Gene-resolved metagenomics reveals partitioned nitrogen transformation potential in a landfill leachate sequencing batch reactor microbiome

Landfill leachate can contain high concentrations of ammonia and organic nitrogen requiring treatment before discharge. Sequencing batch reactors (SBRs) promote biological nitrogen removal through alternating aerobic and anoxic conditions, but taxonomic profiling alone cannot resolve how nitrogen transformation and competing retention pathways are distributed across microbial communities. This study applied gene-resolved metagenomics to a sludge sample collected from a UK landfill leachate SBR. Nitrogen-metabolism genes were linked to contig-level taxonomic assignments and metagenome-assembled genome bins to examine nitrification, denitrification, dissimilatory nitrate reduction to ammonium (DNRA), and assimilatory nitrate reduction (ANR). Nitrogen-transformation potential was partitioned across dominant and minor taxa. Nitrospirota and Pseudomonadota were the most abundant classified phyla, whereas Actinomycetota represented only 4% of the community but included Actinomycetes, which showed the highest within-taxon proportion of nitrogen metabolism associated genes. amoABC was assigned only to Nitrosomonas and Nitrososphaerota, whereas denitrification genes were more widely distributed but frequently occurred as partial pathways. nosZ was detected only in the minor phyla Chloroflexota, Acidobacteriota and Myxococcota. DNRA and ANR-associated genes occurred across multiple lineages; nirA in Nitrospira and DNRA-associated nirBD elsewhere within Nitrospirota suggested nitrifier-associated lineages may also encode nitrogen retention potential. Gene-resolved metagenomics identified candidate low-abundance contributors, restricted distributions of genes associated with ammonia oxidation and nitrous oxide reduction, and nitrogen-retention potential not apparent from taxonomic composition alone. Because the study examined DNA-level potential in a single sludge sample, expression, reactor chemistry and longitudinal data are needed to establish pathway activity and operational relevance.