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We applied nested clade phylogeographical analysis (NCPA) to the mitochondrial DNA phy-logeographical data of the barbel Barbus barbusto assess the historical biogeography scenario suggested for this species by a traditional phylogeographical approach. Major previously inferred historical events received support from the NCPA: i) twofold range fragmentation, an ancient one between the central European and Balkan/Anatolian populations and a more recent one ascribed to the survival of the central European lineage in two refugia during the latest glacial, and ii) contiguous range expansion from the Danubian refuge into the more northwestern river basins. Likely due to insufficient genetic variation, the hypothesized dispersion from the more western central European refuge was not detected by the NCPA as was not the hypothesized expansion throughout the Balkans and Anatolia. The NCPA interpretation of the significant pattern within the Danube river basin as reflecting a recurrent gene flow restricted through isolation by distance should be taken with caution. Similar patterns can reflect non-equilibrium conditions, such as population growth, which seems a plausible alternative interpretation given the star-like genealogy of the Danubian population, and its presumably short period of demographic stability. [ABSTRACT FROM AUTHOR]