Sensitivity of Echinochloa muricata and Echinochloa crus-galli to HPPD- and ALS-inhibiting herbicides in corn

Until recently Echinochloa muricata var. microstachya Wiegand (rough barnyardgrass), an alien species native to North America, was completely overlooked in Belgium due to its close morphological resemblance to Echinochloa crus-galli (L.) P. Beauv. (barnyardgrass). E. muricata var. microstachya has gradually spread and is now locally naturalized and abundant in and along maize fields. One of the possible reasons for its expansion in maize fields, besides e.g. the lack of crop rotation, might be a lower sensitivity to postemergence herbicides acting against panicoid grasses, in particular 4-hydroxyphenyl pyruvate dioxygenase (HPPD)-inhibiting herbicides and acetolactate synthase (ALS) inhibiting herbicides. Dose-response pot experiments were conducted in the greenhouse to evaluate the effectiveness of four HPPD-inhibitor herbicides [topramezone (ARIETTA), mesotrione (CALLISTO), tembotrione (LAUDIS), sulcotrione (MIKADO) and the ALS-inhibitor herbicide nicosulfuron (KELVIN) for controlling local populations of E. crus-galli and E. muricata. Pots were planted with 25 seeds, thinned afterwards to 5 plants (one week after sowing) and irrigated by overhead sprinklers. Herbicides were applied at the 3-4 leaf stage (BBCH stage 13-14). Fresh biomass was harvested 28 d after treatment. In another dose-response pot experiment, the influence of leaf stage at time of herbicide application on efficacy of topramezone for (rough) barnyardgrass control was evaluated. Sensitivity to HPPD-inhibitor herbicides topramezone and sulcotrione was significantly lower for E. muricata populations than for E. crus-galli populations. However, nicosulfuron sensitivity of both species was similar. Compared to E. crus-galli, sensitivity of E. muricata to topramezone was more dependent on leaf stage. Due to the intragenus variability in sensitivity to HPPD-inhibitor herbicides, higher awareness is required for presence of E. muricata plants in maize fields in order to avoid insufficient "barnyardgrass" control.     

The effect of cultivation system and soil factors on the performance of herbicides against Alopecurus myosuroides

Aspects of chemical and cultural control of Alopecurus myosuroides (blackgrass) in winter wheat were studied on a naturally-occurring infestation over a 3-yr period. In two experiments a comparison was made between ploughing and direct drilling, whilst in the third experiment direct drilling alone was used. Straw was spread and burnt in each year. Herbicide treatments were: chlortoluron and isoproturon in all three experiments; diclofop-methyl and pendimethalin in two experiments; metoxuron, terbutryne, tri-allate, carbetamide, propyzamide, chlorsulfuron + methabenthiazuron and trifluralin + linuron in one experiment. Herbicides were applied either pre-emergence, early post-emergence or late postemergence to separate sub-plots. Surface soil (0–2.5 cm) samples were collected each year from unsprayed areas and used for soil analyses and for herbicide activity studies in pot experiments. Ploughing resulted in much lower populations of A. myosuroides than direct drilling. Most herbicides gave much better control of A. myosuroides in ploughed than in direct drilled soil in field and pot experiments, regardless of application time. Diclofop-methyl, applied early post-emergence at 1.13 kg a.i./ha, was the only herbicide treatment in field experiments to achieve over 90% weed control in direct drilled soil. Differences in adsorption between soils given different cultivations, determined as Kd values for chlortoluron and isoproturon, were much greater than differences in organic matter. The much greater adsorptive capacity of direct drilled soil, and consequently the poorer performance of all soil-acting herbicides, was attributed to the concentration of burnt straw residues near the soil surface.     

Soil microbial and faunal responses to herbicide tolerant maize and herbicide in two soils

A glasshouse experiment was set up to compare processes and organisms in two soils planted with genetically modified (GM) herbicide tolerant (HT) maize treated with appropriate herbicides. This was part of a wider project (ECOGEN) looking at the consequences of GM cropping systems on soil biology using a tiered approach at laboratory, glasshouse and field scales. Soil for the experiment was taken from field sites where the same maize cultivars were grown to allow comparison between results under glasshouse and field conditions. The maize cultivars T25 (GM HT glufosinate-ammonium tolerant), Orient (non HT near isogenic control for T25) and Monumental (a conventional, non HT variety) were grown in contrasting sandy loam and clay loam soils, half were sprayed with the appropriate herbicide as used in the field and soil samples were taken at the five-leaf and flowering plant growth stage. The main effects on all measured parameters were those of soil type and plant growth stage, with four categories of subsequent interaction: (1) there were no effects of herbicide on plant growth or soil microarthropods: (2) the maize cultivar (but not the GM HT trait) had effects on the decomposition of cotton strips and the nematode community; (3) herbicide application in general altered the community level physiological profile of the microbial community and reduced both soil basal respiration and the abundance of protozoa; and (4) the specific application of glufosinate-ammonium to T25 maize altered soil microbial community structure measured by ester linked fatty acids. The results from this glasshouse experiment support the findings from the field that there are effects of herbicide application on the soil microbial and meso-faunal community but that, compared to other standard agricultural practices, the differences are relatively small.     

Phototransformation of pesticides on plant leaves: the case of sulcotrione

Photochemistry is one of the main ways of pollutants degradation in the environment. There is an obvious lack of data concerning the photostability of agrochemicals on plant foliage. We report here the first photodegradation study of a triketonic herbicide (sulcotrione) used as a substitute of atrazine. To mimic surface Leaves, we used films made of carnauba grey wax or of cuticular wax extracted from maize Leaves. Under simulated solar light, sulcotrione was rapidly phototyzed. The phototysis was much faster on cuticular wax films than in aqueous phase. Moreover, the formulated sulcotrione (Mikado) disappeared even more quickly than the pure active ingredient. Sulcotrione mainly underwent an intramolecular cyclization. To validate these observations, sulcotrione and its main photoproduct were monitored after maize field treatment. The day after the treatment the main photoproduct was detected on the maize leaves and suLcotrione had already significantly disappeared. This experiment demonstrates that phototysis can be a relevant process in real conditions after field treatment.     

Weed species diversity and community composition in cropping practices at two intensity levels – a six-year experiment

Alternative cropping practices can be expected to increase the diversity and change the species composition of weed communities. In a six-year field experiment we compared species diversity and species composition in the weed community of cereal fields cultivated by conventional and by low-input cropping methods. In the conventional cropping both mineral fertilizers and herbicides were applied but in the low-input cropping the fertlizer was manure and herbicides were avoided altogether. The level of nitrogen fertilization was higher in conventional cropping. A five-year crop rotation comprising rye, oat-pea, barley and two-year grasslands was used in both cropping methods. We found that the species pool was larger (42 vs 35 species) and the average number of species higher (20.2 vs 13.1 species) in low-input than in conventional cropping. This was the case both in the data pooled across crops and in each crop separately. Herbicides affected species diversity and species composition more than did nitrogen fertilization. Species susceptible to the herbicides applied declined in abundance, which resulted in more skewed dominance patterns in the communities of conventional fields than in those of low-input fields. The crop affected the species composition of weed communities more than did the cropping method. Our results suggest that weed species diversity could be promoted by using low-intensity cropping practices.     

除草剂对桉树人工林下植物及土壤微生物群落的影响

除草剂在桉树人工林中的应用越来越普遍,但关于除草剂对桉树人工林林下植物和土壤微生物群落的影响知之甚少。通过桉树人工林低剂量高频率(LHF)、中剂量中频率(MMF)、高剂量低频率(HLF)除草剂喷施试验,并与人工除草(MT)为对照,比较分析不同剂量、不同频率除草剂施用对林下植物和土壤微生物群落的影响。结果表明,施用除草剂导致桉树人工林林下植物种类和功能群组成发生显著变化,但并未显著降低林下植物群落物种丰富度和多样性,随除草剂施用频率的降低及恢复时间的增加,物种丰富度及多样性指数呈恢复趋势。除草剂施用也导致土壤养分含量降低。除草剂通过对林下植物群落和土壤养分的负面影响间接影响土壤微生物群落。LHF显著降低藤本植物而显著提高蕨类植物功能群的重要值,从而显著降低了微生物群落、真菌和放线菌的磷脂脂肪酸(PLFA)含量。MMF显著降低木本和藤本植物而显著提高禾草植物功能群的重要值,导致土壤微生物群落和放线菌的PLFA含量显著降低。HLF未显著影响林下植物及土壤微生物群落,但土壤全磷含量显著降低,速效磷含量也大幅下降。施用除草剂显著降低了土壤微生物生物量碳、氮的含量。因此,生产上应减少除草剂的施用,...     

农作物抗除草剂遗传工程研究进展

控制杂草提高农作物产量是农业生产中共同面临的问题,发展抗除草剂农作物将是最经济最方便控制杂草的技术。由于对除草剂的作用模式和除草剂代谢途径的了解,弄清了除草剂的关键靶酶及其基因,因此分离除草剂靶酶基因,克隆能解毒除草剂的酶基因,通过转化技术可获得抗除草剂农作物,大量的抗除草剂转基因农作物大田试验表明,将最有希望在２０００年进入市场。     

Novel bacterial bioassay for a high-throughput screening of 4-hydroxyphenylpyruvate dioxygenase inhibitors

Plant 4-hydroxyphenylpyruvate dioxygenase (HPPD) is the molecular target of a range of synthetic β-triketone herbicides that are currently used commercially. Their mode of action is based on an irreversible inhibition of HPPD. Therefore, this inhibitory capacity was used to develop a whole-cell colorimetric bioassay with a recombinant Escherichia coli expressing a plant HPPD for the herbicide analysis of β-triketones. The principle of the bioassay is based on the ability of the recombinant E. coli clone to produce a soluble melanin-like pigment, from tyrosine catabolism through p-hydroxyphenylpyruvate and homogentisate. The addition of sulcotrione, a HPPD inhibitor, decreased the pigment production. With the aim to optimize the assay, the E. coli recombinant clone was immobilized in sol–gel or agarose matrix in a 96-well microplate format. The limit of detection for mesotrione, tembotrione, sulcotrione, and leptospermone was 0.069, 0.051, 0.038, and 20 μM, respectively, allowing to validate the whole-cell colorimetric bioassay as a simple and cost-effective alternative tool for laboratory use. The bioassay results from sulcotrione-spiked soil samples were confirmed with high-performance liquid chromatography.     

Influence of temperature on phytotoxicity of triazine herbicides to pine seedlings

The effects of temperature, over a range of 5 to 30 C, on phytotoxicity of simazine, atrazine, propazine, prometryne, prometone, and ipazine to young Pinus resinosa seedlings were investigated in growth chambers. Herbicides were applied to the soil surface and then mixed into the soil before pine seeds were planted. Development of recently germinated seedlings was then studied for 7 weeks. High temperatures greatly accelerated herbicide toxicity, but the effects of temperature varied greatly among herbicides. Atrazine and simazine were more toxic than other herbicides tested at all temperatures. Toxicity of simazine and atrazine was apparent early, whereas effects of propazine, prometryne, prometone, and ipazine were somewhat delayed. After 7 weeks maximum dry-weight production of shoots under each herbicide treatment and control occurred at 20 C, with some decreases noted at lower temperatures and marked decreases at progressively higher ones. At 20 C final seedling dry weights following treatment with simazine or atrazine were only one-third as high as in control plants. Growth was also reduced in lesser amounts by propazine, prometryne, prometone, or ipazine. Variations in phytotoxicity of different triazine herbicides appeared to be related more to their structural differences than their solubilities. Under the constant environmental conditions of the experiments, toxicity symptoms in plants treated with triazine herbicides appeared more rapidly and decisively than in previous field experiments under fluctuating environments. The influence of high temperatures in enhancing triazine toxicity appeared to involve complex interactions of physiological activity of plants and temperature effects on herbicide uptake.     

Vegetated Ditches for the Mitigation of Pesticides Runoff in the Po Valley

In intensive agricultural systems runoff is one of the major potential diffuse pollution pathways for pesticides and poses a risk to surface water. Ditches are common in the Po Valley and can potentially provide runoff mitigation for the protection of watercourses. The effectiveness depends on ditch characteristics, so there is an urgent need for site-specific field trials. The use of a fugacity model (multimedia model) can allows recognition of the mitigation main processes. A field experiment was conducted in order to evaluate the mitigation capacity of a typical vegetated ditch, and results were compared with predictions by a fugacity model. To evaluate herbicide mitigation after an extreme runoff, the ditch was flooded with water containing mesotrione, S-metolachlor and terbuthylazine. Two other subsequent floods with uncontaminated water were applied 27 and 82 days later to evaluate herbicides release. Results show that the ditch can immediately reduce runoff concentration of herbicides by at least 50% even in extreme flooding conditions. The half-distances were about 250 m. As a general rule, a runoff of 1 mm from 5 ha is mitigated by 99% in 100 m of vegetated ditch. Herbicides retention in the vegetated ditch was reversible, and the second flood mobilized 0.03-0.2% of the previous one, with a concentration below the drinking water limit of 0.1 μg L^-1. No herbicide was detected in the third flood, because the residual amount in the ditch was too low. Fugacity model results show that specific physical-chemical parameters may be used and a specific soil-sediment-plant compartment included for modelling herbicides behaviour in a vegetated ditch, and confirm that accumulation is low or negligible for herbicides with a half-life of 40 days or less. Shallow vegetated ditches can thus be included in a general agri-environment scheme for the mitigation of pesticides runoff together with wetlands and linear buffer strips. These structures are present in the landscape, and their environmental role can be exploited by proper management.     

Herbicide Selection Promotes Antibiotic Resistance in Soil Microbiomes

Herbicides are one of the most widely used chemicals in agriculture. While they are known to be harmful to nontarget organisms, the effects of herbicides on the composition and functioning of soil microbial communities remain unclear. Here we show that application of three widely used herbicides—glyphosate, glufosinate, and dicamba—increase the prevalence of antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs) in soil microbiomes without clear changes in the abundance, diversity and composition of bacterial communities. Mechanistically, these results could be explained by a positive selection for more tolerant genotypes that acquired several mutations in previously well-characterized herbicide and ARGs. Moreover, herbicide exposure increased cell membrane permeability and conjugation frequency of multidrug resistance plasmids, promoting ARG movement between bacteria. A similar pattern was found in agricultural soils across 11 provinces in China, where herbicide application, and the levels of glyphosate residues in soils, were associated with increased ARG and MGE abundances relative to herbicide-free control sites. Together, our results show that herbicide application can enrich ARGs and MGEs by changing the genetic composition of soil microbiomes, potentially contributing to the global antimicrobial resistance problem in agricultural environments.     

Cytochrome P450 CYP81A10v7 in Lolium rigidum confers metabolic resistance to herbicides across at least five modes of action

Rapid and widespread evolution of multiple herbicide resistance in global weed species endowed by increased capacity to metabolize (degrade) herbicides (metabolic resistance) is a great threat to herbicide sustainability and global food production. Metabolic resistance in the economically damaging crop weed species Lolium rigidum is well known but a molecular understanding has been lacking. We purified a metabolic resistant (R) subset from a field evolved R L. rigidum population. The R, the herbicide susceptible (S) and derived F₂ populations were used for candidate herbicide resistance gene discovery by RNA sequencing. A P450 gene CYP81A10v7 was identified with higher expression in R vs. S plants. Transgenic rice overexpressing this Lolium CYP81A10v7 gene became highly resistant to acetyl-coenzyme A carboxylase- and acetolactate synthase-inhibiting herbicides (diclofop-methyl, tralkoxydim, chlorsulfuron) and moderately resistant to hydroxyphenylpyruvate dioxygenase-inhibiting herbicide (mesotrione), photosystem II-inhibiting herbicides (atrazine and chlorotoluron) and the tubulin-inhibiting herbicide trifluralin. This wide cross-resistance profile to many dissimilar herbicides in CYP81A10v7 transgenic rice generally reflects what is evident in the R L. rigidum. This report clearly showed that a single P450 gene in a cross-pollinated weed species L. rigidum confers resistance to herbicides of at least five modes of action across seven herbicide chemistries.     

Generation and characterization of soybean and marker-free tobacco plastid transformants over-expressing a bacterial 4-hydroxyphenylpyruvate dioxygenase which provides strong herbicide tolerance

Plant 4-hydroxyphenylpyruvate dioxygenase (HPPD) is part of the biosynthetic pathway leading to plastoquinone and vitamin E. This enzyme is also the molecular target of various new bleaching herbicides for which genetically engineered tolerant crops are being developed. We have expressed a sensitive bacterial hppd gene from Pseudomonas fluorescens in plastid transformants of tobacco and soybean and characterized in detail the recombinant lines. HPPD accumulates to approximately 5% of total soluble protein in transgenic chloroplasts of both species. As a result, the soybean and tobacco plastid transformants acquire a strong herbicide tolerance, performing better than nuclear transformants. In contrast, the over-expression of HPPD has no significant impact on the vitamin E content of leaves or seeds, quantitatively or qualitatively. A new strategy is presented and exemplified in tobacco which allows the rapid generation of antibiotic marker-free plastid transformants containing the herbicide tolerance gene only. This work reports, for the first time, the plastome engineering for herbicide tolerance in a major agronomic crop, and a technology leading to marker-free lines for this trait.     

Herbicide safeners and glutathione metabolism

Herbicide safeners are chemicals which protect crop plants from injury by certain herbicides, without affecting weed control efficacy of the herbicides. The protective mechanism of herbicide safeners has not yet been fully elucidated, but there is increasing evidence that safeners act by selectively enhancing herbicide detoxification in crop plants. To date, two main detoxification pathways have been related to the mode of action of herbicide safeners. The first includes oxidation and subsequent glucose conjugation, mediated by cytochrome P450 -dependent monooxygenases and UDP-glucosyltransferases, respectively. This pathway appears to be important predominantly in safener protection to aryloxyphenoxypropionate and sulfonylurea herbicides. The second pathway represents the conjugation of thiocarbamate sulfoxides and chloroacetanilide herbicides with glutathione. This mechanism is accomplished by either elevating the levels of reduced glutathione or the activity of glutathione S-transferase, or both. Since glutathione has been reported to be involved in several stress situations of plants its function associated with safener-induced herbicide tolerance will be discussed in more detail in this review.     

Growth abilities and phenotype stability of a sulcotrione-degrading Pseudomonas sp. isolated from soil

Pseudomonas sp. 1OP, previously isolated from a French agricultural soil, has been described as the first sulcotrione degrading bacteria. Different conditions of initial pH and herbicide concentration in liquid culture were tested to evaluate the growth performances of the isolate and its degrading capacity, with sulcotrione as the sole carbon and/or energy source. Maximal growth rate (μmax) was obtained under initial neutral conditions and with initial concentration of sulcotrione close to 180 μM, and was described, during the exponential phase, by a sigmoidal curve which could be easily fitted to the modified Gompertz equation. Complementary studies carried on the CMBA by-product and on another β-triketone herbicide showed the relative specificity of the strain against sulcotrione. The sulcotrione degrading phenotype of Pseudomonas sp.1OP was shown to be lost under non-selective conditions. Plasmid-Eckardt modified method, consecutively applied for plasmid profiling, showed that this strain carries one large plasmid (>12 kb) bearing putative genes involved in sulcotrione degradation, as demonstrated by curing experiment.     

Outdoor evaluation of herbicide resistant strains of Anabaena variabilis as biofertilizer for rice plants

Application of diazotrophic cyanobacteria, Anabaena variabilis, as biofertilizer for rice cultivation has a beneficial effect on crop productivity and maintenance of soil fertility. However, periodic applications of herbicides used to obtain high crop productivity are not only detrimental to weeds but to biofertilizer strains of cyanobacteria also. Therefore, research was undertaken to isolate four herbicide resistant strains (Arozin-R, Alachlor-R, Butachlor-R and 2,4-D-R) and a multiple herbicide resistant strain (MHR) of natural isolates of A. variabilis exhibiting resistance against these common rice field herbicides. The outdoor survivability of mutant strains and the productivity of rice crop (IR-36) were evaluated by inoculating the wild type and herbicide resistant mutant strains of A. variabilis in the presence and absence of recommended field dosages of test herbicides. No difference in survival and biofertilizer potentials of the herbicide resistant strains was observed in herbicide treated or in untreated conditions. Highest survivability (87%) was exhibited by MHR relative to other mutants. Highest growth and grain yield (76%) were recorded in plants treated with MHR as compared to uninoculated control rice plants. In conclusion, the mutant strains of A. variabilis had stable resistance to herbicides under outdoor conditions in flooded soils. Not only did the herbicide resistance strains increase growth of rice relative to the uninoculated pots, they were more beneficial for rice growth than the wild type strain. Responsible Editor: Richard W. Bell.     

Safener responsiveness and multiple herbicide resistance in the weed black-grass (Alopecurus myosuroides)

Safeners enhance the selectivity of graminicidal herbicides such as fenoxaprop ethyl in cereals, by increasing their rates of detoxification in the crop. While studying the selectivity of fenoxaprop ethyl in wheat, we determined that the safeners mefenpyr diethyl and fenchlorazole ethyl also enhanced herbicide tolerance in the competing weed black-grass ( Alopecurus myosuroides ). Fenoxaprop ethyl was detoxified by conjugation with glutathione in both wheat and black-grass, with the resulting metabolites processed to the respective cysteine derivatives, which were then N -glycosylated. In black-grass, these detoxification pathways were only slightly enhanced by safeners, suggesting that metabolism alone was unlikely to account for increased herbicide tolerance. Instead, it was determined that safening was associated with an accumulation of glutathione and hydroxymethylglutathione and enzymes with antioxidant functions including phi and lambda glutathione transferases, active as glutathione peroxidases and thiol transferases respectively. These safener-induced changes closely mirrored those determined in two independent black-grass populations that had acquired multiple herbicide resistance (MHR) in the field. In addition to enhanced glutathione metabolism, both safener treatment and MHR resulted in elevated levels of flavonoids in the foliage of black-grass plants, notably flavone- C -glycosides and anthocyanins. Our results demonstrate that safening in a grass weed is associated with an inducible activation in antioxidant and secondary metabolism which mirrors the biochemical phenotype exhibited in plants that are resistant to multiple classes of herbicides.     

Comparative study of the effect of selected agrochemical products on Steinernema feltiae (Rhabditida: Steinernematidae)

The effect of three neurotoxic insecticides, three photosynthetic inhibitor herbicides and three enzymatic inhibitor herbicides on infective juveniles (IJs) of Steinernema feltiae Rioja (native) and ENTONEM® (commercial) strains were evaluated after a 48-h exposure at field tank concentrations and overnight treatment in mQ-water, using Spodoptera littoralis as target. Nematode survival was not affected by acetyl-cholinesterase inhibitors chlorpyrifos and pirimicarb, although chlorpyrifos seriously reduced their virulence. Both nematode strains showed differential sensitivity to cypermethrin, which affects the sodium channels of the nerve membrane, with the ENTONEM® strain being more tolerant than Rioja strain. However, these chemicals showed a strong sublethal effect on the nematode reproductive potential, limiting seriously their possible recycling in the field. Herbicides showed differential toxic effects on nematode survival. The commercial strain was tolerant to enzymatic inhibitor herbicides, whereas tribenuron and chlorsulfuron reduced Rioja strain survival. However, photosynthetic inhibitor herbicides severely affected survival of both nematode strains, with the Rioja strain being more sensitive. Sublethal effects on both nematode strains were observed only after exposition to terbutryn+chlortoluron+triasulfuron, increasing the time to kill insect larvae. These results are useful to optimize EPN dosages and to estimate their field recycling.     

A novel approach to the use of genetically modified herbicide tolerant crops for environmental benefit

The proposed introduction of genetically modified herbicide tolerant (GMHT) crops, with claims of improved weed control, has prompted fears about possible environmental impacts of their widespread adoption, particularly on arable weeds, insects and associated farmland birds. In response to this, we have developed a novel weed-management system for GMHT sugar beet, based on band spraying, which exploits the flexibility offered by the broad-spectrum partner herbicides. Here, we show the results from two series of field experiments which, taken together, demonstrate that, by using this system, crops can be managed for enhanced weed and insect biomass without compromising yield, thus potentially offering food and shelter to farmland birds and other wildlife. These results could be applicable widely to other row crops, and indicate that creative use of GMHT technology could be a powerful tool for developing more sustainable farming systems in the future.