Rapid adaptation and enrichment of salt-tolerant anammox via dosing of chemical enhancers in packed-bed biofilm reactor.

The application of anammox-based processes for saline wastewater treatment is constrained by the scarcity of salt-tolerant seed sludge and the lengthy adaptation periods. To overcome this challenge, exogenous chemical enhancers, hydrazine (N2H4, 5 mg/L) and glycine betaine (GB, 30 mg/L), were introduced and evaluated for their roles in facilitating salt-adapted anammox biofilms enrichment from freshwater seed in packed-bed biofilm reactors. Hydrazine addition for 15 days increased the nitrogen removal rate from approximately 50 to 441.1 mg N/(L·d) within 70 days, which was substantially higher than that achieved through natural acclimation (192.2 mg N/(L·d)). When GB was subsequently supplemented for 30 days to the naturally acclimated reactor, its nitrogen removal rate rapidly increased to 1000 mg N/(L·d) within 30 days and further to 3000 mg N/(L·d) within 60 days, catching up the reactor receiving N2H4 from the outset. According to community analysis, performance recovery coincided with immediate shift from Ca. Brocadia to Ca. Kuenenia, with its relative abundance surged ∼15-fold within 20 days, highlighting the remarkable stimulatory effect of enhancers on Ca. Kuenenia's proliferation. Inferred from KEGG pathway studies, N2H4 primarily enhanced oxidative phosphorylation and ATP synthesis, providing energetic support for early recovery of the proton motive force and osmotic balance. In contrast, GB stabilized cellular osmotic conditions and membrane structures, enabling reallocation of metabolic resources toward antioxidant defense, cellular repair, and folate biosynthesis under saline stress. This alleviated the energetic burden associated with ion transport and lipid remodeling, thereby promoting sustained recovery of the anammox community.