The adhesion strength of a bacterial strain on a substrate influences colonization and biofilm development, so the biomolecular analysis of this interaction is a step that allows insights into the development of antifouling surfaces. As peri‐implantitis is the main cause of failure of implant‐supported oral rehabilitations and the dental literature presents gaps in the atomic bacteria/surface interaction, this study aimed to correlate the qualitative variation of roughness, wettability, chemical composition, and electrical potential of Ti‐6Al‐4V and Ti‐35Nb‐7Zr‐5Ta (TNZT) disks obtained by machining (M) and additive manufacturing (AM) on the colonization and adhesion strength of S. aureus quantified by atomic force microscopy (AFM). The samples were evaluated for roughness, electrical potential, and S. aureus colonization and adhesion strength by specific methods in the AFM with subsequent analysis in the NanoScope software analysis, wettability by sessile drop method, and chemical composition by energy dispersive x‐ray spectroscopy (EDX). Qualitative data were correlated with bacterial adhesion strength. The greater adhesion strength of S. aureus was observed in descending order for TNZT AM, TNZT M, Ti‐6Al‐4V AM, and Ti‐6Al‐4V M. This experimental in vitro study allowed us to conclude that for the evaluated groups, the strength adhesion of S. aureus showed a linear relationship with roughness, and nonlinear for wettability, electrical potential, and S. aureus colonization on the surfaces evaluated. As for the two variation factors, type of alloy and manufacturing method, those that promoted the lowest bacterial adhesion strength were Ti‐6Al‐4V and M, possibly attributed to the synergistic modification of the evaluated surface properties. Thus, this study suggests S. aureus preferences for rough, hydrophilic surfaces with a greater electrical potential difference.