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Chemical characterization of soil extract as growth media for the ecophysiological study of bacteria

Abstract

We investigated the composition of soil-extracted solubilized organic and inorganic matter (SESOM) prepared from three different soils. Growth of various bacterial strains in these soil extracts was evaluated to find appropriate conditions for ecophysiological approaches. Analysis of SESOM by 1H-NMR and gas chromatography/mass spectrometry revealed a complex mixture of organic compounds. An oak forest SESOM supported the growth of several gram-positive and gram-negative soil-derived heterotrophic bacteria, whereas beech forest and grassland soil extracts did not. A metabolomic approach was performed by determining the extracellular metabolite profile of Bacillus licheniformis in SESOM. The results demonstrated that determination of the organic composition of SESOM during batch culturing is feasible. This makes SESOM amenable to studying the ecophysiology of a range of soil bacteria growing on soil-dissolved organic matter under more defined laboratory conditions. SESOM may also increase success in isolating previously uncultured or novel soil bacteria. Cell populations and the corresponding extracellular medium can be obtained readily and specific components extracted, paving the way for proteomic, transcriptomic, and metabolomic analyses. The synthetic carbon mixture based on SESOM, which mimics soil abilities, shows a positive impact on higher cell yields and longer cultivation time for biotechnological relevant bacteria.

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References

  1. Allard B (2006) A comparative study on the chemical composition of humic acids from forest soil, agricultural soil and lignite deposit—bound lipid, carbohydrate and amino acid distributions. Geoderma 130(1–2):77–96

  2. Amelung W, Zhang X (2001) Determination of amino acid enantiomers in soils. Soil Biol Biochem 33(4–5):553–562

  3. Bakken LR (1985) Separation and purification of bacteria from soil. Appl Environ Microbiol 49(6):1482–1487

  4. Benndorf D, Balcke GU, Harms H, Von Bergen M (2007) Functional metaproteome analysis of protein extracts from contaminated soil and groundwater. ISME J 1(3):224–234

  5. Bollmann A, Lewis K, Epstein SS (2007) Incubation of environmental samples in a diffusion chamber increases the diversity of recovered isolates. Appl Environ Microbiol 73(20):6386–6390

  6. Bunk B, Kucklick M, Jonas R, Munch R, Schobert M, Jahn D, Hiller K (2006) MetaQuant: a tool for the automatic quantification of GC/MS-based metabolome data. Bioinformatics 22(23):2962–2965

  7. Burke L, Brozel V, Venter S (2008) Construction and evaluation of a gfp-tagged Salmonella Typhimurium strain for environmental applications. Water SA 34(1):19–24

  8. Daniel R (2004) The soil metagenome—a rich resource for the discovery of novel natural products. Curr Opin Biotechnol 15(3):199–204

  9. Davis KE, Joseph SJ, Janssen PH (2005) Effects of growth medium, inoculum size, and incubation time on culturability and isolation of soil bacteria. Appl Environ Microbiol 71(2):826–834

  10. de Souza ML, Wackett LP, Boundy-Mills KL, Mandelbaum RT, Sadowsky MJ (1995) Cloning, characterization, and expression of a gene region from Pseudomonas sp. strain ADP involved in the dechlorination of atrazine. Appl Environ Microbiol 61(9):3373–3378

  11. Ellis RJ (2004) Artificial soil microcosms: a tool for studying microbial autecology under controlled conditions. J Microbiol Methods 56(2):287–290

  12. Fierer N, Jackson RB (2006) The diversity and biogeography of soil bacterial communities. Proc Natl Acad Sci USA 103(3):626–631

  13. Fierer N, Bradford MA, Jackson RB (2007) Toward an ecological classification of soil bacteria. Ecology 88(6):1354–1364

  14. Fischer H, Meyer A, Fischer K, Kuzyakov Y (2007) Carbohydrate and amino acid composition of dissolved organic matter leached from soil. Soil Biol Biochem 39(11):2926–2935

  15. Gans J, Wolinsky M, Dunbar J (2005) Computational improvements reveal great bacterial diversity and high metal toxicity in soil. Science 309(5739):1387–1390

  16. Hafner SD, Groffman PM, Mitchell MJ (2005) Leaching of dissolved organic carbon, dissolved organic nitrogen, and other solutes from coarse woody debris and litter in a mixed forest in New York State. Biogeochemistry 74(2):257–282

  17. Hochgrafe F, Wolf C, Fuchs S, Liebeke M, Lalk M, Engelmann S, Hecker M (2008) Nitric oxide stress induces different responses but mediates comparable protein thiol protection in Bacillus subtilis and Staphylococcus aureus. J Bacteriol 190(14):4997–5008

  18. Hoi LT, Voigt B, Jurgen B, Ehrenreich A, Gottschalk G, Evers S et al (2006) The phosphate-starvation response of Bacillus licheniformis. Proteomics 6(12):3582–3601

  19. Huang Y, Eglinton G, Van der Hage ERE, Boon JJ, Bol R, Ineson P (1998) Dissolved organic matter and its parent organic matter in grass upland soil horizons studied by analytical pyrolysis techniques. Eur J Soil Sci 49(1):1–15

  20. James N (1958) Soil extract in soil microbiology. Can J Microbiol 4(4):363–370

  21. Joseph SJ, Hugenholtz P, Sangwan P, Osborne CA, Janssen PH (2003) Laboratory cultivation of widespread and previously uncultured soil bacteria. Appl Environ Microbiol 69(12):7210–7215

  22. Kaiser K, Guggenberger G, Haumaier L, Zech W (2001) Seasonal variations in the chemical composition of dissolved organic matter in organic forest floor layer leachates of old-growth Scots pine (Pinus sylvestris L.) and European beech (Fagus sylvatica L.) stands in northeastern Bavaria, Germany. Biogeochemistry 55(2):103–143

  23. Kalbitz K, Solinger S, Park JH, Michalzik B, Matzner E (2000) Controls on the dynamics dissolved organic matter in soils: a review. Soil Sci 165(4):277–304

  24. Kovats E (1958) Gas-Chromatographische Charakterisierung Organischer Verbindungen .1. Retentionsindices Aliphatischer Halogenide, Alkohole, Aldehyde Und Ketone. Helv Chim Acta 41(7):1915–1932

  25. Liebeke M, Pother DC, van Duy N, Albrecht D, Becher D, Hochgrafe F et al (2008) Depletion of thiol-containing proteins in response to quinones in Bacillus subtilis. Mol Microbiol 69(6):1513–1529

  26. Luo Y, Vilain S, Voigt B, Albrecht D, Hecker M, Brozel VS (2007) Proteomic analysis of Bacillus cereus growing in liquid soil organic matter. FEMS Microbiol Lett 271(1):40–47

  27. Makita M, Yamamoto S, Kono M (1976) Gas–liquid chromatographic analysis of protein amino acids as N-isobutyloxycarbonylamino acid methyl esters. J Chromatogr 120(1):129–140

  28. Mongodin EF, Shapir N, Daugherty SC, DeBoy RT, Emerson JB, Shvartzbeyn A et al (2006) Secrets of soil survival revealed by the genome sequence of Arthrobacter aurescens TC1. PLoS Genet 2(12):e214

  29. Nicholson JK, Foxall PJ, Spraul M, Farrant RD, Lindon JC (1995) 750 MHz 1H and 1H-13C NMR spectroscopy of human blood plasma. Anal Chem 67(5):793–811

  30. Pace NR (1997) A molecular view of microbial diversity and the biosphere. Science 276(5313):734–740

  31. Paliy O, Gunasekera TS (2007) Growth of E. coli BL21 in minimal media with different gluconeogenic carbon sources and salt contents. Appl Microbiol Biotechnol 73(5):1169–1172

  32. Palmer KL, Mashburn LM, Singh PK, Whiteley M (2005) Cystic fibrosis sputum supports growth and cues key aspects of Pseudomonas aeruginosa physiology. J Bacteriol 187(15):5267–5277

  33. Palmer KL, Aye LM, Whiteley M (2007) Nutritional cues control Pseudomonas aeruginosa multicellular behavior in cystic fibrosis sputum. J Bacteriol 189(22):8079–8087

  34. Pizzeghello D, Zanella A, Carletti P, Nardi S (2006) Chemical and biological characterization of dissolved organic matter from silver fir and beech forest soils. Chemosphere 65(2):190–200

  35. Radajewski S, McDonald IR, Murrell JC (2003) Stable-isotope probing of nucleic acids: a window to the function of uncultured microorganisms. Curr Opin Biotechnol 14(3):296–302

  36. Rappe MS, Giovannoni SJ (2003) The uncultured microbial majority. Annu Rev Microbiol 57:369–394

  37. Reasoner DJ, Geldreich EE (1985) A new medium for the enumeration and subculture of bacteria from potable water. Appl Environ Microbiol 49(1):1–7

  38. Reizer J, Bachem S, Reizer A, Arnaud M, Saier MH Jr, Stulke J (1999) Novel phosphotransferase system genes revealed by genome analysis - the complete complement of PTS proteins encoded within the genome of Bacillus subtilis. Microbiology 145:3419–3429

  39. Sandnes A, Eldhuset TD, Wollebaek G (2005) Organic acids in root exudates and soil solution of Norway spruce and silver birch. Soil Biol Biochem 37(2):259–269

  40. Schleper C, Jurgens G, Jonuscheit M (2005) Genomic studies of uncultivated archaea. Nat Rev Microbiol 3(6):479–488

  41. Schloss PD, Handelsman J (2006) Toward a census of bacteria in soil. PLoS Comput Biol 2(7):e92

  42. Sharma PD (2005) Terrestial environments. In Environmental Microbiology. Alpha Science International, Harrow, Middlesex, UK, pp 27–51

  43. Shivaji S, Suresh K, Chaturvedi P, Dube S, Sengupta S (2005) Bacillus arsenicus sp nov., an arsenic-resistant bacterium isolated from a sidente concretion in West Bengal, India. Int J Syst Evol Microbiol 55:1123–1127

  44. Shivers RP, Dineen SS, Sonenshein AL (2006) Positive regulation of Bacillus subtilis ackA by CodY and CcpA: establishing a potential hierarchy in carbon flow. Mol Microbiol 62(3):811–822

  45. Stotzky G, Burns RG (1982) The soil environment: clay–humus–microbe interactions. In: Burns RG, Slater JH (eds) Experimental microbial ecology. Blackwell Scientific Publishing, Oxford, p 100110

  46. Strobel BW (2001) Influence of vegetation on low-molecular-weight carboxylic acids in soil solution—a review. Geoderma 99(3–4):169–198

  47. Stulke J, Hanschke R, Hecker M (1993) Temporal activation of beta-glucanase synthesis in Bacillus subtilis is mediated by the Gtp pool. J Gen Microbiol 139:2041–2045

  48. Tobisch S, Zuhlke D, Bernhardt J, Stulke J, Hecker M (1999) Role of CcpA in regulation of the central pathways of carbon catabolism in Bacillus subtilis. J Bacteriol 181(22):996–7004

  49. Torsvik V, Øvreås L (2002) Microbial diversity and function in soil: from genes to ecosystems. Curr Opin Microbiol 5(3):240–245

  50. Torsvik V, Goksoyr J, Daae FL (1990) High diversity in DNA of soil bacteria. Appl Environ Microb 56(3):782–787

  51. Torsvik V, Ovreas L, Thingstad TF (2002) Prokaryotic diversity—magnitude, dynamics, and controlling factors. Science 296(5570):1064–1066

  52. Tringe SG, von Mering C, Kobayashi A, Salamov AA, Chen K, Chang HW et al (2005) Comparative metagenomics of microbial communities. Science 308(5721):554–557

  53. Urich T, Lanzen A, Qi J, Huson DH, Schleper C, Schuster SC (2008) Simultaneous assessment of soil microbial community structure and function through analysis of the meta-transcriptome. PLoS ONE 3(6):e2527

  54. van Hees PAW, Jones DL, Finlay R, Godbold DL, Lundstomd US (2005) The carbon we do not see—the impact of low molecular weight compounds on carbon dynamics and respiration in forest soils: a review. Soil Biol Biochem 37(1):1–13

  55. Veith B, Herzberg C, Steckel S, Feesche J, Maurer KH, Ehrenreich P et al (2004) The complete genome sequence of Bacillus licheniformis DSM13, an organism with great industrial potential. J Mol Microbiol Biotechnol 7(4):204–211

  56. Vilain S, Luo Y, Hildreth MB, Brozel VS (2006) Analysis of the life cycle of the soil saprophyte Bacillus cereus in liquid soil extract and in soil. Appl Environ Microb 72(7):4970–4977

  57. Voigt B, Hoi LT, Jurgen B, Albrecht D, Ehrenreich A, Veith B et al (2007) The glucose and nitrogen starvation response of Bacillus licheniformis. Proteomics 7(3):413–423

  58. Watanabe K, Imase M, Aoyagi H, Ohmura N, Saiki H, Tanaka H (2008) Development of a novel artificial medium based on utilization of algal photosynthetic metabolites by symbiotic heterotrophs. J Appl Microbiol 105(3):741–751

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Acknowledgements

We are grateful to S. Seefeld for performing the ICP and IC. We thank K. Surmann for the assistance and Dr. M. Sadowsky for donating the bacterial strains. This research was funded by a grant from the Apotheker-Paul-Marschall-Stiftung to M. Liebeke and by the South Dakota Agricultural Experiment Station and the State of South Dakota. VSB was the recipient of a fellowship from the Stiftung Alfried Krupp Kolleg Greifswald.

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Correspondence to Michael Lalk.

Additional information

Journal series publication 3622 from the South Dakota Agricultural Experiment Station.

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Fig. S1

Growth of Bacillus licheniformis DSM 13 in synthetic media and appending metabolite concentrations [in millimolars] of culture supernatants (determined by 1H-NMR measurements) for M9 medium with a carbon source cocktail in low concentrations (a) and high concentrations (b). Also displayed is growth in M9 medium with glucose in low concentrations (c) and high concentrations (d) as carbon source. All experiments were done in triplicate and a representative experiment is shown (DOC 221 kb)

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Liebeke, M., Brözel, V.S., Hecker, M. et al. Chemical characterization of soil extract as growth media for the ecophysiological study of bacteria. Appl Microbiol Biotechnol 83, 161–173 (2009). https://doi.org/10.1007/s00253-009-1965-0

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Keywords

  • Dissolved organic matter
  • Gas chromatography/mass spectrometry (GC-MS)
  • Nuclear magnetic resonance (NMR)
  • Growth medium
  • SESOM
  • Soil bacteria
  • Metabolomics