Overcoming elemental sulfur bioavailability limitations with sponge iron coupling for enhanced nitrate removal: Novel perspective on electron transfer and iron-nitrogen-sulfur metabolism.

This study developed a novel sponge iron (sFe⁰) and elemental sulfur (S⁰) coupled autotrophic denitrification biofilter (S⁰-sFe⁰AD). At the ratio Fe⁰ to S⁰ of 0.5 and a hydraulic retention time of 1.5 h, the system achieved high nitrate and phosphate removal rates of 969.7 mgN·L^-1·d^-1 and 56.1 mgP·L^-1·d^-1, with efficiencies over 98.5 %. Activated biochemical sulfidogenic pathways enabled in situ regeneration of highly bioavailable FeS. This process not only expanded electron donor pool but also reduced sulfate production by facilitating multi-pathway denitrification. Moreover, FeS-mediated direct extracellular electron transfer promoted iron-sulfur redox cycling. Metagenomic analysis further revealed enhanced iron and energy metabolism within the coupled system. The enriched iron-sulfur redox bacteria (Thiobacillus, Desulfurivibrio and Geothrix) and genes (narB, mtrC, sox, fccAB and sir) facilitated the establishment of a self-sustaining iron-sulfur cycle, thereby extending system longevity. This study provides novel insights for developing efficient iron-sulfur coupled autotrophic denitrification technology for sustainable wastewater treatment.