BACKGROUND: Insect glutathione S-transferases (GSTs) play a crucial role in insecticide detoxification. However, there remains a distinct lack of information regarding the role of GSTs in the detoxification of Tolfenpyrad (TFP) in insects. RESULTS: Real-time quantitative PCR showed significant upregulation of PxGSTs after exposure to TFP for 6 h. An in vitro inhibition assay showed that TFP could inhibit PxGST⊐, PxGSTε and PxGST⊞, and the most pronounced inhibitory effect was on PxGST⊞. Metabolism assays displayed that PxGST⊞ was superior to other test PxGSTs in metabolizing TFP. The molecular docking of TFP and PxGST⊞ revealed that the H-bond provided by the sidechains of Tyr107 and Tyr162 were key to the detoxification of TFP by PxGST⊞. Further tests using mutant PxGST⊞ proteins at the sites of Tyr107 (PxGST⊞Y107A) and Tyr162 (PxGST⊞Y162A) corroborated that the individual replacement of Tyr107 and Tyr162 could greatly weaken the binding and metabolic abilities to TFP. CONCLUSION: Metabolic interactions between the Plutella xylostella (L.) GSTs (PxGSTs) and TFP were deciphered. This study illustrates the molecular metabolism mechanism of PxGST⊞ towards TFP and provides theoretical underpinnings for the design and optimization of novel TFP-like insecticides.

BACKGROUND Acetylcholine (ACh), as a classical neurotransmitter, plays great roles in the nervous system. There is increasing evidence of its non‐neuronal roles in regulating basic cell functions in vertebrates. However, knowledge about the non‐neuronal cholinergic system in insects is scarce. RESULTS A comparative transcriptome analysis was performed to investigate differences in the key molecular components of the cholinergic system between the head and ovary. The results showed that expression levels of most cholinergic system‐related genes were higher in the head than in the ovary, and some cholinergic components were absent in the ovary. ACh contents ranged from 0.1 to 1.3 μg mg−1 of wet weight during the development of the ovary, and weak acetylcholinesterase activity was also detected. Moreover, the ovary has a capacity for ACh synthesis. Bromoacetylcarnitine (BrACar), a specific carnitine acetyltransferase (CarAT) inhibitor, greatly inhibits ACh synthesis by 83.83% in ovary homogenates, but bromoacetylcholine (BrACh), a specific choline acetyltransferase (ChAT) inhibitor, has no effect on ACh synthesis in the ovary. These findings indicate that non‐neuronal ACh in the ovary is only catalyzed by CarAT. CONCLUSION This study reveals the existence of the non‐neuronal cholinergic system in the ovary of M. separata, whose synthesis and release mechanisms are different from those of the head. These results provide novel insights into the non‐neuronal cholinergic system in insects, and will be valuable in the discovery of new target genes and the future development of green pest control. © 2022 Society of Chemical Industry.

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