Mechanistic insights into the antifungal and antibiofilm activity of <i>Pyracantha koidzumii</i> -derived copper nanoparticles against non- <i>albicans Candida</i> isolates

Abstract Non- albicans Candida species pose an increasing challenge in antifungal therapy due to their biofilm-associated tolerance and multidrug resistance. This study investigates the mechanistic basis of the antifungal and antibiofilm activity of Pyracantha koidzumii –derived copper nanoparticles (CuNPs) against clinical isolates of C. glabrata, C. tropicalis , and C. krusei . Physicochemical characterization confirmed the successful synthesis of CuNPs, revealing quasi-spherical nanoparticles (∼2.7 ± 0.59 nm), a negatively charged surface (−28.4 ± 1.2 mV), and a high particle concentration (∼8.7 × 10 11 NPs ml −1 ). CuNPs exhibited species-dependent antifungal activity, as reflected by the minimum fungistatic (MFuC) and fungicidal (MFC) concentrations. MFuC values were 1.9, 31.2, and 15.6 μ g ml −1 , while the corresponding MFC values were 3.8, 62.4, and 31.2 μ g ml −1 for C. krusei, C. tropicalis , and C. glabrata , respectively. Biofilm assays using crystal violet staining demonstrated a marked reduction in biomass formation across all isolates, accompanied by decreased exopolysaccharide production, and increased susceptibility to oxidative stress. Additionally, calcofluor white-propidium iodide staining indicated cell wall disruption at MFuC levels, while exposure to CuNPs further decreased cell viability after 24 h, yielding an IC 50 of 7.81 μ g ml −1 in HepG2 cells. These findings indicate that, despite the relatively narrow in vitro safety margin suggested by the moderate cytotoxicity of CuNPs, they remain promising candidates as alternative antifungal agents.