Variation in auditory tuning across populations of the Bufotes viridis complex

Introduction Acoustic communication in anurans relies on the evolution of signal production and auditory sensitivity. While variation in advertisement calls has been extensively investigated, less is known about variation in auditory tuning and its evolutionary drivers. Methods We investigated auditory system variation in Bufotes viridis complex by comparing 2 pairs of groups significantly different in size: a) diploid and tetraploid Central Asian sympatric populations and b) European diploid populations from mainland Italy with diploids from the island of Sardinia. Using electrophysiological recordings, we estimated the best excitatory frequency (BEF) and thresholds of the basilar papilla (BP). Results While auditory thresholds did not vary significantly among groups or with body size, BP tuning was negatively associated with body size and differed among groups. Group differences were similar in the two sexes, but statistically significant only in females. BEFs differed between sympatric diploid and tetraploid, supporting a potential role for ploidy consistent with a reproductive character displacement scenario. At the same time, similarly strong differences were observed between allopatric mainland and Sardinian samples, suggesting that divergence in auditory tuning can also arise independently of ploidy and sympatry. Conclusion Overall, our results indicate that auditory tuning in green toads is shaped by multiple, potentially interacting processes, including body size, ploidy, and population-specific evolutionary histories. These findings highlight the need for integrative approaches to understand the coevolution of signals and receivers in complex evolutionary systems.