Insights into the plastic degrading potential of microorganisms from hydrophobic habitats

Abstract

Plastics and microplastics (< 5 mm) are ubiquitous across Earth’s biosphere. These high-molecular-weight polymers are persistent in the environment, given their very low bioavailability, attributed to their high hydrophobicity and insolubility. Despite these parameters, some microbes can utilise similar enzymes that break down plant biomass, to degrade synthetic plastic. Culture-based approaches have utilised plastic-contaminated sites to isolate plastic-degrading microbes, which may be enriched by the long-term presence of these polymers. This thesis identifies ‘hydrophobic habitats’; microbial habitats within environments rich in many hydrophobic substrates which may induce stress and/or be an energy-dense carbon sink for microbes. Such substrates include insoluble polymers such as chitin, cellulose, and lignin, which may be derived from decayed plant and animal material. This thesis aims to use these habitats as a source to screen for plastic-degrading microbes, and to characterise them in terms of their potential in bioremediation. One such habitat is a peat bog, from which a polypropylene-degrading microbe, Gordonia terrae iso11, was isolated. Key issues pertaining to biotic plastic degradation, and the current literature, is presented in a literature review. It highlights that microbial attachment to the surface of the polymer plays a key role in the mechanism of degradation, in contrast to a saprotrophic mechanism, where a microbe secretes enzymes which react with the polymer in the extracellular environment.

This thesis identifies factors with are empirically shown to either enhance or retard rates of plastic degradation in a given microbe or enzyme system. These multifaceted factors include polymer oxidation and enzyme engineering (to enhance plastic breakdown) and issues such as oxidative stress (which may retard polymer breakdown). These, along with a first-order kinetic model, are used to derive a Coefficient of Degradability for microbial substrates, which is a standardised measure which can be used to predict the rates of breakdown and environmental persistence of given carbon substrate.

A culture-based approach to isolate plastic-degrading microbes was carried out in a selection of identified hydrophobic habitats. This yielded 39 candidate plastic-degrading microbes, three of which displayed rates of >10% mass loss of polymer over a 28-day incubation period. These isolates had a high cell surface hydrophobicity, which is proposed to aid in approaching and attaching to the polymer (and forming a biofilm). The most-rapid plastic-degrading microbe, G. terrae iso11, was characterised in terms of biofilm formation under varying nutrient conditions, and biosurfactant production. Interestingly, biosurfactant production remained constant under all tested nutrient conditions (with and without the presence of hydrocarbons). It is proposed that, whilst there is no evidence of differential regulation of biosurfactants, these play a critical role in solubilising hydrocarbons, such as plastic.

Whole genome sequencing of the obtained G. terrae iso11 isolate allowed for the identification of a gene coding for alkane-1-monooxygenase, which was overexpressed in Escherichia coli BL21 (DE3). The enzyme was assayed for activity on polypropylene films. Measured increases in the surface free energy of the polymer, and the aerobic incubation environment, provided evidence of polymer oxidation by the enzyme. Currently, there are no known polypropylene-active enzymes. Therefore, this finding has implications for the usage of alkane-1-monooxygenases for the degradation of polyolefins, such as polypropylene.

Thesis is embargoed until 31 July 2026.

Date of Award	Jul 2024
Original language	English
Awarding Institution	Queen's University Belfast
Sponsors	Department of Agriculture, Environment and Rural Affairs
Supervisor	Stephen Kelly (Supervisor) & Julianne Megaw (Supervisor)