Freshwaters are important, interconnected, and imperiled. Aquatic ecosystems, including freshwater fishes, are closely tied to the terrestrial ecosystems they are embedded within, yet available spatially explicit datasets have been underutilized to determine associations between freshwater fishes and forested areas. Here, we determined the spatial co-occurrence between freshwater fish distributions and forests within 2129 watersheds of the conterminous United States. We identified 21% of freshwater fishes as associated with forested areas, and 2% as strictly present only in highly forested areas (75–100% forested). The northern coasts and southeast regions, both heavily forested, showed the largest numbers of forest-associated fishes in highly forested areas and fish species richness. Fish associated with low-forested areas occurred in the southwest and central plains. Imperiled fishes were relatively evenly distributed among percent forest categories, which was distinctly different from patterns for all fishes. The identification of forest-associated fishes provides insights regarding species-specific landscape contexts. Determining these large-scale patterns of freshwater biodiversity is necessary for conservation planning at regional levels, especially in highly impacted freshwater ecosystems.