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The gut microbiome plays a vital role in human health. Disturbances of this microbial system is associated with diseases such as obesity and inflammatory bowel disease. In populations of microbial species, many organisms partake in byproduct cross-feeding interactions, where byproducts from one organism are consumed by other microbes. Using the digital evolution software Avida, I studied the effect of recurring perturbations and imperfect resource conversion on the evolution of byproduct cross-feeding chains in digital communities. To investigate the effect of perturbation and conversion rate on digital organisms, I evolved digital communities for 200,000 updates in an unperturbed environment that could hold 50 different resource types, each produced as a byproduct of consuming another resource. At 200,000 updates, 50 or 60 % of all organisms were removed at various intervals during periods of different lengths, with a conversion rate less than 100 % between resources in the byproduct chain. I found that 0.9 conversion rate caused communities to evolve longer cross-feeding chains. A conversion rate of 0.5 resulted in communities with much shorter chains, more similar in length to byproduct chains in the human gut. Perturbation events seem to affect chain length only under certain conditions when energy is lost between resources, for example when 60 % of all organisms were removed every 50th update on average. It appears that conversion loss makes digital communities more robust against the effects of perturbations, and that it might protect these communities from going extinct.

The human gut microbiome is a complex ecosystem with hundreds of species interacting with each other and the host. One function of the microbiome is to break down undigested nutrients into smaller nutrients, sometimes available for uptake by the host. The digestion of such macromolecules can involve several species where one feeds on another’s byproducts, forming a large cross-feeding network. The method of digital evolution can be of great aid in studying such complex ecosystems by creating models of the studied system. In this study, the digital evolution software Avida was used to study the effects of perturbations in the system on byproduct cross-feeding chain length. Intense perturbations were found to shorten the chain lengths in general whereas weaker perturbations had either a small or no effect. When perturbations ceased, most byproduct chains displayed recovery to lengths similar to the preperturbation lengths. This indicates that byproduct chain lengths may be kept short by common ecological mechanisms alone, explaining why very long chains are rarely observed while still theoretically possible.

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The emerging evidence on the interconnectedness between the gut microbiome and host metabolism has led to a paradigm shift in the study of metabolic diseases such as obesity and type 2 diabetes with implications on both underlying pathophysiology and potential treatment. Mounting preclinical and clinical evidence of gut microbiota shifts, increased intestinal permeability in metabolic disease, and the critical positioning of the intestinal barrier at the interface between environment and internal milieu have led to the rekindling of the “leaky gut” concept. Although increased circulation of surrogate markers and directly measurable intestinal permeability have been linked to increased systemic inflammation in metabolic disease, mechanistic models behind this phenomenon are underdeveloped. Given repeated observations of microorganisms in several tissues with congruent phylogenetic findings, we review current evidence on these unanticipated niches, focusing specifically on the interaction between gut permeability and intestinal as well as extra-intestinal bacteria and their joint contributions to systemic inflammation and metabolism. We further address limitations of current studies and suggest strategies drawing on standard techniques for permeability measurement, recent advancements in microbial culture independent techniques and computational methodologies to robustly develop these concepts, which may be of considerable value for the development of prevention and treatment strategies.

The bacteria in host-associated microbial communities influence host health through various mechanisms, such as immune stimulation or the release of metabolites. However, viruses that target bacteria, called bacteriophages (phages), may also shape the animal microbiome. Most phage lifecycles can be classified as either lytic or temperate. Lytic phages infect and directly kill bacterial hosts and can directly regulate bacterial population size. Temperate phages, in contrast, have the potential to undergo either a lytic cycle or integrate into the bacterial genome as a prophage. As a prophage, the phage may alter bacterial host phenotypes by carrying novel genes associated with auxiliary metabolic functions, virulence-enhancing toxins, or resistance to other phage infections. Lytic phages may also carry certain auxiliary metabolic genes, which are instead used to takeover bacterial host functions to better accommodate the lytic lifecycle. In either case, the ability to alter bacterial phenotypes may have important ramifications on host-associated communities. This dissertation focused on the genetic contributions that phages, and particularly prophages, provide to the bacterial members of two separate host-associated communities: the honey bee (Apis mellifera) gut microbiome and the amphibian skin microbiome. My second chapter surveyed publicly available whole genome sequences of common honey bee gut bacterial species for prophages. It revealed that prophage distribution varied by bacterial host, and that the most common auxiliary metabolic genes were associated with carbohydrate metabolism. In chapter three, this bioinformatic pipeline was applied to the amphibian skin microbiome. Prophages were identified in whole genome bacterial sequences of bacteria isolated from the skin of American bullfrogs (Lithobates catesbeianus), eastern newts (Notophthalmus viridescens), Spring peepers (Pseudacris crucifer) and American toads (Anaxyrus americanus). Prophages were additionally identified in publicly available genomes of non-amphibian isolates of Janthinobacterium lividum, a bacteria found both on amphibian skin and broadly in the environment. In addition to a diverse set of predicted prophages across amphibian bacterial isolates, several Janthinobacterium lividum prophages from both amphibian and environmental isolates appear to encode a chitinase-like gene undergoing strong purifying selection within the bacterial host. While identifying the specific function of this gene would require in vitro isolation and testing, its high homology to chitinase and endolysins suggest it may be involved in the breakdown of either fungal or bacterial cellular wall components. Finally, my fourth chapter revisits the honey bee gut system by investigating the role of geographic distance in bacteriophage community similarity. A total of 12 apiaries across a transect of the United States, from Virginia to Washington, were sampled and honey bee viromes were sequenced, focusing on the lytic and actively lysing temperate community of phages. Although each apiary possessed many unique bacteriophages, apiaries that were closer together did have more similar communities. Each bacteriophage community also carried auxiliary carbohydrate genes, especially those associated with sucrose degradation, and antimicrobial resistance genes. Combined, the results of these three studies suggest that bacteriophages, and particularly prophages, may be contributing to the genetic diversity of the bacterial community through nuanced relationships with their bacterial hosts. / Doctor of Philosophy / The microbial communities of animals, called "microbiomes", play important roles in the health of animals. The bacteria in these microbiomes can help strengthen the immune system, provide resistance to dangerous pathogens, and break down nutrients. However, bacteria are not alone in the microbiome; viruses are also present. Surprisingly, the vast majority of the world's viruses, even those living inside animals, infect bacteria. These viruses, called "bacteriophages" or "phages", can impact the bacterial communities in a microbiome. Phages can be grouped in to two broad categories based on lifecycle. Lytic phages kill the bacterial host directly after infection. Temperate phages, on the other hand, can either immediately kill the host like lytic phages or alternatively, become a part of the bacterial genome and live as prophages. Phages with both lifecycles can sometimes carry genes that, although not essential to the phage, may change the traits of the bacteria during infection. For example, some phages carry toxin genes, which bacteria use to cause disease in animals. Other phages might carry genes that provide antibiotic resistance or alter the metabolism of the infected bacteria. If a phage gene benefits the infected bacteria, the bacteria may begin interacting with its environment in a new way or may even become more abundant. Alternatively, phages that directly kill infected bacteria may have a negative effect on bacterial population sizes. To begin unraveling how phages influence bacterial species in microbiomes, I investigated two different animal systems: the Western honey bee (Apis mellifera) gut microbiome and the amphibian skin microbiome. I first identified prophages of several common bacterial species that reside in the honey bee gut (Chapter 2). Prophages were more common in certain bacterial species than others, and some possessed genes associated with the breakdown of sugars or pollen, suggesting they help honey bees process their food. Using similar techniques, I then identified prophages in bacteria isolated from the skin microbiomes of several amphibian species common in the eastern United States (American bullfrogs, Eastern newts, Spring peepers, and American toads) (Chapter 3). Most notably, the bacteria Janthinobacterium lividum may benefit from prophages that carry genes for potentially antifungal chitinase enzymes that destroy the fungal cell wall. Finally, I returned to the honey bee gut microbiome system by investigating how honey bee bacteriophage communities change over large geographic distances (Chapter 4). This study, which examined honey bees from 12 apiaries sampled from the east to west coast of the United States, looks primarily at lytic phage and temperate phage that are not integrated as prophage, but are instead seeking a bacterial host to infect. I found that nearby apiaries tended to have more similar communities of bacteriophages, compared to apiaries far away. Additionally, most bacteriophage communities carry genes associated with the breakdown of sugars like sucrose. Overall, these three studies show that phages, and especially prophages, contribute to the genetic landscape of the microbiome by broadly providing bacterial hosts with access to a diverse set of genes.

Le microbiome intestinal humain est une importante communauté de microorganismes, spécifique aux individus et aux populations, dont la composition est influencée par de nombreux facteurs, tels que la génétique et les habitudes de vie de son hôte. La diète est cependant un élément majeur façonnant sa structure. Les influences de plusieurs diètes humaines sur le microbiome ont été largement investiguées. Toutefois, l’impact des variations saisonnières inhérentes à certaines diètes est peu connu. La diète traditionnelle inuite est un exemple de régime alimentaire riche en graisses et protéines animales qui varie temporellement en fonction de la disponibilité saisonnière des ressources. Afin d’étudier les dynamiques temporelles du microbiome intestinal inuit en réponse à la diète traditionnelle, des échantillons de papier hygiénique contenant des selles ont été récoltés auprès d’un groupe de volontaire Inuits du Nunavut (Canada) durant huit mois. Un groupe contrôle de Montréalais (Québec, Canada) de descendance européenne, consommant une diète typiquement occidentale, a également été sollicité. La diversité et la composition du microbiome ont été caractérisées par le séquençage de la région V4 de l’ARNr 16s. Les microbiomes obtenus par un échantillonnage de papier hygiénique et de selles ont été comparés. Ces deux méthodes offrent des représentations similaires mais non-identiques du microbiome intestinal. À partir du séquençage d’échantillons de papier hygiénique, nous avons trouvé que les variations inter-individuelles du microbiome sont plus importantes que les variations intra-individuelles au sein de Montréal et du Nunavut. Des différences significatives de la composition du microbiome s’expliqueraient par la consommation différentielle de certains groupes alimentaires. Bien qu’aucune différence saisonnière marquée n’ait été observée, en termes de composition, le microbiome fluctue davantage à travers le temps chez les individus inuits. Ces résultats suggèrent que le microbiome inuit pourrait être façonné par une diète plus variable. Ensemble, nos résultats suggèrent que la diète traditionnelle a encore un impact important sur la composition, la diversité et la stabilité de microbiome inuit, malgré les transitions alimentaires vécues au Nunavut. / The human gut microbiome represents a diverse microbial community specific to individuals and populations, which is heavily influenced by factors such as genetics and lifestyle. Diet is a major force shaping the gut microbiome, and the effects of dietary choices on microbiome composition have been thoroughly investigated. It has been shown that a change in diet also changes the gut microbiome, but the effects of seasonal diets are poorly known. The traditional Inuit diet is primarily based on animal products, which vary seasonally based on prey availability. To investigate the dynamics of the Inuit diet over time, we collected gut microbiome samples from Inuit volunteers living in Resolute Bay (Nunavut, Canada), and compared them to samples collected from individuals of European descent living in Montréal (Québec, Canada) and consuming a typical Western diet. We sequenced the V4 region of the 16S rRNA gene to characterize the diversity and composition of the Inuit microbiome, and surveyed differences among samples collected with toilet paper or from stool. Our results show that these sampling methods provide similar, but non-identical portraits of the microbiome. Based on sequencing from toilet paper samples alone, we found that inter-individual variations of the microbiome community composition were greater than within-individual variations, both in Nunavut and Montreal, with significant differences in microbiome explained by dietary preferences. No defined seasonal shift of microbiome composition was detected in samples collected over time. However, within-individual microbial diversity fluctuated more with time in Nunavut than in Montreal. Together, these results underline that the traditional Inuit diet still has an important impact on the composition, diversity and stability of the Inuit gut microbiome, even if the traditional seasonality of the diet is less pronounced than expected, due to an increasingly westernized diet in Nunavut.