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Neurotropic flaviviruses within the Japanese encephalitis (JE) serogroup possess the distinctive feature of expressing a unique nonstructural protein, NS1\u2032. The NS1\u2032 protein consists of the full NS1 protein with an additional 52 amino acid extension at the C-terminus and has been demonstrated to exhibit virulence in mammalian hosts upon infection. However, the precise role of the NS1\u2032 protein in the mosquito vectors has yet to be elucidated.<\/jats:p>\n<\/jats:sec>\n<jats:sec id=\"sec022\">\n<jats:title>Methodology\/principal findings<\/jats:title>\n<jats:p>In this study, an NS1\u2032-defective virus (rG66A) was engineered, and its effect on the infection of mosquito cells was investigated. The results demonstrated a significant reduction in the infectivity of the rG66A virus in mosquito cells by RT-qPCR, indicating that the absence of the NS1\u2032 protein impedes JEV replication in <jats:italic>Culex<\/jats:italic> mosquitoes. Additionally, this research elucidated the underlying mechanism by which the NS1\u2032 protein enhances viral infection in mosquitoes by RNA-Seq analysis. Specifically, the NS1\u2032 protein was found to facilitate infection through the suppression of antimicrobial peptides (AMPs) regulated by the Toll pathway.<\/jats:p>\n<\/jats:sec>\n<jats:sec id=\"sec023\">\n<jats:title>Conclusions\/significance<\/jats:title>\n<jats:p>Our research demonstrated that the JEV NS1\u2032 protein contributes to immune escape, thereby enhancing viral infection in mosquitoes. This finding offers new insights into the transmission mechanisms of JEV, elucidating novel aspects of viral propagation.<\/jats:p>\n<\/jats:sec>","DOI":"10.1371\/journal.pntd.0012823","type":"journal-article","created":{"date-parts":[[2025,1,27]],"date-time":"2025-01-27T18:50:35Z","timestamp":1738003835000},"page":"e0012823","update-policy":"https:\/\/doi.org\/10.1371\/journal.pntd.corrections_policy","source":"Crossref","is-referenced-by-count":5,"title":["The Japanese encephalitis virus NS1\u2032 protein facilitates virus infection in 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