Metagenomic insights into nitrogen-cycling microbial communities and their relationships with nitrogen removal potential in the Yangtze River.

Nitrogen (N) pollution is a major threat to river ecosystems worldwide. Elucidating the community structure of N-cycling microorganisms in rivers is essential to understanding how ecosystem processes and functions will respond to increasing N inputs. However, previous studies generally focus on limited functional genes through amplicon sequencing or quantitative PCR techniques and cannot cover all N-cycling microorganisms. Here, metagenomic sequencing and genome binning were used to determine N-cycling genes in water, channel sediments, and riparian soils of the Yangtze River, which has been heavily polluted by N. Additionally, the denitrification and anaerobic ammonium oxidation (anammox) rates that reflect N removal potential were measured using N isotope pairing technique. Results showed that functional genes involved in organic N metabolism (i.e., organic degradation and synthesis) and nitrate reduction pathways (i.e., dissimilatory and assimilatory nitrate reduction to ammonium and denitrification) were more abundant and diverse than other N-cycling genes. A total of 121 metagenome-assembled genomes (MAGs) were identified to be involved in N-cycling processes, and the key MAGs were mainly taxonomically classified as Alphaproteobacteria and Gammaproteobacteria. The abundance and diversity of most N-cycling genes were higher in soils and sediments than in water, as well as higher in downstream and midstream than in upstream sites. These spatial variations were explained not only by local environment and vegetation but also by geographical and climatic factors. N removal process (i.e., denitrification and anammox) rates were significantly related to the abundance or diversity of several N-cycling genes, and climate and edaphic factors could regulate denitrification and anammox rates directly and indirectly through their effects on functional genes. Overall, these results provide a new avenue for further understanding the biogeographic patterns and environmental drivers of N-cycling microorganisms in rivers from the metagenomic perspective.