Up-flow dual-loop anammox system treating mature landfill leachate: ultra-efficient nitrogen removal, anammox granules formation, and functional divergence

Abstract

Achieving the effective retention and rapid enrichment of anammox bacteria (AnAOB) remains a challenge in landfill leachate treatment systems. The application of the anammox process is also restricted by an inadequate supply of nitrite and poor resistance to adverse environments. To overcome these obstacles, an up-flow dual-loop anammox (UDLA) system treating mature landfill leachate was established, based on partial nitrification, anammox, and partial denitrification (PNAPD) technology. The nitrogen removal efficiency and nitrogen loading rate reached 92.9 ± 1.5% and 0.4 kg N/m3/d, and isotope tracing tests demonstrated that 88.2 ± 6.5%-93.9 ± 3.9% of the nitrogen loss contributed by anammox pathway. Under the up-flow dual-loop strategy, anammox granules were successfully cultivated when flux ratios of effluent/influent and produced gas/influent were maintained at 1640.0%, and 250.0%, respectively. The proportion of medium granules (500–1000 μm) with the highest PNAPD activity increased from 0 to 24.9%, whereas the percentage of floc (0–200 μm) reduced from 98.4% to 23.3%. The mechanism of anammox granules formation in the UDLA system was revealed by multidimensional analysis. Candidatus_Brocadia_sapporoensis (13.3%-13.5%), Candidatus_Kuenenia_stuttgartiensis (2.0%-4.8%), and Candidatus_Brocadia_sp. (0.9%-1.1%) were the predominant anammox species, enriched in medium and macro granules. Metagenomic sequencing confirmed the functional divergence of nitrogen and carbon metabolism among the size-fractioned granules, possible functions of specific species, and gene distribution in the nitrogen metabolism pathway. The abundant GlnA, GltB(D), GdhA, GudB (GDH2), and GLUD1_2 genes might promote the synthesis of adhesive proteins and further facilitate microbial aggregation. This study enhanced the comprehension of anammox granules formation and showed important implications for leachate treatment in engineering.