山东省绿盲蝽田间种群对六种杀虫剂的敏感性监测

为了解山东地区绿盲蝽对常用杀虫剂的敏感性变化情况,于2010—2012年采用玻璃管药膜法监测山东聊城、菏泽、滨州、德州地区绿盲蝽田间种群对马拉硫磷、毒死蜱、灭多威、丁硫克百威、联苯菊酯和氟虫腈等6种杀虫剂的敏感性。结果显示,相对于2009年的监测数据,2010—2012 年各地绿盲蝽种群对不同杀虫剂表现出不同程度的敏感性变化,但相对毒力比值均小于10倍。其中对毒死蜱、联苯菊酯的敏感性均未降低,菏泽种群表现为敏感性增强,相对毒力比值小于1。2011—2012年德州种群对丁硫克百威、灭多威和氟虫腈的敏感性降低,相对毒力比值大于3倍,其它种群敏感性变化不大,相对毒力比值均小于3倍。3年间菏泽种群对马拉硫磷的敏感性变化不大,相对毒力比值小于3倍,其它种群敏感性均有所降低。因此,毒死蜱、马拉硫磷、联苯菊酯、丁硫克百威等仍是山东棉区防治绿盲蝽的有效药剂。     

试管药膜法测定10种杀虫剂对绿后丽盲蝽若虫的室内毒力*

本文研究了绿后丽盲蝽密度和食物有无对试管药膜法毒力测定结果的影响,建立了试管药膜法测定绿后丽盲蝽毒力的方法,即将20头绿后丽盲蝽的3龄若虫接入内壁分布有均匀药膜的试管中,让其在试管爬行3 h,然后转移到正常饲养条件下,24 h和48 h后统计其死亡率。应用此法测定了10种杀虫剂对绿后丽盲蝽若虫的毒力,24 h后毒力大小顺序分别为:辛硫磷>硫丹>甲维盐>马拉硫磷>毒死蜱>高效氯氰菊酯>吡虫啉>乙酰甲胺磷>啶虫脒>阿维菌素,致死中浓度（LC50）分别为：8.32、14.75、24.25、27.58、51.04、127.45、151.68、194.47、399.26、439.24 mg/L。48 h毒力大小顺序分别为: 辛硫磷>硫丹>甲维盐>马拉硫磷>毒死蜱>乙酰甲胺磷>高效氯氰菊酯>吡虫啉>啶虫脒>阿维菌素,致死中浓度（LC50）分别为：7.08、10.65、12.98、15.97、27.58、33.61、66.31、103.82、176.30、255.97 mg/L。本文结果表明,试管药膜法是测定杀虫剂对绿后丽盲蝽若虫触杀作用的简便易行的方法。     

三种不同来源胆碱酯酶用于农药残留快速检测的比较

本实验以罗非鱼肌肉胆碱酯酶、Sigma C2888乙酰胆碱酯酶及商品化农药残留检测试剂盒为检测酶,使用酶抑制法进行检测,比较了12种不同浓度农药对这三种酶酶活力抑制情况。结果表明,9种有机磷杀虫剂和3种氨基甲酸酯类杀虫剂对罗非鱼肌肉胆碱酯酶都有较强的抑制作用,IC50值均10×10-6μg/m L。敌敌畏对胆碱酯酶的抑制作用最强,其IC50值为0.031×10-6μg/m L;灭多威的抑制能力最弱,IC50值为50.91×10-6μg/m L。8种农药(丁硫克百威、敌敌畏、辛硫磷、对硫磷、甲胺磷、乐果、马拉硫磷、甲基异柳磷)对罗非鱼肌肉胆碱酯酶的抑制作用Sigma C2888,检测试剂盒ACh E对7种农药(呋喃丹、灭多威、辛硫磷、对硫磷、甲胺磷、马拉硫磷、敌百虫)的灵敏度高于Sigma C2888。与罗非鱼肌肉胆碱酯酶比较,呋喃丹、灭多威、辛硫磷、甲胺磷、敌百虫对检测试剂盒胆碱酯酶的抑制更强。     

美洲斑潜蝇幼虫对杀虫剂的敏感性研究

利用浸叶法测定了美洲斑潜蝇幼虫对１１种不同类型杀虫剂的敏感性。实验结果表明,齐墩螨素对美洲斑潜蝇具有较高的毒力,其ＬＣ_５０仅为０.０８１８ｍｇ／Ｌ,其次是菊酯类杀虫剂溴氰菊酯和高效氯氰菊酯。美洲斑潜蝇对有机磷杀虫剂马拉硫磷、敌敌畏、乙酰甲胺磷、氨基甲酸酯类杀虫剂灭多威和丁硫克百威具有较强的耐药性。     

两种蜜蜂头部乙酰胆碱酯酶对杀虫药剂敏感度比较

通过对抑制动力学常数和抑制时间进程曲线的测定,比较了中华蜜蜂Apis cerana cerana Fabricius和意大利蜜蜂Apis mellifera ligustica Spinola头部乙酰胆碱酯酶(acetylcholinesterase,AChE)对几种有机磷和氨基甲酸酯类杀虫药剂的敏感度。抑制时间进程曲线显示,意大利蜜蜂头部AChE对毒扁豆碱、灭多威、敌敌畏的敏感度高于中华蜜蜂,而两种蜜蜂对残杀威、硫双灭多威、甲胺磷及久效磷的敏感度没有明显差异。意大利蜜蜂头部AChE对毒扁豆碱、残杀威、硫双灭多威、克百威以及丁硫克百威的双分子速率常数(Ki)值分别为4.003×106、5.744×104、5.249×104、1.986×106和5.492×104 (mol/L)-1 ·min-1,均高于中华蜜蜂对这几种杀虫药剂的Ki值,后者分别为3.403×106、4.633×104、4.233×104、1.262×106和5.072×104 (mol/L)-1 · min-1。但中华蜜蜂头部AChE对灭多威的Ki值却高于意大利蜜蜂,前者为10.408×104,后者为4.872×104(mol/L)-1 ·min-1。对AChE被抑制后恢复速率(K3)的测定结果表明,中华蜜蜂头部AChE被残杀威和硫双灭多威抑制后恢复的速率显著低于意大利蜜蜂,但两种蜜蜂被毒扁豆碱、灭多威、克百威和丁硫克百威抑制后恢复的速率差异不显著。     

几种杀虫剂对烟草甲幼虫的触杀毒力

利用烟虫幼虫作试虫，测定了不同浓度的灭多威，乙酰甲胺磷，敌敌畏和敌百虫等４种药剂在４８ｈ和７２ｈ后的触杀毒力，发现灭多威对烟草甲幼虫触杀活性最高，药效发挥最快，乙酰甲胺磷次之，敌百虫活性最小，药效发挥最慢。     

5种药剂对不同发育阶段白蜡绵粉蚧Phenacoccus fraxinus的室内毒力

白蜡绵粉蚧Phenacoccus fraxinus Tang是白蜡Fraxinus chinensis Roxb.的一种重要害虫。为筛选出高效防治白蜡绵粉蚧的药剂,本试验采用浸渍法测定了5种药剂对不同龄期白蜡绵粉蚧的毒力。测试的5种药剂对白蜡绵粉蚧1龄若虫致死中浓度为18.48~24.52 mg/L,均表现较高的毒力。对2龄若虫和成虫的致死中浓度分别为18.48~76.25 mg/L和44.00~79.53 mg/L,毒力稍低于1龄若虫。但5种药剂对白蜡绵粉蚧3龄若虫的毒力明显低于其他龄期,除20%丁硫克百威乳油毒力较高(42.00 mg/L)外,其余4种药剂的致死中浓度为97.73~132.24 mg/L。因此,5种药剂对白蜡绵粉蚧1、2龄若虫和成虫毒力较强且稳定,均可作为防治白蜡绵粉蚧低龄若虫和成虫的优良药剂,但3龄若虫最好选用毒力较高的20%丁硫克百威乳油进行防治。     

几种药剂防治棉盲蝽象田间药效试验

为筛选防治棉盲蝽象的高毒农药替代产品．采用田间试验的方法比较甲氨基阿维菌素1％乳油、丁硫克百威20％乳油＋高效氯氰菊酯4．5％乳油、啶虫脒3％乳油3组药剂对棉盲蝽象的防治效果。试验结果表明：以上几种药剂对棉盲蝽象均具有较好的防治效果．且对作物安全,是防治棉盲蝽象的理想药剂。     

丁硫克百威防治白背飞虱和南方水稻黑条矮缩病效果

为了明确丁硫克百威拌种剂对水稻白背飞虱和南方水稻黑条矮缩病的防治效果及其对天敌的影响,用35%丁硫克百威种子处理干粉剂不同剂量对水稻种子拌种。结果表明,用35%丁硫克百威种子处理干粉剂30g/kg拌种,对白背飞虱和南方水稻黑条矮缩病的防治效果较好;对稻田捕食性蜘蛛种群数量有一定影响,但对水稻黑肩绿盲蝽没有显著影响。在水稻生产中,可采用35%丁硫克百威种子处理干粉剂拌种防治秧田期水稻白背飞虱和南方水稻黑条矮缩病。     

太原市不同地区苹果黄蚜抗药性水平监测

本文用FAO推荐的微量点滴法对太原市两地区的苹果黄蚜的抗药性变化进行了监测,结果发现在2010—2013的四年中,不同地区苹果黄蚜对不同药剂的敏感性变化不同。其中清徐地区苹果黄蚜对毒死蜱、乙酰甲胺磷、甲基阿维菌素、氧化乐果、马拉硫磷的敏感性有降低的趋势,其中对氧化乐果的抗性增长最大,为原来的138.94倍,对吡虫啉、啶虫脒、高效氯氰菊酯的敏感性有升高的趋势,其中对啶虫脒的敏感性升高最明显,敏感性升高为原来的27.68倍;阳曲地区黄蚜对吡虫啉、乙酰甲胺磷、甲基阿维菌素、氧化乐果、马拉硫磷的敏感性有降低的趋势,其中对马拉硫磷的抗性增长最快,为原来的151.02倍,对毒死蜱、啶虫脒、高效氯氰菊酯的敏感性有升高的趋势,其中对高效氯氰菊酯的敏感性升高最明显,敏感性升高为原来的1.735 2倍;清徐地区黄蚜对各种药剂的敏感性增长顺序为:氧化乐果乙酰甲胺磷毒死蜱马拉硫磷甲基阿维菌素吡虫啉高效氯氰菊酯啶虫脒,阳曲地区黄蚜对各种药剂的敏感性增长顺序为:马拉硫磷乙酰甲胺磷氧化乐果甲基阿维菌素吡虫啉毒死蜱啶虫脒高效氯氰菊酯。     

药剂对小菜蛾抗性及敏感品系乙酰胆碱酯酶抑制作用比较

采用浸叶法测定了云南通海、元谋和澜沧的小菜蛾plutella xylostella田间种群对常用杀虫剂的抗药性。结果表明,云南上述地区小菜蛾田间种群对各类杀虫剂均产生了不同程度的抗性。对有机磷类药剂的抗药性为1.74~31.1倍;对菊酯类药剂的抗药性为7.41~764倍;对阿维菌素类药剂则产生了 5.60~4.06×104倍的抗性。通过离体和活体试验测定了药剂对小菜蛾头部乙酰胆碱酯酶(AChE)的抑制作用。敌敌畏和灭多威对通海抗性品系AChE离体和活体内的抑制中浓度(I50)分别是敏感品系的209、26.5倍和2.21、2.16倍;敌敌畏对通海小菜蛾种群的离体和活体内抑制中时间(IT50)小于敏感品系,分别是敏感品系的0.32和0.17倍;而灭多威对通海小菜蛾种群的离体和活体内抑制中时间(IT50)则大于敏感品系,分别是敏感品系的1.37和1.74倍。     

Oxidation of chlorpyrifos, azinphos-methyl and phorate by myeloperoxidase

The present paper deals with the investigations of optimal conditions for the myeloperoxidase (MPO) mediated oxidation of chlorpyrifos, azinphos-methyl and phorate, organophosphorous pesticides (OPs) containing phosphorothionate group, from thio- to oxo-forms, in the presence of hydrogen peroxide. The aim of the work was to apply this oxidation method in the AChE based bioanalytical tests for OPs determination. The maximum concentration of oxo-forms for all tested pesticides was achieved after 10 min incubation of OPs in 50 mM phosphate buffer (pH 6.0) with 100 nM MPO in the presence of 50 μM H₂O₂. Optimal temperature for obtaining maximal concentration of oxo-forms was 37 °C. Only the parent compounds and their oxo-forms were identified chromatographically in the OPs samples after their exposure to MPO. Moreover, no hydrolysis products were detected in the time interval of 1 h after the MPO catalyzed reaction was stopped by catalase. The efficiency of OPs transformation from thio- to oxo-forms was measured using acethylcholinesterase (AChE) test, by comparison of percent of AChE inhibition before and after exposure to the oxidized sample.     

λ-cyhalothrin and cypermethrin induced in vivo alterations in the activity of acetylcholinesterase in a freshwater fish, Channa punctatus (Bloch)

In the present study, the in vivo effects of λ-cyhalothrin and cypermethrin on the activity of acetylcholinesterase (AChE, EC 3.1.1.7) were evaluated for 96 h in brain, muscle and gills of Channa punctatus. Both compounds exhibited tissue specific as well as dose dependent decrease in the activity of AChE. The treated fish showed a significant decrease in the activity of AChE in brain and a lesser inhibition in muscle and gills in response to the increasing concentrations of λ-cyhalothrin as well as cypermethrin. Our results indicated that the brain was the main target organ for both insecticides, followed by muscle and gills, as determined by AChE inhibition study. However, these organs showed variations in the degree of AChE inhibition for separate treatments of both insecticides. The λ-cyhalothrin was a more potent AChE inhibitor as compared to cypermethrin. These findings indicated that apart from the established mechanism of delayed closure of sodium ion channels, these pyrethroids inhibit the activity of AChE in C. punctatus which could further aggravate their neurotoxic effects.     

Cloning of the acetylcholinesterase 1 gene and identification of point mutations putatively associated with carbofuran resistance in Nilaparvata lugens

Molecular mechanisms of carbofuran resistance in the brown planthopper, Nilaparvata lugens Stål, were investigated. A carbofuran-resistant strain (CAS) showed approximately 45.5- and 15.1-fold resistance compared with a susceptible strain (SUS) and a non-selected field strain (FM), respectively. Activities of the esterase and mixed-function oxidase were approximately 2.8- and 1.6-fold higher, respectively, in the CAS strain than in the SUS strain, suggesting that these enzymes play a minor role in carbofuran resistance. Interestingly, the insensitivity of acetylcholinesterase (AChE) to carbofuran was approximately 5.5- and 3.7-fold higher in the CAS strain compared to the SUS and FM strains, respectively, indicating that AChE insensitivity is associated with carbofuran resistance. Western blot analysis identified two kinds of AChEs, of which the type-1 AChE (encoded from Nlace1, which is paralogous to the Drosophila AChE gene) was determined to be the major catalytic AChE in N. lugens. The open reading frame of Nlace1 is composed of 1989 bp (approximately 74 kD) and revealed 52.5% and 24.3% amino acid sequence identities to those of Nephotettix cincticeps and Drosophila melanogaster, respectively. Screening of point mutations identified four amino acid substitutions (G119A, F/Y330S, F331H and H332L) in the CAS strain that likely contribute to AChE insensitivity. The frequencies of these mutations were well correlated with resistance levels, confirming that they are associated with reduced sensitivity to carbofuran in N. lugens. These point mutations can be useful as genetic markers for monitoring resistance levels in field populations of N. lugens.     

绿盲蝽交配干扰剂丁酸己酯微囊的制备及田间防治效果

以乙基纤维素为壁材,丁酸己酯为芯材,采用相分离法制备了绿盲蝽Apolygus lucorum交配干扰剂丁酸己酯微囊。通过正交试验研究了乳化剂700#、聚乙烯醇、乙酸乙酯及控释剂正十二烷4个因素对丁酸己酯微囊形成的影响,得到最佳制备工艺条件。利用生物显微镜对微囊的形态进行了观察,同时测定了在最优条件下制备的丁酸己酯微囊的载药量、包封率和释放速率等指标,并进行了田间药效试验。结果表明:在乙基纤维素3.0 g、丁酸己酯3.0 g、乳化剂700#1.5 g、聚乙烯醇2.0 g、乙酸乙酯30 m L和正十二烷2.0 g,以及滴加速率为5 m L/min、转速为1 000 r/min条件下制备的丁酸己酯微囊的载药量和包封率分别为21.5%和91.9%,外观较完整,粒径分布较均匀,平均粒径约为301.85μm,缓释效果较好(能持续释放35 d以上)。田间试验结果显示,调查期间悬挂丁酸己酯微囊的样区诱捕到绿盲蝽224头,与对照区相比少442头,表明丁酸己酯微囊对绿盲蝽交配具有干扰作用,可明显减少绿盲蝽繁殖的数量。     

Analysis of two acetylcholinesterase genes in Bombyx mori

Two acetylcholinesterases (AChE, EC 3.1.1.7) cDNAs were identified and cloned from silkworm, Bombyx mori. One of those, BmAChE-o cDNA, is comprised of 3197 nucleotides which encode 638 amino acids, having an amino acid sequence homology of 72% with Drosophila melanogaster Ace-orthologous AChE (AO-AChE). In some species, another AChE group based on the sequence, Drosophila Ace-paralogous AChE (AP-AChE) has been recognized in relation to organophosphate- or carbamate-resistance, but there have been few reports of AP-AChE among lepidopteran species. However, we isolated the AP-AChE from lepidopteran silkworm, and cloned full ORF as BmAChE-p, which cDNA consisted of 2465 nucleotides that encode 683 amino acids. The homologies with other AP-AChEs were over 60% when compared. Although silkworm is not a target of pesticides, the genomic information obtained in this study will contribute to insecticide-resistance study on lepidopteran pest species.     

Synergism of resistance to phosalone and comparison of kinetic properties of acetylcholinesterase from four field populations and a susceptible strain of Colorado potato beetle

The susceptibility to phosalone and biochemical characteristics of acetylcholinesterase (AChE) were compared between susceptible (SS) strain and four field populations of Colorado potato beetle (CPB) collected from commercial potato fields of Hamedan Province in west of Iran. Bioassays involving topical application of phosalone to fourth instars revealed up to 252 fold resistance in field populations compared with the SS strain. Synergism studies showed that although esterase and/or glutathione S-transferase metabolic pathways were present and active against phosalone, they were not selected for and did not have a major role in resistance. It is likely that piperonyl butoxide (PBO) reduced phosalone toxicity by inhibiting bio-activation of phosalone. The affinity (K_m) and hydrolyzing efficiency (V_max) of AChE to selected substrates, namely, acetylthiocholine iodide (ATC), propionylthiocholine iodide (PTC), and butyrylthiocholine iodide (BTC) were examined. AChE inhibition by higher substrate concentration was evident only in the SS strain. In resistant field populations, Aliabad (Aa), Bahar (B) and Dehpiaz (Dp), substrate inhibition at higher concentrations was not seen. There was no definite optimal concentration found for any of the substrates examined. When ATC, PTC, and BTC were used as substrate, the reaction rates of AChE from Yengijeh (Yg) population increased as the concentration of all three substrates were increased, but were almost constant at concentration of ATC ? 3.98, PTC ? 2.8, and BTC ? 5 mM. The susceptible form of AChE had the most efficient ATC hydrolysis but very low BTC hydrolysis activity. In contrast, AChEs from field populations elicited relatively reduced ATC hydrolysis, but relatively increased BTC hydrolysis. The in vitro inhibition potency of some organophosphates (OPs), on AChEs of the field populations and SS strain was determined. The rank order from the most potent inhibitor to the least as determined by their bimolecular reaction constants (K_i) was ethyl paraoxon > diazoxon > methyl paraoxon for AChE from Aa, B, Dp, and Yg populations, respectively, whereas the rank order for the susceptible strain was methyl paraoxon > ethyl paraoxon > diazoxon.     

Characterization of acephate resistance in the diamondback moth Plutella xylostella

Decreased acetylcholinesterase (AChE) sensitivity and metabolic detoxification mediated by glutathione S-transferases (GSTs) were examined for their involvement in resistance to acephate in the diamondback moth, Plutella xylostella. The resistant strain showed 47.5-fold higher acephate resistance than the susceptible strain had. However, the resistant strain was only 2.3-fold more resistant to prothiofos than the susceptible strain. The resistant strain included insects having the A298S and G324A mutations in AChE1, which are reportedly involved in prothiofos resistance in P. xylostella, showing reduced AChE sensitivity to inhibition by methamidophos, suggesting that decreased AChE1 sensitivity is one factor conferring acephate resistance. However, allele frequencies at both mutation sites in the resistant strain were low (only 26%). These results suggest that other factors such as GSTs are involved in acephate resistance. Expression of GST genes available in P. xylostella to date was examined using the resistant and susceptible strains, revealing no significant correlation between the expression and resistance levels.