The taxonomic identity of endangered populations of the Least Tern (Sternula antillarum) has long been debated. Their current conservation status provides even more impetus to examine the taxonomic distinctness of these groups. We used rapidly evolving mitochondrial DNA control-region sequences (840 base pairs; n = 188) and microsatellite DNA data (7 loci; n = 417) to examine genetic structure within and among three subspecies that occur within the United States: California Least Tern (S. a. browni), Interior Least Tern (S. a. athalassos), and Eastern Least Tern (S. a. antillarum). Although significant genetic structure was observed among breeding populations from across the species' range, our data indicated little evidence of genetic structure within traditional subspecific groups. Isolation-by-distance analyses, however, identified subtle patterns that may reflect sex-specific differences in dispersal behavior. Our analyses likewise demonstrated little population subdivision among subspecific groups, which raises questions regarding the taxonomic status of traditionally defined subspecies. Our findings can therefore be used to consider a reevaluation of Least Tern subspecies by the American Ornithologists' Union's Committee on Taxonomy and Nomenclature. We further emphasize the need for studies of range-wide breeding-site fidelity and natal philopatry to better understand interpopulation movements of individuals throughout the annual cycle.