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Research has shown that our gut microbiota confers many beneficial functions, including aiding the development of the immune system, metabolism, modulating stress reactivity and behaviour. The diverse population of the gut microbiota has been shown to be heterogeneous between individuals, with host genetic factors emerging as a contributor to gut microbiota composition. Recent work suggests that microRNA may act as a mediator of communication between the host and resident gut microbiota. Here we explore host genetic differences in gut microbiota composition and fecal miRNA profiles in two inbred mouse strains BALB/C and C57BL/6, in relation to gastrointestinal homeostasis. Furthermore, we evaluate the role of host genetics in response to perturbation of the gut microbiota using broad-spectrum antibiotic treatment. Distinct differences in the gut microbiota composition evaluated by fecal 16s rRNA gene sequencing between BALB/c and C57BL/6 mice were found with notable significant differences in genera Prevotella, Alistipes, Akkermansia and Ruminococcus. Significant host genetic differences were also observed in fecal miRNA profiles evaluated using the nCounter Nanostring platform. A BLASTn analysis was used to identify conserved fecal miRNA target regions in bacterial metagenomes, which identified numerous bacterial gene targets. Of those miRNA targets that were conserved in our dataset, 14 significant correlations were found between fecal miRNA and predicted taxa relative abundance. Treatment with broad-spectrum antibiotics for a period of 2 weeks resulted in BALB/c mice exhibiting a decrease in barrier permeability while C56BL/6 barrier permeability remained unchanged, demonstrating a host-specific physiological response to antibiotics at the gastrointestinal barrier. Differential response to antibiotics was also observed in the expression of barrier regulating genes in both host strains. Individual taxa were found to respond differentially by host strain, with Parabacteroides and Bacteroides associating with changes in barrier function. Together these findings suggest that host genetics play a role in determining the host-microbe relationship in both healthy homeostatic conditions and altered microbial conditions. / Thesis / Master of Science (MSc)

The human gut and brain have a bidirectional communication that has shown to play a pivotal role in our health and disease. Literature has shown that microbiota composition and diversity can be influenced by both genetic and environmental factors, contributing to shaping an individual’s microbial composition. The current work includes analysis of the microbiome of several mouse models to better understand how gene-environmental interactions during early development can influence the composition of microbiota within the gut. Here, male and female mice from several strains (C57BL/6, Balb/C, FVB, CD1) and genetically modified mice including T-cell receptor knock out mice (TCRβ-/-δ-/-) and Fragile-X-mice (FMR1-KO) were exposed to early life stressors including lipopolysaccharide (LPS) injection on postnatal day 3 (P3) and/or overnight maternal separation on P9. Fecal samples were collected at P24 and microbiota composition was determined by amplifying the 16s rRNA gene variable 3(v3) region and sequenced using the MiSeq Illumina platform. DADA2, was used to analyze this data in R software. Among the group, strain was found to be significant among alpha and beta diversity metrics while sex and stress were found to contribute to within strain variation, which demonstrated that both genetic and environmental factors are important in shaping an individual’s microbial composition. Secondly, we also explored the role of gut microbiota on the development of the immune system in TCRβ-/-δ-/- and C57BL/6 mice. Mice that lack T-lymphocytes were found to have a lower alpha diversity, as well as separated from their wild-type controls by beta diversity. Several bacterial taxa were found to be influenced by the immune system, demonstrating a bidirectional communication between the gut and T-cells. Lastly, the influence of litter, an environmental factor on microbial composition was explored within inbred mouse strains, C57BL/6 and Balb/C. Litter was found to influence alpha diversity, in which litters among C57BL/6 exhibited the greatest variation in such diversity. Beta diversity was also found to be influenced by litter, as related litters were found to cluster together. Differences in bacterial taxa between the inbred strains were observed and a subset of those taxa were found to be influenced by litter. Hierarchical clustering and co-occurrence analysis revealed different clusters of co-occurring taxa between both strains. These findings demonstrate that environmental factors can contribute to influence the composition of microbiota. / Thesis / Master of Science (MSc)

京都大学 / 新制・課程博士 / 博士(農学) / 甲第24906号 / 農博第2569号 / 新制||農||1102(附属図書館) / 京都大学大学院農学研究科森林科学専攻 / (主査)教授 大村 和香子, 教授 髙野 俊幸, 教授 飛松 裕基 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DGAM

A healthy gut microbiome is crucial for the development, growth, and health of dairy calves; however, diarrhea in pre-weaned calves is highly prevalent, difficult to treat, and causes detrimental effects to the dairy industry. This study characterized early gut microbiota using longread-based 16S rRNA gene sequencing and investigated its associations with calf diarrhea and colostrum microbiota. The full-length 16S rRNA gene was amplified and sequenced on a Nanopore sequencer. We identified shared bacterial species in colostrum and calf feces, whose abundance in calf feces reduced with age. Diarrheic calves exhibited differing gut diversity before, during, and after diarrhea, and harbored increased bacteria resistant to the Cefotaxime antibiotic. Several bacterial species were associated with age and calf health. Additionally, a machine learning model identified bacteria to predict diarrhea. This study will be useful for the goal of reducing antibiotic use to promote gut health and prevent and treat neonatal calf diarrhea.

The relationship between measures of temperament, growth performance, and social hierarchy in finishing beef cattle were explored in two experiments. In experiment 1, high OCS (objective chute score) steers had periods of significantly higher ADG (P < 0.01), but OCS had no relationship with dominance ranking (P > 0.47). Conversely, slow exit velocity (EV) correlated with higher ranking (P ≤ 0.06), but EV had no relationship with performance (P > 0.37) in a competitive environment. Rank showed no relationship with performance (P > .58). In experiment 2, steers with fast EV had periods of decreased growth (P ≤ .06), intake (P ≤ .06), and gain:feed (G:F; P = 0.02). There were no interactions between EV, OCS, and monensin or between EV and monensin. Monensin and EV together, however, significantly impacted overall (days 0 – end) G:F (P = 0.02) and gain (P = 0.05). Overall, these studies further confirm the idea that EV affects performance as does OCS in concert with monensin. Moreover, it further confirms that different measures of temperament correlated to different aspects of performance and should not be lumped together under the general term “temperament” when describing its relationship with performance.

Η εντερική χλωρίδα θα μπορούσε να περιγραφεί ως ένα μικροβιακό όργανο, μείζονος σημασίας για την ομοιόσταση του οργανισμού, εξαιτίας της συμμετοχής του σε πολλαπλές και διαφορετικές λειτουργιές όπως η πέψη, η απορρόφηση των θρεπτικών συστατικών, η αποβολή των άχρηστων ουσιών και η φυσική ανοσία. Η σύστασή της έχει παρατηρηθεί ότι διαφέρει από άνθρωπο σε άνθρωπο. Ενώ οι κυριότεροι παράγοντες που διαμορφώνουν το τελικό μικροβιακό περιεχόμενο περιλαμβάνουν την ηλικία, το περιβάλλον, τις διαιτητικές συνήθειες, το γενετικό υπόβαθρο, την καταγωγή, την χρήση αντιβιοτικών, πρεβιοτικών ή προβιοτικών, την έκθεση σε ποικιλλία μικροβίων και τις χειρουργικές επεμβάσεις. Η ανθρώπινη εντερική χλωρίδα αποτελείται κατά κύριο λόγο από μόλις τέσσερα φύλα βακτηρίων, τα Bacteroidetes (23%), τα Firmicutes (64%), τα Actinobacteria (3%), και τα Proteobacteria (8%). Τα τελευταία χρόνια αρκετό είναι το ερευνητικό ενδιαφέρον στην μελέτη της σχέσης μεταξύ της σύνθεσης της εντερικής χλωρίδας και της παχυσαρκίας. Η έρευνα των πιθανών μηχανισμών αλληλεπίδρασης του μικροβιακού περιεχομένου και του ξενιστή εκτελείσεται τόσο με την χρήση πειραματοζώων όσο και σε ανθρώπινο επίπεδο. Έχει αποδειχθεί ότι σε παχύσαρκα άτομα υπάρχει χαμηλότερο ποσοστό Bacteroidetes και μεγαλύτερο Firmicutes, σε σύγκριση με άτομα κανονικού βάρους. Αν και έχουν επίσης διατυπωθεί αντίθετα αποτελέσματα. Οι διαιτητικές συνήθεις είναι ένας από τους κύριους παράγοντες που επηρεάζουν την σύσταση της εντερικής χλωρίδας. Παράλληλα, είναι γνωστό ότι το μοντέλο της οικογένειας και το γονεϊκό πρότυπο είναι καθοριστικής σημασίας στην διαμόρφωση διαιτητικών και διατροφικών επιλογών. Στην παρούσα μελέτη στόχος ήταν να ερευνήσουμε την επιρροή του οικογενειακού περιβάλλοντος, της μεσογειακής διατροφής και της παχυσαρκίας στην σύσταση της εντερικής χλωρίδας. Στην έρευνα συμμετείχαν 35 άτομα, από 12 οικογένειες, ηλικίας από 18 ετών έως και 77 ετών. Όλοι είναι μέλη οικογενειών που κατοικούν στην περιοχή της Πάτρας, και αφού πληροφορήθηκαν εκτενώς για τον σκοπό και την μεθεδολογιά της, συνένεσαν εθελοντικά. Μετρήθηκαν τα ανθρωπομετρικά χαρακτηριστικά κάθε εθελοντή, και υπολογίστηκε ο δείκτης μάζας σώματος (ΔΜΣ), καταγράφηκαν, μέσω ερωτηματολογίων, δημογραφικά, κοινωνικο-οικονομικά στοιχεία και οι διαιτητικές-διατροφικές τους συνήθειες. Ακόμα έγινε συλλογή κοπράνων, από όπου απομονώθηκε DNA και αναλύθηκε με την χρήση Real Time PCR. Μετά την στατιστική επεξεργασία, δεν επιβεβαιώθηκε η αρχική εκτίμηση για παρόμοια βακτηριακή κατανομή μεταξύ των μελών της κάθε οικογένειας και τελικά δεν σχετίστηκε στατιστικά σημαντικά η υιοθέτηση της Μεσογειακής διατροφής (MedDiet score) με την σύσταση της εντερικής χλωρίδας. / The intestinal flora is a microbial organ of major importance for the homeostasis of human organism, because of its participation in multiple and diverse functions such as digestion, absorption of nutrients, elimination of waste products and immunity. Gut flora is affected by various factors, such as the age, environment, dietary habits, host’s genotype, origin, using of antibiotics, prebiotics or probiotics, exposure to a variety of microbes and surgeries. Human intestinal flora consists primarily of four bacterial genders: the Bacteroidetes (23%), the Firmicutes (64%), the Actinobacteria (3%), and the Proteobacteria (8%). In the last years, many scientists study the correlation between the gut microbiota and obesity. Although, there are conflicting results, there is evidence that obese people have lower percentage of Bacteroides and higher percentage of Firmicutes, when compared with normal weight. It is known that dietary habits affect the composition of gut flora. Furthermore, it has also been proved that type of family and/or parenting model are crucial factors in shaping dietary and nutritional choices. The objective of this study was to investigate the correlation between the Mediterranean diet, obesity and family’s environment with gut microbiota. The survey involved 35 people, from 12 different families, aged from 18 years to 77 years. We measured anthropometric characteristics of each volunteer, calculated body mass index (BMI) and then demographic, socio-economic data and dietary-nutritional habits were recorded through questionnaires. We collected stool sample from every partitipant, DNA was isolated and analyzed using Real-Time PCR. Statistical analysis did not confirm the initial estimate for similar bacterial distribution among the members of each family, and eventually the adherence to Mediterranean Diet (MedDiet score) was not significally related with changes of the intestinal flora.

Gut microbial communities exist in the vast majority of animals, and often form complex symbioses with their hosts that affect their host's biology in numerous ways. To date, the majority of studies of these complex interactions have focused on the nutritional benefits provided by the microbiota; however, the natural microbiota can also influence development, immunity, and the metabolism of its host. Apis mellifera, the honey bee, harbors a distinctive bacterial community that is present in individuals from distant locations around the world; however, the basis of the bee-microbiota association is unknown. This dissertation explores properties of the bacterial microbiota within bees, including its persistence of this association, mechanisms of transmission, localization through host ontogeny, and basic metabolic capabilities that define and maintain the symbiotic relationship. Apis and Bombus species (honey and bumble bees) share a distinct bacterial microbiota that is not present in other bees and wasps. Close analysis of the A. mellifera microbiota revealed consistent communities in adult worker gut organs and a general lack of bacteria in larvae. Contact between workers and with hive materials were identified as major routes of transmission for bacterial communities, showing the importance of social behavior in this association. Genomic analysis of a gut bacterium co-sequenced with the Bombus impatiens genome revealed it as a divergent lineage of Gammaproteobacteria, and deletions of conserved metabolic pathways, reduction in genome size, and its low GC content all suggest that the bacterial species has had a long association with its host.

Variations in drug metabolism may alter drug efficacy and cause toxicity; better understanding of the mechanisms and risks shall help to practice precision medicine. At the 21st International Symposium on Microsomes and Drug Oxidations held in Davis, California, USA, in October 2-6, 2016, a number of speakers reported some new findings and ongoing studies on the regulation mechanisms behind variable drug metabolism and toxicity, and discussed potential implications to personalized medications. A considerably insightful overview was provided on genetic and epigenetic regulation of gene expression involved in drug absorption, distribution, metabolism, and excretion (ADME) and drug response. Altered drug metabolism and disposition as well as molecular mechanisms among diseased and special populations were presented. In addition, the roles of gut microbiota in drug metabolism and toxicology as well as long non-coding RNAs in liver functions and diseases were discussed. These findings may offer new insights into improved understanding of ADME regulatory mechanisms and advance drug metabolism research. (C) 2017 Chinese Pharmaceutical Association and Institute of Materia Medica, Chinese Academy of Medical Sciences. Production and hosting by Elsevier B.V.

It is well established that replacing dietary saturated fatty acids with unsaturated fatty acids reduces cardiovascular disease (CVD) risk. Although epidemiological and clinical evidence indicate health benefits of consuming various fatty acid classes including n-9, n-6, and short- and longer-chain n-3 fatty acids, current dietary recommendations fall short of providing the optimal amounts of these fatty acids in daily diets. In addition, significant knowledge gaps remain in our understanding of the effects of, and mechanisms underpinning the action of, the various fatty acid classes on risk factors for CVD. The objective of this research was to contribute to the evaluation of health benefits of using different dietary oils, and determine how these benefits may play a role in improving public health and decreasing CVD risk. Additionally, this research examined effects of diet-gene interactions, endogenous fatty acid ethanolamides (FAEs) on body fat mass distribution as well as changes in the composition of gut microbiota following consumption of dietary oil treatments. The Canola Oil Multicenter Intervention Trial (COMIT) was conducted as a 5-phase randomized, controlled, double-blind, cross-over clinical trial. Each 4-wk treatment period was separated by 4-wk washout intervals. A total of 130 volunteers with abdominal obesity consumed each of 5 identical weight-maintaining, fixed-composition diets with one of the following treatment oils (60 g/3000 kcal) in the form of beverages: 1) conventional canola oil (Canola; n-9 rich), 2) high–oleic acid canola oil with docosahexaenoic acid (CanolaDHA; n-9 and n-3 rich), 3) a blend of corn and safflower oil (25:75) (CornSaff; n-6 rich), 4) a blend of flax and safflower oils (60:40) (FlaxSaff; n-6 and short-chain n-3 rich), and 5) high–oleic acid canola oil (CanolaOleic; highest in n-9). At endpoints, plasma fatty acid levels reflected the differences in fatty acid composition of five dietary treatments. All diets lowered total cholesterol (TC) compared with baseline. TC was lowest after the FlaxSaff phase and highest after CanolaDHA. The CanolaDHA treatment improved HDL-C, triglycerides, and blood pressure thereby reducing Framingham risk scores compared with other oils varying in unsaturated fatty acid composition. Homozygotes minor allele carriers of rs174583 (TT) on FADS2 gene showed lower (P<0.01) plasma EPA and DPA levels across all diets, but no differences were observed in DHA concentrations after the CanolaDHA feeding. In addition, plasma FAE levels were positively associated with plasma fatty acid profiles. Minor allele A carriers of rs324420 of FAAH gene showed a higher (P<0.05) plasma FAE levels compared with major allele C carriers across all diets, and showed higher (P=0.0002) docosahexaenoylethanolamide levels in response to the CanolaDHA diet. Impacts of consuming 60 g of five dietary oil treatments on gut microbiota composition were relatively minor at the phylum level and mainly at the genus level, while BMI contributed to a significant shift at the phylum level. In conclusion, consumption of a novel DHA-enriched canola oil improved blood lipid profile and largely reduced CVD risk. Diet-gene interactions might help identify sub-populations who appear to benefit from increased consumption of DHA and oleic acid. The metabolic and physiological responses to dietary fatty acids may be influenced via circulating FAEs, while the altered microbiota profile by shifts in MUFA and/or PUFA may be associated with specific physiological effect. Personalized diets varying in unsaturated fatty acids composition based on specific lifestyles, environmental factors, psychosocial factors, and genetic make-ups will become the future “healthy eating” recommendations to prevent CVD risk. / May 2016