Critical role of extracellular DNA in the establishment and maintenance of anammox biofilms.

Anaerobic ammonium oxidation (anammox) has been widely used for the sustainable removal of nitrogen from wastewater. Extracellular DNA (exDNA), as one of the main components of biofilms, not only determines the initial formation process, but also allows the three-dimensional structure to be maintained. Since the effects of exDNA on anammox biofilm formation are still poorly understood, this study elucidated the effects of exDNA on different stages of anammox biofilm establishment and maintenance under static conditions and its mechanism. The results revealed that exDNA mainly affected the maintenance stage of anammox biofilm formation. Compared with the absence of exDNA, nitrogen removal efficiency in the presence of exDNA was 6.17 % higher; the number of bacteria cells attached to the carrier was 2.23 times that in the absence of exDNA. The spatiotemporal distribution of bacteria was revealed by fluorescence in situ hybridization. After 30 days, the relative abundances of anammox in biofilms were 6.19 % and 0.4 % in the presence and absence of exDNA, respectively, indicating its positive role in anammox bacteria (AnAOB) adhesion and biofilm formation. The presence of exDNA in extracellular polymeric substances (EPS) promotes the synthesis of proteins and soluble microbial products. According to the extended Derjaguin-Landau-Verwey-Overbeek (X - DLVO) theory, the presence of exDNA also reduced the Lewis acid-base interaction energy and created favorable thermodynamic conditions for AnAOB adhesion. These findings advance our understanding of the role of exDNA in anammox-mediated biofilm formation and offer insights into the mechanism of exDNA in the establishment and maintenance stages.