Dongdong Xu

Partial nitritation and anammox (PN/A) process has been proposed as a sustainable biotechnology for mainstream nitrogen removal. However, it remains challenging to sustain high activities of ammonia-oxidizing bacteria (AOB) and anammox bacteria and suppress the growth of nitrite-oxidizing bacteria (NOB). In this study, a novel dual-hydrogel system comprising two reactors with distinct immobilized microbial communities was developed to integrate acidic PN and anammox for low-strength mainstream wastewater treatment. The system achieved a total inorganic nitrogen removal efficiency of 81.7% at a short overall hydraulic retention time of 5.5 h. In the acidic PN reactor, hydrogel beads effectively immobilized and enriched acid-tolerant AOB (“Candidatus Nitrosoglobus”), while creating a localized low-pH niche with sufficient free nitrous acid microenvironment to selectively inhibit NOB growth. This enabled high-rate PN without strict dissolved oxygen limitation. A pH-based feeding strategy allowed the bulk pH of the acidic PN reactor to be self-maintained at ∼6.1, achieving an appropriate effluent ratio of nitrite to ammonium and minimizing acid inhibition of the downstream anammox process. In the anammox reactor, the beads maintained a relatively higher internal pH, forming a favorable niche for retained anammox bacteria (“Candidatus Brocadia”) and facilitating high-rate nitrogen removal. The hydrogel beads also exhibited excellent structural stability during long-term operation (over 300 days). These results demonstrate an effective approach for simultaneously sustaining AOB and anammox activities while suppressing NOB, providing a promising approach for stable PN/A to treat mainstream wastewater.

The granule-based anammox process is a promising biotechnology for nitrogen removal; however, it is challenging to achieve a high rate and efficiency simultaneously. In this study, we addressed this challenge by coupling two granule types with complementary kinetics in an anammox sludge bed reactor. Bright red granules (GR) exhibited higher maximum specific anammox activity (SAA) but lower substrate affinity; they predominantly occupied the reactor's lower section, where substrate concentrations are high, forming a high-capacity layer. Yellow-brown granules (GY) showed higher substrate affinity but lower maximum SAA; they mainly populated the upper low-substrate zone, forming an efficient substrate-scavenging layer. This self-organized granule stratification enabled both a high nitrogen removal rate (14.5 kg-N/(m³·d)) and high efficiency (90.2%). Physicochemical property and microbial community analyses of granules showed that the stratification of "affinity" was mainly governed by the granular structure and mass transfer rather than intrinsic microbial traits. By actively withdrawing excessive sludge from the fast-growing lower zone while preserving biomass in the upper zone, such stratification can be stably maintained. Collectively, these findings advance our understanding of granule heterogeneity in anammox sludge bed bioreactors and offer a practical strategy for performance optimization.

Anammox is a sustainable biotechnology for nitrogen removal from wastewater. The granule-based anammox process provides excellent biomass retention and is widely applied for the treatment of high-strength wastewater, but its efficacy declines significantly when treating sulfate-rich high-salinity wastewater. This study evaluated the impacts of mass transfer limitations caused by sulfate-induced crystallization in the micropores of anammox granules. Long-term bioreactor operation revealed that salts containing chloride and sulfate inhibited total-nitrogen removal rates by 58% more than chloride salts alone. Through a combination of batch activity assays, nanocomputed tomography (nano-CT), Raman spectroscopy, and numerical simulations, we identified the in situ crystallization of Ca3(SO3)2.12(SO4)0.88 as a key stressor in sulfate-containing systems. This crystallization, facilitated by localized Ca2+ enrichment on EPS-polysaccharides and the microporous architecture of granules, preferentially occurred in narrow pores and high-curvature regions. The resulting pore blockage accounted for ∼40% of the observed decline in anammox activity, primarily through restricted substrate diffusion. By highlighting the roles of sulfate and calcium in restricting mass transfer in anammox granules, this work provides a mechanistic basis for optimizing the performance of anammox systems.