BACKGROUND: 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.
RESULTS:The glyphosate-resistant population (A) exhibited particularly high parameters of resistance (GR 50 = 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 epsps transcription 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. CONCLUSION: 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. /journal/ps estimation of plant fitness costs associated with herbicide-resistance genes. Weed Sci 63:203-216 (2015). 42 Schönbrunn E, Eschenburg S, Shuttleworth WA, Schloss JV, Amrhein N, Evans JN, et al., Interaction of the herbicide glyphosate with its target enzyme 5-enolpyruvylshikimate 3-phosphate synthase in atomic detail. Proc Natl Acad Sci 98:1376-1380 (2001). 43 Pollegioni L, Schonbrunn E and Siehl D, Molecular basis of glyphosate resistance-different approaches through protein engineering.
BACKGROUND: Amaranthus palmeri S. Watson, a problematic weed infesting summer crops in Argentina, has developed multiple herbicide resistance. Resistance to acetolactate synthase (ALS)-inhibiting herbicides is particularly common, with highlevel resistance mostly caused by different mutations in the ALS enzyme. Six versions of the enzyme were identified from a resistant A. palmeri population, carrying substitutions D376E, A205V, A122S, A282D, W574L and S653N. This work aims to provide a comparative analysis of these mutants and the wild-type (WT) enzyme to fully understand the herbicide resistance. Thus, all the versions of the ALS gene from A. palmeri were heterologously expressed and purified to evaluate their kinetics and inhibitory response against imazethapyr, diclosulam, chlorimuron-ethyl, flucarbazone-sodium and bispyribac-sodium.RESULTS: A decrease in catalytic efficiency was detected in the A205V, A122S-A282D, W574L and S653N ApALS enzymes, whereas only A205V and W574L substitutions also produced a decrease in the substrate affinity. In vitro ALS inhibition assays confirmed cross-resistance to almost all the herbicides tested, with the exception of A282D ApALS, which was as susceptible as WT ApALS. Moreover, the results confirmed that the novel substitution A122S provides cross-resistance to at least one herbicide within each of the five families of ALS inhibitors, and this property could be explained by a lower number of hydrophobic interactions between the herbicides and the mutant enzyme.CONCLUSION: This is the first report to compare various mutations in vitro from A. palmeri ALS. Our data contribute to understanding the impacts of herbicide resistance in this species.
Amaranthus palmeri S. Waston is currently one of the most problematic weeds worldwide. Biotypes with resistance to herbicides such as glyphosate and ALS inhibitors are now present in almost all Argentinian cultivable areas. In this work, we studied glyphosate resistance in three different populations, some of them previously characterized as resistant to ALS inhibitors. Dose-response curves were conducted in order to assess the effect of glyphosate on the survival and dry biomass of the populations. Subsequently, the presence of target-site resistance (TSR) was studied. Results confirmed the glyphosate resistance in the three populations, showing different levels of resistance, being R2 and R3 significantly more resistant than r1 population. A high prevalence of the P106S substitution was detected in the three resistant populations, while none increase in the relative EPSPS copy number was noticed. Some surviving plants without any of the TSR mechanisms for glyphosate were detected in R3 population, suggesting the presence of non-target-site resistance (NTSR).
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