In this study, antioxidant and antimicrobial potentials of Usnea intermedia, U. filipendula, and U. fulvoreagens and their stictic and usnic acid contents were investigated. Antioxidant activity and total phenolic contents were evaluated in acetone, ethanol, and methanol extracts of these three species. Antioxidant activity was measured by ABTS [2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid)] method and total phenolic contents were measured by Folin-Ciocalteu method. High-performance liquid chromatography (HPLC) was used for the determination of lichen acids. It can be concluded from stictic and usnic acids contents that the order of solvent efficiency is acetone > ethanol > methanol and acetone > methanol > ethanol, respectively. Broth microdilution method was performed to determine the minimum inhibitory concentration (MIC) of the methanol extracts of three Usnea species. The MIC values of all the extracts ranged from 64 µg/ml to 512 µg/ml for all the bacterial strains that were tested in this study, and all the Fluoro quinolone-resistant Escherichia coli isolates (except for E101) were sensitive to the methanol extracts of the three Usnea species. This paper is the first study to determine the stictic acid content in U. intermedia and U. filipendula. Our findings indicate that these three Usnea species could be used as antioxidant and antimicrobial agents.