Abstract
Ammonia oxidizers are key players in the global nitrogen cycle, yet little is known about their ecological performances and adaptation strategies for growth in saline terrestrial ecosystems. This study combined 13C-DNA stable-isotope probing (SIP) microcosms with amplicon and shotgun sequencing to reveal the composition and genomic adaptations of active ammonia oxidizers in a saline-sodic (solonetz) soil with high salinity and pH (20.9 cmolc exchangeable Na+ kg−1 soil and pH 9.64). Both ammonia-oxidizing archaea (AOA) and bacteria (AOB) exhibited strong nitrification activities, although AOB performed most of the ammonia oxidation observed in the solonetz soil and in the farmland soil converted from solonetz soil. Members of the Nitrosococcus, which are more often associated with aquatic habitats, were identified as the dominant ammonia oxidizers in the solonetz soil with the first direct labeling evidence, while members of the Nitrosospira were the dominant ammonia oxidizers in the farmland soil, which had much lower salinity and pH. Metagenomic analysis of “Candidatus Nitrosococcus sp. Sol14”, a new species within the Nitrosococcus lineage, revealed multiple genomic adaptations predicted to facilitate osmotic and pH homeostasis in this extreme habitat, including direct Na+ extrusion/H+ import and the ability to increase intracellular osmotic pressure by accumulating compatible solutes. Comparative genomic analysis revealed that variation in salt-tolerance mechanisms was the primary driver for the niche differentiation of ammonia oxidizers in saline-sodic soils. These results demonstrate how ammonia oxidizers can adapt to saline-sodic soil with excessive Na+ content and provide new insights on the nitrogen cycle in extreme terrestrial ecosystems.
Similar content being viewed by others
Log in or create a free account to read this content
Gain free access to this article, as well as selected content from this journal and more on nature.com
or
Data availability
Raw 16S rRNA and amoA gene sequences were deposited in the NCBI Sequence Read Archive (SRA) database under the BioProject ID PRJNA641227. Metagenomics and metagenome-assembled genomes (MAGs) data are available at MG-RAST under the study names DAAN_WGS and DAAN_MAGs, respectively.
References
Kuypers MMM, Marchant HK, Kartal B. The microbial nitrogen-cycling network. Nat Rev Microbiol. 2018;16:263–76.
Stein LY, Klotz MG. The nitrogen cycle. Curr Biol. 2016;26:R94–R98.
Erguder TH, Boon N, Wittebolle L, Marzorati M, Verstraete W. Environmental factors shaping the ecological niches of ammonia-oxidizing archaea. FEMS Microbiol Rev. 2009;33:855–69.
Nicol GW, Leininger S, Schleper C, Prosser JI. The influence of soil pH on the diversity, abundance and transcriptional activity of ammonia oxidizing archaea and bacteria. Environ Microbiol. 2008;10:2966–78.
Lehtovirta-Morley LE, Ge C, Ross J, Yao H, Nicol GW, Prosser JI. Characterisation of terrestrial acidophilic archaeal ammonia oxidisers and their inhibition and stimulation by organic compounds. FEMS Microbiol Ecol. 2014;89:542–52.
Lehtovirta-Morley LE, Stoecker K, Vilcinskas A, Prosser JI, Nicol GW. Cultivation of an obligate acidophilic ammonia oxidizer from a nitrifying acid soil. Proc Natl Acad Sci USA. 2011;108:15892–7.
Hayatsu M, Tago K, Uchiyama I, Toyoda A, Wang Y, Shimomura Y, et al. An acid-tolerant ammonia-oxidizing γ-proteobacterium from soil. ISME J. 2017;11:1130–41.
Prosser JI, Nicol GW. Archaeal and bacterial ammonia-oxidisers in soil: the quest for niche specialisation and differentiation. Trends Microbiol. 2012;20:523–31.
Gubry-Rangin C, Hai B, Quince C, Engel M, Thomson BC, James P, et al. Niche specialization of terrestrial archaeal ammonia oxidizers. Proc Natl Acad Sci USA. 2011;108:21206–11.
Aigle A, Prosser JI, Gubry-Rangin C. The application of high-throughput sequencing technology to analysis of amoA phylogeny and environmental niche specialisation of terrestrial bacterial ammonia-oxidisers. Environ Microbiome. 2019;14:3.
Antony CP, Kumaresan D, Hunger S, Drake HL, Murrell JC, Shouche YS. Microbiology of Lonar Lake and other soda lakes. ISME J. 2013;7:468–76.
Montanarella L, Chude V, Yagi K, Krasilnikov P, Panah SKA, Mendonca-Santos MDL, et al. Status of the World’s Soil Resources (SWSR) - Main Report. 2015.
Vera-Gargallo B, Chowdhury TR, Brown J, Fansler SJ, Durán-Viseras A, Sánchez-Porro C, et al. Spatial distribution of prokaryotic communities in hypersaline soils. Sci Rep. 2019;9:1769.
Hollister EB, Engledow AS, Hammett AJM, Provin TL, Wilkinson HH, Gentry TJ. Shifts in microbial community structure along an ecological gradient of hypersaline soils and sediments. ISME J. 2010;4:829–938.
Metternicht GI, Zinck JA. Remote sensing of soil salinity: potentials and constraints. Remote Sens Environ. 2003;85:1–20.
Shi YL, Liu XR, Zhang QW. Effects of combined biochar and organic fertilizer on nitrous oxide fluxes and the related nitrifier and denitrifier communities in a saline-alkali soil. Sci Total Environ. 2019;686:199–211.
Konneke M, Bernhard AE, de la Torre JR, Walker CB, Waterbury JB, Stahl DA. Isolation of an autotrophic ammonia-oxidizing marine archaeon. Nature. 2005;437:543–6.
Bayer B, Vojvoda J, Offre P, Alves RJE, Elisabeth NH, Garcia JAL, et al. Physiological and genomic characterization of two novel marine thaumarchaeal strains indicates niche differentiation. ISME J. 2016;10:1051–63.
Santoro AE, Dupont CL, Richter RA, Craig MT, Carini P, McIlvin MR, et al. Genomic and proteomic characterization of “Candidatus Nitrosopelagicus brevis”: An ammonia-oxidizing archaeon from the open ocean. Proc Natl Acad Sci USA. 2015;112:1173–8.
Pan KL, Gao JF, Li DC, Fan XY. The dominance of non-halophilic archaea in autotrophic ammonia oxidation of activated sludge under salt stress: a DNA-based stable isotope probing study. Bioresour Technol. 2019;291:8.
Nejidat A. Nitrification and occurrence of salt-tolerant nitrifying bacteria in the Negev desert soils. FEMS Microbiol Ecol. 2005;52:21–29.
Ward BB, O’Mullan GD. Worldwide distribution of Nitrosococcus oceani, a marine ammonia-oxidizing gamma-proteobacterium, detected by PCR and sequencing of 16S rRNA and amoA genes. Appl Environ Micro. 2002;68:4153–7.
Koops HP, Böttcher B, Möller UC, Pommerening-Röser A, Stehr G. Description of a new species of Nitrosococcus. Arch Microbiol. 1990;154:244–8.
Fumasoli A, Bürgmann H, Weissbrodt DG, Wells GF, Beck K, Mohn J, et al. Growth of Nitrosococcus-related ammonia oxidizing bacteria coincides with extremely low pH values in wastewater with high ammonia content. Environ Sci Technol. 2017;51:6857–66.
Olivera NL, Prieto L, Bertiller MB, Ferrero MA. Sheep grazing and soil bacterial diversity in shrublands of the Patagonian Monte, Argentina. J Arid Environ. 2016;125:16–20.
Pérez-Hernandez V, Hernandez-Guzman M, Serrano-Silva N, Luna-Guido M, Navarro-Noya YE, Montes-Molina JA, et al. Diversity of amoA and pmoA genes in extremely saline alkaline soils of the former lake Texcoco. Geomicrobiol J. 2020;37:785–97.
Picone N, Pol A, Mesman R, van Kessel MAHJ, Cremers G, van Gelder AH. et al. Ammonia oxidation at pH 2.5 by a new gammaproteobacterial ammonia-oxidizing bacterium. ISME J. 2020;15:1150–64.
Pan H, Liu HY, Liu YW, Zhang QC, Luo Y, Liu XM, et al. Understanding the relationships between grazing intensity and the distribution of nitrifying communities in grassland soils. Sci Total Environ. 2018;634:1157–64.
Santos JP, Mendes D, Monteiro M, Ribeiro H, Baptista MS, Borges MT, et al. Salinity impact on ammonia oxidizers activity and amoA expression in estuarine sediments. Estuar Coast Shelf Sci. 2018;211:177–87.
Ye L, Zhang T. Ammonia-oxidizing bacteria dominates over ammonia-oxidizing archaea in a saline nitrification reactor under low DO and high nitrogen loading. Biotechnol Bioeng. 2011;108:2544–52.
Luo S, Wang S, Tian L, Shi S, Xu S, Yang F, et al. Aggregate-related changes in soil microbial communities under different ameliorant applications in saline-sodic soils. Geoderma. 2018;329:108–17.
Wang WJ, He HS, Zu YG, Guan Y, Liu ZG, Zhang ZH, et al. Addition of HPMA affects seed germination, plant growth and properties of heavy saline-alkali soil in northeastern China: comparison with other agents and determination of the mechanism. Plant Soil. 2011;339:177–91.
Xia W, Zhang C, Zeng X, Feng Y, Jia Z. Autotrophic growth of nitrifying community in an agricultural soil. ISME J. 2011;5:1226–36.
Francis CA, Roberts KJ, Beman JM, Santoro AE, Oakley BB. Ubiquity and diversity of ammonia-oxidizing archaea in water columns and sediments of the ocean. Proc Natl Acad Sci USA. 2005;102:14683–8.
Holmes AJ, Costello A, Lidstrom ME, Murrell JC. Evidence that participate methane monooxygenase and ammonia monooxygenase may be evolutionarily related. FEMS Microbiol Lett. 1995;132:203–8.
Fowler SJ, Palomo A, Dechesne A, Mines PD, Smets BF. Comammox Nitrospira are abundant ammonia oxidizers in diverse groundwater-fed rapid sand filter communities. Environ Microbiol. 2018;20:1002–15.
Zhao ZR, Huang GH, He SS, Zhou N, Wang MY, Dang CY, et al. Abundance and community composition of comammox bacteria in different ecosystems by a universal primer set. Sci Total Environ. 2019;691:145–55.
Alves RJE, Minh BQ, Urich T, von Haeseler A, Schleper C. Unifying the global phylogeny and environmental distribution of ammonia-oxidising archaea based on amoA genes. Nat Commun. 2018;9:1517.
Richter M, Rossello-Mora R. Shifting the genomic gold standard for the prokaryotic species definition. Proc Natl Acad Sci USA. 2009;106:19126–31.
Konstantinidis KT, Rosselló-Móra R, Amann R. Uncultivated microbes in need of their own taxonomy. ISME J. 2017;11:2399–406.
Luo C, Rodriguez-R LM, Konstantinidis KT. MyTaxa: an advanced taxonomic classifier for genomic and metagenomic sequences. Nucleic Acids Res. 2014;42:e73.
Chaumeil P-A, Mussig AJ, Hugenholtz P, Parks DH. GTDB-Tk: a toolkit to classify genomes with the Genome Taxonomy Database. Bioinformatics. 2020;36:1925–7.
Kuroda T, Mizushima T, Tsuchiya T. Physiological roles of three Na+/H+ antiporters in the halophilic bacterium Vibrio parahaemolyticus. Microbiol Immunol. 2005;49:711–9.
Daebeler A, Kitzinger K, Koch H, Herbold CW, Steinfeder M, Schwarz J, et al. Exploring the upper pH limits of nitrite oxidation: diversity, ecophysiology, and adaptive traits of haloalkalitolerant. Nitrospira ISME J. 2020;14:2967–79.
Padan E, Venturi M, Gerchman Y, Dover N. Na+/H+ antiporters. Biochim Biophys Acta. 2001;1505:144–57.
Kraegeloh A, Amendt B, Kunte HJ. Potassium transport in a halophilic member of the bacteria domain: identification and characterization of the K+ uptake systems TrkH and TrkI from Halomonas elongata DSM 2581T. J Bacteriol. 2005;187:1036–43.
Becker EA, Seitzer PM, Tritt A, Larsen D, Krusor M, Yao AI, et al. Phylogenetically driven sequencing of extremely halophilic archaea reveals strategies for static and dynamic osmo-response. PloS Genet. 2014;10:e1004784.
Cardoso FS, Castro RF, Borges N, Santos H. Biochemical and genetic characterization of the pathways for trehalose metabolism in Propionibacterium freudenreichii, and their role in stress response. Microbiology. 2007;153:270–80.
Sadeghi A, Soltani BM, Nekouei MK, Jouzani GS, Mirzaei HH, Sadeghizadeh M. Diversity of the ectoines biosynthesis genes in the salt tolerant Streptomyces and evidence for inductive effect of ectoines on their accumulation. Microbiol Res. 2014;169:699–708.
Ngugi DK, Blom J, Alam I, Rashid M, Ba-Alawi W, Zhang G, et al. Comparative genomics reveals adaptations of a halotolerant thaumarchaeon in the interfaces of brine pools in the Red Sea. ISME J. 2015;9:396–411.
Spang A, Poehlein A, Offre P, Zumbragel S, Haider S, Rychlik N, et al. The genome of the ammonia-oxidizing Candidatus Nitrososphaera gargensis: insights into metabolic versatility and environmental adaptations. Environ Microbiol. 2012;14:3122–45.
Glover HE. The relationship between inorganic nitrogen oxidation and organic carbon production in batch and chemostat cultures of marine nitrifying bacteria. Arch Microbiol. 1985;142:45–50.
Lehtovirta-Morley LE, Ross J, Hink L, Weber EB, Gubry-Rangin C, Thion C, et al. Isolation of ‘Candidatus Nitrosocosmicus franklandus’, a novel ureolytic soil archaeal ammonia oxidiser with tolerance to high ammonia concentration. FEMS Microbiol Ecol. 2016;92:fiw057.
Kits KD, Sedlacek CJ, Lebedeva EV, Han P, Bulaev A, Pjevac P, et al. Kinetic analysis of a complete nitrifier reveals an oligotrophic lifestyle. Nature. 2017;549:269–72.
Hink L, Gubry-Rangin C, Nicol GW, Prosser JI. The consequences of niche and physiological differentiation of archaeal and bacterial ammonia oxidisers for nitrous oxide emissions. ISME J. 2018;12:1084–93.
Shen JP, Zhang LM, Zhu YG, Zhang JB, He JZ. Abundance and composition of ammonia-oxidizing bacteria and ammonia-oxidizing archaea communities of an alkaline sandy loam. Environ Microbiol. 2008;10:1601–11.
Jia Z, Conrad R. Bacteria rather than archaea dominate microbial ammonia oxidation in an agricultural soil. Environ Microbiol. 2009;11:1658–71.
Millero FJ, Feistel R, Wright DG, McDougall TJ. The composition of Standard Seawater and the definition of the Reference-Composition Salinity Scale. Deep-Sea Res Part I-Oceanogr Res Pap. 2008;55:50–72.
Mosier AC, Allen EE, Kim M, Ferriera S, Francis CA. Genome sequence of “Candidatus Nitrosopumilus salaria” BD31, an ammonia-oxidizing archaeon from the San Francisco bay estuary. J Bacteriol. 2012;194:2121–2.
Matsutani N, Nakagawa T, Nakamura K, Takahashi R, Yoshihara K, Tokuyama T. Enrichment of a novel marine ammonia-oxidizing archaeon obtained from sand of an eelgrass zone. Microbes Environ. 2011;26:23–29.
Park BJ, Park SJ, Yoon DN, Schouten S, Damste JSS, Rhee SK. Cultivation of autotrophic ammonia-oxidizing archaea from marine sediments in coculture with sulfur-oxidizing bacteria. Appl Environ Micro. 2010;76:7575–87.
Parada AE, Fuhrman JA. Marine archaeal dynamics and interactions with the microbial community over 5 years from surface to seafloor. ISME J. 2017;11:2510–25.
Wu YJ, Whang LM, Fukushima T, Chang SH. Responses of ammonia-oxidizing archaeal and betaproteobacterial populations to wastewater salinity in a full-scale municipal wastewater treatment plant. J Biosci Bioeng. 2013;115:424–32.
Cardarelli EL, Bargar JR, Francis CA. Diverse Thaumarchaeota dominate subsurface ammonia-oxidizing communities in semi-arid floodplains in the western United States. Micro Ecol. 2020;80:778–92.
Wang HT, Gilbert JA, Zhu YG, Yang XR. Salinity is a key factor driving the nitrogen cycling in the mangrove sediment. Sci Total Environ. 2018;631-2:1342–9.
Oren A. Thermodynamic limits to microbial life at high salt concentrations. Environ Microbiol. 2011;13:1908–23.
Ito M, Guffanti AA, Oudega B, Krulwich TA. mrp, a multigene, multifunctional locus in Bacillus subtilis with roles in resistance to cholate and to Na+ and in pH homeostasis. J Bacteriol. 1999;181:2394–402.
Krulwich TA, Sachs G, Padan E. Molecular aspects of bacterial pH sensing and homeostasis. Nat Rev Microbiol. 2011;9:330–43.
Swartz TH, Ikewada S, Ishikawa O, Ito M, Krulwich TA. The Mrp system: a giant among monovalent cation/proton antiporters? Extremophiles. 2005;9:345–54.
Oren A. Bioenergetic aspects of halophilism. Microbiol Mol Biol R. 1999;63:334–48.
Mackay MA, Norton RS, Borowitzka LJ. Organic osmoregulatory solutes in Cyanobacteria. J Gen Microbiol. 1984;130:2177–91.
Sadler M, McAninch M, Alico R, Hochstein LI. The intracellular Na+ and K+ composition of the moderately halophilic bacterium, Paracoccus halodenitrificans. Can J Microbiol. 1980;26:496–502.
Brown AD. Compatible solutes and extreme water stress in eukaryotic micro-organisms. Adv Micro Physiol. 1978;17:181–243.
Reed RH, Warr SRC, Richardson DL, Moore DJ, Stewart WDP. Multiphasic osmotic adjustment in a euryhaline cyanobacterium. FEMS Microbiol Lett. 1985;28:225–9.
Welsh DT, Herbert RA. Osmoadaptation of Thiocapsa roseopersicina OP-1 in batch and continuous culture: Accumulation of K+ and sucrose in response to osmotic stress. FEMS Microbiol Ecol. 1993;13:151–7.
Sauvage D, Hamelin J, Larher F. Glycine betaine and other structurally related compounds improve the salt tolerance of Rhizobium meliloti. Plant Sci Lett. 1983;31:291–302.
Campbell MA, Chain PSG, Dang H, Sheikh EI, Norton AF, Ward JM, et al. MG. Nitrosococcus watsonii sp. nov., a new species of marine obligate ammonia-oxidizing bacteria that is not omnipresent in the world’s oceans: calls tovalidate the names’Nitrosococcus halophilus’ and ‘Nitrosomonas mobilis’. FEMS Microbiol Ecol. 2011;76:39–48.
Arguelles JC. Physiological roles of trehalose in bacteria and yeasts: a comparative analysis. Arch Microbiol. 2000;174:217–24.
Widderich N, Czech L, Elling FJ, Könneke M, Stöveken N, Pittelkow M, et al. Strangers in the archaeal world: osmostress-responsive biosynthesis of ectoine and hydroxyectoine by the marine thaumarchaeon Nitrosopumilus maritimus. Environ Microbiol. 2016;18:1227–48.
Bursy J, Pierik AJ, Pica N, Bremer E. Osmotically induced synthesis of the compatible solute hydroxyectoine is mediated by an evolutionarily conserved ectoine hydroxylase. J Biol Chem. 2007;282:31147–55.
Kol S, Merlo ME, Scheltema RA, de Vries M, Vonk RJ, Kikkert NA, et al. Metabolomic characterization of the salt stress response in Streptomyces coelicolor. Appl Environ Micro. 2010;76:2574–81.
Csonka LN. Physiological and genetic responses of bacteria to osmotic stress. Microbiol Rev. 1989;53:121–47.
Saum SH, Sydow JF, Palm P, Pfeiffer F, Oesterhelt D, Muller V. Biochemical and molecular characterization of the biosynthesis of glutamine and glutamate, two major compatible solutes in the moderately halophilic bacterium Halobacillus halophilus. J Bacteriol. 2006;188:6808–15.
Ventosa A, Nieto JJ, Oren A. Biology of moderately halophilic aerobic bacteria. Microbiol Mol Biol R. 1998;62:504–44.
Mahan MJ, Csonka LN. Genetic analysis of the proBA genes of Salmonella typhimurium: physical and genetic analysis of the cloned proB+A+ genes of Escherichia coli and of a mutant allele that confers proline overproduction and enhanced osmotolerance. J Bacteriol. 1983;156:1249–62.
Empadinhas N, Pereira PJB, Albuquerque L, Costa J, Sa-Moura B, Marques AT, et al. Functional and structural characterization of a novel mannosyl-3-phosphoglycerate synthase from Rubrobacter xylanophilus reveals its dual substrate specificity. Mol Microbiol. 2011;79:76–93.
Santos H, da Costa MS. Compatible solutes of organisms that live in hot saline environments. Environ Microbiol. 2002;4:501–9.
Koops HP, Purkhold U, Pommerening-Röser A, Timmermann G, Wagner M. The Lithoautotrophic Ammonia-Oxidizing Bacteria. In: Dworkin M, Falkow S, Rosenberg E, Schleifer KH, Stackebrandt E (eds). The Prokaryotes: a Handbook on the Biology of Bacteria, 3rd edn. New York, USA: Springer Science+Business Media; 2006, pp 778–811.
Acknowledgements
We thank Dr. Wei Gao for the help on soil sample collection. We are also grateful to Profs. Brett Baker and Yuanfeng Cai for discussion on metagenomics analysis. We thank Dr. Yong-Xin Liu and the WeChat subscription ID “meta-genome” for some metagenomic analysis methods. We also gratefully acknowledge the helpful comments and careful corrections from Prof. Graeme Nicol, and the technical support from Rong Huang, Zhiying Guo, Yufang Sun, Deling Sun and Ruhai Wang of the Analytical Center of the Institute of Soil Science. This study was supported by the National Natural Science Foundation of China (41530857), the Strategic Priority Research Program of Chinese Academy of Sciences (XDA28020203) and the Key Deployment Project of the Chinese Academy of Sciences (KFZD-SW-112).
Author information
Authors and Affiliations
Contributions
XS and ZJ designed the study. XS performed the experiments and analyzed the data. JZ, XZ, and WX helped data mining. QB constructed the metagenome-assembled genomes. BZ helped acquire soil data. ZJ and J-BZ supervised the project and approved the final version. XS wrote the manuscript with input from all authors.
Corresponding author
Ethics declarations
Competing interests
The authors declare no competing interest.
Additional information
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary information
Rights and permissions
About this article
Cite this article
Sun, X., Zhao, J., Zhou, X. et al. Salt tolerance-based niche differentiation of soil ammonia oxidizers. ISME J 16, 412–422 (2022). https://doi.org/10.1038/s41396-021-01079-6
Received:
Revised:
Accepted:
Published:
Version of record:
Issue date:
DOI: https://doi.org/10.1038/s41396-021-01079-6
This article is cited by
-
Contrasting responses of microbial communities in nonsaline and saline aquatic ecosystems: Insights into biogeography, co-occurrence patterns, and assembly processes
Aquatic Sciences (2026)
-
Unveiling soil microbial dynamics: insights into bacterial community responses to solonetz dealkalinization
Journal of Soils and Sediments (2025)
-
Achieving stable partial nitrification through synergistic inhibition of free ammonia and salinity on nitrite-oxidizing bacteria
Frontiers of Environmental Science & Engineering (2025)
-
Nitrifying niche in estuaries is expanded by the plastisphere
Nature Communications (2024)
-
15N-DNA stable isotope probing reveals niche differentiation of ammonia oxidizers in paddy soils
Applied Microbiology and Biotechnology (2024)
