Biosynthesized ZnO nanoparticles enhance salt tolerance in Brassica Napus through antioxidant and gene regulatory pathways

Soil salinity is a major constraint to crop establishment and yield, particularly during germination and early seedling development of Brassica napus. Here, we report that zinc oxide nanoparticles (ZnONPs) biosynthesized using Pseudomonas aeruginosa alleviate salinity stress through coordinated physiological, biochemical, and molecular mechanisms. Comprehensive physicochemical analyses confirmed the formation of hexagonal wurtzite ZnONPs with a plate-like morphology (thickness ~ 54.5 nm) and an optical band gap of 3.01 eV. Canola seeds were primed with ZnONPs (25-100 mg L⁻¹) under control or 150 mM NaCl stress and subsequently grown in hydroponics. Salt stress reduced germination (41%), biomass, and vigor, while elevating lipid peroxidation and reactive oxygen species. ZnONPs, particularly at 50 mg L⁻¹, restored germination to control levels, enhanced shoot (52 cm) and root (35 cm) elongation, and nearly doubled salt-stressed biomass. ZnONP treatment suppressed malondialdehyde and H₂O₂ accumulation below control values, while up-regulating superoxide dismutase and catalase activity. Furthermore, ZnONPs reduced osmolyte (proline and glycine betaine) accumulation, increased chlorophyll content, lowered Na⁺, and elevated K⁺ and Zn uptake, thereby improving ion homeostasis. Transcript analysis revealed that salinity strongly induced the stress-responsive kinase BnSRK2D (~ 86-fold) and repressed auxin-responsive genes, whereas 50 mg L⁻¹ ZnONPs normalized these responses, down-modulating stress signaling and restoring auxin pathways. Collectively, these findings demonstrate that biogenic ZnONPs mitigate salinity stress in B. napus through integrated regulation of antioxidant defense, ion balance, and hormone-mediated gene expression, highlighting their potential as sustainable nanopriming agents for crop improvement under saline conditions.