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Title: Detection of Organohalide-Respiring Enzyme Biomarkers at a Bioaugmented TCE-Contaminated Field Site

Abstract

RNA-based biomarkers have been successfully detected at field sites undergoing in situ bioremediation, but the detection of expressed enzymes is a more direct way to prove activity for a particular biocatalytic process of interest since they provide evidence of potential in situ activity rather than simply confirming presence and abundance of genes in a given population by measurement of DNA copies using qPCR. Here we successfully applied shotgun proteomics to field samples from a trichloroethene (TCE)-contaminated industrial site in southern Ontario, Canada that had been bioaugmented with the commercially available KB-1 TM microbial culture. The KB-1 TM culture contains multiple strains of Dehalococcoides mccartyi (D. mccartyi) as well as an organohalide respiring Geobacter species. The relative abundances of specific enzymatic proteins were subsequently compared to corresponding qPCR-derived levels of DNA and RNA biomarkers in the same samples. Samples were obtained from two wells with high hydraulic connectivity to the KB-1 TM-bioaugemented enhanced in situ bioremediation system, and two control wells that showed evidence of low levels of native organohalide respiring bacteria (OHRB), Dehalococcoides and Geobacter. Enzymes involved in organohalide respiration were detected in the metaproteomes of all four field samples, as were chaperonins of D. mccartyi, chemotaxis proteins, and ATPases.more » The most highly expressed RDase in the bioaugmentation culture (VcrA) was the most highly detected enzyme overall in the bioaugmented groundwater samples. In one background groundwater well, we found high expression of the Geobacter pceA RDase. The DNA and RNA biomarkers detected using qPCR-based assays were a set of orthologs of Dehalococcoides reductive dehalogenases (VcrA, TceA, BvcA, dehalogenase “DET1545”), and the Ni-Fe uptake hydrogenase, HupL. Within a sample, RNA levels for key enzymes correlated with relative protein abundance. These results indicate that laboratory observations of TCE-bioremediation biomarker protein expression are recapitulated in field environmental systems and that both RNA and protein biomarker monitoring hold promise for activity monitoring of in situ populations of OHRB.« less

Authors:
 [1];  [1];  [2]; ORCiD logo [2];  [1];  [3];  [1]
  1. Cornell Univ., Ithaca, NY (United States). School of Civil and Environmental Engineering
  2. Pacific Northwest National Lab. (PNNL), Richland, WA (United States). Biological Sciences Division
  3. Univ. of Toronto, ON (Canada). Dept. of Chemical Engineering and Applied Chemistry
Publication Date:
Research Org.:
Pacific Northwest National Lab. (PNNL), Richland, WA (United States)
Sponsoring Org.:
USDOE
OSTI Identifier:
1567199
Report Number(s):
PNNL-SA-147432
Journal ID: ISSN 1664-302X
Grant/Contract Number:  
AC05-76RL01830
Resource Type:
Accepted Manuscript
Journal Name:
Frontiers in Microbiology
Additional Journal Information:
Journal Volume: 10; Journal ID: ISSN 1664-302X
Publisher:
Frontiers Research Foundation
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; reductive dehalogenase, organohalide respiration, Dehalococcoides, Trichloroethene, biomarkers, Geobacter, proteomics, KB-1

Citation Formats

Heavner, Gretchen L., Mansfeldt, Cresten B., Wilkins, Michael J., Nicora, Carrie D., Debs, Garrett E., Edwards, Elizabeth A., and Richardson, Ruth E. Detection of Organohalide-Respiring Enzyme Biomarkers at a Bioaugmented TCE-Contaminated Field Site. United States: N. p., 2019. Web. doi:10.3389/fmicb.2019.01433.
Heavner, Gretchen L., Mansfeldt, Cresten B., Wilkins, Michael J., Nicora, Carrie D., Debs, Garrett E., Edwards, Elizabeth A., & Richardson, Ruth E. Detection of Organohalide-Respiring Enzyme Biomarkers at a Bioaugmented TCE-Contaminated Field Site. United States. doi:10.3389/fmicb.2019.01433.
Heavner, Gretchen L., Mansfeldt, Cresten B., Wilkins, Michael J., Nicora, Carrie D., Debs, Garrett E., Edwards, Elizabeth A., and Richardson, Ruth E. Thu . "Detection of Organohalide-Respiring Enzyme Biomarkers at a Bioaugmented TCE-Contaminated Field Site". United States. doi:10.3389/fmicb.2019.01433. https://www.osti.gov/servlets/purl/1567199.
@article{osti_1567199,
title = {Detection of Organohalide-Respiring Enzyme Biomarkers at a Bioaugmented TCE-Contaminated Field Site},
author = {Heavner, Gretchen L. and Mansfeldt, Cresten B. and Wilkins, Michael J. and Nicora, Carrie D. and Debs, Garrett E. and Edwards, Elizabeth A. and Richardson, Ruth E.},
abstractNote = {RNA-based biomarkers have been successfully detected at field sites undergoing in situ bioremediation, but the detection of expressed enzymes is a more direct way to prove activity for a particular biocatalytic process of interest since they provide evidence of potential in situ activity rather than simply confirming presence and abundance of genes in a given population by measurement of DNA copies using qPCR. Here we successfully applied shotgun proteomics to field samples from a trichloroethene (TCE)-contaminated industrial site in southern Ontario, Canada that had been bioaugmented with the commercially available KB-1TM microbial culture. The KB-1TM culture contains multiple strains of Dehalococcoides mccartyi (D. mccartyi) as well as an organohalide respiring Geobacter species. The relative abundances of specific enzymatic proteins were subsequently compared to corresponding qPCR-derived levels of DNA and RNA biomarkers in the same samples. Samples were obtained from two wells with high hydraulic connectivity to the KB-1TM-bioaugemented enhanced in situ bioremediation system, and two control wells that showed evidence of low levels of native organohalide respiring bacteria (OHRB), Dehalococcoides and Geobacter. Enzymes involved in organohalide respiration were detected in the metaproteomes of all four field samples, as were chaperonins of D. mccartyi, chemotaxis proteins, and ATPases. The most highly expressed RDase in the bioaugmentation culture (VcrA) was the most highly detected enzyme overall in the bioaugmented groundwater samples. In one background groundwater well, we found high expression of the Geobacter pceA RDase. The DNA and RNA biomarkers detected using qPCR-based assays were a set of orthologs of Dehalococcoides reductive dehalogenases (VcrA, TceA, BvcA, dehalogenase “DET1545”), and the Ni-Fe uptake hydrogenase, HupL. Within a sample, RNA levels for key enzymes correlated with relative protein abundance. These results indicate that laboratory observations of TCE-bioremediation biomarker protein expression are recapitulated in field environmental systems and that both RNA and protein biomarker monitoring hold promise for activity monitoring of in situ populations of OHRB.},
doi = {10.3389/fmicb.2019.01433},
journal = {Frontiers in Microbiology},
number = ,
volume = 10,
place = {United States},
year = {2019},
month = {6}
}

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Figures / Tables:

Figure 1 Figure 1: DNA biomarkers detected in filtered groundwater collected from monitoring well PM2A2, extraction well EW1, and background wells O-BH9-A1 and O-BH10-A1. The data in this figure are from DNA extracted off the filters (as opposed to the pre-filters). The detection limit is approximately 100 copies/mL. tceA was below detectionmore » in all samples. Error bars represent average standard deviations of replicate samples (n = 3–6).« less

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    Works referencing / citing this record:

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    journal, September 2006

    • Schefe, Jan H.; Lehmann, Kerstin E.; Buschmann, Ivo R.
    • Journal of Molecular Medicine, Vol. 84, Issue 11
    • DOI: 10.1007/s00109-006-0097-6

    Relationship between Dehalococcoides DNA in ground water and rates of reductive dechlorination at field scale
    journal, September 2006


    Growth and Yields of Dechlorinators, Acetogens, and Methanogens during Reductive Dechlorination of Chlorinated Ethenes and Dihaloelimination of 1,2-Dichloroethane
    journal, April 2007

    • Duhamel, Melanie; Edwards, Elizabeth A.
    • Environmental Science & Technology, Vol. 41, Issue 7
    • DOI: 10.1021/es062010r

    Correlation of Respiratory Gene Expression Levels and Pseudo-Steady-State PCE Respiration Rates in Dehalococcoides ethenogenes
    journal, January 2008

    • Rahm, Brian G.; Richardson, Ruth E.
    • Environmental Science & Technology, Vol. 42, Issue 2
    • DOI: 10.1021/es071455s

    Monitoring Biodegradation of Ethene and Bioremediation of Chlorinated Ethenes at a Contaminated Site Using Compound-Specific Isotope Analysis (CSIA)
    journal, January 2012

    • Mundle, Scott O. C.; Johnson, Tiffany; Lacrampe-Couloume, Georges
    • Environmental Science & Technology, Vol. 46, Issue 3
    • DOI: 10.1021/es202792x

    Relating Chloroethene Respiration Rates in Dehalococcoides to Protein and mRNA Biomarkers
    journal, August 2012

    • Rowe, Annette R.; Heavner, Gretchen L.; Mansfeldt, Cresten B.
    • Environmental Science & Technology, Vol. 46, Issue 17
    • DOI: 10.1021/es300996c

    Bioremediation of Chlorinated Ethenes in Fractured Bedrock and Associated Changes in Dechlorinating and Nondechlorinating Microbial Populations
    journal, May 2014

    • Pérez-de-Mora, Alfredo; Zila, Anna; McMaster, Michaye L.
    • Environmental Science & Technology, Vol. 48, Issue 10
    • DOI: 10.1021/es404122y

    Dehalococcoides ’ Gene Transcripts As Quantitative Bioindicators of Tetrachloroethene, Trichloroethene, and cis -1,2-Dichloroethene Dehalorespiration Rates
    journal, July 2008

    • Rahm, Brian G.; Richardson, Ruth E.
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    Concurrent Ethene Generation and Growth of Dehalococcoides Containing Vinyl Chloride Reductive Dehalogenase Genes During an Enhanced Reductive Dechlorination Field Demonstration
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    • Scheutz, Charlotte; Durant, Neal d.; Dennis, Philip
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    Statistical Analysis of Membrane Proteome Expression Changes in Saccharomyces c erevisiae
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    Large-scale analysis of the yeast proteome by multidimensional protein identification technology
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    Cleaning up with genomics: applying molecular biology to bioremediation
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    journal, January 2018

    • Türkowsky, Dominique; Jehmlich, Nico; Diekert, Gabriele
    • FEMS Microbiology Ecology, Vol. 94, Issue 3
    • DOI: 10.1093/femsec/fiy013

    Experimental validation of novel and conventional approaches to quantitative real-time PCR data analysis
    journal, July 2003


    Organic cofactors in the metabolism of Dehalococcoides mccartyi strains
    journal, April 2013

    • Schipp, Christian J.; Marco-Urrea, Ernest; Kublik, Anja
    • Philosophical Transactions of the Royal Society B: Biological Sciences, Vol. 368, Issue 1616
    • DOI: 10.1098/rstb.2012.0321

    Overview of organohalide-respiring bacteria and a proposal for a classification system for reductive dehalogenases
    journal, April 2013

    • Hug, Laura A.; Maphosa, Farai; Leys, David
    • Philosophical Transactions of the Royal Society B: Biological Sciences, Vol. 368, Issue 1616
    • DOI: 10.1098/rstb.2012.0322

    Genomic, transcriptomic and proteomic analyses of Dehalobacter UNSWDHB in response to chloroform
    journal, August 2016

    • Jugder, Bat-Erdene; Ertan, Haluk; Wong, Yie Kuan
    • Environmental Microbiology Reports, Vol. 8, Issue 5
    • DOI: 10.1111/1758-2229.12444

    Transcription and mass-spectroscopic proteomic studies of electron transport oxidoreductases in Dehalococcoides ethenogenes
    journal, September 2006


    Absolute quantification of Dehalococcoides proteins: enzyme bioindicators of chlorinated ethene dehalorespiration
    journal, October 2009


    Expression of Reductive Dehalogenase Genes in Dehalococcoides ethenogenes Strain 195 Growing on Tetrachloroethene, Trichloroethene, or 2,3-Dichlorophenol
    journal, May 2007

    • Fung, J. M.; Morris, R. M.; Adrian, L.
    • Applied and Environmental Microbiology, Vol. 73, Issue 14
    • DOI: 10.1128/aem.00215-07

    Growth of a Dehalococcoides-Like Microorganism on Vinyl Chloride and cis-Dichloroethene as Electron Acceptors as Determined by Competetive PCR
    journal, July 2003

    • Cupples, A. M.; Spormann, A. M.; McCarty, P. L.
    • Applied and Environmental Microbiology, Vol. 69, Issue 7
    • DOI: 10.1128/aem.69.7.4342.2003

    Transcriptional Expression of the tceA Gene in a Dehalococcoides-Containing Microbial Enrichment
    journal, November 2005


    Comparative metagenomics of three Dehalococcoides-containing enrichment cultures: the role of the non-dechlorinating community
    journal, January 2012


    Metabolism of trichloroethylene.
    journal, May 2000

    • Lash, L. H.; Fisher, J. W.; Lipscomb, J. C.
    • Environmental Health Perspectives, Vol. 108, Issue suppl 2
    • DOI: 10.1289/ehp.00108s2177

    Localized Plasticity in the Streamlined Genomes of Vinyl Chloride Respiring Dehalococcoides
    journal, November 2009


    Biomarkers’ Responses to Reductive Dechlorination Rates and Oxygen Stress in Bioaugmentation Culture KB-1TM
    journal, February 2018


      Figures/Tables have been extracted from DOE-funded journal article accepted manuscripts.