Genetic structure and GWAS in an international pomegranate collection including Sardinian germplasm

Pomegranate (Punica granatum L.) is a strategic crop for Mediterranean and Mediterranean-like environments. We assembled an integrated panel of 163 accessions (109 international; 54 Sardinian) resequenced at ~20×, yielding >20,000 high-quality SNPs. Population-structure analyses (Ward, DAPC, network) resolved 10-11 clusters. The Sardinian set included a low-diversity cluster comprising the majority of Sardinian accessions, many of which formed clone-like pairs. Inter-cluster divergence was moderate (AMOVA FST=0.150). Sliding-window scans (1 Mb) identified highly differentiated regions (FST>0.50) on Chr1, Chr2 and Chr6. Mean values for fruit colour, aril colour and seed hardness differed significantly among clusters, with seed hardness being the most discriminant trait. As a proof of concept, a preliminary multilocus GWAS for seed hardness and taste (BLINK; FarmCPU), correcting for structure, revealed multiple loci, including a robust cross-method signal for taste on chromosome 7 and additional model-dependent signals for seed hardness (Chr1, Chr2, Chr5, and Chr7) and taste (Chr3 and Chr6). A representative subset has been established as a common garden in Sardinia for long-term validation and deployment. These results illustrate how integrating regional landraces with international germplasm can support conservation and guide breeding for fruit-quality targets in Mediterranean contexts.