There are two main limitations for sprayable RNA pesticide development: delivery efficiency and synthetic cost of double-stranded RNA (dsRNA). We previously constructed a nanocarrier-based transdermal dsRNA delivery system and a novel bacteria-based hairpin RNA (hpRNA) expression system to solve these challenges. Herein, as a subsequent exploration of RNA pesticide (sprayable ds/hpRNA for pest control), we performed a greenhouse application of bacteria-expressed and nanocarrier-delivered RNA pesticide on green peach aphid. The nanoparticle SPc could combine and deliver dsRNA across the aphid cuticle and V-type proton ATPase subunits d (ATP-d) and G (ATP-G) were selected as the potential RNA interference (RNAi) targets. Our plasmid-Escherichia coli system simultaneously expressing ATP-d and ATP-G hairpin RNAs (hpRNAs) was constructed for mass production of hpRNA. The expressed hpRNA was mixed with SPc and detergent to form RNA formulation, which showed a certain insecticidal activity through the spray application in the greenhouse. Total control efficacy of our RNA pesticide could reach 61% on 3 d and maintained at 50% until the sixth day. To our knowledge, our study is the first attempt to apply the bacteria-expressed and nanocarrier-delivered RNA pesticides for pest control in the greenhouse trial, which is beneficial for promoting the development of RNA pesticides.

喷雾 RNA 农药的开发存在两个主要限制：双链 RNA (dsRNA) 的递送效率和合成成本。我们之前构建了一个基于纳米载体的透皮 dsRNA 递送系统和一个新的基于细菌的发夹 RNA (hpRNA) 表达系统来解决这些挑战。在此，作为对 RNA 农药（用于害虫防治的可喷洒 ds/hpRNA）的后续探索，我们对绿桃蚜虫进行了细菌表达和纳米载体递送的 RNA 农药的温室应用。纳米颗粒 SPc 可以结合并传递 dsRNA 穿过蚜虫角质层，选择V 型质子 ATP 酶亚基 d (ATP-d)和G (ATP-G)作为潜在的 RNA 干扰 (RNAi) 目标。我们的质粒——大肠杆菌构建了同时表达ATP-d和ATP-G发夹 RNA (hpRNA) 的系统用于大规模生产 hpRNA。将表达的hpRNA与SPc和去污剂混合形成RNA制剂，在温室内喷施后显示出一定的杀虫活性。我们的 RNA 农药的总防治效果在第 3 天可以达到 61%，并保持在 50%，直到第 6 天。据我们所知，我们的研究是首次尝试将细菌表达和纳米载体传递的RNA农药应用于温室试验中的害虫防治，这有利于促进RNA农药的发展。