Impact of Stepwise Salinity Elevation on Nitrogen Removal and Microbial Properties of Morphologically Distinct Anammox Sludge

The anaerobic ammonium oxidation (anammox) process offers potential for saline wastewater treatment but is hindered by salt inhibition. This study investigates the salt tolerance mechanisms of granular (R1), biofilm-carrier (R2), and floccular (R3) sludge in up-flow anaerobic sludge blanket (UASB) reactors under 0–20 g/L NaCl. Granular sludge outperformed other biomass types, maintaining >90% ammonia nitrogen (NH4+-N) removal at 20 g/L NaCl due to structural stability and extracellular polymeric substances (EPS) adaptation (shift from hydrophobic proteins to polysaccharides). Microbial analysis revealed a transition from Planctomycetes/Proteobacteria to salt-tolerant Pseudomonadota, with Candidatus_Kuenenia replacing Candidatus_Brocadia as the dominant anaerobic ammonium oxidation bacteria (AnAOB) (reaching 14.5% abundance in R1). Genetic profiling demonstrated coordinated nitrogen metabolism: Hzs/Hdh inhibition (>85%) and NirBD/NrfAH activation (0.23%) elevated NH4+-N, while NarGIV/NapA decline (1.10%→0.58%) increased nitrate nitrogen (NO3−-N). NxrB/NirSK maintained low nitrite nitrogen (NO2−-N), and GltBD upregulation (0.43%) enhanced osmoregulation. These findings underscore the superior resilience of granular sludge under high salinity, linked to microbial community shifts and metabolic adaptations. This study provides critical insights for optimizing anammox processes in saline environments, emphasizing the importance of biomass morphology and microbial ecology in mitigating salt inhibition.