AbstractAnalysis of plant pollen can provide valuable insights into the existing spectrum of microorganisms in the environment. When harvesting bee-collected pollen as a dietary supplement for human consumption, timely preservation of the freshly collected pollen is fundamental for product quality. Environmental microorganisms contained in freshly collected pollen can lead to spoilage by degradation of pollen components. In this study, freshly collected bee pollen was sampled at different locations and stored under various storage conditions to examine the hypothesis that storage conditions may have an effect on the composition of microorganisms in pollen samples. The samples were analyzed using 16S and 18S amplicon sequencing and characterized by palynological analysis. Interestingly, the bacterial communities between pollen samples from different locations varied only slightly, whereas for fungal community compositions, this effect was substantially increased. Further, we noticed that fungal communities in pollen are particularly sensitive to storage conditions. The fungal genera proportion Cladosporium and Mycosphaerella decreased, while Zygosaccharomyces and Aspergillus increased during storage. Aspergillus and Zygosaccharomyces fractions increased during storage at 30 °C, which could negatively impact the pollen quality if it is used as a dietary supplement.