Hydrogel interfaces for applications in microbial biotechnology

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Abstract

Hydrogels are three-dimensional, water-swollen, highly crosslinked polymers that can be designed to provide biocompatible and biofunctional interfaces for cells and biomolecules. With facile fabrication and precise control over chemistry, pore size, and mechanical properties, hydrogels have been studied extensively in various areas of biomedical and bioengineering, particularly in drug delivery and tissue engineering applications. However, hydrogels have not been well-studied or well-applied to many emerging applications in microbiology. This thesis explores two new applications involving hydrogel interfaces: (1) photodegradable hydrogels for high-throughput screening and isolation of rare bacteria and (2) hydrogels for protection of electroactive biofilms from environmental shocks in microbial electrolysis cell systems. The initial portion of this thesis focuses on the use of photodegradable hydrogels for microbial cell screening and rare cell isolation. The photodegradable hydrogel used here was formed with polyethylene glycol (PEG) o-nitrobenzyl acrylate and PEG-tetrathiol macromers, which form three-dimensional hydrogels through thiol-acrylate addition reactions to encapsulate heterogenous populations of bacterial cells. The individual entrapped cells can be cultured into clonal microcolonies due to the suitable hydrogel mesh size for nutrient transport to the cells. Cells are monitored en masse and rare cells showing unique growth phenotypes are identified and extracted from the hydrogel interface using a high-resolution light patterning tool. The optimum experimental setup for achieving high throughput observation and clean extraction was developed. Release kinetics with light dose, the effect of light pattern on cell morphology, and the DNA quality of the extracted cells after exposure to 365 nm light patterns was also investigated. We demonstrated the use of this approach as a screening interface by rapidly screening a mutant library of the Gram-negative bacteria Agrobacterium tumefaciens to identify, isolate, and genetically characterize strains with rare growth profiles. The reported method offers an inexpensive and practical approach to cell screening and cell sorting and can be applied to a wide range of applications where isolating phenotypically pure cells from complex, heterogenous mixtures is essential. This includes applications in microbiology, microbial therapeutics, and biomedical diagnostics. The next section of this thesis focuses on developing PEG-based hydrogels that are designed to protect electroactive biofilms from harsh environmental stressors. The coating was fabricated using PEG-tetrathiol and PEG-divinyl sulfone macromers that form hydrogels with crosslinks resistant to degradation from acid or base hydrolysis, while still promoting nutrient diffusion and electron transport. Methods of fabricating anodes containing electroactive biofilms with the hydrogels are first reported, followed by investigation of the hydrolytic stability of the coatings. Transport of a carbon source (acetate) through the coating is then modeled, and the long-term stability and compatibility of the coating over the biofilm is investigated. Lastly, the effect of the coating on the biofilm recovery from an environmental shock (ammonium exposure) is demonstrated to emphasize the potential benefit of the coating. Finally, the future directions of hydrogels in these applications are recommended, which include discussion on developing a hydrogel chemistry that is degradable on exposure to a near-infrared (NIR) light source as well as discussion on chemical and biological hydrogel additives that will improve its performance.

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Keywords

Hydrogels, Cell isolation, Photodegradable hydrogels

Graduation Month

August

Degree

Doctor of Philosophy

Department

Department of Chemical Engineering

Major Professor

Ryan R. Hansen

Date

2021

Type

Dissertation

Citation