Staphylococcus aureus (S. aureus) is one of the bacterial species capable of forming multilayered biofilms on implants. Such biofilms formed on implanted medical devices often require the removal of the implant in order to avoid sepsis or, in the worst case, even the death of the patient. To address the problem of unwanted S. aureus biofilm formation, its first step, i.e., adhesion, must be understood and prevented. Thus, the development of adhesion-reducing surface coatings for implant materials is of utmost importance. In this work, we used single-cell force spectroscopy to analyze the adhesion of the biofilm-forming S. aureus strain SA113 on naive and protein-coated silicon surfaces (SiO2). In addition to the wild type, we used the SA113 ΔdltA knockout mutant to further investigate the effect of d-alanylation of lipoteichoic acids of the cell wall. In order to examine how the surface charge affects adhesion, we coated silanized SiO2 surfaces with amphiphilic class II hydrophobins. The naturally occurring hydrophobin HFBI was used as well as the HFBI variant D40Q/D43N, which is less negatively charged at physiological pH due to the exchange of two acidic aspartate residues. These two types of hydrophobin-coated surfaces resemble each other in roughness and wettability but differ only in charge. By measurement of the forces with which each S. aureus strain binds to hydrophobin-coated surfaces, we show that the adhesion of S. aureus at surfaces can be influenced by the charges exposed by the target surfaces. Therefore, in addition to hydrogen bonding, electrostatic interactions between the cell and the hydrophilic surface govern adhesion on these surfaces. Moreover, we found that for both HFBI coatings, the adhesion strength of S. aureus is reduced by nearly a factor of 30 compared to silanized SiO2 surfaces. Therefore, hydrophobin coatings are of great interest for further use in the field of biomedical surface coating.