Root exudates recruit beneficial microbes to promote anammox-driven nitrogen cycling in wetland

Anammox bacteria serve as a major biological sink in nitrogen (N) cycling within wetland, yet the hydrophyte root exudates-mediated microbial interplay mechanism that sustain their activity and ecosystem function remain unclear. To address this gap, we established flow-controlled microcosms planted with Iris pseudacorus, combined with ¹⁵N stable isotope tracing and metagenome-assembled genomes (MAGs) analysis. Our findings revealed that root exudates significantly enhanced in-situ anammox rates (rhizosphere: 5.9 ± 2.0 mg N/(m³·d), non-rhizosphere: 0.4 ± 0.02 mg N/(m³·d), p < 0.001), leading to a remarkable enrichment of anammox bacteria in the rhizosphere (6.5 × 10⁷ copies/g dry sludge, p < 0.001). We further uncovered a previously overlooked partial denitrification pathway that supplied nitrite, substantially increasing anammox contributions to rhizosphere N removal (16.6 ± 4.1%). Key bioactive components, flavonoids and amino acids, selectively recruited beneficial rhizobacteria affiliated to Pseudomonadota and Bacteroidota. MAGs-based analysis revealed that these microbial taxa encoded pathways for producing essential substrates (nitrite loop) and metabolites (cofactor, biotin) supporting anammox metabolism. The symbiotic interaction facilitated the survival and metabolic activity of anammox bacteria in the oligotrophic rhizosphere habitat. These findings unveil a natural plant-microbiota interaction that effectively enhances the sustainability of N cycling and provide new insights for optimizing nitrogen removal strategies in engineered wetland systems.