Anaerobic ammonium oxidation (anammox) promoted by pyrogenic biochar: Deciphering the interaction with extracellular polymeric substances (EPS).

Efficient biological nitrogen removal (BNR) by anaerobic ammonium oxidation (anammox) can be achieved with presence of redox-active pyrogenic biochar that potentially acting as an insoluble electron acceptor. Anammox bacteria and other symbiotic consortia are surrounded by extracellular polymeric substances (EPS) forming aggregate architecture, which also contains electrochemical-active biomolecules such as aromatic proteins and humic substances. Therefore, understanding the role of EPS is necessary in biochar-promoting anammox process. Herein, we investigated the influence of biochar with granular-sized (GP) and micrometer-sized (MP) particle sizes on microbiology and characteristics of EPS in anammox sludge. Addition of GP and MP biochar not only improved the BNR efficiency by 17.5% and 34.6%, respectively, but also increased the relative abundance of Candidatus Brocadia. The bulk and bound EPS contents substantially decreased in biochar-amended groups, while more slime EPS was produced. Spectroscopic (FTIR, Raman, and circular dichroism) and electrochemical (voltammetry and impedance spectrum) analyses revealed that biochar addition enhanced the structural integrity and electron-transfer capability of anammox sludge. EPS depletion led to a steep decrease in BNR efficiency (21.5% vs 83.0% with EPS-retained sludge), whereas it resumed to 42.1% in the presence of MP biochar. Electron transport system activity data showed that biochar replenished the loss of anaerobic respiration metabolism due to EPS depletion. In summary, these results suggested that EPS possibly work as transient mediator for shuttling electrons from ammonium oxidation to soluble (nitrite) and insoluble electron acceptors (redox-active biochar).