CONICET | Buscador de Institutos y Recursos Humanos

Crop cultivation under no-till systems associated with different glyphosate resistant crop varieties has changed the weed flora, including herbicide resistant biotypes. Glyphosate is an inhibitor of the enzyme EPSPS (5-enolpyruvylshikimate-3-phosphate synthase). The accumulation of shikimate from leaf disks can be used to diagnose glyphosate resistance in weeds. The objective of this study was to evaluate the shikimate accumulation in two Johnsongrass (Sorghum halepense (L.) Pers.) biotypes. Leaf disks with 4 mm diameter were extracted from plants (GR) originated from Salta (Argentina), where glyphosate applications have failed to control Johnsongrass., and from Tucumán (Argentina), without history of use of this herbicide. Leaf disks were incubated in each well of a 96-well microtiter plate containing glyphosate concentrations ranging from 2 to 500 microM. Shikimate was determined spectrophotometrically to an optical density of 380 nm. A shikimate standard curve was determined from the known herbicide concentration, compared to the dishes without herbicide. The results demonstrate the accumulation of shikimate as the product concentration increased. The maximum shikimate concentration on the susceptible plants (GS) was 25.16 microg/ml and was obtained at the concentration of 166 microM of the herbicide. The resistant plants continued accumulating shikimate until the concentration of 500 microM, with maximum shikimate value of 15.10 microg/ml. For all glyphosate concentration range, GS plants accumulate greater amount of shikimate than the GR, which indicates the presence of the resistance. The values of IC50 (glyphosate concentration needed for 50% of reduction of the shikimate levels) were of 43.79 and 61.86 microM for the GS and GR plants, respectively. The resistance factor (IC50 for GR/ IC50 for GS) was 1.41. Altogether, these results suggest that resistance may be caused either by an alteration in EPSPS enzyme or a higher EPSPS activity in the resistant biotype. This method confirms preliminary data from herbicide dose-response curves obtained with plants by our research group, which confirmed glyphosate resistance in Johnsongrass. This research method is a quick diagnosing tool for the detection of glyphosate resistance.