Functional Characterization of Trypsin Genes in Spodoptera litura and Their RNA Silencing in Combination with Insecticides

Trypsin is one of the major digestive proteinases secreted by the insect midgut. It plays important roles in digestion, molting, diapause, zymogen activation, innate immunity, reproduction and insecticide resistance. However, the functions of trypsin and its potential as a target for the control of Spodoptera litura, a polyphagous agricultural pest, remain poorly understood. In this study, seven trypsin genes related to S. litura development were characterized. Temporal-spatial expression results showed that the seven trypsin genes were predominantly expressed during the larval stage and mainly localized in the gut. Their expression was positively correlated with juvenile hormone titer. Oral RNAi using star polycation (SPc) as a nanocarrier showed that silencing Sltrypsin1, Sltrypsin5, and Sltrypsin7 significantly reduced larval weight and food conversion efficiency and prolonged larval development. In a field-simulating trial, when dsSltrypsin1/SPc or dsSltrypsin7/SPc was sprayed on soybean or maize leaves, it significantly delayed S. litura larval development, decreasing their pupation and eclosion rates by 37.1–42.3% and 12.7–30.4%, respectively. The overall control efficacy ranged from 48.1% to 57.4%, suggesting that Sltrypsin1 and Sltrypsin7 play important roles in the digestion of soybean and maize. Feeding on artificial diets containing dsSltrypsin1/SPc or dsSltrypsin7/SPc significantly reduced LC50 values of lufenuron, spinosad, RH-5849, and chlorantraniliprole, with synergism ratios exceeding 1.5-fold. These results suggested that Sltrypsin1 and Sltrypsin7 play important roles in food digestion in S. litura feeding on soybean and maize. They could be co-applied with insecticides to increase control efficiency, reduce the application amount and mitigate resistance to conventional insecticides.