Scholarship & Research
Permanent URI for this communityhttps://scholarworks.montana.edu/handle/1/1
Browse
9 results
Search Results
Item Rock powered life in the Samail ophiolite: an analog for early Earth(Montana State University - Bozeman, College of Agriculture, 2021) Fones, Elizabeth Marie; Chairperson, Graduate Committee: Eric Boyd; Daniel R. Colman, Emily A. Kraus, Daniel B. Nothaft, Saroj Poudel, Kaitlin R. Rempfert, John R. Spear, Alexis S. Templeton and Eric S. Boyd were co-authors of the article, 'Physiological adaptations to serpentinization in the Samail ophiolite, Oman' in the journal 'The International Society for Microbial Ecology journal' which is contained within this dissertation.; Daniel R. Colman, Emily A. Kraus, Ramunas Stepanauskas, Alexis S. Templeton, John R. Spear and Eric S. Boyd were co-authors of the article, 'Diversification of methanogens into hyperalkaline serpentinizing environments through adaptations to minimize oxidant limitation' in the journal 'The International Society for Microbial Ecology journal' which is contained within this dissertation.; David W. Mogk, Alexis S. Templeton and Eric S. Boyd were co-authors of the article, 'Endolithic microbial carbon cycling activities in subsurface mafic and ultramafic igneous rock' which is contained within this dissertation.Serpentinization is a geochemical process wherein the oxidation of Fe(II)-bearing minerals in ultramafic rock couples with the reduction of water to generate H 2, which in turn can reduce inorganic carbon to biologically useful substrates such as carbon monoxide and formate. Serpentinization has been proposed to fuel a subsurface biosphere and may have promoted life's emergence on early Earth. However, highly reacted waters exhibit high pH and low concentrations of potential electron acceptors for microbial metabolism, including CO 2. To characterize how serpentinization shapes the distribution and diversity of microbial life, direct cell counts, microcosm-based activity assays, and genomic inferences were performed on environmental rock and water samples from the Samail Ophiolite, Oman. Microbial communities were shaped by water type with cell densities and activities generally declining with increasing pH. However, cells inhabiting highly reacted waters exhibited adaptations enabling them to minimize stresses imposed by serpentinization, including preferentially assimilating carbon substrates for biomolecule synthesis rather than dissimilating them for energy generation, maintaining small genomes, and synthesizing proteins comprised of more reduced amino acids to minimize energetic costs and maximize protein stability in highly reducing waters. Two distinct lineages of a genus of methanogens, Methanobacterium, were recovered from subsurface waters. One lineage was most abundant in high pH waters exhibiting millimolar concentrations of H2, yet lacked two key oxidative [NiFe]-hydrogenases whose functions were presumably replaced by formate dehydrogenases that oxidize formate to yield reductant and CO 2. This allows cells to overcome CO 2/oxidant limitation in high pH waters via a pathway that is unique among characterized Methanobacteria. Finally, gabbro cores from the Stillwater Mine (Montana, U.S.A) were used to develop methods for detecting the activities of cells inhabiting mafic to ultramafic igneous rocks while controlling for potential contaminants. Optimized protocols were applied to rock cores from the Samail Ophiolite, where rates of biological formate and acetate metabolism were higher in rocks interfacing less reacted waters as compared with more extensively reacted waters, and in some cases may greatly exceed activities previously measured in fracture waters. This dissertation provides new insights into the distribution, activities, and adaptations exhibited by life in a modern serpentinizing environment.Item Structure and function of a prokaryotic argonaute from Pseudomonas aeruginosa(Montana State University - Bozeman, College of Agriculture, 2020) Erickson, Reece Sheridan; Chairperson, Graduate Committee: Blake WiedenheftArgonautes (Ago) are structurally and functionally diverse proteins present in all domains of life. A common feature of these ancient proteins is their ability to bind nucleic acid guides that target the protein to complementary sequences. Although eukaryotic argonautes (eAgo) have been well-studied, we still know very little about the function of prokaryotic argonautes (pAgo) in bacterial and archaeal species. To address this gap in our knowledge, my thesis focused on determining the biochemical properties as well as the cellular functions of a pAgo from the organism Pseudomonas aeruginosa PACS2 (PaAgo). Here, we show that PaAgo plays a role in regulating the expression of transposons within PACS2. I also present results indicating that deletion of the PaAgo gene and its neighboring genes causes toxicity to P. aeruginosa. Finally, I provide evidence that PaAgo and a neighboring protein are binding partners and form a multi-protein complex. Future work will focus on copurifying and sequencing PaAgo nucleic acid guides as well as clarifying the mechanisms guide acquisition and biological function.Item Virus host interactions at the single cell level in hot springs of Yellowstone National Park(Montana State University - Bozeman, College of Letters & Science, 2019) Munson-McGee, Jacob Hampton; Chairperson, Graduate Committee: Mark J. Young; Jamie C. Snyder and Mark J. Young were co-authors of the article, 'Introduction to archaeal viruses' in the journal 'Genes' which is contained within this dissertation.; Ross Hartman was an author and Mark J. Young were co-authors of the article, 'vFish for the quantification of viral infection in natural environments' submitted to the journal 'Environmental microbiology' which is contained within this dissertation.; Erin K. Field, Mary Bateson, Colleen Rooney, Ramunas Stepanauskas and Mark J. Young were co-authors of the article, 'The identification and characterization of a nanoarchaeota, its cellular host and a nanoarchaeal virus across Yellowstone National Park hot springs' which is contained within this dissertation.; Colleen Rooney and Mark J. Young were co-authors of the article, 'An uncultivated virus infecting a nanoarchaeal parasite in the hot springs of Yellowstone National Park' submitted to the journal 'Virology' which is contained within this dissertation.; Shengyun Peng, Samantha Dewerff, Ramunas Stepanauskas, Rachel J. Whitaker, Joshua Weitz and Mark J. Young were co-authors of the article, 'A virus or more in (nearly) every cell: ubiquitous networks of virus-host interactions in extreme environments' in the journal 'The ISME journal' which is contained within this dissertation.Viruses are the most abundant biological entities on the planet and virus-host interactions are some of the most important factors in shaping microbial community structure and function and global chemical cycling. The high temperature low pH hot spring of Yellowstone National Park contain simplified microbial communities of 8-10 Archaeal species, and comparatively simple viral communities. These idealized communities that contain only viruses and their Archaeal hosts represent a model natural environment for the study of viruses and their hosts. This work presented here builds on previous population level studies of the viral and microbial communities to examine virus-host interactions at the single cell level. The identification of viral infection has long been a scourge of environmental virologist. In order to identify viral infection in natural environments we have adapted Fluorescent in situ hybridization (FISH) techniques to directly identify viral sequences. We further advance this technique to be compatible with flow cytometry analysis for the rapid quantification of viral infection of uncharacterized viruses in natural environments. This technique is used to quantify viral infection of two different viruses, previously only characterized by metagenomic sequencing analysis, in four geographically separate low pH high temperature hot springs of Yellowstone National Park. Finally, we combine viral and cellular metagenomics with cellular transcriptomics and single cell genomics to identify virus host interactions at the single cell level and identify viruses that are replicating in the hot springs. This work suggests that a majority of cells in the hot springs are interacting with viruses and that a majority of the cells are interacting with multiple viruses at any given time. We also identify RNA sequences from a majority of the viral types present in the hot springs suggesting that viral replication is occurring and is an important force in determining the structure and function of the microbial communities in these hot springs. Together these works represent a significant advancement of our understanding of virus host interactions in natural environments as well as new techniques to be used in future studies.Item Partitioning of reactive oxygen species via the re-oxidation of electron transfer flavoprotein(Montana State University - Bozeman, College of Letters & Science, 2019) Austvold, Chase Kennor; Chairperson, Graduate Committee: Edward DratzThe biology of Reactive Oxygen Species are poorly understood. Within a healthy cell, Reactive Oxygen Species behave as signaling molecules, although overproduction leads to oxidative damage. In order to understand when the overproduction of Reactive Oxygen Species takes place, or leads to oxidative damage, the elementary step of quantification becomes necessary. Electron Transfer Flavoprotein is a known Reactive Oxygen Species producing enzyme and was studied. Electron Transfer Flavoprotein is a key-player within the production of energy within the eukaryotic mitochondria. The redox nature of Electron Transfer Flavoprotein's catalytic cofactor, flavin adenine dinucleotide produced two types of ROS; the superoxide anion (O 2 °-) and hydrogen peroxide (H 2 O 2). Electron Transfer Flavoprotein produced roughly five-fold more O 2 °-compared to H 2 O 2 as the enzyme became oxidized. It has been put forward that the production of these two Reactive Oxygen Species is dictated by the formation of a radical pair between the flavin adenine dinucleotide of Electron Transfer Flavoprotein and molecular oxygen. Two types of radical pairs can be formed, either in a triplet or singlet state, and the rate in which these states occur can be influenced by a static magnetic field. Therefore, the effect of a magnetic field on these products was also studied. Upon the suppression of magnetic field strength, the production of H 2 O 2 decreased and a proportional increase of O 2 °-was observed.Item Investigation of a control strategy for manipulation and prevention of Pseudomonas aeruginosa PAO1 biofilms in metalworking fluids(Montana State University - Bozeman, College of Engineering, 2018) Ozcan, Safiye Selen; Chairperson, Graduate Committee: Christine Foreman; Markus Dieser, Albert E. Parker, Narayanaganesh Balasubramanian and Christine M. Foreman were co-authors of the article, 'Quorum sensing inhibition as a promissing method to control biofilm growth in metalworking fluids' submitted to the journal 'Environmental science & technology' which is contained within this thesis.Microbial contamination in metalworking fluid (MWF) circulation systems is a serious problem. Particularly water based MWFs promote microbial colonization despite the use of biocides. Inhibiting the quorum sensing mechanism (i.e. cell-cell communication) in bacteria is a promising approach to control and prevent biofilm formation. The objective of this study was (i) to determine the microbial community in MWFs from operational machining shops, (ii) to investigate the effect of well-known quorum sensing inhibitors on controlling biofilm formation, and (iii) to implement experimental data from selected enzymes to a computer simulation biofilm accumulation model (BAM). Planktonic and biofilm samples from two local machining shops in Bozeman, MT, were collected to determine the extent of microbial colonization. In both operations, microbial communities were dominated by Pseudomonadales (60.2-99.7%). Rapid recolonization was observed even after dumping spent MWFs and cleaning. Considering the dominance of Pseudomonadales in MWFs, the model organism Pseudomonas aeruginosa PAO1 was selected for testing the effects of quorum sensing inhibitor compounds on biofilm formation. From a variety of enzymes, natural, and chemical compounds screened for quorum sensing inhibition, Patulin (40microns) and Furanone C-30 (75microns), were found to be effective in reducing biofilm formation in MWFs when applied as single compound amendments and in combination with the polysaccharide degrading enzyme alpha-amylase from Bacillus amyloliquefaciens. Particularly Furanone C-30, as a single amendment and in combination with alpha-amylase decreased biofilm formation by 76% and 82% after 48 hours. Putatively identified homoserine lactones in MWFs treated with Furanone C-30 provided evidence for quorum sensing inhibition on biofilm formation. BAM was employed to study the effect of alpha-amylase (3 Units mL -1) on P. aeruginosa PAO1 biofilms in batch reactors for 24 and 48 hours. In the absence of alpha-amylase, biofilm thickness was predicted to be 23.11 and 31.37 microns, while its presence reduced thickness to 10.47 and 13.07 microns after 24 and 48 hours, respectively. The results presented herein highlight the potential effectiveness of quorum sensing inhibition as a strategy to reduce biofilms in MWFs.Item Effects of carboxin, an oxathiin systemic fungicide, on cell permeability and DNA dependent RNA synthesis(Montana State University - Bozeman, College of Agriculture, 1971) Shively, Owen DanielItem Regulation of leukocyte L-selectin expression(Montana State University - Bozeman, College of Agriculture, 1993) Palecanda, Aiyappa MuthannaItem Demonstration of the leukocyte-endothelial cell adhesion cascade involved in lymphocyte recirculation and neutrophil and lymphocyte localization to sites of inflammation(Montana State University - Bozeman, College of Agriculture, 1994) Bargatze, Robert FrostItem Risk assessment of plant-based pharmaceuticals and biologics(Montana State University - Bozeman, College of Agriculture, 2006) Shama, Leslie Marie; Chairperson, Graduate Committee: Robert K. D. Peterson.Biotechnology is evolving to produce pharmaceutical proteins in plants. Plantbased pharmaceutical production creates concerns of exposure in an open environment and contamination of the food supply. Consequently, quantitative human health and ecological risk assessments were conducted for aprotinin, gastric lipase, and LT-B expressed in maize. A comparative, qualitative risk assessment was conducted for conventionally derived and plant-cell derived Newcastle disease virus vaccine. Effect and exposure scenarios were modeled for each quantitative risk assessment and each scenario was based on a tiered approach in which inadvertent exposure through ingestion were examined to determine a risk characterization of the plant-based pharmaceuticals. The qualitative Newcastle disease virus vaccine risk assessment characterized risks based on the potential exposures to the poultry being vaccinated, to humans administering the vaccine, and to non-target birds. For the human-health risk assessment, the dietary exposure evaluation model (DEEM) was used to estimate the inadvertent dietary intake of the pharmaceutical proteins in food. The ecological risk assessment used Monte Carlo simulations to evaluate the exposure of each protein in maize for four receptor species.