The Gram-negative Pseudomonas aeruginosa bacterial pathogen is reputed for its resistance to multiple antibiotics, and this property is strongly associated with the development of biofilms. Bacterial biofilms form by aggregation of microorganisms on a solid surface and secretion of an extracellular polysaccharide substances that acts as a physical protection barrier for the encased bacteria. In addition, the P. aeruginosa quorum-sensing system contributes to antibiotic resistance by regulating the expression of several virulence factors, including exotoxin A, elastase, pyoverdin and rhamnolipid. The organosulfur compound allicin, derived from garlic, has been shown to inhibit both surface-adherence of bacteria and production of virulence factors. In this study, the effects of allicin on P. aeruginosa biofilm formation and the production of quorum-sensing controlled virulence factors were investigated. The results demonstrated that allicin could inhibit early bacterial adhesion, reduce EPS secretion, and down-regulate virulence factors’ production. Collectively, these findings suggest the potential of allicin as a therapeutic agent for controlling P. aeruginosa biofilm.