Amaranthus palmeri a New Invasive Weed in Spain with Herbicide Resistant Biotypes

Amaranthus palmeri is the most prominent invasive weed in agricultural land from North America, partly due to its propensity to evolve resistance to multiple herbicide sites of action. In the last two decades, reports of this species have increased throughout the American continent and occasionally in other continents. In 2007, A. palmeri populations were found in three localities in northeastern Spain, and they are still present today. To determine whether these three populations resulted from a common or independent introduction events¿and when and from where they could have occurred¿research was carried out aiming to characterize the resistance profile and mechanisms to 5-enolpyruvylshikimate-3-phosphate synthase-and acetolactate synthase (ALS)-inhibiting herbicides and to analyze the relationship between these three populations using inter simple sequence repeat DNA fingerprinting. Dose-response trials confirmed that the three populations were susceptible to glyphosate but resistant to nicosulfuron-methyl. Resistance to ALS inhibitors was due to several amino acid substitutions in positions Pro197, Trp574 and Ser653. Moreover, the substitutions Ser653Ile and Pro197Thr are described for the first time in this species. At field-labeled rates, all populations were fully controlled with alternative herbicides with other sites of action. Amaranthus palmeri individuals were clustered in three groups based on unweighted pair group method with arithmetic mean analysis, which corresponded to the three sampled populations, with a 67% of genetic relationship among them. Considering this high genetic variability and the different positions and amino acid substations found between populations, it was hypothesized that different colonization events occurred from the American continent probably prior to the introduction of glyphosate resistant crops. Prevention from new introductions is warranted because new herbicide resistance traits could arrive, complicating the management of this invasive weed species, while managing or eradicating the already established populations.This work was funded by Spanish Government through the project AGL2017-83325-C4-2-R (AEI/FEDER/UE). Joel Torra acknowledges support from the Spanish Ministry of Science, Innovation and Universities (grant Ramon y Cajal RYC2018-023866-I)

An Asp376Glu substitution in ALS gene and enhanced metabolism confers high tribenuron-methyl resistance in Sinapis alba

Acetolactate synthase (ALS) inhibiting herbicides (group 2) have been widely applied for the last 20 years to control Sinapis alba in cereal crops from southern Spain. In 2008, a tribenuron-methyl (TM) resistant (R) S. alba population was first reported in a cereal field in Malaga (southern Spain). In 2018, three suspected R S. alba populations (R1, R2 and R3) to TM were collected from three different fields in Granada (southern Spain, 100 km away from Malaga). The present work aims to confirm the putative resistance of these populations to TM and explore their resistance mechanisms. Dose–response assays showed that the R1, R2 and R3 populations ranging between 57.4, 44.4 and 57.1 times more resistance to TM than the susceptible population (S). A mutation in the ALS gene (Asp376Glu) was detected in the Rs S. alba populations. 14C-metabolism studies show that metabolites and TM were changing significantly faster in the R than in the S plants. Alternative chemical control trials showed that 2,4-D and MCPA (auxin mimics), glyphosate (enolpyruvyl shikimate phosphate synthase,EPSPS, inhibitor-group 9), metribuzin (PSII inhibitors/Serine 264 Binders, -group 5) and mesotrione (hydroxyphenyl pyruvate dioxygenase, HPPD, inhibitor-group 27) presented a high control of the four populations of S. alba tested, both S and R. Based on these results, it is the first case described where the Asp376Glu mutation and P450-mediated metabolism participates in resistance to TM in S. alba. Comparing these results with those found in the S. alba population in Malaga in 2008, where the resistance was TSR type (Pro197Ser), we can suggest that despite the geographical proximity (over 100 km), the resistance in these cases was due to different evolutionary events

Tribenuron-methyl metabolism and the rare Pro197Phe double mutation together with 2,4-D metabolism and reduced absorption can evolve in Papaver rhoeas with multiple and cross herbicide resistance to ALS inhibitors and auxin mimics

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.This work has been supported by the Asociacion de Agroquimicos y Medio Ambiente, Spain. Joel Torra acknowledges support from the Spanish Ministry of Science, Innovation, and Universities (grant Ramon y Cajal RYC2018–023866-I) and by the Spanish State Research Agency, Spain (AEI) and the European Regional Development Fund, EU (ERDF) through the projects AGL2017-83325-C4-2-R and PID2020-113229RB- C42. The field surveys made in Portugal were supported by Foundation for Science and Technology through the project UIDB/05064/2020 (VALORIZA). Jorge Lozano-Juste group is funded by grants RYC2020- 029097-I and PID2021-128826OA-I00 from Ministerio de Ciencia e Innovaci ́on (MCIN, Spain), AEI and the ERDF

Enhanced detoxification via Cyt-P450 governs cross-tolerance to ALS-inhibiting herbicides in weed species of centaurea

Centaurea is a genus of winter weeds with a similar life cycle and competitive traits, which occurs in small-grains production fields in the central-southern of the Iberian Peninsula. However, most of herbicides recommended for weed management in wheat show poor control of Centaurea species. This study summarizes the biology, herbicide tolerance to acetolactate synthase (ALS) inhibitors, and recommended chemical alternatives for the control of Centaurea species. Four species (C. cyanus L., C. diluta Aiton, C. melitensis L. and C. pullata L. subsp. baetica Talavera), taxonomically characterized, were found as the main important broadleaf weeds in small- grains production fields of the Iberian Peninsula. These species showed innate tolerance to tribenuron-methyl (TM), showing LD50 values (mortality of 50% of a population) higher than the field dose of TM (20 g ai ha 1). The order of tolerance was C. diluta (LD50 =702 g ha 1) ≫ C. pullata (LD50 =180 g ha 1) ≫ C. cyanus (LD50 =65 g ha 1) >C. melitensis (LD50 =32 g ha 1). Centaurea cyanus and C. melitensis presented higher foliar retention (150–180 μL herbicide solution), absorption (14–28%) and subsequent translocation (7–12%) of TM with respect to the other two species. Centaurea spp. plants were able to metabolize 14C-TM into non-toxic forms (hydroxylated OH-metsulfuron-methyl and conjugated-metsulfuron-methyl), with cytochrome P450 (Cyt-P450) monooxygenases being responsible for herbicide detoxification. Centaurea cyanus and C. mellitensis metabolized up to 25% of TM, while C. diluta and C. pullata metabolized more than 50% of the herbicide. Centaurea species showed 80–100% survival when treated with of florasulam, imazamox and/or metsulfuron-methyl, i.e., these weeds present cross-tolerance to ALS inhibitors. In contrast, auxin mimics herbicides (2,4-D, clopyralid, dicamba, fluroxypir and MCPA) efficiently controlled the four Centaurea species. In addition, the mixture of ALS-inhibitors and auxin mimics also proved to be an interesting alternative for the control of Centaurea. These results show that plants of the genus Centaurea found in the winter cereal fields of the Iberian Peninsula have an innate tolerance to TM and cross-resistance to other ALS-inhibiting herbicides, governed by reduced absorption and translocation, but mainly by the metabolization of the herbicide via Cyt-P450