Background ion shift induces microbial compensatory and structural remodeling in anammox granules

Stable municipal anammox treatment requires the retention of slow-growing anammox bacteria within structurally robust granules. However, the role of background cation availability in maintaining mineral-associated granule stability remains unclear. Here, parallel continuous-flow anammox reactors were operated under contrasting cation conditions to determine how influent hydrochemistry affects granule persistence and reactor performance. The control reactor (R1) maintained stable and efficient removal of nitrogen (over 80%), whereas the cation-limited reactor (R2) showed progressive deterioration, with the total nitrogen removal efficiency (NRE) declining to a minimum of 49%. The performance deterioration of R2 was not attributed to impaired anammox metabolic activity, as evidenced by the improved specific anammox activity and consistently maintained bacterial viability. Instead, cation deficiency destabilized the mineral-associated structure, reduced surface hydrophobicity and settleability, finally triggering biomass washout. Anammox granules initiated compensatory responses characterized by upregulated EPS accumulation, and PICRUSt2 predictions suggested potential adjustments in EPS turnover and ion transport pathways. However, these adaptations were insufficient to counteract structural collapse. Community succession featured a decline in the relative abundance of Candidatus Kuenenia and the enrichment of heterotrophic nitrate-reducers in R2. Moreover, reduced background ion availability was associated with lower microbial network connectivity and a tendency toward greater contribution of stochastic processes to community assembly. Background water chemistry therefore governs whether metabolically competent anammox biomass can remain structurally retained as durable granules and sustain stable reactor function, highlighting ionic compatibility as a key consideration for robust mainstream anammox operation.