The role of fungal endophyte infection in the evolution of Lolium multiflorum resistance to diclofop-methyl

Diclofop-methyl resistance was evaluated in populations of Italian ryegrass ( Lolium multiflorum Lam.) infected and uninfected with fungal endophyte ( Neotyphodium ). Survival was tested in susceptible populations using herbicide screening. The results served as inputs to a model for investigating the role of endophyte infection in the evolution of L. multiflorum resistance to herbicide. The tolerance of infected plants varied depending on the origin of the population and the herbicide dose. Only in some populations and at some diclofop-methyl doses did plants infected with these endophytes have higher survivorship than endophyte-free plants. The model demonstrated that endophytes might indeed play an important role in the evolution of herbicide-resistant weeds, delaying the appearance of herbicide resistance.     

An aspartate to glycine change in the carboxyl transferase domain of acetyl CoA carboxylase and non‐target‐site mechanism(s) confer resistance to ACCase inhibitor herbicides in a Lolium multiflorum population

BACKGROUND: The increasing use of ACCase‐inhibiting herbicides has resulted in evolved resistance in key grass weeds infesting cereal cropping systems worldwide. Here, a thorough and systematic approach is proposed to elucidate the basis of resistance to three ACCase herbicides in a Lolium multiflorum Lam. (Italian rye grass) population from the United Kingdom (UK24). RESULTS: Resistance to sethoxydim and pinoxaden was always associated with a dominant D2078G (Alopecurus myosuroides Huds. equivalent) target‐site mutation in UK24. Conversely, whole‐plant herbicide assays on predetermined ACCase genotypes showed very high levels of resistance to diclofop‐methyl for all three wild DD2078 and mutant DG2078 and GG2078 ACCase genotypes from the mixed resistant population UK24. This indicates the presence of other diclofop‐methyl‐specific resistance mechanism(s) yet to be determined in this population. The D2078G mutation could be detected using an unambiguous DNA‐based dCAPS procedure that proved very transferable to A. myosuroides, Avena fatua L., Setaria viridis (L.) Beauv. and Phalaris minor Retz. CONCLUSION: This study provides further understanding of the molecular basis of resistance to ACCase inhibitor herbicides in a Lolium population and a widely applicable PCR‐based method for monitoring the D2078G target‐site resistance mutation in five major grass weed species. Copyright © 2010 Society of Chemical Industry     

Differences in the molecular basis of resistance to the cyclohexanedione herbicide sethoxydim in Lolium multiflorum

Resistance to the cyclohexanedione (CHD) herbicide sethoxydim was investigated in two UK Lolium multiflorum populations, Yorks A2 and PYL. Resistance screening experiments demonstrated a qualitative difference in the responses of the two populations to sethoxydim, suggesting that the molecular basis of resistance between them was different. After treatment, Yorks A2 plants were either alive (78% of sample tested) or dead (22% tested) but plants of the PYL population showed two further intermediate categories of response. The level of acetyl-CoA carboxylase (ACCase) insensitivity directly correlated with the degree of resistance at the whole plant level, indicating that the molecular basis of resistance is associated with differences in ACCase sensitivity in each population. Direct sequencing of the carboxyl transferase domain of the ACCase gene showed that an Ile-418-Leu substitution in the L. multiflorum chloroplastic ACCase (GenBank accession number AY710293 ) confers resistance to sethoxydim in Yorks A2. This corresponds to amino acid residue 1781 in the Alopecurus myosuroides full ACCase sequence. This is the first report of this mutation in this L. multiflorum, which has also been reported in four other grass-weeds, including L. rigidum. However, no amino acid substitutions were found to be specifically associated with the resistant phenotypes in the PYL population and the molecular basis of resistance in this population remains to be resolved.     

Non‐target‐site mechanism of glyphosate resistance in Italian ryegrass (Lolium multiflorum)

In Shizuoka Prefecture, Japan, glyphosate‐resistant Lolium multiflorum is a serious problem on the levees of rice paddies and in wheat fields. The mechanism of resistance of this biotype was analyzed. Based on LD₅₀, the resistant population was 2.8–5.0 times more resistant to glyphosate than the susceptible population. The 5‐enolpyruvyl‐shikimate‐3‐phosphate synthase (EPSPS) gene sequence of the resistant biotype did not show a non‐synonymous substitution at Pro106, and amplification of the gene was not observed in the resistant biotype. The metabolism and translocation of glyphosate were examined 4 days after application through the direct detection of glyphosate and its metabolite aminomethylphosphonic acid (AMPA) using liquid chromatograph‐tandem mass spectrometer (LC‐MS/MS). AMPA was not detected in either biotype in glyphosate‐treated leaves or the other plant parts. The respective absorption rates of the susceptible and resistant biotypes were 37.90 ± 3.63% and 41.09 ± 3.36%, respectively, which were not significantly different. The resistant biotype retained more glyphosate in a glyphosate‐treated leaf (91.36 ± 1.56% of absorbed glyphosate) and less in the untreated parts of shoots (5.90 ± 1.17%) and roots (2.76 ± 0.44%) compared with the susceptible biotype, 79.58 ± 3.73%, 15.77 ± 3.06% and 4.65 ± 0.89%, respectively. The results indicate that the resistance mechanism is neither the acquisition of a metabolic system nor limiting the absorption of glyphosate but limited translocation of the herbicide in the resistant biotype of L. multiflorum in Shizuoka Prefecture.     

Molecular identification of weedy glyphosate-resistant Lolium (Poaceae) in California

The origins of invasive weed populations can involve multiple species, introductions and genotypes. In California, USA, self-incompatible (SI) Lolium species are highly successful agricultural weeds. During the last decade, resistance to glyphosate has evolved in California populations of Lolium . However, the species identity of glyphosate-resistant populations is unclear. Occurrence of three SI species, L. multiflorum , L. rigidum and L. perenne , and the lack of reproductive barriers between them, suggests that any or all species may be contributing to resistant populations. To elucidate the identity and evolutionary history of resistant Lolium , we assessed genetic variation of known species and unknown California individuals using five simple sequence repeat loci, a nuclear DNA sequence and a chloroplast DNA sequence. Employing two analytical approaches, we identified three major genetic groups within the accessions of SI species that were similar, but not identical, to the taxonomic species identities. California glyphosate-resistant and susceptible plants were most closely related to the L. multiflorum group. However, a few glyphosate-resistant plants and a third of the glyphosate-susceptible plants also identified with L. rigidum and L. perenne . While L. rigidum and L. perenne contribute to invasive populations within California, the majority of glyphosate-resistant individuals tested were L. multiflorum .     

砜吡草唑与氟噻草胺协同作用防治多花黑麦草及其对小麦的安全性

多花黑麦草是近来危害严重,抗药性发生普遍的一种恶性杂草,对我国小麦安全生产造成了严重威胁。为筛选对多花黑麦草有效的防除药剂,本文采用盆栽法进行了除草剂室内配方配比筛选试验,并对筛选的配方进行了室内安全性评价。结果表明,麦田常用茎叶处理除草剂精噁唑禾草灵、炔草酯、甲基二磺隆、啶磺草胺、氟唑磺隆对多花黑麦草防除效果均不理想,多花黑麦草对其均产生了不同程度的抗性。而土壤处理除草剂中砜吡草唑和氟噻草胺对多花黑麦草防除效果较好。进一步通过室内配方配比筛选试验,得出砜吡草唑和氟噻草胺1∶1.2混合对多花黑麦草的ED₅₀为3.46 g/hm~2,共毒系数(CTC)为137.40,表明二者1∶1.2混配对多花黑麦草防除有增效作用。安全性试验表明,砜吡草唑和氟噻草胺1∶1.2混配下对小麦的ED₁₀为381.98 g/hm~2,对多花黑麦草和小麦的选择性指数为56.93,表明砜吡草唑和氟噻草胺混配选择性最高,能有效防除多花黑麦草且对小麦安全。     

Diclofop‐resistant Lolium rigidum from northern Greece with cross‐resistance to ACCase inhibitors and multiple resistance to chlorsulfuron

Repeated use of ACCase‐ and ALS‐inhibiting herbicides in northern Greece has resulted in the evolution of a population of Lolium rigidum resistant to diclofop and chlorsulfuron. The biotype from Athos was highly resistant to diclofop and also exhibited differential cross‐resistance to clodinafop, fluazifop, tralkoxydim and sethoxydim. Assay of ACCase activity confirmed that the resistant biotype was tenfold more resistant to diclofop than the susceptible biotype, suggesting that the resistance mechanism could involve an altered target site. The diclofop‐resistant biotype has also exhibited multiple resistance to chlorsulfuron and the mechanism for this is unknown. Seed‐bioassay was found to be a rapid, cheap and reliable method to identify populations of L rigidum resistant to ACCase inhibitors and chlorsulfuron. Moreover, root elongation in the seed bioassay was more sensitive to ACCase inhibitors and chlorsulfuron than shoot elongation. © 2000 Society of Chemical Industry     

Resistance to ACCase-inhibiting herbicides and isoproturon in UK populations of Lolium multiflorum: mechanisms of resistance and implications for control

Herbicide-resistant Lolium multiflorum (Italian rye-grass) was first reported in the UK in 1993 and had been confirmed on 25 farms by 1999. In this study, resistance to five herbicides belonging to the aryloxyphenoxypropionate, cyclohexanedione and phenyl-urea classes was determined in six populations of L multiflorum from the UK under glasshouse and simulated field conditions. Glasshouse conditions tended to exaggerate the degree of resistance, but experiments performed in both environments detected resistance in four populations of L multiflorum. Four populations (Essex A1, Lincs A1, Wilts B1, Yorks A2) were resistant to diclofop-methyl, fluazifop-P-butyl, tralkoxydim and partially resistant to isoproturon, but only the population from Yorkshire (Yorks A2) showed resistance to cycloxydim. Biochemical analyses of acetyl coenzyme A carboxylase (ACCase) activity, oxygen consumption by thylakoids, diclofop metabolism and glutathione S-transferase activity showed that, in three of the resistant populations, an enhanced rate of herbicide metabolism conferred resistance. This is the first report world-wide of an enhanced metabolism mechanism of diclofop resistance in L multiflorum. In the Yorks A2 population, an insensitive ACCase was detected (target-site resistance) which also conferred cross-resistance to all of the other ACCase inhibitors investigated.     

Diclofop-methyl resistance in populations of Lolium spp. from central Italy

Populations of Lolium spp. collected in central Italy were screened for resistance to acetyl-coenzyme A carboxylase (ACCase)-inhibiting herbicides and compared with known susceptible and resistant Lolium rigidum (Gaud.) populations from Australia. Populations Roma'94 and Tuscania'97 were up to 8- and 7.5-fold more resistant to diclofop-methyl, respectively, than susceptible populations in pot experiments. However, populations Tarquinia'97 and Vetralla'94 were not resistant. Diclofop-methyl resistance levels in the Italian populations were lower than in the Australian populations SLR31 and WLR96 (16.5 and > 64 times more resistant than S respectively). In an agar germination test, Tuscania'97 showed low levels of cross-resistance to fluazifop-p-butyl, whereas no cross-resistance was found in the Roma'94 population.     

氟噻草胺与氟唑磺隆混配协同作用及在小麦田杂草防治中的应用

本文通过室内生物测定研究了氟噻草胺与氟唑磺隆对小麦及多花黑麦草的活性,田间试验测定了二者混配对阔叶杂草的防效及对小麦分蘖数的影响。室内水培法测定结果表明,氟噻草胺、氟唑磺隆对多花黑麦草有效抑制中剂量(ED50)分别为0.37mg/L和105.91mg/L,氟噻草胺与氟唑磺隆混配比例为8∶2、12∶2、16∶2、20∶2时,共毒系数(CTC)分别为94.3、239.7、198.1、156.5,表明氟噻草胺与氟唑磺隆混用具有相加或增效作用,其中以12∶2、16∶2比例混配对多花黑麦草防治增效最为显著。盆栽试验结果表明,苗前以12∶2和16∶2比例土壤封闭喷施对小麦与多花黑麦草的选择性指数为1.51和1.43;苗后3d及苗后15d茎叶喷施对多花黑麦草与小麦选择性较差,选择性指数低于1。田间测定结果表明,氟噻草胺与氟唑磺隆混配对阔叶杂草荠菜、播娘蒿、婆婆纳防效优于氟噻草胺单独使用,混配条件下株防效为85.45%~100%,鲜重防效为89.57%~100%,对小麦分蘖数无显著影响。     

Simulating evolution of glyphosate resistance in Lolium rigidum I: population biology of a rare resistance trait

Despite frequent use for the past 25 years, resistance to glyphosate has evolved in few weed biotypes. The propensity for evolution of resistance is not the same for all herbicides, and glyphosate has a relatively low resistance risk. The reasons for these differences are not entirely understood. A previously published two‐herbicide resistance model has been modified to explore biological and management factors that account for observed rates of evolution of glyphosate resistance. Resistance to a post‐emergence herbicide was predicted to evolve more rapidly than it did to glyphosate, even when both were applied every year and had the same control efficacy. Glyphosate is applied earlier in the growing season when fewer weeds have emerged and hence exerts less selection pressure on populations. The evolution of glyphosate resistance was predicted to arise more rapidly when glyphosate applications were later in the growing season. In simulations that assumed resistance to the post‐emergence herbicide did not evolve, the evolution of glyphosate resistance was less rapid, because post‐emergence herbicides were effectively controlling rare glyphosate‐resistant individuals. On their own, these management‐related factors could not entirely account for rates of evolution of resistance to glyphosate observed in the field. In subsequent analyses, population genetic parameter values (initial allele frequency, dominance and fitness) were selected on the basis of empirical data from a glyphosate‐resistant Lolium rigidum population. Predicted rates of evolution of resistance were similar to those observed in the field. Together, the timing of glyphosate applications, the rarity of glyphosate‐resistant mutants, the incomplete dominance of glyphosate‐resistant alleles and pleiotropic fitness costs associated with glyphosate resistance, all contribute to its relatively slow evolution in the field.     

Glyphosate resistance in four different populations of Lolium rigidum is associated with reduced translocation of glyphosate to meristematic zones

Weed populations with resistance to glyphosate have evolved over the last 7 years, since the discovery of the first glyphosate‐resistant populations of Lolium rigidum in Australia. Four populations of L. rigidum from cropping, horticultural and viticultural areas in New South Wales and South Australia were tested for resistance to glyphosate by dose–response experiments. All populations required considerably more glyphosate to achieve 50% control compared with a known susceptible population, indicating they were resistant to glyphosate. Translocation of glyphosate within these resistant populations was examined by following the movement of radiolabelled glyphosate applied to a mature leaf. All resistant plants translocated significantly more herbicide to the tip of the treated leaf than did susceptible plants. Susceptible plants translocated twice as much herbicide to the stem meristematic portion of the plant compared with resistant plants. These different translocation patterns suggest an association between glyphosate resistance in L. rigidum and the ability of glyphosate to accumulate in the shoot meristem.