Human population growth and economic development threaten the integrity of freshwater ecosystems globally, reducing their ability to support biodiversity and provide ecosystem services. However, our knowledge of freshwater biodiversity is fragmented due to bias in conservation research toward primarily terrestrial or charismatic taxonomic groups. Here, we utilize the most comprehensive assessment of freshwater biodiversity for an entire continent to examine the implications of this shortfall. Results indicate that groups that have been the focus of most conservation research are poor surrogates for patterns of both richness and threat for many freshwater groups, and that the existing protected area network underrepresents freshwater species. Areas of highest species richness and threat are congruent with areas where reliance on ecosystem services by humans and pressures placed on freshwater ecosystems are high. These results have implications for targets to reduce biodiversity loss and safeguard associated ecosystem services on which millions of people depend globally.