Stingless bees are abundant and diverse key actors in several plant-pollinator networks in the neotropics, but little is known about their natural history and ecology. This study aims to contribute to knowledge about the diversity and dispersion of stingless bees and discusses the importance of nesting substrates. It was carried out in the Araguari river valley in Minas Gerais, Brazil, where a nest site survey was conducted in an area of 100 ha during 11 alternate months from 2006 to 2008, for a total of 1,200 observation hours. Sixty-nine nests were found, belonging to 12 genera and 20 different species. Nests ofMelipona rufiventriswere by far the most abundant. Stingless bees nested more frequently in hollows of live trees (64%), and 11 different substrates were identified. Seventeen plant species were used as nesting substrates andAnadenanthera macrocarpa(Fabaceae) was the main host, encompassing 23% of the surveyed nests. The area studied is important for the maintenance of stingless bees because it provides nesting sites for them. Without nesting sites the reproductive division of colonies is compromised, affecting the nests' survival.