Nanostructured Ti-6Al-4V Reduces Adhesion of Several Bacterial Species: An In Vitro Study

Biomaterial-associated infections (BAIs) and insufficient early cellular response remain critical challenges for orthopedic implants. We introduce a comprehensive study that bridges current knowledge gaps by examining an early-stage antimicrobial effect on the clinically relevant alloy Ti-6Al-4V. It combines pathogenic strains with parallel osteoblast assays and tilted-view SEM analysis to obtain a qualitative understanding of the adhesion mechanisms. Detailed physicochemical characterization revealed a progressive increase in nanoscale roughness and oxide layer thickness, accompanied by selective Al/V depletion and pronounced hydrophilization. To evaluate biological responses, we used standardized in vitro models with Staphylococcus aureus, Staphylococcus epidermidis, and Escherichia coli. Bacterial adhesion was quantified by SYTO9 staining, a GFP-expressing strain as a viability control, and SEM imaging. Nanostructured (R _q ≤ 40 nm) surfaces significantly reduced early bacterial attachment compared to polished nanoflat controls. In parallel, osteoblast-like SaOs-2 cells showed stable adhesion and spreading, confirmed by phalloidin/DAPI staining and LDH cytotoxicity assay. Together, these results demonstrate that NaOH-etched Ti-6Al-4V surfaces can impair early microbial adhesion based on physical action and preserve osteoblast compatibility. By integrating advanced materials characterization with microbiological and cell biological assays, we provide a framework for topography-driven surface design toward infection-resistant orthopedic implants that support favorable early osteoblast-surface interactions.