External redox couple enhanced anammox sludge activity at low temperature: Insight into intracellular resource...

Anaerobic ammonium oxidation (anammox), an energy-efficient deamination biotechnology, faces operational challenges in low-temperature environments. Enhancing the metabolic activity of anammox bacteria (AnAOB) is pivotal for advancing its application in mainstream municipal wastewater treatment. Inspired by the metabolic adaptability of AnAOB and based on our previous findings, this work investigated the enhancement of intracellular ATP and NADH synthesis through the exogenous supply of reduced humic acid (HA_red) and H₂O₂ redox couple, aiming to augment AnAOB activity under low-temperature conditions. Our experimental setup involved continuous dosing of 0.0067 μmol g^-1 volatile suspended solid of H₂O₂ and 10 mg g^-1 volatile suspended solid of HA_red into a mainstream anammox reactor operated at 15 °C with an influent TN content of 60 mg/L. The results showed that HA_red / H₂O₂ couple succeeded in maintaining the effluent TN at 10.72 ± 0.91 mg l^-1. The specific anammox activity, ATP and NADH synthesis levels of sludge increased by 1.34, 2.33 and 6.50 folds, respectively, over the control setup devoid of the redox couple. High-throughput sequencing analysis revealed that the relative abundance of Candidatus Kuenenia after adding HA_red / H₂O₂ couple reached 3.65 % at the end of operation, which was 5.14 folds higher than that of the control group. Further metabolomics analysis underscored an activation in the metabolism of amino acids, nucleotides, and phospholipids, which collectively enhanced the availability of ATP and NADH for the respiratory processes. These findings may provide guidance on strategy development for improving the electron transfer efficiency of AnAOB and underscore the potential of using redox couples to promote the mainstream application of anammox technology.