Doctor of Philosophy, The Ohio State University, 2022, Biochemistry Program, Ohio State

The bacterial phylum, Bacteroidetes, is a remarkably diverse and ubiquitous lineage whose members are profoundly integral to the ecological niches they occupy such as the mammalian gut, the rhizosphere, and the ocean microbiome. They also possess a wealth of notable features that distinguishes them from other groups of bacteria. Despite their apparent ecological importance and the intriguing characteristics that they possess, they remain a poorly studied group. This deficiency in characterization is readily evident with regards to translation initiation in the Bacteroidetes, which is contrary to our common understanding of initiation in more well-studied bacteria. Among prokaryotic genomes, many protein coding genes have a Shine Dalgarno (SD) motif proximal to the start codon which serves to recruit ribosomes to that site. Translation initiation is often facilitated by pairing of the SD sequence to its motific counterpart, the anti-SD (ASD), on the 16S rRNA of the ribosome. This SD-ASD interaction was thought to be the predominant mechanism of translation initiation among bacteria and in many cases indispensable. Yet, SD sequences are greatly underrepresented in the genomes of Bacteroidetes, suggesting that the SD-ASD mechanism has become largely irrelevant. In the absence of SD sequences, alternative translational determinants must substitute and be more prominent in the Bacteroidetes. In this study, we identify translational determinants in the Bacteroidetes that supplant the SD motif. We observe similar alternative determinants in other organisms such as Escherichia coli that do use the SD sequences, but less so in organisms such as B. subtilis which relies heavily on SD sequences. However, what remains uncertain is why the Bacteroidetes lineage has largely dispensed with the SD sequence. Another intriguing aspect of the Bacteroidetes is that their ribosomes virtually retain the complete ASD sequence. The increasing irrelevance of the SD sequence should lessen t (open full item for complete abstract)

Committee: Kurt Fredrick (Advisor) Subjects: Biochemistry; Genetics; Microbiology; Molecular Biology

Doctor of Philosophy, The Ohio State University, 2018, Microbiology

Organic carbon in anoxic ecosystems flows in a cascade from complex plant material to more labile sugars, and ultimately to short-chain fatty acids (SCFA) and gasses like carbon dioxide and methane. Microbial communities, groups of microorganisms that interact with one another, facilitate this process. Microbial anaerobic carbon degradation is exemplified in ruminants. These animals harness energy from plant material using the power of interacting microorganisms, which break down plant carbon into SCFA under largely anoxic conditions in the rumen. Because microbial SCFA can provide up to 80% of the animal's energy, understanding microbial carbon degradation mechanisms in the rumen is important for many agricultural industries including the production of meat, milk, leather, and wool. Beyond domesticated ruminants, there are over 75 million wild ruminants that are fundamental members in ecosystems from Alaska to Australia. Furthermore, the microbial enzymes that break down plant material in the rumen have industrial applications for modifying enzymatic cocktails in biofuel production. The research presented here uses cultivation-independent and laboratory approaches to assign carbon degradation capabilities to specific members of the microbial community in the moose rumen. Moose, animals that naturally forage on woody biomass, were selected to provide access to natural rumen microbial communities that are especially adapted to a high lignocellulose diet. We sampled rumen fluid from moose in the spring, summer, and winter, along a seasonal gradient in lignocellulose. Rumen fluid was sampled via the rumen cannula, offering access into the active microbial interactions mediating complex carbon degradation. From these rumen fluid samples, we performed high-throughput shotgun metagenomics and metaproteomics, coupled to multiple methods for metabolite quantification (1H NMR, sequential fiber analyses, and carbohydrate microarray polymer profiling (CoMPP)). We binned hun (open full item for complete abstract)

Committee: Kelly Wrighton Ph.D. (Advisor); Venkat Gopalan Ph.D. (Committee Member); Jeffrey Firkins Ph.D. (Committee Member); Daniel Wozniak Ph.D. (Committee Member) Subjects: Microbiology

MS, University of Cincinnati, 2006, Engineering : Environmental Engineering

Recently, 16S rDNA Bacteroidetes-targeted PCR assays were developed to discriminate between ruminant and human fecal pollution. These assays are rapid and relatively inexpensive but have been used in a limited number of environmental applications. In this study, we evaluated the efficacy of human- and ruminant-specific 16S rDNA Bacteroidetes assays in determining the primary sources of fecal pollution in Plum Creek (NE) watershed. The sensitivity and specificity, as well as the temporal and spatial application of these assays were challenged against feces from different animals, water, and sediment samples from this watershed. Phylogenetic analyses of 981 fecal and environmental 16S rDNA clones were also performed to study the diversity of Bacteroidetes in this watershed. On average, the host specific assays indicated that ruminant feces were present in more than one-third of the water samples and in all sampling seasons, with increasing frequency in downstream sites along Plum Creek. The human-targeted assay indicated that only 5% of the water samples were positive for human fecal signals, although a higher percentage of human-associated signals (i.e., 24%) were detected in sediment samples. Phylogenetic analysis of Bacteroidetes 16S rDNA sequences derived from fecal and environmental samples demonstrated the presence of a high level of sequence diversity, with nearly half of the recovered sequences showing less than 97% identity to sequences found in publicly available databases. Approximately, 60% of all clones clustered with yet to be cultured Bacteroidetes species associated with sequences obtained from ruminant feces, further supporting the prevalence of ruminant contamination in this watershed. The preponderance of clones affiliated with uncultured Bacteroidetes strongly suggests that the bacterial diversity of this group is not sufficiently represented by the culturable members for which sequence data are available. The sequencing data also indicated that ot (open full item for complete abstract)

Committee: Dr. Daniel Oerther (Advisor) Subjects: Environmental Sciences