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Journal of Soils and Sediments

, Volume 19, Issue 1, pp 91–96 | Cite as

Assessing soil extracellular DNA decomposition dynamics through plasmid amendment coupled with real-time PCR

  • Fang Wang
  • Rongxiao Che
  • Zhihong Xu
  • Yanfen Wang
  • Xiaoyong CuiEmail author
Soils, Sec 1 • Soil Organic Matter Dynamics and Nutrient Cycling • Short Original Communication
  • 101 Downloads

Abstract

Purpose

Determining soil extracellular DNA decomposition dynamics is essential to assessing lateral gene transfer possibility, nutrient-cycling efficiency, and the reliability of DNA-based methods for examining microbes in soils. The existing methods based on competent cell transformation and stable isotope probes are generally inefficient and not strictly quantitative. Therefore, this study aimed to establish a rigorously quantitative and efficient approach to monitor the decomposition dynamics of the soil extracellular DNA.

Materials and methods

A soil was collected from a Tibetan alpine meadow. Extracellular DNA was simulated by modified exogenous plasmids. The plasmid solution was sprayed onto the fresh soil and thoroughly homogenized. Then, the soil was incubated for 4 weeks, during which they were sampled and immediately stored at − 20 °C on days 0, 0.5, 1, 2, 4, 8, 16, and 28 of the incubation. Finally, the total soil DNA was extracted, and the exogenous plasmid copies remained in the soils were determined using real-time PCR. Additionally, another similar experiment was conducted with a sterilized soil to assess the abiotic influences on the changes in the exogenous plasmid copies.

Results and discussion

In the fresh soil, the exogenous plasmid DNA copies decreased quickly in the first 12 h of the incubation, remained stable in the following 36 h, and gradually dropped to 1.10–5.20% of the initial plasmid copies at sampling time 0 after being incubated for 4 weeks. The variations in the soil plasmid DNA copies fitted well with the modified exponential decay model. As for the sterilized soil, the exogenous plasmid copies remained stable during the 16 day’s incubation. However, they dramatically dropped after being incubated for 28 days, which was probably elicited by the recolonization of microbes in the soil. Collectively, the decrease in the exogenous plasmid DNA copies could be mainly attributed to biological activities.

Conclusions

Extracellular DNA can persist in soil for more than 4 weeks. Exogenous plasmid amendment coupled with real-time PCR provides a convenient and rigorously quantitative approach for monitoring extracellular DNA degradation in soils.

Keywords

Antibiotic resistance genes DNA degradation Quantitative approach Environmental DNA eDNA 

Notes

Funding information

This work was supported by the National Key Research and Development Program of China (2016YFC0501800), the Strategic Priority Research Program (A) of the Chinese Academy of Sciences (XDA20050103), and the National Natural Science Foundation of China (31570518).

Supplementary material

11368_2018_2176_MOESM1_ESM.docx (22 kb)
ESM 1 (DOCX 18 kb)

References

  1. Agnelli A, Ascher J, Corti G, Ceccherini MT, Nannipieri P, Pietramellara G (2004) Distribution of microbial communities in a forest soil profile investigated by microbial biomass, soil respiration and DGGE of total and extracellular DNA. Soil Biol Biochem 36:859–868CrossRefGoogle Scholar
  2. Alef K, Nannipieri P (1995) Methods in applied soil microbiology and biochemistry. Academic Press, LondonGoogle Scholar
  3. Ascher J, Ceccherini MT, Pantani OL, Agnelli A, Borgogni F, Guerri G, Nannipieri P, Pietramellara G (2009) Sequential extraction and genetic fingerprinting of a forest soil metagenome. Appl Soil Ecol 42:176–181CrossRefGoogle Scholar
  4. Barnard RL, Osborne CA, Firestone MK (2015) Changing precipitation pattern alters soil microbial community response to wet-up under a Mediterranean-type climate. ISME J 9:946–957CrossRefGoogle Scholar
  5. Berendonk TU, Manaia CM, Merlin C, Fatta-Kassinos D, Cytryn E, Walsh F, Burgmann H, Sorum H, Norstrom M, Pons MN, Kreuzinger N, Huovinen P, Stefani S, Schwartz T, Kisand V, Baquero F, Martinez JL (2015) Tackling antibiotic resistance: the environmental framework. Nat Rev Microbiol 13:310–317CrossRefGoogle Scholar
  6. Blum SAE, Lorenz MG, Wackernagel W (1997) Mechanism of retarded DNA degradation and prokaryotic origin of DNases in nonsterile soils. Syst Appl Microbiol 20:513–521CrossRefGoogle Scholar
  7. Cai P, Huang Q, Zhang X, Chen H (2006) Adsorption of DNA on clay minerals and various colloidal particles from an Alfisol. Soil Biol Biochem 38:471–476CrossRefGoogle Scholar
  8. Che RX, Deng YC, Wang F, Wang WJ, Xu ZH, Wang YF, Cui XY (2015) 16S rRNA-based bacterial community structure is a sensitive indicator of soil respiration activity. J Soils Sediments 15:1987–1990CrossRefGoogle Scholar
  9. Che RX, Wang WJ, Zhang J, Nguyen TTN, Tao J, Wang F, Wang YF, Xu ZH, Cui XY (2016) Assessing soil microbial respiration capacity using rDNA- or rRNA-based indices: a review. J Soils Sediments 16:2698–2708CrossRefGoogle Scholar
  10. Che RX, Wang F, Wang WJ, Zhang J, Zhao X, Rui YC, Xu ZH, Wang YF, Hao YB, Cui XY (2017) Increase in ammonia-oxidizing microbe abundance during degradation of alpine meadows may lead to greater soil nitrogen loss. Biogeochemistry 136:341–352CrossRefGoogle Scholar
  11. Che RX, Deng YC, Wang F, Wang WJ, Xu ZH, Hao YB, Xue K, Zhang B, Tang L, Zhou HK, Cui XY (2018a) Autotrophic and symbiotic diazotrophs dominate nitrogen-fixing communities in Tibetan grassland soils. Sci Total Environ 639:997–1006CrossRefGoogle Scholar
  12. Che RX, Deng YC, Wang WJ, Rui YC, Zhang J, Tahmasbian I, Tang L, Wang SP, Wang YF, Xu ZH, Cui XY (2018b) Long-term warming rather than grazing significantly changed total and active soil procaryotic community structures. Geoderma 316:1–10CrossRefGoogle Scholar
  13. Che RX, Qin JL, Tahmasbian I, Wang F, Zhou ST, Xu ZH, Cui XY (2018c) Litter amendment rather than phosphorus can dramatically change inorganic nitrogen pools in a degraded grassland soil by affecting nitrogen-cycling microbes. Soil Biol Biochem 120:145–152CrossRefGoogle Scholar
  14. Finkel SE, Kolter R (2001) DNA as a nutrient novel role for bacterial competence gene homologs. J Bacteriol 183:6288–6293CrossRefGoogle Scholar
  15. Frostegård Å, Courtois S, Ramisse V, Clerc S, Bernillon D, Le Gall F, Jeannin P, Nesme X, Simonet P (1999) Quantification of bias related to the extraction of DNA directly from soils. Appl Environ Microbiol 65:5409–5420Google Scholar
  16. Griffiths RI, Whiteley AS, O'Donnell AG, Bailey MJ (2003) Influence of depth and sampling time on bacterial community structure in an upland grassland soil. FEMS Microbiol Ecol 43:35–43CrossRefGoogle Scholar
  17. Gulden RH, Lerat S, Hart MM, Powell JR, Trevors JT, Pauls KP, Klironomos JN, Swanton CJ (2005) Quantitation of transgenic plant DNA in leachate water: real-time polymerase chain reaction analysis. J Agric Food Chem 53:5858–5865CrossRefGoogle Scholar
  18. Hannan S, Ready D, Jasni AS, Rogers M, Pratten J, Roberts AP (2010) Transfer of antibiotic resistance by transformation with eDNA within oral biofilms. FEMS Immunol Med Microbiol 59:345–349CrossRefGoogle Scholar
  19. Harris D, Paul EA (1994) Measurement of bacterial growth rates in soil. Appl Soil Ecol 1:277–290CrossRefGoogle Scholar
  20. Herzog S, Wemheuer F, Wemheuer B, Daniel R (2015) Effects of fertilization and sampling time on composition and diversity of entire and active bacterial communities in German grassland soils. PLoS One 10:e0145575CrossRefGoogle Scholar
  21. Hobbie SE, Eddy WC, Buyarski CR, Adair EC, Ogdahl ML, Weisenhorn P (2012) Response of decomposing litter and its microbial community to multiple forms of nitrogen enrichment. Ecol Monogr 82:389–405CrossRefGoogle Scholar
  22. Kandhavelu M, Vennison SJ (2008) Persistence of plasmid DNA in different soils. Afr J Biotechnol 7:2543–2546Google Scholar
  23. Kunito T, Ihyo Y, Miyahara H, Seta R, Yoshida S, Kubo H, Nagaoka K, Sakai M, Saeki K (2016) Soil properties affecting adsorption of plasmid DNA and its transformation efficiency in Escherichia coli. Biol Fertil Soils 52:223–231CrossRefGoogle Scholar
  24. Lim J, Shin SG, Lee S, Hwang S (2011) Design and use of group-specific primers and probes for real-time quantitative PCR. Front Environ Sci Eng 5:28–39CrossRefGoogle Scholar
  25. Mao D, Luo Y, Mathieu J, Wang Q, Feng L, Mu Q, Feng C, Alvarez PJ (2014) Persistence of extracellular DNA in river sediment facilitates antibiotic resistance gene propagation. Environ Sci Technol 48:71–78CrossRefGoogle Scholar
  26. Mazzoleni S, Carteni F, Bonanomi G, Senatore M, Termolino P, Giannino F, Incerti G, Rietkerk M, Lanzotti V, Chiusano ML (2015) Inhibitory effects of extracellular self-DNA: a general biological process? New Phytol 206:127–132CrossRefGoogle Scholar
  27. Mićić M, Whyte JD, Karsten V (2016) High performance bead beating based lysing, homogenization and grinding for DNA, RNA and proteins extraction with FastPrep® systems, sample preparation techniques for soil, plant, and animal samples. Springer Protocols Handbooks. Humana Press, New York, pp 99–116Google Scholar
  28. Morrissey EM, McHugh TA, Preteska L, Hayer M, Dijkstra P, Hungate BA, Schwartz E (2015) Dynamics of extracellular DNA decomposition and bacterial community composition in soil. Soil Biol Biochem 86:42–49CrossRefGoogle Scholar
  29. Paget E, Monrozier LJ, Simonet P (1992) Adsorption of DNA on clay minerals: protection against DNaseI and influence on gene transfer. FEMS Microbiol Lett 97:31–39CrossRefGoogle Scholar
  30. Paget E, Lebrun M, Freyssinet G, Simonet P (1998) The fate of recombinant plant DNA in soil. Eur J Soil Biol 34:81–88CrossRefGoogle Scholar
  31. Peng C, Huang QY, Lu YD, Chen WL, Jiang DH, Wei LA (2007) Amplification of plasmid DNA bound on soil colloidal particles and clay minerals by the polymerase chain reaction. J Environ Sci 19:1326–1329CrossRefGoogle Scholar
  32. Pietramellara G, Ascher J, Ceccherini MT, Nannipieri P, Wenderoth D (2007) Adsorption of pure and dirty bacterial DNA on clay minerals and their transformation frequency. Biol Fertil Soils 43:731–739CrossRefGoogle Scholar
  33. Pietramellara G, Ascher J, Borgogni F, Ceccherini MT, Guerri G, Nannipieri P (2009) Extracellular DNA in soil and sediment: fate and ecological relevance. Biol Fertil Soils 45:219–235CrossRefGoogle Scholar
  34. R Core Team (2018) R: a language and environment for statistical computing. R Foundation for Statistical Computing, Vienna http://www.R-project.org/ Google Scholar
  35. Romanowski G, Lorenz MG, Sayler G, Wackernagel W (1992) Persistence of free plasmid DNA in soil monitored by various methods, including a transformation assay. Appl Environ Microbiol 58:3012–3019Google Scholar
  36. Romanowski G, Lorenz MG, Wackernagel W (1993a) Use of polymerase chain reaction and electroporation of Escherichia coli to monitor the persistence of extracellular plasmid DNA introduced into natural soils. Appl Environ Microbiol 59:3438–3446Google Scholar
  37. Romanowski G, Lorenz MG, Wackernagel W (1993b) Plasmid DNA in a groundwater aquifer microcosm-adsorption, DNAase resistance and natural genetic transformation of Bacillus subtilis. Microb Ecol 2:171–181Google Scholar
  38. Trevors JT (1996) Sterilization and inhibition of microbial activity in soil. J Microbiol Methods 26:53–59CrossRefGoogle Scholar
  39. Tuominen L, Kairesalo T, Hartikainen H (1994) Comparison of methods for inhibiting bacterial activity in sediment. Appl Environ Microbiol 60:3454–3457Google Scholar
  40. Udikovic-Kolic N, Wichmann F, Broderick NA, Handelsman J (2014) Bloom of resident antibiotic-resistant bacteria in soil following manure fertilization. Proc Natl Acad Sci U S A 111:15202–15207CrossRefGoogle Scholar
  41. Vasi F, Travisano M, Lenski RE (1994) Long-term experimental evolution in Escherichia coli. II. Changes in life-history traits adaptation to a seasonal environment. Am Nat 144:432–456CrossRefGoogle Scholar
  42. Vishnivetskaya TA, Layton AC, Lau MCY, Chauhan A, Cheng KR, Meyers AJ, Murphy JR, Rogers AW, Saarunya GS, Williams DE, Pfiffner SM, Biggerstaff JP, Stackhouse BT, Phelps TJ, Whyte L, Sayler GS, Onstott TC (2014) Commercial DNA extraction kits impact observed microbial community composition in permafrost samples. FEMS Microbiol Ecol 87:217–230CrossRefGoogle Scholar
  43. WRB (2006) World reference base for soil resources 2006. FAO/ISRIC/ISSS, ItalyGoogle Scholar
  44. Yan H, Wang DD, Dong B, Tang FF, Wang BC, Fang H, Yu YL (2011) Dissipation of carbendazim and chloramphenicol alone and in combination and their effects on soil fungal:bacterial ratios and soil enzyme activities. Chemosphere 84:634–641CrossRefGoogle Scholar
  45. Yu WH, Li N, Tong DS, Zhou CH, Lin CX, Xu CY (2013) Adsorption of proteins and nucleic acids on clay minerals and their interactions: a review. Appl Clay Sci 80–81:443–452CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.College of Life SciencesUniversity of Chinese Academy of SciencesBeijingChina
  2. 2.Environmental Futures Research Institute, School of Environment and ScienceGriffith UniversityBrisbaneAustralia
  3. 3.Institute of International Rivers and Eco-securityYunnan UniversityKunmingChina

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