## 3 Results and Discussion
### Surface Properties  
Since bacterial adhesion and subsequent biofilm formation are influenced by surface properties, such as roughness, topography, and hydrophobicity [63; 64], the three tested materials (SST, PVC, and SIL) were first characterized concerning these parameters.  
The surface topography and roughness of the materials were determined by AFM analysis (Figure 2 and Table 2). SIL exhibited considerably more irregularities and higher roughness values compared to SST and PVC materials (_Ra_\(\mathrm{SIL}\) = 565.0 \(\pm\) 61.2 nm versus _Ra_\(\mathrm{SST}\) = 43.5 \(\pm\) 9.2 nm and _Ra_\(\mathrm{PVC}\) = 31.7 \(\pm\) 12.0 nm). In turn, the average roughness obtained for SST and PVC was similar.  
Surface characterization was complemented by SEM analysis to evaluate surface morphology (Figure 3). SIL displayed more pronounced protuberances compared to SST and PVC, which contributed to the higher roughness values observed in the AFM measurements.  
Some studies have reported that increased surface roughness promotes bacterial adhesion by creating favorable sites for colonization [65,66]. However, other studies have shown that the association between surface roughness and bacterial adhesion depends on the size, shape, and density of the microscale features [67] and the intrinsic bacterial surface properties [66].  
Surface hydrophobicity also plays an important role in bacterial adhesion. The hydrophobicity of the tested materials was determined by measuring contact angles (Table S1 in Supplementary Material) and applying the Van Oss approach [51]. The free energy of interaction (\(\Delta G_{i\pi i}\)) values are presented in Table 2. All surface materials displayed \(\Delta G_{i\pi i}<0\) m\(\cdot\)m\({}^{-2}\), indicating that they are hydrophobic. Among the tested surfaces, SIL  
\begin{table}
\begin{tabular}{c c c} \hline \hline \multirow{2}{*}{**Surface**} & **Roughness** & **Hydrophobicity** \\ \cline{2-3}  & \(R_{a}\) **(nm)** & \(\Delta G_{i\pi i}\) **(m\(\cdot\)m\({}^{-2}\))** \\ \hline SST & 43.5 (\(\pm\)9.2) & \(-\)10.9 \\ PVC & 31.7 (\(\pm\)12.0) & \(-\)12.3 \\ SIL & 565.0 (\(\pm\)61.2) & \(-\)75.0 \\ \hline SST, stainless steel; PVC, polyvinyl chloride; SIL, polyvinyl chloride. & & \\ \end{tabular}
\end{table}
Table 2: Average roughness (\(R_{a}\)) and free energy of interaction (\(\Delta G_{i\pi i}\)) values determined for the tested materials. Roughness results are presented as mean \(\pm\) standard deviation (SD), while the \(\Delta G_{i\pi i}\) values were estimated using Equation (4).  
Figure 3: SEM images of (**a**) SST, (**b**) PVC, and (**c**) SIL (magnification of 500\(\times\)).  
Figure 2: Representative two-dimensional AFM images of (**a**) SST, (**b**) PVC, and (**c**) SIL. Note that the scanning area for (**a**) SST and (**b**) PVC was 20 μm \(\times\) 20 μm, whereas the scanning area for (**c**) SIL had to be considerably larger (100 μm \(\times\) 100 μm) due to the extreme irregularity of this surface. The color bar corresponds to the \(z\)-range of the respective AFM image.  
showed the lowest free energy of interaction value, thus being the more hydrophobic surface. Some studies suggest that hydrophobic surfaces are prone to bacterial adhesion [68].