Browsing by Subject "Antibiotic resistance"
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Item The MAR1 transporter of Arabidopsis thaliana has roles in aminoglycoside antibiotic transport and iron homeostasis(2009-08) Conte, Sarah Schorr; Lloyd, Alan M.Widespread antibiotic resistance is a major public health concern, and plants represent an emerging antibiotic exposure route. Recent studies indicate that crop plants fertilized with antibiotic-laden animal manure accumulate antibiotics, however, the molecular mechanisms of antibiotic entry and subcellular partitioning within plant cells remain unknown. Here we report that mutations in the Arabidopsis locus Multiple Antibiotic Resistance (MAR1) confer resistance, while MAR1 overexpression causes hypersensitivity to multiple aminoglycoside antibiotics. Resistance is highly specific for aminoglycosides and does not extend to antibiotics of other classes, including the aminocyclitol, spectinomycin. Yeast expressing MAR1 are hypersensitive to the aminoglycoside, G418, but not to chloramphenicol or cycloheximide. MAR1 encodes a protein with 11 putative transmembrane domains with low similarity to ferroportin1 from Danio rerio. A MAR1:YFP fusion protein localizes to the chloroplast, and chloroplasts from plants overexpressing MAR1 accumulate more of the aminoglycoside, gentamicin, while mar1-1 mutant chloroplasts accumulate less than wild type. MAR1 overexpression lines are slightly chlorotic, and this chlorosis is rescued by application of exogenous iron. MAR1 expression is also downregulated by low iron. Taken together, these data suggest that MAR1 is a plastid transporter that is likely to be involved in cellular iron homeostasis, and allows opportunistic entry of multiple antibiotics into the chloroplast. mar1 mutants represent an interesting example of plant antibiotic resistance that is based on the restriction of antibiotic entry into a subcellular compartment. Knowledge about this process – and other processes of antibiotic entry – could enable the production of crop plants that are incapable of antibiotic accumulation, aid in development of phytoremediation strategies for decontamination of water and soils polluted with antibiotics, and further the development of new plant-based molecular markers. The work described here also contributes to our understanding of how plants interact with the antibiotics they encounter, both in the laboratory (where aminoglycosides such as kanamycin are used heavily to select for transgenics) and in the natural environment.Item Metagenomic investigation of the antibiotic resistance in coastal marine ecosystemsTallman, James Joseph IIIItem Microfabricated environments for the study of bacterial group behavior(2015-12) Fitzpatrick, Mignon Denise; Shear, Jason B.; Whiteley, MarvinThis thesis describes the application of micro three dimensional printing (µ3DP) techniques to create protein microstructures for the study of bacterial group behavior in small populations. Studies involving aggregates of ~10^1 to ~10^5 cells have shown extensive and complex communication and spatial organization. Multiphoton lithography (MPL) provides a means to quickly design and execute the fabrication of microscale structures with submicron resolution from a variety of biocompatible polymers. Using this technique, intricate spatial arrangements of bacteria can be achieved while maintaining small population sizes at high cell density (≥10^8 cells/mL), providing in vitro culture conditions which better simulate in vivo settings. As a result, valuable information can be obtained about bacterial social interactions through the coupling of additional analytical techniques to detect the presence or absence of extracellular signaling molecules. While quorum sensing (QS) remains the most extensively studied means of bacterial communication, it is becoming increasingly apparent that additional factors are necessary to effect certain changes in population-wide genetic expression which can lead to increased virulence, pathogenicity, and the development of antibiotic resistance. The work presented in this thesis addresses the influences of cell density, chemical heterogeneity of the environment within cell aggregates, and level of cell surface attachment as mechanisms to induce or influence the development of antibiotic resistance. Building upon previous work presented by members of the Shear lab, BSA-gelatin protein microstructures were used to study the behavior and response of the opportunistic pathogen Pseudomonas aeruginosa under these conditions. Antibiotic resistance was observed in low cell number/high density populations in agreement with previous work presented by the Shear lab. In addition, it was found that localized regions of oxygen depletion do not correlate directly with antibiotic resistance development, as the population size required for depletion far exceeded that for development of resistance. Finally, a new technique directed at simultaneous biofilm inhibition and cell removal from solution was explored.