Study On The Resistance Level And Mechanism Of Echinochloa Colona To Diuron In Sugarcane Field Of Guangxi

Echinochloa Colona is the main weed in sugarcane field of Guangxi.It has strong ability of tillering,large amount of seeds,strong adaptability under adverse environment,etc.It is the key and difficult point of weed control in sugarcane field.Diuron,as a substitute for urea herbicide,is a common weed control variety in sugarcane field of Guangxi.Through field investigation,it was found that the control effect of diuron on E.Colona was obviously decreased,and it was suspected that the population in the field had resistance to diuron.In order to provides scientific basis for the control of resistance of E.Colona and the scientific and reasonable use of diuron.In this study,we determined and evaluated the resistance level of E.Colona to diuron,determined the spectrum of resistant population,and systematically studied the resistance mechanism of E.Colona to diuron.The main results as follows:（1）34 field samples of E.Colona were collected from five sugarcane producing areas in Nanning,Laibin,Liuzhou,Qinzhou and Chongzuo of Guangxi,and the sensitivity of these populations to diuron was determined by single dose screening method.16 suspected resistant populations were found,accounting for 47.06%.Then,we tested the resistance level of 16 suspected resistant populations.The results showed that 4 populations were in middle resistance level,accounting for 11.76%,the resistance times between 6.27-8.27,12 populations were in low resistance level,the resistance times between2.44-5.71.Among them,population LBXB6 showed the highest resistance to diuron with the GR₅₀ of 1206.30 g a.i./hm²（RI=8.74）.The GR₅₀values of other populations varied from 336.90 g a.i./hm²-1074.30 g a.i./hm²（RI=2.44-7.78）.The GR₅₀value of population CZJZ2 was only 138.00 g a.i./hm²,which was used as sensitive population.（2）To determine the target resistance mechanism of E.Colona to diuron,the target gene psb A of diuron was amplified and sequenced,and the sequence of resistant population was compared with the relatively sensitive population.The results showed that the sequence length was 839 bp.The psb A gene sequences generated from this study were based on BLASTn searches of the Gen Bank database.We found that the similarity was 99.28%,99.52%and99.52%with the sequences of E.crus-galli、E.frumentacea and E.colona,respectively.These results indicated that this sequence is psb A gene sequence of E.Colona,and the primer can amplify psb A gene well.After comparing the resistant population with the sensitive population,it was found that there was no mutation in the reported 8 amino acid sites of ser-264-gly etc,which were associated with diuron resistance.This indicated that the resistance of E.Colona to diuron might not be caused by target mutation.（3）To find out whether the resistance of E.Colona is caused by the difference of absorption to diuron,high performance liquid chromatography（HPLC）was used to determine the absorption rate of diuron in the leaves of resistant and relatively sensitive populations of E.Colona.48 hours after the treatment,the average absorption rates of the two resistant populations and the relatively sensitive population were 68.06%,61.28%and 63.97%,respectively.There was no significant difference between the resistant population and the relatively sensitive population in the absorption of diuron.This indicated that the resistance of E.Colona to diuron might not be caused by the difference of absorption.（4）The metabolism of diuron in resistant and relatively sensitive populations of E.Colona was determined.48 hours after the treatment,the average metabolic rates of the two resistant populations and the relatively sensitive populations were 63.50%,56.16%and 22.02%,respectively.The average metabolic rates to diuron of the resistant population were significantly higher than that of the relatively sensitive population.This indicated that the main reason of resistance of E.Colona to diuron was the increase of metabolism of it.In order to further explore the role of metabolic enzymes in resistance of E.Colona,the synergistic effects of malathion,4-chloro-7-nitrobenzofurazan（NBD-Cl）and piperonyl butyl ether（PBO）on diuron were determined.Malathion,an inhibitor of MFO,had a significant synergistic effect on diuron in the treatment of resistant populations.The synergistic effect of 5 resistant populations was 1.78-1.92 times.Among them,NNXXT1 population had the highest efficiency multiple（1.92 times）;4-chloro-7-nitrobenzofuran（NBD-Cl）,a glutathione-S-transferase（GST）inhibitor,had a significant synergistic effect on diuron in the treatment of resistant population,The synergistic effect of 5resistant populations was 1.85-2.25 times.Among them,LBXB6 population had the highest efficiency multiple（2.25 times）.However,malathion and NBD-Cl had no significant synergistic effect on diuron in the relatively sensitive population.These results showed that MFO and GST were involved in metabolism resistance of diuron.However,PBO had no significant synergistic effect on diuron in resistant and relatively sensitive populations.The GSTs activity of resistant and relatively sensitive populations of E.Colona was measured.It was found that the GST activity of resistant population was significantly higher than that of relatively sensitive population.Day 3 after application,the GSTs activity of CZJZ2 was 102 mol/min/g,and the GSTs activity of 5 resistant populations was 2.06-2.60 times of that of the relative sensitive population.Among them,the GSTs activity of LBXB6 population was the highest,265.67 mol/min/g,2.60 times of that of the relative sensitive population.It can be seen that the resistance of resistant population to diuron is related to the increase of GSTs activity.（5）Through indoor bioassay,the phenomenon of cross resistance and multi resistance of E.Colona resistant population was proved.The results showed that resistant populations LBWX10,LBXB3,LBXB4,LBXB5 and LBXB6 were at risk of cross-resistance to atrazine,and resistant populations LBXB4 was at risk of multi-resistance to glyphosate.