Abstract The essential oil isolated from plantlets of Thymus mastichina L. ssp. mastichina cultured in vitro was analyzed by GC and GC/MS. The oil was characterized by the richness of 1,8-cineole (55.5%) and linalool (24.5%). These results are in accordance with the chemical composition previously reported for this taxon. The oil showed antifungal activity against eight pathogenic fungi of die genus Fusarium with MICs and MFCs ranging from 1500 to 2100 Î¼g/mL and from 2.0 to 2.4 mg/mL, respectively.