Besides surface roughness and morphology, the hydrophobicity and surface free energy (SFE) of an implant surface are known to influence bacterial adhesion [42, 43]. Physico-chemical interactions (non-specific) are composed of van der Waals forces, electrostatic interactions, and acid-based interactions, which in turn define the surface free energy of a substratum [44]. The surface free energy can be calculated by contact angle measurement of different liquids with differing hydrophobicities [25] or by measuring the wettability by determining water contact angles [45]. Results from different studies that relate surface free energy and hydrophobicity to microbial adhesion are conflicting [44, 46]. However, it has become apparent that, according to the thermodynamic model of microbial adhesion, hydrophobic materials are preferentially colonized by hydrophobic bacteria and vice versa [39, 44, 47,48,49]. Consequently, the adhesion properties of different bacteria are affected by the hydrophobicity of the bacterial cell surface [11, 44]. Both S. epidermidis and S. sanguinis are known to be rather hydrophobic; therefore, hydrophobic surfaces are preferable [44, 49]. Accordingly, Drake et al. reported that titanium samples with hydrophobic surfaces have higher levels of bacterial colonization of S. sanguinis than titanium samples with hydrophilic surfaces [50]. Surface roughness itself is known to influence hydrophobicity [51], but many studies have also clearly shown that minor variations in surface roughness do not significantly affect hydrophobicity values [12]. In the present study, different specimens with varying hydrophobicity but similar surface roughness were selected to eliminate the influence of surface roughness. To our knowledge, this is the first in vitro study to vary surface roughness and hydrophobicity in well-defined patterns to define the predominant factor for the two single-species biofilms tested. For S. sanguinis, no significant difference could be found with regard to bacterial adhesion between the hydrophobic and hydrophilic surfaces of zirconia and titanium. In contrast, S. epidermidis showed higher initial adhesion on hydrophobic than on hydrophilic surfaces; this finding can be attributed to the hydrophobic properties of S. epidermidis and explained by the thermodynamic model of microbial adhesion.