Effects of Nitrogen Input on Community Structure of the Denitrifying Bacteria with Nitrous Oxide Reductase Gene (nosZ I): A Long-Term Pond Experiment

Abstract Excessive nitrogen (N) input is an important factor influencing aquatic ecosystems and has received increasing public attention in the past decades. It remains unclear, however, how N input affects the denitrifying bacterial communities that play a key role in regulating N cycles in various ecosystems. To test our hypothesis – that the abundance and biodiversity of denitrifying bacterial communities decrease with increasing N – we compared the abundance and composition of denitrifying bacteria having nitrous oxide reductase gene (nosZ I) from sediments (0-20 cm) in five experimental ponds with different nitrogen fertilization treatment (TN10, TN20, TN30, TN40, TN50) using quantitative PCR and pyrosequencing techniques. We found that: 1) N addition significantly decreased nosZ I gene abundance, 2) the Invsimpson and Shannon indices (reflecting biodiversity) first increased significantly along with the increasing N loading in TN10~TN40 followed by a decrease in TN50, 3) the beta diversity of the nosZ I denitrifier was clustered into three groups along the TN concentration levels: Cluster I (TN50), Cluster II (TN40), and Cluster III (TN10-TN30), 4) the proportions of Alphaproteobacteria and Betaproteobacteria in the high-N treatment (TN50) were significantly lower than in the lower N treatments (TN10-TN30). 5). The TN concentration was the most important factor driving the alteration of denitrifying bacteria assemblages. Our findings shed new light on the response of denitrification-related bacteria to long-term N loading at pond scale and on the response of denitrifying microorganisms to N pollution.