Morphological, vocal and genetic studies have shown that the Madeira River and its right bank tributaries delimit populations of primates and birds. We sequenced the cytochrome b gene (approx. 950 bp) for individuals of three suboscine passerine bird species, Glyphorynchus spirurus (Furnariidae), Willisornis poecilinotus (Thamnophilidae) and Schiffornis turdina (Tityridae), on opposite banks of the Madeira River and two of its right-bank tributaries, the Aripuana and Jiparana rivers. Phylogenetic hypotheses (parsimony, maximum likelihood and Bayesian analysis) revealed clades that have over 3.1% genetic differentiation on opposite banks of the Madeira River for G. spirurus, W. poecilinotus and S. turdina, suggesting that this river restricts gene flow among populations of these three species. The Jiparana and Aripuana rivers apparently separate distinct populations of G. spirurus, the smallest species we examined, but not those of the other two heavier bodied species, W. poecilinotus and S. turdina. In G. spirurus four clades with high levels of genetic differentiation (3.2–5.5%) were found to be delimited by the three rivers evaluated, whereas in W. poecilinotus and S. turdina no genetic structure across the Jiparana and Aripuana rivers was detected. In general, birds that are known to show population structure across the Madeira tributaries (Glyphorynchus spirurus, Hemitriccus minor, Hypocnemis rondoni, Herpsilochmus stotzi, and Hylophylax naevius) have body masses smaller than those of both Willisornis poecilinotus and Schiffornis turdina, but some exceptions are discussed. Future studies controlling for several variables are necessary to determine the extent to which body mass is a useful predictor of genetic population structure in understory suboscine passerines.