<p>Herbicide resistance poses a threat to sustainable vegetation management. Recently, the first report of 2,4-D resistance in buckhorn plantain (<i>Plantago lanceolata</i> L.) as well as the first report of 2,4-D resistance in turf was published. Additional 2,4-D resistant buckhorn plantain ecotypes have been reported in Indiana, Ohio, Pennsylvania, Georgia, and Virginia in the short time since. Thus, the aims of this research were to investigate the mechanism(s) of 2,4-D resistance in a resistant ecotype of buckhorn plantain, screen other potentially resistant ecotypes and measure them for fitness penalties, and identify effective turfgrass cultural control practices for managing buckhorn plantain. </p><p><br></p><p>A radiolabeled 2,4-D experiment was conducted to investigate absorption and translocation, and a 2,4-D dose-response experiment was conducted using malathion as a cytochrome P450 inhibitor to assess the potential mechanism of 2,4-D resistance in buckhorn plantain. The clearest difference between the resistant (IN-GW) and susceptible ecotype (IN-WL) was the interaction between ecotype and harvest period for [¹⁴C]2,4-D in the non-treated shoots. After 192 hr, the susceptible ecotype had a higher amount of [¹⁴C]2,4-D in the non-treated shoots (16.1%) than the resistant ecotype at any of the harvest periods (5.5-7.3%); the amount of [¹⁴C]2,4-D in the non-treated shoots was similar across all three harvest periods for the resistant ecotype. Thus, reduced translocation plays an apparent role in 2,4-D resistance in buckhorn plantain. Malathion pre-treatment did not fully revert the resistant ecotype back to susceptible. Thus, if cytochrome P450 metabolism is part of the 2,4-D resistance mechanism of this buckhorn plantain ecotype, it is likely a contributor and not the sole mechanism of resistance. </p><p><br></p><p>In total, this research identified four 2,4-D resistant buckhorn plantain ecotypes from Indiana and one from Ohio. Only one report of a failure to control buckhorn plantain was confirmed to be a susceptible ecotype. When compared to susceptible ecotypes in a garden study, no major fitness penalties were identified in resistant ecotypes. </p><p><br></p><p>Given that no specific cultural or biological control methods of buckhorn plantain have been recognized to date, two field trials were conducted to investigate the influence of 1) mowing height and nitrogen rate on buckhorn plantain coverage and 2) mowing frequency on buckhorn plantain coverage and seed production. Nitrogen fertilization and low mowing reduced buckhorn plantain coverage after 3 yr, but low mowing also increased crabgrass and dandelion as well as reduced turf quality. Frequent mowing reduced viable seed production, but that did not translate into a reduction in buckhorn plantain coverage after 2 yr. </p><p><br></p><p>This research demonstrates the complex mechanism of action of 2,4-D, as the resistance mechanism for buckhorn plantain was not fully elucidated. It also highlights the importance of utilizing best management practices for managing weeds in turf, including rotating herbicide chemistries, high and frequent mowing, and nitrogen fertilization.</p>