Spodoptera litura is a destructive agricultural pest and has evolved resistance to multiple insecticides, especially pyrethroids. At present, the resistance mechanism to pyrethroids remains unclear. Four field-collected populations, namely CZ, LF, NJ and JD, were identified to have high resistance to pyrethroids comparing to pyrethroid-susceptible population (GX), with resistant ratio ranging from 11.5- to 9123.5-fold. To characterize pyrethroid resistance mechanism, the transcriptomes between two pyrethroid-resistant (LF and NJ) and a pyrethroid-susceptible (GX) populations were compared by RNA-sequencing. Results showed that multiple differentially expressed genes were enriched in metabolism-related GO terms and KEGG pathways. 35 up-regulated metabolism-related unigenes were selected to verify by qRT-PCR and 15 unigenes, including 4 cytochrome P450s (P450s), 5 glutathione S-transferase (GSTs), 1 UDP-glycosyltransferase (UGT), 4 carboxylesterases (COEs) and 1 and ATP-binding cassette transporters (ABC), were all up-regulated in the four pyrethroid-resistant populations. The expression levels of CYP3 and GST3, which were annotated as CYP6A13 and GSTE1, respectively, showed positive correlation with their pyrethroid resistance levels among the four pyrethroid-resistant populations. While the expression levels of CYP5, CYP12, COE4 and ABC5 showed good correlation with their pyrethroid resistance levels in at least three populations. UGT5 had the highest expression level among the tested UGT genes in the four pyrethroid-resistant populations. RNAi mediated silencing of CYP6 increased the cumulative mortality treated by beta cypermethrin and cyhalothrin significantly, while silencing of GST3 increased the cumulative mortality treated by fenvalerate significantly. CYP3, CYP5, CYP6, CYP12, GST3, COE4, UGT5 and ABC5 play important roles in pyrethroid resistance among the four pyrethroid-resistant populations. Our work provides a valuable clue for further study of pyrethroid resistance mechanisms in S. litura.

中文翻译：

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斜纹夜蛾转录组分析揭示拟除虫菊酯抗性的分子机制

斜纹夜蛾是一种破坏性农业害虫，已进化出对多种杀虫剂，尤其是拟除虫菊酯的抗性。目前，对拟除虫菊酯的耐药机制尚不清楚。与拟除虫菊酯易感种群（GX）相比，野外采集的四个种群，即 CZ、LF、NJ 和 JD 被确定对拟除虫菊酯具有高抗性，抗性比范围为 11.5 至 9123.5 倍。为了表征拟除虫菊酯抗性机制，通过 RNA 测序比较了两个拟除虫菊酯抗性（LF 和 NJ）和拟除虫菊酯易感 (GX) 种群之间的转录组。结果表明，多个差异表达的基因富含代谢相关的 GO 术语和 KEGG 通路。选择35个上调代谢相关的unigenes进行qRT-PCR验证，15个unigenes，包括 4 种细胞色素 P450s (P450s)、5 种谷胱甘肽 S-转移酶 (GSTs)、1 种 UDP-糖基转移酶 (UGT)、4 种羧酸酯酶 (COEs) 和 1 种和 ATP 结合盒式转运蛋白 (ABC)，在四种拟除虫菊酯抗性人群。CYP3和GST3的表达水平，分别标注为CYP6A13和GSTE1，在4个拟除虫菊酯抗性人群中与它们的拟除虫菊酯抗性水平呈正相关。而 CYP5、CYP12、COE4 和 ABC5 的表达水平在至少三个人群中与它们的拟除虫菊酯抗性水平显示出良好的相关性。UGT5 在四个拟除虫菊酯抗性群体中测试的 UGT 基因中具有最高的表达水平。RNAi 介导的 CYP6 沉默显着增加了 β 氯氰菊酯和氯氟氰菊酯治疗的累积死亡率，而 GST3 的沉默显着增加了氰戊菊酯治疗的累积死亡率。CYP3、CYP5、CYP6、CYP12、GST3、COE4、UGT5 和 ABC5 在四个拟除虫菊酯抗性人群中在拟除虫菊酯抗性中起重要作用。我们的工作为进一步研究斜纹夜蛾的拟除虫菊酯抗性机制提供了有价值的线索。