Aim: Factors that isolate populations and reduce gene flow are considered key drivers of speciation and possibly diversification. Here we analyze the diversification rates of nearly 80% of the actinopterygian fish families in relation to biological traits and habitat factors associated with isolation and fragmentation levels. Location: Global Methods: Net diversification rate for each family was estimated using the method-of-moments estimator for stem-group ages. Phylogenetic generalized least-squares analysis (PGLS), controlling for the non-independence between clades due to phylogeny, was applied with diversification rate as the response variable to test the effects of mean body size, proportions of strictly freshwater, reef-associated and migratory species and including the median latitudinal distribution and range of each family. Results: After accounting for the phylogenetic relatedness of families and for their latitudinal distribution, we found strong support in agreement with our isolation and fragmentation hypotheses: predominance of freshwater dependence, reef-association, small body size or non-migratory behavior in families is related to more rapid rates of diversification. We also found a highly significant and positive effect of latitudinal range and no clear effect of median latitude. Main conclusions: This analysis suggests that factors related to the physical fragmentation of habitats and to lower dispersal ability of species have played an important role in the diversification processes of the most diverse group of vertebrates.