The introduction of glyphosate-resistant (GR) crops revolutionized weed management; however, the improper use of this technology has selected for a wide range of weeds resistant to glyphosate, referred to as superweeds. We characterized the high glyphosate resistance level of an Amaranthus hybridus population (GRH)—a superweed collected in a GR-soybean field from Cordoba, Argentina—as well as the resistance mechanisms that govern it in comparison to a susceptible population (GSH). The GRH population was 100.6 times more resistant than the GSH population. Reduced absorption and metabolism of glyphosate, as well as gene duplication of 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) or its overexpression did not contribute to this resistance. However, GSH plants translocated at least 10% more ¹⁴C-glyphosate to the rest of the plant and roots than GRH plants at 9 h after treatment. In addition, a novel triple amino acid substitution from TAP (wild type, GSH) to IVS (triple mutant, GRH) was identified in the EPSPS gene of the GRH. The nucleotide substitutions consisted of ATA¹⁰², GTC¹⁰³ and TCA¹⁰⁶ instead of ACA¹⁰², GCG¹⁰³, and CCA¹⁰⁶, respectively. The hydrogen bond distances between Gly-101 and Arg-105 positions increased from 2.89 Å (wild type) to 2.93 Å (triple-mutant) according to the EPSPS structural modeling. These results support that the high level of glyphosate resistance of the GRH A. hybridus population was mainly governed by the triple mutation TAP-IVS found of the EPSPS target site, but the impaired translocation of herbicide also contributed in this resistance.

Multiple resistance mechanisms to ALS inhibitors and auxin mimics in two Papaver rhoeas populations were investigated in wheat fields from Portugal. Dose-response trials, also with malathion (a cytochrome P450 inhibitor), cross-resistance patterns for ALS inhibitors and auxin mimics, alternative herbicides tests, 2,4-D and tribenuron-methyl absorption, translocation and metabolism experiments, together with ALS activity, gene sequencing and enzyme modelling and ligand docking were carried out. Results revealed two different resistant profiles: one population (R1) multiple resistant to tribenuron-methyl and 2,4-D, the second (R2) only resistant to 2,4-D. In R1, several target-site mutations in Pro197 and enhanced metabolism (cytochrome P450-mediated) were responsible of tribenuron-methyl resistance. For 2,4-D, reduced transport was observed in both populations, while cytochrome P450-mediated metabolism was also present in R1 population. Moreover, this is the first P. rhoeas population with enhanced tribenuron-methyl metabolism. This study reports the first case for P. rhoeas of the amino acid substitution Pro197Phe due to a double nucleotide change. This double mutation could cause reduced enzyme sensitivity to most ALS inhibitors according to protein modelling and ligand docking. In addition, this study reports a P. rhoeas population resistant to 2,4-D, apparently, with reduced transport as the sole resistance mechanism.

The evolution of herbicide resistance in weeds has emerged as one of the most serious threats to sustainable food production systems, which necessitates the evaluation of herbicides to determine their efficacy. The first herbicide resistance case in the Iberian Peninsula was reported about 50 years ago, wherein Panicum dichotomiflorum was found to be resistant (R) to atrazine in Spanish maize fields. Since then, herbicide resistance has evolved in 33 weed species, representing a total of 77 single-herbicide-resistance cases in this geographic area: 66 in Spain and 11 in Portugal. Changes in agricultural practices, namely the adoption of non-tillage systems and the increased use of herbicides, led to the selection of weed biotypes resistant to a wide range of herbicides. Nowadays the most important crops in Spain and Portugal (maize, winter cereals, rice, citrus, fruits, and olive orchards) are affected, with biotypes resistant to several mechanisms of action (MoAs), namely: ALS inhibitors (20 species), ACCase inhibitors (8 species), PS II inhibitors (18 species), and synthetic auxin herbicides (3 species). More recently, the fast increase in cases of resistance to the EPSPS-inhibiting herbicide glyphosate has been remarkable, with 11 species already having evolved resistance in the last 10 years in the Iberian Peninsula. The diversity of resistance mechanisms, both target-site and non-target-site, are responsible for the resistance to different MoAs, involving point mutations in the target site and enhanced rates of herbicide detoxification, respectively. More serious are the 13 cases reported with multiple-herbicide resistance, with three cases of resistance to three–four MoAs, and one case of resistance to five MoAs. Future research perspectives should further study the relationship between management strategies and the occurrence of TSR and NTSR resistance, to improve their design, develop monitoring and diagnostic tools for herbicide resistance, and deepen the study of NTSR resistance.

The levels of resistance to glyphosate of 13 barnyard grass (Echinochloa crus-galli) populations harvested across different agriculture areas in the Southern Iberian Peninsula were determined in greenhouse and laboratory experiments. Shikimate accumulation fast screening separated the populations regarding resistance to glyphosate: susceptible (S) E2, E3, E4, and E6 and resistant (R) E1, E5, E7, E8, E9, E10, E11, E12, and E13. However, resistance factor (GR₅₀ E1–E13/GR₅₀ E6) values separated these populations into three groups: (S) E2, E3, E4, and E6, (R) E1, E5, E7, E8, and E9, and very resistant (VR) E10, E11, E12, and E13. ¹⁴C-glyphosate assays performed on two S populations (E2 and E6) showed greater absorption and translocation than those found for R (E7 and E9) and VR (E10 and E12) populations. No previous population metabolized glyphosate to amino methyl phosphonic acid (AMPA) and glyoxylate, except for the E10 population that metabolized 51% to non-toxic products. The VR populations showed two times more 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) activity without herbicide than the rest, while the inhibition of the EPSPS activity by 50% (I₅₀) required much higher glyphosate in R and VR populations than in S populations. These results indicated that different target-site and non-target-site resistance mechanisms were implicated in the resistance to glyphosate in E. crus-galli. Our results conclude that resistance is independent of climate, type of crop, and geographic region and that the level of glyphosate resistance was mainly due to the selection pressure made by the herbicide on the different populations of E. crus-galli studied.

The conclusion of the dam project located in Alqueva, in Southern Portugal, has resulted in a significant increase of new irrigated areas, since 2006. This has meant that, in recent years, there have been progressive flora changes in farming systems traditionally implemented in the Alentejo region. The present work has analyzed the weed flora in an early stage of these changes, and the impact of environmental factors on the distribution of natural vegetation under Mediterranean climate conditions in the influence area of Alqueva. In 2007, 105 floristic surveys were carried out in autumn-winter crop plots or other soil use, and 264 species were identified. Families with higher expression were: Asteraceae, Poaceae, and Fabaceae. Only three species have been identified in more than half of farms, Avena sterilis, Phalaris minor and Lolium rigidum and they were part of the 15 species that revealed high and very high infestation degrees. Soil texture and extractable phosphorus have been determined as active ecological factors, according to the method of ecological profiles and Mutual Information. Therefore, these factores were those with the greatest influence on the species distribution. L. rigidum distribution showed to be associated with medium soil texture and A. sterilis distribution also showed to be associated with medium and fine soil texture soils, without showing ecological preference by extractable phosphorus. The distribution of P. minor was not related to the soil texture but showed preference for soils with medium phosphorus content.