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• Reinforcement is the process through which prezygotic reproductive barriers evolve in sympatry due to selection against hybridization between co-occurring, closely related species. The role of self-fertilization in reinforcement and reproductive isolation is uncertain in part because its efficiency as a barrier against heterospecific mating can depend on the timing of autonomous selfing. • To investigate whether increased autonomous selfing has evolved as a mechanism for reinforcement, we compared Phlox cuspidata populations across their native Texas range using both estimates of genetic diversity and experimental manipulation with morphological measurements. Specifically, we investigated patterns of variation in floral traits and timing of selfing between individuals from allopatric populations of P. cuspidata and from populations sympatric with the closely related species, P. drummondii. • We infer intermediate rates of selfing across field-collected individuals with no significant difference between allopatric and sympatric populations. Among greenhouse grown plants, we find no differences in timing of selfing or other floral traits including anther dehiscence timing, anther-stigma distances, autonomous selfing rate and self-seed count between allopatric and sympatric populations. However, our statistical analyses indicate that P. cuspidata individuals sympatric with P drummondii seem to have generally larger flowers compared to allopatric individuals. • Despite strong evidence of costly hybridization with P. drummondii, we find no evidence of trait divergence due to reinforcement in P. cuspidata. Although we document nearly complete autonomous self-seed set in the greenhouse, estimates of selfing rates from genetic data imply realized selfing is much lower in nature suggesting an opportunity for reinforcing selection to act on this trait. 

Abstract PremiseA central goal of pollination biology is to connect plants with the identity of their pollinator(s). While predictions based on floral syndrome traits are extremely useful, direct observation can reveal further details of a species' pollination biology. The wildflowerPhlox drummondiihas a floral syndrome consistent with pollination by Lepidoptera. We tested this prediction using empirical data. MethodsWe observed each step of pollination inP. drummondii. First, we observed 55.5 h of floral visitation across the species range. We used temporal pollinator exclusion to determine the contribution of diurnal and nocturnal pollination to reproductive output. We then quantifiedP. drummondiipollen transfer by the dominant floral visitor,Battus philenor. Finally, we tested the effect ofB. philenorvisitation onP. drummondiireproduction by quantifying fruit set following single pollinator visits. ResultsBattus philenoris the primary pollinator ofP. drummondii. Pollination is largely diurnal, and we observed a variety of lepidopteran visitors during the diurnal period. However,B. philenorwas the most frequent visitor, representing 88.5% of all observed visits. Our results show thatB. philenoris an extremely common visitor and also an effective pollinator by demonstrating that individuals transfer pollen between flowers and that a single visit can elicit fruit set. ConclusionsOur data are consistent with the prediction of lepidopteran pollination and further reveal a single butterfly species,B. philenor, as the primary pollinator. Our study demonstrates the importance of empirical pollinator observations, adds to our understanding of pollination mechanics, and offers a specific case study of butterfly pollination.