This study aimed to examine the antibacterial activity of extracts of Amazonian Agaricomycetes macrofungi. The sample comprised 168 specimen collected at the Zoobotanical Park of the Federal University of Acre (Brazil), of which 32 (19%) were isolated. Extracts of the 32 isolates and 28 Agaricomycetes basidiomata were prepared and tested for activity against Staphylococcus aureus, Streptococcus pneumoniae, Escherichia coli, and Klebsiella pneumoniae bacteria, using the disc diffusion method. Of the 32 metabolite extracts of the isolated Basidiomycetes, 11 (34.4%) exhibited antibacterial activity especially against Gram-negative E. coli and K. pneumoniae bacteria, while of the 28 basidiomata extracts, 6 (21.4%) exhibited antibacterial activity mainly against the Gram-positive S. aureus bacteria. This study is the first to report the antibacterial activity of the Agaricomycetes species Coriolopsis caperata, Cyclomyces iodinus, Cymatoderma sp., Favolus tenuiculus, and Tyromyces cf. polyporoides, and contributes as the first investigation about the antibacterial potential of Basidiomycetes grown in Southwest Amazonia.