Methyl jasmonate-loaded chitosan nanoparticles and biochar improve maize thermotolerance

Heat stress is a major abiotic constraint that severely limits maize (Zea mays L.) productivity under changing climate conditions. This study explored a novel integrative strategy to enhance thermotolerance in maize through the combined application of methyl jasmonate-loaded chitosan nanoparticles (MJNPs) and eucalyptus-derived biochar (EBB). Methyl jasmonate was nano-encapsulated using the ionic gelation method and characterized by SEM, TEM, and FTIR analyses, which confirmed uniform spherical nanoparticles and effective surface functionalization. A greenhouse experiment was conducted under controlled heat stress (40 °C) to evaluate physiological, biochemical, nutrient uptake, yield, and gene expression responses across eight treatments. Relative to non-stressed control plants, heat stress alone reduced plant height by 37%, photosynthetic rate (PN) by 46%, relative water content (RWC) by 25%, and grain number and grain weight by 25% and 6%, respectively. However, the combined MJNPs + biochar treatment under heat stress (HEMN) markedly alleviated these adverse effects. Compared with heat-stressed plants without amendments, HEMN increased plant height by 39%, RWC by 8%, membrane stability index (MSI) by 14%, and PN by 21%. In addition, grain number and seed weight increased by 10% and 6%, respectively, relative to heat-stressed plants, while water-use efficiency (WUE) improved by 13% under the same comparison. Nutrient uptake of phosphorus, magnesium, and iron increased by 15-22% in HEMN-treated plants compared with heat-stressed controls. Gene expression analysis revealed pronounced upregulation of stress-responsive genes, including HSP70, DHN3, and LEA-1, as well as auxin biosynthesis-related genes (TAA1, ZmYUC1, CYP79B2) and aquaporins in HEMN-treated plants relative to heat stress alone, indicating activation of coordinated molecular defense mechanisms. Furthermore, principal component analysis (PCA) and hierarchical clustering of gene-expression heatmaps confirmed strong multivariate associations between enhanced physiological performance and transcriptional activation, supporting the integrated nature of thermotolerance regulation. These findings demonstrate that the synergistic application of MJNPs and biochar significantly enhances maize thermotolerance relative to heat stress alone by improving water relations, nutrient homeostasis, photosynthetic performance, and molecular stress responses. This integrated nano-biochar strategy represents a scalable and environmentally sustainable approach for mitigating climate-induced heat stress and improving crop resilience in future agricultural systems.