Y4667D Mutation in the Ryanodine Receptor Confers High Level Resistance to Diamide Insecticides in the Rice Stem Borer, Chilo suppressalis Walker (Lepidoptera: Crambidae)

Chilo suppressalis is a major rice pest with escalating resistance to diamide insecticides, threatening sustainable management. However, the precise molecular mechanisms underlying this resistance remain unclear. In this study, we assessed the sensitivity of 71 C. suppressalis field populations in China to chlorantraniliprole during 2023–2024 and investigated target-site mutations associated with resistance. The proportion of highly resistant populations increased to 80% in 2023 (RR = 111.6–2706.4) and 90.3% in 2024 (RR = 160–1794.7). Multiple RyR mutations, including Y4667D, were identified in highly resistant populations. Introgressing the Y4667D mutation into a laboratory strain generated the 4667D strain, which exhibited high resistance to chlorantraniliprole and other diamides. Resistance showed autosomal inheritance with incomplete dominance. Modeling and molecular docking revealed that Y4667D reduced CsRyR binding affinity for chlorantraniliprole. Furthermore, Y4667D conferred significant fitness costs such as longer larval duration and reduced reproductive output. These findings provide insights into the molecular mechanisms of diamide resistance, inform pesticide management strategies, and aid the development of novel resistance-breaking pesticides.