The Chondrostoma genus is widespread in Europe, with numerous endemic species in northern Mediterranean rivers. We reconstructed the phylogenetic relationships of this genus, using the whole cytochrome b sequence and compared the two freshwater fish dispersion hypotheses: (1) dispersion around the Mediterranean Sea during the Lago Mare phase of the Messinian salinity crisis (Bianco’s hypothesis) and (2) an older and more gradual colonization of the Mediterranean rivers (Banarescu’s hypothesis). All phylogenetic analyses identified two levels of divergences, implying two radiation events in the Chondrostoma genus. The first radiation mainly concerned Mediterranean species, whereas the second one includes Danubian and Mesopotamian species. This phylogeographic pattern was already observed for the genus Squalius, which exhibits a similar geographic range distribution in Europe and probably is shared with several other Mediterranean genera, such as Scardinius, Rutilus, and Telestes. Furthermore, assuming a molecular clock of 1% per million years, the first radiation appears consistent with a Messinian dispersion during the Lago Mare, 5.3 million years ago, whereas the second one may correspond to a Mesopotamian dispersion through the Black Sea to the Danube system. According to our results, the Lago Mare theory is strengthened, and a more recent and pre-Pleistocene colonization of the Danube from Mesopotamian freshwater fishes is suggested
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