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Abstract:
The microbial communiCes that inhabit the mammalian gut form a crucial component that affects
health and disease. Established knowledge points to predictable changes in the composiCon of the
microbiome at the species level as a result of diseases. However, there is a gap in knowledge of how
strain-level changes in commensal bacteria occur during and as a result of disease-related physiological
changes. In the first chapter of this thesis, we describe the evoluCon of an Escherichia coli strain in the
mouse gut during chronic intesCnal inflammaCon. As the healthy and inflamed intesCnal condiCons
would be different, I hypothesized that bacteria would also adapt differently to the two condiCons. I
isolated and cultured evolved and ancestral populaCons of E. coli from gnotobioCc mice, and then
performed metabolic tesCng to idenCfy phenotypes that were different between the populaCons that
evolved in the healthy and inflamed mice. In parallel, genome sequencing was used to idenCfy
inflammaCon-specific genotypes. Based on the idenCfied phenotypes and genotypes, the principles of
evoluConary medicine were used to propose novel therapeuCc strategies against chronic inflammatory
bowel disease. Such experiments could help us elucidate how changes in physiology and strain-level
changes in bacterial communiCes are inter-connected, which in turn could potenCally guide the
development of novel therapeuCc strategies.
Another area of focus in microbiome research is the role of bacteria-bacteria interacCons in shaping
the microbiome and thus affecCng health and disease. The evoluConary history shared between the
microbes and host suggests a high level of inter-dependence. In the second and third chapters of the
thesis, we explore the interacCons between naturally co-exisCng Bacteroidetes in the wild mouse gut.
First, we newly isolated strains from the cecal content of wild-derived mice, and I phenotyped bacterial
metabolism to characterize the metabolism of a novel Bacteroides species (chapter II). Second, we
used in vitro assays to determine the nature of interacCons between these strains (chapter III). I
performed protein family clustering and comparison between strains, as well as comparison of SNPs
between strains to idenCfy geneCc differences that could explain the interacCons observed. We found
that inter-species interacCons are host-dependent, i.e., strains of the same two species show different
types of interacCon based on their host origin, suggesCng that host-microbe co-evoluCon may also
influence the nature of microbe-microbe interacCons. Studying how microbes interact with each other
at different taxonomic levels and with the host will allow us to predict how the introducCon of
species/strains into the microbiome could change its composiCon. Such knowledge in turn could help
guide the development of novel techniques to manipulate the microbiome to improve health.
An important mechanism of bacterial interacCon involves the use of secreCon systems to transport
“effector” proteins into the extracellular medium or directly into adjoining cells. The type VI secreCon
system (T6SS) is one such molecular machine that many gram-negaCve bacteria use to translocate
effectors into other bacteria, host cells, or the environment. In the fourth chapter of this thesis, I
idenCfy and geneCcally characterize the T6SS of Phocaeicola sartorii from the wild mouse gut
microbiome. By studying the differences between the T6SS of this species across different strains, I
also idenCfy putaCve novel T6SS effectors and immunity proteins and speculate on the evoluConary
history of the T6SS in this species. In chapter V, I take a broader look at the types and prevalence of
T6SS among bacteria across phyla. With an extensive literature review, I speculate on the ecological
and evoluConary factors that determine the observed presence-absence paiern of T6SS across
bacterial taxa. The presence and absence of T6SS among bacteria within a community can have notable effects on the community structure and funcCon, as bacteria oYen use the T6SS as a weapon to inhibit,
kill, or outcompete nearby bacteria. Furthermore, the T6SS, owing to its modular nature and specific
acCvity, can be a potent tool to manipulate the microbiome precisely and accurately. Therefore,
studying the presence-absence paierns and ecological funcCons of T6SS among natural microbial
communiCes could be useful to develop novel methods for manipulaCng the microbiome composiCon.
