Spatially resolved denitrification coupled with methane and arsenite oxidation at the millimeter-scale straw-soil...

Straw return reshapes the biogeochemical processes in paddy soils by driving microbial transformation of key elements. Despite growing awareness of these individual processes, the integration of these processes under millimeter-scale spatiotemporal heterogeneity remains unclear. Combining high-resolution geochemical profiling with multiomics, we revealed that straw addition altered the depth-dependent dynamics of arsenic, carbon, and nitrogen, establishing a sophisticated three-layer microbial stratification. We identified 1) an 18 mm organic matter (OM)-rich layer extending from the straw layer, which serves as a methanogenic epicenter co-occurring with active nitrogen fixation microbes; 2) an overlying layer dominated by aerobic methane oxidation and denitrification microbes; and 3) a deeper substraw layer dominated by anaerobic arsenite oxidation and denitrification microbes. Significantly positively correlated abundances of transcribed mcrA with nifH genes and pmoA or aioA/arxA with denitrification genes were identified. Corroboratively, intensified co-occurrence patterns of mcrA with nifH, pmoA with denitrification, and aioA/arxA with denitrification genes were observed in the OM-rich, upper, and lower layers, respectively. Moreover, the co-occurred mcrA-nifH and aioA-nirS/arxA-narG genes in different metagenome-assembled genomes presented 80.6 to 260.8- and 1.55 to 6.85-fold greater transcriptional activity in the OM-rich and lower layers than in the other layers, respectively. Our results demonstrated that straw incorporation established a dynamic soil redox zone, restructuring millimeter-scale microbial networks and promoting potentially coupled denitrification with arsenite or methane oxidation, as well as methanogenesis with nitrogen fixation. These findings provide a mechanistic basis for optimizing subsurface straw placement and nitrate application to enhance nutrient cycling and mitigate environmental risks.