Abstract
We first measured the content of chlorfenapyr and tralopyril in silkworm larvae using HPLC, revealing that chlorfenapyr can be biotransformed into tralopyril in silkworms. Then, a differential transcriptomic database of small RNA was constructed through Illumina RNA-Sequencing. qRT-PCR was conducted to determine the expression levels of Bmo-miR-6497-5p and the target CYP450 gene, and Bmo-miR-6497-5p was significantly upregulated in the L3 silkworm larvae 24, 48, and 72 h after they were treated with chlorfenapyr. Furthermore, the target P450 gene CYP337A2 was downregulated at these time points. Dual-luciferase validation revealed that the luciferase activity significantly decreased after Bmo-miR-6497-5p bound to CYP337A2. In addition, miRNA mimics/inhibitor injection and bioassays of chlorfenapyr and tralopyril revealed that the mortality of third silkworm larvae injected with the antagomir of Bmo-miR-6497-5p was significantly increased after exposure to a sublethal concentration of chlorfenapyr. These results imply that Bmo-miR-6497-5p targets CYP337A2, regulating its expression. Also, silkworms increase their tolerance to chlorfenapyr by upregulating Bmo-miR-6497-5p expression, thereby inhibiting the biotransformation of chlorfenapyr to toxic tralopyril catalyzed by CYP337A2. The present study reveals the function of microRNA in silkworm tolerance to chlorfenapyr and improves understanding regarding insecticide resistance in Lepidopteran insects.