Resolution of the systematic relationships of the New England Boneset, Eupatorium novae-angliae, has been elusive. This rare species, known from only 15 sites in Massachusetts and Rhode Island, has been demonstrated to be male-sterile and agamospermous, and thus inferred to be polyploid, but its progenitor diploids have not been identified clearly. In a study that hinged on a combination of fieldwork and morphological study together with molecular analysis, we have demonstrated that E. novae-angliae contains ITS repeats characteristic of two sexual diploid species of the genus. One is the widespread E. perfoliatum, the second is a previously unrecognized endemic to clay-based Carolina bay and depression meadow habitats in the Carolinas, that had been included in E. leucolepis and is now recognized as a separate species, E. paludicola. The molecular data highlight the distinctiveness of E. novae-angliae and underscore the need for efforts to continue to protect it in its native habitat.