Effect of Different Ammonia Concentrations on Community Succession of Ammonia-oxidizing Microorganisms in a Simulated Paddy Soil Column Hu Baolan, Liu Shuai, Shen Lidong, Zheng Ping, Xu Xiangyang, Lou Liping* Department of Environmental Engineering, Zhejiang University, Hangzhou, Zhejiang, China Abstract Ammonia oxidation is performed by both ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA). To explore the effect of ammonia concentration on the population dynamic changes of ammonia-oxidizing microorganisms, we examined changes in the abundance and community composition of AOA and AOB in different layers. Most of the archaeal amoA sequences were Nitrosotalea-related and the proportion that Nitrosotalea cluster occupied decreased in the surface layer and increased in the deep layer during the cultivation process. Nitrosopumilus-related sequences were only detected in the deep layer in the first stage and disappeared later. Both phylogenetic and quantitative analysis showed that there were increased Nitrosomonas-related sequences appeared in the surface layer where the ammonia concentration was the highest. Both AOA and AOB OTU numbers in different layers decreased under selective pressure and then recovered. The potential nitrification rates were 25.06 mg?N?L21?g21 dry soil?h21 in the mid layer which was higher than the other two layers. In general, obvious population dynamic changes were found for both AOA and AOB under the selective pressure of exogenous ammonia and the changes were different in three layers of the soil column. Citation: Baolan H, Shuai L, Lidong S, Ping Z, Xiangyang X, et al. (2012) Effect of Different Ammonia Concentrations on Community Succession of Ammonia- oxidizing Microorganisms in a Simulated Paddy Soil Column. PLoS ONE 7(8): e44122. doi:10.1371/journal.pone.0044122 Editor: Matthias Horn, University of Vienna, Austria Received February 15, 2012; Accepted July 30, 2012; Published August 31, 2012 Copyright: 2012 Baolan et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: The authors wish to thank the National Natural Science Foundation of China (40801198), National Key Technologies Research and Development program of China (2012BAJ25B00) and the Natural Science Foundation of Zhejiang province (Y507227). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: baolanhz@hotmail.com Introduction Nitrification is critical in the global nitrogen cycle and is a unique pathway linking reduced inorganic nitrogen with oxidised inorganic nitrogen in nature [1]. Ammonia oxidation is the first and rate-limiting step of nitrification. It was assumed that all ammonia oxidisers were bacteria until ammonia-oxidizing archaea were found. The findings of Venter et al. [2] and Treusch et al. [3] indicated the posibility of archaea involving in ammonia oxidation and the exist of ammonia-oxidation archaea was finally certified by the enrichments and pure cultures of several strains of AOA [4– 10]. Recent research has demonstrated that ammonia-oxidizing archaea may form a separate and deep-branching phylum, the Thaumarchaeota [11–13]. Molecular biological research based on the functional gene amoA and the 16SrRNA gene has revealed a widespread distribution of AOA [14–17]. Quantitative PCR analyses found that AOA normally outnumbered their substrate competitors, such as AOB, in both soil [17] and water [18], but the relative contributions of AOA and AOB in the soil nitrogen cycle remained undetermined. Some researchers believed that AOB played a more important role than AOA in soil nitrification. Jia and Conrad [19] found that changes in potential nitrification rates were only correlated with the number of AOB amoA genes when they added substrate (NH3) or inhibitor (C2H2) of ammonia oxidation to the soil. Subsequently, Di et al. [20] discovered the same phenomenon in New Zealand grassland. In contrast, other researchers regarded AOA as the main drivers of ammonia oxidation in soil. Gubry-Rangin et al. [21] reported that the potential nitrification rates were positively correlated with the number and transcriptional activity of AOA amoA genes. Zhang et al. [22] confirmed the dominant role of AOA in soil nitrification by means of stable isotope probe and the quantitative analysis of archaral amoA genes. Offre et al. [23] also demonstrated that the growth of only archaeal but not bacterial ammonia oxidizers occurred in microcosms with active nitrification. Differences in cell size, specific cell activity and other physiological characteristics may explain the different contributions of AOA and AOB to soil nitrification. In addition, AOA and AOB may compete mutually or exhibit functional redundancy under some conditions, whereas under other conditions the fundamental physiological differences between these organisms may lead to niche separation of AOA and AOB [24]. Therefore, environmental factors are important in determining the different nitrification activities and relative contributions of AOA and AOB. Among all environmental factors, ammonia is the substrate of ammonia oxidation for which concentrations will directly affect nitrification activity. It was found that AOA were able to grow well and the growth of AOA was coupled with soil nitrification when the concentration of ammonia was relatively low or the supply of ammonia was through the mineralization of organic matter; however, AOB were more competitive in soil nitrification and the number of AOB amoA gene copies was greater than that of AOA PLOS ONE | www.plosone.org 1 August 2012 | Volume 7 | Issue 8 | e44122