The orchid collection of the ELTE Botanical Garden, Budapest, Hungary was monitored for airborne fungi using viable and non-viable air-sampling methods (Hirst-type and a 3-stage Andersen sampler) with three different culture media. A new culture method was also applied to identify fungal spores from Hirst-type samples. The aim of this study was to determine the diversity, human-and phytopathological potential of the air spora. To find out sources of airborne fungi, samples were collected from the air in an adjacent greenhouse and outdoors, and from necrotic plants. A total of 58 genera were found in the air samples. Cladosporium and Penicillium spp. were common members of the airborne biota. A high proportion (27.5%) of identified genera may be presented as a member of microbial consortium associated with the orchids. Airborne fungi potentially pathogenic to humans were also detected. One species, Zygosporium masonii, was new to Hungary. Statistical analysis indicated that conditions of sampling had significant effects. The principal component analysis elucidated the three principal components representing 75.34% of the total variance; the clusters of variables were related to the three types of culture media. Relative abundance of small-sized spores was high, presumably because of the fungal species composition and accelerated sedimentation of large spores in still air. Apparently, in the studied orchid greenhouse, a specific mycobiota developed due to the climate and hosts (Orchideaceae) grown there.