Bottleneck legacies and expansion consequences: the Italian wolf genomic make-up and the potential effect of anthropogenic hybridization on its evolutionary pathway

In the face of accelerating environmental changes driven by human activities, integrating demographic, genetic and genomic perspectives is crucial for the conservation of wild populations. Population bottlenecks not only reduce population size, but also erode genetic diversity, increase inbreeding, and elevate genetic load, impacting long-term adaptability. Conversely, population expansion can introduce founder effects and genetic structuring, requiring targeted conservation measures. Additionally, anthropic activities strongly influence demographic and evolutionary processes. For example, anthropogenic hybridization between wild species and their domestic counterparts can alter evolutionary trajectories through dog introgression, potentially leading to genetic swamping or behavioral modifications, which may be particularly relevant in humandominated landscapes. The Italian wolf (Canis lupus italicus) exemplifies these challenges. This predator recovered from a severe bottleneck after the 1970s, expanded across Italy reaching the Alps, and is increasingly subjected to wolf-dog hybridization with domestic dogs. In this thesis, we applied whole-genome approaches to assess bottleneck legacies and examined recovery dynamics using temporal and spatial population genetics analyses. We estimated the naïve prevalence of wolf-dog hybridization in critical recovery areas and assessed its contribution to neutral genetic variability. Using whole-genome data, we investigated behavioral trait-related genes, introgression, and selection signatures. Finally, we developed a pilot panel of genetic markers to detect admixture and potential behavioral alterations in Italian wolves. We found that, despite demographic recovery, Italian wolves still exhibit relatively low genetic diversity, a small effective population size, signatures of inbreeding, and a non-negligible genetic load. While genetic diversity appears to slightly increase at the neutral level, population substructuring and unidirectional gene flow raise concerns about potential inbreeding depression at the local scale. Wolf-dog hybridization was particularly significant in Central and Southern Apennine recovery areas, and we demonstrated its contribution to both neutral and adaptive variability. Furthermore, we detected recent and ancient dog introgression, which may influence wolf behavior and interactions with anthropic environments. Our findings highlight the necessity of integrating key genetic parameters to design appropriate longterm conservation management plans, rather than relying solely on demographic assessments. As wolf-dog hybridization may influence the Italian wolf genomic make-up and behavior – potentially reducing wariness and increasing human-wildlife conflicts – cost-effective diagnostic panels of markers could serve as a tool for targeted management. Addressing these genetic and behavioral consequences is critical for ensuring the long-term viability and adaptability of the Italian wolf population in a changing and human-dominated environment.