细胞色素P450s酶系催化除草剂代谢的研究进展

综述了细胞色素P450s酶系催化的单加氧反应机理,细胞色素P450s酶系在酰胺类、三氮苯类、磺酰脲类、脲类、苯氧羧酸类等除草剂的活性或降解代谢中的催化反应。讨论了研究细胞色素P450s酶系代谢作用在除草剂选择性、抗药性机理,抗除草剂作物的培育以及除草剂安全剂的解毒机理等方面的意义。     

绿磺隆、醚苯磺隆及其体外代谢物对乙酰乳酸合成酶的离体抑制作用

利用异源表达于酵母细胞中的小麦细胞色素P450cDNA(CYP71C6v1)研究了磺酰脲类除草剂绿磺隆、醚苯磺隆的代谢作用。结果表明,代谢产物5-羟基绿磺隆和5-羟基醚苯磺隆能够抑制乙酰乳酸合成酶(ALS酶)活性,且代谢产物与母体化合物绿磺隆、醚苯磺隆抑制ALS酶活性的IC50值差异小,但是代谢产物在茎叶喷雾小麦和菜豆时,均未表现出活性。绿磺隆及其代谢产物抑制小麦ALS酶活性的IC50值分别为7.1×10-9和7.9×10-9mol/L,抑制菜豆ALS酶活性的IC50分别为3.6×10-9和4.1×10-9mol/L;醚苯磺隆及其代谢产物抑制小麦ALS酶活性的IC50分别为4.6×10-9和5.3×10-9mol/L,抑制菜豆ALS酶活性的IC50分别为4.7×10-9和4.9×10-9mol/L。结果表明,在磺酰脲类分子苯环5位上进行结构改造,有可能得到高活性的化合物。     

稻稗HJHL-715种群对五氟磺草胺的抗药性水平及抗性机理分析

为明确东北稻区稻稗Echinochloa oryzoides HJHL-715种群对五氟磺草胺的抗性水平及抗性机制,采用整株生物测定法测定稻稗种群对五氟磺草胺的敏感性,明确抗性种群的交互抗性和多抗性情况,研究3种细胞色素P450抑制剂对其敏感性的影响;并应用分子生物学方法进行稻稗的乙酰乳酸合酶（acetolactate synthase,ALS）离体活性测定、ALS基因序列分析及其表达量测定。结果表明:在东北稻区,五氟磺草胺对稻稗HJHL-715种群鲜重的抑制中剂量GR50为62.53 g/hm²;稻稗HJHL-715的ALS基因序列中未发现氨基酸突变,其ALS离体活性与敏感种群的ALS离体活性无显著性差异,ALS基因表达量显著低于敏感种群。1-氨基苯并三唑（1-aminobenzotriazole,ABT）、胡椒基丁醚（piperomyl butoxide,PBO）、马拉硫磷3种P450抑制剂显著提高了稻稗HJHL-715种群对五氟磺草胺的敏感性,使其对五氟磺草胺的GR50由原来的62.53 g/hm²分别下降到5.78、5.02、3.53 g/hm²。表明东北稻区已经出现了对五氟磺草胺具有高水平抗性的稻稗种群,稻稗HJHL-715种群对五氟磺草胺的抗性很可能是由细胞色素P450介导的代谢增强所致。     

杂草对乙酰乳酸合成酶抑制剂抗药性研究进展

乙酰乳酸合成酶(ALS)抑制剂类除草剂已经成为一类广泛使用的除草剂。综述了杂草对ALS抑制剂类除草剂抗药性的产生与发展、抗性机理、抗性基因应用等方面的研究进展。其抗性产生机理主要有杂草对除草剂代谢能力增强、ALS基因突变导致对除草剂敏感性降低和ALS含量提高等。     

宁夏地区稗草种群对五氟磺草胺的抗性机制研究

为了明确宁夏稻区稗草对五氟磺草胺抗性水平及抗性机制。采用整株生物测定法测定了宁夏地区6个稗的原变种Echinochloa crus-galli var.crus-galli种群对五氟磺草胺的抗性水平,并测定了每个种群的乙酰乳酸合酶基因(ALS)序列和ALS酶离体活性,以及P450抑制剂马拉硫磷对稗草种群抗性水平的影响。结果显示,与敏感种群相比,5个疑似抗性种群对五氟磺草胺表现出不同程度的抗性(10.18倍～32.71倍),其中抗性种群N14,N22,N27和N51的ALS 574位色氨酸突变为亮氨酸,N53的197位脯氨酸突变为亮氨酸,敏感种群N43没有发现突变位点,五氟磺草胺对抗性种群ALS酶的IC₅₀均明显高于敏感种群,马拉硫磷对五氟磺草胺有增效作用,可提高稗草种群对五氟磺草胺的敏感性。综上所述,稗草种群对五氟磺草胺产生抗性是由于靶标基因ALS突变,同时稗草种群对五氟磺草胺的抗性也可能与细胞色素P450介导的代谢增强有关。     

抗阿维菌素小菜蛾的细胞色素P450酶系研究

通过对不同发育时期敏感和抗阿维菌素小菜蛾品系细胞色素P450含量的测定,以及使用不同模式底物对P450单加氧酶活性的比较研究发现:除成虫期外,不同发育时期抗性品系小菜蛾中P450和细胞色素b5的含量都高于敏感品系;抗性品系还原型辅酶Ⅱ(NADPH)-细胞色素P450还原酶活性是敏感品系的1.97倍;同时发现抗性品系中甲氧试卤灵-O-脱甲基酶(MROD)、乙氧试卤灵-O-脱乙基酶(EROD)、乙氧基香豆素-O-脱乙基酶(ECOD)以及对硝基苯甲醚-O-脱甲基酶(PNOD)的活性均明显高于敏感品系,分别为敏感品系的9.41、4.15、1.67和2.94倍。研究结果表明,细胞色素P450含量和单加氧酶活性的增高是小菜蛾对阿维菌素产生抗性的一个重要机制。     

Response of Arabidopsis thaliana to 22 ALS inhibitors: baseline toxicity and cross-resistance of csr1-1 and csr1-2 resistant mutants

The baseline toxicity of 22 acetolactate synthase (ALS)-inhibiting herbicides and the cross-resistance patterns of chlorsulfuron- and imazapyr-resistant (R) lines on these 22 ALS-inhibiting herbicides were investigated using the model species Arabidopsis thaliana. The 22 herbicides consisted of 18 sulfonylureas (SU), three imidazolinones (IMI) and one triazolopyrimidine (TP). The ED₅₀ values (doses of herbicides required to reduce dry matter by 50%) of the post-emergence-treated Col and Ler susceptible (S) lines ranged from 22 to 4822 mg ha⁻¹ and from 17 to 3143 mg ha⁻¹ respectively. The csr1-1 chlorsulfuron-resistant line (substitution of Pro₁₉₇ to Ser) conferred a high resistance to the only TP tested as well as to nine SU herbicides (R:S ratio ≥30), a low resistance to two SU herbicides (R:S≥5 and <30) and little or no resistance to the three IMI and seven other SU herbicides (R:S <5). This result contradicts the expectation that an ALS mutation selected by an SU herbicide confers high cross-resistance to other SU herbicides. We found that the efficacy of specific ALS inhibitors was different for different species and therefore could not be predicted from our results with A. thaliana; however, the cross-resistance patterns in A. thaliana were highly correlated with cross-resistance patterns in unrelated species with the same resistance mutation. These results have implications for resistance management.     

杂草对除草剂抗性机理研究进展

随着除草剂持续、广泛地使用,杂草的抗性不断增强,新的抗性机理不断被发现。本文总结了近10年来杂草在靶标抗性 (target-site resistance, TSR) 和非靶标抗性 (non-target-site resistance, NTSR) 机理方面的重要研究进展。TSR机理主要包括乙酰乳酸合成酶、乙酰辅酶A羧化酶和5-烯醇丙酮酰莽酸-3-磷酸合酶等多个除草剂靶标酶上发生的新抗性突变,以及应对更强除草剂选择压力的靶标酶多位点突变和多靶标同工酶突变等。此外,本文也针对细胞色素P450 (CYP81) 酶、醛酮还原酶和ABC转运蛋白 (EcABCC8) 等NTSR机理在对草甘膦抗性中的作用进行了详细描述。本文不仅有助于全面、深入理解杂草抗性分子机理和抗性进化规律,在科学使用除草剂和创制新作用机理的除草剂等方面也具有一定的启示作用。     

Amaranthus hybridus populations resistant to triazine and acetolactate synthase-inhibiting herbicides

Amaranthus hybridus L. populations (A, B and C) obtained from escapes in Massac County and Pope County fields in southern Illinois, USA were subjected to greenhouse and laboratory experiments to measure multiple resistance to triazine and acetolactate synthase (ALS)‐inhibiting herbicides and cross‐resistance between sulfonylurea and imidazolinone herbicides. Phytotoxicity responses of the three populations revealed that only population B exhibited multiple resistances to triazine and ALS‐inhibiting herbicides. This population was >167‐, >152‐ and >189‐fold resistant to atrazine, imazamox and thifensulfuron, respectively, at the whole plant level compared with the susceptible population. Population A was only resistant to triazines and population C was only resistant to ALS‐inhibiting herbicides. Results from in vivo ALS enzyme and chlorophyll fluorescence assays confirmed these findings and indicated that an altered site‐of‐action mediated resistance to both triazine and ALS‐inhibiting herbicides. Gene sequencing revealed that a glycine for serine substitution at residue 264 of the D1 protein, and a leucine for tryptophan substitution at residue 574 of ALS were the causes of resistance for the three populations.