Root-Driven Reactive Oxygen Species Controls Multidimensional Arsenic Speciation in the Rice Rhizosphere

Arsenic (As) contamination in agroecosystems poses significant risks to food security and human health. The mechanisms of As speciation change in soil within wetland rhizospheres are understood, but their location-precise importance within heterogeneous root-associated microbiomes is uncertain. While microbial processes are often considered dominant drivers of As redox transformations, the role of abiotic factors such as reactive oxygen species (ROS) remains underexplored due to limited in situ evidence. Here, we combined multiple high-resolution in situ techniques to map microscale distributions of As(III)/As(V) and key environmental parameters in rice rhizospheres across three paddy soils. A novel ratiometric fluorescent approach was developed for in situ visualization of ROS. Strong spatial correspondence was observed between ROS hotspots and decreased As(III) (R² = 0.797), exceeding that for O₂ (R² = 0.348). Integration of imaging with functional gene analysis (aioA, arsC), sterilization and ROS-quenching experiments, and structural equation modeling indicates that ROS-driven processes play a crucial role under the studied conditions. However, as gene abundance reflects microbial potential rather than activity, microbial contributions cannot be excluded. These findings highlight ROS as a key regulator of As speciation and provide new insights into coupled abiotic-biotic processes in rhizosphere environments.