The soundscape composition of temperate freshwater habitats is poorly understood. Our goal was to document the occurrence of biological and anthropogenic sounds in freshwater habitats over a large (46,000 km2) area along the geographic corridors of five major river systems in North America (Connecticut, Kennebec, Merrimack, Presumpscot, and Saco). The underwater soundscape was sampled in 19 lakes, 17 ponds, 20 rivers and 20 streams, brooks and creeks that were grouped into broad categories (brook/creek, pond/lake, and river). Over 7,000 sounds were measured from 2,750 minutes of recording in 173 locations over a five-week period in the spring of 2008. Sounds were classified into major anthropophony (airplane, boat, traffic, train and other noise) and biophony (fish air movement, also known as air passage, other fish, insect-like, bird, and other biological) categories. The three most significant findings in this study are: 1) freshwater habitats in the New England region of North America contain a diverse array of unidentified biological sounds; 2) fish air movement sounds constitute a previously unrecognized important component of the freshwater soundscape, occurring at more locations (39%) and in equal abundance than other fish sounds; and 3) anthropogenic noises dominate the soundscape accounting for 92% of the soundscape by relative percent time. The high potential for negative impacts of the anthropophony on freshwater soundscapes is suggested by the spectral and temporal overlap of the anthropophony with the biophony, the higher received sound levels of the anthropophony relative to the biophony, and observations of a significant decline in the occurrence, number, percent time, and diversity of the biophony among locations with higher ambient received levels. Our poor understanding of the biophony of freshwater ecosystems, together with an apparent high temporal exposure to anthropogenic noise across all habitats, suggest a critical need for studies aimed at identification of biophonic sound sources and assessment of potential threats from anthropogenic noises.