Evolution and Biochemical Mechanisms of Multiple resistance to Diclofop-methyl and Chlorsulfuron in Lolium rigidum from Northern Greece

Transcription

1 Evolution and Biochemical Mechanisms of Multiple resistance to Diclofop-methyl and Chlorsulfuron in Lolium rigidum from Northern Greece Eleni Kotoula-Syka Democritus University of Thrace, Greece

2 Introduction The widespread use of herbicides over the past decades for weed control, has exposed huge weed populations to strong selection pressures for herbicide resistant traits. The aryloxyphenoxypropionate (AOPP) and cycloexanedione (CHD) herbicides, are two important groups of post-emergence herbicides, used to control grass weeds in grass and dicot crops that share a common mode of action (ACCase inhibitors)

3 Introduction Chlorsulfuron the first acetolactate synthase (ALS) inhibiting herbicide, is a cereal-selective herbicide and has become widely used mainly for broad-leaf weeds and L. rigidum control. Resistance to ALS inhibitors in broad-leaf weeds has been mostly due to an altered ALS rather, than to differences in herbicide uptake, translocation or metabolism

4 Introduction A polymorphic and obligate out-crosser weed, such as L. rigidum, has the ability to rapidly evolve resistance to different herbicides, leading to cases of multiple resistance. L. rigidum is a major weed in winter wheat in northern Greece, and since the mid-1970s diclofop-methyl has been used to control this weed quite effectively. Later other AOPP and CHD herbicides were progressively introduced throughout the 1980s.

5 Introduction Since late 1980s, chlorsulfuron, has also become widely used, mainly for broad-leaved weeds and L. rigidum control. These herbicides have enabled most farms to practice minimum tillage and in many locations were the only herbicides used for L. rigidum control. Recently, failures of diclofop-methyl and chlorsulfuron in controlling L. rigidum were reported by farmers in northern Greece.

6 Objectives The objectives of the present study were to confirm the evolution of resistance to ACCase inhibitors and multiple resistance to chlorsulfuron and to elucidate the mechanisms of resistance in a L. rigidum biotype from Northern Greece.

7 Materials and Methods L. rigidum seeds were collected from winter wheat fields treated repeatedly for more than five consecutive years with diclofop-methyl and chlorsulfuron and from a field that had never been treated with herbicides. Pot experiments were conducted outdoors and commercial formulations of the herbicides were applied pre- or post-emergence at the 3-4 leaf stage, at several rates.

8 Materials and Methods Plant growth was evaluated by determining shoot fresh weight per pot (two weeks after post-emergence application of AOPP and CHD herbicides or 25 days after pre-emergence application of chlorsulfuron) The ACCase extraction and its activity was assayed using the procedure described by Tal el al. (1996). The concentration of herbicide causing 50% inhibition of ACCase activity (IC 50 ) was estimated for the concentration response curve. To elucidate the mechanism of resistance to chlorsulfuron, L. rigidum seedlings were treated postemergence with commercial formulation of chlorsulfuron with and without pretreatment of malathion.

9 Results and Discussion Resistance to diclofop-methyl and cross- resistance to other AOPP and CHD herbicides. Analysis of shoot fresh weight vs dose response curves indicated that population C was highly resistant to diclofop and differed 60-fold from populations A, B and D. For further tests, population C was chosen as the resistant biotype (R) and population D as the susceptible biotype (S ).

10 Results and Discussion

11 Results and Discussion In addition to resistance to diclofop, the R biotype showed differential cross-resistance to other ACCase inhibiting herbicides. The highest R/S value was recorder for diclofop and considerably smaller value for clodinafor, 97.5 and 33.9 respectively. Much lower R/S values of 6.9, 6.0 and 10.2 respectively were detected for fluazifop, tralkoxydim and sethoxydim.

12 Results and Discussion Table 1. Response of resistant (R) and susceptible (S) Lolium rigidum biotypes to various AOPP and CHD herbicides. ED 50 Herbicide Group Biotype (g AI ha -1 ) R/S ED 50 a Diclofop AOPP R S Clodinafop AOPP R S Fluazifop AOPP R S Tralkoxydim CHD R S Sethoxydim CHD R S a Shoot fresh weight ED 50 in the resistant biotype/shoot fresh weight ED 50 in the susceptible biotype.

13 Results and Discussion Differential patterns of cross-resistance to various AOPP and CHD herbicides in a given species have been shown by other workers. The phenomenon is very common in many resistant grass weeds and has serious implications on the planning of herbicide/crop rotations.

14 Results and Dicsussion Differences in ACCase activity. ACCase activity in the R biotype was less affected by increasing concentrations of diclofop-acid than in the S biotype, resulting in IC 50 values of 100 and 10 μm, respectively.

15 Results and Dicsussion

16 Results and Discussion The results with extracted ACCase enzyme are in agreement with the whole-plant results (Fig. 1 and Table 1) and suggest that it is a target-site-based resistance. Other enzyme inhibition studies, suggested several distinct mutations in the ACCase gene, conferring different levels of resistance to various ACCase inhibitors.

17 Results and Discussion Multiple resistance to chlorsulfuron. Dose response curves of L. rigidum populations showed that the population C was also highly resistant to chlorsulfuron and differed significantly from populations A and D, while population B showed an intermediate level of resistance to chlorsulfuron Multiple resistance for diclofop and chlorsulfuron in L. rigidum has been reported by others and suggestions for the mechanism of this were also made.

18 Results and Discussion

19 Results and Discussion Mechanism of resistance to chlorsulfuron. Pretreatment of R L. rigidum plants with malathion, a known inhibitor of P 450 monooxygenases, increased their sensitivity to chlorsulfuron, whereas the response of the S plants did not change. This data indicate that the chlorsulfuron resistance is based on enhanced detoxification of the herbicide.

20 Results and discussion Table 2. Response of resistant (R) and susceptible (S) L. rigidum biotypes to chlorsulfuron applied following pretreatment with different rates of malathion Malathion ( g ai ha -1 ) Chlorsulfuron ED 50 R ( g ai ha -1 ) S

21 Conclusions The diclofop-resistance in the R biotype of L. rigidum was due to a less sensitive ACCase (target site resistance) The diclofop-resistant R biotype was also multiple resistant to chlorsulfuron due to enhanced detoxification of the herbicide (metabolic resistance).

22 Thank you for your attention

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