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QUIK STATS (last updated Dec 10, 2013 )
NOTES ABOUT THIS BIOTYPE
the last year we`ve been working with several A. quitensis biotypes wich
were not controlled with usual dosis of glyphosate in agricultural fields.
We performed greenhouse studies comparing them with a known susceptible biotipe
and we found a very high level of resistance in three of the suspected
dose response experiments were carried on using a wide range of doses from 34
g. a.e/ha up to 17280 g e.a/ha. In all the suspected biotypes we
registered surviving plants even at the highest dose tested.
Contributors: Daniel Tuesca, Juan Carlos Papa; Sergio Morichetti
and Nicolás Montero Bulacio.
A triple target-site mutation confers high levels of resistance to glyphosate in an Amaranthus quitensis population from Argentina Sarah-Jane Hutchingsa, Eddie McIndoea, Ryan Carlinb, Wenjin Yub, Anushka Howella, Weining Gub, Wenling Wangb, Mike Langforda, Jo Mattocksa and Shiv-Shankar Kaunduna a Syngenta, Jealott’s Hill International Research Centre, RG42 6EY Bracknell, UK b Syngenta, Research Triangle Park, NC USA E:mail: firstname.lastname@example.org
Resistance to glyphosate has evolved and is quickly spreading in Amaranthus quitensis due to excessive use of the EPSPS-inhibiting herbicide in Argentinian soybean production systems. Here, we confirmed resistance to glyphosate and determined the mechanism involved in an A. quitensis population (AMAQU-R) from Santa Fe Province, Argentina. AMAQU-R plants survived glyphosate rates as high as 48 kg ai/ha, whereas individuals from the sensitive populations (AMAQU-S1 and AMAQU-S2) were killed at 750 g ai/ha or below. Differential glyphosate uptake, movement or metabolism was not associated with resistance in AMAQU-R and only a small average relative increase (1.94) in EPSPS gene copy number/expression was detected in AMAQU-R. Three linked EPSPS mutations were identified around the binding site of glyphosate in AMAQU-R. These comprised T102I and P106S amino acid substitutions documented to confer high levels of glyphosate resistance, as well as a novel A103V target-site mutation. A 3.5-fold more tolerance to glyphosate was observed for the TIAVPS mutant compared to the double TIPS strain following heterologous gene expression and enzyme analysis. The triple glyphosate resistance mutations may explain the rapid spread of A. quitensis in Argentina and could be a real threat for neighbouring countries under similar glyphosate selection pressure.
A triple mutation in the epsps gene is responsable of high levels of resistance to glyphosate in Amaranthus hybridus from Argentina
Valeria Perotti, Alvaro Larran, Valeria Palmieri y Hugo Permingeat.
Traditionally, the smooth pigweed (Amaranthus hybridus, previously known as A. quitensis) has been one of the most problematic weeds in the summer crops of our country, not only for its physiology (extraordinary growth and reproduction rates) but also for the appearance of subpopulations with multiple herbicide resistance. The finding of a subpopulation resistant to ALS inhibitors and to glyphosate in Canals (Córdoba) gave us the material to fulfill an objective: to elucidate the molecular basis of the resistance, in this case, to glyphosate. After the confirmation of the agronomic resistance by dose-response assays (where LD50 and GR50 showed to be higher than 17 Kg ha-1), the accumulation of shikimate in leaf disks of single plants was determined. Thus, we obtained values from 4 to 7 ng of shikimate per microliter for sensitive plants (S), and close to 0 for resistant plants (R), suggesting a target-site mechanism. Subsequently, the sequencing of the epsps gene region enclosing the Proline 106 (a position which mutated is responsible for glyphosate resistance in different weeds) was carried out. The PCR products of 12 R plants and 2 S plants were sequenced, finding the so-called TIPS double mutation (corresponding to the substitutions T102I and P106S) in 8 R plants, confirming a target-site mechanism. A third mutation was found to cause an alanine-valine substitution at position 103 of the amino acid sequence (taking as reference the EPSPS sequence of maize). This replacement represents a very interesting finding from a biotechnological point of view, since it could be involved in the unusually high glyphosate resistance levels observed in this subpopulation. Further research is necessary to confirm this hypothesis. Finally, the comparative estimation of the epsps gene copy number was performed using the als gene as a reference. These results showed that the subpopulation of Canals has an average increase of 3.6 times relative to the subpopulation S. Thus, the present work allowed to elucidate for the first time the mechanism endowing glyphosate resistance in A. hybridus species.
Key words: smooth pigweed, EPSPS, triple mutation, Amaranthus quitensis
CONTRIBUTING WEED SCIENTISTS
The evolution of herbicide‐resistant weeds is one of the most important concerns of global agriculture. Amaranthus hybridus L. is a competitive weed for summer crops in South America. In this article, we intend to unravel the molecular mechanisms by which an A. hybridus population from Argentina has become resistant to extraordinarily high levels of glyphosate.
The glyphosate‐resistant population (A) exhibited particularly high parameters of resistance (GR50 = 20 900 g ai ha−1, Rf = 314), with all plants completing a normal life cycle even after 32X dose application. No shikimic acid accumulation was detected in the resistant plants at any of the glyphosate concentrations tested. Molecular and genetic analyses revealed a novel triple substitution (TAP‐IVS: T102I, A103V, and P106S) in the 5‐enol‐pyruvylshikimate‐3‐phosphate synthase (EPSPS) enzyme of population A and an incipient increase on the epsps relative copy number but without effects on the epspstranscription levels. The novel mechanism was prevalent, with 48% and 52% of the individuals being homozygous and heterozygous for the triple substitution, respectively. In silico conformational studies revealed that TAP‐IVS triple substitution would generate an EPSPS with a functional active site but with an increased restriction to glyphosate binding.
The prevalence of the TAP‐IVS triple substitution as the sole mechanism detected in the highly glyphosate resistant population suggests the evolution of a new glyphosate resistance mechanism arising in A. hybridus. This is the first report of a naturally occurring EPSPS triple substitution and the first glyphosate target‐site resistance mechanism described in A. hybridus.