title: The theoretical and experimental exploration of the use of predatory bacteria to control biofilms
Creators: name: Ogundero, Ayo
Publisher: name: University of Glasgow
Year: 2023

description: Membrane based technologies are widely used for treating drinking water in sparsely populated areas, but their effectiveness is significantly diminished by the growth of biofilms and biofouling. Preventing or removing biofilms can increase the life span of membranes and, thus, improve their economic viability. Most cleaning methods involve physical disruption or antimicrobial chemicals and, therefore, require an interruption in the membrane operation. Bdellovibrio, a group of predatory bacteria, are a potential alternative to antimicrobials or physical disruption because of its ability to kill a large range of gram-negative bacterial prey and the inability of their prey to develop genetic resistance. However, the use of Bdellovibrio in industrial application has not been widespread in part due to the lack of understanding of the dynamics between Bdellovibrio and their prey. To compound this, many of the previous investigations into Bdellovibrio and biofilm ecology are limited by inaccurate, uninformative, and labour-intensive methods to quantify the population dynamics, which makes it difficult to build comprehensive models to exploit Bdellovibrio as a control to biofilms in systems like drinking water membranes. This thesis aims to develop a set of novel methods and technologies to accurately investigate Bdellovibrio and the effect they have on dynamics of their prey; Pseudomonas sp, a key gram negative biofilm forming species. This research develops the first protocol to use flow cytometry to accurately and rapidly quantify Bdellovibrio and Pseudomonas sp growth, which makes recording high resolution population dynamics feasible. The protocol was used for the development and experimental validation of mathematical models which aimed to predict Bdellovibrio dynamics in batch and chemostat systems. We show the first experimental observation of Bdellovibrio-prey oscillations, a key component of predation dynamics and a desired phenomenon for the use of Bdellovibrio as a self-sustaining biocontrol. To extend the models for application to systems where biofilms prevail, we demonstrated a new method of deploying flow cytometry and fluorescent assays to quantify and characterise the effect of nutrients on biofilm growth and predation. The findings suggest that extracellular polymeric substances (EPS) play a vital role in the attachment and persistence of biofilm when under Bdellovibrio predation. Thus, in biofilm research, the simple density dependent predator-prey interactions need to be augmented by representing the spatial heterogeneities in biofilm processes and properties such as its detachment, EPS and presence of metabolically damaged cells. For a more nuanced analysis of predator-prey interactions, at the resolution of individual organisms, this research develops a novel microfluidic device to observe Bdellovibrio predation on a 1-D biofilm. This thesis describes both the rationale and novel protocols for combining electron-beam lithography with, the more commonly used, photolithography to create an array of high-resolution channels to constrain biofilms and challenge them with predators. The research demonstrates the opportunities and the technical challenges in using microfluidics. Ultimately, if we are to develop mathematical models that can be parameterised and used effectively in designing strategies for controlling biofilms using predatory bacteria, then observations at the individual scale in microfluidic devices will be invaluable.