Differences in style length confer prezygotic isolation between two dioecious species of Silene in sympatry.

One fundamental signature of reinforcement is elevated prezygotic reproductive isolation between related species in sympatry relative to allopatry. However, this alone is inadequate evidence for reinforcement, as traits conferring reproductive isolation can occur as a by-product of other forces. We conducted crosses between Silene latifolia and S. diclinis, two closely related dioecious flowering plant species. Crosses with S. latifolia mothers from sympatry exhibited lower seed set than mothers from five allopatric populations when S. diclinis was the father. However, two other allopatric populations also exhibited low seed set. A significant interaction between style length and sire species revealed that seed set declined as style length increased when interspecific, but not intraspecific, fathers where used. Moreover, by varying the distance pollen tubes had to traverse, we found interspecific pollen placement close to the ovary resulted in seed set in both long- and short-styled S. latifolia mothers. Our results reveal that the long styles of S. latifolia in sympatry with S. diclinis contribute to the prevention of hybrid formation. We argue that forces other than reinforcing selection are likely to be responsible for the differences in style length seen in sympatry.