Environmental Science and Pollution Research

, Volume 23, Issue 5, pp 4207–4217 | Cite as

Functional and structural responses of soil N-cycling microbial communities to the herbicide mesotrione: a dose-effect microcosm approach

  • Olivier CrouzetEmail author
  • Franck Poly
  • Frédérique Bonnemoy
  • David Bru
  • Isabelle Batisson
  • Jacques Bohatier
  • Laurent Philippot
  • Clarisse Mallet
Research Article


Microbial communities driving the nitrogen cycle contribute to ecosystem services such as crop production and air, soil, and water quality. The responses to herbicide stress of ammonia-oxidizing and ammonia-denitrifying microbial communities were investigated by an analysis of changes in structure-function relationships. Their potential activities, abundances (quantitative PCR), and genetic structure (denaturing gradient gel electrophoresis) were assessed in a microcosm experiment. The application rate (1 × FR, 0.45 μg g−1 soil) of the mesotrione herbicide did not strongly affect soil N-nutrient dynamics or microbial community structure and abundances. Doses of the commercial product Callisto® (10 × FR and 100 × FR) or pure mesotrione (100 × FR) exceeding field rates induced short-term inhibition of nitrification and a lasting stimulation of denitrification. These effects could play a part in the increase in soil ammonium content and decrease in nitrate contents observed in treated soils. These functional impacts were mainly correlated with abundance shifts of ammonia-oxidizing Bacteria (AOB) and Archaea (AOA) or denitrifying bacteria. The sustained restoration of nitrification activity, from day 42 in the 100 × FR-treated soils, was likely promoted by changes in the community size and composition of AOB, which suggests a leading role, rather than AOA, for soil nitrification restoration after herbicide stress. This ecotoxicological community approach provides a nonesuch multiparameter assessment of responses of N-cycling microbial guilds to pesticide stress.


Herbicide Soil microcosms Ammonia oxidizers Denitrifiers Microbial ecotoxicology 



This work was supported by a Fellowship from the French Ministère de l’Education, de la Recherche et de la Technologie. We thank S. Leininger for providing plasmids with bacterial—amoA and archaeal—amoA DNA inserts used for qPCR standards and to G. Borrel for providing the plasmids containing archaeal—16S rDNA genes used for qPCR standards. We would also like to thank J. Watts, English native translator, for the English review of this paper.

Supplementary material

11356_2015_4797_MOESM1_ESM.ppt (904 kb)
Fig. S1 (PPT 904 kb)
11356_2015_4797_MOESM2_ESM.ppt (274 kb)
Table S1 (PPT 274 kb)


  1. Alexander M (2000) Aging, bioavailability, and overestimation of risk from environmental pollutants. Environ Sci Technol 34:4259–4265CrossRefGoogle Scholar
  2. Andersch I, Anderson JPE (1991) Influence of pesticides on nitrogen transformations in soils. Toxicol Environ Chem 30:153–158CrossRefGoogle Scholar
  3. Attard E, Recous S, Chabbi A, De Berranger C, Guillaumaud N, Labreuche J, Philippot L, Schmid B, Le Roux X (2011) Soil environmental conditions rather than denitrifier abundance and diversity drive potential denitrification after changes in land uses. Glob Chang Biol 17:1975–1989CrossRefGoogle Scholar
  4. Baker GC, Smith JJ, Cowan DA (2003) Review and re-analysis of domain-specific 16S primers. J Microbiol Methods 55:541–555CrossRefGoogle Scholar
  5. Beigel C, Charnay MP, Barriuso E (1999) Degradation of formulated and unformulated triticonazole fungicide in soil: effect of application rate. Soil Biol Biochem 31:525–534Google Scholar
  6. Bending GD, Rodriguez-Cruz MS, Lincoln SD (2007) Fungicide impacts on microbial communities in soils with contrasting management histories. Chemosphere 69:82–88CrossRefGoogle Scholar
  7. Bru D, Ramette A, Saby NPA, Dequiedt S, Ranjard L, Jolivet C, Arrouays D, Philippot L (2011) Determinants of the distribution of nitrogen-cycling microbial communities at the landscape scale. ISME J 5:532–542CrossRefGoogle Scholar
  8. Carlisle SM, Trevors JT (1986) Effect of the herbicide glyphosate on nitrification, denitrification and acetylene reduction in soil. Water Air Soil Pollut 29:189–203CrossRefGoogle Scholar
  9. Chaabane H, Vulliet E, Calvayrac C, Coste C-M, Cooper J-F (2008) Behaviour of sulcotrione and mesotrione in two soils. Pest Manag Sci 64:86–93CrossRefGoogle Scholar
  10. Chang Y-J, Akma H, Stephen JR, Mullen MD, White DC, Peacockt A (2001) Impact of herbicides on the abundance and structure of indigenous β-subgroup ammonia-oxidizer communities in soil microcosms. Environ Toxicol Chem 20:2462–2468CrossRefGoogle Scholar
  11. Crouzet O, Batisson I, Besse-Hoggan P, Bonnemoy F, Bardot C, Poly F, Bohatier J, Mallet C (2010) Response of soil microbial communities to the herbicide mesotrione: a dose-effect microcosm approach. Soil Biol Biochem 42:193–202CrossRefGoogle Scholar
  12. Crouzet O, Wiszniowski J, Donnadieu F, Bonnemoy F, Bohatier J, Mallet C (2013) Dose-dependent effects of the herbicide mesotrione on soil cyanobacterial communities. Arch Environ Contam Toxicol 64:23–31CrossRefGoogle Scholar
  13. Dyson JS, Beulke S, Brown CD, Lane MCG (2002) Adsorption and degradation of the weak acid Mesotrione in soil and environmental fate. J Environ Qual 31:613–618CrossRefGoogle Scholar
  14. Erguder TH, Boon N, Wittebolle L, Marzorati M, Verstraete W (2009) Environmental factors shaping the ecological niches of ammonia-oxidizing archaea. FEMS Microbiol Rev 33:855–869CrossRefGoogle Scholar
  15. Fromin N, Hamelin J, Tarnawski S, Roesti D, Jourdain-Miserez K, Forestier N, Teyssier-Cuvelle S, Gillet F, Aragno M, Rossi P (2002) Statistical analysis of denaturing gel electrophoresis (DGE) fingerprinting patterns. Environ Microbiol 4:634–643CrossRefGoogle Scholar
  16. Hammer Ø, Harper DAT, Ryan PD (2001) PAST: paleontological statistics software package for education and data analysis. Palaeontol Electron 4:4–9Google Scholar
  17. Henry S, Bru D, Stress B, Hallet S, Philippot L (2006) Quantitative detection of the nosZ gene, encoding nitrous oxide reductase, and comparison of the abundances of 16S rRNA, narG, nirK, and nosZ genes in soils. Appl Environ Microbiol 72:5181–5189CrossRefGoogle Scholar
  18. Hernandez M, Jia ZJ, Conrad R, Seeger M (2011) Simazine application inhibits nitrification and changes the ammonia-oxidizing bacterial communities in a fertilized agricultural soil. FEMS Microbiol Ecol 78:511–519Google Scholar
  19. Kandeler E, Deiglmayr K, Tscherko D, Bru D, Philippot L (2006) Abundance of narG, nirS, nirK, and nosZ Genes of Denitrifying Bacteria during Primary Successions of a Glacier Foreland. Appl Environ Microb 72: 5957–5962Google Scholar
  20. Kara E, Arli M, Uygur V (2004) Effects of the herbicide Topogard on soil respiration, nitrification, and denitrification in potato-cultivated soils differing in pH. Biol Fertil Soils 39:474–478CrossRefGoogle Scholar
  21. Kowalchuk GA, Stephen JR (2001) Ammonia-oxidizing bacteria: a model for molecular microbial ecology. Annu Rev Microbiol 55:485–529CrossRefGoogle Scholar
  22. Le Roux X, Poly F, Currey P, Commeaux C, Hai B, Nicol GW, Prosser JI, Schloter M, Attard E, Klumpp K (2008) Effects of aboveground grazing on coupling among nitrifier activity, abundance and community structure. ISME J 2:221–232CrossRefGoogle Scholar
  23. Leininger S, Urich T, Schloter M, Schwark L, Qi J, Nicol GW, Prosser JI, Schuster SC, Schleper C (2006) Archaea predominate among ammonia-oxidizing prokaryotes in soils. Nature 442:806–809CrossRefGoogle Scholar
  24. Li X, Zhang H, Wu M, Su Z, Zhang C (2011) Impact of acetochlor on ammonia-oxidizing bacteria in microcosm soils. J Environ Sci 20:1126–1131CrossRefGoogle Scholar
  25. McCarty GW (1999) Modes of action of nitrification inhibitors. Biol Fertil Soils 29:1–9CrossRefGoogle Scholar
  26. MEA (2005) Millennium Ecosystem Assessment. Ecosystems and human well-being, synthesis. Island Press, Washington, DCGoogle Scholar
  27. Mertens J, Broos K, Wakelin SA, Kowalchuck GA, Springael D, Smolders E (2009) Bacteria, not archaea, restore nitrification in a zinc-contaminated soil. ISME J 3:916–923CrossRefGoogle Scholar
  28. Mitchell G, Bartlett DW, Fraser TEM, Hawkes TR, Holt DC, Townson JK, Wichert RA (2001) Mesotrione: a new selective herbicide for use in maize. Pest Manag Sci 57:120–128CrossRefGoogle Scholar
  29. Muñoz-Leoz B, Ruiz-Romera E, Antigüedad I, Garbisu C (2011) Tebuconazole application decreases soil microbial biomass and activity. Soil Biol Biochem 43:2176–83CrossRefGoogle Scholar
  30. Muyzer G, de Waal EC, and Uitterlinden AG (1993) Profiling of complex microbial populations by denaturing gradient gel electrophoresis analysis of polymerase chain reaction-amplified genes coding for 16S rRNA. Appl Environ Microbiol 59:695–700Google Scholar
  31. Ochsenreiter T, Selezi D, Quaiser A, Bonch-Osmolovskaya L, Schleper C (2003) Diversity and abundance of Crenarchaeota in terrestrial habitats studied by 16S RNA surveys and real time PCR. Environ Microbiol 5:787–797CrossRefGoogle Scholar
  32. Pell MB, Stenberg B, Torstensson L (1998) Potential denitrification and nitrification tests for evaluation of pesticide effects in soil. Ambio 27:24–28Google Scholar
  33. Philippot L, Hallin S (2005) Finding the missing link between diversity and activity using denitrifying bacteria as a model functional community. Curr Opin Microbiol 8:234–239CrossRefGoogle Scholar
  34. Philippot L, Cregut M, Chèneby D, Bressan M, Dequiet S, Martin-Laurent F, Ranjard L, Lemanceau P (2008) Effect of primary mild stresses on resilience and resistance of the nitrate reducer community to a subsequent severe stress. FEMS Microbiol Lett 285:51–57CrossRefGoogle Scholar
  35. Prosser JI, Nicol GW (2012) Archaeal and bacterial ammonia oxidisers in soil: the quest for niche specialisation and differentiation. Trends Microbiol 20:523–531CrossRefGoogle Scholar
  36. Puglisi E, Vasileiadis S, Demiris C, Bassi D, Karpouzas DG, Capri E, Cocconceli PS, Trevisan M (2012) Fungicides impact on the diversity and function of non-target ammonia oxidizing microorganisms residing in a litter soil cover. Microb Ecol 64:692–701CrossRefGoogle Scholar
  37. Robinson DE (2008) Atrazine accentuates carryover injury from mesotrione in vegetable crops. Weed Technol 22:641–645CrossRefGoogle Scholar
  38. Rotthauwe J-H, Witzel K-P, Liesack W (1997) The ammonia monooxygenase structural gene amoA as a functional marker: molecular fine-scale analysis of natural ammonia-oxidizing populations. Appl Environ Microbiol 63:4704–4712Google Scholar
  39. Rouchaud J, Neus O, Cools K, Bulcke R (2000) Dissipation of the triketone mesotrione herbicide in the soil of corn crops grown on different soil types. Toxicol Environ Chem 77:31–40CrossRefGoogle Scholar
  40. Ruyters S, Nicol GW, Prosser JI, Lievens B, Smolders E (2013) Activity of the ammonia oxidising bacteria is responsible for zinc tolerance development of the ammonia oxidising community in soil: a stable isotope probing study. Soil Biol Biochem 58:244–247CrossRefGoogle Scholar
  41. Seghers D, Verthé K, Reheul D, Bulcke R, Siciliano SD, Verstraete W, Top EM (2003) Effect of long-term herbicide applications on the bacterial community structure and function in an agricultural soil. FEMS Microbiol Ecol 46:139–146CrossRefGoogle Scholar
  42. Stephen JR, Chang YJ, Macnaughton SJ, Kowalchuk GA, Leung KT, Flemming CA, White DC (1999) Effect of toxic metals on indigenous soil b-subgroup proteobacterium ammonia oxidizer community structure and protection against toxicity by inoculated metal-resistant bacteria. Appl Environ Microb 65:95–101Google Scholar
  43. Stratton GW (1990) Effects of the herbicide glyphosate on nitrification in four soils from Atlantic Canada. Water Air Soil Pollut 51:373–383Google Scholar
  44. Throbäck I, 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 49:401–417CrossRefGoogle Scholar
  45. Tiedje JM, Simkins S, Groffman PM (1989) Perspectives on measurement of denitrification in the field including recommended protocols for acetylene based methods. Plant Soil 115:261–284CrossRefGoogle Scholar
  46. Tourna M, Freitag TE, Nicol GW, Prosser JI (2008) Growth, activity and temperature responses of ammonia oxidising archaea and bacteria in soil microcosms. Environ Microbiol 10:1357–1364CrossRefGoogle Scholar
  47. Treusch AH, Leininger S, Kletzin A, Schuster SC, Klenk H-P, Schleper C (2005) Novel genes for nitrite reductase and Amo-related proteins indicate a role of uncultivated mesophilic crenarchaeota in nitrogen cycling. Environ Microbiol 7:1985–1995CrossRefGoogle Scholar
  48. Tsui MT, Chu LM (2003) Aquatic toxicity of glyphosate-based formulations: comparison between different organisms and the effects of environmental factors. Chemosphere 52:1189–1197CrossRefGoogle Scholar
  49. Vienneau DM, Sullivan CA, House SK, Stratton GW (2004) Effects of the herbicide hexazinone on nutrient cycling in a low-pH blueberry soil. Environ Toxicol 19:115–122CrossRefGoogle Scholar
  50. Wainwright M (1978) A review of the effects of pesticides on microbial activity in soils. Eur J Soil Sci 29:287–298CrossRefGoogle Scholar
  51. Watanabe K, Kodama Y, Harayama S (2001) Design and evaluation of PCR primers to amplify 16S ribosomal DNA fragments used for community fingerprinting. J Microbiol Methods 44:253–262CrossRefGoogle Scholar
  52. Wessen E, Hallin S (2011) Abundance of archaeal and bacterial ammonia oxidizers—possible bioindicator for soil monitoring. Ecol Indic 11:1696–1698CrossRefGoogle Scholar
  53. Wittebolle L, Marzorati M, Clement L, Balloi A, Daffonchio D, Heylen K, De Vos P, Verstraete W, Boon N (2009) Initial community evenness favours functionality under selective stress. Nature 458:623–626CrossRefGoogle Scholar
  54. Yeomans JC, Bremner JM (1985) Denitrification in soil: effects of herbicides. Soil Biol Biochem 17:447–452CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • Olivier Crouzet
    • 1
    Email author
  • Franck Poly
    • 4
    • 5
  • Frédérique Bonnemoy
    • 2
    • 3
  • David Bru
    • 6
  • Isabelle Batisson
    • 2
    • 3
  • Jacques Bohatier
    • 2
    • 3
  • Laurent Philippot
    • 6
  • Clarisse Mallet
    • 2
    • 3
  1. 1.INRA UR 251 PESSAC, Centre Versailles-GrignonVersailles cedexFrance
  2. 2.CNRS UMR 6023 LMGEAubière cedexFrance
  3. 3.Clermont Université, Université Blaise PascalClermont-FerrandFrance
  4. 4.Ecologie MicrobienneINRA USC 1193 - CNRS UMR 5557VilleurbanneFrance
  5. 5.Ecologie MicrobienneUniversité de Lyon, Université Lyon 1VilleurbanneFrance
  6. 6.AgroécologieINRA, UMR 1347Dijon cedexFrance

Personalised recommendations