Reciprocal transport of carbon and nitrogen between plants and the hyphosphere microbiome in a temperate grassland

Despite its relevance for plant nutrient acquisition, the role of the hyphosphere in structuring fungal-bacterial networks and mediating carbon-nitrogen (C-N) exchange under field conditions remains unclear. We therefore examined reciprocal C-N fluxes along the plant-fungi-bacteria continuum and their link to microbial community assembly across grassland soil microhabitats. Sterilized compartmentalized ingrowth cores were exposed to soil in the field for 3 months. Plants were pulse-labeled with ¹³CO₂ and ¹³C was traced into microbial biomarkers over 7 d. ¹⁵N-labeled urea was supplied to a hyphal compartment and isotope recovery in plants was quantified. We reconstructed microbial networks and visualized microscale isotope distributions. Plant-derived ¹³C was rapidly incorporated into fungi within 1 d and into hyphosphere bacteria after 7 d. Conversely, 16.9% of the initially added 150 μg ¹⁵N was transferred from the hyphal compartment to plants. Microbial diversity declined from the rooted toward the hyphal-dominated compartments, while networks reorganized with relatively more positive fungal-bacterial associations. Arbuscular mycorrhizal fungi act as primary channels of reciprocal C-N exchange, rapidly transporting newly fixed plant C into root-distant soil zones. More broadly, soil fungi act as ecological filters shaping bacterial assembly and may foster increasingly positive associations with bacteria in hyphal-dominated habitats.