AbstractInvasive plants along transportation corridors can significantly threaten ecosystems and biodiversity if they spread beyond anthropogenic environments. Rapid evolution may increase the ability of invading plant populations to establish in resident plant communities over time, posing a challenge to invasion risk assessment. We tested for adaptive differentiation in Dittrichia graveolens (stinkwort), an invasive species of ruderal habitat in California that is increasingly spreading away from roadsides into more established vegetation. We collected seeds from eight pairs of vegetated sites and their nearest (presumed progenitor) roadside population. We assessed differentiation between populations in roadside and vegetated habitat for germination behavior and for response to competition in a greenhouse experiment. We also tested for increased performance in vegetated habitat with a grassland field experiment including a neighbor removal treatment. Germination rates were slightly reduced in seeds from vegetated sites, which may indicate lower seed viability. Otherwise, plants did not show consistent differences between the two habitat types. Competition strongly reduced performance of D. graveolens in both the greenhouse and in the field, but plants originating from vegetated sites did not show enhanced competitive ability. Our findings show no evidence of adaptive differentiation between D. graveolens populations from roadside and vegetated habitats to date, suggesting that invasiveness in grasslands has not been enhanced by rapid evolution in the 40 + years since this species was introduced to California. Evolutionary constraints or potentially high levels of gene flow at this small scale may limit adaptation to novel habitats along roadsides.