Technical Abstract: Diversification of phytophagous insects is often associated with changes in the use of host taxa and host parts. We focus on a group of newly discovered neotropical tephritids in the genus Blepharoneura, and report the discovery of an extraordinary number of sympatric, morphologically cryptic species all feeding as larvae on flowers of a single host species at a single field site in eastern Ecuador. The host plant, Gurania spinulosa, is functionally dioecious and highly sexually dimorphic. Molecular analyses of the mitochondrial cytochrome oxidase I gene from flies reared from flowers of G. spinulosa reveal six distinct haplotype groups that differ by 7.2-10.1% (uncorrected pairwise distances). Haplotype groups correspond to six distinct and well-supported clades. Flies in all six clades feed as larvae on calyx tissue, but members of five clades specialize on the calyces of flowers of a particular sex: three clades comprise male flowers specialists; two clades comprise female flower specialists; the sixth clade comprises generalists reared from male and female flowers. The six clades occupy significantly different morphological spaces defined by wing pigmentation patterns; however, diagnostic morphological characters were not discovered. Behavioral observations suggest specific courtship behaviors may play a role in maintaining reproductive isolation among sympatric species.