AbstractDespite the large number of dams in most important rivers of Brazil, little is known about fish vertical distribution in neotropical reservoirs, especially deep ones. This study aims to understand how fish were vertically distributed in the deepest reservoir in Brazil, located in the Jequitinhonha River. Three areas of the reservoir were sampled quarterly using an echosounder. The deepest fish detected was at 115 m, but most were shallower than 25 m. Diel vertical migration of fish was detected, but not in all the sampling areas and campaigns. Fish distribution was determined by several limnological factors of the reservoir, but mainly by thermal variation in water column. These results can provide guidance for environmental and fisheries managers and ecologists to improve sampling techniques in deep reservoirs since vertical distribution and its relation with habitat use are determinant to propose management measures in similar ecosystems.