Instituto de Microbiología y Zoología Agrícola (IMyZA), Centro de investigaciones en Ciencias Agronómicas y Veterinarias (CICVyA), Instituto Nacional de Tecnología Agropecuaria (INTA), Buenos Aires, Argentina.
Instituto de Agrobiotecnología y Biología Molecular (IABIMO), Instituto Nacional de Tecnología Agropecuaria (INTA), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET) and Lab de Agrobiotecnología DFBMC, Facultad de Ciencias Exactas y Naturales, Univ de Buenos Aires, Buenos Aires, Argentina.
Neotrop Entomol. 2021 Feb;50(1):121-128. doi: 10.1007/s13744-020-00819-1. Epub 2020 Oct 6.
The "cotton boll weevil" (Anthonomus grandis Boheman) is a key pest in America whose larval stage develops within the cotton flower bud. During its development, the larva uses the flower bud as food and as a shelter from predators. This behavior limits the effective control through conventional insecticide applications and biocontrol techniques. Increasing genetic information from insects has allowed the development of new control technologies based on the use of RNA interference (RNAi) to design orally delivered double-stranded RNA (dsRNA) strategies. In this study, we evaluated the effect of continuous oral administration of six specific dsRNA in order to identify an effective target gene for RNAi-mediated control of cotton boll weevil. First, six selected A. grandis gene fragments were amplified and cloned to perform in vivo synthesis of the specific dsRNA, and subsequently, larvae and adults were fed with this dsRNA for 2 weeks. Larvae mortality ranged from 40 to 60% depending on the targeted gene sequence. Indeed, α-amylase and cytochrome p450 dsRNAs were the most effective. Oral administration in adults caused smaller but still significant death rates (15-30%). Thus, the results demonstrated RNAi responses depend on life stages and target genes. The dsRNA ingestion was capable of providing knockdown mRNA levels in cotton boll weevil midgut and this effect was significantly higher in the larval stage. In this study, we present a new report of silencing of midgut genes in A. grandis larva induced by continuously feeding with dsRNA. This potential new tool should be further evaluated in cotton boll weevil control strategies.
棉铃象鼻虫（Anthonomus grandis Boheman）是美洲的一种重要害虫，其幼虫阶段在棉花花蕾内发育。在其发育过程中，幼虫以花蕾为食，并将其作为躲避捕食者的庇护所。这种行为限制了通过常规杀虫剂应用和生物防治技术进行有效控制。昆虫的遗传信息不断增加，使得基于 RNA 干扰（RNAi）的新技术得以发展，用于设计口服双链 RNA（dsRNA）策略。在这项研究中，我们评估了连续口服六种特定 dsRNA 的效果，以确定用于 RNAi 介导的棉铃象鼻虫控制的有效靶基因。首先，扩增和克隆了六个选定的 A. grandis 基因片段，以进行特定 dsRNA 的体内合成，随后用该 dsRNA 喂养幼虫和成虫 2 周。幼虫死亡率因靶基因序列而异，范围为 40%至 60%。实际上，α-淀粉酶和细胞色素 p450 dsRNA 的效果最显著。在成虫中的口服给药虽然死亡率较小，但仍具有显著的效果（15-30%）。因此，结果表明 RNAi 反应取决于生命阶段和靶基因。dsRNA 的摄入能够在棉铃象鼻虫的中肠中提供靶基因 mRNA 的敲低水平，在幼虫阶段效果更为显著。在这项研究中，我们报告了通过持续喂养 dsRNA 可在 A. grandis 幼虫中沉默中肠基因，这是一种新的潜在工具，应该在棉铃象鼻虫的控制策略中进一步评估。
