Design, Synthesis, and Antibiofilm Efficacy of Ag/Se-Doped CuO-Chitosan Nanocomposites as Potent Antifungal Agents Against Candida albicans.

In this study, we report the fabrication, characterization, and antimicrobial evaluation of copper (II) oxide CuO (TS1) and (CuO)-based nanocomposites incorporating chitosan (Ch) (TS2), silver (Ag) (Ch-CuO-Ag; TS3), and selenium (Se) (Ch-CuO-Se; TS4) nanoparticles. X-ray diffraction (XRD) confirmed the monoclinic crystalline structure of CuO with an average crystallite size of ∼12 nm, and successful integration of Ch, Ag, and Se without phase impurities. FTIR analysis revealed strong interactions between CuO and Ch functional groups, evidenced by spectral shifts and peak broadening, suggesting hydrogen bonding and structural modifications. Morphological analysis via electron microscopy showed a transition from aggregated nanosheets to homogeneous nanospheres upon doping with Ag and Se. Composite TS1 (CuO) exhibited superior antimicrobial activity, surpassing the standard drugs imipenem and fluconazole (FLZ) in terms of inhibition zones and minimum inhibitory concentrations (MICs). Sub-MIC concentrations of the composites did not affect the growth of Candida albicans, validating their use in biofilm inhibition assays. Notably, TS1 and TS4 (Ch─CuO─Se) significantly enhanced the efficacy of FLZ, reducing its MIC and MBEC values by up to 8-fold. Biofilm inhibition reached 82.2% with TS1-FLZ synergy. Gene expression analysis revealed marked downregulation of biofilm-associated virulence genes (SAP2, SAP6, PLB1, PLB2, LIP4, and LIP5).