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The dinitroaniline herbicides (particularly trifluralin) have been globally used in many crops for selective grass weed control. Consequently, trifluralin resistance has been documented in several important crop weed species and has recently reached a level of concern in Australian Lolium rigidum populations. Here, we report novel mutations in the L. rigidum a-tubulin gene which confer resistance to trifluralin and other dinitroaniline herbicides. Nucleotide mutations at the highly conserved codon Arg-243 resulted in amino acid substitutions of Met or Lys. Rice calli transformed with the mutant 243- Met or 243-Lys a-tubulin genes were 4- to 8-fold more resistant to trifluralin and other dinitroaniline herbicides (e.g., ethalfluralin and pendimethalin) compared to calli transformed with the wild type a-tubulin gene from L. rigidum. Comprehensive modeling of molecular docking predicts that Arg-243 is close to the trifluralin binding site on the a-tubulin surface and that replacement of Arg-243 by Met/Lys-243 results in a spatial shift of the trifluralin binding domain, reduction of trifluralin-tubulin contacts, and unfavorable interactions. The major effect of these substitutions is a significant rise of free interaction energy between a-tubulin and trifluralin, as well as between trifluralin and its whole molecular environment. These results demonstrate that the Arg-243 residue in a-tubulin is a determinant for trifluralin sensitivity, and the novel Arg-243-Met/Lys mutations may confer trifluralin resistance in L. rigidum. [ABSTRACT FROM AUTHOR]

A novel glyphosate resistance double point mutation (T102I/P106S, TIPS) in the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene has been recently identified for the first time only in the weed species Eleusine indica. Quantification of plant resistance cost associated with the TIPS and the often reported glyphosate resistance single P106S mutation was performed. A significant resistance cost (50% in seed number currency) associated with the homozygous TIPS but not the homozygous P106S EPSPS variant was identified in E. indica plants. The resistance cost associated with the TIPS mutation escalated to 85% in plants under resource competition with rice crops. The resistance cost was not detected in nonhomozygous TIPS plants denoting the recessive nature of the cost associated with the TIPS allele. An excess of 11-fold more shikimate and sixfold more quinate in the shikimate pathway was detected in TIPS plants in the absence of glyphosate treatment compared to wild type, whereas no changes in these compounds were observed in P106S plants when compared to wild type. TIPS plants show altered metabolite levels in several other metabolic pathways that may account for the expression of the observed resistance cost. [ABSTRACT FROM AUTHOR]

Twenty-two amino acid substitutions at seven conserved amino acid residues in the acetohydroxyacid synthase (AHAS) gene have been identified to date that confer target-site resistance to AHAS-inhibiting herbicides in biotypes of field-evolved resistant weed species. However, the effect of resistance mutations on AHAS functionality and plant growth has been investigated for only a very few mutations. This research investigates the effect of various AHAS resistance mutations in Lolium rigidum on AHAS functionality and plant growth. The enzyme kinetics of AHAS from five purified L. rigidum populations, each homozygous for the resistance mutations Pro-197-Ala, Pro-197-Arg, Pro-197-Gln, Pro-197-Ser or Trp-574-Leu, were characterized and the pleiotropic effect of three mutations on plant growth was assessed via relative growth rate analysis. All these resistance mutations endowed a herbicide-resistant AHAS and most resulted in higher extractable AHAS activity, with no-to-minor changes in AHAS kinetics. The Pro-197-Arg mutation slightly (but significantly) increased the Km for pyruvate and remarkably increased sensitivity to feedback inhibition by branched chain amino acids. Whereas the Pro-197-Ser and Trp-574-Leu mutations exhibited no significant effects on plant growth, the Pro-197-Arg mutation resulted in lower growth rates. It is clear that, at least in L. rigidum, these five AHAS resistance mutations have no major impact on AHAS functionality and hence probably no plant resistance costs. These results, in part, explain why so many Pro-197 AHAS resistance mutations in AHAS have evolved and why the Pro-197-Ser and the Trp-574-Leu AHAS resistance mutations are frequently found in many weed species. [ABSTRACT FROM PUBLISHER]

Abstract: In two different locations of the Western Australian “wheatbelt”, Lolium rigidum (rigid ryegrass) seeds were collected from organic fields (no herbicide use) and neighbouring conventional fields (persistent herbicide use), the latter infested with herbicide-resistant plants, to investigate the occurrence of gene flow among field populations as revealed by herbicide resistance gene transfer. Herbicides targeting acetyl-CoA carboxylase (ACCase) or acetolactate-synthase (ALS) were used to detect herbicide-resistant plants. Overall, the frequency of plants resistant to ACCase- or ALS-inhibiting herbicides was, respectively, 21% and 74% in the conventional fields and 2% and 37% in neighbouring organic fields. Mutant, herbicide-resistant ACCase and ALS alleles were detected in 16% and 38% of plants from conventional fields and in 0.53% and 3.7% of plants from organic fields. Identical mutant, herbicide-resistant ALS haplotypes were detected both in conventional and organic fields, supporting the occurrence of gene flow between L. rigidum populations in different fields. Gene flow can thus substantially increase the frequency of herbicide-resistant plants in unselected L. rigidum populations. Although gene flow cannot be prevented, it can be limited or managed. Hygiene tactics such as clean crop seed, weed seed removal at harvest and seed destruction post-harvest should be considered in order to minimize gene transfer among farms. [Copyright &y& Elsevier]