MtDNA of common vole from Grotta della Ferrovia site
The recent developments in palaeoecological reconstruction methods, collagen extraction of small bone samples and ancient DNA analyses led us to test new approaches to enhance the chronological resolution of past climate reconstructions inferred from small mammal assemblages.  Grotta della Ferrovia (Fabriano, Ancona, Italy, 40°25'36''N, 13°0'11''E, 215 m a.s.l.) is a small cave that opens on the southern bank of the Esino river, around 45 km west of Ancona. Discovered in 1966, it is famous among scholars because it is the first locality where the fossil birch mouse was recorded in Italy and because the lithic assemblage found in the cave was related to the Late Epigravettian. Across the small mammal sequence, divided into seven layers (from GDF7 to GDF2), two main phases can be recognised: in the first phase, from GDF7 to GDF5, Microtus arvalis is dominant over Microtus agrestis, in a poorly diversified assemblage. The sequence gradually changes in GDF4 and a second phase can be observed in GDF3 and GDF2, where Apodemus gr. sylvaticus-flavicollis dominates, but combined with a higher number of taxa. From GDF7 to GDF3, few individuals of Sicista cf. subtilis and Alexandromys oeconomus are recorded. Environmental and climatic reconstructions calculated with the Habitat Weighting and the Bioclimatic methods show a dominance of open meadows (GDF7 to GDF4) that were gradually replaced by closed or semi-closed forests (GDF3 and GDF2) in a context of gradually increasing temperature, going from a cold phase to conditions similar to the current ones in the Grotta della Ferrovia area. Direct radiocarbon dates on 18 rodent bones and aDNA analyses on three Microtus arvalis teeth helped us to better understand the significance of the oscillation inferred from the small mammal sequences. A robust series of 9 radiocarbon measurements from GDF7 to GDF5 shows that this part of the sequence accumulated between ~24,600 - 19,600 cal BP (95.4% probability), which includes the final phases of the Last Glacial Maximum. The ancient DNA from Microtus arvalis confirmed the chronological framework and the presence of italic populations in this area during the LGM, similar to the slightly more ancient Grotta del Sambuco on the western side of the central Peninsula. The small mammal radiocarbon dates from GDF4 and above are less consistent with their stratigraphic position, probably related to reworking of the sediment, but suggest the last part of the sequence accumulated during the Late Glacial. Our results show that it is now possible to validate and enhance paleoclimatic and paleoenvironment inferences from small mammal sequence studies with direct radiocarbon dates and ancient DNA analyses, using these integrated methods as a high resolution tool for studying the past.