Dissimilatory iron reduction (DIR) is significant to the biogeochemical cycling of iron, carbon and other elements, and may be applied to bioremediation of organic pollutants, toxic metals, and radionuclides; however, the mechanism(s) of DIR and factors controlling its kinetics are still unclear. To provide insights into these questions, the interaction between a common dissimilatory iron reducing bacterium (DIRB)was studied, Shewanella oneidensis MR-1, and ferric iron (Fe(III)) (oxy)(hydr)oxide mineral surfaces. Firstly, atomic force microscopy was used to study how S. oneidensis MR-1 dissolved Fe(III) (oxy)(hydr)oxides and compared it to two other cases where Fe(III) (oxy)(hydr)oxides were either dissolved by a chemical reductant or by a mutant with an electron shuttling compound. Without the electron shuttling compound, the mutant is unable to respire on Fe(III) (oxy)(hydr)oxides, but with the electron shuttling compound, it can. It was found that the cells of S. oneidensis MR-1 formed microcolonies on mineral surfaces and dissolved the minerals in a non-uniform way which was consistent with the shape of microcolonies, whereas Fe(III) (oxy)(hydr)oxides were uniformly dissolved in both of the other cases. Secondly, confocal microscopy was used to study the adhesion behavior of S. oneidensis MR-1 cells on Fe(III) (oxy)(hydr)oxide surfaces across a broad range of bulk cell densities. While the cells were evenly distributed under low bulk cell densities, microcolonies were observed at high bulk cell densities. This adhesion behavior was modeled by a new, two-step adhesion isotherm which fit better than a simple Langmuir or Freundlich isotherm. The results of these studies suggest that DIR is in-part transport limited and the surface cell density may control DIR.

Previous research into microbial colonies and biofilms shows a significant gap in our current understanding of how bacterial structures develop. Despite the huge body of research undertaken into the formation, genetic makeup, composition, and optimal growth conditions of colonies, no study has been successful in identifying all individual bacteria in a colony in three-dimensions as a function of time. This lack of bacterial cell lineage in such a simple class of organisms is conspicuous in the light of what is known about other organisms, such as Caenorhabditis elegans [1]. In this thesis I show that using laser scanning confocal microscopy in conjunction with developments in sample preparation and post acquisition image analysis, it is possible to fully reconstruct all individual bacteria within an Escherichia coli (E. coli ) microcolony grown in viscoelastic media. Additionally, I show that by further pushing the resolution of confocal microscopes, commercial systems are capable of extracting three-dimensional information on protein structures inside bacteria at early stages of growth. This thesis is in three parts. The first part shows that by pushing the resolution of a commercial laser scanning confocal microscope system it is possible to achieve single cell resolution of a bacterial colony growing in three dimensions in a viscoelastic medium (agarose) from a seed bacterium. The growth of individual bacteria is examined as the concentration of agarose in the media is altered. Results show there is a nonlinear dependence between the rate of growth of a bacterium and the concentration of the agarose in the media with a peak in growth rate at 3% (weight) concentrations of agarose in M9 media. The second part of this work presents a study of how an initially two-dimensional colony growing between a glass slide and agarose gel suddenly invades the third spatial dimension by buckling. The results show that the cells within the centre of the colony flex and buckle, due to confinement by their neighbours, creating additional layers. Indeed, flexing is not limited to the buckling event but occurs throughout the early growth cycle of a colony. The final part of this thesis shows that by further pushing the resolution of confocal microscopes, commercial systems are capable of extracting three-dimensional information about the temporal evolution of the spatial distribution of the FtsZ septation ring within the cell. As the bacterial colony grows from a seed bacterium to a microcolony, the error in placing the division accurately at the cell centre is seen to increase as the number of bacteria within the colony increases and spatial confinement occurs.

This is a two-part project. Proton pump inhibitors (PPIs) have been associated with increased risk of C. difficile infections and increased toxin production when combined with antimicrobial therapy. The first part of this project involved characterization of a hypervirulent NAP1 C. difficile strain, including genome sequencing and assembly, and the development of methods to study its transcriptomics using RNA-Seq, which will enable future researchers to study different expression patterns when toxigenic C. difficile is challenged with PPIs and/or antimicrobials in vitro. The second part of this project involved characterizing a clinical isolate of a NAP1 C. difficile displaying a markedly elevated MIC to metronidazole (MIC = 16 mg/mL), which initially exhibited MIC of 32 mg/mL. A method of obtaining a metronidazole-susceptible revertant from this isolate was developed and a revertant was obtained. The genomes of both isolates were sequenced, assembled, and aligned, then compared to each other for polymorphisms.

This is a two-part project. Proton pump inhibitors (PPIs) have been associated with increased risk of C. difficile infections and increased toxin production when combined with antimicrobial therapy. The first part of this project involved characterization of a hypervirulent NAP1 C. difficile strain, including genome sequencing and assembly, and the development of methods to study its transcriptomics using RNA-Seq, which will enable future researchers to study different expression patterns when toxigenic C. difficile is challenged with PPIs and/or antimicrobials in vitro. The second part of this project involved characterizing a clinical isolate of a NAP1 C. difficile displaying a markedly elevated MIC to metronidazole (MIC = 16 mg/mL), which initially exhibited MIC of 32 mg/mL. A method of obtaining a metronidazole-susceptible revertant from this isolate was developed and a revertant was obtained. The genomes of both isolates were sequenced, assembled, and aligned, then compared to each other for polymorphisms.