Microfibers (MFs), primarily originating from sewage sludge and laundry effluents, are the most prevalent form of microplastics (MPs) in agricultural soils. While their ecological effects have been explored, the visualization, crop-level accumulation, and potential transport mechanisms of MFs within soil–plant systems remain poorly understood. This study combines 1,3,6,8-pyrene tetrasulfonic acid (PTSA) fluorescent staining with a sequential multimodal microscopy workflow to effectively track the distribution, adsorption, accumulation, and uptake of MFs under realistic soil cultivation conditions. Three edible vegetables—lettuce, Chinese cabbage, and cherry radish—were used to evaluate species-specific response patterns. The results revealed clear differences in MF interactions across species: lettuce exhibited strong MF adsorption on root surfaces and subsequent penetration via crack-entry and apoplastic pathways without entering cells. In contrast, Chinese cabbage and cherry radish showed limited MF adsorption and no uptake. These patterns were associated with root permeability and antioxidative capacities, indicating that plant functional traits play a critical role in determining the transport capacity of MPs. Beyond introducing a novel method for MF visualization in complex terrestrial matrices, this study provides new insights into the risks posed by MFs to soil-plant systems. The findings also highlight potential threats to food safety and underscore the need to establish plant-specific thresholds and pollution mitigation strategies to support sustainable agriculture and protect public health
