Understanding how new species arise remains a persistent challenge in evolutionary biology. Extensive studies on a range of organisms have shown that the speciation process is complex and involves the development of several mechanisms for reproductive isolation. Recently, it has been hypothesized that postcopulatory-prezygotic isolation, which occurs in the female reproductive tract, may play an important role in speciation. Empirical evidence of specific incompatibilities, however, remains sparse. Here we use matings between two related species of Drosophila, D. mojavensis and D. arizonae, to demonstrate the role of reproductive tract interactions in speciation and to identify aberrant phenotypes in heterospecifically mated females.