Rhizosphere micro-environment restructuring by foliar 24-epibrassinolide enhances wheat resilience to polyethylene nanoplastics.

INTRODUCTION: The stress effects of nanoplastics (NPs) on wheat growth have been increasingly reported. The plant growth regulator 24-epibrassinolide (EBL) shows potential in mitigating abiotic stress, but the mechanisms by which it alleviates NPs toxicity-particularly through root metabolites, rhizosphere metabolites, and rhizosphere microbial communities-remain unclear. OBJECTIVES: This study aimed to investigate how EBL mitigates polyethylene nanoplastics (PE-NPs) stress in wheat, focusing on plant growth, root metabolism, rhizosphere metabolites, and rhizosphere microbial communities. METHODS: Wheat seedlings were treated with PE-NPs with or without exogenous EBL. Growth parameters, photosynthesis, oxidative damage, and antioxidant enzyme activities were measured. Integrated metabolomics and mass spectrometry imaging (MSI) technologies were employed to characterize metabolic alterations and to spatially visualize metabolite distributions. Rhizosphere metabolites were analyzed using triple quadrupole mass spectrometry, and rhizosphere microbial communities were characterized through high-throughput sequencing, with functional profiles inferred based on 16S rRNA data. RESULTS: EBL treatment significantly alleviated PE-NPs toxicity, increasing shoot and root lengths by 6 % and 14 % and enhancing fresh weights by 16 % and 32 %, respectively. It improved photosynthetic efficiency, reduced malondialdehyde (MDA) accumulation in shoots (29 %) and roots (25 %), and enhanced antioxidant activity and increased jasmonic acid levels. Metabolomics and spatial metabolomics revealed changes in flavonoid biosynthesis, starch/sucrose metabolism, and purine metabolism in wheat roots. Rhizosphere metabolite analysis showed enrichment of galactose, starch/sucrose, and alpha-linolenic acid metabolism. Additionally, EBL enhanced the abundance of Actinobacteriota and Acidobacteriota and increased predicted plastic-degradation-related functions based on FAPROTAX analysis, improved soil physicochemical properties, and increased available nitrogen and phosphorus levels. CONCLUSION: Exogenous EBL alleviated PE-NPs toxicity in wheat by improving photosynthetic performance, reprogramming root metabolism, and reshaping rhizosphere metabolite and microbial profiles. These findings provide new insights into EBL-mediated tolerance to PE-NPs and support further evaluation of foliar EBL as a plant-centered strategy for improving wheat resilience under nanoplastic stress.