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Mestrado em Engenharia Agronómica - Proteção de Plantas - Instituto Superior de Agronomia This dissertation reports the study of two resistance mechanisms in a glyphosate-resistant Conyza canadensis population (B15), comparatively with a susceptible population (B), from intensive olive orchards in Alentejo (south of Portugal). Initially, the presence of resistance was confirmed by a shikimate leaf disc assay. Then, the sequence of the gene encoding the enzyme 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) was determined and, lastly, were analysed the expression levels of the gene encoding this enzyme. The shikimate leaf disc assay showed a higher accumulation of shikimate in the population B, relatively to population B15, 24 hours after being exposed to glyphosate. The EPSPS-2 cDNA sequencing didn’t reveal differences in enzyme’s constitution between the two populations. The analysis of gene expression also revealed no differences in expression levels of this enzyme, 24 hours after glyphosate application. These results suggest that the studied population has no EPSPS enzyme mutation or overexpression as a mechanism of resistance to glyphosate

Mestrado em Engenharia Agronómica - Proteção de plantas - Instituto Superior de Agronomia The objective of this study was to assess efficacy of the glyphosate in three populations of Conyza canadensis, two suspected of resistance (B15-1 and B15-2) and a susceptible one (B), from intensive olive orchards in Alentejo, south of Portugal. There were performed: seedling-Petri plate assays, greenhouse dose-response assays and a shikimate accumulation assay. The seedling-petri plate assays showed that B population had significant higher germination (60-80%) than B15-1 (31.3%) and B15-2 (52.7%) populations. There was evidence of resistance in the B15-1 population and susceptibility of the B15-2 population. Mortality was the key parameter to distinguish between populations by the discriminating concentration: 0.3 mM glyphosate. The dose-response assays revealed a resistance level of 0.52 and ED50 of 59.2 g e.a. ha-1 (Bioassay I) and 87.2 g e.a. ha-1 (Bioassay II) for B15-2 population. The B population showed an ED50 of 113.3 g e.a. ha-1. The shikimate accumulation assay indicated susceptibility of both populations (B and B15-2), although the accumulation was higher in the B population (0.69 μg mg-1) than in the B15-2 population (0.49 μg mg-1), six days after application (720 g ha-1). The results showed evidence of resistance in the B15-1 population and confirmed the susceptibility of B and B15-2 populations.

Mestrado em Engenharia Agronómica - Protecção das Plantas - Instituto Superior de Agronomia The objective of this study was to determine the efficacy of this herbicide in two populations of Conyza bonariensis, one suspected of resistance (B8) and a susceptible one (C), from olive groves and citrus orchards in Alentejo, south of Portugal. Assays were performed on germination agar medium, glyphosate dose-response assays in petri dishes with different concentrations of glyphosate and a greenhouse (1) and laboratory (2) study were conducted (1) to confirm and characterize glyphosate resistance and (2) to determine whether resistant and susceptible populations have differential shikimate accumulation. Population B8 showed significant higher germination and emergence than population C but lower mortality rate. Dose–response studies revealed that the suspected resistant population was two- to three-fold less sensitive to glyphosate than the susceptible population. The dose-response flazasulfuron assay with whole plant in the rosette stage revealed the doses were completely effective on both populations. Shikimate accumulation was higher in resistant population at 72-h after treatment, for the recommended concentration of glyphosate. However, it was lower for concentrations of 360 g ha-1. These results pointed that population B8 is resistant to glyphosate and flazasulfuron is an option for the chemical control of resistant populations.