背景:地中海果蝇(medfly),头状炎Wiedemann,是影响全球水果和蔬菜生产的主要害虫,其控制主要基于杀虫剂。双链RNA(dsRNA)能够下调内源性基因,从而通过RNA干扰(RNAi)影响害虫和病原体的基本生命功能,被设想为传统杀虫剂的更具体和更环保的替代品。然而,这种策略还没有在medfly中探索。
结果:这里,我们通过注射体外转录的成虫基因特异性dsRNA发夹,筛选了7个候选靶基因。几个基因被显著下调,与用靶向绿色荧光蛋白(GFP)cDNA的对照dsRNA处理的苍蝇相比,导致昆虫死亡率增加。三个dsRNA,与ATP合酶的β亚基(ATPsynbeta)同源,液泡ATP酶(V-ATP酶),和核糖体蛋白S13(RPS13),能够在注射后仅48小时内将存活概率减半。然后,我们使用基于两个自身剪接-内含子的表达系统在大肠杆菌中作为环状分子生产了这三种dsRNA和GFP对照的新版本,并测试了它们作为口服递送的杀虫化合物对抗medfly成虫。我们观察到V-ATPase和RPS13mRNA的显着下调(约30%和90%,分别)与治疗三天后的对照组相比。在medfly中没有记录到明显的死亡率,但是通过沉默V-ATPase和RPS13可以实现产卵和孵化减少。
结论:总而言之,我们报道了dsRNA分子作为medfly口服杀虫剂的潜力。本文受版权保护。保留所有权利。
BACKGROUND: The Mediterranean fruit fly (medfly), Ceratitis capitata Wiedemann, is a major pest affecting fruit and vegetable production worldwide, whose control is mainly based on insecticides. Double-stranded RNA (dsRNA) able to down-regulate endogenous genes, thus affecting essential vital functions via RNA interference (RNAi) in pests and pathogens, is envisioned as a more specific and environmentally-friendly alternative to traditional insecticides. However, this strategy has not been explored in medfly yet.
RESULTS: Here, we screened seven candidate target genes by injecting in adult medflies gene-specific dsRNA hairpins transcribed in vitro. Several genes were significantly down-regulated, resulting in increased insect mortality compared to flies treated with a control dsRNA targeting the green fluorescent protein (GFP) complementary DNA (cDNA). Three of the dsRNAs, homologous to the beta subunit of adenosine triphosphate (ATP) synthase (ATPsynbeta), a vacuolar ATPase (V-ATPase), and the ribosomal protein S13 (RPS13), were able to halve the probability of survival in only 48 h after injection. We then produced new versions of these three dsRNAs and that of the GFP control as circular molecules in Escherichia coli using a two-self-splicing-intron-based expression system and tested them as orally-delivered insecticidal compounds against medfly adults. We observed a significant down-regulation of V-ATPase and RPS13 messenger RNAs (mRNAs) (approximately 30% and 90%, respectively) compared with the control medflies after 3 days of treatment. No significant mortality was recorded in medflies, but egg laying and hatching reduction was achieved by silencing V-ATPase and RPS13.
CONCLUSIONS: In sum, we report the potential of dsRNA molecules as oral insecticide in medfly. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.