Delivery of Methoprene-Tolerant dsRNA to Improve RNAi Efficiency by Modified Liposomes for Pest Control

Abstract

The application of RNA interference (RNAi) technology for pest control is environmentally friendly and accurate. However, the efficiency of RNAi is often inconsistent and unreliable, and finding a suitable carrier element is considered critical to success in overcoming biotic and abiotic barriers to reach the target site. The fall armyworm, Spodoptera frugiperda (FAW), which is one of most important global agricultural pests, has recently spread rapidly to other parts of the world. In this study, a method to improve the stability and RNAi efficiency of the dsRNA carrier complex was reported. Methoprene-tolerant gene (Met) was selected as a target, a gene which is critical to the growth and development of FAW. Biomaterials nanoliposomes (LNPs) were modified with polyethylenimine (PEI) to deliver the dsRNA of Met. The synthesized Met3@PEI@LNPs reached a size of 385 nm and were found to load dsRNA effectively. Through stability and protection assays, it was found that LNPs provided reliable protection. In addition, the release curve also demonstrated that LNPs were able to prevent premature release under alkaline condition of the insect midgut but accelerate the release after entering the acidic environment of the target cells. The cell transfection efficiency of the prepared LNPs reached 96.4%. Toxicity tests showed that the use of LNPs could significantly improve the interference efficiency, with 91.7% interference efficiency achieved when the concentration of dsRNA in LNPs was only 25% of that of the control. Successful interference of Met demonstrated it could significantly shorten the larval period and make the larvae pupate earlier, thus achieving the purpose of control. In this study, we have demonstrated the use of nanotechnology to provide a novel RNAi delivery method for pest control.

The detoxification role of P450 genes implicated in adaptation of Helicoverpa armigera to walnut phytochemistry

“…The qPCR validation was performed using AceQ® qPCR SYBR® Green Master Mix kit (Vazyme Biotech, Nanjing, China) following the manufacturer's protocols (Huang et al 2023). The qPCR ampli cation conditions were 95°C for 5 min, followed by 40 cycles of 95°C for 15 s and 60°C for 30 s. The actin gene of cotton bollworm was used as a candidate reference gene (Su et al 2023). All primers used for qPCR are listed in Table S2.…”

Insecticidal activity and underlying molecular mechanisms of a phytochemical plumbagin against Spodoptera frugiperda

“…Eggs of S. frugiperda used in this study were supplied by the Institute of Plant Protection, Chinese Academy of Agricultural Sciences, which were collected from maize fields in Jiangcheng, Yunnan Province, China. The larvae were reared on artificial diet in a growth room at 26 °C ± 1 °C, 70% ± 5% relative humidity, with a photoperiod of 14:10 (L:D) as described previously (Su et al, 2023).…”

“…The qPCR validation was carried out based on SuperReal PreMix Plus (SYBR Green) reagent kit (TianGen, Beijing, China), according to the manufacturer's instructions (Huang et al, 2018) and using the same samples used for RNA-seq. The qPCR programme was 95 °C for 10 min, followed by 40 cycles of 95 °C for 15 s and 60 °C for 30 s. The gene β-actin was used as a reference gene as described previously (Su et al, 2023). The primers for qPCR were listed in Supplementary Table S1.…”

Transcript Level and Sequence Matching Are Key Determinants of Off-Target Effects in RNAi

“…10 Therefore, dsRNA nucleic acid pesticides are considered as the most promising fourth-generation pesticides offering effective solutions for common issues associated with traditional pesticides, such as chemical pesticide residue pollution, pest resistance, and pest resurgence. 11,12 The research and application of dsRNA pesticides originated from the discovery of RNAi. 2 In the research of insects, scientists have carried out extensive and in-depth studies in a variety of insects using RNAi technology.…”

“…Double-stranded RNA (dsRNA) pesticides can be used to control agricultural pests. , The mechanism of action involves the utilization of specific dsRNA or small interfering RNA (siRNA) to recognize mRNA of the target gene through gene base complementarity, recruit the RNA-induced silencing complex, and cause post-transcriptional silencing of target genes in pest insects. , dsRNAs could disrupt the normal growth and development of pests resulting in lethal effects, ultimately achieving the goal of pest control and plant protection. , dsRNA pesticides possess numerous advantages including target specificity, high efficiency, broad applicability, low development cost, convenient resistance management, and relative safety for higher animals . Therefore, dsRNA nucleic acid pesticides are considered as the most promising fourth-generation pesticides offering effective solutions for common issues associated with traditional pesticides, such as chemical pesticide residue pollution, pest resistance, and pest resurgence. , …”