Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

Submerged macrophytes shape the abundance and diversity of bacterial denitrifiers in bacterioplankton and epiphyton in the Shallow Fresh Lake Taihu, China

  • 540 Accesses

  • 5 Citations


nirK and nirS genes are important functional genes involved in the denitrification pathway. Recent studies about these two denitrifying genes are focusing on sediment and wastewater microbe. In this study, we conducted a comparative analysis of the abundance and diversity of denitrifiers in the epiphyton of submerged macrophytes Potamogeton malaianus and Ceratophyllum demersum as well as in bacterioplankton in the shallow fresh lake Taihu, China. Results showed that nirK and nirS genes had significant different niches in epiphyton and bacterioplankton. Bacterioplankton showed greater abundance of nirK gene in terms of copy numbers and lower abundance of nirS gene. Significant difference in the abundance of nirK and nirS genes also existed between the epiphyton from different submerged macrophytes. Similar community diversity yet different community abundance was observed between epiphytic bacteria and bacterioplankton. No apparent seasonal variation was found either in epiphytic bacteria or bacterioplankton; however, environmental parameters seemed to have direct relevancy with nirK and nirS genes. Our study suggested that submerged macrophytes have greater influence than seasonal parameters in shaping the presence and abundance of bacterial denitrifiers. Further investigation needs to focus on the potential contact and relative contribution between denitrifiers and environmental factors.

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5



Dissolved oxygen


Total nitrogen


Total phosphorus

NO3 –N:

Nitrate nitrogen

NO2 –N:

Nitrite nitrogen

NH4 +–N:

Ammoniacal nitrogen


Quantitative PCR


Operational taxonomic units


Principal coordinates analysis


Detrended correspondence analysis


Redundancy analysis


Spearman’s rank correlation coefficient


Dry weight


Ammonia-oxidizing archaea


Ammonia-oxidizing bacteria


  1. Abell GC, Revill AT, Smith C, Bissett AP, Volkman JK, Robert SS (2010) Archaeal ammonia oxidizers and nirS-type denitrifiers dominate sediment nitrifying and denitrifying populations in a subtropical macrotidal estuary. ISME Journal 4:286–300. doi:10.1038/ismej.2009.105

  2. Antonyuk SV, Strange RW, Sawers G, Eady RR, Hasnain SS (2005) Atomic resolution structures of resting-state, substrate- and product-complexed Cu-nitrite reductase provide insight into catalytic mechanism. Proc Natl Acad Sci U S A 102:12041–12046

  3. Attard E et al (2011) Soil environmental conditions rather than denitrifier abundance and diversity drive potential denitrification after changes in land uses. Glob Chang Biol 17:1975–1989

  4. Barta J, Melichova T, Vanek D, Picek T, Santruckova H (2010) Effect of pH and dissolved organic matter on the abundance of nirK and nirS denitrifiers in spruce forest soil. Biogeochemistry 101:123–132

  5. Bowen JL, Babbin AR, Kearns PJ, Ward BB (2014) Connecting the dots: linking nitrogen cycle gene expression to nitrogen fluxes in marine sediment mesocosms. Front Microbiol 5:429. doi:10.3389/fmicb.2014.00429

  6. Braker G, Fesefeldt A, Witzel KP (1998) Development of PCR primer systems for amplification of nitrite reductase genes (nirK and nirS) to detect denitrifying bacteria in environmental samples. Appl Environ Microbiol 64:3769–3775

  7. Braker G, Zhou J, Wu L, Devol AH, Tiedje JM (2000) Nitrite reductase genes (nirK and nirS) as functional markers to investigate diversity of denitrifying bacteria in pacific northwest marine sediment communities. Appl Environ Microbiol 66:2096–2104

  8. Braker G, Schwarz J, Conrad R (2010) Influence of temperature on the composition and activity of denitrifying soil communities. FEMS Microbiol Ecol 73:134–148

  9. Bremer C, Braker G, Matthies D, Reuter A, Engels C, Conrad R (2007) Impact of plant functional group, plant species, and sampling time on the composition of nirK-Type denitrifier communities in soil. Appl Environ Microbiol 73:6876–6884

  10. Bruns A, Nubel U, Cypionka H, Overmann J (2003) Effect of signal compounds and incubation conditions on the culturability of freshwater bacterioplankton. Appl Environ Microbiol 69:1980–1989

  11. Burke C, Thomas T, Lewis M, Steinberg P, Kjelleberg S (2011) Composition, uniqueness and variability of the epiphytic bacterial community of the green alga Ulva australis. ISME Journal 5:590–600. doi:10.1038/ismej.2010.164

  12. Cai X, Gao G, Tang X, Dong B, Dai J, Chen D, Song Y (2013) The response of epiphytic microbes to habitat and growth status of Potamogeton malaianus Miq. in Lake Taihu. J Basic Microbiol 53:828–837

  13. Coci M, Bodelier PLE, Laanbroek HJ (2008) Epiphyton as a niche for ammonia-oxidizing bacteria: detailed comparison with benthic and pelagic compartments in shallow freshwater lakes. Appl Environ Microbiol 74:1963–1971

  14. Coci M, Nicol GW, Pilloni GN, Schmid M, Kamst-van Agterveld MP, Bodelier PLE, Laanbroek HJ (2010) Quantitative assessment of ammonia-oxidizing bacterial communities in the epiphyton of submerged macrophytes in Shallow Lakes. Appl Environ Microbiol 76:1813–1821

  15. Crump BC, Koch EW (2008) Attached bacterial populations shared by four species of aquatic angiosperms. Appl Environ Microbiol 74:5948–5957. doi:10.1128/AEM.00952-08

  16. Dandie CE et al (2011) Abundance, diversity and functional gene expression of denitrifier communities in adjacent riparian and agricultural zones. FEMS Microbiol Ecol 77:69–82

  17. Enwall K, Philippot L, Hallin S (2005) Activity and composition of the denitrifying bacterial community respond differently to long-term fertilization. Appl Environ Microbiol 71:8335–8343. doi:10.1128/AEM.71.12.8335-8343.2005

  18. Enwall K, Throback IN, Stenberg M, Soderstrom M, Hallin S (2010) Soil resources influence spatial patterns of denitrifying communities at scales compatible with land management. Appl Environ Microbiol 76:2243–2250

  19. Eriksson PG (2001) Interaction effects of flow velocity and oxygen metabolism on nitrification and denitrification in biofilms on submersed macrophytes. Biogeochemistry 55:29–44. doi:10.1023/a:1010679306361

  20. Eriksson PG, Weisner SEB (1997) Nitrogen removal in a wastewater reservoir: the importance of denitrification by epiphytic biofilms on submersed vegetation. J Environ Qual 26:905–910

  21. GB11893-1989: Water quality- determination of total phosphorus- ammonium molybdate spectrophotometric method. http://www.mep.gov.cn/image20010518/3655.pdf

  22. Gordon-Bradley N, Lymperopoulou DS, Williams HN (2014) Differences in bacterial community structure on Hydrilla verticillata and Vallisneria americana in a freshwater spring. Microbes Environments JSME 29:67–73

  23. Hai B et al (2009) Quantification of key genes steering the microbial nitrogen cycle in the rhizosphere of sorghum cultivars in tropical agroecosystems. Appl Environ Microbiol 75:4993–5000. doi:10.1128/AEM.02917-08

  24. He D, Ren LJ, Wu QL (2012) Epiphytic bacterial communities on two common submerged macrophytes in Taihu Lake: diversity and host-specificity. Chin J Oceanol Limnol 30:237–247. doi:10.1007/s00343-012-1084-0

  25. He D, Ren LJ, Wu QLL (2014) Contrasting diversity of epibiotic bacteria and surrounding bacterioplankton of a common submerged macrophyte, Potamogeton crispus, in freshwater lakes. FEMS Microbiol Ecol 90:551–562

  26. Hempel M, Blume M, Blindow I, Gross EM (2008) Epiphytic bacterial community composition on two common submerged macrophytes in brackish water and freshwater. BMC Microbiol 8:58. doi:10.1186/1471-2180-8-58

  27. Hempel M, Grossart HP, Gross EM (2009) Community composition of bacterial biofilms on two submerged macrophytes and an artificial substrate in a pre-alpine lake. Aquat Microb Ecol 58:79–94

  28. Hilt S, Gross EM (2008) Can allelopathically active submerged macrophytes stabilise clear-water states in shallow lakes? Basic Appl Ecol 9:422–432

  29. HJ/T199-2005: Water quality- determination of total-nitrogen gas-phase molecular absorption spectrometry. http://kjs.mep.gov.cn/hjbhbz/bzwb/shjbh/sjcgfffbz/200601/W020110127355110870522.pdf

  30. Hou J, Cao X, Song C, Zhou Y (2013) Predominance of ammonia-oxidizing archaea and nirK-gene-bearing denitrifiers among ammonia-oxidizing and denitrifying populations in sediments of a large urban eutrophic lake (Lake Donghu). Can J Microbiol 59:456–464. doi:10.1139/cjm-2013-0083

  31. Jezberova J, Jezbera J, Brandt U, Lindstrom ES, Langenheder S, Hahn MW (2010) Ubiquity of Polynucleobacter necessarius ssp. asymbioticus in lentic freshwater habitats of a heterogeneous 2000 km area. Environ Microbiol 12:658–669. doi:10.1111/j.1462-2920.2009.02106.x

  32. Kasalicky V, Jezbera J, Hahn MW, Simek K (2013) The diversity of the Limnohabitans genus, an important group of freshwater bacterioplankton, by characterization of 35 isolated strains. PLoS One 8:e58209. doi:10.1371/journal.pone.0058209

  33. Körner S (1999) Nitrifying and denitrifying bacteria in epiphytic communities of submerged macrophytes in a treated sewage channel. Acta Hydrochim Hydrobiol 27:27–31

  34. Kraft B, Tegetmeyer HE, Meier D, Geelhoed JS, Strous M (2014) Rapid succession of uncultured marine bacterial and archaeal populations in a denitrifying continuous culture. Environ Microbiol 16:3275–3286. doi:10.1111/1462-2920.12552

  35. Leps J, Smilauer P (2003) Multivariate analysis of ecological data using CANOCO. United States of America by Cambridge University Press, New York

  36. Petersen DG, Blazewicz SJ, Firestone M, Herman DJ, Turetsky M, Waldrop M (2012) Abundance of microbial genes associated with nitrogen cycling as indices of biogeochemical process rates across a vegetation gradient in Alaska. Environ Microbiol 14:993–1008

  37. Philippot L et al (2009) Mapping field-scale spatial patterns of size and activity of the denitrifier community. Environ Microbiol 11:1518–1526

  38. Rich JJ, Myrold DD (2004) Community composition and activities of denitrifying bacteria from adjacent agricultural soil, riparian soil, and creek sediment in Oregon, USA. Soil Biol Biochem 36:1431–1441

  39. Saarenheimo J, Rissanen AJ, Arvola L, Nykanen H, Lehmann MF, Tiirola M (2015) Genetic and environmental controls on nitrous oxide accumulation in lakes. PLoS One 10:e0121201. doi:10.1371/journal.pone.0121201

  40. Santoro AE, Boehm AB, Francis CA (2006) Denitrifier community composition along a nitrate and salinity gradient in a coastal aquifer. Appl Environ Microbiol 72:2102–2109. doi:10.1128/AEM.72.3.2102-2109.2006

  41. Scheffer M, Hosper SH, Meijer ML, Moss B, Jeppesen E (1993) Alternative equilibria in shallow lakes. Trends Ecol Evol 8:275–279. doi:10.1016/0169-5347(93)90254-M

  42. Smith JM, Ogram A (2008) Genetic and functional variation in denitrifier populations along a short-term restoration chronosequence. Appl Environ Microbiol 74:5615–5620. doi:10.1128/AEM.00349-08

  43. Su MX, Kleineidam K, Schloter M (2010) Influence of different litter quality on the abundance of genes involved in nitrification and denitrification after freezing and thawing of an arable soil. Biol Fertil Soils 46:537–541

  44. Szukics U, Abell GCJ, Hodl V, Mitter B, Sessitsch A, Hackl E, Zechmeister-Boltenstern S (2010) Nitrifiers and denitrifiers respond rapidly to changed moisture and increasing temperature in a pristine forest soil. FEMS Microbiol Ecol 72:395–406

  45. Throback IN, Enwall K, Jarvis A, Hallin S (2004) Reassessing PCR primers targeting nirS, nirK and nosZ genes for community surveys of denitrifying bacteria with DGGE. FEMS Microbiol Ecol 49:401–417. doi:10.1016/j.femsec.2004.04.011

  46. Trias R, Garcia-Lledo A, Sanchez N, Lopez-Jurado JL, Hallin S, Baneras L (2012) Abundance and composition of epiphytic bacterial and archaeal ammonia oxidizers of marine red and brown macroalgae. Appl Environ Microbiol 78:318–325. doi:10.1128/aem.05904-11

  47. Tujula NA, Crocetti GR, Burke C, Thomas T, Holmstrom C, Kjelleberg S (2010) Variability and abundance of the epiphytic bacterial community associated with a green marine Ulvacean alga. ISME Journal 4:301–311. doi:10.1038/ismej.2009.107

  48. Vila-Costa M, Bartrons M, Catalan J, Casamayor EO (2014) Nitrogen-cycling genes in epilithic biofilms of oligotrophic high-altitude lakes (central Pyrenees, Spain). Microb Ecol 68:60–69. doi:10.1007/s00248-014-0417-2

  49. Wang Z, Zhang XX, Lu X, Liu B, Li Y, Long C, Li A (2014) Abundance and diversity of bacterial nitrifiers and denitrifiers and their functional genes in tannery wastewater treatment plants revealed by high-throughput sequencing. PLoS One 9:e113603. doi:10.1371/journal.pone.0113603

  50. Wertz S et al (2006) Maintenance of soil functioning following erosion of microbial diversity. Environ Microbiol 8:2162–2169

  51. Wolsing M, Prieme A (2004) Observation of high seasonal variation in community structure of denitrifying bacteria in arable soil receiving artificial fertilizer and cattle manure by determining T-RFLP of nir gene fragments. FEMS Microbiol Ecol 48:261–271. doi:10.1016/j.femsec.2004.02.002

  52. Wu QL, Zwart G, Wu J, Agterveld MPKV, Liu S, Hahn MW (2007a) Submersed macrophytes play a key role in structuring bacterioplankton community composition in the large, shallow, subtropical Taihu Lake, China. Environ Microbiol 9:2765–2774

  53. Wu QL, Zwart G, Wu J, Kamst-van Agterveld MP, Liu S, Hahn MW (2007b) Submersed macrophytes play a key role in structuring bacterioplankton community composition in the large, shallow, subtropical Taihu Lake. China Environ Microbiol 9:2765–2774. doi:10.1111/j.1462-2920.2007.01388.x

  54. You SJ (2005) Identification of denitrifying bacteria diversity in an activated sludge system by using nitrite reductase genes. Biotechnol Lett 27:1477–1482

  55. Yuan Q, Liu P, Lu Y (2012) Differential responses of nirK- and nirS-carrying bacteria to denitrifying conditions in the anoxic rice field soil. Environ Microbiol Rep 4:113–122. doi:10.1111/j.1758-2229.2011.00311.x

  56. Zeng J, Bian YQ, Xing P, Wu QLL (2012) Macrophyte species drive the variation of bacterioplankton community composition in a shallow freshwater lake. Appl Environ Microbiol 78:177–184

  57. Zhang JX, Yang YY, Zhao L, Li YZ, Xie SG, Liu Y (2015) Distribution of sediment bacterial and archaeal communities in plateau freshwater lakes. Appl Microbiol Biotechnol 99:3291–3302

  58. Zumft WG (1997) Cell biology and molecular basis of denitrification. Microbiol Mol Biol Rev 61:533–616

Download references


The authors are grateful to Ms. Ya-peng Zhang and Mr. Xun Cao for their valuable help in field investigation and sampling. This work is financially supported by the National Natural Science Foundation of China (41173078 & 41403064), the Natural Science Foundation of Jiangsu Province, China (BK20140922), and by the Natural Science Foundation of the Jiangsu Higher Education Institutions of China (14KJB610007).

Author information

Correspondence to Guo-xiang Wang.

Additional information

Responsible editor: Robert Duran

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Fan, Z., Han, R., Ma, J. et al. Submerged macrophytes shape the abundance and diversity of bacterial denitrifiers in bacterioplankton and epiphyton in the Shallow Fresh Lake Taihu, China. Environ Sci Pollut Res 23, 14102–14114 (2016). https://doi.org/10.1007/s11356-016-6390-1

Download citation


  • Denitrifier
  • Epiphytic bacteria
  • Bacterioplankton
  • Submerged macrophytes
  • Taihu Lake