The phylogeographic structure of 15 genera of Amazonian marsupials and rodents is summarized based on comparative sequence of the mitochondrial cytochrome b gene. The data are limited in geographical coverage, with samples widely scattered throughout Amazonia from the base of the Andes in Peru to the Guianan coast and eastern Brazil. We use this approach to define species boundaries, based minimally on the principle of reciprocal monophyly, in conjunction with morphological or other genetic discontinuities. The taxa so defined are older than previously appreciated, with many lineages dating from 1 to more than 3 Myr, and thus apparently predating the early Pleistocene. We relate patterns of concordant geographical shifts with underlying tectonic history and to current positions of major rivers. Finally, we provide comments on the utility of these data and patterns to conservation, articulating a need to incorporate phylogeographic information as part of the rationale in establishing conservation priorities at the organismal and geographical area levels.