Underlying mechanisms of ANAMMOX bacteria adaptation to salinity stress

  • Han Wang
  • Han-Xiang Li
  • Fang FangEmail author
  • Jin-song GuoEmail author
  • You-Peng Chen
  • Pen Yan
  • Ji-Xiang Yang
Environmental Microbiology - Original Paper


Dealing with nitrogen-rich saline wastewater produced by industries remains challenging because of the inhibition of functional microorganisms by high salinity. The underlying mechanisms of anaerobic ammonium-oxidizing bacteria (AnAOB) exposed to salinity stress should be studied to investigate the potential of anaerobic ammonium oxidation (ANAMMOX) for applications in such wastewater. In this study, the total DNA from granular sludge was extracted from an expanded granular sludge bed (EGSB) reactor operated at 0, 15 and 30 g/L salinity and subjected to high-throughput sequencing. The nitrogen removal performance in the reactor could be maintained from 86.2 to 88.0% at less than 30 g/L salinity level. The microbial diversity in the reactor under saline conditions was lower than that under the salt-free condition. Three genera of AnAOB were detected in the reactor, and Candidatus Kuenenia was the most abundant. The predictive functional profiling based on the Clusters of Orthologous Groups of proteins (COGs) database showed that the inhibition of AnAOB under saline conditions was mainly characterised by the weakening of energy metabolism and intracellular repair. AnAOB might adapt to salinity stress by increasing their rigidity and intracellular osmotic pressure. The predictive functional profiling based on the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway database revealed that the inhibition of AnAOB was mainly manifested by the weakening of intracellular carbohydrate and lipid metabolism, the blockage of intracellular energy supply and the reduction of membrane transport capacity. AnAOB might adapt to salinity stress by strengthening wall/membrane synthesis, essential cofactors (porphyrins) and energy productivity, enhancing intracellular material transformation and gene repair and changing its structure and group behaviour. The stability of the nitrogen removal performance could be maintained via the adaptation of AnAOB to salinity and their increased abundance.


Mechanisms ANAMMOX bacteria (AnAOB) High-throughput sequencing Predictive functional profiling Salinity stress 



This study was funded by the National Natural Science Foundation of China (Grant nos. 51878091 and 21876016).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no competing interests.

Research involving human and animal participants

This article does not involve any studies with human participants or animals performed by any of the authors.

Supplementary material

10295_2019_2137_MOESM1_ESM.doc (1.6 mb)
Supplementary material 1 (DOC 1677 kb)


  1. 1.
    Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P (2002) Molecular biology of the cell, 4th edn. Garland Science, New YorkGoogle Scholar
  2. 2.
    APHA, AWWA, WEF (1995) Standard methods for the examination of water and wastewater, 19th edn. American Public Health Association, Washington, DCGoogle Scholar
  3. 3.
    Cao S, Du R, Li B, Ren N, Peng Y (2016) High-throughput profiling of microbial community structures in an ANAMMOX–UASB reactor treating high-strength wastewater. Appl Microbiol Biot 100:6457–6467. CrossRefGoogle Scholar
  4. 4.
    Da Costa MS, Santos H, Galinski EA (1998) An overview of the role and diversity of compatible solutes in Bacteria and Archaea. Biotechnology of extremophiles. Springer, New York, pp 117–153CrossRefGoogle Scholar
  5. 5.
    Dapena-Mora A, Campos J, Mosquera-Corral A, Méndez R (2006) Anammox process for nitrogen removal from anaerobically digested fish canning effluents. Water Sci Technol 53:265–274. CrossRefGoogle Scholar
  6. 6.
    Diepold A, Armitage JP (2015) Type III secretion systems: the bacterial flagellum and the injectisome. Philos Trans R Soc Lond B Biol Sci 370:20150020. CrossRefGoogle Scholar
  7. 7.
    Epstein W (2003) The roles and regulation of potassium in bacteria. Prog Nucleic Acid Res Mol Biol 75:293–320. CrossRefGoogle Scholar
  8. 8.
    Fan X-Y, Gao J-F, Pan K-L, Li D-C, Dai H-H (2017) Temporal dynamics of bacterial communities and predicted nitrogen metabolism genes in a full-scale wastewater treatment plant. RSC Adv 7:56317–56327. CrossRefGoogle Scholar
  9. 9.
    Fang F, Yang M-M, Wang H, Yan P, Chen Y-P, Guo J-S (2018) Effect of high salinity in wastewater on surface properties of anammox granular sludge. Chemosphere 210:366–375. CrossRefGoogle Scholar
  10. 10.
    Field A (2013) Discovering statistics using IBM SPSS statistics. Sage, New YorkGoogle Scholar
  11. 11.
    Gill SR, Pop M, DeBoy RT, Eckburg PB, Turnbaugh PJ, Samuel BS, Gordon JI, Relman DA, Fraser-Liggett CM, Nelson KE (2006) Metagenomic analysis of the human distal gut microbiome. Science 312:1355–1359. CrossRefGoogle Scholar
  12. 12.
    Giustinianovich EA, Campos J-L, Roeckel MD, Estrada AJ, Mosquera-Corral A, del Río ÁV (2018) Influence of biomass acclimation on the performance of a partial nitritation-anammox reactor treating industrial saline effluents. Chemosphere 194:131–138. CrossRefGoogle Scholar
  13. 13.
    Gonzalez-Silva BM, Rønning AJ, Andreassen IK, Bakke I, Cervantes FJ, Østgaard K, Vadstein O (2017) Changes in the microbial community of an anammox consortium during adaptation to marine conditions revealed by 454 pyrosequencing. Appl Microbiol Biot 101:5149–5162. CrossRefGoogle Scholar
  14. 14.
    Green ER, Mecsas J (2016) Bacterial secretion systems—an overview. Microbiol Spectr. Google Scholar
  15. 15.
    Henderson JF, Paterson ARP (2014) Nucleotide metabolism: an introduction. Academic, CambridgeGoogle Scholar
  16. 16.
    Jacobs K, Lebel L, Buizer J, Addams L, Matson P, McCullough E, Garden P, Saliba G, Finan T (2016) Linking knowledge with action in the pursuit of sustainable water-resources management. Proc Natl Acad Sci USA 113:4591–4596. CrossRefGoogle Scholar
  17. 17.
    Jin R-C, Ma C, Mahmood Q, Yang G-F, Zheng P (2011) Anammox in a UASB reactor treating saline wastewater. Process Saf Environ Prot 89:342–348. CrossRefGoogle Scholar
  18. 18.
    Jin R-C, Yang G-F, Yu J-J, Zheng P (2012) The inhibition of the Anammox process: a review. Chem Eng J 197:67–79. CrossRefGoogle Scholar
  19. 19.
    Kartal B, de Almeida NM, Maalcke WJ, Op den Camp HJ, Jetten MS, Keltjens JT (2013) How to make a living from anaerobic ammonium oxidation. FEMS Microbiol Rev 37:428–461. CrossRefGoogle Scholar
  20. 20.
    Kartal B, Koleva M, Arsov R, van der Star W, Jetten MS, Strous M (2006) Adaptation of a freshwater anammox population to high salinity wastewater. J Biotechnol 126:546–553. CrossRefGoogle Scholar
  21. 21.
    Kartal B, Kuenen JV, Van Loosdrecht M (2010) Sewage treatment with anammox. Science 328:702–703. CrossRefGoogle Scholar
  22. 22.
    Langille MG, Zaneveld J, Caporaso JG, McDonald D, Knights D, Reyes JA, Clemente JC, Burkepile DE, Thurber RLV, Knight R (2013) Predictive functional profiling of microbial communities using 16S rRNA marker gene sequences. Nat Biotechnol 31:814. CrossRefGoogle Scholar
  23. 23.
    Lewinson O, Livnat-Levanon N (2017) Mechanism of action of ABC importers: conservation, divergence, and physiological adaptations. J Mol Biol 429:606–619. CrossRefGoogle Scholar
  24. 24.
    Li Z-X, Zhang L, Chen X-B, Li H, Li D (2015) Acclimation of a highly-efficient and seawater tolerant anammox sludge. J Environ Sci (China) 35:748–756Google Scholar
  25. 25.
    Liu C, Yamamoto T, Nishiyama T, Fujii T, Furukawa K (2009) Effect of salt concentration in anammox treatment using non woven biomass carrier. J Biosci Bioeng 107:519–523. CrossRefGoogle Scholar
  26. 26.
    Ma C, Jin R-C, Yang G-F, Yu J-J, Xing B-S, Zhang Q-Q (2012) Impacts of transient salinity shock loads on Anammox process performance. Bioresour Technol 112:124–130. CrossRefGoogle Scholar
  27. 27.
    Meng Y, Yin C, Zhou Z, Meng F (2018) Increased salinity triggers significant changes in the functional proteins of ANAMMOX bacteria within a biofilm community. Chemosphere 207:655–664. CrossRefGoogle Scholar
  28. 28.
    Nelson DL, Cox MM (2008) Glycolysis, gluconeogenesis, and the pentose phosphate pathway. Lehninger principles of biochemistry, 4th edn. W.H. Freeman, New York, pp 521–559Google Scholar
  29. 29.
    Paredes D, Kuschk P, Mbwette T, Stange F, Müller R, Köser H (2007) New aspects of microbial nitrogen transformations in the context of wastewater treatment—a review. Eng Life Sci 7:13–25. CrossRefGoogle Scholar
  30. 30.
    Parks DH, Tyson GW, Hugenholtz P, Beiko RG (2014) STAMP: statistical analysis of taxonomic and functional profiles. Bioinformatics 30:3123–3124. CrossRefGoogle Scholar
  31. 31.
    Qiao S, Bi Z, Zhou J, Cheng Y, Zhang J (2013) Long term effects of divalent ferrous ion on the activity of anammox biomass. Bioresour Technol 142:490–497. CrossRefGoogle Scholar
  32. 32.
    Rønning AJ (2013) Adaptation of anaerobic ammonium oxidizing (anammox) bacteria to salinity in a continuous reactor. MS thesis. Institutt for bioteknologiGoogle Scholar
  33. 33.
    Sheetz MP, Sable JE, Döbereiner H-G (2006) Continuous membrane-cytoskeleton adhesion requires continuous accommodation to lipid and cytoskeleton dynamics. Annu Rev Biophys Biomol Struct 35:417–434. CrossRefGoogle Scholar
  34. 34.
    Shinohara T, Qiao S, Yamamoto T, Nishiyama T, Fujii T, Kaiho T, Bhatti Z, Furukawa K (2009) Partial nitritation treatment of underground brine waste with high ammonium and salt content. J Biosci Bioeng 108:330–335. CrossRefGoogle Scholar
  35. 35.
    Speth DR, Lagkouvardos I, Wang Y, Qian P-Y, Dutilh BE, Jetten MS (2017) Draft genome of Scalindua rubra, obtained from the interface above the discovery deep brine in the Red Sea, sheds light on potential salt adaptation strategies in anammox bacteria. Microb Ecol 74:1–5. CrossRefGoogle Scholar
  36. 36.
    Tang C-J, Zheng P, Mahmood Q, Chen J-W (2009) Start-up and inhibition analysis of the Anammox process seeded with anaerobic granular sludge. J Ind Microbiol Biotechnol 36:1093. CrossRefGoogle Scholar
  37. 37.
    Uygur A (2006) Specific nutrient removal rates in saline wastewater treatment using sequencing batch reactor. Process Biochem 41:61–66. CrossRefGoogle Scholar
  38. 38.
    Van de Graaf AA, de Bruijn P, Robertson LA, Jetten MS, Kuenen JG (1996) Autotrophic growth of anaerobic ammonium-oxidizing micro-organisms in a fluidized bed reactor. Microbiol 142:2187–2196. CrossRefGoogle Scholar
  39. 39.
    Van de Graaf AA, Mulder A, de Bruijn P, Jetten M, Robertson LA, Kuenen JG (1995) Anaerobic oxidation of ammonium is a biologically mediated process. Appl Environ Microbiol 61:1246–1251Google Scholar
  40. 40.
    Van Der Star WR, Miclea AI, Van Dongen UG, Muyzer G, Picioreanu C, Van Loosdrecht MC (2008) The membrane bioreactor: a novel tool to grow anammox bacteria as free cells. Biotechnol Bioeng 101:286–294. CrossRefGoogle Scholar
  41. 41.
    Van Teeseling MC, Mesman RJ, Kuru E, Espaillat A, Cava F, Brun YV, VanNieuwenhze MS, Kartal B, Van Niftrik L (2015) Anammox Planctomycetes have a peptidoglycan cell wall. Nat Commun 6:6878. CrossRefGoogle Scholar
  42. 42.
    Ventosa A, Nieto JJ, Oren A (1998) Biology of moderately halophilic aerobic bacteria. Microbiol Mol Biol Rev 62:504–544Google Scholar
  43. 43.
    Wadhams GH, Armitage JP (2004) Making sense of it all: bacterial chemotaxis. Nat Rev Mol Cell Biol 5:1024. CrossRefGoogle Scholar
  44. 44.
    Wang K, Ye X, Zhang H, Chen H, Zhang D, Liu L (2016) Regional variations in the diversity and predicted metabolic potential of benthic prokaryotes in coastal northern Zhejiang, East China Sea. Sci Rep 6:38709. CrossRefGoogle Scholar
  45. 45.
    Wei Q, Kawagoshi Y, Huang X, Hong N, Van Duc L, Yamashita Y, Hama T (2016) Nitrogen removal properties in a continuous marine anammox bacteria reactor under rapid and extensive salinity changes. Chemosphere 148:444–451. CrossRefGoogle Scholar
  46. 46.
    Xie W, Wang F, Guo L, Chen Z, Sievert SM, Meng J, Huang G, Li Y, Yan Q, Wu S (2011) Comparative metagenomics of microbial communities inhabiting deep-sea hydrothermal vent chimneys with contrasting chemistries. ISME J 5:414. CrossRefGoogle Scholar
  47. 47.
    Xing H, Wang H, Fang F, Li K, Liu L, Chen Y, Guo J (2017) Effect of increase in salinity on ANAMMOX–UASB reactor stability. Environ Technol 38:1184–1190. CrossRefGoogle Scholar
  48. 48.
    Yang J, Zhang L, Hira D, Fukuzaki Y, Furukawa K (2011) Anammox treatment of high-salinity wastewater at ambient temperature. Bioresour Technol 102:2367–2372. CrossRefGoogle Scholar
  49. 49.
    Zhang Z-Z, Ji Y-X, Cheng Y-F, Jin R-C (2018) Increased salinity improves the thermotolerance of mesophilic anammox consortia. Sci Total Environ 644:710–716. CrossRefGoogle Scholar
  50. 50.
    Zhou E, Trepat X, Park C, Lenormand G, Oliver M, Mijailovich S, Hardin C, Weitz D, Butler J, Fredberg J (2009) Universal behavior of the osmotically compressed cell and its analogy to the colloidal glass transition. Proc Natl Acad Sci USA 106:10632–10637. CrossRefGoogle Scholar

Copyright information

© Society for Industrial Microbiology and Biotechnology 2019

Authors and Affiliations

  1. 1.College of Urban Construction and Environmental EngineeringChongqing UniversityChongqingPeople’s Republic of China
  2. 2.Chongqing Institute of Green and Intelligent TechnologyChinese Academy of SciencesChongqingPeople’s Republic of China

Personalised recommendations