Polyploids are thought to harbor more genetic diversity than diploids, despite the genetic bottleneck they experience upon formation. Identifying the means by which polyploids become more genetically variable can advance our understanding of the evolutionary trajectories of polyploid lineages. Gene flow is one such mechanism that can contribute to polyploid genetic diversity. Here, we used microsatellite markers to measure genetic diversity and to estimate gene flow among closely related diploid and tetraploid species of the North American plant genus Houstonia section Amphiotis. We found that tetraploid populations were more genetically variable when compared to diploid populations and that gene flow (admixture) occurred more often among tetraploid populations than among diploids. In addition, we provide evidence to support previous reports that autopolyploidy is the likely mode of polyploid formation in this group. We suggest that gene flow among closely related species, particularly among tetraploid populations, has likely contributed to the genetic diversity levels of tetraploid lineages of Houstonia species.