Scedosporium apiospermum is an emerging pathogen colonizing the airways of patients with cystic fibrosis and causing severe infections in immunocompromised hosts. In order to improve our knowledge on the pathogenic mechanisms of this fungus, we investigated the production of siderophores. Cultivation on CAS medium and specific assays for different classes of siderophores suggested the secretion of hydroxamates. A maximal production was obtained by cultivation of the fungus at alkaline pH in an iron-restricted liquid culture medium. Siderophores were then extracted from the culture filtrate by liquid/liquid extraction, and separated by reverse phase high performance liquid chromatography. Two siderophores, dimerumic acid and N α-methyl coprogen B, were identified by electrospray ionization-mass spectrometry and MS–MS fragmentation. Finally, comparison of various strains suggested a higher production of N α-methyl coprogen B by clinical isolates of respiratory origin. Studies are initiated in order to determine the potential usefulness of these siderophores as diagnostic markers of scedosporiosis.