Systematic and Applied Microbiology 36 (2013) 359–367 Contents lists available at SciVerse ScienceDirect Systematic and Applied Microbiology jo ur nal ho mepage: www.elsevier.de/syapm Community structure and population dynamics of ammonia oxidizers in composting processes of ammonia-rich livestock waste  Takeshi Yamada a,b , Shinya Araki a , Wakako Ikeda-Ohtsubo a , Keiko Okamura c , Akira Hiraishi b,c , Hideyo Ueda d , Yasuichi Ueda d , Keisuke Miyauchi a , Ginro Endo a,∗ a Faculty of Engineering, Tohoku Gakuin University, Tagajo, Miyagi 983-8537, Japan b Department of Environmental and Life Sciences, Toyohashi University of Technology, Toyohashi, Aichi 441-8580, Japan c Electronic-Inspired Interdisciplinary Research Institute, Toyohashi University of Technology, Toyohashi, Aichi 441-8580, Japan d Institute of Hyperthermophiles, Japan Life Center Co., Nakizin, Okinawa 905-0212, Japan a r t i c l e i n f o Article history: Received 12 October 2012 Received in revised form 10 February 2013 Accepted 10 February 2013 Keywords: Livestock manure Compost Ammonia oxidation Ammonia-oxidizing bacteria amoA a b s t r a c t This study investigated the relationship between the population dynamics of ammonia-oxidizing bacte- ria (AOB) and archaea (AOA), and changes in the concentrations of nitrogenous compounds during ammonia-rich livestock waste-composting processes. The data showed that ammonia in beef and dairy cow livestock waste-composting piles was slowly oxidized to nitrite and nitrate after approximately 21–35 days under thermophilic or moderately thermophilic and mesophilic conditions. Real-time quan- titative PCR (qPCR) assays showed a relative abundance of betaproteobacterial AOB during ammonia oxidation but did not detect AOA in any composting stage. Furthermore, real-time qPCR and terminal- restriction fragment length polymorphism (T-RFLP) analyses for the AOB in two composting processes (beef and dairy cow livestock waste) out of the three studied found that thermophilic or moderately ther- mophilic uncultured betaproteobacterial AOB from the “compost AOB cluster” contributed to ammonia oxidation during hot composting stages. Non-metric multidimensional scaling analyses of the data from T-RFLP showed that only a few analogous species predominated during composting of beef, dairy cow and pig livestock wastes, and thus, the AOB community structures in the three composting piles operating under different conditions were similar. AOB-targeted clone library analyses revealed that uncultured members of the “compost AOB cluster”, which could be clearly distinguished from the authentic species of the genus Nitrosomonas, were the major constituents of the AOB populations. These results suggested that a limited and unique species of AOB played a role in ammonia oxidation during the composting of ammonia-rich livestock waste. © 2013 Elsevier GmbH. All rights reserved. Introduction Composting can be used to convert livestock farming waste into nutrient-rich biofertilizers that are effectively accessible to plants. Fresh livestock waste contains a large amount of organic nitroge- nous compounds (e.g. proteins, nucleic acid, and amino acids), which are easily mineralized to produce ammonia via hydrolysis of urea, proteins and nucleic acids, and/or deamination of unincorpo- rated nitrogenous organic compounds in the composting process. Ammonia produced in composting piles tends to change the pH  All the bacterial amoA and 16S rRNA gene sequences obtained in this study were deposited in the DDBJ/EMBL/GenBank databases (accession numbers: AB713935 to AB713951). In addition, two archaeal amoA gene sequences from the clones used as standards in quantitative real-time PCR assays were deposited in the DDBJ/EMBL/GenBank databases (accession numbers: AB720058 to AB720059). ∗ Corresponding author. Tel.: +81 22 368 7493; fax: +81 22 368 7070. E-mail address: gendo@tjcc.tohoku-gakuin.ac.jp (G. Endo). to alkaline, and is easily vaporized as free ammonia (NH 3 ) during the high-temperature stage (>70 ◦ C) and under high pH conditions (Fukumoto et al. 2003; Osada and Fukumoto 2001). Once NH 3 is released into the environment, it can be dissolved in rainwater and can cause eutrophication in closed water areas (Schulze et al. 1989). Several technologies have been used to diminish NH 3 emission dur- ing composting processes, including biofiltration (Jun and Wenfeng 2009; Yasuda et al. 2009), the use of chemical scrubbers (Miller and Macauley 1988), and hyperthermophilic pre-treatment (Yamada et al. 2007, 2008). Residual ammonia in composting piles is utilized in a microbial nitrification–denitrification process, such as assimila- tion, or for biomass formation in the composting processes. Chemolithotrophic nitrification consists of oxidation of ammonia to nitrite and the subsequent oxidation of nitrite to nitrate, and its efficient progress enhances the production of good compost fertilizer with well-balanced nitrogenous compounds. However, incomplete nitrification leads to an imbalance of nitrogenous com- pounds in compost fertilizer, and N 2 O is generated as a by-product 0723-2020/$ – see front matter © 2013 Elsevier GmbH. All rights reserved. http://dx.doi.org/10.1016/j.syapm.2013.02.001