Historically for bats, the southwestern Brazilian Amazon has had scant biological data available, which compromised large-scale comparisons and macroecological studies that could support conservation initiatives in the area. We tested faunal similarity among 26 well-sampled bat assemblages distributed throughout the Amazon, including our database from surveys in the upper Madeira River region, a core area of the southwestern Brazilian Amazon. To document bats we conducted nocturnal mist-net sampling under standardized Rapid Assessments for Long-term Ecological Research (RAPELD) protocols in forests and farmlands, and diurnal search of roosts in rocky outcrops located along the riverbed of the Madeira River. We captured 2930 bats representing 66 species, 20 of which were previously unreported for the region. Thirty-four species recorded were exclusive to forests, and two to the rocky outcrops of the Madeira River. Frugivores outnumbered the other trophic guilds, followed by the gleaning insectivores in forests, and then aerial insectivores in the farmlands. The southwestern Brazilian Amazon fauna is more similar to others from the western Amazon and less similar to the bat fauna from the eastern Amazon and the Guianas. This geographic association is undoubtedly related to a more recent history of formation of the terrestrial ecosystems of the western Amazon lowlands. With 87 species currently known, the southwestern Brazilian Amazon is now one of the richest areas in the world for bats, which helps to understand the limits of the distributional patterns between eastern and western Amazon bat faunas.