{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,30]],"date-time":"2026-09-30T09:38:50Z","timestamp":1790761130228,"version":"4.1.0"},"reference-count":94,"publisher":"MDPI AG","issue":"10","license":[{"start":{"date-parts":[[2024,5,10]],"date-time":"2024-05-10T00:00:00Z","timestamp":1715299200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001655","name":"German Academic Exchange Service","doi-asserted-by":"publisher","award":["57507871"],"award-info":[{"award-number":["57507871"]}],"id":[{"id":"10.13039\/501100001655","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>In this study, we report the complexities and challenges associated with achieving robust RNA interference (RNAi)-mediated gene knockdown in the mosquitoes Aedes\u00a0aegypti and Aedes albopictus, a pivotal approach for genetic analysis and vector control. Despite RNAi\u2019s potential for species-specific gene targeting, our independent efforts to establish oral delivery of RNAi for identifying genes critical for mosquito development and fitness encountered significant challenges, failing to reproduce previously reported potent RNAi effects. We independently evaluated a range of RNAi-inducing molecules (siRNAs, shRNAs, and dsRNAs) and administration methods (oral delivery, immersion, and microinjection) in three different laboratories. We also tested various mosquito strains and utilized microorganisms for RNA delivery. Our results reveal a pronounced inconsistency in RNAi efficacy, characterized by minimal effects on larval survival and gene expression levels in most instances despite strong published effects for the tested targets. One or multiple factors, including RNase activity in the gut, the cellular internalization and processing of RNA molecules, and the systemic dissemination of the RNAi signal, could be involved in this variability, all of which are barely understood in mosquitoes. The challenges identified in this study highlight the necessity for additional research into the underlying mechanisms of mosquito RNAi to develop more robust RNAi-based methodologies. Our findings emphasize the intricacies of RNAi application in mosquitoes, which present a substantial barrier to its utilization in genetic control strategies.<\/jats:p>","DOI":"10.3390\/ijms25105218","type":"journal-article","created":{"date-parts":[[2024,5,14]],"date-time":"2024-05-14T08:03:08Z","timestamp":1715673788000},"page":"5218","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":18,"title":["Challenges of Robust RNAi-Mediated Gene Silencing in Aedes Mosquitoes"],"prefix":"10.3390","volume":"25","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-0739-5597","authenticated-orcid":false,"given":"Lucas Henrique","family":"Figueiredo Prates","sequence":"first","affiliation":[{"name":"Department of Insect Biotechnology in Plant Protection, Justus Liebig University Giessen, 35394 Giessen, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jakob","family":"Fiebig","sequence":"additional","affiliation":[{"name":"Department of Insect Biotechnology in Plant Protection, Justus Liebig University Giessen, 35394 Giessen, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0009-1217-0692","authenticated-orcid":false,"given":"Henrik","family":"Schlosser","sequence":"additional","affiliation":[{"name":"Department of Insect Biotechnology in Plant Protection, Justus Liebig University Giessen, 35394 Giessen, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0000-7175-8618","authenticated-orcid":false,"given":"Eleni","family":"Liapi","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Biotechnology, University of Thessaly, 41500 Larissa, Greece"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4173-9816","authenticated-orcid":false,"given":"Tanja","family":"Rehling","sequence":"additional","affiliation":[{"name":"Department of Insect Biotechnology in Plant Protection, Justus Liebig University Giessen, 35394 Giessen, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3440-8939","authenticated-orcid":false,"given":"C\u00e9lia","family":"Lutrat","sequence":"additional","affiliation":[{"name":"ASTRE, CIRAD, 34398 Montpellier, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1913-416X","authenticated-orcid":false,"given":"Jeremy","family":"Bouyer","sequence":"additional","affiliation":[{"name":"ASTRE, CIRAD, 34398 Montpellier, France"},{"name":"ASTRE, CIRAD, INRAE, Univ. Montpellier, Plateforme Technologique CYROI, 97491 Sainte-Clotilde, La R\u00e9union, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Qiang","family":"Sun","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, MI 48824, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1820-3220","authenticated-orcid":false,"given":"Han","family":"Wen","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, MI 48824, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7786-012X","authenticated-orcid":false,"given":"Zhiyong","family":"Xi","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, MI 48824, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9217-394X","authenticated-orcid":false,"given":"Marc F.","family":"Schetelig","sequence":"additional","affiliation":[{"name":"Department of Insect Biotechnology in Plant Protection, Justus Liebig University Giessen, 35394 Giessen, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4022-4853","authenticated-orcid":false,"given":"Irina","family":"H\u00e4cker","sequence":"additional","affiliation":[{"name":"Department of Insect Biotechnology in Plant Protection, Justus Liebig University Giessen, 35394 Giessen, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2024,5,10]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"S68","DOI":"10.1016\/j.jip.2012.07.012","article-title":"RNAi: Future in insect management","volume":"112","author":"Burand","year":"2013","journal-title":"J. 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