Mechanisms of Resistance to Aryloxyphenoxypropionate Herbicides in Two Resistant Biotypes ofAlopecurus myosuroides(blackgrass): Herbicide Metabolism as a Cross-Resistance Mechanism

The mechanisms of AOPP herbicide resistance in twoAlopecurus myosuroidesbiotypes were investigated. Resistant biotype Peldon A1, which is highly resistant to the phenyl-urea chlorotoluron, is moderately resistant to the AOPP herbicides diclofop-methyl, fenoxaprop-ethyl, fluazifop-P-butyl, and the CHD tralkoxydim. Resistant biotype Lincs. E1, which is only moderately resistant to chlorotoluron, is highly resistant to the AOPP herbicide fenoxaprop-ethyl, and moderately resistant to diclofop-methyl, fluazifop-P-butyl, and the CHD tralkoxydim. There is no clear evidence of resistance to the CHD sethoxydim in either biotype. Both Peldon A1 and Lincs. E1 exhibited moderately enhanced metabolism of diclofop-methyl. The approximate half life of diclofop was 8 and 9 HAT, respectively, compared to 17 HAT for the susceptible Rothamsted biotype. Peldon A1 showed moderately enhanced metabolism of fenoxaprop-P-ethyl. However, in the highly resistant Lincs. E1, fenoxaprop-P-ethyl metabolism rates were intermediate between Peldon A1 and the susceptible biotype. Fenoxaprop-P-ethyl metabolism inA. myosuroideswas not significantly reduced by inhibitors of cytochrome P450: PBO, tetcyclasis, or ABT. While enhanced herbicide metabolism can account for the moderate AOPP/CHD resistance observed in Peldon A1in vivo, it cannot account in total for fenoxaprop-ethyl resistance in Lincs. E1. Lincs. E1 may possess one or more additional resistance mechanism.     

Characterisation of Multiple Glutathione Transferases Containing the GST I Subunit with Activities toward Herbicide Substrates in Maize (Zea mays)

The glutathione transferases (GSTs) of maize with activities toward chloroacetanilide herbicides are relatively well characterised, but their range of substrate specificities has not been determined in detail. GST activities toward an extensive range of chemically diverse xenobiotic substrates, including the herbicides atrazine, alachlor, metolachlor and fluorodifen, have been determined in crude and purified preparations from the roots and shoots of dark-grown maize seedlings treated with and without the herbicide-safener dichlormid. With the exception of the activity toward atrazine, specific activities were higher in the roots than in the shoots in all cases. In untreated shoots activities were in the order atrazine=alachlor=metolachlor>fluorodifen with safener–treatment selectively increasing the activity toward the chloroacetanilides and fluorodifen. In the roots the highest GST activities toward herbicides were toward the chloroacetanilides. Dichlormid treatment resulted in an increase in activities toward all four herbicides in the roots of one maize cultivar (Pioneer 3394) but only enhanced the activities toward the chloroacetanilides and fluorodifen in cultivar Artus. Using the non-herbicide 1-chloro-2,4-dinitrobenzene (CDNB) as substrate, anion-exchange chromatography showed that the roots and shoots contained a similar range of GST isoenzymes. All of these isoenzymes were enhanced in response to safeners, though the extent of this induction was organ-dependent. GST isoenzymes containing the GST I subunit were purified from safener-treated roots by a combination of hydrophobic interaction chromatography and affinity chromatography using Orange A agarose. Three isoenzymes could then be purified following resolution by anion-exchange chromatography. The three GSTs were termed GST I/I, GST I/II and GST I/III with GST I, II and III referring to the presence of 29 kDa, 27 kDa and 26 kDa subunits respectively. This revised nomenclature for the maize GSTs was considered necessary in view of the continued discovery of new isoenzymes, such as GST I/III, composed of subunits which have been previously described. GST I/I had measurable activity toward atrazine, low activities toward the other herbicides and appreciable activities toward a range of other xenobiotic substrates. GST I/II and the novel GST I/III isoenzymes both showed high activities toward the chloroacetanilides and fluorodifen but lower activities toward the other substrates and negligible activities toward atrazine. The GST II subunit of GST I/II also had activity as a glutathione peroxidase. Our results show that the GST I subunit can form dimers with the GST III subunit in addition to the GST I and GST II subunits and that the degree of specificity toward herbicide substrates of the respective isoenzymes is greater than previously reported. Our results also suggest that the safener-inducible GST II subunit has additional activities as a glutathione peroxidase. © 1997 SCI.     

Glutathione Transferase Activities and Herbicide Selectivity in Maize and Associated Weed Species

The role of glutathione transferases (GSTs) in the selectivity of the herbicides alachlor, atrazine, fluorodifen and metolachlor, which are detoxified by glutathione conjugation in plants, was determined in seedlings of maize (Zeamays L.) and the associated weed species Abutilon theophrasti Medic., Digitariasanguinalis (L.) Scop. Echinochloa crus-galli (L.) Beauv., Panicum miliaceum (L.), Setaria faberi Herrm. and Sorghum bicolor (L.) Moench. The availability of glutathione was also determined in all species and tissue concentrations were found to be in the range 120–160 μm for all species except D. sanguinalis and S. bicolor, which contained half this amount. GST activities toward the herbicides were determined in crude protein extracts from the plants using HPLC to quantify the biosynthesis of the herbicide conjugates. The specific activities of the GSTs toward the substrates were in the order alachlor>fluorodifen> atrazine>metolachlor in all species except A. theophrasti, where fluorodifen was a better substrate than alachlor. In most cases there was a good correlation between GST activities and the selectivity of the herbicides applied pre-emergence. In the case of atrazine, GST activities were also related to the relative rates of herbicide conjugation in vivo. In contrast, there was no simple relationship between glutathione availability and the selectivity of the herbicides. However, with alachlor there was evidence that glutathione availability was limiting GST activity and influencing tolerance.     

Glutathione transferases and herbicide detoxification in suspension-cultured cells of giant foxtail (Setaria faberi)

Glutathione transferases (GSTs) catalysing the conjugation of 1-chloro-2,4-dinitrobenzene, the chloro-s-triazine herbicide atrazine, the chloroacetanilide herbicides metolachlor and alachlor and the diphenyl ether herbicide fluorodifen have been identified in suspension-cultured cells derived from the grass weed giant foxtail (Setaria faberi Herrm.). In contrast to suspension-cultured cells of maize, where atrazine-conjugating GSTs are lost during de-differentiation, the GSTs active toward this herbicide in S. faberi plants were also expressed in cultures, suggesting that these isoenzymes are subject to different regulation in the crop and weed. As a result, glutathione conjugation was the major route of atrazine metabolism in S. faberi cultures. Activities of these GSTs were maximal three days after sub-culturing when the cells were dividing most actively, when they were determined to be in the order CDNB>alachlor>metolachlor= fluorodifen>atrazine. This indicated that GSTs which are enhanced during cell division can metabolise herbicides. On the basis of activity per mg protein, GST activities in the cultures were between 20 and 60-fold higher than those determined in the foliage of S. faberi seedlings. The GSTs with activity towards CDNB were resolved into three peaks following anion-exchange chromatography at pH 7·8 using Q-Sepharose. Peak 1 GSTs were not retained, while peak 2 and peak 3 were sequentially resolved with an increasing concentration of salt. Peak 1 GSTs showed activity toward metolachlor and atrazine but showed little activity toward fluorodifen. Peak 2 and peak 3 GSTs were active toward atrazine and metolachlor, with peak 3 being particularly associated with activity toward fluorodifen. The GSTs in these peaks were then further purified using S-hexyl-glutathione-agarose affinity chromatography. In each case, the affinity-bound fraction of the GSTs consisted of 28 kDa and 26 kDa polypeptides, suggesting that the GST isoenzymes in S. faberi cultures are composed of related subunits. Our results demonstrate that the GST isoenzymes involved in herbicide metabolism in suspension cultures of a grass weed show a similar level of complexity to that determined in maize cell cultures. © 1998 SCI     

Glutathione transferase activities toward herbicides used selectively in soybean

Using extracts from suspension-cultured cells of soybean (Glycine max cv. Mandarin) as a source of active enzymes, the activities of glutathione transferases (GSTs) catalysing the conjugation of 1-chloro-2,4-dinitrobenzene (CDNB) and selective herbicides were determined to be in the order CDNB≫ fomesafen>metolachlor=acifluorfen>chlorimuron-ethyl. GST activities showed a thiol dependence in a substrate-specific manner. Thus, GST activities toward acifluorfen and fomesafen were greater when homoglutathione (hGSH), the endogenously occurring thiol in soybean, was used as the co-substrate rather than glutathione (GSH). Compared with GSH, hGSH addition either reduced or had no effect on GST activities toward other substrates. In the absence of enzyme, the rates of hGSH conjugation with acifluorfen, chlorimuron-ethyl and fomesafen were negligible, suggesting that rapid hGSH conjugation in soybean must be catalysed by GSTs. GST activities were subsequently determined in 14-day-old plants of soybean and a number of annual grass and broadleaf weeds. GST activities of the plants were then related to observed sensitivities to post-emergence applications of the four herbicides. When enzyme activity was expressed on a mg^-1 protein basis, all grass weeds and Abutilon theophrasticontained considerably higher GST activity toward CDNB than soybean. With fomesafen as the substrate, GST activities were determined to be in the order soybean≫Echinochloa crus-galli>Digitaria sanguinalis>Sorghum halepense=Setaria faberi with none of the broadleaf weeds showing any activity. This order related well to the observed selectivity of fomesafen, with the exception of A. theophrasti, which was partially tolerant to the herbicide. Using metolachlor as the substrate the order of the GST activities was soybean>A. theophrasti≫S. halepense>Amaranthus retroflexus>Ipomoea hederacea, with the remaining species showing no activity. GST activities toward metolachlor correlated well with the selectivity of the herbicide toward the broadleaf weeds but not toward the grass weeds. Acifluorfen and chlorimuron-ethyl were selectively active on these species, but GST activities toward these herbicides could not be detected in crude extracts from whole plants. © 1997 SCI     

New, quick tests for herbicide resistance in black-grass (Alopecurus myosuroides Huds) based on increased glutathione S-transferase activity and abundance

Black-grass (Alopecurus myosuroides Huds) is a major grass weed in winter cereals in Europe. It reduces yields and can act as a secondary host for a range of diseases. Herbicide resistance in this species was first detected in the UK in the early 1980s, and has now been reported in thirty counties. To successfully manage herbicide resistance it is vital that suspect populations are tested so that appropriate action can be taken. Ideally, a test will be quick, cheap and easy to use. Furthermore, it should provide an unequivocal result before post-emergence herbicides are to be applied, allowing alternative strategies to be adopted where necessary. This paper reports the development of new tests for herbicide resistance based on our observation that the resistant black-grass biotype Peldon contains approximately double the activity of the enzyme glutathione S-transferase (GST) compared with susceptible biotypes. Data are presented on the production of a monoclonal antiserum to a novel 30 kDa GST polypeptide purified from the biotype Peldon. An ELISA using this antiserum is described and the utility of this assay to detect resistant black-grass biotypes in plants grown under glass and in the field is presented. In addition, a microtitre assay for GST activity is described, which allows the rapid assessment of GST activities of plants. Both abundance and activity of GSTs are discussed as markers for herbicide resistance in black-grass.     

Purification,characterization and comparison of glutathione S-transferases from black-grass (Alopecurus myosuroides Huds) biotypes

In the UK biotypes of black-grass (Alopecurus myosuroides Huds) showing resistance to both chlorotoluron (CTU) and aryloxyphenoxypropionate graminicides are increasingly being observed. Although the precise mechanisms involved in this resistance have yet to be identified, increased herbicide metabolism has been implicated as being involved in at least some cases of resistance. Glutathione S-transferases (GSTs) are a group of enzymes which have been demonstrated to metabolise herbicides in some plants, and the resistant black-grass biotype Peldon contains approximately double the GST activity towards 1-chloro-2,4-dinitrobenzene (CDNB) of susceptible biotypes. To investigate further the possible role of GSTs in herbicide resistance in black-grass, a purification procedure has been developed for these enzymes. A 27.5 kDa polypeptide possessing GST activity was purified from the susceptible biotype Herbiseed. Purification of GSTs from the resistant biotype Peldon also identified this polypeptide along with an additional 30 kDa polypeptide. An in-vitro kinetic study of both crude and purified GST extracts, and western blot analysis using antisera raised against the 27.5 kDa polypeptide, suggest that the 30 kDa polypeptide may possess GST activity, and is not a precursor of the 27.5 kDa polypeptide. These results are discussed and compared to GST profiles for other weeds and crops demonstrating herbicide resistance or tolerance. © 1999 Society of Chemical Industry     

Phytotoxicity of trifluoromethyl- and methyl-substituted dinitroaniline herbicides on resistant and susceptible populations of black-grass (Alopecurus myosuroides)

A Petri-dish assay was used to determine the phytotoxicities of the dinitroaniline herbicides pendimethalin, trifluralin, benfluralin and ethalfluralin, and the methyl-substituted analogues of the last three herbicides, on susceptible (Rothamsted) and multiple-herbicide resistant (Peldon) populations of blackgrass Alopecurus myosuroides Huds. The Peldon population showed a high degree of resistance to pendimethalin, which possesses a 3,4-dimethyl substitution. The two populations were equally sensitive to trifluralin, benfluralin and ethalfluralin which do not possess ring-methyl groups but contain a 4-trifluoromethyl group. Substitution of the 4-trifluoromethyl with a methyl group, as demonstrated by the analogues, reduced phytotoxicity, but to a much greater degree in Peldon than in the Rothamsted population. The study indicates that resistance to pendimethalin in the Peldon population is attributable to an oxidative degradation of the 4-methyl group, analogous to that which occurs with resistance to chlorotoluron. The lack of cross-resistance to the other dinitroaniline herbicides appears to be due to the absence of ring-methyl or other groups which are vulnerable to oxidative degradation, trifluoromethyl groups being particularly resistant to this form of reaction.     

Recombinant glutathione transferases from flufenacet-resistant black-grass (Alopecurus myosuroides Huds.) form different flufenacet metabolites and differ in their interaction with pre- and post-emergence herbicides

BACKGROUND

Black-grass (Alopecurus myosuroides Huds.) has become a problematic weed in cereals in Europe. Besides resistance to post-emergent herbicides becoming increasingly widespread, enhanced metabolism of inhibitors of the synthesis of very-long-chain fatty acids (VLCFAs), such as flufenacet, is evolving. Yet, cross-resistance patterns and evolution of this resistance remains poorly understood.

RESULTS

The cDNA sequences of five glutathione transferases (GSTs) upregulated in flufenacet resistant black-grass were identified and used for recombinant protein expression. Moderate to slow detoxification of flufenacet was verified for all candidate GSTs expressed in E. coli, and the most active protein produced flufenacet-alcohol instead of a glutathione conjugate, in the presence of reduced glutathione (GSH). Moreover, cross-resistance to other VLCFA-inhibitors e.g., acetochlor and pyroxasulfone and the ACCase inhibitor fenoxaprop was verified in vitro. Various other herbicides of different modes of action including VLCFA-inhibitors were not detoxified by the candidate GSTs.

CONCLUSIONS

As several in planta upregulated GSTs detoxified flufenacet in vitro, the shift in sensitivity observed in black-grass populations, is likely a result of an additive effect. The polygenic character and the relatively low turnover rate of the individual GSTs may explain the slow evolution of flufenacet resistance. In addition, flufenacet resistance was accompanied by cross-resistance with some, but not all, herbicides of the same mode of action, and furthermore to the ACCase inhibitor fenoxaprop-ethyl. Hence, not only the rotation of herbicide modes of action, but also of individual active ingredients is important for resistance management. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.     

Activity of glutathione S-transferase toward some herbicides and its regulation by benoxacor in non-embryogenic callus and in vitro regenerated tissues of Zea mays

A procedure was developed to obtain non-embryogenic callus and regenerated lines from root segments of Zea mays grown in aseptic conditions. The activity of glutathione-S-transferases (GSTs), for non-embryogenic callus, was determined toward 1-chloro-2,4-dinitrobenzene (CDNB) and it was compared with that obtained for corn seedlings grown without hormones. For the callus masses, increases of specific activity toward CDNB and the kinetic parameter V_max were observed with respect to corn seedlings. The procedure permitted the regenerating of tissues from callus explants, therefore the GST(CDNB) activity and the effect of the safener benoxacor on its expression were investigated for the regenerated tissues grown in agarized substrate and in liquid medium. These explants showed a constitutive GST(CDNB) activity higher than corn seedlings and this activity was increased, for both tissues, in response to the presence of the safener benoxacor in the growing medium. The GST activity for the above tissues was also assayed toward benoxacor and terbuthylazine, metolachlor and fluorodifen herbicides. Measurable GST activity was found toward some of the above chemicals and it was found to be significantly enhanced in response to benoxacor treatment.     

不同种群小菜蛾乙酰胆碱酯酶性质研究

小菜蛾乙酰胆碱酯酶(AChE)的性质研究表明,不同种群小菜蛾成虫与幼虫AChE活性相比,差异极显著,成虫是幼虫的5～7倍;武汉种群小菜蛾成虫和幼虫AChE与底物的亲和力最小。小菜蛾谷胱甘肽鄄S鄄转移酶(GST)的性质研究表明,宣化、武汉、云南3个田间种群GST活性较高,明显高于台湾敏感品系;北京种群GST活性较低,与台湾敏感品系近似。此研究为进一步分析小菜蛾不同种群、不同虫态抗药性差异提供了依据。     

Multiple origins for black-grass (Alopecurus myosuroides Huds) target-site-based resistance to herbicides inhibiting acetyl-CoA carboxylase

We have investigated the process of evolution of target-site-based resistance to herbicides inhibiting acetyl-CoA carboxylase (ACCase) in nine French populations of black-grass (Alopecurus myosuroides Huds). To date, two different ACCase resistant alleles are known. One contains an isoleucine-to-leucine substitution at position 1781, the second contains an isoleucine-to-asparagine substitution at position 2041. Using phylogenetic analysis of ACCase sequences, we showed that 1781Leu ACCase alleles evolved from four independent origins in the nine black-grass populations studied, while 2041Asn ACCase alleles evolved from six independent origins. No geographical structure of black-grass populations was revealed. This implies that these populations, although geographically distant, are, or have until recently been, connected by gene flows. Comparison of biological data obtained from herbicide sensitivity bioassay and molecular data showed that distinct resistance mechanisms often exist in a single black-grass population. Accumulation of different resistance mechanisms in a single plant was also demonstrated. We conclude that large-scale evolution of resistance to herbicides in black-grass is a complex phenomenon, resulting from the independent selection of various resistance mechanisms in local black-grass populations undergoing contrasted herbicide and agronomical selection pressures, and connected by gene flows whose parameters remain to be determined.     

Control by glyphosate and its alternatives of glyphosate‐susceptible and glyphosate‐resistant Echinochloa colona in the fallow phase of crop rotations in subtropical Australia

Echinochloa colona is the most common grass weed of summer fallows in the grain‐cropping systems of the subtropical region of Australia. Glyphosate is the most commonly used herbicide for summer grass control in fallows in this region. The world's first population of glyphosate‐resistant E. colona was confirmed in Australia in 2007 and, since then, >70 populations have been confirmed to be resistant in the subtropical region. The efficacy of alternative herbicides on glyphosate‐susceptible populations was evaluated in three field experiments and on both glyphosate‐susceptible and glyphosate‐resistant populations in two pot experiments. The treatments were knockdown and pre‐emergence herbicides that were applied as a single application (alone or in a mixture) or as part of a sequential application to weeds at different growth stages. Glyphosate at 720 g ai ha^?1 provided good control of small glyphosate‐susceptible plants (pre‐ to early tillering), but was not always effective on larger susceptible plants. Paraquat was effective and the most reliable when applied at 500 g ai ha^?1 on small plants, irrespective of the glyphosate resistance status. The sequential application of glyphosate followed by paraquat provided 96–100% control across all experiments, irrespective of the growth stage, and the addition of metolachlor and metolachlor + atrazine to glyphosate or paraquat significantly reduced subsequent emergence. Herbicide treatments have been identified that provide excellent control of small E. colona plants, irrespective of their glyphosate resistance status. These tactics of knockdown herbicides, sequential applications and pre‐emergence herbicides should be incorporated into an integrated weed management strategy in order to greatly improve E. colona control, reduce seed production by the sprayed survivors and to minimize the risk of the further development of glyphosate resistance.     

Effect of herbicide antidotes on glutathione content and glutathione S-transferase activity of sorghum shoots

The effects of the herbicide antidotes CGA-92194 (α-[(1,3-dioxolan-2-yl-methoxy)-imino]benzeneacetonitrile), flurazole [phenylmethyl 2-chloro-4-(trifluoromethyl)-5-thiazolecarboxylate], dichlormid (2,2-dichloro-N,N-di-2-propenylacetamide), and naphthalic anhydride (1H,3H-naphtho(1,8-cd)-pyran-1,3-dione) on nonprotein thiol content, glutathione content, and glutathione S-transferase (GST) activity in etiolated sorghum (Sorghum bicolor L.) Moench) shoots were examined. CGA-92194 and naphthalic anhydride had no effect on nonprotein thiol or reduced glutathione (GSH) content of sorghum shoots. In contrast, dichlormid and flurazole increased nonprotein thiol content of sorghum shoots by 24 and 48%, respectively. These increases were largely attributable to an increase in GSH. The antidotes increased GST activity less than twofold when using CDNB (1-chloro-2,4-dinitrobenzene) as a substrate. In contrast, when using metolachlor [2-chloro-N-(2-ethyl-6-methylphenyl)-N-(2-methoxy-1-methylethyl)acetamide] as a substrate, the increase in GST activity in response to antidote treatment was much greater: flurazole (30-fold), CGA-92194 (20-fold), naphthalic anhydride (17-fold), dichlormid (5-fold). The degree of protection from metolachlor injury conferred by a particular antidote was strongly correlated (R² = 0.95) with its ability to enhance GST activity, as evaluated with metolachlor as substrate. A comparison of GST activity in untreated and CGA-92194-treated seedlings, over a range of metolachlor concentrations (0.5–500 μM), indicated that the relative enhancement of enzyme activity by CGA-92194 was greater at lower metolachlor concentrations. The rate of nonenzymatic conjugation of metolachlor and GSH in vitro was much less (on a gram fresh weight equivalent basis) than the enzymatic rate. These results are consistent with the hypothesis that the above antidotes protect sorghum by enhancing GST activity which results in accelerated detoxification of metolachlor via GSH conjugation.     

The effect of temperature on glutathione S-transferase activity and glutathione content in Alopecurus myosuroides (black grass) biotypes susceptible and resistant to herbicides

Herbicide‐resistant populations of Alopecurus myosuroides (black grass) have become widespread throughout the UK since the early 1980s. Previous observations in this laboratory have demonstrated that natural climatic fluctuations caused increases in endogenous glutathione S‐transferase (GST) enzyme activity in A. myosuroides plants as they mature, which is thought to be linked to herbicide resistance in this species. The present study has investigated the effects of plant growth at 10°C and 25°C, and reports GST specific activity and glutathione (GSH) pool size in resistant and susceptible A. myosuroides biotypes. Findings demonstrate differences in GST activity between resistant and susceptible populations, which are transient at lower growth temperatures. The GSH pool size was elevated at lower growth temperature in both biotypes. We speculate that these endogenous responses are part of a natural mechanism of acclimation to environmental change in this species and suggest that resistant plants are more able to adapt to environmental stress, as indicated in this instance by temperature change. These observations imply that the control of resistant A. myosuroides by graminicides may be more effective when applied at lower temperatures and at earlier growth stages.     

Sensitivity of Echinochloa crus‐galli populations to maize herbicides: a comparison between cropping systems

Echinochloa crus‐galli is an important maize weed with significant variation in herbicide sensitivity. This differential response may reflect differences in selection pressure caused by years of cropping system‐related herbicide usage. The herbicide sensitivity of E. crus‐galli populations from three divergent cropping systems was evaluated in dose–response pot experiments. Populations were collected from sandy fields with (i) a long‐term organic cropping system, (ii) a conventional cropping system with maize in the crop rotation or (iii) a conventional cropping system with long‐term monocropping of maize. Each cropping system was represented by six E. crus‐galli populations. The effectiveness of three foliar‐applied maize herbicides (nicosulfuron, cycloxydim and topramezone) and two soil‐applied maize herbicides (S‐metolachlor and dimethenamid‐P) was tested at three doses and two runs. Foliar‐applied herbicides were applied at the three true leaves stage. Soil‐applied herbicides were applied immediately after sowing. The foliage dry weight per pot was determined 4 weeks after treatment. Plant responses were expressed as biomass reduction. Herbicide sensitivity was consistently lowest for populations from maize monocropping systems. Compared with populations from organic cropping systems, populations from monocropping systems showed 6.9%, 9.8% and 29.3% lower sensitivity to cycloxydim, topramezone and nicosulfuron respectively. Populations from the conventional crop rotation system showed intermediate sensitivity levels, which did not significantly differ from sensitivity levels of populations from the other cropping systems. Sensitivity to dimethenamid‐P and S‐metolachlor was not affected by cropping system. Environmental conditions influenced herbicidal response . This study indicated that integrated weed management may be necessary to preserve herbicide efficacy over the long term.     

Non-enzymatic conjugation of fenoxaprop-ethyl with glutathione and cysteine in several grass species

Laboratory studies have shown that the amounts of glutathione (GSH) and cysteine are higher in grass species that are moderately tolerant, such as wheat (Triticum aestivum L., cv. Fredrick), and moderately susceptible, such as barley (Hor deum vitlgare L., cv. Legér) and triticale (cv. OAC Trillium), to fenoxaprop-ethyl (FE) than in species that are very susceptible to the her bicide, such as oat (Avena saliva L., cv. OAC Woodstock), wild oat (Avena fatua L.), yellow foxtail (Setaria glanca (L.) Bcauv.), large crab grass (Digitaria sanguinalis (L.) Scop.) and bar nyard grass (Echinochloa crus-galli (L.) P.B.). The safener, fenchlorazole-ethyl (FCE) was found to increase and decrease, respectively, the amounts of GSH and cysteine in the moderately tolerant and moderately susceptible species but had no effect on the susceptible species. It is sug gested that in the moderately tolerant and moderately susceptible species, especially following FCE treatment, more GSH is available to detoxify the herbicide. Glutathione-S-tranferase activity (GST) for FE was found to be very low in all of the species tested. In vitro experiments at physio-logical pH. demonstrated that FE may conjugate with GSH nonenzymatically. Therefore, it is suggested that nonenzymatic conjugation of fenoxaprop-ethyl with glutathione may be an important mechanism for tolerance of some grasses to this herbicide.     

冬小麦田大穗看麦娘种群动态及对小麦产量的影响

为明确麦田杂草大穗看麦娘Alopecurus myosuroides的田间种群发生动态规律,采用固定样方和随机样方取样方法于2013—2015年在山东省济南市研究了冬小麦田大穗看麦娘田间种群的萌发和生长动态,并分析了不同密度大穗看麦娘对小麦产量及其构成因素的影响。结果表明:在济南市小麦播后20~40 d为大穗看麦娘出苗高峰期,至11月下旬出苗量占全年出苗总量的97.1%。11月上旬,大穗看麦娘开始出现分蘖,至翌年4月上中旬结束,分蘖数平均每株为6.3个,比小麦多3.1个。3月下旬大穗看麦娘开始拔节,4月中旬平均株高超过小麦,5月下旬平均株高达到75.6 cm,高出小麦2.5 cm。5月上旬大穗看麦娘单茎平均鲜重达到最大为1.7 g,小麦为12.6 g。大穗看麦娘出苗、株高、分蘖和鲜重的变化与小麦变化趋势一致,与时间、温度密切相关。随着大穗看麦娘密度的增加,小麦产量急剧下降,当密度达到420株/m~2时,小麦产量损失率高达65.8%。大穗看麦娘对小麦产量构成因素中的穗密度影响最大,其次是穗粒数,对千粒重影响不显著,当密度为420株/m~2时,小麦穗密度减少55.2%,穗粒数减少26.4%。     

Molecular cloning, expression, and characterization of a phi-type glutathione S-transferase from Oryza sativa

The structural gene for glutathione S-transferase in Oryza sativa was successfully cloned from a cDNA library by the polymerase chain reaction method. The deduced amino acid sequence of this gene showed 44-66% similarity to the sequences of the class phi GSTs from Arabidopsis thaliana and Zea mays. This gene was expressed in Escherichia coli with the pET vector system and the gene product was purified to homogeneity by GSH-Sepharose affinity column chromatography. The expressed OsGSTF3-3 was a homo-dimer composed of 24 kDa subunit and its pI value was approximately 7.3. The OsGSTF3-3 was retained on GSH affinity column and its K_m value for GSH was 0.28 mM. The OsGSTF3-3 displayed high activity toward 1-chloro-2,4-dinitrobenzene, a general GST substrate and also had high activities towards acetanilide herbicides, alachlor, and metolachlor. The OsGSTF3-3 was highly sensitive to inhibition by benastatin A and S-hexyl-GSH. From these results, the expressed OsGSTF3-3 is a phi class GST and seems to play an important role in the conjugation of the chloroacetanilide herbicides.     

Antioxidant role of propolis extract against oxidative damage of testicular tissue induced by insecticide chlorpyrifos in rats

Pesticides induce oxidative stress leading to generate free radicals and alternate the antioxidant or oxygen free radical scavenging enzyme system. This study was conducted to investigate the oral toxicity of chlorpyrifos toward male rat and the oxidative stress of the sub-lethal dose (9 mg/kg; 1/25 LD₅₀) on the lipid peroxidation level (LPO), reduced glutathione content (GSH) and antioxidant enzymes; catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione-S-transferase (GST) activities of testicular tissue. Also, the protective effects of propolis extract (50 mg/kg b.w.) alone or in combination with chlorpyrifos were investigated. The oral administration of chlorpyrifos significantly caused elevation in LPO level by 1.79-fold as compared to control. The activities of antioxidant enzymes including CAT, SOD, GPx and GST were decreased significantly (23.66%, 27.75%, 29.13% and 11.52%) as well as the level of GSH decreased by 21.97% in testicular tissue as compared to control animals. Co-administration of propolis extract with chlorpyrifos or alone in male rats decreased LPO level, normalized CAT, SOD GPx and GST activities, while GSH content was increased in testicular tissue. We conclude that propolis extract significantly reduces chlorpyrifos-induced oxidative stress in rat testis and the protective effect of the pre-treatment with propolis extract as attenuating agent could be due to its antioxidant properties.