Extracellular DNA from dead microorganisms can persist in soil for weeks to years1–3. Although it is implicitly assumed that the microbial DNA recovered from soil predominantly represents intact cells, it is unclear how extracellular DNA affects molecular analyses of microbial diversity. We examined a wide range of soils using viability PCR based on the photoreactive DNA-intercalating dye propidium monoazide4. We found that, on average, 40% of both prokaryotic and fungal DNA was extracellular or from cells that were no longer intact. Extracellular DNA inflated the observed prokaryotic and fungal richness by up to 55% and caused significant misestimation of taxon relative abundances, including the relative abundances of taxa integral to key ecosystem processes. Extracellular DNA was not found in measurable amounts in all soils; it was more likely to be present in soils with low exchangeable base cation concentrations, and the effect of its removal on microbial community structure was more profound in high-pH soils. Together, these findings imply that this ‘relic DNA’ remaining in soil after cell death can obscure treatment effects, spatiotemporal patterns and relationships between microbial taxa and environmental conditions.
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The authors thank S. Grandy and J. Schnecker at the University of New Hampshire, C. Bueno de Mesquita and L. Vimercati at the University of Colorado Boulder, E. Skokan at Black Cat Farms and K. McLachlan at Kansas State University for soil collection or access to collection sites. The authors also thank J. Henley, R. Hacker-Cary and K. Vaccarello for assistance with DNA extractions. Sequencing was performed at the University of Colorado BioFrontiers Institute's Next-Gen Sequencing Core Facility. Funding to support this work was provided by the National Science Foundation (DEB 0953331, EAR 1331828, DUE 1259336 and EAR 1461281) and a Visiting Postdoctoral Fellowship award to P.C. from the Cooperative Institute for Research in Environmental Sciences.
The authors declare no competing financial interests.
Supplementary Figures 1-11. (PDF 950 kb)
Full dataset for Fig. 1; mean percent of each taxa per soil, per treatment; and amplicon copies per replicate per gram of soil. (XLSX 90 kb)
List of soil sample locations, ecosystem descriptions, edaphic characteristics, mean community dissimilarity after relic DNA removal, mean percent of relic DNA for each soil, mean richness, and mean 16S and ITS amplicon abundances for both untreated and PMA-treated soils. (XLSX 56 kb)
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Carini, P., Marsden, P., Leff, J. et al. Relic DNA is abundant in soil and obscures estimates of soil microbial diversity. Nat Microbiol 2, 16242 (2017). https://doi.org/10.1038/nmicrobiol.2016.242
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