Collaborative Research: Stable isotope-based differentiation of vinyl chloride assimilators from cometabolizers in contaminated groundwater

1233087/1233154 Mattes/Cupples氯乙烯（VC）是一种已知的人类致癌物质，也是一种常见的地下水污染物。本研究项目的重点是确定和表征参与VC生物修复现场的关键微生物。VC羽流是在厌氧条件下通过氯化溶剂的还原脱氯产生的，但通常会逃逸到向下梯度的好氧地下水区域。好氧，生长耦合VC氧化（即VC同化）细菌很容易从VC污染的网站，支持的想法，他们发挥了作用的降解。然而，直接证据表明，VC同化细菌的存在和活性在原位仍然难以捉摸。此外，目前还不清楚VC同化分离株是地下的优势微生物，或者它们的培养是富集偏倚的结果。此外，VC同化物不能与乙烯氧化物的较大相关组区分开来，乙烯氧化物是偶然的VC氧化物。为了解决这些知识差距，本研究将使用稳定同位素探测（SIP）来区分VC同化细菌和偶然氧化VC的细菌（即共代谢者）。SIP涉及在生长偶联生物降解期间将13 C标记的底物同化成核酸。之后，分析13 C标记的核酸以鉴定负责污染物降解的生物体。中心的假设是，SIP技术，当与现有的分子工具结合使用时，将区分ethenetrophs和VC同化在实验室和现场应用。研究涉及以下具体目标：i）使用来自VC污染场地的土壤和/或地下水样品，并应用DNA-SIP来揭示VC同化微生物的存在、身份和活性，ii）使用VC同化和共代谢实验室培养物来开发基于SIP的测定，该测定区分这些密切相关的微生物组，以及iii）提供VC-同化细菌在原位存在并活跃。有毒污染物对地下水的污染是一个严重的社会问题，威胁着人类健康和环境。原地生物修复实践有望将地下水恢复到污染前的水平。旨在更好地了解生物降解微生物群落的存在、丰度和活动的研究将有助于在基础科学和生物修复实践之间架起桥梁。这项研究的长期目标是通过开发分子工具来表征环境中降解污染物的微生物，推进生物修复领域。这项研究将涉及对环境工程本科生和研究生的培训，使他们能够将分子生物学工具应用于环境问题。这项工作将增加代表性不足的少数民族学生的人数，还将涉及发展K-12外联活动。该研究将通过解决地下水污染物造成的水质人为恶化而造福社会。

1233087/1233154Mattes/CupplesVinyl chloride (VC) is a known human carcinogen and a common groundwater contaminant. This research project focuses on identifying and characterizing the key microorganisms involved in VC bioremediation at field sites. VC plumes are generated under anaerobic conditions via reductive dechlorination of chlorinated solvents, but often escape into downgradient aerobic groundwater zones. Aerobic, growth-coupled VC-oxidizing (i.e. VC assimilating) bacteria are readily isolated from VC-contaminated sites, supporting the idea that they play a role in the degradation. However, direct evidence that VC-assimilating bacteria are present and active in situ remains elusive. Further, it is unclear if VC-assimilating isolates are the dominant microbes in the subsurface or if their cultivation is a result of enrichment bias. Also, VC-assimilators cannot be distinguished from the larger related group of ethene-oxidizers, which are fortuitous VC oxidizers. To address these knowledge gaps, this research will use stable isotope probing (SIP) to differentiate VC-assimilating bacteria from bacteria that fortuitously oxidize VC (i.e. the cometabolizers). SIP involves microbial assimilation of 13C labeled substrates into nucleic acids during growth-coupled biodegradation. Following this, 13C labeled nucleic acids are analyzed to identify the organisms responsible for contaminant degradation. The central hypothesis is that SIP techniques, when used in conjunction with existing molecular tools, will differentiate between etheneotrophs and VC-assimilators in both laboratory and field-based applications. The research involves the following specific aims: i) to use soil and/or groundwater samples from VC-contaminated sites andapply DNA-SIP to reveal the presence, identity, and activity of VC-assimilating microorganisms, ii) to use both VC-assimilating and cometabolizing laboratory cultures to develop a SIP-based assay that differentiates between these closely related microbial groups and iii) to provide direct evidence that VC-assimilating bacteria are present and active in situ. Groundwater contamination by toxic pollutants is a major societal problem that threatens human health and the environment. In situ bioremediation practice hold promise for restoring groundwater to pre-contamination levels. Studies aimed at better understanding the presence, abundance and activity of biodegrading microbial communities will serve to bridge fundamental science and the practice of bioremediation. The long-term goal of this research is to advance the bioremediation field by developing molecular tools to characterize pollutant-degrading microorganisms in the environment. The research will involve the training of undergraduate and graduate environmental engineering students to apply molecular biology tools to environmental problems. The work will increase the number of underrepresented minority students and will also involve the development of K-12 outreach activities. The research will benefit society by addressing anthropogenic deterioration of water quality by groundwater pollutants.