Since the beginning of the nineteenth century, rich non-marine molluscan faunas have been recognized in the Plio-Pleistocene Upper Valdarno Basin (Tuscany, central Italy). Further research has improved the palaeoecological and biochronological meaning of these faunas, which are often associated with significant Villafranchian vertebrate remains. The Upper Valdarno Basin fill consists of four unconformity-bounded units: Castelnuovo dei Sabbioni, Montevarchi, Fosso Salceto (occurring in the Palazzolo sub-basin) and Torrente Ciuffenna synthems. Rich molluscan assemblages of alluvial and palustrine environment are recorded from the Lower Pleistocene Montevarchi Synthem deposits. Extinct and archaic freshwater gastropods and bivalves characterising the Middle and Upper Villafranchian thermophilous assemblages of intermountain basins of central Italy, such as the Plio-Pleistocene Tiberino Basin, occur. The new record of the extinct genus NeumayriaDe Stefani, 1877 is reported. A humid-temperate climatic phase is recognized in concomitance with the major expansion of these assemblages in the Upper Valdarno Basin. The fluvial deposits of the Torrente Ciuffenna Synthem, Middle Pleistocene in age, yielded poor mixed assemblages composed of terrestrial and freshwater species. The recorded land species, represented by Pupillidae and Vertiginidae associated with a dry exposed environment, point to an arid or semiarid cool climatic phase.