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Antimicrobial resistance (AMR) is set to be the next global pandemic with AMR related deaths set to outgrow cancer deaths by the year 2050. Increasing resistance to the traditional arsenal of antibiotics is expanding at an alarming rate and novel antimicrobial therapeutics are urgently required to tackle this emerging issue. Proteus mirabilis and Pseudomonas aeruginosa are two bacterial species commonly associated with urinary tract infections in humans. To contribute to the progression of novel antimicrobials against P. mirabilis and P. aeruginosa, this thesis was based on the isolation and utilisation of bacteriophages and bacteriophage derived products to help treat biofilms caused by both bacteria. The isolation of phage PM-CJR showed promising antimicrobial potential against P. mirabilis due to the production of depolymerase enzymes. The thesis describes the identification and utilisation of both the whole phage and its tail-spike protein with the pectate lyase activity to treat P. mirabilis biofilms. The phage and the tail-spike protein were assessed in a range of different in vivo and in vitro assays to demonstrate their antimicrobial and antivirulence properties against P. mirabilis biofilms. This thesis also studies the combinational treatment of phage and cold atmospheric plasma for the treatment of P. aeruginosa biofilms and establishes the most effective order or application of each antimicrobial for the most effective treatment option against P. aeruginosa biofilms. We successfully showed that the use of CAP followed by exposure to P. aeruginosa phages is the most effective order of application for eradication of P. aeruginosa biofilms.

The enormity of bacterial diversity is such that taxonomy is necessary for classification. Taxonomy has historically used an array of different classification techniques, from GC content, to morphology, biochemical capability, and more recently,16S rDNA and whole genome analyses. Currently, there is no standard approach to placing a bacterium taxonomically, leading to a large degree of subjectivity. Average nucleotide identity (ANI) analyses of 71 culture-based genomes and 64 metagenome-assembled genomes revealed vast genetic diversity, with the genomes clustering into 59 genera and 75 species. Where these species formed groups of four or more genomes, pangenomic analyses were carried out on both these groups, and the original taxa they had been assigned to (Butyrivibrio fibrisolvens, Butyrivibrio proteoclasticus, Butyrivibrio hungatei, Pseudobutyrivibrio ruminis, Pseudobutyrivibrio xylanivorans, and an additional unnamed Butyrivibrio sp group). Core genes only formed 0.66-8.50% of the genomes of the original taxa, revealing a wealth of accessory genes. In contrast, the core genes for the groups formed by ANIm analysis composed 38.48-76.67% of the genomes. Many of the accessory genes were identified as carbohydrate active enzymes, belonging to glycosyl hydrolase families 2, 3, 5, 13, and 43. Cloning and expression of a selection of these genes was attempted, but success was limited due to time constraints. The culture-based genomes were bioprospected for linoleate isomerases, and putative enzymes identified. Two of these were selected for expression and purification. The resulting proteins were incubated with linoleic acid and the incubations analysed via gas chromatography. Whilst peaks could be seen at appropriate retention times, it is inconclusive as to whether this is as a result of these proteins catalysing the breakdown of linoleic acid. This thesis provides the largest pangenomic study on Butyrivibrio and Pseudobutyrivibrio to date. As such, it provides novel insight into the functional and genomic capacity of these dominant rumen bacteria.

This work aimed to examine whether halophilic archaea may act as a reservoir for antimicrobial resistance genes in extreme environments and their potential to be a source of new antimicrobial agents. The extremely halophilic archaea Halorubrum saccharovorum CSM-52, isolated from Kilroot salt mine in Northern Ireland, was investigated for the presence of resistance mechanisms, specifically active efflux pumps and the potential of the involvement of these pumps in their innate resistance to antimicrobials. Separately, H. saccharovorum was serially exposed to sublethal concentrations of ciprofloxacin in order to examine its adaptive response and ability to develop antibiotic resistance under antimicrobial stress. The metagenome of stalactite structures formed within the Triassic halite of Kilroot salt mine were subjected to metagenomic mining using a novel bioinformatic pipeline, AMPLY, for the discovery antimicrobial peptide sequences within this unique environment. The top hit sequences were synthesized and examined for their antimicrobial activity against clinically relevant bacteria, Candida albicans and representative haloarchaea.