Microsatellite DNA provided evidence that Atlantic cod overwintering in inshore Newfoundland are genetically distinguishable from cod overwintering offshore. We compared variation in five loci in samples from inshore locations around Trinity Bay, Newfoundland, and from an offshore region on the northern Grand Bank (North Cape). Cod collected inshore were divided into two groups on the basis of antifreeze level in blood: those with high antifreeze levels presumed to have overwintered in cold ( 2°C) waters. Fish overwintering inshore differed from offshore fish in allele sharing and in (δμ) 2 distances, while fish with low antifreeze levels did not. Subpopulation structure ( Rst) was detected when offshore cod were compared with inshore cod with high levels of antifreeze but not when compared with those with low levels of antifreeze. These results suggest that cod overwintering inshore are genetically distinct from offshore cod. Inshore and offshore cod from the areas studied remain genetically distinct despite the fact that individuals from the two populations intermingle inshore during the summer and fall feeding migration. Thus, we found evidence of population structure at a finer geographical scale than has been shown to date for this species.