PLFA Profiling of Coal Mine Spoil: An Integrated Approach for the Assessment of Ecological Restoration


  • Payal Agrawal Department of Biotechnology and Bioinformatics, Sambalpur University, At/po- Jyoti Vihar, Burla– 768019, Odisha
  • Jitesh Kumar Maharana Department of Biotechnology and Bioinformatics, Sambalpur University, At/po-Jyoti Vihar, Burla– 768019, Odisha
  • Amiya Kumar Patel Department of Biotechnology and Bioinformatics, Sambalpur University, At/po- Jyoti Vihar, Burla– 768019, Odisha



Coal Mine Spoil, Microbial Community Structure, Mine Spoil Genesis, PLFA


Coal mine overburden spoil created aftermath of mining activities represents disequilibrated geomorphic system. The pedodiversity including its link with biodiversity and landscape ecology describe the spatial diversity has emerged as functional determinants of ecosystem processes. Being the driving force mediating soil processes, ecosystem restoration through mine spoil genesis is monitored based on the shift in microbial community structure in different age series coal mine spoil. Phospholipid fatty acid analysis is culture-independent approach, which provides a set of molecular markers to determine microbial community composition and discriminate microbial communities of different origin. PLFAs are synthesized during microbial growth, rapidly degraded following cell death and reliably reflect living microbial communities. Relative distribution of 51 PLFAs revealed significant variation in microbial community structure across the sites with Shannon diversity index varies from 1.5265 (OB0) to 2.0139 (OB15) and Pielous evenness index from 0.4110 (OB0) to 0.5260 (OB15). Fungal to bacterial ratio exhibited an increasing trend from OB0 (0.055) to OB15 (0.348) over time, which revealed the sign of mine spoil genesis. The principal component analysis and redundancy analysis discriminate different age series coal mine spoil into independent clusters, which evaluated the broad scale patterns of microbial community structure influencing the pace and progress of mine spoil genesis.


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Alavi R, Alinejad-Rokny H, Zadeh MS. Prioritizing coercive plant species in Choghart iron mine desert region. Australian Journal of Basic and Applied Sciences. 2011; 5(12):1075–78.

Kujur M, Patel AK. PLFA profiling of soil microbial community structure and diversity in different dry tropical ecosystems of Jharkhand. International Journal of Current Microbiology and Applied Sciences. 2014; 3(3):556–75.

Kelly JJ, Haggblom MM, Tate RL. Effects of heavy metal contamination and remediation on soil microbial communities in the vicinity of a zinc smelter as indicated by analysis of microbial community phospholipid fatty acid profiles. Biology and Fertility of Soils. 2003; 38:65–71.

Veresoglou SD, Mamolos AP, Thornton B, Voulgari OK, Sen R, Vereogou DS. Medium term fertilization of grassland plant communities masks plant species linked effects on soil microbial community structure. Plant Soil. 2011; 344:187– 96.

Frostegard A, Tunlid A, Baath E. Changes in microbial community structure during long term incubation in two soils experimentally contaminated with metals. Soil Biology and Biochemistry. 1996; 28:55–63.

Frostegard A, Tunlid A and Baath E. Use and misuse of PLFA measurements in soil. Soil Biology and Biochem. 2011; 43(8):1621–25.

Zelles L, Bai QY, Beck T, Beese F. Signature fatty acids in phospholipids and lipo polysaccharides as indicators of microbial biomass and community structure in agricultural soils. Soil Biology and Biochem. 1992; 24:317–23.

Bardgett RD, McAlister E. The measurement of soil fungal: bacterial biomass ratios as an indicator of ecosystem self-regulation in temperate meadow grasslands. Biology and Fertility of Soils. 1999; 29:282–90.

White DC, Stair JO, Ringelberg DB. Quantitative comparisons of in situ microbial biodiversity by signature biomarker analysis. Journal of Industrial Microbiology. 1996; 17:185–96.

Heipieper HJ, Meulenbeld G, Oirschot QV, De Bont JAM. Effect of environment factors on trans/cis ratio of unsaturated fatty acids in Pseudomonas putida S12. Applied and Environmental Microbiology. 1996; 62:2773–7. PMid:16535373. PMCid:PMC1388911

Frostegard A, Baath E, Tunlip A. Shifts in the structure of soil microbial communities in limed soils as revealed by phospholipid fatty acid analysis. Soil Biology and Biochemistry. 1993; 25:723–30.

Kaur A, Chaudhary A, Kaur A, Choudhary R, Kaushik R. Phospholipid fatty acid, a bio-indicator of environment monitoring assessment in soil ecosystem. Current Sciences. 2005; 89:1103–12.

Vestal JR, White DC. Lipid analysis in microbial ecology quantitative approaches to the study of microbial communities. Bioscience. 1989; 39:535–41. PMid:11542183

Zelles L. Fatty acid patterns of phospholipids and lipopolysaccharides in the characterization of microbial communities in soil: A review. Biology and Fertility of Soils. 1999; 29:111–29.

White DC. Is there anything else you to understand about the microbiota that cannot be derived from analysis of nucleic acid? Microbial Ecology. 1994; 28:163–6. BF00166805. PMid:24186442

Dickens SJM, Allen EB, Santiago LS, Crowley D. Exotic annuals reduce soil heterogeneity in coastal sage scrub soil chemical and biological characteristics. Soil Biology and Biochemistry. 2013; 58:70–81.

Lores M, Gomez-Brandon M, Dominguez J. Tracking down microbial communities via fatty acid and analysis: Analytical strategy for solid organic samples. Current Research, Technology and Education Topics in Applied Microbiology and Microbial Biotechnology. 2010:1502–8.

Zelles L. Phospholipid fatty acid profiles in selected members for soil microbial communities. Chemosphere. 1997; 35:275–94.

Morgan JAW, Winstanley C. Microbial biomarkers. Modern Soil Microbiology; 1997. 331–48.

Frostegard A, Baath E. The use of phospholipid fatty acid to estimate bacterial and fungal biomass in soil. Biology Fertility of Soils. 1996; 22:59–65.

Olsson PA, Van Aarle IM, Gavito ME, Bengtson P, Bengtsson G. 13C incorporation into signature fatty acids as an assay for carbon allocation in arbuscular mycorrhiza. Applied and Environmental Microbiology. 2005; 71:2592–9. PMid:15870350. PMCid:PMC1087529

Olsson PA. Signature fatty acids provide tools for determination of the distribution and interactions of mycorrhizal fungi in soil. FEMS Microbiology Ecology. 1999; 29:303–15.

Hill GT, Mitkowski NA, Aldrich-Wolfe L, Emele LR, Jurkonie DD, Ficke A, Maldonado- Ramirez S, Lynch ST, Nelson EB. Methods for assessing the composition and diversity of soil microbial communities. Applied Soil Ecology. 2000; 15:25–36.

Frostegard A, Tunlid A, Baath E. Microbial biomass measured as total lipid phosphate in soils of different organic content. Journal of Microbiological Methods. 1991; 14:151–63.

Zhong S, Wu Y, Xu J. Phosphorus utilization and microbial community in response to lead/iron addition to a waterlogged soil. Journal of Environmental Science. 2009; 21:1415–23.

Bowman JP, Skerratt JH, Nichols PD, Sly LI. Phospholipid fatty acid and lipopolysaccharide fatty acid signature lipids in methane utilizing bacteria. FEMS Microbiology Ecology. 1991; 85:15–22.

Robie JV, White DC. Lipid analysis in microbial ecology: Quantitative approaches to the study of microbial communities. Bioscience. 1989; 39(8):535–41. PMid:11542183

Chowdhury N, Marschner P, Burns R. Response of microbial activity and community structure to decreasing soil osmotic and metric potential. Plant Soil. 2011; 344:241–54.

Claassens S, Riedel KJ, Rensburg L Van, Bezuidenhout JJ, Rensburg Jansen van PJ. Microbial community function and structure on coal mine discard under rehabilitation. South African Journal of Plant Soil. 2006; 23(2):105–12. /10.1080/02571862.2006.10634739

White DC, Davies WM, Nickels JS, King JD, Bobbie RJ. Determination of the sedimentary microbial biomass by extractable lipid phosphate. Oecologia. 1979; 40:51–62. PMid:28309603

Fierer N, Schimel JP, Holden PA. Variation in microbial community composition through two soil depth profile. Soil Biology and Biochemistry. 2003; 35:167–76.

Fraterrigo JM, Balser TC, Turner MG. Microbial community variation and its relationship with nitrogen mineralization in historically altered forests. Ecology. 2006; 87:70–579. PMid:16602287

Harris JA. Measurements of the soil microbial community for estimating the success of restoration. European Journal of Soil Science. 2003; 54:801–8.

Fang C, Rhadosevich M, Fuhrmann J. Characterization of rhizosphere microbial community structure in five similar grass species using FAME and BIOLOG analyses. Soil Biology and Biochemistry. 2001; 33:679–82.

Hossain Z, Sugiyama SI. Geographical structure of soil microbial communities in northern Japan: effects of distance, land use type and soil properties. European Journal of Soil Biology. 2011; 47(2):88–94.

Willers C, Jansen Van Rensberg PJ, Claassens S. Phospholipid fatty acid profiling of microbial communities-A review of interpretations and recent applications. Journal of Applied Microbiology. 2015; 119:1207–18. jam.12902. PMid:26184497

Zhang QC, Shamsi IH, Xu DT, Wang GH, Lin XY, Jilani G, Hussain N, Chaudhry AN. Chemical fertilizer and organic manure inputs in soil exhibit a vice versa pattern of microbial community structure. Applied Soil Ecology. 2012; 57:1–8. https://

Ferrari AE, Ravnskov S, Larsen J, Tønnersen T, Maronna RA, Wall LG. Crop rotation and seasonal effects on fatty acid profiles of neutral and phospholipids extracted from no?till agricultural soils. Soil Use and Management. 2015; 31(1):165–75.

Hassan N, Anesio AM, Rafiq M, Holtvoeth J, Bull I, Haleem A, Shah AA, Hasan F. Temperature driven membrane lipid adaptation in glacial psychrophilic bacteria. Frontiers in Microbiology. 2020; 11:824. PMid:32477293. PMCid:PMC7240044

Schäpe SS, Krause JL, Masanetz RK, Riesbeck S, Starke R, RolleKampczyk U, et al. Environmentally relevant concentration of bisphenol s shows slight effects on SIHUMIx. Microorganisms. 2020; 8(9):1436. PMid:32961728. PMCid:PMC7564734

Yang W, Jeelani N, Cai A, Cheng X, An S. Coastal reclamation alters soil microbial communities following different land use patterns in the Eastern coastal zone of China. Scientific Reports. 2021; 11(1):1–4. PMid:33790383. PMCid:PMC8012362

Liao M, Chen CL, Huan CY. Effects of heavy metals on soil microbial activity and diversity in a reclaimed mining wasteland of red soil area. Journal of Environmental Sciences. 2005; 17:832–7.

Trippe KM, Manning VA, Reardon CL, Klein AM, Weidman C, Ducey TF, et al. Phytostabilization of acidic mine tailings with biochar, biosolids, lime, and locally-sourced microbial inoculum: Do amendment mixtures influence plant growth, tailing chemistry, and microbial composition?. Applied Soil Ecology. 2021; 165.

Moore-Kucera J, Dick RP. PLFA profiling of microbial community structure and seasonal shift in soils of a Douglas-fir chronosequences. Microbial Ecology. 2008; 55:500–11. https:// PMid:17786504

Yu S, Ehrenfeld JG. Relationships among plants, soils and microbial communities along a hydrological gradient in the New Jersey Pinelands, USA. Annals of Botany. 2010; 10:185– 96. PMid:19643908. PMCid:PMC2794054

Cairney JWG, Meharg AA. Interaction between ectomycorrhizal fungi and soil saprotrophs: Implications for decomposition of organic matter in soils and degradation of organic pollutants in the rhizosphere. Canadian Journal of Botany. 2002; 80:803–9.

Hackl E, Pfeffer M, Donat C, Bachmann G, ZechmeisterBoltenstern S. Composition of the microbial communities in the mineral soil under different types of natural forest. Soil Biology and Biochemistry. 2005; 37:661–71.

Rajapaksha RMCP, Tobor-Kaplon MA, Baath E. Metal toxicity affects fungal and bacterial activities in soil differently. Applied and Environmental Microbiology. 2004; 70(50):2966–73. https:/ PMid:15128558. PMCid:PMC404458

Peacock AD, Macnaughton ST, Cantu JM, Dale VH, White DC. Soil microbial biomass and community composition along an anthropogenic disturbance gradient with a long leaf pine habitat. Ecological Indicators. 2001; 1:113–21.

Bailey VL, Smith JL, Bolton HJ. Fungal to bacterial ratios in soils investigated for enhanced Carbon sequestration. Soil Biology and Biochemistry. 2002; 34:997–1007.

Hogberg MN, Hogberg P, Myrold DD. Is microbial community composition in boreal forest soils determined by pH, C-to-N ratio, the trees or all three?. Oecologia. 2007; 150:590–601, PMid:17033802

Van der Wal A, van Veen JA, Smant W, Boschker HTS, Bloem J, Kardol P, et al. Fungal biomass development in a chronosequence of land abandonment. Soil Biology and Biochemistry. 2006;


Waldrop MP, Balser TC, Firestone MK. Linking microbial community composition to function in a tropical soil. Soil Biology and Biochemistry. 2000; 32(13):1837–46.

Lauber CL, Strickland MS, Bradford MA, Fierer N. The influence of soil properties on the structure of bacterial and fungal communities across land use types. Soil Biology and Biochemistry. 2008; 40:2407–15.

Baath E, Anderson TH. Comparison of soil fungal/bacterial ratios in a pH gradient using physiological and PLFA based techniques. Soil Biology and Biochemistry. 2003; 35:955–63.

Zhou W, Hui D, Shen W. Effects of soil moisture on the temperature sensitivity of soil heterotrophic respiration: A laboratory incubation study. PLoS One. 2014; 9(3):92531–40. PMid:24647610. PMCid:PMC3960259

Anderson JPE, Domsch KH. Quantities of plant nutrients in the microbial biomass of selected soils. Soil Science. 1980; 130:211–16.

Ludwig JA, Reynolds JF. Statistical Ecology: A primer in method and computing. John Wiley and Sons;1998. p. 337.

Urbanova M, Kopecky J, Valaskova V, Mareckova MS, Elhottova D, Kyselkova M, et al. Development of bacterial community during spontaneous succession on spoil heaps after brown coal mining. FEMS Microbiology Ecology. 2011; 78:59–69. PMid:21707674

Yadav SK, Banerjee A, Jhariya MK, Meena RS, Khan N, Raj A. Eco-restoration of bauxite mining: An ecological approach. In: Natural Resources Conservation and Advances for Sustainability;

p. 173–93.




How to Cite

Agrawal, P., Maharana, J. K., & Patel, A. K. (2022). PLFA Profiling of Coal Mine Spoil: An Integrated Approach for the Assessment of Ecological Restoration. Journal of Ecophysiology and Occupational Health, 22(2), 77–87.