Display omitted •Exploration of a new sterilization method.•Reduced bacterial adhesion by synergistic structural and physical washing effects.•Fabrication of the surface structure by femtosecond laser.•LIPSS surface and ultrasonic washing greatly decreased S. aureus adhesion. Staphylococcus aureus infection is common in the clinical environment. It has been shown that the presence of micro/nano structures on material surfaces promote bacterial adhesion resistance. Herein, we assessed the S. aureus adhesion properties on laser micro/nano structured stainless-steel (316 L) surfaces after mechanical rotation and ultrasonic washing. The interaction force between S. aureus and structured surfaces was evaluated. A high concentration S. aureus solution was used to evaluate the bacterial sterilization efficiency after film formation on the stainless-steel surface. After 24 h of incubation, S. aureus films were formed on material surfaces. The comparison of static washing, surface mechanical rotation, and ultrasonic washing showed a decrease of S. aureus adhesion on the polished and laser induced periodic surface structures. However, S. aureus adhesion on the micro/nanoparticle surface after mechanical rotation washing did not display any obvious change compared to the polished one. Additionally, specimens after ultrasonic cleaning showed clear antibacterial adhesion than mechanical rotation. After the ultrasonic sterilization process, the laser induced periodic laser surface sample showed optimal bacterial adhesion inhibition. Finally, in vitro tests showed that the biocompatibility of the laser-induced structured surface did not change significantly from the polished surface one.