Residual ammonia shapes nitrite-oxidizing activity and microbial spatial dynamics in gel-immobilized anammox biomass under mainstream conditions

Abstract

Cell immobilization is a promising strategy for compact and efficient mainstream anammox operations. However, maintaining process stability remains challenging because of the recurring unintended activation of nitrite-oxidizing bacteria, which can outcompete aerobic ammonia-oxidizing and anammox bacteria. Therefore, a deeper understanding of the microbial interactions within immobilized anammox communities is essential for optimizing system performance; however, this remains limited. This study examined nitrogen-metabolizing microbial groups' activity and spatial dynamics in polyvinyl alcohol cryogel carriers used to treat low-strength wastewater (60 mg NH4+-N/L). While a single-stage partial nitritation-anammox process was successfully established, NH4+ depletion triggered NO3- accumulation. Analysis of nitrogen-metabolizing activities under varying operational conditions highlighted the necessity of maintaining residual NH4+ concentrations above a threshold (approximately 5 mg N/L) to suppress NO2- oxidation and thereby promote anammox. Nitrospira, identified as the sole nitrite-oxidizing genus, persisted within the gel carriers even when NO2- oxidation was effectively suppressed, and subsequent NH4+ depletions led to its reactivation and proliferation, ultimately displacing Candidatus Jettenia, the sole identified anammox genus. Microbial niche stratification within gel carriers played a central role in shaping the competition dynamics. The gel surface, dominated by aerobic ammonia-oxidizing Nitrosomonas, regulated the availability of key substrates (NH4+, NO2-, and O-2), governing the competition between Nitrospira and Ca. Jettenia in the inner layers. These findings provide valuable insights into microbial interactions in immobilized anammox systems, guiding efforts to enhance microbial synergy and advance sustainable mainstream anammox applications.