The higher education sector represents a unique environment and it acts as a work environment, a learning environment for students, and frequently, also a home environment. The aim of this study was to determine the microbial contamination (SARS-CoV-2, fungi, and bacteria) in Higher Education Facilities (HEI) by using active and passive sampling methods and combining culture-based methods with molecular tools targeting Aspergillus section Fumigati. In addition, the resistance to azole profile was also assessed. Surface samples showed a range of total bacterial contamination between 1 × 10³ to 3.1 × 10⁶ CFU·m⁻², while Gram-negative bacteria ranged from 0 to 1.9 × 10⁴ CFU·m⁻². Fungal contamination ranged from 2 × 10³ to 1.8 × 10⁵ CFU·m⁻² on MEA, and from 5 × 10³ to 1.7 × 10⁵ CFU·m⁻² on DG18. The most prevalent species found on both media was Cladosporium sp. (47.36% MEA; 32.33% DG18). Aspergillus genera was observed on MEA (3.21%) and DG18 (14.66%), but not in the supplemented media used for the azole screening. Aspergillus section Fumigati was detected in 2 air samples (2.22%, 2 out of 90 samples) by qPCR. When testing for SARS-CoV-2 all results were negative. The present study showed that although cleaning and disinfection procedures are done regularly due to the COVID-19 pandemic, being effective in eliminating SARS-CoV-2, surfaces were often contaminated with microorganisms other than SARS-CoV-2. This can be a result of increasing resistance to biocides, and to the wide range of environmental factors that can contribute to the dissemination of microbial contamination indoors.

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