Scholarship & Research
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Item Single cell encapsulation, detection, and sorting of Pseudomonas syringae using drop-based microfluidics(Montana State University - Bozeman, College of Engineering, 2023) Lindsay, Travis Carson; Chairperson, Graduate Committee: Abigail Richards; Connie Chang (co-chair)Bacteria can survive antibiotic or bactericidal treatment through genetic mutations. Even within bacterial populations that are fully susceptible to treatment, a small proportion of cells can have enhanced survival capacity in a phenomenon called persistence. Traditional microbiology methods can fail to identify or isolate these persister cells present within the population. A novel method for high-throughput single cell analyses of microbial populations is that of drop-based microfluidics, in which individual cells can be isolated within picoliter-sized drops. In this work, fluorescent detection and dielectrophoresis-based sorting of drops was developed for isolating Pseudomonas syringae persister cells following antimicrobial treatment. We demonstrate: (1) the dielectrophoresis-based sorting of dye-filled 25 micron drops based upon two colors, (2) differences between laser-induced fluorescent detection of dyes compared to single bacterial cells, (3) single-cell isolation of P. syringae into 25 micron droplets with ~10% of droplets containing singlecells, and (4) the treatment, staining, and fluorescent characterization of P. syringae at 0.5x, 5x, and 50x the minimum inhibitory concentration of carbonyl cyanide m-chlorophenyl hydrazone (CCCP), an antibiotic which resulted in 6.2%, 10.2%, and 88.6% cell death of the population, respectively. These results provide the groundwork for studying antibiotic-treated P. syringae and the isolation of surviving cells that will lend insight into the molecular basis of persistence for preventing recurrent infections and decreasing the likelihood of antibiotic resistance.Item The influence of growth rate and cell concentration on bacterial attachment to surfaces in a continuous flow system(Montana State University - Bozeman, College of Engineering, 1983) Nelson, Christopher HenryItem Etiology and characterization of two Pseudomonas syringae pathovars causing two bacterial kernel blights of barley(Montana State University - Bozeman, College of Agriculture, 1994) Martinez-Miller, ConcepcionItem Reactor optimization of volatilized p-xylene metabolism(Montana State University - Bozeman, College of Engineering, 1993) Vaughn, Barbara ChristineItem Epidemiology of epiphytic Pseudomonas syringae on barley(Montana State University - Bozeman, College of Agriculture, 1987) Georgakopoulos, Dimitrios G.Item Plant-associated fluorescent pseudomonads : their systematic analysis, microbial antagonism and iron interaction(Montana State University - Bozeman, College of Agriculture, 1982) Hemming, Bruce ClarkItem Microbial diversity and zinc toxicity to Pseudomonas sp. from Coeur d'Alene River sediment(Montana State University - Bozeman, College of Engineering, 2007) Barua, Sutapa; Chairperson, Graduate Committee: Brent M. PeytonCoeur d'Alene River (CDAR) in northern Idaho is one of the metal contaminated rivers in US. The sediments of the river are enriched with As, Cd, Cu, Pb, and Zn which are toxic metals to humans and animals. It is hypothesized that microorganisms living in this river sediment can remove the metals and thus detoxify their environment. The objective of this work is to investigate the microbial communities existing in CDAR sediment using 16S ribosomal RNA (rRNA) gene sequencing and 16S rRNA gene microarray (PhyloChip) analysis. According to our phylogenetic analysis, the CDAR clones fell into 13 distinct phylogenetic classes including 2 environmental samples, 1 uncultured bacterium, and an unclassified Chloroflexi. The major representative genera found were Thiobacillus (7 of 91), Azoarcus (7/91), Acidobacterium (6/91), Burkholderia (5/91), Flavobacterium (5/91) and Janthinobacterium (5/91). PhyloChip data showed the presence of 1551 operational taxonomic units (OTUs). 97% of the clone library sequences matched at various taxonomic levels with the microarray results.Item Molecular aspects of uranium toxicity : speciation and physiological targeting(Montana State University - Bozeman, College of Engineering, 2009) VanEngelen, Michael Robert; Chairperson, Graduate Committee: Brent M. Peyton; Robin Gerlach (co-chair)Uranium (U), as the uranyl ion (UO 2 ²+), is a widely distributed contaminant at several Department of Energy (DOE) sites, former war zones, and across the globe. Although many U remediation efforts depend on U-bacterial interactions, little information regarding U-bacterial interactions resolved at the molecular level exist. In this study, experiments were performed aimed at understanding the effect of molecular UO 2 ²+ speciation on bacterial bioaccumulation and toxicity using an environmental Pseudomonas sp. isolate. Results showed that the charge and stability of UO 2 ²+ species largely controlled the extent of UO 2 ²+ bioaccumulation and UO 2 ²+ toxicity, respectively. Further experimentation, including a combination of in vivo, in vitro, and in silico studies, revealed a specific mechanism of UO 2 ²+ toxicity, the first to be reported. This mechanism involves the binding of UO 2 ²+ to pyrroloquinoline quinone (PQQ), a cofactor present in a number of bacterial dehydrogenase enzymes. Based on the specific binding mode of UO 2 ²+ to PQQ, it is hypothesized that the present work has direct implications for UO 2 ²+ inhibition of flavoproteins, potentially extending the application of the findings of this work to eukaryotic systems. Recent trends suggest that U-related activity will increase in the near future, and therefore understanding fundamental interactions between UO 2 ²+ and living systems is both an environmental and human health imperative.