Dunlun Song

Quantification of γ-aminobutyric acid in the heads of houseflies (Musca domestica) and diamondback moths (Plutella xylostella (L.)), using capillary electrophoresis with laser-induced fluorescence detection.

A novel method was developed for quantifying the levels of γ-aminobutyric acid (GABA) in the heads of houseflies (Musca domestica) and diamondback moths (Plutella xylostella (L.)), using capillary electrophoresis with laser-induced fluorescence detection (CE-LIF). The GABA in sample was derivatized with 4-chloro-7-nitro-2,1,3-benzoxadiazole (NBD-Cl) prior to CE-LIF analysis. In total, 32 mmol/L borate buffer, at pH 9.2 and containing 5.3 mmol/L β-cyclodextrin (β-CD) and 10.4 mmol/L sodium dodecyl sulfate (SDS), was determined to be the optimum CE background electrolyte (BGE) for GABA analysis. The detection limit of GABA was 0.016 μmol/L. The relative standard deviations (RSDs) of the migration time and peak area of GABA were 1.78 and 4.93%, respectively. The average recoveries of 0.97, 3.88, and 5.83 μmol/L of GABA, each added to the head sample of housefly, ranged from 88.9 to 110.5%. This method is simple and applicable to GABA assays of the heads of insects. With this newly developed CE-LIF method, the amounts of GABA in the heads of houseflies (M. domestica) and diamondback moths (P. xylostella (L.)) were measured. The results are relevant to the understandings of some insecticides and insecticide-resistance mechanisms in pests.

Duplication of acetylcholinesterase gene in diamondback moth strains with different sensitivities to acephate

This study examined the acetylcholinesterase 1 gene (AChE1) in Plutella xylostella strains with different sensitivities to acephate. Multiple haplotypes of the gene were found in the field-collected strains including distinct haplotypes carrying one or both previously reported mutations (A298S and G324A). Moreover, sequencing results indicated the presence of duplicated copies of the gene in the field-collected strains. No correlation was found between copy numbers of AChE1 and levels of resistance to acephate suggesting that extensive AChE1 duplication is not a major resistance factor at least in some P. xylostella strains. Proportions of the A298S and G324A mutations showed no correlation with levels of resistance to acephate. This suggests that acephate resistance of P. xylostella is complex and cannot be evaluated based on the AChE1 copy number or proportions of the resistance mutations alone.

Both point mutations and low expression levels of the nicotinic acetylcholine receptor β1 subunit are associated with imidacloprid resistance in an Aphis gossypii (Glover) population from a Bt cotton field in China

Aphis gossypii Glover is a destructive pest of numerous crops throughout the world. Although the expansion of Bt cotton cultivation has helped to control some insect pests, the damage from cotton aphids has not been mitigated. The evolution of aphid resistance to imidacloprid has made its chemical control more difficult since its introduction in 1991. Field populations of A. gossypii that were collected from different transgenic (Bt) cotton planting areas of China in 2014 developed different levels of resistance to imidacloprid. The IMI_R strain has developed high resistance to imidacloprid with the resistance ratio >1200-fold. Compared with the susceptible IMI_S strain, the IMI_R strain also developed a high level cross resistance to sulfoxaflor and acetamiprid. The limited synergism with either PBO or DEF suggests that resistance may be due to the site mutation of molecular target rather than to enhanced detoxification. Three target-site mutations within the nicotinic acetylcholine receptor (nAChR) β1 subunit were detected in the IMI_R strain. The R81T mutation has been reported to be responsible for imidacloprid resistance in A. gossypii and M. persicae. Both V62I and K264E were first detected in A. gossypii. These point mutations are also present in field populations, suggesting that they play a role in the resistance to imidacloprid. Furthermore, the expression level of transcripts encoding β1 subunit was decreased significantly in the IMI_R strain compared with the IMI_S strain, suggesting that both point mutations and the down-regulation of nAChR β1 subunit expression may be involved in the resistance mechanism for imidacloprid in A. gossypii. These results should be useful for the management of imidacloprid-resistant cotton aphids in Bt cotton fields in China.

Omethoate-Induced Changes of (+)-δ-Cadinene Synthase Activity and Gossypol Content in Cotton Seedlings

Abstract The gene expression and activity of (+)-δ-cadinene synthase during cotton development and in response to stress, as well as the spatial and temporal pattern of sesquiterpene biosynthesis, constitute one of chemical defense mechanisms in cotton plants. In order to explore the effects of omethoate on the cotton defense in relation to (+)-δ-cadinene synthase and gossypol, effects of omethoate treatments on activity of (+)-δ-cadinene synthase and gossypol content in cotton seedlings were investigated. Cotton seedlings treated with 400 mg L−1 omethoate exhibited a significant decrease in the specific activity of (+)-δ-cadinene synthase from 12 to 120 h after treating when compared to the untreated control; significantly lower (+)-δ-cadinene synthase activity was also observed in cotton seedlings treated with 200 mg L−1 omethoate from 72 to 120 h after treating; but for cotton seedlings treated with 100 mg L−1 omethoate, from 12 to 120 h after treating, no significant changes were observed for activity of (+)-δ-cadinene synthase. The gossypol content in cotton seedlings treated with 100, 200 or 400 mg L−1 omethoate for different time periods showed no significant changes compared to that of the control. These results indicated that the activity of (+)-δ-cadinene synthase in cotton seedlings in responses to exposure of omethoate at three concentrations for different time periods followed dose- and time-dependent responses to omethoate exposure. With omethoate as a chemical stress factor for cotton seedlings, the cotton defense in relation to the activity of (+)-δ-cadinene synthase in cotton seedlings may be affected by omethoate application.

Enantioseparation of Methyl 2-Hydroxypropionate with Two Peracylated β -Cyclodextrin Derivatives as CGC Chiral Stationary Phases

Two peracylated β-cyclodextrin derivatives, 2,3,6-tri-O-valeryl-β-cyclodextrin and 2,3,6-tri-O-octanoyl-β-cyclodextrin, were synthesized and used as capillary gas chromatography (CGC) chiral stationary phases. The two peracylated β-cyclodextrin derivatives showed enantioseparation abilities to some chiral compounds tested, such as enantiomers of methyl 2-chloropropionate, 5-hydroxy-4, 4-dimethyl-dihydrofuran-2-one, and methyl 2-hydroxypropionate. As CGC chiral stationary phases, the two peracylated β-cyclodextrin derivatives showed excellent enantioseparation abilities to methyl 2-hydroxypropionate (i.e., methyl lactate). On 2,3,6-tri-O-valeryl-β-cyclodextrin, the enantiomer separation factor and resolution of methyl lactate were 1.21 and 6.08, respectively, and on 2,3,6-tri-O-octanoyl-β-cyclodextrin, 1.22 and 5.49, respectively. The enantioseparation results of methyl lactate on the two peracylated β-cyclodextrin derivatives were better than on most reported β-cyclodextrin derivatives, including a commercial chiral column of permethylated β-cyclodextrin tested in this study. Results suggest that CGC methods using 2,3,6-tri-O-valeryl-β-cyclodextrin or 2,3,6-tri-O-octanoyl-β-cyclodextrin as chiral stationary phase could be used for enantioseparation and evaluating the enantiomeric optical purity of methyl lactate.

Pyrethroid resistance associated with M918 L mutation and detoxifying metabolism in Aphis gossypii from Bt cotton growing regions of China

BACKGROUND The cotton aphid, Aphis gossypii (Glover), is a destructive pest that affects numerous crops throughout the world. Pyrethroid resistance has become endemic in A. gossypii populations in the cotton growing areas of China due to wide- spread application of insecticides. To assess the extent of pyrethroid resistance, bioassays were conducted on field populations collected from several cotton cultivation areas from 2010 to 2015. The frequency of a known resistance-associated sodium channel mutation (M918 L) in A. gossypii was evaluated and the bioassay of bifenthrin with or without the synergist was performed to illuminate the mechanisms underlying resistance to pyrethroids. RESULTS The field populations exhibited very high levels of resistance to both beta-cypermethrin and deltamethrin. Pretreatment with synergists, DEF and PBO, significantly increased the toxicity of bifenthrin to cotton aphid populations collected from Bt cotton fields in China. Further, 96.8-100% of individuals with the M918 L mutation (including both RR and RS individuals) were observed in various populations, and only 2.8-3.2% of individuals with wild-type homozygotes (SS) were detected. CONCLUSION The mutation M918 L in the voltage-gated sodium channel along with detoxifying metabolism was contributed to the pyrethroid resistance in the field populations of Aphis gossypii from cotton growing regions of China. And insecticides with different modes of action should be recommended for the control of A. gossypii in the future.

A P-glycoprotein gene serves as a component of the protective mechanisms against 2-tridecanone and abamectin in Helicoverpa armigera

P-glycoprotein (P-gp) exists in animals, fungi and bacteria and likely evolved as a defense mechanism against harmful substances. Here a cDNA (4054bp) encoding a putative P-glycoprotein gene from Helicoverpa armigera was cloned and named HaPgp1. This putative HaPgp1 sequence encoded a protein of 1253 amino acids with a molecular mass of approximately 137kDa. qPCR analyses demonstrated that the expression of HaPgp1 was significantly higher in 4th instar larvae when compared to other developmental stages. HaPgp1 transcripts were more abundant in the head and fat bodies than in other tissues. Compared with the control, the expression of HaPgp1 reach a peak at 12h after the treatment by 2-tridecanone in all tissues. However, the expression of HaPgp1 increased from 12h to 48h after treatment with abamectin in all tissues. Immunohistochemistry analyses also verified that 2-tridecanone and abamectin can induce the increase of HaPgp1 expression. RNAi of HaPgp1 significantly raised the mortality rate of larvae treated by 2-tridecanone and abamectin, as compared to control larvae fed with GFP dsRNA. These results illustrate the possible involvement of HaPgp1 as a component of the protective mechanisms to plant secondary chemicals such as 2-tridecanone and to certain classes of insecticides, like abamectin.