The purpose of this study was to examine the impact of hydrophobicity, surface chemistry, and topography on bacterial and microalgal adhesion. To this end, the effects of surface plasma treatments (Argon and Tetrafluorocarbon) of polycarbonate substrates on bioadhesion were investigated in vitro. Surfaces were characterized by goniometry, atomic force microscopy (AFM), and X-ray photoelectron spectroscopy (XPS). Two marine bacterial strains, a hydrophobic Pseudoalteromonas sp. 5M6 and a hydrophilic Paracoccus sp. 4M6 and two microalgae (Cylindrotheca closterium and Porphyridium purpureum) involved in marine biofouling were selected. Their adhesion on the substrates was observed and quantified using confocal laser scanning microscopy-microfluidic flow-cells. It was demonstrated that the combination of three parameters, namely surface energy, fluorination, and nanotopography, significantly decreases the adhesion of three microorganisms out of four (Pseudoalteromonas sp. 5M6, Cylindrotheca Closterium, and Porphyridium purpureum) whereas one parameter on its own is insufficient.