Seabuckthorn (Hippophae rhamnoides L.) is a small fruit tree belonging to the family Elaeagnaceae. Because seabuckthorn fruit is rich in unsaturated fatty acids and vitamins, this plant has potential as a food and medicinal crop. Here, we focused on symbiotic nitrogen fixation that could aid in the cultivation of this species. Microscopic observations showed that seabuckthorn root nodules have a standard morphology characteristic of Frankiaactinorhizal root nodules. Under nitrogen-free conditions, seabuckthorn seedlings inoculated with a homogenate of root nodules grew normally, and the fresh weight of root nodules was positively correlated with plant growth. In the field, nitrogenase activity in root nodules was high from May to September, when air temperatures were high and photosynthesis was active. Inhibition of nitrogen fixation by nitrate has been well documented in the root nodules of legumes. Therefore, we investigated the effect of nitrate on nitrogenase activity in seabuckthorn root nodules. Nitrogenase activity in seabuckthorn root nodules was not inhibited by the addition of high concentrations (up to 30 mM) of nitrate over a short term (5 days), but was apparently inhibited by long-term (20-30 days) treatments with 5 and 10 mM of nitrate.