Abstract：
Insect pest control can be achieved by the application of RNA interference (RNAi), a key molecular tool in functional genomics. Whereas most RNAi research has focused on insect pests, few studies have been performed on natural enemies. Validating the efficacy of RNAi in natural enemies is crucial for assessing its safety and enabling molecular research on these organisms. Here, we assessed the efficacy of RNAi in the ladybird beetle Eriopis connexa Germar (Coleoptera: Coccinellidae), focusing on genes related to reproduction, such as vitellogenin (Vg) and its receptor (VgR). In the transcriptome of E. connexa, we found one VgR (EcVgR) and two Vg genes (EcVg1 and EcVg2). These genes have been validated by in silico analyses of functional domains and evolutionary relationships. Five-day-old females were injected with 500 ng/µL of a specific double-stranded RNA (dsRNA) (dsEcVg1, dsEcVg2, or dsEcVgR) for RNAi tests, while nonspecific dsRNA (dsGFP or dsAgCE8.1) was used as a control. Interestingly, dsEcVg2 was able to knockdown both Vg genes, while dsEcVg1 could silence only EcVg1. Additionally, the viability of the eggs was significantly reduced when both Vg genes were knocked down at the same time (after treatment with dsEcVg2 or \"dsEcVg1+dsEcVg2\"). Ultimately, malformed, nonviable eggs were produced when EcVgR was silenced. Interestingly, no dsRNA treatment had an impact on the quantity of eggs laid. Therefore, the feasibility of RNAi in E. connexa has been confirmed, suggesting that this coccinellid is an excellent Neotropical model for molecular research on natural enemies and for studying RNAi nontarget effects.
摘要：
通过RNA干扰（RNAi）的应用可以实现害虫的控制，功能基因组学的关键分子工具。尽管大多数RNAi研究都集中在害虫上，对天敌的研究很少。验证RNAi在天敌中的功效对于评估其安全性和对这些生物体进行分子研究至关重要。这里,我们评估了RNAi在瓢虫EriopisconnexaGermar（鞘翅目：球藻科）中的功效，关注与生殖相关的基因，如卵黄蛋白原(Vg)及其受体(VgR)。在E.connexa的转录组中，我们发现了一个VgR（EcVgR）和两个Vg基因（EcVg1和EcVg2）。这些基因已通过功能域和进化关系的计算机模拟分析得到验证。五天大的雌性被注射500ng/µL的特定双链RNA（dsRNA）（dsEcVg1，dsEcVg2或dsEcVgR）用于RNAi测试，而非特异性dsRNA(dsGFP或dsAgCE8.1)用作对照。有趣的是,dsEcVg2能够敲除两个Vg基因，而dsEcVg1只能使EcVg1静默。此外,当同时敲除两个Vg基因时（用dsEcVg2或“dsEcVg1dsEcVg2”处理后），卵的活力显着降低。最终,畸形,当EcVgR沉默时，产生了无法存活的卵。有趣的是,没有dsRNA处理对产卵的数量有影响。因此,RNAi在E.connexa中的可行性已经得到证实，表明这种球虫是天敌分子研究和研究RNAi非靶标作用的极好的新热带模型。
