Potential basis of glyphosate resistance in Lolium perenne from Douro vineyards

Glyphosate resistant Lolium perenne populations have been identified in Douro vineyards – north Portugal whose resistance mechanisms were not yet fully elucidated. Therefore, we studied the resistance of these two biotypes under greenhouse and laboratory conditions. The objective of this study was to confirm whether target site resistance could explain the differences between sensitive and resistant L. perenne biotypes. In order to confirm this hypothesis, shikimate leaf disc assay, sequencing of 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene and the estimation of its number of copies were carried out. After being exposed to glyphosate, the GR population showed a lower accumulation of shikimate compared to a reference susceptible population. The sequencing of cDNA sequence encoding the EPSPS enzyme revealed that the most frequent point mutations responsible for glyphosate resistance [Pro106Ser/Thr/Gly/Cys/Ala/Ile/Val/Met/Leu] were not present. The estimation of the relative EPSPS gene copy number, by means of quantitative real time-PCR and ΔΔCt value, revealed a two-fold amplification of the EPSPS gene. The overexpression of the EPSPS gene had contributed as a mechanism of resistance to glyphosate. Further studies on glyphosate translocation are in course