LAUSR

Excessive nitrogen (N) input is an important factor influencing aquatic ecosystems and has received increasing public attention in the past decades. It remains unclear 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 nos Z 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.