Development of improved cultivation methods for environmental microorganisms
File(s)
Author(s)
Morrison, Andrew
Type
Thesis
Abstract
Cultivation of bacteria remains an essential prerequisite for numerous research and biotechnological applications. Despite their ubiquity, only a minority of environmental bacteria are cultivable using standard techniques. Therefore, a vast microbial ‘dark matter’ awaits exploration for valuable therapeutics or research potential. Despite innovations in enhanced cultivation techniques, limitations remain including throughput, dependencies on environmental factors or abrupt transitions of microorganisms from native to laboratory conditions. This thesis addresses these challenges with the development, optimisation, and experimental assessment of two novel cultivation methodologies. The first methodology allows for the gradual transition and acclimatisation of microorganisms from native environments to culture media using the novel Enhanced Domestication (EDEN) device. Compared to the instantaneous transition, acclimatisation of pond water bacteria to R2A culture media significantly enhanced cultivation diversity, colony yield and greater taxonomic range of isolates including those previously reported to be recalcitrant to cultivation. Moreover, likely novel taxa cultivated with EDEN exhibited antimicrobial activity against methicillin-resistant Staphylococcus aureus. The second methodology involves the development of a cell encapsulation apparatus, termed Bacterial Encapsulation and Containment (BEAD), enabling microinjection and cultivation of bacteria within alginate hydrogel beads using an automated pneumatic pump system. The performance of in situ cultivation with BEAD-encapsulated bacteria were assessed using the newly developed cultivation cassettes growth chambers. The findings propose enhanced cultivation strategies to unlock the biosynthetic potential of uncultured environmental bacteria.
Version
Open Access
Date Issued
2024-01-02
Date Awarded
2024-05-01
Copyright Statement
Creative Commons Attribution NonCommercial Licence
License URL
Advisor
Low, Harry
Buck, Martin
Publisher Department
Infectious Disease
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)