Document Type

Dissertation

Date of Degree

Fall 2016

Access Restrictions

Access restricted until 02/23/2018

Degree Name

PhD (Doctor of Philosophy)

Degree In

Civil and Environmental Engineering

First Advisor

Timothy E. Mattes

Abstract

Vinyl chloride (VC) is a human carcinogen and common groundwater contaminant in the United States. Some of the indigenous bacteria can utilize VC for their growth, which is important for bioremediation. As previous studies have been majorly focused on VC-degrading bacteria in pure cultures, we initiated the study to investigate the microbial community structure and interactions in more complicated systems, such as mixed-pure cultures and groundwater enrichment cultures. Finally, we extended our study into the field by investigating the diversity and abundance of functional genes in VC-assimilating pathways at six contaminated sites.

In our first study, Nocardioides was found to be the most dominant genus in Carver groundwater enrichment cultures via stable-isotope probing and 16S rRNA gene amplicon Illumina sequencing. As cross-feeding was observed, in the second study, mixed-pure culture experiment was conducted to explore the potential effects of VC-assimilating Nocardioides on other bacteria, which showed VC cometabolizer Mycobacterium strain JS622 would take up carbon from VC to sustain their growth when mixed with VC-assimilating Nocardioides sp. strain JS614. The third study was conducted with a different groundwater source from Fairbanks, AK, which again showed Nocardioides is dominant in the microbial community. A novel VC-assimilating Nocardioides sp. bacteria was isolated, named XL1. The putative genome of XL1 extracted from enrichment culture metagenome was 99% to 100% identical to strain JS614, with a plasmid genome bin similar to strain JS614 plasmid pNOCA01, though the morphology of strain XL1 was distinct from strain JS614. About 90% of the plasmid contigs could be mapped onto Nocardioides sp. strain JS614 plasmid with 100% identity, containing known aerobic ethene and VC degradation pathway genes encoding alkene monooxygenase and epoxyalkane: coenzyme M transferase (EaCoMT). Glutathione synthase and glutathione S-transferase genes, possibly involved in epoxide detoxification, were found in Polaromonas, Mesorhizobium and Pseudomonas-affiliated genome bins. The study also showed cultures adapted to VC faster after amended with ethene. The in-situ study (the fourth study) revealed 192 different EaCoMT T-RFs from six chlorinated ethene contamination sites via T-RFLP analysis, implicating higher EaCoMT diversity than previously known. Phylogenetic analysis revealed that a majority of the 139 cloned sequences (78.4%) grouped with EaCoMT genes found in VC- and ethene-assimilating Mycobacterium strains and Nocardioides sp. strain JS614. EaCoMT gene abundance was positively correlated with VC and ethene concentrations at the sites studied.

Keywords

Adaptation, Bacteria Diversity, Bioremediation, Groundwater, Metagenomics, Vinyl Chloride

Pages

xviii, 234 pages

Bibliography

Includes bibliographical references (pages 222-234).

Copyright

Copyright © 2016 Xikun Liu

Available for download on Friday, February 23, 2018

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