Alterations of oxidative status induced by amino acid synthesis-inhibiting herbicides in sensitive and target-site resistantpopulations of Amaranthus palmeri
Authors:   , Vicente Eceiza Mikel Gonzalo 
Published: June 2023

Among the available methods to control weeds, herbicides stand as quick and effective tools, and conform the most extended method to control weeds. Two of the most prominent herbicide groups are 5-enolpyruvylshikimate-3-phosphate-synthase (EPSPS) and acetolactate synthase (ALS) inhibitors. The only known EPSPS inhibitor is glyphosate, the most used herbicide in the world, while ALS inhibitors include many active substances. EPSPS and ALS are important enzymes involved in the synthesis of aromatic amino acids and branched-chain amino acids, respectively. How these amino acid biosynthetic pathways are disrupted after the application of glyphosate or an ALS inhibitor is well studied, but the sequence of physiological events from herbicide application to plant death is not completely understood. Currently established physiological effects are quite similar between both herbicide groups (despite the different target enzymes). Some studies show that oxidative stress is among these physiological effects induced by glyphosate and ALS inhibitors, but both its origin (whether it is linked to EPSPS or ALS inhibition or is a separate side effect) and the role of oxidative damage and alterations of the antioxidant systems (such as glutathione) in the physiological downfall that ends with plant death are unknown. In this context, the present study mainly aimed to evaluate the role of the oxidative stress and the metabolism of glutathione in the mode of action of glyphosate or ALS inhibitors, trying to unravel if they are related to the mechanism of action of both groups of herbicides. Four populations of the weed Amaranthus palmeri S. Wats. were grown: glyphosate-sensitive (GFS), glyphosate-resistant (GFR), ALS-inhibitor sensitive (AIS), and ALS-inhibitor resistant (AIR). In both resistant populations, resistance was conferred by a target-site resistance (TSR) mechanism. GFS and GFR populations were treated with different glyphosate doses, while AIS and AIR populations were treated with different nicosulfuron (a sulfonylurea, ALS inhibitor) doses. Several physiological analyses were performed and compared in leaves of these populations: reactive oxygen species (ROS) accumulation, oxidative damage, antioxidant systems (with special emphasis on glutathione metabolism), and hormonal signalling. There were virtually no differences between untreated sensitive and resistant plants in the measured parameters evidencing that TSR is not accompanied by a different basal oxidative pattern in any of both cases. GFS and AIS plants treated with any dose of glyphosate or nicosulfuron died, while all resistant plants survived the herbicide treatments. Treated GFS and AIS plants showed a higher ROS production and accumulation than untreated plants, an effect that did not take place in resistant plants. This great ROS overproduction in treated sensitive plants led to a herbicide dose-proportional lipid peroxidation, an unmistakable sign of oxidative damage and to changes of antioxidant systems. Only treated plants of both sensitive populations showed increase of glutathione S-transferase (GST) activity and gene expression, indicating the involvement of glutathione as an antioxidant. The presence of oxidative stress only in sensitive plants treated with glyphosate and an ALS inhibitor and its absence in TSR plants indicate that oxidative stress is related to EPSPS or ALS inhibition, although the exact mechanisms remain elusive. Oxidative damage was evidenced as another common effect of both types of herbicides, but it was too mild to provoke plant death by itself, evidencing that oxidative stress is not the main cause of plant death. Apart from that, both herbicides affected hormonal profile, but induced different hormones. In this way, the changes detected in the hormonal profile show a different effect of herbicidal action. The mechanisms by which these hormonal changes are triggered remain unknown, but the induction of certain hormones may be related to oxidative stress.