Seaside Sparrows (Ammospiza maritima) are tidal salt marsh endemic passerines found along the Atlantic and Gulf coasts of North America. Currently, there are 7 described subspecies, and “MacGillivray’s” Seaside Sparrow (A. m. macgillivraii) is the name given to the Atlantic coast subspecies breeding from North Carolina to northern Florida. In 2019 the US Fish and Wildlife Service received a petition to list this subspecies under the Endangered Species Act due to shrinking populations and loss of marsh habitat, which necessitated a Species Status Assessment. As part of the Species Status Assessment, we analyzed genetic (microsatellite and mitochondria) data from 464 Seaside Sparrows from Connecticut to Florida, USA, to infer population connectivity (gene flow) among Atlantic coast populations, and to assess the concordance of population genetic data with the putative ranges of the subspecies. Bayesian cluster analysis (program Structure) indicates three genetically distinct population segments: (1) Florida + Georgia, (2) Charleston, South Carolina, and (3) North Carolina to Connecticut. The population in Charleston, South Carolina was the most strongly differentiated based on microsatellite data, and harbored a unique mitochondrial haplotype not shared by other sampling locations, possibly reflecting long-standing isolation. Our results indicate population genetic discordance with currently described ranges of the subspecies of Seaside Sparrow and provide grounds for the consideration of separate management plans for the three populations.