Root Water Relations Drive Polystyrene Microplastic-Induced Phytotoxicity in Cucurbita moschata

Microplastics (MPs) are emerging contaminants of agricultural soils but their phytotoxic mechanisms under realistic terrestrial conditions remain poorly understood because most studies have been conducted in hydroponic systems. In this study, Cucurbita moschata was grown in a soil-like substrate and exposed to 100 nm Polystyrene microplastics (PSMPs), with different doses (0, 0.1, 0.3, and 0.5 mg mL⁻¹) to investigate their effects on plant growth, water relations, stomatal characteristics, photosynthetic pigments, oxidative stress, antioxidant responses, secondary metabolism, and root ultrastructure. PSMP exposure caused a concentration-dependent decline in relative water content (RWC), stomatal pore width, chlorophyll content, and plant growth. Scanning electron microscopy revealed progressive particle accumulation on the root surface at lower concentrations, with vascular localization only at the highest concentration, indicating increasing impairment of root water uptake. Oxidative stress increased with PSMP exposure, as reflected by elevated H₂O₂ and lipid peroxidation, while antioxidant enzymes and phenylpropanoid metabolites were progressively induced but became insufficient to prevent oxidative damage under severe stress. Multivariate data analysis model, Partial Least Squares–Discriminant Analysis (PLS-DA) further supported these responses. It identified water status, antioxidant defense, and growth as the principal processes differentiating PSMP-treated plants from controls. Taken together, the findings identified disruption of root water relations as the primary physiological driver of PSMP phytotoxicity which initiated a cascade of stomatal regulation, photosynthetic dysfunction, oxidative imbalance, and growth inhibition. This study provides a mechanistic framework for predicting the impacts of microplastic contamination on crop productivity under realistic agricultural conditions.