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Abstract Aim Patterns of genetic variation within freshwater fish populations may reflect the historical impact of climate change on either sea‐level or environmental conditions. Past sea‐level changes enlarged palaeodrainages and so connected currently isolated rivers, whereas changes in environmental conditions reduced forest cover and may have constrained the movement of fish specialized to this habitat. We assayed genetic variation in Hollandichthys multifasciatus , a freshwater fish endemic to the Atlantic Forest of coastal Brazil, to test the relative importance of these factors in shaping current patterns of genetic divergence. Location River drainages along the south‐eastern Brazilian coast. Methods A GIS was used to reconstruct palaeodrainages during the Last Glacial Maximum ( LGM ). Niche modelling was used to infer areas of stability for the southern Atlantic Forest sensu stricto (present and LGM ). The contribution of river connections inside or outside areas of stability was evaluated using a calibrated phylogeny, analyses of molecular variance, and Bayesian skyline plots from two mt DNA loci. Results Analyses of 182 individuals from 26 populations and 12 palaeodrainages indicated that structure associated with palaeodrainages explains 75% of the genetic variation among populations, with estimated divergence times occurring within the Pleistocene. The variation explained by palaeodrainages and estimated population sizes was unrelated to the ecological stability of the region. Main conclusions This study demonstrates the importance of Pleistocene palaeodrainages in structuring genetic divergence patterns. The analyses suggest that past connections due to sea‐level retreat played a significant role in the diversification of the ichthyofauna along the Brazilian coastal drainages. Moreover, the lack of a signature of habitat stability in structuring genetic variation suggests that refugia may be less important in structuring genetic diversity for freshwater species than for terrestrial species. In addition, our work highlights the utility of a GIS ‐based approach to recover past connections among coastal basins. Understanding these connections is crucial for studying diversification of riverine organisms and for identifying areas of conservation priority.
Thomaz et al. (Sun,) studied this question.