Myrtus communis L. İs a shrub widespread in the Mediterranean area. The interest in this species is growing, mainly due to its pharmacological and aromatic properties. The overexploitation of wild populations induced increasing degradation of plant cover with serious risk of loss of genetic diversity. This research explored the morphological, chemical, and genetic diversity of wild myrtle populations in Sicily, with the aim to provide a first characterization of a core collection of 36 accessions from 7 localities for future domestication programs. Amplified fragment length polymorphism fingerprinting generated 152 polymorphic fragments. STRUCTURE analysis identified three genetic clusters (A, B, and C) corresponding to specific geographical origin. Analysis of molecular variance estimated a quite high overall fixation index (FST = 0.332). Misilmeri and Ispica were the more divergent populations (FST = 0.502), while M. Pellegrino and Scopello revealed the lowest FST (0.153). The relationships between genetic, morphological, and biometric data were investigated. Significant correlation between genetic clusters and bush shape/plant growth behavior was found (P < 0.005). Moreover, morphological traits such as leaf, fruit, and seed size were significantly correlated to Clusters B and C. Leaves' secondary metabolite profiles were evaluated based on antioxidant activity and total tannin and phenol concentrations. High antioxidant activity differences were recorded using DPPH (21.4-35.5 mmol Trolox/100 g DW) and ABTS (24.2-39.5 mmol Trolox/100 g DW) methods. A low variability was observed among populations regarding phenol (2466-3800 mg catechin equivalents/100 g DW) and total tannin contents (93.9-262.3 mg catechin equivalents/100 g DW). Results indicated that multiple approaches based on genetic, morphological, and chemical traits might allow the characterization of natural myrtle diversity.