Author

Mouzhong Xu

Date

September 2013

Document Type

Dissertation

Degree Name

Ph.D.

Institution

Oregon Health & Science University

Abstract

Anthropogenic perturbation of global nitrogen cycles, from fossil fuel combustion to excessive N-fertilizer use, has led to a number of environmental problems including eutrophication, acidification, loss of biodiversity, and increased emission of greenhouse gases. The cycling of reactive nitrogen (Nr) is governed by a variety of microbially mediated processes, including nitrification, denitrification, ammonification, dissimilatory nitrate reduction to ammonium (DNRA) and anaerobic ammonia oxidation (anammox).For over a century, aerobic ammonia oxidation, the first and rate-limiting step in nitrification, was thought to be carried out only by ammonia oxidizing bacteria (AOB).Recent studies, however, suggest that ammonia oxidizing archaea (AOA), assigned to the newly designated phylum Thaumarchaeota, are actively involved in global nitrogen and carbon cycles. While Thaumarchaeota are abundant and ubiquitous across diverse ecosystems and appear to have substantive roles in nitrification in marine environments, their contributions to nitrification in terrestrial environments and their related metabolic properties are less well understood. This dissertation examines metabolic properties, phylogenetic diversity, and ecological functions of Thaumarchaeota in terrestrial environments using both cultivation and cultivation-independent molecular techniques. In particular, two terrestrial systems were investigated: freshwater sediments and the plant rhizosphere. In studies on Columbia River freshwater sediments, I employed a combination of analytical and molecular approaches to characterize the biogeochemical gradients and populations of ammonia oxidizers at a fine vertical scale, for the purpose of determining the environmental factors shaping the distribution of AOA and AOB (Chapter 3). Depth profiles of ammonia monooxygenase subunit A (amoA) gene abundance and expressionin AOA and β-AOB, and archaeal amoA diversity were analyzed. AOA generally had both higher abundance of amoA genes and gene transcripts than β-AOB, regardless of season and depth, suggesting potentially important roles of AOA in nitrification. Composition changes in putative AOA populations across steep O[subscript 2] and E[subscript h] gradients were observed, and higher AOA diversity corresponded to steep E[subscript h] gradients. A combination of environmental factors including oxygen, nitrite and nitrate concentrations, and E[subscript h] showed significant correlations with archaeal amoA abundance. Furthermore, to determine potential nitrification activity by AOA and AOB in freshwater sediments, nitrification-coupled growth assays were performed in sediment slurry incubations with the addition of an ammonia oxidation inhibitor allylthiourea (ATU) and a variety of organic substrates (Chapter 4). Results showed that β-AOB were the dominant ammonia oxidizers in sediment incubations with either organic or inorganic amendments, despite the fact that AOA were more abundant than β-AOB in natural sediments. These results suggested that some Thaumarchaeota might not be autotrophic ammonia oxidizers, and may have alternative ecological strategies, such as heterotrophic or mixotrophic metabolisms, to maintain their high abundance in freshwater sediments. Using a plant-root enrichment culture system, we characterized the diversity and metabolic potential of mesophilic soil Thaumarchaeota selected with organic and inorganic amendments (Chapter 5). Comparative analysis of 16S rRNA and amoA genes indicated that specific archaeal clades were selected under different conditions, whereas AOB were not detected in this enrichment. Three amoA-containing clades were identified, while a fourth clade identified by 16S rRNA gene analysis alone, designated the “rootclade,” yielded no corresponding amoA gene with the primers used. Analysis of archaeal community composition by PCR-single stranded conformation polymorphism (PCRSSCP) under different culture conditions revealed that the root clade was present only in media with organic amendment, while amoA–containing clades were present in media with either organic or inorganic amendment. We hypothesized that the root clade archaea were heterotrophs, assimilating organic carbon in root extract as substrates. Furthermore, results from gene abundance and expression analyses, together with potential nitrification activity assays suggested differential contributions by the clades to nitrification in our system. Taken together, our results indicate diverse metabolic lifestyles in soil Thaumarchaeota.

Identifier

doi:10.6083/M4125QNX

Division

Institute of Environmental Health

School

School of Medicine

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