Glyphosate is the world's most widely used herbicide. A potential substitute for glyphosate in some use patterns is the herbicide paraquat. Following many years of successful use, neither glyphosate nor paraquat could control a biotype of the widespread annual ryegrass (Lolium rigidum), and here the world's first case of multiple resistance to glyphosate and paraquat is confirmed. Dose-response experiments established that the glyphosate rate causing 50% mortality (LD(50)) for the resistant (R) biotype is 14 times greater than for the susceptible (S) biotype. Similarly, the paraquat LD(50 )for the R biotype is 32 times greater than for the S biotype. Thus, based on the LD(50 )R/S ratio, this R biotype of L. rigidum is 14-fold resistant to glyphosate and 32-fold resistant to paraquat. This R biotype also has evolved resistance to the acetyl-coenzyme A carboxylase (ACCase) inhibiting herbicides. The mechanism of paraquat resistance in this biotype was determined as restricted paraquat translocation. Resistance to ACCase-inhibiting herbicides was determined as due to an insensitive ACCase. Two mechanisms endowing glyphosate resistance were established: firstly, a point mutation in the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene, resulting in an amino acid substitution of proline to alanine at position 106; secondly, reduced glyphosate translocation was found in this R biotype, indicating a co-occurrence of two distinct glyphosate resistance mechanisms within the R population. In total, this R biotype displays at least four co-existing resistance mechanisms, endowing multiple resistance to glyphosate, paraquat and ACCase herbicides. This alarming case in the history of herbicide resistance evolution represents a serious challenge for the sustainable use of the precious agrochemical resources such as glyphosate and paraquat.

Glyphosate resistance evolution in weeds is a growing problem in world agriculture. Here, we have investigated the mechanism(s) of glyphosate resistance in a Lolium rigidum population (DAG1) from South Africa. Nucleotide sequencing revealed the existence of at least three EPSPS homologues in the L. rigidum genome and identified a novel proline 106 to leucine substitution (P106L) in 52% DAG1 individuals. This mutation conferred a 1.7-fold resistance increase to glyphosate at the whole plant level. Additionally, a 3.1-fold resistance increase, not linked to metabolism or translocation, was estimated between wild-type P106-DAG1 and P106-STDS sensitive plants. Point accepted mutation analysis suggested that other amino acid substitutions at EPSPS position 106 are likely to be found in nature besides the P106/S/A/T/L point mutations reported to date. This study highlights the importance of minor mechanisms acting additively to confer significant levels of resistance to commercial field rates of glyphosate in weed populations subjected to high selection pressure.

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The first case of field-evolved paraquat resistance in a population of Lolium rigidum Gaud. (from the Western Cape, South Africa) was confirmed and the mechanism of resistance investigated. The LD50 for the resistant population (R) was 404 g ha–1, some 14 times greater than for the herbicide-susceptible (S) population (30 g ha–1). In addition, the R population was found to be more resistant to paraquat when kept at low temperature (15°C) than when kept at 30°C after paraquat treatment. The R population is normally affected by herbicides with other modes of action. No differences were found in the interaction of paraquat with Photosystem I in thylakoids isolated from the R and the S populations. Constitutive levels of the antioxidative enzymes superoxide dismutase (SOD) and ascorbate peroxidase (APX) did not differ significantly between the two populations and these enzymes responded similarly to paraquat treatment. When [14C] paraquat was applied as droplets to intact plant leaves, similar leaf uptake of [14C] paraquat occurred in the R and S populations. However, quantification data and phosphor imaging revealed restricted translocation of [14C] paraquat to untreated leaves in the R compared to S population. The results of this study with this resistant L. rigidum population from South Africa resemble those found in R biotypes of Hordeum spp. from Australia. The resistance is suggested to be primarily due to sequestration of paraquat, limiting its translocation within the plants. The exact site and mechanism of paraquat sequestration remains to be determined.

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