Dams are obstructing rivers worldwide, impairing habitat and migration opportunities for many freshwater fish species; however, global data linking dam and fish distributions have been limited. Here, we quantify dam obstruction at the biogeographic scale of freshwater ecoregion, which provides the spatial framework necessary to assess the risk of fish species loss due to dams and allows us to identify both ecoregions and genera at risk. Nearly 50% of the 397 assessed freshwater ecoregions are obstructed by large- and medium-size dams, and approximately 27% face additional downstream obstruction. A synthesis of obstruction data and fish traits indicates that taxa such as lampreys (Lampetra spp.), eels (Anguilla spp.), and shads (Alosa spp.) are at particular risk of species loss. Threatened ecoregions with heavy dam obstruction and above-average counts of total, diadromous, or endemic species are found on all continents and include the Murray—Darling Province, Southern Italy, the Lower and Middle Indus Basin, West Korea, the South Atlantic region of the United States, the Upper Paraná, and Mobile Bay ecoregions.