Whole-genome sequencing reveals novel ryanodine receptor mutations P4568L and Y4701D conferring high-level chlorantraniliprole resistance in Spodoptera litura (Lepidoptera: Noctuidae).

BACKGROUND Spodoptera litura is a significant insect pest that attacks many agricultural crops, including horticultural, cotton, and legume crops. It has developed resistance to various insecticides, including chlorantraniliprole, posing significant challenges for pest management. Recently, a field-collected population of S. litura (BM28x) from northwestern Australia was found to be more than 1000-fold less susceptible to chlorantraniliprole than a laboratory susceptible strain. Therefore, understanding the mechanism of this insecticide resistance and developing DNA-based molecular diagnostic assays are critical for developing and implementing sustainable pest control strategies for S. litura. RESULTS We mapped the chlorantraniliprole resistance gene to chromosome 26 using whole-genome sequencing and linkage analysis. A combination of the novel mutations P4568L and Y4701D in the ryanodine receptor (RyR) gene was shown to cause high-level resistance to chlorantraniliprole in the BM28x strain. A molecular diagnostic assay was developed to monitor these resistance mutations in field populations. Mutations P4568L and Y4701D were widely distributed in field populations across Western Australia and the Northern Territory. CONCLUSION This research elucidates the genetic mechanisms underlying chlorantraniliprole resistance in S. litura and provides the first evidence of the Y4701D mutation conferring diamide resistance in Noctuidae. These findings enable the development of rapid molecular diagnostic assays which are vital for proactive resistance management. Given that the resistance is recessive in nature it can remain undetected by simple conventional bioassays until it becomes widespread. The findings significantly advance the field of resistance management by facilitating timely interventions to prevent resistance development, thereby supporting more effective integrated pest management. © 2025 The Author(s). Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.