2D geometric morphometrics reveals temporal patterns in the evolution of modern and Pleistocene Arvicola (Rodentia, Cricetidae) in Europe

This study applies a comprehensive two-dimensional (2D) geometric morphometric framework to investigate dental morphological variation in modern and Pleistocene populations of the genus Arvicola.A reference dataset of 390 first lower molars (m/1) representing four extant species (A.amphibius, A. sapidus, A. italicus, and A. persicus) was analysed using Elliptic Fourier Analysis combined with multivariate statistics (PCA, MANOVA, LDA, CDA, Random Forest, and Mahalanobis distances).All modern species of Arvicola exhibit highly significant morphological differences, demonstrating that m/1 outlines shape reliably discriminates species-specific variation.This validated classification framework was applied to fossil assemblages from five well-dated localities spanning the late Middle Pleistocene to Upper Pleistocene (MIS 6 to MIS 3).The MIS 6 assemblage from Lazaret Cave displays a heterogenous and intermediate morphospace distribution, reflecting the presence of archaic lineages attributed to Arvicola ex gr.cantianus/amphibius and Arvicola ex gr.cantianus/sapidus.By MIS 5.5 (Baume Moula-Guercy), Arvicola amphibius-like morphologies dominate, although several specimens retain plesiomorphic traits.In contrast, MIS 3 assemblages (Abauntz Cave, Cova Eirós, and Trou Al'Wesse) exhibit predominantly modern morphologies, with high classification agreement (up to 100%) and clear species-level assignment.A temporal trend emerges in which discrimination accuracy and morphological distinctiveness increase from the late Middle Pleistocene to MIS 3, documenting the evolutionary transition from archaic Arvicola forms to fully differentiated modern species.The results further suggest a complex biogeographic history of Arvicola sapidus, involving multiple Middle and early Late Pleistocene dispersal events into Western Europe.Overall, this study demonstrates that 2D outline-based geometric morphometrics of the m/1 is a robust and informative tool for resolving Arvicola systematics and for reconstructing evolutionary and palaeoenvironmental patterns in the Pleistocene of Europe.