Abstract: 【Objective】 The objective of this study is to provide a new candidate gene for pest management through studying the function of pesticide detoxification of carboxylesterase gene LmCesF1 in Locusta migratoria. 【Method】The cDNA sequence of LmCesF1 was got from L. migratoria transcriptome database. Amino acid sequence was deduced by using the ExPASy proteomics website software. The molecular mass (MM) and isoelectric point (pI) were predicted by using ExPASy tools. To identify the catalytic triad and substrate binding pocket presented in deduced amino acid sequence, the BLASTp analyses were undertaken in NCBI conserved domain database (CDD). Signal peptide was predicted by SignalP4.0 web tools. The NetNGlyc1.0 Server was used to predict potential N-glycosylation sites of LmCesF1. Phylogenetic tree of LmCesF1 and carboxylesterases from other insect species was constructed with neighbor-joining method. Stage- (including egg, 1-5 instar nymphs and adult) and tissue-dependent expression patterns of LmCesF1 were conducted by reverse transcription quantitative PCR (RT-qPCR). The efficiency of RNAi was assessed by RT-qPCR. RNA interference followed by bioassay was applied to reveal the roles of LmCesF1 in insecticide detoxification. The insecticide was applied at 24 h after ds LmCesF1 injection, dsGFP was used as control. Each nymph was topically applied with 3 µL droplet of insecticide solution (240 ng for malathion, 15 ng for chlorpyrifos, 0.6 ng for deltamethrin and 51 ng for carbaryl) onto the abdomen between the 2nd and 3rd sterna. The doses of these insecticides were predetermined to be approximately LD30 by bioassay. Mortality was recorded at 24 h after topical application of the insecticides. 【Result】 The full-length cDNAs of LmCesF1 (3 121 bp) was obtained from L. migratoria transcriptome database. The open reading frame (ORF) of LmCesF1 was 2 490 bp, encoded 830 amino acid residues. LmCesF1 had a signal peptide at N-terminus. The deduced amino acid sequence of LmCesF1 had the catalytic triad G-P-N. Phylogenetic tree analysis showed that LmCesF1 was clustered to the F clade (nonlepidopteran juvenile hormone esterases). LmCesF1 showed lower expression at egg and early nymph stages, then reached a peak in the 4th-instar nymphs and kept high expression until the adult day 5. The results of tissue-dependent expression of LmCesF1 suggested that the highest expressions of LmCesF1 were found in the foregut, hindgut and fat bodies, but the lowest expressions were in the gastric caeca and midgut. The remaining transcript levels of LmCesF1 were 22.6% and 30% in the nymphs injected with dsLmCesF1 as compared with those in the control (injected with dsGFP) at 12 h and 24 h, respectively. These results indicated an effective silencing of LmCesF1 by RNAi at 12 h and 24 h. Insecticide bioassay showed that the nymph mortality in response to carbaryl treatment increased by 32.9% after the expression of LmCesF1 was suppressed. In contrast, similar exposures of the locusts to malathion, chlorpyrifos, and deltamethrin after the suppression of LmCesF1 did not show significant effects on the susceptibility of the locusts to these insecticides. These results indicated that LmCesF1 most likely to be involved in detoxification of carbaryl in L. migratoria. 【Conclusion】 A juvenile hormone esterase (JHE) like gene LmCesF1 was identified from L. migratoria. This gene was clustered in clade F. The catalytic triad of LmCesF1 has been changed to G-P-N. The transcripts of LmCesF1 were detectable in all tested tissues. LmCesF1 might play a significant role in detoxification of carbaryl in L. migratoria.

Key words: Locusta migratoria , carboxylesterases , reverse transcription quantitative PCR , RNA interference