Evolutionary relationships and genetic structure in Mediterranean Hypericum sect. Adenosepalum with focus on H. scruglii

• Molecular phylogenetics and networks clarify Mediterranean Hypericum relationships • Nuclear and plastid markers reveal significant evolutionary incongruence • Hypericum scruglii shows unique plastid haplotypes confined to Sardinia • Analyses indicate stronger plastid than nuclear differentiation among populations • Findings support recent diversification and conservation relevance of insular taxa Understanding the evolutionary relationships and genetic structure within Mediterranean plant lineages is essential for both systematic and conservation. The genus Hypericum (Hypericaceae) presents an ideal model system due to its ecological versatility, pharmacological relevance, and remarkable diversity across biodiversity hotspots. Yet, the placement of Mediterranean endemics remains poorly resolved. In this study, we investigated phylogeographic patterns and genetic differentiation among three species of section Adenosepalum : the Sardinian endemic H. scruglii , and the more widespread H. tomentosum and H. pubescens . A multilocus dataset comprising nuclear ribosomal cistron (ITS1, 5.8S and ITS2) and two plastid intergenic spacers [ ycf 6- psb M and trn S (GCU) – trn G (UCC) ] was generated from 71 individuals across 17 populations spanning Italy, Spain, France, and Malta. Phylogenetic reconstructions and phylogeographical network analyses were combined to elucidate evolutionary relationships. Congruence testing revealed significant topological incongruence between nuclear and plastid datasets, suggesting complex evolutionary histories involving incomplete lineage sorting and potential historical gene flow. A combination of ribotype and haplotype network analyses revealed distinct geographic clustering, with H. scruglii exhibiting three private plastid haplotypes restricted to Sardinia, while H. pubescens and H. tomentosum showed broader but non-overlapping ribotype sets. Population-level analysis confirmed higher genetic differentiation in plastid markers ( Fst > 0.6) than in nuclear markers ( Fst < 0.4), supporting a scenario of recent Mediterranean diversification with restricted gene flow. Overall, our results provide robust molecular evidence for the taxonomic distinctiveness of H. scruglii and contribute to a refined phylogeographic framework for Mediterranean Hypericum section Adenosepalum , with direct implications for conservation prioritization of insular endemics.