Exploring the diversity and ecology of bacteriophages in the dairy farm environment

Cook, Ryan (2023) Exploring the diversity and ecology of bacteriophages in the dairy farm environment. PhD thesis, University of Nottingham.

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Bacteriophages, viruses that obligately infect bacteria, represent the most abundant and diverse biological entities on the planet, with key and complex ecological roles in all environments where they have been studied. Agricultural wastes and manures (i.e., cattle slurry) are economically important fertilisers that are applied to land. Despite the widespread use of slurries, there is a paucity of knowledge regarding the microbial composition within them.

The first part of this thesis sought to optimise viral metagenomics, for the study of viral communities in nature. As the study of uncultivated viral genomes is underpinned by known complete viral genomes, I assessed the current extent of viral sequencing to provide the most complete reference database possible in an updated and reproducible fashion. This led to the INPHARED database, now published in PHAGE and available on GitHub; a community resource that provides genomes and annotation files to aid in viral genomic and metagenomic analyses. Furthermore, I investigated biases in the current collection of phage genomes and demonstrated that clear biases towards phages of a small subset of clinically relevant bacteria. Subsequently, I sought to benchmark sequencing technologies and assembly approaches for the recovery of viral genomes from viral metagenomes. This work, in part published in Microbiome and under review in Microbial Genomics, demonstrated that choice of sequencing technology and assembly algorithm will have significant impacts on downstream analyses and estimates of viral diversity. Overall, these analyses demonstrated that a combination of long and short read sequencing approaches performed best at recovering viral genomes in a mixed community.

The second part of this thesis applied the understandings described above to the dairy farm environment. I utilised long- and short-read sequencing to characterise the viral community of agricultural slurry in a longitudinal study, as well as modelled slurry tanks that contained agricultural antimicrobials, and the dairy cattle gut across life stages. Analysis of the cattle slurry virome, now published in Microbiome, revealed a diverse and novel community that was stable over time, despite constant influx and efflux of material. Notably, there was widespread phage carriage of a virulence determinant— VapE—that is associated with bovine mastitis-causing pathogens such as Streptococcus spp. Subsequent experiments with mock slurry tanks revealed the slurry virome may be influenced by the presence of footwash, although the reasons for this remain unclear. Analysis of the dairy cow virome uncovered 1,338 predicted complete phage genomes, the most of a single virome study to date. The phages within the dairy cow gut were largely novel, and their community composition changed over key life stages.

The results within this thesis have advanced the methodology of viral metagenomics approaches in general, and show that viruses likely play important ecological roles within agricultural environments, including augmenting the virulence of relevant veterinary pathogens.

Item Type: Thesis (University of Nottingham only) (PhD)
Supervisors: Jones, Michael
Millard, Andrew
Stekel, Dov
Hobman, Jon
Hudson, Chris
Keywords: Bacteriophages; Viral metagenomics; Phage genomes; Sequencing technologies; Viral genomes; Agricultural slurry; Dairy cows; Gut phages
Subjects: Q Science > QR Microbiology > QR355 Virology
Faculties/Schools: UK Campuses > Faculty of Medicine and Health Sciences > School of Veterinary Medicine and Science
Item ID: 73524
Depositing User: Cook, Ryan
Date Deposited: 19 Jul 2023 04:40
Last Modified: 01 Jun 2024 04:30
URI: https://eprints.nottingham.ac.uk/id/eprint/73524

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