Genetics and mechanism of resistance to deltamethrin in a field population of Spodoptera litura (Lepidoptera: Noctuidae)

BACKGROUND: Spodoptera litura (F.) causes enormous losses in many economically important crops. The genetics of insecticide resistance has been extensively studied in several insect pests, but there is a lack of information on S. litura. Therefore, the genetics and mechanisms of the resistance of S. litura to deltamethrin were investigated. RESULTS: Bioassays at generation G1 gave resistance ratios of 9, 5, 41, 52 and 49 for deltamethrin, cypermethrin, profenofos, chlorpyrifos and triazofos respectively, when compared with the susceptible Lab-PK strain. Bioassays at G4 with a deltamethrin-selected population (Delta-SEL) showed that selection gave resistance ratios of 63 and 7 for deltamethrin when compared with the Lab-PK and UNSEL strains respectively. Cross-resistance to other insecticides tested was observed in the selected population. A notable feature of the Delta-SEL strain was that resistance to deltamethrin, cypermethrin, profenofos and chlorpyrifos did not decline over the course of five generations. Synergism tests with microsomal oxidase (MO) and esterase-specific inhibitors indicated that the deltamethrin resistance was associated with MO and, possibly, esterase activity. Reciprocal crosses between the Delta-SEL and Lab-PK strains indicated that resistance was autosomal and incompletely dominant. A direct test of monogenic inheritance suggested that resistance to deltamethrin was controlled by more than one locus. CONCLUSION: Stability and dominance of resistance and cross-resistance suggest that insecticides with different modes of action should be recommended to reduce pyrethroid selection pressure.     

玉米弯孢叶斑病菌NADPH氧化酶生物信息学分析和ATMT敲除载体的构建

从JGI数据库玉米弯孢叶斑病菌(Curvularia lunata)基因组中获得2个编码NADPH氧化酶基因ClNox1和ClNox2。采用生物信息学方法对2个基因编码的蛋白质进行性质、结构和功能等方面的预测。结果表明,ClNox1和ClNox2的分子量分别为6.35和6.41 KD,等电点PI为8.87和8.93,不稳定指数为44.50和39.45;ClNox1是亲水性膜蛋白,包含6个明显的跨膜结构域;ClNox2也是膜蛋白,包含8个明显的跨膜结构域。在亚细胞水平上,ClNox1定位于线粒体上,而ClNox2定位于细胞质膜上;在氨基酸序列方面,ClNox1和ClNox2都含有多个苏氨酸、丝氨酸和酪氨酸激酶磷酸化位点;在功能方面,ClNox1和ClNox2都包含NADPH氧化酶特征结构域,并且与Cochliobolus heterostrophus的NADPH氧化酶基因具有较高的同源性。根据ClNox1和ClNox2序列设计特异性引物扩增目的片段,与标记基因NPTII和HphGFP分别连入骨架载体pPZP100,构建完成ClNox1和ClNox2基因的敲除载体pPZP100NPTIIClNox1和pPZP100HphGFPClNox2,为根癌农杆菌介导的遗传转化提供基础材料。     

NADPH Oxidases Act as Key Enzyme on Germination and Seedling Growth in Barley (Hordeum vulgare L.)

Reactive oxygen species (ROS) play an important role in seed germination. Although hydrogen peroxide (H₂O₂), a type of ROS, enhances the germination rate of various plant seeds, little is known about the mechanism. NADPH oxidases catalyze the production of superoxide anion (O₂^-) that is one of the ROS and the enzymes regulate plant development. We, therefore, investigated the role of NADPH oxidases in seed germination and seedling growth in barley (Hordeum vulgare L.). The production of O₂^- was observed both in embryo and aleurone layers in barley seeds treated with distilled water (DW). However, it was suppressed in seeds treated with diphenylene iodonium (DPI) chloride, NADPH oxidase inhibitor. Moreover, DPI markedly delayed germination and remarkably suppressed α-amylase activity in barley seeds, indicating the importance of NADPH oxidases in germination of barley seeds. The gene expression and the enzyme activity of NADPH oxidases gradually increased after imbibition, and the enzyme activities were closely correlated with seedling growth after imbibition. Besides, DPI markedly suppressed the seedling growth. These results indicated that NADPH oxidases perform a crucial function in germination and seedling growth in barley. These facts clearly reveal that O₂^- produced by NADPH oxidases after imbibition regulates seed germination and seedling growth in barley.     

多胺氧化降解在莴苣种子萌发过程中的作用(摘要)(英文)

[目的]了解多胺氧化降解在莴苣种子萌发过程中的作用。[方法]用1.0mmol/L多胺氧化酶专性抑制剂氨基胍(AG)在24℃黑暗环境下对莴苣浸种4h,播种于含0.5%琼脂的固体培养基上用于种子萌发率的测定,另播种于用处理试剂湿润的纱布上用于内源多胺、多胺氧化酶(PAO)和二胺氧化酶(DAO)活性以及H2O2含量测定。种子萌发率采用目测法,以胚根突破种皮大于1mm为准。以测定平均值±SE(n≥200)表示种子萌发率的大小;DAO、PAO活性测定参照Su等的方法;多胺高效液相色谱测定参照Flores和Galstonl的方法;H2O2含量测定参照Brennan和Frenkel的方法。[结果]AG可强烈抑制种子的萌发,12h后种子萌发率与对照相比差异就达极显著水平。随着莴苣种子的萌发,PAO和DAO活性发生显著的变化,2种酶活性都是先上升后下降,在24和48h分别测得DAO和PAO活性高峰。AG对DAO和PAO活性有强烈的抑制作用,培养24和36h后就检测不到DAO和PAO活性。在种子萌发过程中,内源Put在前24h快速降低,24～60h降低趋缓;Spd在萌发前期降低缓慢,48h后急剧下降;Spm含量低但略呈增加趋势;多胺总量逐渐降低。AG处理可显著增加内源多胺尤其是Put和Spd含量。在种子萌发过程中,种子H2O2含量逐渐增高,36～48h时出现高峰后一直维持较高水平。AG处理可明显降低种子H2O2含量。[结论]在种子萌发期间,内源多胺和H2O2含量的变化与PAO和DAO活性变化相对应,暗示在莴苣种子萌发期间进行着活跃的多胺氧化降解代谢。     

绿盲蝽和中黑盲蝽解毒代谢酶活性的检测

害虫解毒代谢增强是对杀虫剂产生抗性的重要机制。为此,本研究建立了Bt棉田重要害虫绿盲蝽和中黑盲蝽成虫多功能氧化酶、酯酶和谷胱甘肽S转移酶3种解毒酶活性的测定方法。研究结果表明:以DCNB(1,2二氯4硝基苯)为底物,不能检测到棉盲蝽谷胱甘肽S转移酶的活性;棉盲蝽多功能氧化酶活性也不宜选用动力学法进行测定。同时2种棉盲蝽成虫的日龄对解毒酶活性影响不明显。因此,对由不同日龄成虫组成的棉盲蝽田间种群可以直接进行解毒酶活性的检测。     

马尾松毛虫多功能氧化酶测定正交试验优化研究

应用L16(43)正交试验设计对影响马尾松毛虫多功能氧化酶细胞色素P450测定的因素PMSF浓度、酶原稀释比和离心速度进行优化研究。结果表明供试的16组复配组合中,组合A1B1D1(选用0.2 mol/L PMSF浓度、酶原稀释比为2∶1,离心速度1为5 000 r/min)最优,按照该组合的条件进行马尾松毛虫MFO酶细胞色素P450测定,效果最佳。     

红茶发酵中pH调控对多酚氧化酶活性及茶黄素形成的影响

利用悬浮发酵模拟试验研究了酸处理调控红碎茶发酵．结果表明,发酵系统pH值与茶黄素（TF）和茶红素（TR）呈极显著负相关（r=-0．967,p〈0．001;r=-0．864,p〈0．005）,而与茶褐素（TB）呈极显著正相关（r=＋0．826,p〈0．005）;TF形成主要受二羟基多酚氧化酶（DPPO）与三羟基多酚氧化酶（TPPO）相对活性影响;酸处理可促进PP向TF的有效转化,TF转化率比对照提高71．2％．通过综合分析研究,提出了红茶发酵中pH调控提高TF含量的作用机制．     

不同处理对欧榛硬枝扦插生根的影响及生根过程中相关氧化酶活性的变化

以欧榛品种巴塞罗娜为材料,进行基质和生长素的对比试验,研究生根过程中过氧化物酶(POD)、多酚氧化酶(PPO)和吲哚乙酸氧化酶(IAAO)酶活性的动态变化.结果表明:在基质试验中,只有混合基质能够提高插穗生根率;在生长素试验中,IBA处理的插穗生根率达到60%,α-NAA 和ABT1处理的插穗生根率只有15%和20%,而对照不生根.榛子硬枝扦插生根过程可分为诱导期、表达期和伸长期3个阶段.POD活性在诱导期和表达期呈上升趋势,伸长期下降,在第20天和60天各有1个高峰;PPO活性在诱导期和表达期上升,伸长期下降;IAAO活性在诱导期缓慢上升,表达期急剧上升,伸长期下降.     

壳聚糖铜固定化多酚氧化酶的研究

通过壳聚糖上的－ＮＨ２与Ｃｕ＾２＋生成不饱合络合物,以高分子配位键法,对多酚氧化酶（ＰＰＯ）固定化,实验结果表明这种方法是可行的。固定化多酚氧化酶的最佳ｐＨ值为６．２４,在７０℃放置２５ｍｉｎ后酶活力保留３１．１％,。讨论了固定化酶的重复使用性、米氏常数及底物对固定化酶的影响。     

固定化酶膜催化茶多酚形成茶黄素反应条件优选

用SAS统计软件中的响应面分析法探讨了固定化多酚氧化酶酶膜催化儿茶素生产茶黄素的最佳反应条件,确定了酶膜法生产高纯度茶黄素的五个重要因素及最佳组合。五个因素包括反应时间、酶与底物之比、通气量、底物浓度和pH值。最佳反应条件为：反应时间49βmin、酶与底物之比为1:128.7、通气量为23.81L/min 、底物浓度5.95βmg/ml和pH值4.3。该条件下产物茶黄素浓度的理论值为0.766βmg/ml,实测值为0.754±0.017βmg/ml,表明优化模型合理,条件筛选结果准确。