It is uncertain whether the same ecological forces that structure plant and animal communities also shape microbial communities, especially those residing in soil. We sought to uncover the relative importance of present-day environmental characteristics, climatic variation, and historical contingencies in shaping soil actinobacterial communities in a long-term chronosequence. Actinobacteria communities were characterized in surface soil samples from four replicate forest stands with nearly identical edaphic and ecological properties, which range from 9500 to 14,000 years following glacial retreat in Michigan. Terminal restriction fragment length polymorphism (TRFLP) profiles and clone libraries of the actinobacterial 16S rRNA gene were constructed in each site for phenetic and phylogenetic analysis to determine whether dispersal limitation occurred following glacial retreat, or if community composition was determined by environmental heterogeneity. At every level of examination, actinobacterial community composition most closely correlated with distance, a surrogate for time, than with biogeochemical, plant community, or climatic characteristics. Despite correlation with leaf litter C:N and annual temperature, the significant and consistent relationship of biological communities with time since glacial retreat provides evidence that dispersal limitation is an ecological force structuring actinobacterial communities in soil over long periods of time.