转Bt基因玉米对棉铃虫的抗性评价

棉铃虫Helicoverpa armigem（Htlbner）是常年危害玉米的重要害虫。转Bt基因抗虫玉米为害虫的防治提供了新的途径。采用室内离体生物测定方法测定了孟山都转Bt基因抗虫玉米Yield—Gard不同组织对棉铃虫初孵幼虫的杀虫活性以及田间人工接虫抗性鉴定。Bt玉米花丝、苞叶和幼嫩雌穗对棉铃虫具有较高的杀虫活性，取食Bt玉米花丝和幼嫩雌穗（籽粒和穗轴）的棉铃虫初孵幼虫不能存活和完成幼虫期发育，而对照分别有51．0％和22．0％个体完成幼虫期发育。以Bt玉米雌穗苞叶饲养棉铃虫初孵幼虫，5天后幼虫全部死亡，而非Bt玉米对照仅为16．0％，差异显著。田间花丝期人工接种棉铃虫抗性鉴定试验结果表明，Bt玉米雌穗被害率为3．5％，危害级别为0．007，百株存活幼虫14．3头。而非Bt玉米雌穗被害率达到82．5％，危害级别为2．51，平均百株存活幼虫79．6头，显著高于Bt玉米。这些结果说明转Bt基因玉米穗期对棉铃虫的防治效果明理．     

新疆棉铃虫种群对三种化学药剂的抗性

棉铃虫Helicoverpa armigera是经济作物和粮食作物主要害虫之一。为有效防治棉铃虫,新疆维吾尔自治区(简称新疆)大面积种植Bt棉,同时采用化学药剂进行防治,但是化学药剂的大量长期使用势必使棉铃虫产生抗性。武万峰等(2010)研究结果显示新疆棉铃虫种群已对三氟氯氰菊酯和甲维盐表现低水平抗性。为全面掌握新疆棉铃虫种群对这2种药剂及对氯虫苯甲酰胺新型化学药剂的抗性水平,本研究于2015-2018年自新疆阿克苏、博乐和哈密市等5个市采集棉铃虫,监测该害虫对这3种化学药剂的抗性,以期为Bt棉有效利用和化学药剂的合理施用提供理论基础。     

新疆南部转基因棉区棉铃虫种群长期动态研究

（Bacillus thuringiensis）基因抗虫棉在新疆南部推广已超过10 a,为探索Bt棉大面积种植对棉铃虫（Helicoverpa armigera）种群动态的影响,于1999-2010年使用诱虫灯监测了Bt棉大面积种植区域（麦盖提）和非Bt棉大面积种植区域（阿瓦提）棉铃虫的种群动态。结果表明：新疆南部地区20世纪90年代末发生棉铃虫害较重,2000-2004年种群数量保持较高水平;自2005年Bt棉大面积推广以后,Bt棉区棉铃虫的种群数量显著下降,棉铃虫种群数量随Bt棉种植比例的上升而下降（P<0.05）;]随着Bt棉大面积推广年数的增加,Bt棉区棉铃虫各代种群数量均逐渐下降,第二代种群数量和高峰期蛾量下降速率均较越冬代和第一代快,且第二代棉铃虫种群相对丰富度也逐渐下降。因此,新疆地区Bt棉的大面积种植能较好地控制棉铃虫的种群数量,而且对第二代棉铃虫种群的控制效果最好。     

新疆与华北棉区棉铃虫对氯氟氰菊酯kdr型抗性频率比较

通过单对杂交结合热探针法 ,研究比较了新疆与华北棉区田间种群棉铃虫对氯氟氰菊酯 kdr型抗性个体频率、抗性基因型频率和抗性基因频率的差异。结果表明 :新疆种群、河北冀州种群和河南西华种群的 kdr型抗性个体频率分别为 1.0 0 %、2 8.4 3%、2 4 .4 3% ;新疆XF1种群、冀州 JF1、JF2 种群的 kdr型抗性基因频率分别为 0、2 0 .83%、11.11%。分别从群体与个体水平上说明了新疆棉铃虫对菊酯类农药尚处于敏感阶段 ,而华北种群 kdr型抗性已达较高水平 ,并且抗性个体主要以杂合子形式存在。     

棉铃虫对三氟氯氰菊酯不同抗性种群的田间适合度

借鉴昆虫自然种群生命表的研究方法 ,在田间网笼中比较了棉铃虫三氟氯氰菊酯抗性、敏感品系及其杂交 F1代的主要生物学特征和净增殖率 (R0 )指标。结果表明 :在田间自然状态下 ,抗性品系的每头雌蛾平均产卵量、卵孵化率和幼虫发育历期分别为 314粒、2 0 .1%和 2 2 .9天 ,敏感品系分别为 4 38粒、2 8.8%和19.0天 ,两品系间差异显著 ,说明抗性品系在这些特征上存在较明显的不利性。 F1代的幼虫发育历期介于抗性和敏感品系之间。与敏感品系相比 ,抗性品系的相对适合度为 0 .4 6 ,杂交 F1代为 1.4 6。     

棉铃虫Bt抗性种群的RAPD-PCR初步分析

经室内筛选获得了棉铃虫对Bt杀虫剂、Bt毒蛋白和转Bt基因棉的抗性种群。利用RAPD技术 ,成功地扩增得到 105条多态性条带 ,经过聚类分析发现 ,棉铃虫对Bt产生抗性后在基因水平发生了变异。RAPD技术不仅可以用来鉴定棉铃虫对Bt是否产生抗性 ,而且可以区分不同的Bt抗性种群     

华北和新疆棉铃虫对三类农药抗性发展趋势分析

1996－1999年监测了河北冀州,河南西华,山东高密和新疆库尔勒棉铃虫对氯氟氰菊酯,久效磷和灭多威的抗性动态变化,结果表明,新疆棉铃虫抗性水平变化很少,尚处于敏感状态或低抗水平,而华北棉铃虫已处于中到高抗水平,不同年份比较,对氯氟氰菊酯,冀州棉铃虫抗性呈下降趋势,而西华和高密棉铃虫在1996－1998年呈上升趋势,但自1998年起华北地区大量种植Bt棉,使得1999年棉铃虫抗性有所下降,对久效磷和灭多威,华北各地棉铃虫的抗性在1996－1999年均呈上升趋势。棉铃虫抗性个体频率变化趋势与上述结果基本一致。     

一种监测棉铃虫对Bt杀虫晶体蛋白抗性的技术

研究了以诊断试剂盒监测棉铃虫对苏云金杆菌(Bt)杀虫晶体蛋白(ICP)抗性的技术。将1日龄幼虫在含诊断剂量的饲料中(每毫升饲料含Bt ICP0.01mg)取食7天,以生长至3龄以上的个体作为抗性个体的判断指标;筛选出人工饲料中防腐剂剂量的最佳值为每350毫升饲料中含丙酸2.5ml。经对多个棉铃虫不同抗性种群的验证试验,证明诊断试剂盒检测抗性个体频率较为精确且应用简便。     

我国棉铃虫对Bt蛋白Cry1Ac抗性现状及分子机制研究进展

棉铃虫Helicoverpa armigera是一种全球性的重要农业害虫,主要为害棉花、玉米和大豆等作物。长期种植单价Bt棉花（表达Cry1Ac蛋白）会使棉铃虫田间种群承受单一、持续的选择压力,必然会导致棉铃虫对Cry1Ac的抗性发生演化。该文概述我国棉铃虫田间种群对Cry1Ac的抗性现状、自然庇护所对棉铃虫Cry1Ac抗性演化的延缓作用以及棉铃虫对Cry1Ac抗性的遗传多样性,并对今后我国关于棉铃虫Bt抗性的治理对策进行了展望。     

抗Bt杀虫蛋白棉铃虫种群的相对适合度

在室内条件下,比较了对苏云金杆菌（Bacillus thuringinesis,Bt)杀虫蛋白（ICP）有20倍抗性的棉铃虫（Helicoverpa armigera Huebner)种群（R种群）与同源对照种群（S种群）之间的相对适合度,结果表明,与S种群相比,R种群的产卵量,卵孵化率和化蛹率明显降低,幼虫历期和蛹历期明显延长,R种群相对于S种群的适合度为0．65。杂交F1代RS种群相对于S种群的适合度为1．6,显示出杂交优势,正反交F1代种群对Bt ICP敏感性的差异不显,表明棉铃虫对Bt ICP的抗性遗传不存在性连锁,为常染色体遗传。     

DNA-based screening for an intracellular cadherin mutation conferring non-recessive Cry1Ac resistance in field populations of Helicoverpa armigera

A number of cadherin mutants conferring resistance to Bt toxin Cry1Ac have been reported in three major lepidopteran pests, including Helicoverpa armigera. Unlike most of the cadherin mutants conferring recessive resistance, an allele (r₁₅) with a 55aa deletion in the intracellular domain of cadherin (HaCad) was previously identified to cause non-recessive resistance to Cry1Ac in H. armigera. In the present study, a DNA-based PCR method was developed to screen the r₁₅ allele from field populations of H. armigera collected from the main cotton planting areas of China in 2011 and 2012. Three heterozygous r₁₅ alleles were detected from 562 moths collected from northern China (with intensive Bt cotton planting), and r₁₅ allele frequency was estimated to be 0.0027. However, no r₁₅ allele was detected from 314 moths collected from Xinjiang (with limited Bt cotton use). Although all the r₁₅ alleles have the same deletion in the cDNA sequence, at least four different indels causing loss of exon 32 have been detected in the genomic DNA sequences flanking exon 32 of HaCad. Multiple origins of the r₁₅ alleles illustrate parallel genotypic adaption of H. armigera to the selection pressure of Bt cotton.     

Interaction of Cry1Ac toxin (Bacillus thuringiensis) and proteinase inhibitors on the growth, development, and midgut proteinase activities of the bollworm, Helicoverpa zea

Potential resistance development to Bt cotton in certain lepidopterans has prompted research to develop strategies that will preserve this environmental-friendly biotechnology. Proteinase inhibitors are potential candidates for enhancing Bt toxicity against lepidopteran pests and for expanding the spectrum of control for other insects. Interactions of Bt toxin from Bacillus thuringiensis and proteinase inhibitors were investigated by monitoring growth, development, and gut proteinase activities of the bollworm, Helicoverpa zea. Several proteinase inhibitors were combined with Bt protoxin Cry1Ac in artificial diet and fed to newly molted 3rd-instar bollworm larvae to determine effects on larval body weight and length, pupation progress, and mortality rate. Major midgut proteinase activities, including caseinase, tryptic, and chymotrypsin activities, were examined after treatment. A concentration of Bt at a level causing minimal mortality (<10%), was mixed with the following proteinase inhibitors: benzamidine, phenylmethylsulfonyl fluoride (PMSF), and N-α-tosyl-l-lysine chloromethyl ketone (TLCK). When compared with controls, the synergistic effect of Bt toxin and proteinase inhibitors caused significant decreases in mean larval weight and length over time. Midgut samples tested against the substrates azocasein, α-benzoyl-dl-arginine-p-nitroanilide (BApNA), and N-succinyl-alanine-alanine-proline-phenylalanine-p-nitroanilide (SAAPFpNA) showed significant decreases in the protease activity of larvae fed Bt plus inhibitor versus control. Interaction of Bt and proteinase inhibitors significantly retarded larval growth and resulted in developmental delay and up to 20% mortality.     

Biochemical and physiological changes on Bacillus thuringiensis cotton after imidacloprid foliar spray

Bacillus thuringiensis cotton is a variety of cotton genetically modified to contain a gene derived from B. thuringiensis bacteria; which results in expression of toxin protein that confers resistance to bollworm complex (the most destructive pest of cotton). Introduction of Bt cotton lowered the need of insecticides, but still a number of insecticides are used for other insects like jassids, whitefly, aphids and tobacco caterpillar to which Bt gene does not provide effective control. Imidacloprid (tradename Imidacel 17.8 SL) is an insecticide designed for control of these major sucking/piercing insects that affect cotton. So in the present work we studied the post effect of imidacloprid insecticide on plant health of three Bt cotton hybrids (RCH-134, JKCH-1947, NCEH-6R) as there are reports of this insecticide causing growth and yield enhancements in absence of insect pests. Imidacloprid was first sprayed at recommended concentration (40 ml/acre) on 3 months old plants sown in randomly designed plots with three replications of each hybrid. The spray was repeated three times at 10 days interval. The level of B. thuringiensis gene expression, peroxidase activity and total phenols was measured on third day after every spray in leaves along with growth and yield of plants. The insecticide has shown to increase the level of B. thuringiensis protein, peroxidase enzyme activity, total phenols, height, number of bolls retained on plants and yield. These observations suggested that the imidacloprid treated plants showed better growth and development, thereby imidacloprid has growth enhancing effect on Bt cotton plants in addition to its insecticidal properties.     

红铃虫对Bt棉花抗性治理的研究进展

综述了Bt棉花对红铃虫的抗性效率和红铃虫对Bt棉花的抗性机理、抗性遗传、抗性监测及治理对策的研究进展.研究结果表明,不同种植区Bt棉花对红铃虫的抗性水平有较大差别.红铃虫中肠内Bt毒素结合位点数量减少是产生抗性的主要机制.抗性由具1个主效应的多位点基因控制,高水平抗性的产生与编码钙蛋白基因变异有关.避难所策略是红铃虫抗性治理的主要措施.抗性监测表明,自1997年以来,美国亚历桑那州红铃虫田间种群对Bt棉花的抗性频率无显著性变化.     

Toxicity of chlorantraniliprole to Cry1Ac-susceptible and resistant strains of Helicoverpa armigera

Transgenic Bt cotton expressing Cry1Ac is important in controlling various agricultural pests, including Helicoverpa armigera. Especially for transgenic crops that are cultivated in large expanses, avoiding resistance development is a key for ensuring sustainability of Bt technologies. Integrated pest management, in which transgenic crops are strategically combined with rational pesticide use, may help to prevent H. armigera resistance acquisition in Bt cotton. In this study, we evaluated the toxicity of a novel insecticide (chlorantraniliprole) on Cry1Ac-susceptible and resistant individuals of H. armigera. More specifically, we assessed the effect of chlorantraniliprole on the activity of two enzymes and conducted laboratory bioassays to determine its toxicity on H. armigera larvae. Chlorantraniliprole increased esterase and glutathione-S-transferase activities in Cry1Ac susceptible and resistant populations of H. armigera. Cry1Ac resistant populations XJ-F (Cry1Ac resistance ratio 21.8-fold), XJ-10.0 (95.8-fold) and BTR (3536.5-fold) did not show cross-resistance to chlorantraniliprole, with LC₅₀ values of 0.0733 (μg/mL) in XJ-F, 0.0545 (μg/ml) in XJ-10.0 and 0.0731 (μg/mL) in BTR, which were close to that in the susceptible strain 96S (0.0954 μg/mL). Our work shows that chlorantraniliprole could be considered to be integrated in Bt cotton management schemes to delay the H. armigera resistance development.     

基于风险评估的新疆抗虫棉种植分析

基于新疆棉铃虫生物学特性和多年的种群动态趋势,并在收集棉花种植面积、产量、棉花市场价格及棉铃虫防治费用等相关数据的基础上,采用种群模拟模型(CLIMEX模型)并结合随机模拟方法(@RISK软件),评估不同场景下棉铃虫对新疆棉花产业造成的潜在经济损失.棉铃虫种群模拟表明:随着未来气候变化,新疆棉铃虫的周增长指数(GIw)...     

Variation of acephate susceptibility and correlation with esterase and glutathione S-transferase activities in field populations of the tarnished plant bug, Lygus lineolaris

The tarnished plant bug (TPB) has increasingly become an economically important pest of cotton. Heavy dependence on insecticides, particularly organophosphates and pyrethroids, for TPB control facilitated resistance development to multiple classes of insecticides. To better understand resistance and explore ways to monitor resistance in field populations, this study examined acephate susceptibility and the activities of two major detoxification enzymes in nine field populations collected in the Delta region of Mississippi and Arkansas in 2010. Two Arkansas populations from Reed and Backgate had 3.5- and 4.3-fold resistance to acephate, as compared to a susceptible laboratory strain. Extensive planting of cotton and heavy chemical sprays is a major driving force for resistance development to acephate in Mid-south cotton growing areas. Reduced susceptibility to acephate was highly correlated with elevated esterase activities. The acephate-resistant populations from Backgate, Lula, and Reed consistently had higher (up to 5.3-fold) esterase activities than susceptible populations. Regression analysis of LC₅₀s with kinetic esterase activities revealed a significant polynomial quadratic relationship with R² up to 0.89. Glutathione S-transferase (GST) also had elevated activity in most populations, but the variations of GST activities were not significantly correlated with changes of acephate susceptibility. Finally, examination of esterase and GST inhibitors indicated that suppression rates (up to 70%) by two esterase inhibitors in 2010 were slightly lower than those detected in 2006, and ethacrynic acid (EA) inhibited GST effectively in both years. Two other GST inhibitors (sulfobromophthalein and diethyl maleate) displayed significantly lower suppression rates in 2010 than those detected in 2006, suggesting a potential genetic shift in pest populations and a necessity of continued monitoring for insecticide resistance with both bioassay and biochemical approaches. Results indicated that using major detoxification enzyme activities for resistance monitoring may provide insight into acephate resistance in field populations of TPB.     

棉花叶片茸毛性状与绿盲蝽抗性的关系

为明确棉花叶片茸毛性状与其对绿盲蝽抗性的关系,于2008-2009年连续两年系统研究了不同供试棉花品种(系)对绿盲蝽的抗性水平,并在室内测定了棉花中脉和叶片上茸毛的类型、密度和长度.结果表明,不同棉花品种(系)对绿盲蝽的抗性水平存在差异;且不同棉花品种(系)叶片各类型茸毛密度和茸毛长度分别表现显著差异(P<0.05);叶片茸毛密度与棉花对绿盲蝽的抗性呈显著负相关(y=3.5482-0.0007x1-0.0089x2,R2=0.5741,P=0.0007),叶片茸毛类型和茸毛长度与棉花对绿盲蝽的抗性没有显著相关性.选育叶片无毛或少毛的棉花品种(系)可用于棉花绿盲蝽的治理.     

棉铃虫对Cry1Ac抗性的稳定性及其对适合度的影响

为明确棉铃虫Helicoverpa armigera（Hübner）对苏云金芽胞杆菌Bacillus thuringiensis（Bt）Cry1Ac毒蛋白抗性的稳定性及其适合度变化,利用生物测定的方法研究了Cry1Ac抗性品系棉铃虫转到正常饲料饲养后的抗性衰退及再次筛选后抗性的恢复情况,并比较了敏感、抗性和抗性衰退后各品系间的适合度差异。在失去选择压的情况下,高抗品系棉铃虫对Cry1Ac的抗性迅速衰退,经过4代后抗性水平由最初的3626.67倍下降到1436.67倍;到第12代时抗性水平已低于10倍,随后品系保持较稳定的低抗水平;当重新进行抗性再筛选时,其抗性水平可快速恢复,抗性倍数快速回升,5代后恢复到1123.33倍。与敏感品系相比,高抗棉铃虫品系的适合度明显降低,相对适合度仅为0.33,但转到正常饲料连续饲养14代后,棉铃虫适合度明显上升,相对适合度为0.87,主要表现为卵孵化率和幼虫存活率等显著提高。