The effect of chlorsulfuron on growth, infective ability, and symbiotic performance of legume bacteria was investigated. Bacterial growth in pure culture was unaffected by the addition of 0.55 and 5.5 μM chlorsulfuron. Early root hair infections of alfalfa by bacteria were inhibited by chlorsulfuron at 0.28 pM but not at 0.0028 pM in the root media. Inhibition of infection resulted from herbicidal effects on the root hair development. When inoculated red clover and alfalfa plants were grown in aseptic cultures in the presence of 0, 5.5, or 55 μM of chlorsulfuron, only 55 μM of herbicide inhibited the development and nodulation of plants. Early emergence and growth of alfalfa plants in soil supplemented with 2 × 10-6, 2 × 10-3, 2, 4, and 8 g/ha of chlorsulfuron were unaffected. However, 5 to 8 days after emergence plants grown in soil supplemented with 4 or 8 g/ha of chlorsulfuron were severely damaged, with no nodules developed. Nodulation occurred in plants grown in soil containing 2 × 10-3, 3 × 10-6, and 2 g/ha chlorsulfuron, but the nitrogenase activity of the nodules of these plants was less than for control plants. Plants grown in soil containing 2 g/ha of chlorsulfuron developed normally only after an initial growth inhibition of 5 to 6 weeks. The inhibition of nodulation and nitrogenase activity of nodules grown in the presence of chlorsulfuron is probably due to adverse effects of the herbicide on plant growth and development rather than on the rhizobia.