http://hdl.handle.net/10379/703 Sun, 29 Oct 2017 23:35:38 GMT 2017-10-29T23:35:38Z http://hdl.handle.net/10379/6827 Bioreactor scalability: laboratory-scale bioreactor design influences performance, ecology, and community physiology in expanded granular sludge bed bioreactors Connelly, Stephanie; Shin, Seung Gu; Dillon, Robert J.; Ijaz, Umer Zeeshan; Quince, Christopher; Sloan, William; Collins, Gavin Studies investigating the feasibility of new, or improved, biotechnologies, such as wastewater treatment digesters, inevitably start with laboratory-scale trials. However, it is rarely determined whether laboratory-scale results reflect full-scale performance or microbial ecology. The Expanded Granular Sludge Bed (EGSB) bioreactor, which is a high-rate anaerobic digester configuration, was used as a model to address that knowledge gap in this study. Two laboratory-scale idealizations of the EGSB-a one-dimensional and a three-dimensional scale-down of a full-scale design-were built and operated in triplicate under near-identical conditions to a full-scale EGSB. The laboratory-scale bioreactors were seeded using biomass obtained from the full-scale bioreactor, and, spent water from the distillation of whisky from maize was applied as substrate at both scales. Over 70 days, bioreactor performance, microbial ecology, and microbial community physiology were monitored at various depths in the sludge-beds using 16S rRNA gene sequencing (V4 region), specific methanogenic activity (SMA) assays, and a range of physical and chemicalmonitoringmethods. SMA assays indicated dominance of the hydrogenotrophic pathway at full-scale whilst a more balanced activity profile developed during the laboratory-scale trials. At each scale, Methanobacterium was the dominant methanogenic genus present. Bioreactor performance overall was better at laboratory-scale than full-scale. We observed that bioreactor design at laboratory-scale significantly influenced spatial distribution of microbial community physiology and taxonomy in the bioreactor sludge-bed, with 1-D bioreactor types promoting stratification of each. In the 1-D laboratory bioreactors, increased abundance of Firmicutes was associated with both granule position in the sludge bed and increased activity against acetate and ethanol as substrates. We further observed that stratification in the sludge-bed in 1-D laboratory-scale bioreactors was associated with increased richness in the underlying microbial community at species (OTU) level and improved overall performance. Sun, 01 Jan 2017 00:00:00 GMT http://hdl.handle.net/10379/6827 2017-01-01T00:00:00Z http://hdl.handle.net/10379/6820 Determination of survival of wildtype and mutant Escherichia coli in soil Somorin, Yinka; O'Byrne, Conor E. coli resides in the gastrointestinal tract of humans and other warm-blooded animals but recent studies have shown that E. coli can persist and grow in various external environments including soil. The general stress response regulator, RpoS, helps E. coli overcome various stresses, however its role in soil survival was unknown. This soil survival assay protocol was developed and used to determine the role of the general stress response regulator, RpoS, in the survival of E. coli in soil. Using this soil survival assay, we demonstrated that RpoS was important for the survival of E. coli in soil. This protocol describes the development of the soil survival assay especially the recovery of E. coli inoculated into soil and can be adapted to allow further investigations into the survival of other bacteria in soil. Thu, 20 Jul 2017 00:00:00 GMT http://hdl.handle.net/10379/6820 2017-07-20T00:00:00Z http://hdl.handle.net/10379/6665 Manipulation of intestinal epithelial cell function by the cell contact-dependent type III secretion systems of Vibrio parahaemolyticus O'Boyle, Nicky; Boyd, Aoife Vibrio parahaemolyticus elicits gastroenteritis by deploying Type III Secretion Systems (TTSS) to deliver effector proteins into epithelial cells of the human intestinal tract. The bacteria must adhere to the human cells to allow colonization and operation of the TTSS translocation apparatus bridging the bacterium and the host cell. This article first reviews recent advances in identifying the molecules responsible for intercellular adherence. V parahaemolyticus possesses two TTSS, each of which delivers an exclusive set of effectors and mediates unique effects on the host cell. TTSS effectors primarily target and alter the activation status of host cell signaling proteins, thereby bringing about changes in the regulation of cellular behavior. TTSS1 is responsible for the cytotoxicity of V parahaemolyticus, while TTSS2 is necessary for the enterotoxicity of the pathogen. Recent publications have elucidated the function of several TTSS effectors and their importance in the virulence of the bacterium. This review will explore the ability of the TTSS to manipulate activities of human intestinal cells and how this modification of cell function favors bacterial colonization and persistence of V parahaemolyticus in the host. Fri, 10 Jan 2014 00:00:00 GMT http://hdl.handle.net/10379/6665 2014-01-10T00:00:00Z http://hdl.handle.net/10379/6246 Engineering recombinant antibodies for polymer biofunctionalization Hortigüela, María J.; Lucie Aumailley, Lucie; Srivastava, Akshay; Cunningham, Clare; Anandakumar, Soshee; Robin, Sylvain; Pandit, Abhay; Hu, Xuejun; Wall, Gerard The attachment of recognition elements such as antibody fragments to polymeric substrates can be used to mediate cell- or protein-specific interactions. In this work, single-chain Fv (scFv) antibody fragments were isolated against two cell types of interest and expressed in an Escherichia coli expression platform. The scFvs were engineered at their C-terminus to incorporate a cysteine-containing linker, for reaction with maleimide-linked polymers, or a heptasaccharide glycan for complexation with surface amine moieties. Antigen binding of the modified scFvs was unchanged, and expression yields of the glyco-engineered scFvs were similar to the unmodified molecules, while cys-tagged scFv yields varied between scFv variants. Targeted immobilization of the scFvs via either modification resulted in three-to five-fold higher binding of ligands over adsorbed molecules. The study demonstrates a simple and efficient antibody engineering and modification approach for effective targeted immobilization on polymeric substrates. Copyright (C) 2015 John Wiley & Sons, Ltd. Tue, 18 Aug 2015 00:00:00 GMT http://hdl.handle.net/10379/6246 2015-08-18T00:00:00Z