不同地区田旋花种群对草甘膦耐受水平的差异

在温室条件下,研究了草甘膦对北京、河北等地田旋花种群生长及其莽草酸含量变化的影响。结果表明,不同地区田旋花种群对草甘膦具有普遍的耐受能力,其中河北元氏的田旋花种群对草甘膦的耐受能力最强,GR50值为4 014.92 g a.i./hm2;北京海淀田旋花种群对草甘膦相对敏感,GR50值为957.65 g a.i./hm2。经922.5 ga.i./hm2草甘膦处理后,耐药性最高的河北元氏田旋花种群体内莽草酸含量累积缓慢,处理后14 d所受伤害能自行恢复并保持正常生长;敏感的北京海淀田旋花种群体内莽草酸迅速积累,在处理后7 d莽草酸含量达到最高值,植株生长受到一定程度的抑制。田旋花体内莽草酸含量与其耐药性高低呈负相关,因此可以通过检测田旋花经草甘膦处理后体内莽草酸的含量来明确不同地区田旋花对草甘膦耐受水平的高低。     

草甘膦对田旋花和打碗花体内莽草酸含量的影响

草甘膦处理后14d,对田旋花、打碗花、大豆体内莽草酸积累量进行分析,可明确田旋花和打碗花对草甘膦的敏感性.草甘膦840ga.i./hm2、420ga.i./hm2处理后,不同植物体内莽草酸的积累量依次为大豆>打碗花>田旋花.田旋花对草甘膦的耐药能力最强,并且随着生长时间的延长对草甘膦的抗药能力增强.打碗花只是在草甘膦剂量比较低时具有一定的耐药能力.     

三种杂草对草甘膦的敏感性及处理后植株体内莽草酸积累量差异

为明确不同杂草对草甘膦的敏感性,以稗 Echinochloa crusgalli 、马唐 Digitaria sanguinalis 、藜 Chenopodium album 为供试材料,采用生物测定法和吸光光度法分别测定了草甘膦对3种杂草的抑制中浓度(GR50),以及不同剂量处理后杂草体内莽草酸积累量的变化。经410 g/hm2(有效成分)的草甘膦处理后,稗体内莽草酸积累量呈上升-下降-上升趋势,而马唐和藜则表现为缓慢上升,根据此剂量处理下莽草酸积累趋势得出,3种杂草对草甘膦的敏感性由高到低依次为稗、藜和马唐,与生测法的结果一致。经820~3 280 g/hm2(有效成分)的草甘膦处理后,3种杂草体内莽草酸积累量从第2 d开始急剧升高,增长速率随着草甘膦处理剂量的增加而加大;处理后稗、马唐和藜体内莽草酸积累量最高值差异显著,分别为1 137.9、4 989.7和2 084.2 μ g/g,为各自对照水平的16.7、23.7和82.9倍。该研究结果可为系统检测杂草对草甘膦的敏感性提供依据。     

田旋花EPSPS基因表达特征及草甘膦对其表达的影响

采用实时荧光定量RT-PCR测定了田旋花不同组织、不同叶龄的EPSPS基因mRNA的相对表达量以及草甘膦对EPSPS基因相对表达量的影响。结果表明:田旋花EPSPS基因在不同组织表达量差异显著,在叶的表达量高于茎和根;该基因在9叶期的表达量最高,是3叶期的1.5倍;在草甘膦处理后,田旋花EPSPS基因的表达量先升高后降低,在处理后24h达最大值。随草甘膦剂量增加,该基因的表达量升高。研究结果可为深入解析田旋花对草甘膦耐药性机理提供参考。     

甲磺隆和草甘膦对空心莲子草乙酰乳酸合酶活性和莽草酸含量的影响

为研究甲磺隆和草甘膦对空心莲子草的作用机理,探索其最佳的使用技术,采用有效成分为30 g/hm2和60 g/hm2 的甲磺隆处理空心莲子草,能明显抑制其茎和根乙酰乳酸合酶的比活性;有效成分为1537.5g/ hm2和3075g/ hm2 的草甘膦处理则能明显抑制空心莲子草莽草酸含量的积累;施用甲磺隆和草甘膦对空心莲子草生长的抑制作用随药剂浓度的提高而增大.二次施用草甘膦(间隔30天)的结果表明,低剂量处理对抑制空心莲子草根组织莽草酸含量的积累更明显.     

高锰胁迫下草甘膦对空心莲子草体内莽草酸含量和抗氧化酶活性的影响

鉴于高锰胁迫下空心莲子草Alternanthera philoxeroides对草甘膦的耐药性增强,在水培条件下研究了不同浓度锰条件下草甘膦处理后该草体内莽草酸的积累和主要抗氧化酶系统的响应。次高浓度锰(0.31 mmol/L)条件下培养120 d后空心莲子草体内过氧化氢酶(CAT)活性显著高于常规浓度锰处理(0.009 1 mmol/L,对照);高浓度锰(2.45 mmol/L)条件下超氧化物歧化酶(SOD)活性升高,CAT活性下降。草甘膦(按草甘膦酸68 g/hm2)茎叶处理后6 d内,常规锰浓度培养的空心莲子草体内莽草酸含量比用草甘膦刚处理时增加了31.9％~226.0%,且显著高于同一时间次高锰和高锰的处理;不同锰浓度下培养的空心莲子草体内过氧化物酶(POD)和CAT、SOD活性均为先升高后再逐渐下降,但次高锰处理的该3种酶活性均高于对照,高锰处理的SOD和POD活性高于对照,而CAT活性与对照相当。上述结果表明,在较高锰浓度下空心莲子草能启动抗氧化酶系统而能有效地清除自由基;在草甘膦处理后初期,高锰条件下空心莲子草体内莽草酸途径受抑制程度较轻,抗氧化酶活性较高,这可能是空心莲子草耐高锰和高锰条件下该草耐草甘膦的部分机制。     

藜、铁苋菜和苘麻对草甘膦的耐受性及其与莽草酸含量的关系

采用整株生物测定法研究了藜、铁苋菜和苘麻对草甘膦的耐受性,药后14 d测定结果表明,草甘膦对上述3种杂草的ED50分别为215.27、954.34、1 522.54 g a.i./hm~2。通过比较杂草植株地上部莽草酸积累量的变化发现,草甘膦1 230 g a.i./hm~2处理后,藜、铁苋菜和苘麻莽草酸积累量最大值分别为1 400.65、1247.19、581.28μg/g,莽草酸积累量越少的杂草对草甘膦耐受性也越强。药后5 d,对杂草不同部位莽草酸积累量的比较显示,3种杂草顶部叶片莽草酸积累量明显大于根部,敏感种和耐受种相比,顶部叶片莽草酸积累量的差异更为明显,该部位可以准确地评价杂草对草甘膦的耐受程度。     

抗草甘膦基因的克隆及在原核表达系统中的功能分析

为了克隆抗草甘膦基因并在原核表达系统中分析其抗性水平,利用200 μmol/L草甘膦平板从草甘膦严重污染的土壤中筛选到1株抗草甘膦菌株G6PP,经电镜和16S rDNA鉴定为恶臭假单胞菌;以该菌株基因组DNA为模板,通过PCR方法扩增出5-烯醇丙酮莽草酸-3-磷酸酯合成酶(EPSPS)基因,该基因编码440个氨基酸,亚克隆到原核表达载体pEET-28a中构建原核表达载体pET-G6;将重组表达载体转化大肠杆菌BL21 (DE3)中,经IPTG诱导,表达出46 ku的融合蛋白;携带pET-G6质粒的大肠杆菌BL21 (DE3)在液体M63培养基中能耐受150 μmol/L草甘膦的抑制.本研究结果表明克隆到的G6基因具有一定的抗草甘膦活性,对抗草甘膦作物的培育具有实践意义.     

南繁基地牛筋草对草甘膦的抗药性研究

牛筋草是一年生禾本科恶性杂草, 在我国黄淮海流域及长江以南地区的农田危害严重。草甘膦是一种优良的非选择性除草剂, 随着生物育种产业化的推进, 草甘膦会逐步在玉米、大豆等作物田登记应用。育种基地抗草甘膦杂草的产生是其快速传播的潜在因素。为明确三亚一育种基地牛筋草种群对草甘膦的敏感性, 本研究利用生物测定、分子生物学等方法检测了待测种群的抗性水平, 并分析了可能的分子机制。结果发现, 草甘膦对牛筋草种群的生长抑制中量为2 053.0 g/hm2(有效成分用量), 抗性指数(RI)为5.0; 靶标基因EPSPS的保守区域无突变, 但相对表达量是敏感种群的47.4倍; 抗性植株中EPSPS蛋白的浓度是敏感植株的17.1倍。以上结果表明, 该牛筋草种群对草甘膦产生了中等水平抗性, 靶标基因过量表达是其抗性机制之一。     

赤霉素缓解Mn~(2+)对草甘膦拮抗效应的研究

为探索能够减轻或消除Mn2+对草甘膦拮抗效应的方法,以高羊茅为试材,将赤霉素(GA3)与草甘膦和硫酸锰(Mn2+质量分数为0.1%)混用,研究了赤霉素对Mn2+降低草甘膦药效的缓解作用。结果表明:赤霉素+草甘膦+硫酸锰处理组高羊茅比同剂量草甘膦+硫酸锰处理组叶色更黄,萎蔫更严重,与同剂量草甘膦单剂处理组比较接近,其中加入50 mg/L赤霉素处理组缓解草甘膦拮抗效应的效果最好。赤霉素+草甘膦+硫酸锰处理组高羊茅的干、鲜重及叶绿素含量均低于草甘膦+硫酸锰处理组,而丙二醛和莽草酸含量均明显高于草甘膦+硫酸锰处理组。处理后第6天,赤霉素+草甘膦+硫酸锰组莽草酸含量分别比草甘膦+硫酸锰组增加了49.8%(加入50 mg/L赤霉素)和28.8%(加入30 mg/L赤霉素),差异显著;处理后第2天,赤霉素+草甘膦+硫酸锰组丙二醛含量分别比草甘膦+硫酸锰组增加了54.1%(加入50 mg/L赤霉素)和52.9%(加入30 mg/L赤霉素),差异显著。研究表明,将赤霉素与锰肥和草甘膦混合喷施,将有可能在一定程度上缓解Mn2+对草甘膦的拮抗效应,保证草甘膦的除草效果。     

Elevated antioxidant response and induction of tau-class glutathione S-transferase after glyphosate treatment in Vigna radiata (L.) Wilczek

Glyphosate (N-(phosphonomethyl) glycine) is a broad-spectrum herbicide, acting on the shikimic acid pathway inhibiting 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS), thus obstructing the synthesis of tryptophan, phenylalanine, tyrosine and other secondary products. It has also been reported to generate oxidative stress which influences the antioxidant response of target plants. The effect of glyphosate application on total protein, CAT, POD and GST activities was investigated and elevated expression of the oxidative stress enzymes was obtained after glyphosate treatment.Tau-class GSTs are plant-specific, and are chiefly involved in xenobiotics and oxidative stress metabolisms. Many herbicides and safeners have been known to selectively induce tau-class GSTs in different plant species. Here we also report the induction of tau-class GSTs after glyphosate treatment in the seedling roots of two Vigna radiata varieties (PDM11 and PDM54). GSH-agarose affinity chromatography and mass spectrometry revealed that the tau-class GSTs induced in the two varieties were different; the tau-class GSTs present in the untreated controls were also different in the two varieties. The present study highlights the elevated antioxidant response, the induction of tau-class GST and the genotypic variation in the type of tau-GST in control and glyphosate treated varieties of V. radiata.     

A target-site mutation is present in a glyphosate-resistant Lolium rigidum population

Annual ryegrass (Lolium rigidum) is the only weed species to have evolved resistance to the broad‐spectrum herbicide glyphosate in Australia. A population that had failed to be controlled by glyphosate was collected from a vineyard in the Adelaide Hills region of South Australia. Dose–response experiments on this population (SLR 77) showed that it was glyphosate resistant, with an LD₅₀ that was 1.9–3.4 times higher than that of a susceptible population (VLR 1). The movement of radiolabelled glyphosate within SLR 77 plants showed that this population did not have the differential glyphosate translocation mechanism of resistance common to several other Australian glyphosate‐resistant populations. Subsequent analysis of shikimic acid accumulation within the plant after glyphosate treatment showed that this population accumulated significantly less shikimic acid than a susceptible population, but more than a glyphosate‐resistant population with the translocation mechanism, indicating the possible involvement of another mechanism of resistance. Sequencing of a portion of the SLR 77 5‐enolpyruvylshikimate‐3‐phosphate synthase gene was carried out and a mutation causing an amino acid change at position 106 from proline to threonine was identified. This mutation is likely to be responsible for glyphosate resistance in this population, as mutations in this position have been found to be responsible for glyphosate resistance in goosegrass (Eleusine indica) from Malaysia. This paper represents the first report of target‐site glyphosate resistance in L. rigidum and provides evidence that this species has at least two mechanisms of glyphosate resistance present in Australia.     

Absorption,translocation and metabolism of 14C-glyphosate in several weed species*

The pattern and extent of ¹⁴C-glyphosate [N-(phosphonomethyl)glycine] translocation from the treated leaf and metabolism of ¹⁴C-glyphosate were studied in field bindweed (Convolvulus arvensis L.), hedge bindweed (Convolvulus sepium L.). Canada thistle [Cirsium arvense (L.) Scop.] tall morning glory [lpomoea purpurea (L.) Roth.] and wild buckwheat (Polygonum convolvulus L.). ¹⁴C was translocated throughout the plants within 3 days with accumulation in the meristematic tips of the roots and shoots evident. Cross and longitudinal sections of stems and roots showed that the ¹⁴C was localized in the phloem. Field bindweed translocated 3–5% of the applied ¹⁴C from the treated leaf, hedge bindweed 21.6%, Canada thistle 7.8%, tall morningglory 6.5%, and wild buckwheat 5%. Field bindweed, Canada thistle, and tall morningglory metabolized the parent glyphosate to aminomethylphosphonic acid to a limited extent. This metabolite made up less than 15% of the total 14C. Of the total ¹⁴C applied to excised leaves, 50% had disappeared within 25 days.     

Glyphosate toxicity in the shoot apical region of the tomato plant: I. Plastid swelling is the initial ultrastructural feature following in vivo inhibition of 5-enolpyruvylshikimic acid 3-phosphate synthase

Glyphosate induces swelling and eventual bursting of the plastids in the young tissue of the shoot apical region of tomato plants. This rapid and specific effect parallels the concentration of glyphosate in the tissues and the degree of in vivo inhibition of 5-enolpyruvylshikimic acid 3-phosphate synthase as measured by the accumulation of shikimic acid and shikimic acid 3-phosphate. The chloroplasts of the young apical leaves begin to swell between 16 and 20 hr after treatment of the plants with a sublethal glyphosate dose and burst after 4 days. Glyphosate-induced swelling of the proplastids begins much later (at 2 days) in the apical meristem itself than in the apical leaves. The meristem recovers 5 days after glyphosate treatment because the cells containing the damaged proplastids become displaced toward the rib meristem according to the inherent pattern of cell division.     

Glyphosate applied at low doses can stimulate plant growth

BACKGROUND: Glyphosate blocks the shikimic acid pathway, inhibiting the production of aromatic amino acids and several secondary compounds derived from these amino acids. Non-target plants can be exposed to low doses of glyphosate by herbicide drift of spray droplets and contact with treated weeds. Previous studies have reported that low doses of glyphosate stimulate growth, although these data are very limited. The objective of this study was to determine the effects of low glyphosate doses on growth of a range of plant species. RESULTS: Growth of maize, conventional soybean, Eucalyptus grandis Hill ex Maiden, Pinus caribea L. and Commelia benghalensis L. was enhanced by 1.8-36 g glyphosate ha(-1). Growth of glyphosate-resistant soybean was unaffected by any glyphosate dose from 1.8 to 720 g AE ha(-1). The optimum doses for growth stimulation were distinct for plant species and tissue evaluated. The greatest stimulation of growth was observed for C. benghalensis and P. caribea. Shikimic acid levels in tissues of glyphosate-treated soybean and maize were measured and found to be elevated at growth-stimulating doses. CONCLUSION: Subtoxic doses of glyphosate stimulate the growth of a range of plant species, as measured in several plant organs. This hormesis effect is likely to be related to the molecular target of glyphosate, since the effect was not seen in glyphosate-resistant plants, and shikimate levels were enhanced in plants with stimulated growth.     

Influence of glyphosate on Rhizoctonia and Fusarium root rot in sugar beet

This study tests the effect of glyphosate application on disease severity in glyphosate-resistant sugar beet, and examines whether the increase in disease is fungal or plant mediated. In greenhouse studies of glyphosate-resistant sugar beet, increased disease severity was observed following glyphosate application and inoculation with certain isolates of Rhizoctonia solani Kuhn and Fusarium oxysporum Schlecht. f. sp. betae Snyd. & Hans. Significant increases in disease severity were noted for R. solani AG-2-2 isolate R-9 and moderately virulent F. oxysporum isolate FOB13 on both cultivars tested, regardless of the duration between glyphosate application and pathogen challenge, but not with highly virulent F. oxysporum isolate F-19 or an isolate of R. solani AG-4. The increase in disease does not appear to be fungal mediated, since in vitro studies showed no positive impact of glyphosate on fungal growth or overwintering structure production or germination for either pathogen. Studies of glyphosate impact on sugar beet physiology showed that shikimic acid accumulation is tissue specific and the rate of accumulation is greatly reduced in resistant cultivars when compared with a susceptible cultivar. The results indicate that precautions need to be taken when certain soil-borne diseases are present if weed management for sugar beet is to include post-emergence glyphosate treatments.     

Glyphosate reduces urediniospore development and Puccinia psidii disease severity on Eucalyptus grandis

BACKGROUND: Recent studies have shown the effects of glyphosate drift on decreasing rust intensity on Eucalyptus grandis plants. However, the effects of the herbicide on Puccinia psidii initial development are unknown. In this study the systemic action of glyphosate on rust severity was evaluated on Eucalyptus plants maintained under greenhouse conditions. Urediniospore germination and apressorium formation on detached leaves and on water agar medium, previously treated with glyphosate, were also evaluated. RESULTS: Rust severity and the number of urediniospores per leaf area were significantly reduced with increasing glyphosate doses, even on branches not directly treated with the herbicide, indicating a systemic effect of glyphosate on pathogen development. Similarly, higher glyphosate doses also reduced germination and apressorium formation on detached Eucalyptus leaves, regardless of the direct application of the product on the leaf limb or on the petiole base. Puccinia psidii urediniospore germination in water agar medium also decreased with increasing herbicide doses. CONCLUSIONS: Reductions in germination and apressorium formation of P. psidii urediniospores with increasing glyphosate dose indicate that a lower severity and intensity of the disease may perhaps be due to blockage of the shikimic acid pathway in the fungal metabolic system. Copyright © 2011 Society of Chemical Industry     

Isolation and biochemical diagnosis of cell lines of groundnut (Arachis hypogaea L) selected on glyphosate†

Three cell lines of groundnut (Arachis hypogaea L), an important oilseed legume, were selected on glyphosate using in-vitro culture techniques. The cell lines isolated through single as well as stepwise selection procedures showed c 20-fold increase in glyphosate tolerance as compared to the unselected control cell line. Studies on the biochemical mechanism of glyphosate tolerance in these cell lines showed a significant increase in the total extractable activity of the target enzyme, 5-enolpyruvyl shikimate-3-phosphate (EPSP) synthase (EC 2.5.1.19), which was further confirmed with immunological data. The over-expressed EPSP synthase activity was, however, subject to inhibition by glyphosate in vitro. Two other key regulated enzymes of the shikimic acid pathway, 3-deoxy-D -arabino heptulosonate 7-phosphate (DAHP) synthase (EC 4.1.2.15) and chorismate mutase (CM) (EC 5.4.99.5) did not show any change in specific activity in the selected cell lines. The enhanced activity of EPSP synthase in the tolerant cell lines was found to be stably inherited in the absence of selection pressure. © 1999 Society of Chemical Industry