Currently, there is limited understanding of microbial diversity and safety considerations associated with the use of natural substances in pelotherapy, especially concerning fungal diversity and presence. ITS sequencing was used to assess the fungal diversity in natural peloids (peat and sapropel) used in pelotherapy in Lithuania. Fungal diversity and dominance varied across different types of peloids, primarily determined by the environment from which they were sourced. Fungi from the genera Aspergillus, Fusarium, and Mucorales were identified in peloids sourced from peatland but were absent in sapropel peloids obtained from lakes. These fungi are ubiquitous in peatland, where they play a role in decomposing organic matter, however, they can also act as potential pathogens. The presence of potentially pathogenic fungi should be carefully considered when using peatland-derived peloids in pelotherapy. However, further research is needed to evaluate the role of fungi in the beneficial properties of peloids with different compositions at various stages of preparation and use, as well as to assess potential risks.