Fischell Department of Bioengineering Theses and Dissertations
Permanent URI for this collectionhttp://hdl.handle.net/1903/6628
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Item ENABLING RAPID PHENOTYPIC DETECTION OF CEPHALOSPORIN RESISTANCE BEYOND THE CENTRAL LABORATORY(2019) Nguyen, Hieu Thuong; White, Ian; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)The so-called bacterial “superbugs” are largely resistant to some of the most commonly prescribed antibiotics, including a drug class known as cephalosporins used to treat many hospital and community-acquired infections. This major public health threat has been acknowledged for decades by the Centers for Disease Control (CDC) as a major concern; yet, the detection of superbugs has not been made routine since standard testing practices have been limited to specialized “central” laboratories with sophisticated yet bulky and expensive equipment and highly trained personnel. As a result, the lack of simpler testing methods that can be used in everyday clinics and doctor’s offices can be viewed as a source of error contributing to incorrect antibiotic treatment and poorer patient outcomes, factors that drive even more advanced resistance, depleting our drugs or last resort. In this dissertation, we explore new strategies for simplified methods to test for cephalosporin resistance in order to give higher accessibility in the timely detection of superbugs to support the improvement of patient care. To do this, we take an organic chemistry and biochemical approach to develop new detection molecules that report resistance activity in bacteria expressing extended-spectrum β-lactamase (ESBL) enzymes, one of the most prolific resistance strategies used by superbugs. Next, we describe methods of integrating these detection molecules into practical testing methods, and detail the engineering of simpler assays that allow for rapid readout of ESBL phenotypes using commonplace laboratory plate readers, portable Raman devices, and even handheld personal glucose meters (used for diabetes monitoring) purchased from the drugstore.Item Programmable Biomolecule Assembly and Activity in Prepackaged BioMEMS(2008-10-21) Luo, Xiaolong; Rubloff, Gary W.; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Antibiotic resistance is an increasing public health concern and few new drugs for bacterial pathogenesis have been obtained without addressing this resistance. Quorum sensing (QS) is a newly-discovered system mediated by extracellular chemical signals known as "autoinducers", which can coordinate population-scale changes in gene regulation when the number of cells reaches a "quorum" level. The capability to intercept and rewire the biosynthesis pathway of autoinduer-2 (AI-2), a universal chemical signaling molecule, opens the door to discover novel antimicrobial drugs that are able to bypass the antibiotic resistance. In this research, chitosan-mediated in situ biomolecule assembly has been demonstrated as a facile approach to direct the assembly of biological components into a prefabricated, systematically controlled bio-microelectromechanical system (bioMEMS). Our bioMEMS device enables post-fabricated, signal-guided assembly of labile biomolecules such as proteins and DNA onto localized inorganic surfaces inside microfluidic channels with spatial and temporal programmability. Particularly, the programmable assembly and enzymatic activity of the metabolic pathway enzyme Pfs, one of the two AI-2 synthases, have been demonstrated as an important step to reconstruct and interrogate the AI-2 synthesis pathway in the bioMEMS environment. Additionally, the bioMEMS has been optimized for studies of metabolic pathway enzymes by implementing a novel packaging technique and an experimental strategy to improve the signal-to-background ratio of the site-specific enzymatic reactions in the bioMEMS device. I envision that the demonstrated technologies represent a key step in progress toward a bioMEMS technology suitable to support metabolic engineering research and development.