Currently, a problem related to public health is resistance to antibiotics because bacteria have been identified to be resistant. It has been reported that bacteria have developed resistance mechanisms to evade the effect of drugs, especially antibiotics. A resistance mechanism to antibiotics is biofilm formation. The biofilms are microbial communities embedded in an extracellular polymeric matrix and are ubiquitous in the microbial world. In recent years, there has been a special interest in studying new antimicrobial strategies to solve the great problem of resistance to antibiotics; thus, the use of essential oils could be an alternative to fight infections caused by biofilm forming bacteria. It has also been reported that essential oils have antiviral and antibacterial properties. In the present work, the effect of Thymus vulgaris on growth and biofilm formation of uropathogenic Escherichia coli was studied. This study demonstrated the strong effect of T. vulgaris essential oil on the growth and biofilm formation of uropathogenic E. coli.