Non Target Site Tolerance Mechanisms Describe Tolerance to Glyphosate in Avena sterilis

Sterile wild oat (Avena sterilis L.) is an autogamous grass established in warm climate regions. This species has been used as a cover crop in Mediterranean perennial crops during the spring period prior to initiating competition with the main crop for water and nutrients. However, such cover crops need to be controlled (by glyphosate or tillage) before the beginning of summer period (due to the possibility of intense drought stress). In 2011, the olive grove farmers of southern Spain expressed dissatisfaction because of the ineffective control with glyphosate on A. sterilis. Experiments were conducted to determine whether the continued use of glyphosate over a 5 year period had selected a new resistant or tolerant species. The GR50 values obtained for A. sterilis were 297.12 and 245.23 g ae ha-1 for exposed (E) and un-exposed (UE) glyphosate accessions, respectively. The spray retention and shikimic acid accumulation exhibited a non-significant difference between the two accessions. The results of 14C- glyphosate absorption was the same in the two accessions (E and UE), while the translocation from the treated leaf to the rest of the shoots and roots was similar in A. sterilis accessions. Glyphosate metabolism to aminomethylphosphonic acid (AMPA) and glyoxylate was similar in both accessions, but increased after treatment with glyphosate, indicating that metabolism plays an important role in tolerance. Both A. sterilis accessions, present similarity in the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) activity enzyme with different glyphosate concentrations and without glyphosate, confirming that both accessions present the same genomic characteristics. The above-mentioned results indicate that innate tolerance to glyphosate in A. sterilis is probably and partly due to reduced herbicide absorption and translocation and metabolism compared to the susceptibility of other grasses weeds like Chloris inflata, Eleusine indica and Lolium rigidum

From tolerance to resistance: mechanisms governing the differential response to glyphosate in Chloris barbata

Susceptibility and the mechanism (s) governing tolerance/resistance to glyphosate were characterized in two putative-glyphosate-resistant Chloris barbata populations (R1 and R2), collected in Persian lime orchards from Colima State, Mexico, comparing them with one non-treated population (referred to as S). Glyphosate doses required to reduce fresh weight or cause mortality by 50% were 4.2–6.4 times higher in resistant populations than in the S population. The S population accumulated 4.3 and 5.2 times more shikimate than the R2 and R1 popu- lations, respectively. There were no differences in 14 C-glyphosate uptake between R and S populations, but the R plants translo- cated at least 12% less herbicide to the rest of plant and roots 96 h after treatment. Insignificant amounts of glyphosate were metabolized to aminomethyl phosphonate and glyoxylate in both R and S plants. The 5-enolpyruvylshikimate-3-phosphate syn- thase gene of the R populations contained the Pro106-Ser mutation, giving them a resistance 12 (R2) and 14.7 (R1) times greater at target-site level compared with the S population. The Pro106-Ser mutation governs the resistance to glyphosate of the R1 and R2 C barbata populations, but the impaired translocation could contribute to the resistance. These results confirm the first case of glyphosate resistance evolved in this species

The intensity of non-target site mechanisms influences the level of resistance of sourgrass to glyphosate

ABSTRACT Non-target site mechanisms are involved in the resistance of sourgrass (Digitaria insularis) to glyphosate. Studies on the 14C-glyphosate absorption and translocation as well as the detection of glyphosate and its metabolites in sourgrass plants were carried out under controlled conditions to investigate if the differential response of resistant sourgrass biotypes (R1 and R2) is derived from the intensity of non-target site mechanisms involved in the resistance to glyphosate. Different pattern of absorption was observed between S (susceptible) and R2 from 12 up to 48 hours after treatment with glyphosate (HAT), and between S and R1 just at 12 HAT. The initial difference in glyphosate absorption among the biotypes did not maintained at 96 HAT and afterwards. Smaller amount of herbicide left the treated leaf into the rest of shoot and roots in R2 (25%) than in S (58%) and R1 (52%). In addition, slight difference in glyphosate translocation was observed between S and R1. We found high percentage (81%) of glyphosate in the S biotype up to 168 HAT, while just 44% and 2% of glyphosate was recovered from R1 and R2 plant tissues. In addition, high percentage of glyphosate metabolites was found in R2 (98%) and R1 (56%) biotypes, while a very low percentage (11%) was found in the S biotype. As previous studies indicated resistant factors of 3.5 and 5.6 for R1 and R2, respectively, we conclude that the differential response of sourgrass biotypes is derived from the intensity of the non-target site mechanisms involved in the resistance to glyphosate

First confirmation and characterization of target and non-target site resistance to glyphosate in Palmer amaranth (Amaranthus palmeri) from Mexico

Following the introduction of glyphosate-resistant (GR)-cotton crops in Mexico, farmers have relied upon glyphosate as being the only herbicide for in-season weed control. Continuous use of glyphosate within the same year and over multiple successive years has resulted in the selection of glyphosate resistance in Palmer amaranth (Amarantus palmeri). Dose-response assays confirmed resistance in seven different accessions. The resistance ratio based on GR50 values (50% growth reduction) varied between 12 and 83. At 1000 μM glyphosate, shikimic acid accumulation in the S-accession was 30- to 2-fold higher at compared to R-accessions. At 96 h after treatment, 35–44% and 61% of applied 14C-glyphosate was taken up by leaves of plants from R- and S-accessions, respectively. At this time, a significantly higher proportion of the glyphosate absorbed remained in the treated leaf of R-plants (55–69%) compared to S-plants (36%). Glyphosate metabolism was low and did not differ between resistant and susceptible plants. Glyphosate was differentially metabolized to AMPA and glyoxylate in plants of R- and S-accessions, although it was low in both accessions (<10%). There were differences in 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) enzyme activity by 50% (I50) between R- and S-accessions. However, no significant differences were found in the basal EPSPS activity (μmol inorganic phosphate μg−1 total soluble protein min−1) between R- and S-accessions. A point mutation Pro-106-Ser was evidenced in three accessions. The results confirmed the resistance of Palmer amaranth accessions to glyphosate collected from GR-cotton crops from Mexico. This is the first study demonstrating glyphosate-resistance in Palmer amaranth from Mexico

First resistance mechanisms characterization in glyphosate-resistant Leptochloa virgata.

Leptochloa virgata (L.) P. Beauv. is an annual weed common in citrus groves in the states of Puebla and Veracruz, Mexico limiting their production. Since 2010, several L. virgata populations were identified as being resistant to glyphosate, but studies of their resistance mechanisms developed by this species have been conducted. In this work, three glyphosate-resistant populations (R8, R14 and R15) collected in citrus orchards from Mexico, were used to study their resistance mechanisms comparing them to one susceptible population (S). Dose-response and shikimic acid accumulation assays confirmed the glyphosate resistance of the three resistant populations. Higher doses of up to 720 g ae ha-1 (field dose) were needed to control by 50% plants of resistant populations. The S population absorbed between 7 and 13% more 14C-glyphosate than resistant ones, and translocated up to 32.2% of 14C-glyphosate to the roots at 96 h after treatment (HAT). The R8, R14 and R15 populations translocated only 24.5, 26.5 and 21.9%, respectively. The enzyme activity of 5-enolpyruvyl shikimate-3-phosphate synthase (EPSPS) was not different in the S, R8 and R14 populations. The R15 Population exhibited 165.9 times greater EPSPS activity. Additionally, this population showed a higher EPSPS basal activity and a substitution in the codon 106 from Proline to Serine in the EPSPS protein sequence. EPSPS gene expression in the R15 population was similar to that of S population. In conclusion, the three resistant L. virgata populations show reduced absorption and translocation of 14C-glyphosate. Moreover, a mutation and an enhanced EPSPS basal activity at target-site level confers higher resistance to glyphosate. These results describe for the first time the glyphosate resistance mechanisms developed by resistant L. virgata populations of citrus orchards from Mexico

Target and Non-Target Site Mechanisms Developed by Glyphosate-Resistant Hairy beggarticks (Bidens pilosa L.) Populations from Mexico

In 2014 hairy beggarticks (Bidens pilosa L.) has been identified as being glyphosate-resistant in citrus orchards from Mexico. The target and non-target site mechanisms involved in the response to glyphosate of two resistant populations (R1 and R2) and one susceptible (S) were studied. Experiments of dose-response, shikimic acid accumulation, uptake-translocation, enzyme activity and EPSPS gene sequencing were carried out in each population. The R1 and R2 populations were 20.4 and 2.8-fold less glyphosate sensitive, respectively, than the S population. The resistant populations showed a lesser shikimic acid accumulation than the S population. In the latter one, 24.9% of 14C-glyphosate was translocated to the roots at 96 h after treatment; in the R1 and R2 populations only 12.9 and 15.5%, respectively, was translocated. Qualitative results confirmed the reduced 14C-glyphosate translocation in the resistant populations. The EPSPS enzyme activity of the S population was 128.4 and 8.5-fold higher than the R1 and R2 populations of glyphosate-treated plants, respectively. A single (Pro-106-Ser), and a double (Thr-102-Ile followed by Pro-106-Ser) mutations were identified in the EPSPS2 gene conferred high resistance in R1 population. Target-site mutations associated with a reduced translocation were responsible for the higher glyphosate resistance in the R1 population. The low-intermediate resistance of the R2 population was mediated by reduced translocation. This is the first glyphosate resistance case confirmed in hairy beggarticks in the world

Pool of Resistance Mechanisms to Glyphosate in Digitaria insularis

Digitaria insularis biotypes resistant to glyphosate have been detected in Brazil. Studies were carried out in controlled conditions to determine the role of absorption, translocation, metabolism, and gene mutation as mechanisms of glyphosate resistance in D. insularis. The susceptible biotype absorbed at least 12% more C-14-glyphosate up to 48 h after treatment (HAT) than resistant biotypes. High differential C-14-glyphosate translocation was observed at 12 HAT, so that >70% of the absorbed herbicide remained in the treated leaf in resistant biotypes, whereas 42% remained in the susceptible biotype at 96 HAT. Glyphosate was degraded to aminomethylphosphonic acid (AMPA), glyoxylate, and sarcosine by >90% in resistant biotypes, whereas a small amount of herbicide (up to 11%) was degraded by the susceptible biotype up to 168 HAT. Two amino acid changes were found at positions 182 and 310 in EPSPS, consisting of a proline to threonine and a tyrosine to cysteine substitution, respectively, in resistant biotypes. Therefore, absorption, translocation, metabolism, and gene mutation play an important role in the D. insularis glyphosate resistance.Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq