Genetic structure may be highly variable across seabird species, and particularly among those that are distributed over large geographical areas. The Adélie penguin (Pygoscelis adeliae) is a numerically dominant Antarctic seabird that is considered to be a key species in coastal ecosystems. Since the Last Glacial Maximum, penguin colonization of the Antarctic coastline occurred at varying geographical and temporal scales, contributing to an incomplete understanding of how modern colonies relate to each other at local or regional scales. We assessed the population genetic structure of Adélie penguins (n = 86 individuals) from three adjacent colonies along the Victoria Land coast using molecular genetic markers (i.e. seven microsatellite loci isolated through next-generation sequencing). Our results indicate meta-population dynamics and possibly relationships with habitat quality. A generally low genetic diversity (Nei's index: 0.322–0.667) was observed within each colony, in contrast to significant genetic heterogeneity among colonies (pairwise FST = 0.071–0.148), indicating that populations were genetically structured. Accordingly, an assignment test correctly placed individuals within the respective colonies from which they were sampled. The presence of inter-colony genetic differentiation contrasts with previous studies on this species that showed a lack of genetic structure, possibly due to higher juvenile or adult dispersal. Our sampled colonies were not panmictic and suggest a lower migration rate, which may reflect relatively stable environmental conditions in the Ross Sea compared to other regions of Antarctica, where the ocean climate is warming.