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Deep-Proteome Analysis of Bioremediation Cultures Containing Dehalococcoides: from Gene Network Elucidation to Biomarker Quantification in Field Samples


EMSL Project ID
40006

Abstract

The prevalence of chlorinated organic compounds as environmental pollutants, in addition to their potential detoxification through biological pathways, has led to the development of bioremediation strategies that focus on facilitating in situ health and activity of populations of Dehalococcoides (DHC) bacteria. All known DHC strains are obligate dehalorespirers (using halogenated organics as electron acceptors), and several have been shown to transform these pollutants to non-toxic end products (e.g. ethene from perchloroethene and trichloroethene). However, the different strains appear to have distinct substrate ranges--likely due to variability in presence and expression of different reductive dehalogenase genes. Often, DHC activity in the field stalls despite excess addition of electron donor to biostiumulate the populations. As there are still many unknowns with respect to DHC biology and the microbial ecology of the communities in which they robustly grow (mixed cultures support ten-fold higher DHC cell densities than pure cultures) better understanding of these organisms could lead to more effective treatment and biostimulation strategies. Our main aim in the proposed work is to improve the understanding of DHC systems biology and to leverage this understanding to develop a suite of robust biomarkers to monitor DHC health and activity in environmental systems. Over the past two years, our lab has conducted continuous-feed experiments on two DHC-containing mixed cultures including KB1, a commercially-available bioaugmentation culture that has been deployed at dozens of bioremediation sites. Lab cultures are grown under varying combinations of chlorinated electron acceptor, rate of electron acceptor feed, electron donor-to-acceptor feed ratios, and cell stressors (oxygen and pH being the most field-pertinent). We propose to utilize EMSL facilities to conduct deep metaproteome characterization of these culture samples via the accurate mass and time (AMT) tag approach. Combined with our lab's existing heterologous datasets for DNA, RNA, metabolites, respiration rate and metagenome sequences the new proteome datasets will be analyzed using probabilistic network inference approaches (to be partially performed with EMSL computational resources) to highlight key gene networks that correlate with specific respiration reactions, respiration rates, electron donor limitation, pH stress and oxygen stress (the phenotypes of interest). Using the inferences made by these models, we will then select proteins with strong correlation with phenotypes of interest for development as field biomarkers. This suite of biomarkers will then be quantified in cultures using a technique we recently leveraged from the small molecule quantification field (multiple reaction monitoring (MRM)) using EMSL Triple Quadrupole MS resources. These MRM assays will ultimately be tested on biomass samples from field sites where a range of bioremediation strategies are being employed. Ultimately, these biomarkers could be used by site managers to document in situ bioremediation to federal regulators and, if bioremediation is stalling, suggest which specific environmental stressors need to be mitigated.

Project Details

Project type
Large-Scale EMSL Research
Start Date
2010-10-01
End Date
2013-09-30
Status
Closed

Team

Principal Investigator

Ruth Richardson
Institution
Cornell University

Team Members

Benjamin Logsdon
Institution
Cornell University

Gretchen Heavner
Institution
Cornell University

Annette Rowe
Institution
Cornell University

Related Publications

Heavner GL, AR Rowe, CB Mansfeldt, JK Pan, JM Gossett, and RE Richardson. 2013. "Molecular Biomarker-Based Biokinetic Modeling of a PCE-Dechlorinating and Methanogenic Mixed Culture." Environmental Science & Technology 47(8):3724-3733. doi:10.1021/es303517s
Heavner G.L., C.B. Mansfeldt, G.E. Debs, S.T. Hellerstedt, A.R. Rowe, and R.E. Richardson. 2018. "Biomarkers’ Responses to Reductive Dechlorination Rates and Oxygen Stress in Bioaugmentation Culture KB-1TM." Microorganisms 6, no. 1:13. doi:10.3390/microorganisms6010013
Heavner G.L., C.B. Mansfeldt, M.J. Wilkins, C.D. Nicora, G.E. Debs, E.A. Edwards, and R.E. Richardson. 2019. "Detection of Organohalide-Respiring Enzyme Biomarkers at a Bioaugmented TCE-Contaminated Field Site." Frontiers in Microbiology 10. PNNL-SA-147432. doi:10.3389/fmicb.2019.01433
Rowe AR, CB Mansfeldt, GL Heavner, and RE Richardson. 2015. "Relating mRNA and Protein Biomarker Levels in a Dehalococcoides and Methanospirillum-containing Community." Applied Microbiology and Biotechnology 99(5):2313-2327. doi:10.1007/s00253-014-6220-7