Background: A growing incidence of invasive fungal infections, especially among immunocompromised patients, has given increased significance to microbiological diagnostics of yeast-like fungi. More accurate and faster fungi identification methods that can compete with classical methods are being searched for. In this paper, classical microbiological methods are compared to MALDI–TOF MS (matrix-assisted laser desorption/ionization time-of-flight mass spectrometry). Methods: The diagnostic material was collected from buccal mucosa from 98 adults, including 69 with HIV. Only positive cultures were included in the study. Results: Matching results were obtained in 45 samples, and there were nonmatching results in 35 samples, with the majority of these in the study group, constituting 50% of identifications within this group. A particularly common mistake resulting from the use of classical methods is the false identification of C. dubliniensis as C. albicans. Additionally, C. tropicalis proves to be difficult to identify. Conclusions: Our results and literature data suggest that MALDI–TOF MS should be considered an effective alternative to classical methods in terms of fungi identification, especially among HIV-positive patients, due to the different morphology of fungal colonies.

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