AbstractOnychomycosis, a nail fungal infection, is normally caused by dermatophytes. However, yeasts and non‐dermatophyte moulds (NDM) are among pathogens that cause nail disease. Regarding, this study aimed to describe the molecular epidemiology of Fusarium onychomycosis in the North of Iran. Two hundred and fifty seven nail samples collected from the patients clinically suspected of onychomycosis were subjected to direct microscopy, calcofluor white staining and culture. Fusarium isolates were identified at a species level through determination of multi‐locus sequences for internal transcribed spacer and translation elongation factor 1 alpha. Based on the findings, Fusarium species were isolated from onychomycosis patients (n = 27). According to a previous partial genes analysis, the species in the recent study belonged to the members of F. fujikuroi species complex (n = 14), Fusarium incarnatum‐equiseti species complex (n = 1) and F. solani species complex (n = 12). In this study, F. proliferatum was the dominant Fusarium species collected from the samples. The correct identification of Fusarium species is essential regarding the increased prevalence of Fusarium onychomycosis and the inherent resistance of these agents to a wide spectrum of antifungals. The obtained results indicated variation in the epidemiology of Fusarium species isolated from onychomycosis. Moreover, the minimum inhibitory concentration (MIC) of luliconazole and lanoconazole was in the range of 0.001–1 μg/ml, with the geometric mean of MICs obtained at 0.0103 and 0.0343 μg/ml against Fusarium species, respectively. These findings can increase researchers' knowledge regarding diversity of species, distribution of onychomycosis and the choice of a proper treatment.