Abstract

RNA interference (RNAi)-based biopesticides have emerged as a revolutionary tool for sustainable agriculture. However, their efficacy is hampered by the instability of RNAs and multilayered barriers in pests, and particularly RNAi biopesticides are most unsusceptible to lepidopterans pest. Although the use of nanocarriers has enhanced their stability and RNAi efficiency, some inherent drawbacks remain, such as formulation complexity and the uncertainty of nanocarriers. Herein, we utilize the single-stranded RNA self-assembly technique to construct three-dimensional (3D) tetrahedral-shaped RNA origami, an RNA biopesticide with potential for sprayable formulation，termed T-RiOBs. The T-RiOBs display superior foliar wettability, efficient absorption in plants. Notably, excellent penetration performances of T-RiOB are observed in the midguts and body walls of the larvae of Aedes albopictus and lepidopteran pest, Spodoptera exigua ( S. exigua ), indicating T-RiOB not only exhibits enhanced stability in the intestine but promotes body accumulation. These findings strongly suggest that T-RiOB shows potential for field spraying applications. Most importantly, without the assistance of any additional carrier, T-RiOB demonstrates control potential against S. exigua , by effectively silencing the vacuolar-type ATPase ( V-ATPase ) gene, significantly inhibiting larval growth, and markedly increasing mortality. This technique presents a universal, and applicable platform for advancing RNA biopesticides in crop protection.