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Document Type

Open Access Dissertation

Degree Name

Doctor of Philosophy (PhD)

Degree Program

Microbiology

Year Degree Awarded

2015

Month Degree Awarded

September

First Advisor

James F Holden

Subject Categories

Environmental Microbiology and Microbial Ecology

Abstract

This dissertation examined the substrate and energetic limitations of hydrogenotrophic thermophiles from deep-sea hydrothermal vents. Thermophilic and hyperthermophilic organisms in diffuse hydrothermal venting are thought to represent a hot subsurface biosphere associated with deep-sea hydrothermal vents, where primary production is dominated by hydrogenotrophy rather than sulfide oxidation as at the vent/seawater interface of hydrothermal sulfide chimneys. Methanogens and sulfur-reducers are known to compete for hydrogen in mesophilic, freshwater systems, and likely do so in deep-sea hydrothermal vent environments as well. However, the exact size and biomass of the subsurface biosphere is difficult to determine through direct sampling.

Firstly, the distribution of thermophilic and hyperthermophilic methanogens, sulfur-reducers, and heterotrophs in diffuse venting fluids at our field site, Axial Volcano (on the Juan de Fuca Ridge), was examined using culture-dependent (Most-Probable-Number) and independent (omics) techniques. It was confirmed that methane production in diffuse venting fluids could be stimulated by the sole addition of hydrogen and incubation at thermophilic and/or hyperthermophilic temperatures, indicating that methanogens in this system are not limited significantly by nutrient or trace element requirements. To determine why one novel hyperthermophilic methanogen from our field site (Methanocaldococcus bathoardescens) appeared to prefer high levels of nitrogen when grown in the lab, its genome was examined for nitrogen assimilation-related genes.

In the laboratory, the growth energies of Methanocaldococcus and Methanothermococcus spp. over their full temperature ranges were measured in order to determine Arrhenius constants for their production of methane. They were also grown in continuous flow chemostat culture to determine their hydrogen limitations at both optimal and sub-optimal temperatures for growth, and the Monod kineticsfor their hydrogen use and methane production were measured. Additionally, the minimum hydrogen and thiosulfate requirements, as well as Monod kinetics, were measured in batch bioreactor culture for a thiosulfate-reducing, hydrogenotrophic, thermophilic Desulfurobacterium sp. isolated from another site on the Juan de Fuca Ridge, the Endeavour Segment, to determine where it might compete with methanogens for hydrogen.

Finally, the geochemical and distribution data from Axial Volcano and laboratory-derived kinetic data for thermophilic and hyperthermophilic methanogens were used to create a one-dimensional reactive transport model (RTM) of hydrogenotrophic methanogenesis in the subsurface at Axial Volcano. In this way, the relative dimensions and biomass of methanogens in the subsurface can be predicted without direct sampling. In future, this type of model could be used make predictions about the thermophilic subsurface at other vent locations, as well as expanded to include competition between different types of hydrogenotrophs (rather than just hydrogenotrophic methanogens with different optimum temperatures) and interactions with other organisms, such as hydrogen-producing hyperthermophilic heterotrophs.

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