Antifungal mechanisms of Trichoderma-derived zinc oxide nanoparticles against Botrytis cinerea: Evidence from ultrastructural and biochemical analyses

Grey mold caused by Botrytis cinerea poses a significant threat to Hydrangea cultivation worldwide and requires the development of sustainable and effective control measures. This study explored the potential of biologically synthesized Zinc Oxide nanoparticles (T-ZnO NPs) as an eco-friendly alternative to conventional fungicides and evaluated their antifungal efficacy along with plant health impact. The ZnO NPs were synthesized using Trichoderma spp. and characterized using multiple analytical techniques. The DLS and XRD analysis revealed a uniform size distribution (20–40 nm with spherical morphology). Furthermore, SEM imaging showed agglomerated particles, while EDX confirmed Zn and O as the primary elements. FTIR spectroscopy indicated the presence of protein capping agents and ZN-O bonds. In vitro studies demonstrated significant antifungal activity, 72% growth inhibition against Botrytis cinerea . Greenhouse trials showed that dose-dependent effects with 0.5 mg/mL T-ZnO NPs treatment resulted in remarkable improvement in plant growth parameters, including 23.7% increase in leaf number, 31.1% in leaf diameter, 30.1% in shoot length, 39.9% in root length, as compared to the control. The disease severity index was decreased by 46.3% while the CO 2 assimilation rates, stomatal conductance, and transpiration rates were successfully revived (~100%, 85.7%, and 83.3%) by T-ZnO NPs under stress. Plant physiological analysis revealed an enhanced chlorophyll content (39.6%) and modulated stress enzymes (superoxide dismutase peroxidase, catalase, and malondialdehyde) activity. DAB and H 2 DCFDA staining of leaf showed that the plants treated with nanoparticles showed no oxidative damage. This study highlights biosynthesized ZnONPs as an eco-friendly alternative for Botrytis cinerea management, offering potential in sustainable agriculture.