Chronic systemic inflammation increases with age and is associated with late life diseases (e.g. sarcopenia, and frailty) but the mechanisms causing systemic inflammation are largely unknown. Our laboratory has shown that the aged microbiome increases intestinal permeability which allows bacterial products into the circulation, thus causing systemic inflammation. We do not, however, know which microbes drive this phenomenon and ultimately impact healthy or unhealthy aging. To determine the degree to which frailty, sarcopenia, and systemic inflammation can be accelerated or exacerbated via the microbiome, we colonized germfree (recipient) mice with ‘young’ (≤6 months) or ‘old’ (≥ 18 months) microbiota from specific pathogen free mice. Initially, we investigated the impact of recipient age by colonizing young and old germfree mice. Differences in sarcopenia and cellular inflammation were driven by recipient age, not microbiota age, after 6 weeks of colonization, while frailty decreased in old mice colonized with young microbiota. To further investigate the impact of the microbiota in aging, we colonized middle-aged (10-14 month) germfree mice and assessed them 6 weeks and 6 months post colonization. The aged microbiota drove an increase in frailty after 6 months of colonization. To understand the differences between young and old microbial compositions we quantified short-chain fatty acids and sequenced 16S rRNA from fecal pellets of young and old mice. We used frailty, sarcopenia, and cellular inflammation data to identify relationships with short-chain fatty acids and the microbial community. We have identified specific microbes that correlate with age, frailty, sarcopenia, and cellular inflammation from Lachnospiraceae, Akkermansiaceae and Rikenellaceae families. By understanding the role of the microbiome in healthy and unhealthy aging we can develop therapeutics to combat chronic systemic inflammation and prevent and/or reverse poor health outcomes. / Thesis / Master of Science (MSc)

The mangrove ecosystem is a marginal and complex ecosystem. Mangrove trees can tolerate heat, desiccation, high salinity, radiation, and anoxic conditions. The physiological features of mangroves help them tolerate these stressors, but their relationship with prokaryotic communities also plays a role in a productive mangrove ecosystem, mainly in nutrient cycling and biogeochemical transformation. In Saudi Arabia, a few studies were conducted to understand the microbial communities residing in the mangrove ecosystem. Most of the studies were focused on the sediments or rhizosphere of the most dominant species in the kingdom, Avicennia marina. In this study, the bacterial composition of two mangrove species (Avicennia marina and Rhizophora mucronata) and the relationship between them was explored using next generation amplicon sequencing of the V3-V4 region of the 16S rRNA. In both species, samples from four compartments were collected (sediments, rhizosphere, roots, and leaves). Both species had a similar microbial composition, with Proteobacteria and Chloroflexi being the most dominant phyla in all compartments. The lack of difference in alpha diversity measures (number of ASVs and Shannon-diversity index) between species highlights the symbiotic relationship between the trees. Previous studies have reported that A. marina has a more diverse microbial community than R. mucronata, however this difference was not significant in our samples. The multivariate analysis showed us that the microbial composition of the leaf and root samples was grouped separately from the microbial composition of sediment and rhizosphere samples, highlighting the specific microbial composition of each compartment. In addition, the enriched strains in each cluster were explored and related to the surrounding environment of the mangrove ecosystem, followed by the exploration of unique strains in each compartment using SIMPER analysis. In conclusion, this study provides the first information on the Red Sea Northern mangrove (Al-Wajh region) tree microbiomes, encompassing roots, leaves, rhizosphere, and sediments. Furthermore, by showing that some bacteria can colonize different plant compartments we contribute to disentangling their propagation channels within plants.

No description available.

This study investigated the microbial colonization and maternal influences on the neonatal calf gut microbiome. Microbiome samples were collected from dams (n = 6) and calves (n = 6) using sterile flocked swabs. The vaginal, oral, and fecal bacterial communities were examined from the dam and the fecal community of calves was examined from birth to 60 d of age. Microbial communities varied by anatomical location and age of the calf. Metagenomic analysis 16s ribosomal DNA revealed ten phyla associated with microbiomes of the dam and the same ten phyla associated with calf feces at various time points: Bacteroidetes, Firmicutes, Proteobacteria, Actinobacteria, Cyanobacteria, Verrucomicrobia, Spirochaetes, Tenericutes, Fibrobacteres, and Lentisphaerae. Overall, the calf meconium and fecal microbiome is influenced by a combination of the maternal vagina, oral, and fecal microbiomes. Further studies will be needed to identify the transference mechanisms of maternal microbes to offspring and the associated host-microbial interactions. / Master of Science in Life Sciences / This study investigated the microbial colonization and maternal influences on the neonatal calf gut microbiome. Microbiome samples were collected from dams (n = 6) and calves (n = 6) using sterile flocked swabs. The vaginal, oral, and fecal bacterial communities were examined from the dam and the fecal community of calves was examined from birth to 60 d of age. Microbial communities varied by anatomical location and age of the calf. Metagenomic analysis 16s ribosomal DNA revealed ten phyla associated with microbiomes of the dam and the same ten phyla associated with calf feces at various time points: Bacteroidetes, Firmicutes, Proteobacteria, Actinobacteria, Cyanobacteria, Verrucomicrobia, Spirochaetes, Tenericutes, Fibrobacteres, and Lentisphaerae. Overall, the calf meconium and fecal microbiome is influenced by a combination of the maternal vagina, oral, and fecal microbiomes. Further studies will be needed to identify the transference mechanisms of maternal microbes to offspring and the associated host-microbial interactions.

The quality of early milk produced by dams is affected by various factors (i.e. breed, age, parity, environment, nutrition, management). The impact of these factors on the quality of milk then have subsequent effects on calf health and development. Producers are responsible for following guidelines in order to ensure that they feed calves optimal quality milk in order to produce a healthy animal. They can also regulate factors such as environment and nutrition of the dam in order to produce better quality early milk. However, even after maximizing these factors there is still high mortality rate among pre-weaned calves, therefore, other factors such as mode of birth and genetics need to be studied to determine impacts on early milk quality and make further improvements to calf health and decrease mortality. Two experiments were conducted in order to study the effects of maternal and genetic factors on early milk production and to determine relationships that exist with calf health. The objective of the first study was to determine the effects that the mode of delivery had on early milk composition, and on the rumen microbiome of calves. We hypothesized that mode of birth would impact early milk composition, and, in turn, influence the microbial phyla in the calf gut. The second study had three objectives: 1) establish phenotypic relationships between colostrum composition traits, milk production traits, and calf health, 2) determine impact of breed and season on colostrum production and 3) ) elucidate the genetic parameters (i.e. heritability, genotypic, and phenotypic correlations) among colostrum production and milk production We hypothesized that colostrum composition and production differ among breeds and by season and that individual components influence calf health. Additionally, we hypothesized that colostrum quality traits (i.e. Brix score and volume) are heritable. For the first study Charolaise (CHAR; n = 23) and Angus (ANG; n = 15) dams were divided into two experimental groups; dams underwent vaginal (VD; n= 25) or cesarean (CD; n= 13) deliveries. Early milk samples were collected and quantified for protein, lactose, somatic cell count, and fatty acid concentrations. After parturition calves were separated based on dams experimental group. Rumen fluid was collected from calves on d 1, 3, and 28 post-partum. Extracted DNA from fluid were used for metagenomic sequencing (ANG calves, n=11; CHAR calves, n=13). Samples were run on the HiSeq 2500 platform as paired end reads according to Ilumina's standard sequencing protocol. A regression analysis was done in SAS using PROC GLM and regressing mode of birth on milk components for d 1,3, and 28. After, milk components found to be significantly impacted by mode of birth were regressed against microbial counts. Results showed that VD dams were more likely to have increased (P  0.05) protein, solids non-fat, and lactose on d 1 and 3, but decreased (P < 0.05) urea concentrations. Similarly, short, medium, and long-chain fatty acids were increased (P  0.05) in VD d 3 milk. Changes in true protein elicited a decrease (P  0.05) in rumen fluid Actinobacteria and Proteobacteria; whereas, both solids non-fat and lactose were associated with an increased (P  0.05) response in d 1 transition milk. No significant results for d 28 of sampling were observed. Based on our results we suggest that mode of birth influences protein concentrations in early milk. However, only a slight impact on the overall dynamics of the calf rumen was observed with the microbiome remaining relatively stable on the phyla level in response to changes in protein concentration. The second study looked into relationships between colostrum composition traits, management practices, and calf health, as well as determined heritability and genetic correlations for colostrum quality traits. Values for test-day milk, protein, fat, and somatic cell count (SCS) for Holstein (HO, n= 250) and Jersey (JE, n=289) cows were obtained from the Animal Genomic and Improvement laboratory server at the USDA. Brix score, colostrum weight, dam age, parity, and 3-month season of calving were also recorded. After, colostrum samples from JE cows were sent to DHIA where compositional measurements were obtained (i.e. true protein, fat, lactose, SCS, solid non-fats). Lactoferrin concentration for JE colostrum samples was also determined via ELISA. Calf blood samples were collected within 72 h post-partum and total serum protein (TSP) quantified to determine success of passive immunity transfer. Additionally, farm staff were instructed to record colostrum source for 1st feeding (i.e. dam, mix, other), freshness for 1st feeding (frozen vs fresh), Brix score of colostrum fed, volume of colostrum fed, and birth weight. A PROC Mixed with LSMEANS was performed in SAS to determine relationships between colostrum components, test day components, and quality traits for season, breed, and the interaction between season and breed. Also, PROC Mixed with LSMEANS was used to determine relationships of calf health with environment, management, and colostrum components. Additionally, a Pearson correlation was used to determine relationships between colostrum components and quality traits. Results for Holstein and Jersey showed that both colostrum Brix and volume (P < 0.001) differed by breed. Colostrum volume (P < 0.001), lactose (P < 0.001), and lactoferrin (P = 0.002) varied significantly by season. Additionally, test day milk (P = 0.046), fat (P = 0.012), and protein (P = 0.003) varied significantly by season. Moreover, a significant season and breed interaction (P = 0.028) was observed solely for colostrum volume. Calf health models indicated that TSP, colostrum total protein and solid non-fats impacted incidence of respiratory illness, but no factor significantly impacted incidence of scours. Results for Pearson correlation indicated strong correlations between true protein and solid non-fats and Brix (r = 0.99; 0.86). Lactoferrin also had moderate negative correlations with volume and lactose (r = -0.35; -0.33). Heritability and repeatability's were calculated using BLUPF90 family of programs. A single-trait repeatability animal model was used and included a 1-vector phenotype (Brix or Colostrum weight), fixed effects (i.e. calving year, parity, 3-month season of calving, and age at calving), additive genetic variance, random permanent environment effects, and random residual effects. A series of bivariate models were used to calculate genetic correlations of Brix score and colostrum weight with test-day compositional traits. Heritability estimates results for Holstein cow Brix and colostrum weight, were 0.25 and 0.15. Jersey cow heritability estimates were 0.36 and 0.47 respectively. We also observed some significant genetic correlations with Holstein Brix score and test-day milk (-0.23), fat (0.54), and SCS (0.29) having moderate correlations. Holstein colostrum weight had a strong correlation with test-day milk (0.96). Jerseys had strong genetic correlation of Brix score with colostrum weight (-0.98). Low to moderately heritability was observed for Brix score and colostrum weight in both breeds making them receptive to genetic selection in order to improve breeding programs. In conclusion, mode of birth significantly impacted colostrum composition which had subsequent effects on abundance of rumen microbiota. Colostrum Brix and volume were impacted by breed, season, and interaction, and calf incidence of disease was impacted by colostrum composition and environment. Additionally, two factors influencing colostrum quality (Brix score and colostrum weight) were found to be low to moderately heritable and have moderate to strong genetic correlations to compositional traits. Strong significant relationships were also found between colostrum compositional traits and colostrum quality traits. Therefore, incorporating quality traits into breeding programs has the potential to influence compositional traits which, in turn, can impact calf health and development by the interactions that exist between composition and microbial abundance in the rumen. / Master of Science in Life Sciences / Factors like breed, age, parity, nutrition, environment, and management can affect the quality of early milk produced. Many of these factors have been studied and guidelines developed in order to ensure producers feed the best quality milk to their calves which will allow for calves to develop properly. However, there is still a high mortality rate in pre-weaned calves and factors like mode of birth and genetics have not been readily studied. The purpose of our studies were to determine mode of birth impacts on composition of early milk and establish relationships between composition and rumen microbial phyla abundance. Additionally, establish relationships between colostrum composition traits, management practices, and calf health, and determine heritability and genetic correlations of colostrum quality traits to test-day composition traits. Our hypothesis was that colostrum quality traits such as Brix score and colostrum weight are heritable. We also hypothesized that mode of birth influences early milk composition and changes to composition has secondary effects to calf rumen microbial abundance. Charolaise (CHAR; n = 23) and Angus (ANG; n = 15) dams were divided into two experimental groups; dams underwent vaginal (VD; n= 25) or cesarean (CD; n= 13) delivery. Early milk samples were collected and sent to DHIA to quantify components. After parturition calves were separated based on dam's experimental group. Rumen fluid was collected from calves on d 1, 3, 28 post-partum and DNA extracted from fluid (ANG calves, n=11; CHAR calves, n=13). Results showed that VD significant differences in composition of VD and CD cows. Dams in VD group were more likely to have increased (P  0.05) protein, solids non-fat, and lactose but decreased (P < 0.05) urea concentrations. Similarly, short, medium, and long-chain fatty acids were increased (P  0.05) in VD. Changes in true protein elicited a decrease (P  0.05) in rumen fluid Actinobacteria and Proteobacteria. Results suggest that mode of birth influences protein concentrations in early milk and induces a slight impact on the overall dynamics of the calf rumen microbiome. A second study was conducted to establish relationships between colostrum components, management and calf health as well as determine genetic parameters of colostrum quality traits. Holstein (HO, n= 250) and Jersey (JE, n=289) cow test-day data was obtained from the Animal Genomic and Improvement laboratory server at the USDA. Brix score, colostrum weight, dam age, parity, and 3-month season of calving were also recorded. Colostrum samples from JE cows were sent to DHIA where compositional measurements were obtained (i.e. true protein, fat, lactose, SCS, solid non-fats). Lactoferrin concentration for JE cow colostrum samples was also determined via ELISA. Calf blood samples were collected within 72 h post-partum and TSP quantified. Farm staff recorded colostrum source for 1st feeding and colostrum freshness for 1st and feeding. A PROC Mixed was performed to determine impact of test-day milk composition traits on colostrum quality traits by breed, PROC Mixed with LSMEANS was used to determine relationships of environment, colostrum management, and colostrum components with incidence of scours and respiratory disease in calves. A Pearson correlation was used to determine relationships between colostrum components and quality traits Heritability and repeatability's were calculated using BLUPF90 family of programs. A series of bivariate models were used to calculate genetic correlations of Brix score and colostrum weight with test-day compositional traits. Results indicated that colostrum Brix and volume were impacted by season, breed, and the interaction of breed and season. Calf incidence of disease was impacted by colostrum components and total serum protein levels. Results for Pearson correlation indicated strong correlations between true protein and solid non-fats and Brix (r = 0.99; 0.86). Lactoferrin also had moderate negative correlations with volume and lactose (r = -0.35; -0.33). Heritability estimates results for Holstein Brix and colostrum weight were 0.25 and 0.15. Jersey cow heritability estimates were 0.36 and 0.47, respectively. We also observed some significant genetic correlations with Holstein Brix score and test-day milk (-0.23), fat (0.54), and SCS (0.29) having moderate correlations. Holstein colostrum weight had a strong correlation with test-day milk (0.96). Jerseys had strong genetic correlation of Brix score with colostrum weight (-0.98). Results indicated a low to moderately heritability for Brix score and colostrum weight in both breeds making them receptive to genetic selection in order to improve breeding programs. Strong significant relationships were also found between colostrum compositional traits and colostrum quality traits.

The human gut is colonized by a vast bacterial community that is currently rather well characterized at the species level. Yet, each of these species harbor a tremendous amount of individual genetic variations. Our understanding of the human gut microbiome, its dynamics, composition and impact on host health requires a deeper characterization of its bacteria. The amount of publicly available shotgun sequencing data as well as development of computational tools allowed to reach strain-level resolution in metagenomic analysis. In this thesis, I present systematic approaches to study the strain-level variation using complementary phylogenetic and pangenomic methods aiming to address fundamental questions about microbiome transmission in early life as well as impact of functions encoded by microbiome strains on host health. Across two different cohorts, I used a recently-developed strain-tracking method to assess the impact of delivery conditions on the initial seeding of the infant gut microbiome. While mode of delivery (vaginal or C-section) had a great impact on the amount of mother strains transmitted to the infant, place of delivery (home or hospital) and breastfeeding duration also had an impact on the ongoing development, strain replacement or persistence over the first year of life. In comparison, the father appeared as a stable source of strains independent of the delivery mode. This initial mother seeding, despite being reduced in C-section delivery, can be compensated by Fecal Microbiota Transfer, demonstrating the need of fecal microbiota exposure in seeding during vaginal delivery. In addition, strain dynamics was shown partially explained by differences in the carbohydrates degrading capacities, especially the ability to feed on Human Milk Oligosaccharides. These differences in metabolism between strains were also observed by their respective empirical growth rate that was seen associated with transmission and persistence in the infant gut. To further systematically assess the differences of metabolic capacities between strains and the impact on hosts, I developed a new method to identify gene groups (PanPhlAn Genomic Islands, PGIs) co-present across conspecific strains in metagenomic samples. By applying this method on a large collection of over 10,000 samples, I was able to build a set of 5,315 PGIs. Deeper characterization of these PGIs revealed horizontal gene transfer across species, high variation in carbohydrate metabolism capacities and association with the host lifestyle and health status. Together, these analyses demonstrated the complementary aspects of strain variation andstressed out the need to encompass both strain phylogeny and gene content to fully understand the microbiome at the strain-level.

Plant-associated microbes (collectively the microbiome) are important contributors to plant health. They are known to play roles in increasing yield via improving stress tolerance, promoting growth, and suppressing the activity of plant pathogens. We investigated the wheat seed-head microbiome (phytomicrobiome) as a potential source of resistance to Fusarium head blight (FHB), or scab. FHB is a devastating disease in wheat, and other cereal grains, that causes losses in both quantity, through reduced yield, and quality of grain, through the production of toxins such as Deoxynivalenol. Efforts to combat FHB have focused primarily on breeding cultivars with resistance and applying fungicides. However, new resources for combatting FHB may lie in microbiome-plant interactions. To explore host-microbiome-pathogen interactions, we used field trials to characterize the seed head bacterial community (16S rRNA gene amplicons) across planting locations, host resistance genotypes, varieties, and plant development stages. We identified bacterial amplicon sequence variants (ASVs) present in each sample and then examined ASV community composition based on our variables. Characterizing bacterial relative abundance across samples, we identified 9,063 ASVs. These ASVs clustered according to plant developmental stages or maturity plant, location, and host genotype, but not by variety or maturity group. First, comparing plants at the pre-flowering versus mature grain-head stage, we found that both bacterial community richness and evenness changed significantly. In addition to these developmental changes, we found that bacterial community structure changes across locations, even between locations. Finally, we found that, in the presence of the pathogen, ASVs cluster by host resistance genotype, and that there are important taxonomic groups that are differentially abundant in the presence of the pathogen. Overall, we found that the wheat grain-head microbiome is shaped by environment-host-pathogen interactions, and that these interactions lead to differential abundance of particular community members that may be important in the management of FHB. / Master of Science in Life Sciences / Plant associated microbes are important contributors to plant health. They are known to play roles in increasing yield via improved stress tolerance, promoting growth, and suppressing plant disease. We investigated the wheat grain-head microbial communities as a source of disease resistance. The disease is called Fusarium Head Blight (FHB) and is caused by Fusarium graminaerum. FHB is a devastating disease in wheat and other cereals, causing losses, through reduced yield and quality through the production of toxins that prohibit use of the grain. To combat FHB, research has focused on developing plants that have resistance and the application of chemical fungicides. However, new resources for combating FHB may lie in the interactions between plants and microbes. This research is focused on identifying microbes that naturally interact with the plant, and how the pathogen, Fusarium, interacts with these beneficial microbes. In field trials, we characterized the microbial community by DNA sequencing technologies across locations, wheat with varying levels of genetic resistance, and wheat developmental stages. First, between the wheat kernel samples of pre-flowering and maturity, we found significant differences in microbial community. Consistent with other studies we found that the largest changes in microbial community composition across different growing locations. Finally, we found an interaction between the grain head microbiome and host resistance state when plants were exposed to the pathogen. Overall, we find that the wheat grain head microbiome is shaped by growing location and through interactions with the plant host and pathogen.

The host microbiome (mainly bacteria) is essential for host immune function, metabolism, and digestion. Alterations in these microbes, known as dysbiosis, generally results in adverse effects to the host, including diseases. Dysbiosis can be induced from exposures to various anthropogenic contaminants including constituents of municipal wastewater treatment effluents (MWWEs), namely, pharmaceuticals, antibiotics, and excess nutrients. Despite MWWEs being one of the largest dischargers to aquatic ecosystems, impacts of these contaminants on exposed organism microbiomes, especially in aquatic insects, is unclear. In addition, some aquatic contaminants may transfer to riparian habitats through predation on emergent insects that were exposed to contaminants as larvae, and subsequently alter microbiomes of terrestrial predators. Our study evaluated whether MWWEs altered microbiomes of freshwater larval and adult insects and their riparian spider predators using effluent-associated bacteria and stable nitrogen isotopes (δ15N) to confirm effluent exposure. We analyzed microbiome compositions through sequencing of the V3-V4 hypervariable region of the 16S rRNA gene and analyzed food web dynamics with stable carbon (δ13C) and nitrogen (δ15N) isotopes. We found that insects and spiders were enriched in δ15N at one site downstream of wastewater outfalls, indicating exposure to effluents and transfer of nutrients to terrestrial ecosystems. Microbiomes of most larval and adult insects were altered downstream of wastewater outfalls and had lower relative abundances of endosymbiont bacteria, shifts in bacterial diversities, increases in abundances of effluent-associated bacteria, and downregulation of some biosynthesis pathways than those collected at upstream sites. However, spider microbiomes had little evidence of dysbiosis, and were distinct from those of adult insects, despite a close association in their isotopic signatures. Overall, this study provides evidence of biological impacts from MWWEs to exposed insects and suggests that changes in microbial communities of invertebrates may be used as an effective indicator of effluent exposure as part of monitoring frameworks. / Thesis / Master of Science (MSc) / Municipal wastewater treatment facilities release contaminants into aquatic ecosystems that may affect the health of exposed organisms, including their microbiome, which contains bacteria essential for host digestion and immune function. Alterations in microbiomes of contaminant-exposed aquatic insects are poorly characterized and such effects may be transferred to terrestrial ecosystems through emergent insects. This study evaluated microbiomes and food web dynamics of freshwater insects and riparian spiders upstream and downstream of wastewater treatment facilities in the Bow River, AB. Results indicate that microbiomes of some downstream aquatic insects had lower relative abundances of endosymbiont bacteria, shifts in bacterial diversities, and increases in abundances of effluent-associated bacteria than those collected at upstream sites, but no such changes were observed in the spider predators. This study improves our understanding of how freshwater insect microbiomes are altered by municipal wastewater effluents and suggests that directly effluent-exposed organisms are more at risk of dysbiosis.

Epilepsy is one of the most common neurological disorders worldwide. This neurological disorder is characterized by spontaneous recurrent seizures and impacts about 65 million people globally. As there is no cure for epilepsy, the treatment goal for patients is seizure management, and ultimately seizure freedom. The first line of defense in seizure management is anti-epileptic drugs, which aim to restore the excitatory and inhibitory balance in the brain. Unfortunately, about 30% of people with epilepsy are drug resistant, a number which has remained unchanged despite the increasing amount of anti-epileptic drugs. This leads patients to seek alternative treatments, which include surgery, vagus nerve stimulation, or diet alterations such as the ketogenic diet. Due to the invasiveness of surgeries, difficulty to maintain specialty diets, or lack of effectiveness of these treatments in some patients, additional therapies are needed. The gut-brain axis is a bidirectional communication network connecting the central and enteric nervous systems. Part of this network includes communication via the gut microbiota. The gut microbiota consists of all the microorganisms living in the gut, including bacteria, viruses, and fungi. It is involved in aiding nutrient absorption, promoting the maturation of immune cells and functions, and protection against pathogens. There is growing interest in the role of the gut microbiome in human health and disease. Studies have shown that patients with epilepsy have altered gut microbiomes compared to healthy controls, and that gut microbiome alteration can impact seizure frequencies. These exciting findings have ignited research on the potential therapeutic role of the gut microbiome in epilepsy. Although studies have explored the impact of alterations in the gut microbiome on seizure activity, they have not studied how anti-epileptic drugs may contribute to this relationship. Thus, this dissertation explores the role of the commonly prescribed anti-epileptic drug topiramate on the gut microbiome. Fecal samples of mice treated with topiramate were analyzed using 16S ribosomal RNA gene sequencing. Analysis revealed that topiramate ingestion increased the probiotic bacteria Lactobacillus johnsonii in the gut microbiome. In addition, cotreatment of topiramate and Lactobacillus johnsonii reduced seizure susceptibility in a pentylenetetrazol-kindling seizure model. Moreover, cotreatment increased the butyrate producing family Lachnospiraceae and subsequently increased the neuroprotective SCFA, butyrate in the gut microbiome. Importantly, cotreatment also resulted in an increased GABA/glutamate ratio in the cortex of mice that underwent pentylenetetrazol-kindling. These results are the first to demonstrate that the anti-seizure effect of topiramate may be facilitated by the modulation of the gut microbiota via increasing butyrate and altering the GABA/glutamate ratio in the cortex. Lastly, this work highlights the potential for probiotics as an adjuvant therapy in seizure management. / Doctor of Philosophy / Epilepsy is one of the most common neurological disorders worldwide. This neurological disorder is characterized by spontaneous recurrent seizures and impacts about 65 million people globally. As there is no cure for epilepsy, the treatment goal for patients is seizure management, and ultimately seizure freedom. The first line of defense in seizure management is anti-epileptic drugs, which aim to restore the excitatory and inhibitory balance in the brain. Unfortunately, about 30% of people with epilepsy are drug resistant, a number which has remained unchanged despite the increasing amount of anti-epileptic drugs. Due to this unmet need, epilepsy patients utilize alternative treatments, which include surgery, vagus nerve stimulation, or diet modifications such as the ketogenic diet. Due to the invasiveness of surgeries, difficulty to maintain specialty diets, or lack of effectiveness of these treatments in some patients, additional therapies are needed. The gut microbiota consists of all the microorganisms living in the gut, including bacteria, viruses, and fungi, which can be both harmful and helpful. In healthy individuals, the gut microbiota coexists in a balance that prevents diseases and helps the host, however, disruptions in this balance can lead to susceptibility to several diseases. As a result, researchers are increasingly interested in the role of the gut microbiota in human health and disease. In epilepsy, the gut microbiome is altered compared to healthy individuals, and gut microbiome alterations can impact seizure activity. This has led researchers to investigate the potential therapeutic role of the gut microbiome in epilepsy. Although studies have explored the impact of alterations in the gut microbiome on seizure activity, they have not studied how anti-epileptic drugs may contribute to this relationship. Thus, this dissertation explores the role of the commonly prescribed anti-epileptic drug topiramate on the gut microbiome. The results demonstrate that topiramate increases probiotic bacteria in the gut microbiome of mice. Moreover, this probiotic bacterium facilitates topiramate in reducing the susceptibility to seizures in a mouse model by resulting in a beneficial gut microbiome and restoring excitatory and inhibitory balance to the brain. These results are the first to demonstrate that the anti-seizure effect of topiramate may be facilitated by the gut microbiome. Lastly, this work highlights the potential for probiotics as an adjuvant therapy in seizure management.

Alkyl dimethyl benzyl ammonium chloride (ADBAC) and didecyl dimethyl ammonium chloride (DDAC) are common quaternary ammonium compounds used as disinfectants in households, medical, and restaurant settings. They cause occupational skin and respiratory hazards in humans, and developmental and reproductive toxicity in mice. They also cause increased secretions of proinflammatory cytokines in cell lines and vaginal inflammation in porcine models; but have not been evaluated for developmental immunotoxicity. We assessed immunotoxicity in-vitro with J774A.1 murine macrophage cell line by analyzing cytokine production and phagocytosis; and evaluated developmental immunotoxicity in CD-1 mice by analyzing antibody production. Additionally, because of the associations between gut microbiome dysbiosis and immune disease, we monitored changes in the microbiome as a result of ADBAC+DDAC exposure. Production of cytokines TNF-alpha and IL-6 increased at low ADBAC+DDAC concentrations, and IL-10 decreased in the murine macrophages with ADBAC+DDAC exposure. The phagocytic function of macrophages was also severely decreased. ADBAC+DDAC altered the mouse microbiome by decreasing the relative abundance of Bacteroides and increases in Clostridia in F0 and F1 generations. IgG primary and secondary responses were altered in F1 male mice; and IgA and IgM production were decreased in secondary response in F2 male mice. Since ADBAC+DDAC show signs of immunotoxicity in mice, further studies are needed to reassess risk for human exposure as ADBAC+DDAC may be contributing to immune disease. / Master of Science / Disinfectants are used every day in households, hospitals, and restaurants. Two common ingredients in disinfectants are alkyl dimethyl benzyl ammonium chloride (ADBAC) and didecyl dimethyl ammonium chloride (DDAC). These chemicals can cause asthma and allergic dermatitis in humans. In animals, they cause reduced fertility, altered development, and tissue inflammation. Disinfectant exposure could potentially alter bacterial populations in the gut. Altered microbial populations are associated with many inflammatory diseases. This study evaluated ADBAC and DDAC for their ability to alter immune function and change bacterial populations in the gut. Exposure to ADBAC and DDAC caused inflammation and altered antibody production for two generations. ADBAC and DDAC exposure also significantly altered bacterial communities in the gut. Both changes in the immune function and changes in the gut bacteria could contribute to inflammatory disease. Humans are exposed frequently to ADBAC and DDAC. If these chemicals alter immune function in humans, they could be contributing significantly to human disease.