Evolution of anammox granular sludge performance during long-term preservation: Insights from double exponential...

Because of serious shortage, the preservation of anammox granular sludge (AnGS) has become necessary for the practical application of anammox technology. However, limited information is available about the decay kinetics and biological mechanisms of AnGS during long-term preservation. In this study, AnGS was preserved at 4°C for 600 days to investigate the nitrogen removal activity (NRA), settling performance, and microbial community dynamics. Results showed that the NRA of AnGS demonstrated a distinct biphasic decay, i.e., the initial fast decay and the subsequent slow decay. It could be perfectly fit by a double exponential model (R² = 0.9725), with decay rates of 20.30^-1·d^-1 and 167.50^-1·d^-1 respectively. The settling performance of AnGS displayed good stability, with slight reductions in particle size (23.8%) and settling velocity (25.0%). This stability could be ascribed to the less protein degradation in extracellular polymeric substances (EPS). Microbial analysis revealed that the core anammox bacteria (AnAOB) populations were well retained although their functional gene expression (hzsA) was significantly downregulated. Additionally, sensitive low-abundance heterotrophic populations exhibited dynamic shifts, with significant decrease over 600 days. Mechanistically, the fast decay phase was primarily driven by the transcriptional downregulation of functional genes (hzsA) and sensitive cell death. While the slow decay phase was sustained by the deep dormancy of tolerant AnAOB. This work provides quantitative insights into long-term preservation performance of AnGS, and is expected to offer valuable guidance for improving preservation and resuscitation technologies.