Application of efficient antimicrobial surfaces has been estimated to decrease both, the healthcare-associated infections and the spread of antibiotic-resistant bacteria. In this paper, we prepared ZnOand ZnO/Ag nanoparticle covered surfaces and evaluated their antimicrobial efficacy towards a Gram-negative bacterial model (Escherichia coli), a Gram-positive bacterial model (Staphylococcus aureus) anda fungal model (Candida albicans) in the dark and under UVA illumination. The surfaces were preparedby spin coating aliquots of ZnO and ZnO/Ag nanoparticle suspensions onto glass substrates. Surfaces contained 2 or 20 g Zn/cm2 and 0–0.02 g Ag/cm2. No significant antimicrobial activity of the surfaces,except of those with the highest Ag or Zn content was observed in the dark. On the other hand, UVAilluminated surfaces containing 20 g Zn/cm2 and 2 g Zn plus 0.02 g Ag/cm2caused >3 log decreasein the viable counts of E. coli and S. aureus in 30 min. As proven by brilliant blue FCF dye degradationand elemental analysis of the surfaces, this remarkable antimicrobial activity was a combined result of photocatalytic effect and release of Zn and Ag ions from surfaces. Surfaces retained significant antibacterial and photocatalytic properties after several usage cycles. Compared to bacteria, yeast C. albicans was significantly less sensitive to the prepared surfaces and only about 1 log reduction of viable count was observed after 60 min UVA illumination. In conclusion, the developed ZnO/Ag surfaces exhibit not only high antibacterial activity but also some antifungal activity.