, and GSTs, compared to the susceptible STE2 strain. Notably, esterase E4 and venom carboxylesterase-6 were significantly overexpressed, up to 70-fold, compared to the laboratory strain. Functional enrichment analysis revealed a significant overrepresentation of genes associated with catalytic activity under molecular functions and metabolic process under biological process. Using weighted gene co-expression network analysis (WGCNA), we identified two co-expression modules (green and blue) that included 48 genes strongly linked to pyrethroid insecticide resistance. A co-expression network was subsequently built based on the weight values within these modules. Conclusions: This study highlights the role of esterases in the pyrethroid resistance of an An. stephensi population. The identification of candidate genes associated with insecticide resistance will facilitate the development of rapid diagnostic tools to monitor resistance trends.<\/jats:p>","DOI":"10.3390\/genes15121603","type":"journal-article","created":{"date-parts":[[2024,12,16]],"date-time":"2024-12-16T06:48:39Z","timestamp":1734331719000},"page":"1603","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":10,"title":["Esterase-Mediated Pyrethroid Resistance in Populations of an Invasive Malaria Vector Anopheles stephensi from Ethiopia"],"prefix":"10.3390","volume":"15","author":[{"given":"Daibin","family":"Zhong","sequence":"first","affiliation":[{"name":"Joe C. Wen School of Population & Public Health, University of California at Irvine, Irvine, CA 92697, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3518-2372","authenticated-orcid":false,"given":"Teshome","family":"Degefa","sequence":"additional","affiliation":[{"name":"School of Medical Laboratory Sciences, Institute of Health, Jimma University, Jimma MVJ4+R95, Ethiopia"},{"name":"Tropical and Infectious Diseases Research Centre (TIDRC), Jimma University, Jimma MVJ4+R95, Ethiopia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9283-5520","authenticated-orcid":false,"given":"Guofa","family":"Zhou","sequence":"additional","affiliation":[{"name":"Joe C. Wen School of Population & Public Health, University of California at Irvine, Irvine, CA 92697, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ming-Chieh","family":"Lee","sequence":"additional","affiliation":[{"name":"Joe C. Wen School of Population & Public Health, University of California at Irvine, Irvine, CA 92697, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1232-3125","authenticated-orcid":false,"given":"Chloe","family":"Wang","sequence":"additional","affiliation":[{"name":"Joe C. Wen School of Population & Public Health, University of California at Irvine, Irvine, CA 92697, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jiale","family":"Chen","sequence":"additional","affiliation":[{"name":"Joe C. Wen School of Population & Public Health, University of California at Irvine, Irvine, CA 92697, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Delenasaw","family":"Yewhalaw","sequence":"additional","affiliation":[{"name":"School of Medical Laboratory Sciences, Institute of Health, Jimma University, Jimma MVJ4+R95, Ethiopia"},{"name":"Tropical and Infectious Diseases Research Centre (TIDRC), Jimma University, Jimma MVJ4+R95, Ethiopia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Guiyun","family":"Yan","sequence":"additional","affiliation":[{"name":"Joe C. Wen School of Population & Public Health, University of California at Irvine, Irvine, CA 92697, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2024,12,15]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"2498","DOI":"10.3201\/eid2912.230637","article-title":"Detection of Anopheles stephensi mosquitoes by molecular surveillance, Kenya","volume":"29","author":"Ochomo","year":"2023","journal-title":"Emerg. Infect. Dis."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"102","DOI":"10.1016\/j.pt.2023.11.011","article-title":"Anopheles stephensi ecology and control in Africa","volume":"40","author":"Zhou","year":"2024","journal-title":"Trends Parasitol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"218","DOI":"10.1186\/s12936-023-04649-5","article-title":"Resistance of Anopheles stephensi to selected insecticides used for indoor residual spraying and long-lasting insecticidal nets in Ethiopia","volume":"22","author":"Teshome","year":"2023","journal-title":"Malar. J."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"731","DOI":"10.1016\/j.pt.2024.06.008","article-title":"Invasive Anopheles stephensi in Africa: Insights from Asia","volume":"40","author":"Taylor","year":"2024","journal-title":"Trends Parasitol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"43","DOI":"10.4103\/1995-7645.250344","article-title":"Resistance status of main malaria vector, Anopheles stephensi Liston (Diptera: Culicidae) to insecticides in a malaria Endemic Area, Southern Iran","volume":"12","author":"Abbasi","year":"2019","journal-title":"Asian Pac. J. Trop. Med."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"208","DOI":"10.1186\/1475-2875-10-208","article-title":"Combining indoor residual spraying and insecticide-treated nets for malaria control in Africa: A review of possible outcomes and an outline of suggestions for the future","volume":"10","author":"Okumu","year":"2011","journal-title":"Malaria J."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1227","DOI":"10.1016\/S0140-6736(21)02499-5","article-title":"Effectiveness and cost-effectiveness against malaria of three types of dual-active-ingredient long-lasting insecticidal nets (LLINs) compared with pyrethroid-only LLINs in Tanzania: A four-arm, cluster-randomised trial","volume":"399","author":"Mosha","year":"2022","journal-title":"Lancet"},{"key":"ref_8","unstructured":"WHO (2016). Test Procedures for Insecticide Resistance Monitoring in Malaria Vector Mosquitoes, World Health Organization."},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"Muturi, E.J., Dunlap, C., Smartt, C.T., and Shin, D. (2021). Resistance to permethrin alters the gut microbiota of Aedes aegypti. Sci. Rep., 11.","DOI":"10.1038\/s41598-021-93725-4"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"537","DOI":"10.1146\/annurev-ento-010814-020828","article-title":"Insecticide resistance in mosquitoes: Impact, mechanisms, and research directions","volume":"60","author":"Liu","year":"2015","journal-title":"Annu. Rev. Entomol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"105235","DOI":"10.1016\/j.meegid.2022.105235","article-title":"Detection and population genetic analysis of kdr L1014F variant in eastern Ethiopian Anopheles stephensi","volume":"99","author":"Samake","year":"2022","journal-title":"Infect. Genet. Evol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"632","DOI":"10.4269\/ajtmh.20-1357","article-title":"Analysis of the knockdown resistance locus (kdr) in Anopheles stephensi, An. arabiensis, and Culex pipiens sl for insight Into the evolution of target-site pyrethroid resistance in eastern Ethiopia","volume":"106","author":"Carter","year":"2022","journal-title":"Am. J. Trop. Med. Hyg."},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Acford-Palmer, H., Phelan, J.E., Tadesse, F.G., Kristan, M., Collins, E., Spadar, A., Walker, T., Bousema, T., Messenger, L.A., and Clark, T.G. (2023). Identification of two insecticide resistance markers in Ethiopian Anopheles stephensi mosquitoes using a multiplex amplicon sequencing assay. Sci. Rep., 13.","DOI":"10.1038\/s41598-023-32336-7"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"59","DOI":"10.1186\/1475-2875-10-59","article-title":"Knockdown resistance (kdr)-like mutations in the voltage-gated sodium channel of a malaria vector Anopheles stephensi and PCR assays for their detection","volume":"10","author":"Singh","year":"2011","journal-title":"Malaria J."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"315","DOI":"10.1093\/jme\/tjv242","article-title":"Knockdown resistance (kdr) mutations in Indian Anopheles stephensi (Diptera: Culicidae) populations","volume":"53","author":"Dykes","year":"2016","journal-title":"J. Med. Entomol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"180","DOI":"10.1186\/s12936-020-03252-2","article-title":"Insecticide resistance in Anopheles stephensi in Somali Region, eastern Ethiopia","volume":"19","author":"Yared","year":"2020","journal-title":"Malar. J."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"315","DOI":"10.1111\/j.1365-2583.2007.00728.x","article-title":"Transcriptional analysis of insecticide resistance in Anopheles stephensi using cross-species microarray hybridization","volume":"16","author":"Vontas","year":"2007","journal-title":"Insect Mol. Biol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"349","DOI":"10.1186\/1756-3305-7-349","article-title":"ABC transporters are involved in defense against permethrin insecticide in the malaria vector Anopheles stephensi","volume":"7","author":"Epis","year":"2014","journal-title":"Parasit. Vectors"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"37","DOI":"10.1016\/j.actatropica.2017.03.012","article-title":"Gene expression modulation of ABC transporter genes in response to permethrin in adults of the mosquito malaria vector Anopheles stephensi","volume":"171","author":"Mastrantonio","year":"2017","journal-title":"Acta Trop."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"97","DOI":"10.4103\/0972-9062.134808","article-title":"Differential expression of glutathione s-transferase enzyme in different life stages of various insecticide-resistant strains of Anopheles stephensi: A malaria vector","volume":"51","author":"Sanil","year":"2014","journal-title":"J. Vector Borne Dis."},{"key":"ref_21","doi-asserted-by":"crossref","unstructured":"De Marco, L., Sassera, D., Epis, S., Mastrantonio, V., Ferrari, M., Ricci, I., Comandatore, F., Bandi, C., Porretta, D., and Urbanelli, S. (2017). The choreography of the chemical defensome response to insecticide stress: Insights into the Anopheles stephensi transcriptome using RNA-Seq. Sci. Rep., 7.","DOI":"10.1038\/srep41312"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"70","DOI":"10.1186\/s12936-020-3144-9","article-title":"Key to the females of Afrotropical Anopheles mosquitoes (Diptera: Culicidae)","volume":"19","author":"Coetzee","year":"2020","journal-title":"Malaria J."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"23","DOI":"10.1186\/1475-2875-6-23","article-title":"Identification of field caught Anopheles gambiae s.s. and Anopheles arabiensis by TaqMan single nucleotide polymorphism genotyping","volume":"6","author":"Walker","year":"2007","journal-title":"Malar. J."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"36","DOI":"10.3201\/eid2901.220786","article-title":"Molecular tools for early detection of invasive malaria vector Anopheles stephensi mosquitoes","volume":"29","author":"Singh","year":"2023","journal-title":"Emerg. Infect. Dis."},{"key":"ref_25","doi-asserted-by":"crossref","unstructured":"Zhong, D., Hemming-Schroeder, E., Wang, X., Kibret, S., Zhou, G., Atieli, H., Lee, M.C., Afrane, Y.A., Githeko, A.K., and Yan, G. (2020). Extensive new Anopheles cryptic species involved in human malaria transmission in western Kenya. Sci. Rep., 10.","DOI":"10.1038\/s41598-020-73073-5"},{"key":"ref_26","unstructured":"Andrews, S. (2010). FastQC Version 0.12.0: A Quality Control Tool for High Throughput Sequence Data, Babraham Institute."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"2114","DOI":"10.1093\/bioinformatics\/btu170","article-title":"Trimmomatic: A flexible trimmer for Illumina sequence data","volume":"30","author":"Bolger","year":"2014","journal-title":"Bioinformatics"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"139","DOI":"10.1093\/bioinformatics\/btp616","article-title":"edgeR: A Bioconductor package for differential expression analysis of digital gene expression data","volume":"26","author":"Robinson","year":"2009","journal-title":"Bioinformatics"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"289","DOI":"10.1111\/j.2517-6161.1995.tb02031.x","article-title":"Controlling the false discovery rate: A practical and powerful approach to multiple testing","volume":"57","author":"Benjamini","year":"1995","journal-title":"J. R. Stat. Soc. Series B Stat. Methodol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"2628","DOI":"10.1093\/bioinformatics\/btz931","article-title":"ShinyGO: A graphical gene-set enrichment tool for animals and plants","volume":"36","author":"Ge","year":"2020","journal-title":"Bioinformatics"},{"key":"ref_31","doi-asserted-by":"crossref","unstructured":"Langfelder, P., and Horvath, S. (2008). WGCNA: An R package for weighted correlation network analysis. BMC Bioinform., 9.","DOI":"10.1186\/1471-2105-9-559"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"2498","DOI":"10.1101\/gr.1239303","article-title":"Cytoscape: A software environment for integrated models of biomolecular interaction networks","volume":"13","author":"Shannon","year":"2003","journal-title":"Genome Res."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"4288","DOI":"10.1093\/nar\/gks042","article-title":"Differential expression analysis of multifactor RNA-Seq experiments with respect to biological variation","volume":"40","author":"McCarthy","year":"2012","journal-title":"Nucleic Acids Res."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"169","DOI":"10.1042\/bj3300169","article-title":"Evidence that the E4 and FE4 esterase genes responsible for insecticide resistance in the aphid Myzus persicae (Sulzer) are part of a gene family","volume":"330","author":"Field","year":"1998","journal-title":"Biochem. J."},{"key":"ref_35","doi-asserted-by":"crossref","unstructured":"Wang, H.L., Rao, Q., and Chen, Z.Z. (2023). Identifying potential insecticide resistance markers through genomic-level comparison of Bemisia tabaci (Gennadius) lines. Arch. Insect Biochem. Physiol., 114.","DOI":"10.1002\/arch.22034"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"36","DOI":"10.1111\/1748-5967.12489","article-title":"Identification of candidate odorant-degrading enzyme genes in the antennal transcriptome of Aphidius gifuensis","volume":"51","author":"Kang","year":"2021","journal-title":"Entomol. Res."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"621","DOI":"10.1038\/nmeth.1226","article-title":"Mapping and quantifying mammalian transcriptomes by RNA-Seq","volume":"5","author":"Mortazavi","year":"2008","journal-title":"Nat. Methods"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"956","DOI":"10.1126\/science.1160342","article-title":"A global view of gene activity and alternative splicing by deep sequencing of the human transcriptome","volume":"321","author":"Sultan","year":"2008","journal-title":"Science"},{"key":"ref_39","doi-asserted-by":"crossref","unstructured":"Raz, T., Kapranov, P., Lipson, D., Letovsky, S., Milos, P.M., and Thompson, J.F. (2011). Protocol dependence of sequencing-based gene expression measurements. PLoS ONE, 6.","DOI":"10.1371\/journal.pone.0019287"},{"key":"ref_40","doi-asserted-by":"crossref","unstructured":"Mane, S.P., Evans, C., Cooper, K.L., Crasta, O.R., Folkerts, O., Hutchison, S.K., Harkins, T.T., Thierry-Mieg, D., Thierry-Mieg, J., and Jensen, R.V. (2009). Transcriptome sequencing of the Microarray Quality Control (MAQC) RNA reference samples using next generation sequencing. BMC Genom., 10.","DOI":"10.1186\/1471-2164-10-264"},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"259","DOI":"10.1016\/j.ygeno.2010.07.010","article-title":"A comparison between ribo-minus RNA-sequencing and polyA-selected RNA-sequencing","volume":"96","author":"Cui","year":"2010","journal-title":"Genomics"},{"key":"ref_42","doi-asserted-by":"crossref","unstructured":"Takele Assefa, A., Vandesompele, J., and Thas, O. (2020). On the utility of RNA sample pooling to optimize cost and statistical power in RNA sequencing experiments. BMC Genom., 21.","DOI":"10.1186\/s12864-020-6721-y"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"862","DOI":"10.1038\/nrg930","article-title":"DNA pooling: A tool for large-scale association studies","volume":"3","author":"Sham","year":"2002","journal-title":"Nat. Rev. Genet."}],"container-title":["Genes"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4425\/15\/12\/1603\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T16:55:38Z","timestamp":1760115338000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4425\/15\/12\/1603"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,12,15]]},"references-count":43,"journal-issue":{"issue":"12","published-online":{"date-parts":[[2024,12]]}},"alternative-id":["genes15121603"],"URL":"https:\/\/doi.org\/10.3390\/genes15121603","relation":{},"ISSN":["2073-4425"],"issn-type":[{"value":"2073-4425","type":"electronic"}],"subject":[],"published":{"date-parts":[[2024,12,15]]}}}