Root entry pathways dictate type-dependent uptake, translocation and phytotoxicity of microplastics in tobacco

Microplastic pollution increasingly threatens agricultural ecosystems, yet how different microplastic types interact with plants remains largely unknown. Here, we systematically compared the uptake, translocation, and physiological impacts of polystyrene (PS) microspheres and polyacrylonitrile (PAN) microfibers in hydroponically grown tobacco. We identified distinct root entry pathways: PS microspheres penetrated via the root apex and lateral root initiation sites, whereas PAN microfibers infiltrated exclusively through intercellular gaps at lateral root emergence, failing to cross the root apex. Both types subsequently translocated to aerial tissues, with PS microspheres demonstrating significantly higher root uptake, retention, and shoot translocation efficiency than PAN microfibers. Growth responses were strongly type- and concentration-dependent: PS microspheres inhibited growth, photosynthesis, and root development at ≥ 1 mg/L, while PAN microfibers promoted growth over a broader range (0.1–10 mg/L), becoming inhibitory only at ≥ 50 mg/L. PS exposure induced more pronounced disruptions in nitrogen assimilation, carbon metabolism, and key hormone signaling (ethylene, auxin, cytokinin, gibberellin), alongside greater reactive oxygen species (ROS) accumulation and intensified antioxidant responses. Transcriptomic analyses corroborated these physiological findings, revealing stronger activation of stress-responsive genes under PS treatment. This study reveals that structurally distinct microplastics differ not only in uptake and translocation efficiency, but also in root entry pathways and downstream physiological and transcriptomic responses, identifying microplastic morphology as a key determinant of plant phytotoxicity. These findings provide important insights for ecological risk assessment and mitigation of agricultural microplastic pollution. • PS microspheres and PAN microfibers differ in uptake and translocation in tobacco. • Polystyrene microspheres penetrate both the root apex and lateral-root initiation sites, whereas polyacrylonitrile microfibers infiltrate only intercellular spaces at lateral-root initiation and do not cross the apex. • PS microspheres shows higher toxicity at lower concentrations than PAN microfibers. • PS microspheres disrupts nitrogen, carbon metabolism, and hormone signaling. • PS microspheres triggers stronger stress-related gene expression than PAN microfibers.