棉铃虫抗药性基因频率的测定(英文)

本文介绍了用单个棉铃虫测定杀虫药剂抗性基因频率的灵敏方法。该技术是基于比较三个相等量单个棉铃虫的乙酰胆碱酯酶的活性,每个棉铃虫要用微量滴定板同时做3个试验（也可用Enppen－dorf管）（No．1－3）：1）不加抑制剂（No．1）;2）不加酶源（No．2）;3）加一定浓度的残杀威以抑制编码敏感AceS等位基因的乙酰胆碱酯酶,但不能抑制编码抗性AceR等位基因的乙酰胆碱酯酶。如果No．1和No．3的强度一样强,说明残杀威对乙酰胆碱酯酶的活性没有影响,那么基因型应是纯合抗性（AceRR,No3＝No．1）。No．2和地．3的强度一样弱时,说明残杀威完全抑制了乙酰碱酯酶的活性,那么基因型应是纯合敏感（AceSS,No．3＝No．2）。No．3的强度介于No．1（强）和No．2（弱）之间显示出残杀威部分地抑制了乙酰胆碱酯酶的活性;基因型是杂合型（AceSR,No．2＜No.3＜No．1）。作者用该方法测定了1995,1996两年从河北省邯郸和固安县棉田采集的棉铃虫,在邯郸棉铃虫抗药性高发地区,其抗性基因频率AceRR分别是68．3％和65．3％;但在棉铃虫抗药性低发区的固安县其抗性基因频率分别是23．9％和29．4％。     

不同抗性水平棉铃虫地理种群的基因交流对抗性演化的影响

采自我国长江流域棉区的棉铃虫Helicoverpa armigera(Hubner)四川简阳种群、湖北武穴种群、湖南衡阳种群、江苏南京种群和浙江杭州种群对辛硫磷、三氟氯氰菊酯和硫丹的抗性水平分别为8.05～38.80、2.86～290.00和5.87～19.83倍。抗性较高的简阳种群和较低的杭州种群正、反交F₁代对上述3种农药的抗性为13.16和16.69倍、10.43和19.00倍及12.42和10.21倍,抗性遗传的显性系数为-0.2220、-0.2147和0.0700,表明对辛硫磷、三氟氯氰菊酯抗性呈不完全隐性方式遗传,而硫丹抗性则为半隐性遗传。P1自交F2代对辛硫磷、三氟氯氰菊酯和硫丹的抗性水平分别为17.66、7.57和9.03倍,F₁代对杭州种群回交F₂代的抗性倍数分别为8.79、3.Oo和6.61。其结果,高抗区和低抗区棉铃虫种群的基因流动将导致对三种农药抗性水平明显下降。     

THE ROLE OF PHOSPHOTASE AND ACETYLCHOLINESTERASE IN RESISTANCE TO PHOXIM IN HELICOVERPA ARMIGERA

Abstract The resistance mechanism underlying phoxim resistance was investigated through biochemical assays. The results show that there are marked relations between the phosphatase activity and resistance to phoxim. Furthermore, there is a significant difference in the K _i values of AChE between the two strains and it is inferred that the change in ability of binding to phoxim is a mechanism responsible for resistance to phoxim. The metabolic inhibitors Triphenyl Phosphate (TPP) and O,O-diethyl-O-phenyl-thiophosphate (SV₁) displayed significant synergism of phoxim toxicity to the two strains. It implied that esterase might be involved in resistance. However, there is no marked correlation between resistance and CarE.     

磷酸酯酶与乙酰胆碱酯酶在棉铃虫对辛硫磷抗药性中的作用(英文)

利用人工选育的方法 ,获得了棉铃虫对辛硫磷的抗药性品系和相对敏感品系 ,其对辛硫磷的LD50分别为 1 1 .0 574μg/虫和 0 .9790 μg/虫。以生物测定与生化测定的方法 ,比较了两品系的差异。增效剂活体试验测定结果表明 ,磷酸三苯酯、增效磷对棉铃虫三龄幼虫均表现出一定的增效作用 ,但在两品系间的增效作用存在差异 ,对相对敏感品系的增效倍数分别为 2 .2 7倍、2 .1 3倍 ,对抗性品系的增效倍数分别为 6.93倍和 6.43倍。磷酸酯酶活性测定结果表明 ,抗性品系的酶活力分别是相对敏感品系的1 6.67倍 (碱性 )和 1 .89倍 (酸性 ) ,Km 和Vmax则分别为 0 .76倍、2 .64倍 (碱性 )和 1 .38倍、1 .78倍 (酸性 ) ;而羧酸酯酶的活性则差异不大。对靶标酶乙酰胆碱酯酶的活性测定表明 ,与相对敏感品系相比 ,抗性品系酶活性显著增强 ,并且对杀虫剂的亲和力降低 ,说明乙酰胆碱酯酶在棉铃虫对辛硫磷的抗药性中也有着重要的作用     

棉铃虫抗药性的生理生化机制研究

本文报道了棉铃虫Helicoverpa armigera田间抗性种群对杀虫剂抗药性的生理生化机制。抗性种群(HJ-R)5龄幼虫羧酸酯酶、谷胱甘肽转移酶、多功能氧化酶活力均明显高于相对敏感种群(HD-S)。两种群乙酰胆碱酯酶对杀虫剂敏感性没有显著差异。HJ-R种群的腹神经索对氰戊菊酯表现了2-3倍的神经不敏感性。HJ-R种群对氨基甲酸酯类杀虫剂的抗性主要是由代谢机制引起,其中多功能氧化酶可能起主导作用;对菊酯的抗性是由多功能氧化酶、酯酶、以及神经不敏感性几个因子综合作用的结果。     

BIOCHEMICAL ASPECTS OF INSECTICIDE RESISTANCE IN COTTON BOLLWORM FROM HANDAN OF HEBEI PROVINCE*

Abstract The cotton bollworms, Helicoverpa armigera Hübner, collected from Handan of Hebei Province, have evolved high resistance to pyrethroid, organophosphate and carbamate insecticides, The sensitivity of acetylcholinesterase (AChEs) to paraoxon and methomyl varied with the development stages of the cotton bollworm. After the treatments with LD₅ and LD₅₀ of parathion and methomyl to the cotton bollworms, the affinity of AChE to acetylthiocholine (ATCh) and acetyl-β-bmethyl-thio choline (MeTCh) increased significantly except the treatment of parathion using LD₅₀ dosage while the sensitivity of AChEs to paraoxon significantly decreased. The sensitivity of AChEs to methomyl strongly increased in the treatment of parathion using LD₅₀ dosages while strongly decreased in other treatments. The affinity of carboxylesterase to β-naphthyl acetate (β-NA) was higher in groups of treatment with insecticides than in group of control. The glutathione-s-transferase (GST) activity significantly decreased in the induced groups using LD₅ dosages, while increased in the selection groups using LD₅₀ dosages. The effects of parathion and methomyl on the phosphatases of cotton bollworm were related to the dosages of application and the time after treatment and the effect on the alkaline phosphatase was stronger than on acid phosphatase.     

菜缢管蚜、棉铃虫对杀虫混剂及其单剂的抗性遗传力分析

采用阈性状分析法,估算了菜缢管蚜Lipaphis erysimi(Kaltenbach)和棉铃虫Helicoverpa armigera(Hubuer)对菊马混剂及其单剂、棉铃虫对灭铃威混剂及其单剂的抗性现实遗传力,并对抗性风险和混剂延缓抗性的作用进行了评估。结果表明,菜缢管蚜和棉铃虫对混剂的现实抗性遗传力均明显小于各单剂,依次为：拟除虫菊酯>氨基甲酸酯>有机磷>混剂。混剂的使用寿命大于组成其各个单剂的使用寿命之和,抗性风险依次为：拟除虫菊脂>氨基甲酸酯>有机磷>混剂。菊马混剂对菜缢管蚜和棉铃虫,灭铃威混剂对棉铃虫均有延缓抗性发展的作用。灭铃威对棉铃虫抗性发展的延缓作用最显著。     

棉铃虫对Bt生物农药早期抗性及与转Bt基因棉抗虫性的关系

用饲料感染法建立了棉铃虫Helicoverpa rmigera(Hubmer)敏感品系(SUS1)对Bt生物农药的敏感毒力基线和区分剂量,1995年测定了五省六县棉铃虫初孵幼虫对Bt生物农药的敏感性,结果表明：山东阳谷、河北邯郸、河南新乡、安徽萧县及江苏丰县棉铃虫已产生早期抗性,抗性个体百分率为5%～10%,与敏感品系相比,LC₅₀值稍有增加,但斜率b值明显变小;而江苏东台棉铃虫仍属敏感。这是国内外首次诊测到棉铃虫对Bt生物农药抗性。用棉叶喂饲法测定比较了转Bt基因棉花品系对不同种群棉铃虫的抗虫性效果,结果表明：用早期抗性的阳谷和新乡棉铃虫初孵幼虫接虫5d后平均死亡率较敏感品系下降16%～29%,说明棉铃虫对Bt农药与转Bt生物基因棉花品系间存在交互抗性。还讨论了Bt农药的抗性治理对策。     

SUBSTRATE SPECIFICITY AND DEVELOPMENTAL CHANGES OF ACETYLCHOLINESTERASE IN COTTON BOLLWORM*

Abstract The substrate specificity and developmental changes of acetylcholinesterase (AChE) of cotton bollworm, Helicoverpa armigera Hübner, were investigated during 1991 to 1994. The insects were collected from Handan suburbs of Hebei Province and Guan County of Shandong Province. The results show that the specific activity and Michaelis constants (k_m) of AChE toward acetylthiocholine (ATCH) and acetyl-β-methyl-thiocholine (MeTCh) regularly varied with the developmental stage of cotton bollworm. The two peaks of the specific activity were observed respectively in the third instar and sixth instar of larvae. The specific activity of AChE in pupae was the lowest and that in heads of four-days moth was the highest in various developmental stages of cotton bollworms. The tendency of K_m and maximum velocity (V_max) was identical with the change of specific activity in the AChEs of cotton bollworm. The activation energy (Ea) of AChE toward MeTCh in pupae and adults was 3. 9–4. 3 times as much as that of, larvae in cotton bollworms from Handan of Hebei Province. It suggests that the spending energies of AChE for hydrolysing substrate are different in larva, pupa and adult. The optimum conditions of AChE reaction with ATCh in larvae were 50–100 mg of tissue weights for the amount of enzyme, 10–20 minutes for the reaction time, 35°C for the reaction temperature and 8. 0 for the reaction pH value.     

棉铃虫对氰戊菊酯等杀虫剂抗性的选育及其生化机理

从用药水平低的棉田采集棉铃虫Helicover pa armigera(Hubner),在室内恒温条件下以人工饲料饲养,用氰戊菊酯等4种杀虫剂和1种混合剂经点滴法分别逐代处理棉铃虫幼虫,以选育其抗药性。用氰戊菊酯选择15代,抗性达311倍;而用灭多威、甲基对硫磷和辛硫磷分别选择13代、14代及13代,抗性仅分别达10.8倍、3.5倍及5.2倍,抗性发展较慢;甲基对硫磷与辛硫磷的混合剂选择12代,抗性也只有4.8倍。氰戊菊酯和溴氰菊酯、三氟氯氰菊酯间存在明显的交互抗性。用生测法测定酶抑制剂和生化法测定酶活性的结果表明,棉铃虫对氰戊菊酯的抗性主要与幼虫体内多功能氧化酶和羧酸酯酶的活性提高有关。     

棉铃虫对拟除虫菊酯抗性稳定性研究

研究棉铃虫Helicoverpa armigera (Hubner)对三种拟除虫菊醌（氰戊菊酯、溴氰菊酯、功夫菊酯)的抗性稳定性及敏感性恢复表明,即使棉铃虫对氰戊菊酯的抗性达到3166.3倍以上,抗性仍不稳定,经14代室内饲养后抗性下降为61.4倍;对一系列田间抗性种群的抗性稳定性研究后发现,棉铃虫对这三种拟除虫菊酯的抗性不稳定,在没有杀虫剂选择的情况下,开始几代抗性下降较快,当下降到一定水平(2～9倍)后,抗性比较稳定,很难完全恢复对拟除虫菊酯的敏感性。     

RESISTANCE ALLEVIATION IN THE LARVAL COTTON BOLLWORM TO FENVALERATE AFTER PRE-TREATMENT WITH BACILLUS THURINGIENSIS

Abstract  Resistance alleviation and mechanism in the 2nd instar larvae of one susceptible and two resistant strains of the cotton bollworm, Helicoverpa armigera to fenvalerate were studied at 24 h after the larvae were treated with Bacillus thuringiensis Berliner ( B.t. ) preparation. The results showed that responses of larvae with B. t pretreatment to fenvalerate were much more sensitive than those without B. t pretreatment. Compared with the susceptible strain (YZ-S), net ratios between LD₅₀s with and without B. t pretreatment in the two resistant strains increased approximate 62. 6% and 80. 9%, respectively) inhibition of specific inhibitors to esterases displayed that efficacy of malaoxon and paraoxon methyl to acetyl-cholinesterase increased from 66.1% to 99. 8% in vitro and in vivo ; dynamic factors of acetylcholinesterase were significantly changed, in which K _m and V _max values decreased from 45. 58% to 68. 62% and K _i values of malaoxon and paraoxon methyl to acethylcholinesterase increased approximate 60%. It suggested that the sensitivity of the 2nd instar larvae of resistant H. armigera to fenvalerate might increase after B. t pretreatment for 24 h, and change of acetylcholinesterase was an important factor during resistance alleviation by B. t pretreatment. The research showed that it is practicable to bring the coordinated use of B. t and chemical insecticides in IPM system.     

CYTOCHROME P450 MONOOXYGENASES ASSOCIATED WITH PYRETHROID RESISTANCE IN HELICOVERPA ARMIGERA (HÜBNER)

Abstract  The relationship between the cytochrome P-450 (cytP⁴⁵⁰ monooxygenase and insecticide resistance mechanisms in Helicoverpa armigera was studied. The level of cytP450 and activity of p -ni-troanisole ( p -NA) O-demethylase in resistant strain were 1. 71 times and 2. 21 times respectively higher as compared with those of the susceptible strain. Aldrin epoxidase activity showed 1. 35 fold strain difference. Therefore aldrin epoxidase might not be important for H. armigera resistance mechanism, the level of cytP450 and activity of p -NA O-demethylase might play a vital role in H. armigera resistance mechanisms. The reason of different resistance mechanisms among different H. armigera resistant strains is discussed.     

棉铃虫对Bt生物农药早期抗性及与转Bt基因棉抗虫性的关系

用饲料感染法建立了棉铃虫Helicoverpa rmigera(Hubmer)敏感品系(SUS1)对Bt生物农药的敏感毒力基线和区分剂量,1995年测定了五省六县棉铃虫初孵幼虫对Bt生物农药的敏感性,结果表明：山东阳谷、河北邯郸、河南新乡、安徽萧县及江苏丰县棉铃虫已产生早期抗性,抗性个体百分率为5%～10%,与敏感品系相比,LC₅₀值稍有增加,但斜率b值明显变小;而江苏东台棉铃虫仍属敏感。这是国内外首次诊测到棉铃虫对Bt生物农药抗性。用棉叶喂饲法测定比较了转Bt基因棉花品系对不同种群棉铃虫的抗虫性效果,结果表明：用早期抗性的阳谷和新乡棉铃虫初孵幼虫接虫5d后平均死亡率较敏感品系下降16%～29%,说明棉铃虫对Bt农药与转Bt生物基因棉花品系间存在交互抗性。还讨论了Bt农药的抗性治理对策。     

苏云金杆菌预处理棉铃虫二龄幼虫对氰戊菊酯抗性缓解及机理研究(英文)

本文研究了Ｂ．ｔ预处理棉铃虫二龄幼虫对氰戊菊酯抗性缓解及其机理。结果表明,Ｂ．ｔ预处理后的幼虫对氰戊菊酯的敏感性提高。与敏感性种群相比较,在Ｂ．ｔ预处理和未处理两个抗性种群的幼虫对氰戊菊酯的ＬＤ_５０之间纯比值分别大约增加了６２．６％和８０．９％。特异性酶抑制剂对乙酰胆碱酯酶的抑制作用显示,在活体内,马拉氧磷和甲基对氧磷对乙酰胆碱酯酶抑制效率提高了６６．１％和９９．８％;在活体外抑制效率（Ｉ_５０） ６０％和６７．５％及５５．６５％和８６．４％。同时,乙酰胆碱酯酶动力学各因子变化显著。在Ｂ．ｔ预处理的抗性棉铃虫幼虫中,Ｋｍ值从４５．５８％减少到６８．６２％;Ｋｉ值分别增加了２１．２％和４０．９％及３８．２％和６１．５％。这些结果证实Ｂ．ｔ预处理２４ｈ抗性棉铃虫二龄幼虫对氰戊菊酯的敏感性提高,乙酸胆碱酯酶的变化是重要原因之一。这一研究为在ＩＰＭ系统中将微生物杀虫剂和化学杀虫剂协调使用缓解棉铃虫对拟除虫菊酯类杀虫剂的抗性提供理论基础。     

棉铃虫对氰戊菊酯抗性遗传分析

本文采用剂量对数-死亡机率值回归线(LD-P线)分析法, 利用室内选育的棉铃虫 Heliccverpa armitera(Hubner)对氰戊菊酯抗性晶系和敏感品系,研究了棉铃虫对氰戊菊酯的抗性遗传规律。结果表明,棉铃虫对氰戊菊酯的抗性为多基因遗传,其主要基因为不完全显性,抗性不属性连锁遗传。利用增效剂对抗性机理进行的研究表明,多功能氧化酶(MFO)是棉铃虫对氰戊菊酯产生抗药性的主要因子,因此,可以推断多功能氧化酶基因为不完全显性。     

棉铃虫的细胞色素P-450单加氧酶系与菊酯类杀虫剂抗药性的关系(英文)

细胞色素Ｐ－４５０单加氧酶系在昆虫抗药性中起重要作用。本文研究了棉铃虫抗性品系与敏感品系间细胞色素Ｐ４５０含量、对－硝基茴香醚Ｏ－脱甲基酶和艾氏剂环氧化酶的差异。结果表明：抗性品系中细胞色素Ｐ－４５０含量、对－硝基茴香醚Ｏ－脱甲基酶活性分别是敏感品系的１．７１倍和２．２１倍,而艾氏剂环氧化酶活性仅为１．３５倍。因此细胞色素Ｐ－４５０含量和对－硝基茴香醚Ｏ－脱甲基酶在棉铃虫抗菊酯品系中起重要作用。进一步讨论了不同抗性品系间抗性机制不同的原因。     

杀虫药剂和植物次生性物质对棉铃虫羧酸酯酶的诱导作用

棉铃虫Helzcoverpa armigera中肠羧酸酯酶(CarEs)比活力随幼虫发育阶段呈有规律的变化。5日龄以前CarEs比活力增长比较缓慢,5日龄以后Cares比活力快速增长,8日龄开始CarEs比活力下降,直到预蛹期,CarEs比活力又开始上升。用聚丙烯酰胺凝胶电泳分离到的9条同功酶带中,E2、E4、E5、E6、E7和E9对β-乙酸萘酯(β-NA)的水解活性高于对a—乙酸萘酯(a-NA),E1、E3、和E8对上述两种底物的水解活性正好相反,对a-NA活性高于对β-NA。低剂量的对硫磷、马拉硫磷、倍硫磷、增效磷(SVl)、灭多威和溴氰菊酯对棉铃虫中肠CarEs比活力和对底物的亲和力具有明显的影响。药剂处理后48 h,Cares的比活力明显降低,对底物的亲和力则与药剂的种类有关。马拉硫磷处理后48h,在观察到的6条主要同功酶带中,E3的活性明显降低,其次是E6、E2和E1;而E5的活性则明显升高,斟的活性也有所升高。用含0.01%的芸香苷、2-十三烷酮和槲皮素的人工饲料饲养棉铃虫经2代或7代后,CarEs比活力均明显升高。     

A COMPARISON OF SENSITIVITY TO INHIBITOR AMONG ACETYLCHOLINESTERASE (AChE) MOLECULAR FORMS OF RESISTANT AND SUSCEPTIBLE STRAINS IN HELICOVERPA ARMIGERA*

Abstract The sensitivity of 2.8s and 8.7s acetylcholinesterase (AChE) to eserine sulfate is significantly lower in resistant (R) strain than in susceptible (S) strain in five AChE forms isolated by sucrose gradient centrifugation from cotton bollworm, Helicoverpa armigera. There are 186 and 85 times of difference in heads of adults and 10¹⁰ and 10⁵ times of difference in heads of larvae based on a comparison of I₅₀ values for 2.8s and 8.7s forms respectively. The sensitivity of 5.3s form of AChE to eserine sulfate shows 123 times of difference between R and S strains in larvae, however no difference in adults. The above results indicate that insensitive 2.8s, 8.7s and 5.3s forms of AChE may play an important role in the resistance of cotton boll‐worm to organophosphate and carbamate insecticides.     

棉铃虫对拟除虫菊酯杀虫剂敏感性的修复及其乙酰胆碱酯酶的变化

利用生物测定和生物化学方法研究了抗药性棉铃虫Helicoverpa armigera(Hubner)对拟除虫菊酯敏感性修复及其体内乙酰胆碱酯酶的变化,结果表明：当去除药剂选择压后,棉铃虫对拟除虫菊酯的抗性为不稳定。随着停止用药的时间加长,敏感性增加。从田间收集的棉铃虫在室内饲养后第1代对拟除虫菊酯的抗性下降速度较迅速。特异性酶抑制剂对敏感和抗性棉铃虫种群的乙酰胆碱酯酶亲和力试验结果证实,乙酰胆碱酯酶在抗性棉铃虫对杀虫剂反应发生变化的过程中发生改变,这种酶的改变可能对棉铃虫抗药性的不稳定性有重要影响。