Topography-mediated antibacterial surfaces that inactivate bacteria by physical contact have gained attention in recent years. Contrary to conventional antibacterial coatings, topography-mediated antibacterial surfaces do not suffer from coating instability and possible toxicity problems. In this study, a one-step hard anodization process has been deployed to fabricate a topography-mediated antibacterial aluminum surface. By optimizing anodization parameters, such as the concentration of the electrolyte, current density, and anodization time, desirable features of micronanoscale morphology were achieved. The optimum conditions of anodized aluminum that provided pores of a diameter of 151 ± 37 nm effectively killed 100% of bacteria.