Resistance Of Keng Stiffgrass To Fenoxaprop-P-ethyl In Winter Wheat Fields

Keng stiffgrass(Pseudosclerochloa kengiana)is a member of the genus sclerochloa in the family Poaceae.It is one of troublesome grass weed species in the wheat fields rotated with rice and is widely distributed in southeast China.Fenoxaprop-P-ethyl,an ACCase-inhibiting herbicide,is commonly used to control grass weeds including keng stiffgrass in wheat fields.Since its introduction in 1998,fenoxaprop-P-ethyl have been used widely in China due to their high efficacy,ease of use and cost-effectiveness.However,Resistance has evolved in many grass weeds under continuously selective pressure of fenoxaprop-P-ethyl.In recent years,growers have noticed that keng stiffgrass was unsuccessfully controlled by fenoxaprop-P-ethyl at the recommended rate.In order to characterize the resistant level and the resistant mechanism to fenoxaprop-P-ethyl in keng stiffgrass from partial regions in China,and provide the theory basis for scientific control of keng stiffgrass,we collected 62 keng stiffgrass populations from Shandong,Jiangsu and Henan provinces,determined their sensitivity to fenoxaprop-P-ethyl and reveal the target-site-resistance mechanisms of resistant keng stiffgrass in this study.The main results were as following:1.All the 62 populations were treated with fenoxaprop-P-ethyl at the rate of 62.1g ai ha-1 and 124.2g ai ha-1 respectively by using whole-plant pot bioassay.The results showed that 25 populations were resistant to fenoxaprop-P-ethyl,and other 37 populations were sensitive.Whole-plant dose-response assay was conducted to determine the resistant level of 25 resistant populations to fenoxaprop-P-ethyl,and the results indicated that all the resistant were were highly resistant to fenoxaprop-P-ethyl.The RI values were arranging from 14.7-to 103.2-fold when compared with the susceptible population.2.Two copies of the gene encoding plastidic ACCase in keng stiffgrass were isolated,and both the two homologous genes(Acc1;1 and Acc1;2)are expressed.3.By comparing the partial ACCase sequences derived from the resistant populations and susceptible populations,the SD-10 population was found to contain a tryptophan-to-leucine mutation at codon position 1999;the SD-12,SD-13,SD-19 and SD-31 populations were found to contain a tryptophan-to-serine mutation at codon position 1999;the JS-16,JS-25,SD-2,SD-3,SD-9,SD-14,SD-15,SD-17,SD-18,SD-22,SD-24 and SD-25 populations were found to contain a tryptophan-to-cysteine mutation at codon position 2027;the JS-5,SD-4,SD-11 and SD-30 populations were found to contain a isoleucine-to-asparagine mutation at codon position 2041;the JS-13,SD-16 and SD-32 poulations were found to contain a asparticacid-to-glycine mutation at codon position 2078;the SD-23 poulation was found to contain a glycine-to-alanine mutation at codon position 2096.4.By comparing the two homoeologous ACCase genes Acc1;1 and Acc1;2 derived from the resistant populations,the Trp-1999-Leu mutation was detected in the Acc1;2 locus;the Trp-1999-Ser and Gly-2096-Ala mutations were detected in the Acc1;1 locus;the Trp-2027-Cys and Asp-2078-Gly mutations were detected in the Acc1;1 or Acc1;2 locus;the Ile-2041-Asn mutation was detected in the Acc1;2 locus,and it was only found in one individual plant of SD-4 population that the Ile-2041-Asn mutation was detected in the Acc1;1 locus.5.The cross-and mutiple-resistance experiments showed that SD-12 and SD-13 populations were highly resistant to quizalofop-P-ethyl,moderately resistant to clodinafop-propargyl and haloxyfop-R-methyl,lowly resistant to fluazifop-P-butyl and pinoxaden,but were sensitive to sethoxydim and clethodim;the JS-25,SD-4 and SD-11 populations exhibited high resistance to clodinafop-propargyl and fluazifop-P-butyl,moderate resistance to sethoxydim and pinoxaden,but susceptible to clethodim;The SD-32 population showed high resistance to sethoxydim,moderate resistance to clodinafop-propargyl,and low resistance to clethodim and pinoxaden;SD-23 population was highly resistant to haloxyfop-R-methyl,moderately resistant to clodinafop-propargyl and quizalofop-P-ethyl,lowly resistant to fluazifop-P-butyl,but were sensitive to sethoxydim,clethodim and pinoxaden;the SD-4,SD-10,SD-11,SD-12,SD-13 and SD-23 populations did not evolved resistance to mesosulfuron-methyl,flucarbazone-sodium,pyroxsulam and isoproturon.6.Five(d)CAPS markers were developed to identify the ACCase Trp1999,Ser1999,Cys2027,Ile2041 and Asn2041 alleles in this study.The results showed that all 350 individual plants of SD-12 population contained Trp-1999-Ser mutation and were heterozygous mutants;318 heterozygous Trp-1999-Ser mutation plants and 32 wild types were detected from the SD-13 population;all 180 individual plants of JS-25 population contained Trp-2027-Cys mutation and were heterozygous mutants;all 180 individual plants of SD-4 population contained Ile-2041-Asn mutation and were heterozygous mutants.7.When exposed to constant temperature,keng stiffgrass germinated over a temperature range of 5 to 15?;in the alternating temperature 10/5?,15/10? and 20/15?,germination values were all greater than 88%.Medium light has no significant effect on seed germination.More than 86% of seeds germinated at pH values ranged from 4 to 10.Seed germination was sensitive to osmotic potential and completely inhibited at osmotic potential of-0.8 Mpa,butVI was quite tolerant to salinity.Highest seedling emergence(>90%)occurred from depths of 0 to 1.0 cm,and when seeds were buried deeper than 7 cm,no seedlings emerged.8.The common herbicide used in wheat fenoxaprop-P-ethyl,clodinafop-propargyl,pinoxaden,mesosulfuron-methyl and pyroxsulam provided more than 80% control of keng stiffgrass when applied at the three-to nine-leaf stages.Isoproturon applied at three-to sixleaf stage provided more than 90% keng stiffgrass control;however,isoproturon applied at nine-leaf stage achieved only 41.8% control.Flucarbazone-sodium applied at any leaf stage provided lower than 75% keng stiffgrass control.This study determined the sensitivity of 62 keng stiffgrass populations collected from the most wheat planting regions in China to fenoxaprop-P-ethyl,and characterized the cross-and multiple-resistance patterns in the most resistant populations.Molecular analyses confirmed that the specific point mutations in the plastid ACCase were the important reason for resistance to fenoxaprop-P-ethyl in keng stiffgrass.We evaluated the effects of environmental factors affecting seed germination and seedling emergence,and evaluated the effect of common herbicides on control of keng stiffgrass.In addition,molecular markers were also developed to rapidly identify mutations at positions 1999,2027 and 2041.