Abstract: During the period from October 2011 to August 2012 a total of 124 patients were clinically examined for mycotic otitis at the outpatient clinic of Otolaryngology Department, Assiut University Hospitals (AUH). The mycological analysis of ear swabs revealed the isolation of 18 fungal species and one variety belonging to 7 genera. Among the 92 mycologically-positive cases, 56 were males (60.9%) and 36 were females (39.1%). The disease was more prevalent among persons between 21-30 years where it was diagnosed in 29 patients matching 31.5% of total positive cases. Aspergillus infection was confirmed in 84.8 % of total cases. Fungi belonging to section Nigri were associated with infection of 39 cases (42.4%). Other species related to sections Flavi and Terrei were involved in 35 and 4 cases (38 % and 4.3% of the total cases respectively). Candida species (C. albicans, C. glabrata, C. krusei and C. parapsilosis) were involved in 16.3% of otomycotic cases. The remaining fungal species comprised Chrysosporium keratinophilum, Cladosporium sphaerospermum, Mucor circinelloides, Phoma epicoccina and Stemphylium vesicarium were rare etiologic agents of otomycosis. Extracellular proteolytic, lipolytic and ureolytic enzymes were produced by 95%, 76.2% and 36.6% of fungal isolates respectively. Aflatoxins were detected in cultures of 72.5% of Aspergillus species belonging to section Flavi. In-vitro sensitivity test showed that Terbinafine, Clotrimazole and Clove oil were the most effective antifungal agents. Key words: Otomycosis, Aspergillus, Candida, protease, lipase, urease, aflatoxins, antifungal agents.