We investigated the phylogeography of the salt water rotifer Brachionus plicatilis, a cyclical parthenogen with passive dispersal mechanisms, using resting eggs recovered from saline lake sediments. Individual resting eggs were obtained from a large selection of lakes which were representative of five endorheic basins and the chain of coastal ponds in the Iberian Peninsula. The novel use of resting eggs allows the integration of seasonal and annual variations as well as the impact of stochastic effects such as drift and local extinction. A 653 bp fragment of the mitochondrial cytochrome oxidase subunit I (COI) gene was sequenced from 98 eggs. Our results revealed a deep phylogeographical structure in this species, with a division into two main lineages with distinct geographical distributions, which probably diverged at the beginning of the Pleistocene period. Most of the mitochondrial DNA haplotypes were restricted to single lakes. Nested clade analysis supported Early Pleistocene fragmentation of populations, low gene flow and some long–distance colonization. These conclusions contrast strongly with previous ideas on rotifer biogeography and this pattern is consistent with a recolonization of the Iberian Peninsula from two glacial refugia. The results provide new insights into the processes responsible for the genetic diversification of passive dispersers, a life–history trait typical of zooplanktonic biotas.