Freshwater biodiversity is declining dramatically, and the current biodiversity crisis requires defining bold goals and mobilizing substantial resources to meet the challenges. While the reasons are varied, both research and conservation of freshwater biodiversity lag far behind efforts in the terrestrial and marine realms. We identify fifteen pressing global needs to support informed global freshwater biodiversity stewardship. The proposed agenda aims to advance freshwater biodiversity research globally as a critical step in improving coordinated action towards its sustainable management and conservation.