Iron Robustly Stimulates Simultaneous Nitrification and Denitrification Under Aerobic Conditions

Simultaneous nitrification and denitrification (SND) is a promising single-reactor biological nitrogen-removal method. Activated sludge with and without iron scrap supplementation (Sludge-Fe and Sludge-C, respectively) was acclimated under aerobic condition. The total nitrogen (TN) content of Sludge-Fe substantially decreased from 25.0 +/- 1.0 to 11.2 +/- 0.4 mg/L, but Sludge-C did not show the TN-removal capacity. Further investigations excluded a chemical reduction of NO3--N by iron and a decrease of NH4+-N by microbial assimilation, and the contribution of SND was verified. Moreover, the amount of aerobic denitrifiers, such as bacteria belonging to the genera Thauera, Thermomonas, Rhodobacter, and Hyphomicrobium, was considerably enhanced, as observed through Miseq Illumina sequencing method. The activities of the key enzymes ammonia monooxygenase (AMO) and nitrite oxidoreductase (NXR), which are associated with nitrification, and periplasmic nitrate reductase (NAP) and nitrite reductase (NIR), which are related to denitrification, in Sludge-Fe were 1.23-, 1.53-, 3.60-, and 1.55-fold higher than those in Sludge-C, respectively. In Sludge-Fe, the quantity of the functional gene NapA encoding enzyme NAP, which is essential for aerobic denitrification, was significantly promoted. The findings indicate that SND is the primary mechanism underlying the removal of TN and that iron scrap can robustly stimulate SND under aerobic environment.