Anti-biofilm Activity of CuO/ZnO Nanoparticles Biosynthesized by Psychrotrophic Bacteria

Abstract

Bacterial biofilms are a major contributor to persistent infections because biofilm-associated cells exhibit increased tolerance to antimicrobial agents. This study aimed to investigate the potential of cold-tolerant bacteria isolated from the Zagros highlands for the extracellular biosynthesis of copper oxide (CuO) and zinc oxide (ZnO) nanoparticles and to evaluate their antibacterial and antibiofilm activities against Staphylococcus epidermidis. CuO and ZnO nanoparticles were synthesized using cell-free supernatants obtained from psychrotrophic bacteria and subsequently characterized using UV–Vis spectroscopy, dynamic light scattering (DLS) and X-ray diffraction (XRD). DLS analysis revealed mean hydrodynamic diameters of 63.21 nm for CuO and 59.16 nm for ZnO, with corresponding polydispersity index (PDI) values of 0.271 and 0.190, respectively. XRD analysis confirmed the crystalline nature of both nanoparticles, with estimated crystallite sizes of 63.21 and 59.19 nm for CuO and ZnO, respectively. Their antimicrobial and biofilm-inhibitory activities were evaluated using crystal violet staining and colony-forming unit assays. ZnO exhibited stronger concentration-dependent activity and completely inhibited S. epidermidis biofilm formation at 250 μg/mL, whereas CuO required a concentration of 500 μg/mL. Both nanoparticles also reduced the number of viable biofilm-associated cells as their concentrations increased. Overall, psychrotrophic bacteria-mediated biosynthesis yielded CuO and ZnO nanoparticles with promising antimicrobial and antibiofilm properties, with ZnO demonstrating greater activity against S. epidermidis biofilms. These findings support their potential application as eco-friendly agents for biofilm control.