Morphological and genetic differentiation of Bufo toads: two cryptic species in Western Europe (Anura, Bufonidae)

The Common toad Bufo bufo sensu lato is a widespread, morphologically conserved taxon. Recent studies have uncovered deep genetic differentiation between population groups, highlighting the need to revise the current taxonomy of the group and recognize additional species. Here we investigate patterns of variation in molecular (a mitochondrial DNA restriction enzyme assay and sequence data for two nuclear DNA fragments totalling 979 bp) and 17 morphological variables in Northern France where two of these groups meet (B. bufo sensu stricto and B. spinosus), in order to delineate their contact zone and uncover characters that would allow discrimination of the two taxa. Mitochondrial DNA data show an abrupt transition from areas where B. bufo is present to those inhabited by B. spinosus, with a narrow area of overlap east of the city of Caen. Morphometric characters, particularly those related to the positioning of the parotoid glands and metatarsal tubercle shape and size, proved useful in discriminating between species (AUC ≥ 0.97, kappa ≥ 0.79). We then used the differentiating character states to allocate over 300 museum specimens from Western Europe to either species with consistent results, including comparable values of AUC and kappa of the identification models, indicating that models could successfully be applied across datasets. We summarize available evidence relevant to the delineation of the distribution of B. bufo and B. spinosus in France and discuss the characters differentiating both species in an evolutionary context. In view of the observed morphological and genetic differentiation and the absence of unequivocal evidence for widespread hybridization we support the view that B. bufo and B. spinosus are best considered different species. Finally, we propose that ‘parotoids in parallel position’ and a thin and smooth skin are derived character states for B. bufo over the northern part of its range.