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Knowledge of factors driving reproductive isolation is essential to understand the process of speciation. To study the reproductive isolation of two closely related species with overlapping flowering seasons, Iris domestica and Iris dichotoma, we compared their reproductive system, floral biology and pollination biology. The results indicated that I. domestica was facultatively xenogamous, whereas I. dichotoma was facultatively autogamous. Although the two species differed significantly in floral colour, floral diameter, floral structure, nectar volume, flower opening and closing times, they shared the same diurnal pollinator, the honeybee Apis cerana. The frequency of pollination by A. cerana did not differ significantly between the two species, but honeybee pollination of I. domestica was more efficient compared with that of I. dichotoma. Despite the difference in floral structure between the two species, both species deposited pollen on the same parts of the body of honeybees. The temporal partitioning of within-day flowering times between I. domestica (from 07.15 to 08.15 h to 18.00 to 19.00 h) and I. dichotoma (from 15.45 to 16.15 h to 22.00 to 23.00 h), together with the time memory of honeybees, meant that the two species did not overlap in the time of their pollination, thus leading to temporal isolation as a major driver of reproductive isolation between the two species.

Within-day temporal isolation of two species of Iris (Iridaceae) sharing the same pollinator