Intergenerational Defense Priming in Wheat Using ZnO Nanoparticles Against Spot Blotch

Defense priming enables plants to adopt a heightened state of alert without directly triggering defense responses, enabling faster and stronger activation upon subsequent stress. In this study evaluated zinc oxide nanoparticles (ZnO NPs) as a priming in wheat against spot blotch caused by Bipolaris sorokiniana . ZnO NPs priming alone did not induce visible defense responses; however, upon pathogen challenge, primed plants exhibited a significant increase in resistance. Among the tested concentrations, 15 mg L⁻¹ concentration was most effective, significantly enhancing shoot and root growth, biomass, and leaf area, while reducing disease severity by ⁓ 66.67% compared to unprimed infected plants. Primed plants also showed elevated chlorophyll and carotenoid contents, increased total ROS scavenging activity (77.01%), and reduced lipid peroxidation (37.37%). Antioxidant enzyme activities (SOD, CAT, APX) and levels of total phenolics (111.11%), ascorbic acid, proline (41.44%), PAL (59.86%), and POX (37.91%) were markedly enhanced, contributing to improved biochemical defense. Yield traits, including spike number, spike length, spikelet fertility and density, as well as thousand grain weight were positively affected. Notably, these benefits persisted in the F1 generation, indicating intergenerational effects. Epigenetic analysis revealed altered cytosine methylation in the promoter regions of PR1 and PR3 , correlating with elevated PR3 expression and suggesting heritable epigenetic reprogramming. Overall, this work provides the first evidence of ZnO NPs functioning as a defense priming agent and its intergenerational effect in wheat, offering a sustainable and heritable approach against spot blotch, with potential implications for crop protection and yield improvement. Graphical representation of the experimental workflow depicting the green synthesis, characterization, and application of ZnO NPs for intergenerational priming in wheat ( Triticum aestivum HUW 510) against spot blotch disease. ZnO NPs were synthesized using potato extract and 0.2 M zinc nitrate Zn(NO₃)₂ at pH 6.5–7.5 and characterized by SEM, TEM, FTIR, and UV-Vis spectroscopy. Wheat seeds were treated with water (negative control), 0.2 M zinc nitrate (positive control), or ZnO NPs at concentrations of 5, 10, 15, and 20 mg/L, and grown under controlled conditions. In the F0 generation, plants were assessed for morphological traits and disease phenotypes. Based on disease response, 15 mg/L ZnO NPs was identified as the most effective concentration and was selected for further analysis of biochemical parameters, epigenetic modifications, gene expression, and yield attributes. Seeds from the F0 generation were then used to raise the F1 generation, which was similarly evaluated to investigate the persistence of priming-induced effects across generations.