Rapid vacuolar sequestration: the horseweed glyphosate resistance mechanism

BACKGROUND: Glyphosate‐resistant (GR) weed species are now found with increasing frequency and threaten the critically important GR weed management system. RESULTS: The reported ³¹P NMR experiments on glyphosate‐sensitive (S) and glyphosate‐resistant (R) horseweed, Conyza canadensis (L.) Cronq., show significantly more accumulation of glyphosate within the R biotype vacuole. CONCLUSIONS: Selective sequestration of glyphosate into the vacuole confers the observed horseweed resistance to glyphosate. This observation represents the first clear evidence for the glyphosate resistance mechanism in C. canadensis. Copyright © 2010 Society of Chemical Industry     

Glyphosate-resistant horseweed made sensitive to glyphosate: low-temperature suppression of glyphosate vacuolar sequestration revealed by 31P NMR

BACKGROUND: Horseweed has been the most invasive glyphosate‐resistant (GR) weed, spreading to 16 states in the United States and found on five continents. The authors have previously reported that GR horseweed employs rapid vacuolar sequestration of glyphosate, presumably via a tonoplast transporter, substantively to reduce cytosolic glyphosate concentrations. 1 It was hypothesized that glyphosate sequestration was the herbicide resistance mechanism. If resistance is indeed endowed by glyphosate sequestration, suppression of sequestration offers the potential for controlling GR horseweed at normal herbicide field‐use rates. RESULTS: Low‐temperature ³¹P NMR experiments performed in vivo with GR cold‐acclimated horseweed showed markedly suppressed vacuolar accumulation of glyphosate even 3 days after glyphosate treatment. [In stark contrast, 85% of the visible glyphosate was sequestered 24 h after spraying warm‐acclimated GR horseweed.] Cold‐acclimated GR horseweed treated at normal use rates and maintained at low temperature succumbed to the lethal effects of glyphosate over a 40 day period. Treatment of GR horseweed in the field when temperatures were cooler showed the predicted positive herbicidal response. CONCLUSIONS: Low temperature markedly diminishes vacuolar sequestration of glyphosate in the GR horseweed biotype, yielding a herbicide response equivalent to that of the sensitive biotype. This supports the recent hypothesis 1 that glyphosate sequestration is the resistance mechanism employed by GR horseweed. Copyright © 2011 Society of Chemical Industry     

Shikimate accumulation in nine weedy species following glyphosate application

Glyphosate is commonly used in a variety of weed control scenarios and plants respond biochemically by continuing to attempt to push carbon through the shikimate pathway. The result is an accumulation of shikimate in sensitive plants in the days immediately after application. This research determined shikimate accumulation dynamics in nine problematic plant species from 0 to 6 days after treatment (DAT). Ambrosia artemisiifolia, Trifolium repens and Conyza canadensis showed rising concentrations through 6 DAT to >4000 mg L^?1. Chenopodium album, Xanthium strumarium and Urochloa platyphylla showed concentrations levelling off to a constant of about 2000 mg L^?1. Ambrosia trifida, Amaranthus palmeri and Polygonum pensylvanicum generally showed lower levels of shikimate accumulation and concentrations were decreasing after 3 DAT. No apparent relationships were evident between shikimate accumulation pattern and plant growth rate or speed of glyphosate activity. There also appeared to be no pattern in the shikimate accumulation trends that would allow for an informed decision as to which of these species would be most likely to develop glyphosate resistance. Shikimate concentrations were in agreement between field and glasshouse conditions for C. canadensis, but did not agree with those from A. palmeri.     

Evolution and spread of glyphosate resistance in Conyza bonariensis in California and a comparison with closely related Conyza canadensis

Glyphosate‐resistant weeds are an increasing problem in perennial cropping systems in the Central Valley of California, USA. To elucidate the evolutionary origins and spatial spread of resistance, we investigated the geographical distribution of glyphosate resistance and the population genetic diversity and structure of Conyza bonariensis and compared the results with previously studied C. canadensis. Thirty‐five populations from orchards and vineyards across the Central Valley were sampled. Population genetic structure was assessed using microsatellite markers. Population‐level resistance was assessed in glasshouse screening of plants grown from field‐collected seed. Bayesian clustering and analyses of multilocus genotypes indicated multiple origins of resistance, as observed in C. canadensis. Pairwise F_ST analysis detected spatial spread of resistance in the south of the Central Valley, also similar to C. canadensis. The results strongly indicate that the southern valley was an environment markedly more suitable than the northern valley for resistance spread and that spread in Conyza species was driven by increased uniformity of strong selection in the southern valley, due to recent regulation on herbicides other than glyphosate. Accordingly, resistant C. canadensis individuals occurred at high frequencies only in the southern valley, but interestingly resistant C. bonariensis occurred at high frequencies throughout the valley. Expression of resistance showed varying degrees of plasticity in C. bonariensis. The lower selfing rate and substantially greater genotypic diversity in C. bonariensis, relative to C. canadensis, indicate greater evolutionary potential over shorter time periods. Interspecific hybridisation was detected, but its role in resistance evolution remains unclear.     

Effects of glyphosate on soil microbial communities and its mineralization in a Mississippi soil

Transgenic glyphosate-resistant (GR) soybean [Glycine max (L.) Merr.] has enabled highly effective and economical weed control. The concomitant increased application of glyphosate could lead to shifts in the soil microbial community. The objective of these experiments was to evaluate the effects of glyphosate on soil microbial community structure, function and activity. Field assessments on soil microbial communities were conducted on a silt loam soil near Stoneville, MS, USA. Surface soil was collected at time of planting, before initial glyphosate application and 14 days after two post-emergence glyphosate applications. Microbial community fatty acid methyl esters (FAMEs) were analyzed from these soil samples and soybean rhizospheres. Principal component analysis of the total FAME profile revealed no differentiation between field treatments, although the relative abundance of several individual fatty acids differed significantly. There was no significant herbicide effect in bulk soil or rhizosphere soils. Collectively, these findings indicate that glyphosate caused no meaningful whole microbial community shifts in this time period, even when applied at greater than label rates. Laboratory experiments, including up to threefold label rates of glyphosate, resulted in up to a 19% reduction in soil hydrolytic activity and small, brief (<7 days) changes in the soil microbial community. After incubation for 42 days, 32-37% of the applied glyphosate was mineralized when applied at threefold field rates, with about 9% forming bound residues. These results indicate that glyphosate has only small and transient effects on the soil microbial community, even when applied at greater than field rates.     

Effects of repeated glyphosate applications on soil microbial community composition and the mineralization of glyphosate

Background: Repeated applications may have a greater impact on the soil microbial community than a single application of glyphosate. Experiments were conducted to study the effect of one, two, three, four or five applications of glyphosate on soil microbial community composition and glyphosate mineralization and distribution of ¹⁴C residues in soil. RESULTS: Fatty acid methyl esters (FAMEs) common to gram‐negative bacteria were present in higher concentrations following five applications relative to one, two, three or four applications both 7 and 14 days after application (DAA). Additionally, sequencing of 16S rRNA bacterial genes indicated that the abundance of the gram‐negative Burkholderia spp. was increased following the application of glyphosate. The cumulative percentage ¹⁴C mineralized 14 DAA was reduced when glyphosate was applied 4 or 5 times relative to the amount of ¹⁴C mineralized following one, two or three applications. Incorporation of ¹⁴C residues into soil microbial biomass was greater following five glyphosate applications than following the first application 3 and 7 DAA. CONCLUSION: These studies suggest that the changes in the dissipation or distribution of glyphosate following repeated applications of glyphosate may be related to shifts in the soil microbial community composition. Copyright © 2009 Society of Chemical Industry     

药液表面张力和黏度对草甘膦药效的影响及其机理研究

通过添加有机硅助剂和丙三醇调节草甘膦药液的表面张力和黏度,测定其对草甘膦药液液滴的物理性状及生物活性的影响。结果表明：降低药液的表面张力,雾滴的铺展直径增加,干燥时间缩短,药液的黏度以及在杂草叶片表面的最大稳定持留量没有显著性变化;草甘膦对杂草的防效表现为先升高后降低,对阔叶杂草最高目测防效和鲜重防效可提高42%和41%,对禾本科杂草防效可提高37%和37%。增加草甘膦的药液黏度,药液在杂草叶片上的最大稳定持留量增加,表面张力降低,对雾滴的干燥时间和铺展直径影响很小;草甘膦对阔叶杂草最高目测防效和鲜重防效可提高42%和41%,对禾本科杂草的防效可提高42%和42%。适当降低草甘膦药液的表面张力或增加其黏度均可提高其对杂草的防除效果。     

Absorption and translocation of glyphosate with conventional and organosilicone adjuvants

This study was conducted to determine the effects of three organosilicone-based and six conventional organic adjuvants on the absorption and translocation of ¹⁴C-glyphosate in guineagrass and redroot pigweed. The organosilicone adjuvants produced rapid absorption of the ¹⁴C-glyphosate into the redroot pigweed leaves, reaching maximum absorption within 0.5–1.0 h after application. The conventional adjuvants produced slower absorption of the ¹⁴C-glyphosate, as the maximum absorption was not achieved until at least 24 h after application in redroot pigweed, remaining similar until 72 h. In guineagrass, the maximum absorption of the glyphosate was earlier than 24 h with the organosilicone-based adjuvants, compared with longer times for the conventional adjuvants. The organosilicone-based adjuvants also increased the glyphosate translocation in redroot pigweed, but not in guineagrass. Organosilicone adjuvants have the potential to provide greater rainfastness to glyphosate on redroot pigweed and, to a lesser extent, on guineagrass.     

草甘膦作用机制和抗性研究进展

草甘膦是迄今为止最为重要、应用最广泛和最优秀的除草剂之一。然而,由于抗草甘膦转基因作物的广泛商业化导致草甘膦使用量迅速增长,杂草抗药性发生,这不仅对草甘膦的药效发挥和未来可持续应用造成了严重影响,而且对现代农业生产安全构成了威胁。本文通过对草甘膦的作用机理、草甘膦抗性杂草发展现状和抗性机制进行系统的总结和分析,以期为我国草甘膦的抗性研究和科学使用提供参考。     

超高效液相色谱-串联质谱法测定柑橘中的草甘膦及其代谢物残留

本研究首次建立了一种超高效液相色谱-串联质谱法(UHPLC-MS/MS)同时测定柑橘中草甘膦及其代谢物N-乙酰草甘膦?氨甲基磷酸(AMPA)和N-乙酰AMPA?样品经过0.5%甲酸水提取, 以十八烷基硅烷键合硅胶柱(C18)净化, 利用水和200 mmol/L碳酸氢铵溶液(含0.1%氨水)作为流动相梯度洗脱, 在多反应监测模式下定量分析?结果表明, 草甘膦及其代谢物在0.05~1 mg/kg范围内线性关系良好, 相关系数均大于0.99; 草甘膦及其3个代谢物在柑橘全果和果肉基质中3个加标水平下回收率为 70.5%~109.5%, 相对标准偏差(RSD)为0.6%~10.1%, 定量限为0.05 mg/kg?本方法的前处理样品无需衍生, 简便?快捷?高效?准确可靠, 可用于柑橘中草甘膦及其代谢物N-乙酰草甘膦?AMPA和N-乙酰AMPA的定量检测?     

Comparison of efficacy, absorption and translocation of three glyphosate formulations on velvetleaf (Abutilon theophrasti)

Growth analysis, absorption and translocation studies were conducted to compare a 1-aminomethanamide dihydrogen tetraoxosulfate (GLY-A) formulation of glyphosate with two isopropylamine (GLY-IPA-1, GLY-IPA-2) formulations of glyphosate on velvetleaf. The two isopropylamine formulations differed by the presence of a surfactant in the formulation, GLY-IPA-1 containing surfactant whereas GLY-IPA-2 did not. Four- to six-leaf velvetleaf was treated with GLY-A and GLY-IPA-1 and GLY-IPA-2 (0, 50, 67, 89, 119, 158, 280, 420, 560 and 840 g AE ha(-1)) with and without ammonium sulfate (AMS; 20 g L(-1)). GLY-A and GLY-IPA-2 included a non-ionic surfactant (2.5 mL L(-1)) in the spray solution at all herbicide concentrations. No additional surfactant was added to GLY-IPA-1. The IC50 value for GLY-A was 88 g AE ha(-1) compared with 346 and 376 g AE ha(-1) for GLY-IPA-1 and GLY-IPA-2 respectively in the absence of AMS. When AMS (20 g L(-1)) was added to the spray solution, the estimated IC50 values were 143, 76 and 60 g AE ha(-1) for GLY-IPA-1, GLY-IPA-2 and GLY-A respectively. Absorption of 14C-glyphosate into the third leaf of five- to six-leaf velvetleaf was three- to sixfold greater 72 h after treatment (HAT) when applied as GLY-A compared with GLY-IPA-1 and GLY-IPA-2 respectively in the absence of AMS. AMS (20 g L(-1)) increased absorption of 14C-glyphosate in all glyphosate formulations two- to threefold, but differences among the formulations remained. Approximately three- and sixfold more 14C-glyphosate applied as GLY-A had translocated out of the treated leaf compared with GLY-IPA-1 and GLY-IPA-2 respectively by 72 HAT. Adding AMS (20 g L(-1)) increased translocation of 14C-glyphosate out of the treated leaf approximately 2.5-fold for all three formulations. The increased efficacy of GLY-A versus GLY-IPA-1 and GLY-IPA-2 on velvetleaf is due to the greater rate of absorption and subsequent translocation of glyphosate out of the treated leaf. AMS increased the efficacy of all three formulations by increasing absorption and translocation of glyphosate in the plant.     

Evaluation of the efficacy of glyphosate plus urea phosphate in the greenhouse and the field

BACKGROUND: Glyphosate is a non‐selective, foliar‐applied, systemic herbicide that kills weeds by inhibiting the synthesis of 5‐enolpyruvylshikimate‐3‐phosphate synthase. Urea phosphate (UPP), made by the reaction of urea with phosphoric acid, was applied as an adjuvant for glyphosate in this study. Experiments in the greenhouse and the field were conducted to determine the effects of UPP by comparing the efficacies of glyphosate plus UPP, glyphosate plus 1‐aminomethanamide dihydrogen tetraoxosulfate (AMADS) and Roundup. RESULTS: The optimum concentration of UPP in glyphosate solution was 2.0% when UPP was used as an adjuvant. The ED₅₀ values for glyphosate‐UPP were 291.7 and 462.4 g AI ha^?1 in the greenhouse and the field respectively, while the values for Roundup were 448.2 and 519.6 g AI ha^?1. The ED₅₀ values at 2 weeks after treatment (WAT) and 3 WAT were lowered when UPP was used as an adjuvant in the greenhouse and field study, and the glyphosate + UPP was absorbed over a 2 week period. UPP may increase the efficacy by causing severe cuticle disruption or accelerating the initial herbicide absorption. The result also showed that UPP could reduce the binding behaviour of Ca²⁺ to glyphosate. CONCLUSION: The application of UPP as an adjuvant could increase the efficacy of glyphosate and make it possible to achieve effective control of weeds with glyphosate at lower dose. Moreover, UPP showed less causticity to spraying tools and presented less of a health hazard. Therefore, UPP is accepted as being a new, effective and environmentally benign adjuvant for glyphosate. Copyright © 2011 Society of Chemical Industry     

Effects of surfactants and simulated rainfall on the efficacy of the Engame formulation of glyphosate in johnsongrass, prickly sida and yellow nutsedge

The effects of surfactants and simulated rain were investigated on the efficacy of Engame and Roundup Ultramax formulations of glyphosate on johnsongrass ( Sorghum halepense L.), prickly sida ( Sida spinosa L.) and yellow nutsedge ( Cyperus esculentus L.). Flame surfactant provided the greatest enhancement of Engame efficacy and the effect was species-dependent. Flame enhanced the activity of Engame on johnsongrass and yellow nutsedge but not on prickly sida. Engame and Engame plus Flame were more active than Roundup Ultramax on a glyphosate acid-equivalent basis on johnsongrass without rain, and on yellow nutsedge with or without rain. The Engame and Roundup Ultramax activities on johnsongrass were similar with rain, and rain occurring between five and 30 min after treatment diminished their activities to < 38% of the control. With the addition of Flame surfactant, Engame activity on johnsongrass increased, such that 50% and 80% of the control were realized, even with rain occurring between five and 15 min after treatment, respectively. Engame and Roundup Ultramax provided better control of prickly sida than of johnsongrass following a rain event. The addition of Flame surfactant to Engame did not enhance the activity on prickly sida. Yellow nutsedge control with Engame and Engame plus Flame was greater than with Roundup Ultramax and rain had little effect on control regardless of the length of the rain-free period. These results demonstrated that the activities of Engame, Engame plus Flame and Roundup Ultramax were species-dependent and surfactant-dependent.     

抗草甘膦杂草及其检测方法发展现状

草甘膦在世界范围的多年大量使用已经引起了抗草甘膦杂草的产生。本文针对全球迄今为止发现的21种抗草甘膦杂草的发生、发展状况进行了论述。探讨了抗草甘膦杂草抗药性检测方法,分别从整株生物测定及生物化学等方面介绍了抗草甘膦杂草检测方法的研究现状,为抗草甘膦杂草检测方法的发展及其抗性监测方法的建立提供参考。     

甘氨酸法合成草甘膦中由甲醛引发的副反应研究

利用核磁共振碳谱、磷谱、氢谱对甘氨酸法合成草甘膦的各阶段反应进行定量跟踪分析, 发现由甲醛引起的副反应是影响草甘膦收率的重要因素之一,证实了羟甲基亚瞵酸、增甘膦、N-甲基草甘膦和亚磷酸是甘氨酸法合成草甘膦的主要副产物,减少或抑制其生成是草甘膦合成工艺优 化的关键。探讨了抑制副反应发生的条件,对甘氨酸法合成草甘膦工艺优化有重要的理论指导意义。     

Mobility and leaching of glyphosate: a review

There is currently concern that glyphosate, a strongly sorbing non-selective herbicide which is widely used in Europe, may be leached from the root zone into drainage water and groundwater. The purpose of this review is to present and discuss the state of knowledge with respect to the mobility and leaching of glyphosate from agricultural soils. Specific attention is given to the adsorption behaviour of glyphosate and the analysis of available studies on glyphosate transport. In addition, there are a number of experimental and numerical studies indicating that other strongly sorbing substances may be transported rapidly to the sub-surface. The experimental studies analysed in the paper encompass column-, lysimeter- and field-scale experiments on glyphosate transport. The experimental findings, combined with transport studies on other strongly sorbing pesticides in the literature, support the hypothesis that transport of glyphosate may be caused by an interaction of high rainfall events shortly after application on wet soils showing the presence of preferential flow paths. Concentrations of glyphosate in European groundwater have been reported occasionally but monitoring is still limited.     

Optimizing foliar activity of glyphosate on Bidens frondosa and Panicum maximum with different adjuvant types

The influence of non-ionic (X-77) and organosilicone (L-77) adjuvants and of methylated seed oil (MSO) on the uptake, translocation and efficacy of glyphosate was investigated in Bidens frondosa L. and Panicum maximum Jacq. In addition, the physicochemical properties of adjuvants and adjuvant + glyphosate aqueous solutions were determined. Significantly lower surface tension and contact angle values were obtained with aqueous solutions of L-77 alone and with glyphosate. Over a 48-h time course, it was observed that >50% of applied ¹⁴C-glyphosate was absorbed within 15 min in B. frondosa with L-77. At 6 h and thereafter, ¹⁴C glyphosate absorption was significantly higher with MSO compared with X-77 in B. frondosa . In P. maximum , uptake and translocation of ¹⁴C-glyphosate + adjuvant were increased in general up to 48 h after treatment application, except with L-77, which showed no improvement in uptake – instead there was a significant reduction compared with no treatment with L-77. This indicated its antagonistic effect on this grass species. The lower values of ¹⁴C-glyphosate in P. maximum also confirmed that adjuvant effects were species specific. In the efficacy studies, glyphosate formulated with L-77 achieved significantly higher control of B. frondosa , while there was no control of P. maximum with this treatment. This confirmed antagonism in glyphosate absorption into P. maximum by L-77. Furthermore, significantly higher control of tested plants was recorded with MSO in comparison to X-77, which confirms the solubilizing or humectant nature of MSO.     

Resistance to glyphosate in Lolium rigidum

Annual ryegrass (Lolium rigidum) is a widespread and important weed of Australia and populations of this weed have developed resistance to most major herbicides, including glyphosate. The possible mechanisms of resistance have been examined in one glyphosate-resistant Lolium population. No major differences were observed between resistant and susceptible biotypes in respect of (i) the target enzyme (EPSP synthase), (ii) DAHP synthase, the first enzyme of the target (shikimate) pathway, (iii) absorption of glyphosate, or (iv) translocation. Following treatment with glyphosate, there was greater accumulation of shikimate (derived from shikimate-3-Pi) in susceptible than in resistant plants. In addition, the resistant population exhibited cross-resistance to 2-hydroxy-3-(1,2,4-triazol-1-yl)propyl phosphonate, a herbicide which, although structurally similar to glyphosate, acts at an unrelated target site. On the basis of these observations we speculate that movement of glyphosate to its site of action in the plastid is involved in the resistance mechanism. © 1999 Society of Chemical Industry     

Expression of a wheat cytochrome P450 monooxygenase in yeast and its inhibition by glyphosate

Glyphosate is a non-selective herbicide which acts by inhibiting 5-enolpyruvylshikimate-3-phosphate synthase. Wheat cytochrome P450 monooxygenase specifically catalyzes the metabolism of some sulfonylurea herbicides such as chlorsulfuron and triasulfuron. Here we report that glyphosate is an inhibitor of a wheat cytochrome (CYP71C6v1), the cDNA of which was amplified by RT-PCR and heterologously expressed in yeast. The microsomal fractions derived from this strain had a Soret peak at 502 nm in the reduced carbon monoxide difference spectrum, which is a typical spectral characteristic. The addition of glyphosate to the microsomal protein resulted in a Type II spectrum indicative of binding via the nitrogen group to haem of cytochrome P450 as a sixth ligand. A spectral dissociation constant, K(s) of 70 micromol litre(-1) was observed and an IC50 of 11 micromol litre(-1) was found for glyphosate inhibition of CYP71C6v1 P450 activity.