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Macrofungi are organisms responsible for the decomposition of all the dead elements in a forest, they recycle the matter and without them the ecosystem would maintain many organic remains without optimal use of it. In Panama, macrofungi are very little studied organisms, for the province of Colón the studies are scarce. That is why the objective of this research was to characterize the macrofungi in a fragment of secondary forest in the Guanche community. The developed methodology allowed sampling (collections) to be carried out in a transect with a length of 1000 m and 10 m of road edge. The macrofungi were identified with specialized literature and were deposited in the Herbarium of the University of Panama (PMA). As a result, 53 specimens were obtained (6 species from the Ascomycota Division and 47 species from the Basidiomycota Division). The Basidiomycota Division represented 88.2%, being the group with the greatest presence in the forest while the Ascomycota Division represented 9.8% of the species. Substrate preference was diverse, including wood (69.2%), leaf litter (25%), manure (4%), and insects (1%). This study provides a valuable contribution to the taxonomic and ecological understanding of fungi in Panama, shedding light on their role in forest ecosystems and providing relevant information for conservation and management strategies within the province of Colón.