Metabarcoding technology through high-throughput sequencing can characterize the species composition of bulk samples, including even degraded DNA. In the present study, we conducted DNA metabarcoding (the nuclear ITS2 region) for indoor fungal samples originating from a nursery school with a suspected mould problem and from a university building. Good-quality sequences were obtained, and the results showed that DNA metabarcoding gives a high resolution in fungal identification and allows a more complete discovery of taxonomic diversity when compared to culture-dependent methods. The numbers of fungal classes, orders, families, and genera per sample varied greatly, between 11–16, 14–33, 15–46, and 26–76, respectively. Comparable ranges of Shannon’s diversity indices were 0.70–1.74, 0.80–2.46, 0.88–2.89, and 0.78–3.42, respectively. When comparing fungal diversities between the nursery school and the university building, mean diversities were closely similar. Proportions of sequences successfully assigned at class, order, family, genus, and species levels equaled 100.0, 99.7, 99.3, 94.7, and 27.9 %. Thus, the method used is highly effective until the genus level. Although differences were observed in the fungal composition of the studied buildings, it is difficult to draw definite conclusions of the causes of supposedly poor indoor air quality (yet, without medical evidence) in the nursery school.