Author's School

Graduate School of Arts & Sciences

Author's Department/Program

Biology and Biomedical Sciences: Computational and Systems Biology

Language

English (en)

Date of Award

Spring 4-26-2013

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Chair and Committee

Jeffrey I Gordon

Abstract

The human gut harbors tens of trillions of microbes belonging to all three domains of life, Bacteria, Archaea, and Eukarya; most are members of Bacteria. These organisms collaborate and compete for functional niches and physical space: habitats), together forming a continuously functioning metabolic organ that influences many aspects of host biology. The factors that drive the assembly, determine the stability, and shape the adaptive responses of the gut microbiota to a variety of perturbations are the subject of intense study as greater appreciation is gained of the importance of this microbial community for human health. My thesis focused on the viral component of the microbiota that had been less characterized than its bacterial component. I first developed and applied a series of experimental and computational tools for metagenomic analyses of viruses purified from frozen fecal samples obtained from healthy adult monozygotic twin pairs and their mothers living in the USA, over the course of a year. The virome in this population was dominated by phages and exhibited high inter-personal variation and contrasting intrapersonal stability, suggesting a prevalent temperate lifestyle rather than a predator-prey relationship that is a feature of marine microbial communities. To further characterize the role of phage in shaping gut community structure, I colonized adult germ-free mice with a defined model human gut microbiota composed of 15 sequenced human gut symbionts, seven of which harbored 10 prophages, one of which: Bacteroides cellulosilyticusWH2) was represented by a library of >25,000 isogenic transposon mutants covering 80% of genes in its genome. Once assembled, this model microbiota was subjected to a staged phage attack with a pool of virus-like particles: VLPs) purified from the fecal microbiota of five humans from the first study. Shotgun sequencing of DNA isolated from the input human VLP preparation, cecal and fecal samples collected over time from these gnotobiotic mice, and VLPs recovered from their fecal samples, revealed a ordered and reproducible sequence of phage attack, allowing me to associate novel phages present in the input VLP preparation with bacterial hosts, and to characterize the dynamics and identify genetic determinants of prophage induction. Finally, I used the tools I developed to characterize the phages and eukaryotic viruses present in the fecal microbiota of healthy and malnourished twins living in Malawi, sampled during their first two years of life. Together, this work provided new perspectives about viral diversity and viral-bacterial host dynamics associated with the human gut microbiota.

DOI

https://doi.org/10.7936/K7K935J0

Comments

Permanent URL: http://dx.doi.org/10.7936/K7K935J0

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