Ammonia-oxidizing archaea are dominant over comammox in soil nitrification under long-term nitrogen fertilization

The complete ammonia oxidizers (comammox) capable of catalyzing nitrification, oxidizing ammonia to nitrate, via activity of only one type of microbes were recently discovered which has updated our knowledge of traditional two-step nitrification. The extent of contribution of comammox and canonical ammonia oxidizers including ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) to soil nitrification, especially in soils with long-term input of nitrogen (N) fertilizers, remains unknown. The transcriptional abundance of amoA gene from comammox, AOA, and AOB in soils fertilized for 29 years was investigated in different seasons and soil layers via quantitative PCR. The results showed that comammox were detected in all soil samples; however, AOA and AOB had significantly higher transcriptional abundance of amoA gene than comammox. Nitrification activity was most significantly correlated with the transcriptional abundance of AOA amoA gene (Pearson correlation, r = 0.217, P < 0.05) suggesting AOA were the dominant contributors to soil potential nitrification. Lower abundances of amoA gene transcripts were observed in July than in April and November. The application of high level of mineral N fertilizer decreased the abundance of both AOA and AOB; however, long-term input of organic manure combined with mineral N fertilizer stabilized the abundances of ammonium-oxidizing microbes in soils. Seasonal variation and fertilization regimes substantially affected the abundance of both AOA and AOB, but AOB were not as sensitive in responding to the seasonal variation and fertilization as AOA. The analysis of RDA and VPA demonstrated that sampling month, soil depth, and fertilization regime explained 30.20%, 11.46%, and 5.40% of the variation in nitrification microorganism amoA gene composition, respectively. Seasonal variation exerted the most influences on the nitrifiers’ composition, and soil depth and fertilization regime were also important factors in shaping the nitrifier communities. According to the correlation analysis, NO3−–N content was the most important soil property in impacting the transcriptional abundance of amoA gene, and the amoA gene transcript abundance decreased with increasing NO3−–N content. The results suggest that the activity of comammox may be more inhibited by the long-term nitrogen fertilization than canonical ammonia oxidizers in agricultural soils. This study provides insights into the different responses of comammox and canonical ammonia oxidizers to fertilization, seasonal variation, and soil depth and their relative contributions to nitrification in agricultural soil.