International Scientific Conference on Probiotics and Prebiotics IPC Proceedings

International Scientific Conference on Probiotics and Prebiotics IPC 2015 Proceedings June 23rd – 25th 2015, Budapest, Hungary Organising Committee...
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International Scientific Conference on Probiotics and Prebiotics IPC 2015 Proceedings

June 23rd – 25th 2015, Budapest, Hungary

Organising Committee Alojz Bomba Peter Kurti Norbert Bomba Michaela Birkusová Beáta Mičianová

President of the IPC 2015, Slovakia Executive Director, Denmark Slovakia Slovakia Slovakia

Scientific Committee Alojz Bomba Sin-Hyeog Im Ajay A. Awati Nadiya Boyko Roy Fuller Eric Guillemard Wilhelm Holzapfel Yasuhiro Koga

President of the IPC 2015, Slovakia Chairman of Scientific Committee, Republic of Korea United Kingdom Ukraine United Kingdom France Republic of Korea Japan

Acknowledgments The Organising Committee wish to express appreciation to the Pavol Jozef Šafárik University, Košice and particularly to the Faculty of Medicine for the scientific support of IPC2015. Special thanks to Japanese Society for Probiotics Science for their encouraging co-operation and support. Particularly we would like to acknowledge our sponsors for their funding and support that enabled the conference. The successful realization of the conference is especially owed to the dedication of Prof. Alojz Bomba, Prof. Sin-Hyeog Im and Prof. Dr. Wilhelm Holzapfel.

LIST OF CONTENTS Conference Programme......................................................................................................................6 Abstracts of Oral Presentations........................................................................................................13 Abstracts of Poster Presentations.....................................................................................................55 Author Index....................................................................................................................................118 Keyword Index.................................................................................................................................121

ISBN 978-80-89589-11-1

Conference Program

IPC 2015

PR O G RAM June 22, 2015

Immunology Master Class WORKSHOP PROGRAMME Chair: Andrew Foey, Co-Chair: Sin-Hyeog Im 13:00 Andrew Foey - Lecturer in Immunology, School of Biomedical & Healthcare Sciences, Plymouth University Peninsula Schools of Medicine & Dentistry Mucosal Immunology – from Outside to Inside 14:00 Andrew Foey - Lecturer in Immunology, School of Biomedical & Healthcare Sciences, Plymouth University Mmmmm! Luminal Tasting & Antigen Presentation 15:00 Coffee Break 15:30 Sin-Hyeog Im -Academy of Immunology and Microbiology, Institute for Basic Science, Pohang University of Science and Technology (POSTECH), Republic of Korea T Cell Biology & Immune Disorders 16:30 Flow Cytometry Application 17:00 Coffee Break 17:30 Sin-Hyeog Im -Academy of Immunology and Microbiology, Institute for Basic Science, Pohang University of Science and Technology (POSTECH), Republic of Korea Microbiome and Immune Regulation 18:30 Q & A: General Discussion / Comments 19:00 Networking Cocktail

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June 22, 2015

Why Nutrition and Health Claim Applications Fail - Common Shortcomings Versus Best Practice WORKSHOP PROGRAMME Chair: Yolanda Sanz, Co-Chair: Elinor McCartney 13:00 Elinor McCartney - European Union Food Legislation Expert Find Out Why Efsa Will Reject Your Next Nutrition and Health Claims Dossier 14:00 Yolanda Sanz - Vice-Chair, Panel of Members on Dietetic Products, Nutrition and Allergies, EFSA EFSA Health Claims Process and Lessons Learnt 15:00 Coffee Break 15:20 Hans Van Der Saag -Bioactor Health Claims for SMEs it’s not Easy, but it Can be Done 15:40 Leen Allegaert - Barry Callebaut Compiling Your Dossier to Build a Solid Health Claim 16:00 Rosalind Miller - British Nutrition Foundation (BNF) Do You Plan to Apply? We Prepare a“Toolkit“ for You 16:20 Stéphane Vidry – ILSI Europe The Approach of Pathway-27 to Support SMSs in the Substantiation of Health Claims 16:40 Rosalind Malcom – University of Surrey Work Package on the REDICLAIM project 17:00 Coffee break 17:20 Interactive Breakout Groups - 5-6 groups working on particular tasks 18:40 Industry Panel - Q&A session with companies experienced in the health claims regulation BioActor Barry Callebaut Cosucra Vésale Pharma WagrAlim 19:10 Networking Cocktail and Discussion

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Conference Program

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June 23, 2015 - CONFERENCE DAY 1 7:00 Registration Desk Open 8:45 Opening Speech, Alojz Bomba, President of the Conference Session 1, Probiotics and Gut Homeostasis: Mechanisms of Action Chair: Jean-Paul Lallès, Co-Chair: Alberto Finamore KEYNOTE Lecture 9:00 Jean-Paul Lallès: Gut Wall Homeostasis: Roles and Regulations of Barrier Function, Inducible Heat Shock Proteins and Intestinal Alkaline Phosphatase 9:40 Alberto Finamore: L. Amylovorus: Mechanisms in Gut Epithelial Cells and Pig Intestine 10:00 Muriel Mercier-Bonin:Lactobacillus Farciminis and Gut Mucus O-Glycosylation 10:20 Coffee Break Session 2, Modulations of the Intestinal Micro-flora Chair: Yolanda Sanz, Co-Chair: Alojz Bomba KEYNOTE Lecture 10:50 Yolanda Sanz: Microbiome’s Influence on Energy Balance and Metabolic Disease: My New Gut Project 11:30 Kumiko Kato: Effect of Ingestion of Yogurt Supplemented with Bifidobacterium Longum on the Gut Microbiota Disturbance Caused by Animal-based Diet 11:50 Billy Hargis: Microbial Properties of GIT and Hatcher Cabinets After in Ovo or Hatcher Spray of Probiotics 12:10 Lunch Session 3, Probiotics for Immune Disorders Stimulation of the Intestinal Immune System Chair:Hiroshi Ohno,Co-Chair:Sin-Hyeog Im PLENARY Lecture I 13:40 Hiroshi Ohno:Integrated Omics Approach for Understanding the Gut Ecosystem 14:30 Elodie Neau:Combination of in Vitro and in Vivo Approaches to Select a Candidate Probiotic Strain against Allergy (YSA nominated) 14:50 Sin-Hyeog Im: Mechanisms of Action of Probiotics in Immune Modulation 15:10 Kseniya Klimina: The Preclinical Testing of Drug Formulation Based on Bifidobacterium Longum, Lactobacillus Rhamnosus and Enterococcus Faecium Strains for Treatment of Inflammatory Bowel Disease (YSA nominated) 15:30 Coffee Break Session 4, Future of Probiotics and Prebiotics – Visions and Opportunities Animal Health Chair: Philippe Thonart, Co-Chair: Ajay Awati KEYNOTE Lecture 15:20 Philippe Thonart: Scale up Procedure for Fermentation and Down Stream Processing of Probiotic Strains: A Challenge for the Industry 16:00 Flore Depeint: Combination of Two in Vitro Models to Investigate the Impact of a Pesticide (Chlorphyrifos) and Prebiotic (Inulin) on Intestinal Microbiota and Permeability of Intestinal Mucosa 16:20 Monika Mueller: Prebiotic Effect of Fructans with Different Structure and Polymerization Degree 16:40 Kanchana Poonsuk: Effect of Synbiotics on Broiler Performance, Villus Morphology and Complete Blood Count (YSA nominated)

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17:00 Karel De Winter: Turning Bulk Sugars into Prebiotics: Highly Efficient Production of Kojibiose from Sucrose and Glucose 17:20 Gordon Stanley Howarth: Factors Derived from Faecalibacterium Prausnitzii Further Promote 5fluorouracil-induced Cell Death but Increase Transepithelial Electrical Resistance in Transformed Colonic Epithelial Cells 17:40 Ajay Awati: Probiotics and Feed Enzymes: Synergy at Play 18:00 Networking bar Poster Visit

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June 24, 2015 - CONFERENCE DAY 2 SESSION 5,Biomarkers of Probiotic Efficacy Chair:Bruno Pot, Co-Chair: Ajay Awati PLENARY Lecture II 9:00 Bruno Pot:Selected Probiotics Can Limit Weight Gain and Insulin Resistance in Diet-induced Obese Mice 9:50 Valeriy Danilenko:Development of New Methods for Screening and Selection of Probiotic Lactobacillus Strains with Antioxidant Properties 10:10 Roman Yunes: Gamma-aminobutyric Acid Producing Lactobacillus and Bifidobacterium Strains Isolated from the Human Body - Candidates for Future Psychobiotics (YSA nominated) 10:30 Coffee Break SESSION 6, Legal Status and Regulatory Issues Chair: Alojz Bomba,Co-Chair: Elinor McCartney KEYNOTE Lecture 11:00 Elinor McCartney: Regulation of Probiotics for Humans in the EU - What’s New? SESSION 7,Probiotics for Intestinal Disorders Chair: Mario Guslandi, Co-Chair:Lynne McFarland 11:40 Mario Guslandi:Probiotics for Chronic Intestinal Disorders 12:05 Lynne McFarland: Probiotics for the Prevention of Antibiotic-associated Diarrhea 12:30 Hania Szajewska: Probiotics for Acute Diarrhea in Children 12:55 Lunch SESSION 8, Models to Study Intestinal Interactions Chair: Wilhelm Holzapfel, Co-Chair:Sin-Hyeog Im 14:20 Marian Maďar: The New Fluorescent Visualization Techniques for Study of the Bacterial Microflora in Gastrointestinal Tract 14:40 Mario Gorenjak: Probiotics May Ameliorate Proatherogenic Effects of the Metabolic Syndrome 15:00 Alex Galanis: Molecular Methodology for Efficient Detection and Identification of Probiotic Lactic Acid Bacteria in Novel Foods 15:20 Iñaki Iturria:Zebrafish as a Promising Model to Evaluate the Efficacy of Probiotics 15:40 Coffee Break SESSION 9,Models to Study Intestinal Interactions Future of Probiotics and Prebiotics – Visions and Opportunities Chair:Nadiya Boyko,Co-Chair:Alojz Bomba 16:10 Nadiya Boyko: Use of Designed Food in Modification of Gut Microbiota and Treatment of Type 2 Diabetes 16:30 Sae Hun Kim:Health Benefits of Probiotics via Serum Cholesterol Reduction and Immune Modulation 16:50 Rajeev Kapila: Immune Regulation by Probiotic Lactobacilli with Relevance to Health During Infancy and Aging 17:10 Robert Schiestl: Intestinal Microbiota Involved in Genetic Instability, Inflammation, Longevity and Latency of Lymphoma in Atm Deficient Mice 17:30 Wilbert Sybesma:Benefits of and Access to Locally Produced Functional Fermented Foods in Africa 17:50 Poster Visit 20:00 Conference Dinner

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June 24, 2015 - CONFERENCE DAY 2 Bacteriocins SYMPOSIUM: From Knowledge to the Innovative Applications Mode of Action of Bacteriocins Chair: Svetoslav Todorov, Co-Chair: Paul Cotter 9:35 Svetoslav Todorov: What ‘s New with these Bacteriocins? Quo Ego Vado, Populus 9:50 Djamel Drider: Potential Antiviral Activity of Lactic Acid Bacteria: Insights and Trends 10:30 Coffee Break Biopreservation and Fermentation Processes Chair: Svetoslav Todorov, Co-Chair: Djamel Drider 11:10 Svetoslav Todorov: Bacteriocinogenic LAB from Cheeses - Application in Biopreservation. Reality or Mythology? 11:50 Luis Augusto Nero: Brazilian Artisanal Food Products as Source of Bacteriocinogenic LAB 12:30 Luana Perin: Microbial Dynamics of a Minas Cheese Produced with Raw Goat Milk and Added of a Nisin Producer Lactococcus Lactis Subsp. Lactis Glc05 (YSA nominated) 12:50 Lunch Application of Bacteriocins Chair: Svetoslav Todorov, Co-Chair: Luis Augusto Nero 14:30 Paul Cotter: Finding 'New' Bacteriocins 15:10 Xing Wan: An Integration Vector for Lactococcus Lactis Counterselection Based on Class Iia Bacteriocin Sensitivity 15:30 John Renye:Bacteriocin Production in Dairy Lactic Acid Bacteria 15:50 Coffee Break 16:20 Stephanie Caroline Gordts: Labyrinthopeptins, a Novel Class of Lantibiotics, Exhibit Broad and Potent Anti-Dengue Virus Activity 16:40 Closing Speech: Svetoslav Todorov

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June 25, 2015 - CONFERENCE DAY 3 Sesion 10, Genetics, Functional Genomics, Metabolomics of Probiotics Mucosal Health and Integrity Chair: Alojz Bomba, Co-Chair:Jean-Paul Lallès 9:00 Gema Pereira-Caro:Ability of Bifidobacterium Longum and Lactobacillus Rhamnosus Probiotics to Catabolize Orange Juice Flavanones 9:20 Margaret Lorraine Britz: Starvation Stress Responses in Labtobacillus Casei and Carbon Scavening: a Proteomic Approach 9:40 Jean-Paul Lallès: Short and Long-term Effects of Early Oral Administration of Lactobacillus Amylovorus on Ileal and Colonic Permeability and Mast Cells in a Pig Model 10:00 Coffee Break SESSION 11,Gut Microbiota, Metabolic Syndrome and Probiotics Chair:Wilhelm Holzapfel,Co-Chair:Won-Jae Lee KEYNOTE Lecture 10:30 Won-Jae Lee: How Gut Microbiomes Impact Host Growth and Immunity: Lesson from a Drosophila Model 11:10 Wilhelm Holzapfel: Metabolic Syndrome, Obesity and Gut Microbiota – Towards Unraveling the Black-box 11:30 Alojz Bomba:Lipid Metabolism, Atherosclerosis and Probiotics 11:50 Susan Joyce: Bile Acid Modifications at the Microbe-host Interface 12:10 Yosep Ji: Probiotics Influence on Murine Gut Microbiota and Relationship with Host Metabolism 12:30 Lunch SESSION 12, Fermented Dairy Products to Modulate the Gut Microbiota and Health Chair: Gwénaël Jan,Co-Chair:Françoise Rul KEYNOTE Lecture 13:15 Gwénaël Jan:Starter SURFace against Inflammation of the Gut : Any Unrevealed Potential in Dairy Starters? 13:55 Françoise Rul : Yoghurt Starters in the Gut : The Story of Unexpected Probiotic Effect 14:15 Tadaaki Miyazaki: Modulation of Autoimmune and Anti-viral Immune Responses by Lactic Acid Bacteria 14:35 Muriel Thomas:The Epithelial Imprinting of Microbiota: How Intestinal Microbes Shape Functional and Structural Features of Intestinal Epithelium 15:00 Coffee Break SESSION 13, The Impact of Nutritional Interventions on Gut Barrier Function Chair:Jerry Wells,Co-Chair: Andrew Foey 15:15 Jerry Wells:The Regulation of Gut Barrier Function and Potential Biomarkers 15:55 Robert Jan Brummer:Intestinal Barrier Function in Relation to the Gut-brain Axis 16:15 Karen Krogfelt:Contribution of E. Coli Pathobionts to Epithelial Barrier Dysfunction in Inflammatory Bowel Disease 16:35 Andrew Foey: Epithelial Cell Production of B-defensin-2: The Inter-relationship between Probiotic Bacteria and Inflammatory Cytokines 16:55 Presenting Young Scientist Award Developing Scientist Poster Award Closing Speech Farewell Drink

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Abstracts of Oral Presentations

Abstracts of Oral Presentations Gut Wall Homeostasis: Roles and Regulations of Barrier Function, Inducible Heat Shock Proteins and Intestinal Alkaline Phosphatase Lallès J. INRA, France Introduction: Tight control of gut barrier function appears crucial in preserving gut homeostasis and host health. Indeed, many non-communicable diseases, including chronic gut inflammatory diseases, metabolic syndrome and obesity are thought to involve gut barrier defects and abnormal enteric entry of pro-inflammatory microbial components (e.g. lipopolysaccharide) into the body. The pathogenesis of these diseases is not fully understood yet but it may involve defects in host-microbiota crosstalk. The gut mucosa is permanently exposed to harsh environment and toxic substances and microbes. Thus, intestinal epithelial cells (IEC) have developed protection systems including notably inducible heat shock proteins (iHSPs) and intestinal alkaline phosphatase (IAP). They both contribute to enhance gut barrier protection. Related diseases may be prevented or treated (at least partially) through dietary approaches, including probiotics.Gut epithelial barrier. Gut barrier is complex, comprising the mucus, the epithelial monolayer and the enteric immune system. Gut epithelium participates in digestion and absorption while regulating body access of deleterious components. Passage of noxious substances across the epithelium is regulated by tight junctions between cells (paracellular permeability) and by transcellular uptake of macromolecules. The enteric nervous system controls this barrier while mast cells and their inflammatory mediators are involved in gut permeability alterations. Methods: Inducible HSPs. iHSPs are a broad family of cellular proteins dedicated to intracellular protein trafficking, with many functional implications, including gut barrier. While HSP25/27 interacts directly with IEC cytoskeleton, HSP70/72 also bears anti-inflammatory properties. In fact, transcription factor HSF1 that controls iHSP induction also controls gene expression of key tight junction proteins. Importantly, iHSPs are induced by diverse microbial components and related metabolites. Intestinal alkaline phosphatase. IAP acts as a major anti-inflammatory enzyme, locally as well as systemically, through two mechanisms: (i) detoxification of pro-inflammatory microbial components by dephosphorylation, and (ii) control of local (and systemic) inflammation through the down-regulation of Toll-like receptor 4 (TLR4)-triggered NF-κB activation and subsequent inflammatory cytokine production. Importantly, IAP is produced by IEC and is found in both intestinal and colonic lumen as well as in IEC cytoplasm and basolateral space and finally blood plasma. IAP has been shown to participate in gut barrier protection, though often indirectly through its anti-inflammatory properties. Probiotics and gut protection. Various probiotics, including many lactobacilli confer gut epithelial protection, at least partially through HSP25/27 and/or HSP70/72 induction. Molecular mediators include bacterial components, metabolites or other secreted molecules (e.g. peptides, polyphosphate). Some probiotics directly produce IAP-like enzymes while others can stimulate host IAP, though probably indirectly though inflammation down-regulation. Results: Conclusion. iHSPs and IAP are major drivers of gut barrier protection. Importantly, they respond to dietary factors, including probiotics and are modulated by luminal microbial signals and inflammation. Both iHSPs and IAP down-regulate inflammation enhance IEC resistance to stress and finally contribute to prevent chronic inflammatory diseases. Keywords: Probiotic, Heat Shock Protein, Intestinal Alkaline Phosphatase, Barrier Function, Gut

L. Amylovorus: Mechanisms in Gut Epithelial Cells and Pig Intestine Finamore A. Council for Agricultural Research and Economics, Italy

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Introduction: The Toll-like receptor (TLR) family plays a critical role in the host defense or in the development of inflammation by recognizing microbe-associated molecular patterns. Among these receptors, TLR4 has been associated with pathogenesis of several diseases despite the established immumodulatory activities of probiotics; their ability to inhibit the TLR signaling pathways remains controversial. Previously, we showed that Lactobacillus amylovorus DSM 16698T, a lactobacillus isolated from unweaned pigs, protected the intestinal cells from enterotoxigenic Escherichia coli (ETEC) K88 infection through cytokine regulation. Here we investigated whether the ability of L. amylovorus to counteract the inflammatory status triggered by ETEC in intestine was elicited through inhibition of the TLR4 signaling pathways. TLR4 detects Gram-negative bacteria, but recent studies identified other molecules able to bind to and activate this receptor, namely the extracellular heat shock proteins (Hsps), such as the extracellular Hsp72 and Hsp90 When released from cells, these Hsps may induce inflammation in a TLR4- and NF-kB-dependent mechanism, and circulating Hsp72 has been found increased in pathological conditions including renal disease, hypertension, atherosclerosis and sickle cell disease. Methods: We used the human intestinal Caco-2/TC7 cells and pig intestinal explants. Differentiated Caco-2 cells and pig explants were treated with ETEC, L. amylovorus or L. amylovorus cell free supernatant, either alone or simultaneously with ETEC. Expression of proteins involved in the TLR4 cascade and heat shock proteins, was evaluated by Western Blot analysis, while inflammatory cytokines IL-1β and IL-8 secretion was analyzed by cytometric bead array inflammatory kit. Results: Western blot analysis show that L. amylovorus and its cell free supernatant suppress the activation of the different steps of TLR4 signaling in Caco2 cells and pig explants by inhibiting the ETEC induced increase in the level of TLR4 and MyD88, the phosphorylation of the IKKα, IKKβ, IκB and NF-κB subunit-p65, as well as the over-production of inflammatory cytokines through modulation of the negative regulators Tollip and IRAK-M. The immunofluorescence analysis confirmed the lack of P-p65 translocation into the nucleus. We also show that L.amylovorus blocks the up-regulation of the extracellular heat shock (Hsp)72 and Hsp90, proteins critical for TLR4 function. By using anti-TLR2 antibody, we demonstrate that TLR2 is required for the suppression of TLR4 signaling activation. Discussion: In conclusion, our results provide advance in the knowledge of the mechanisms of probiotic antiinflammatory activity by demonstrating that L. amylovorus DSM 16698T and its cell free supernatant inhibit the ETEC induced activation of the TLR4 signaling pathway through modulation of the negative regulators Tollip and IRAK-M, as well as down-regulation of the extracellular Hsp72 and Hsp90, which are important for TLR4 functioning, leading to reduced pro-inflammatory cytokine production. In addition, we show that these anti-inflammatory activities are TLR2 dependent. These results may contribute to develop therapeutic interventions using L. amylovorus in intestinal disorders of piglets and humans. Keywords: L. Amylovorus, inflammation, toll-like receptors, epithelial cells, pig

Lactobacillus Farciminis and Gut Mucus O-Glycosylation Mercier-Bonin M. INRA, France Introduction: Despite well-known intestinal epithelial barrier impairment and visceral hypersensitivity in irritable bowel syndrome (IBS) patients and IBS-like animal models, structural and physical changes in the mucus layer remain poorly understood. Similarly, the probiotics-induced effect on the mucus layer in stressed-animals has been poorly investigated. Using a water avoidance stress (WAS) model in rats, we aimed at evaluating whether (i) WAS modified gut permeability, visceral sensitivity, mucin expression, biochemical structure of O-glycans and related mucus physical properties in ileum and colon, (ii) a probiotic treatment with Lactobacillus farciminis prevented these alterations. Methods: Male Wistar rats received orally L. farciminis or vehicle for 14 days. At day 10, they were submitted either to sham or 4-day WAS. Intestinal paracellular permeability and visceral sensitivity were measured in vivo. The 14

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number of goblet cells and Muc2 expression were evaluated by histology and immunohistochemistry, respectively. Adhesion of L. farciminis to the intestinal mucosa and to purified Muc2 was determined. The mucin Oglycosylation profile was obtained by mass spectrometry. Atomic Force Microscopy performed surface imaging of intestinal mucus at nanoscale. Results: A 4-day WAS induced gut hyperpermeability and visceral hypersensitivity but did not modify either the number of goblet cells or Muc2 expression in ileum and colon. In contrast, O-glycosylation of mucins was strongly affected, with the appearance of elongated polylactosaminic-chain containing O-glycan structures. Mucin sialylation and sulfation remained unchanged. These changes were associated with flattening and loss of the mucus layer cohesive properties. L. farciminis bound to the ileal and colonic mucosa, and to intestinal Muc2. Interestingly, it prevented WAS-induced functional alterations and changes in mucin O-glycosylation and mucus physical properties. Discussion: This study (1) showed, for the first time, that intestinal mucus alterations induced by a 4-day WAS were linked to a shift in O-glycosylation rather than to mucin expression changes. O-glycan modifications may influence physico-chemical interactions between mucin fibers, with a direct impact on the morphology and physical properties of the mucus network, leading to a “leaky” mucus barrier. Intestinal barrier function impairment observed under stress conditions resulted from defects of both epithelial and mucus barriers acting in concert, thus leading to visceral hypersensitivity. Furthermore, the probiotic treatment with L. farciminis prevented WAS-induced epithelial barrier disruption and biochemical and physical mucus alterations, contributing to the strengthening of the intestinal barrier function. This study provides further rationale for the use of this probiotic in gut pathologies characterized by an intestinal barrier defect, like IBS. (1) Da Silva S., Robbe-Masselot C., Ait Belgnaoui A., Mancuso A., Mercade-Loubière M., Cartier C., Gillet M., Ferrier L., Loubière P., Dague E., Théodorou V., Mercier-Bonin M.,. 2014, Stress disrupts intestinal mucus barrier in rats via mucin O-glycosylation shift: prevention by a probiotic treatment, Am. J. Physiol. Gastrointest. Liver Physiol., 307, G420–G429. Keywords: water avoidance stress, gut permeability, mucus, adhesion, L. farciminis

Microbiome Influence on Energy Balance and Metabolic Disease: My New Gut Project Sanz Y. IATA-CSIC, Spain Introduction: Obesity and its associated co-morbidities (metabolic syndrome, diabetes, etc.) constitute a major health concern among diet-related diseases since their prevalence is steadily increasing worldwide. Excessive caloric intake in relation to energy expenditure has been considered the main cause of the current obesity epidemic, although nutrient distribution may also play an important role. Emerging evidence suggests gut microbiota-diet interactions also contribute to determining the final energy yield of a diet, the fate and metabolic effects of specific nutrients and eating behavior. The associations established between specific bacterial taxa and these disorders so far are, however, inconsistent across studies and the possible mechanisms of action of the microbiota in these disorders are largely unknown in humans. The role of the microbiota in the breakdown of indigestible carbohydrates is the best characterized to date. This activity leads to the generation of short-chain fatty acids (SCFAs) and gases (e.g. hydrogen) that can also be further metabolized, activating the overall colonic fermentation and, thereby, increasing the energy extraction from the diet. This idea is, however, inconsistent with the beneficial role attributed to fiber intake and to some of the SCFAs produced by its fermentation in obesity (e.g. improved insulin sensitivity, satiety and reduced inflammation). Gut microbiota is also likely to participate in the catabolism of peptides and amino acids contributing to supply amino acids and the generation of SCFAs. The gut microbiota could also participate indirectly in lipid metabolism by interfering with the enterohepatic circulation of bile acids and directly by utilizing dietary fat that escapes digestion. Nevertheless, the understanding of the role of the microbiota in protein and lipid metabolism and their physiological consequences is very limited.

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In the My New Gut project (7th FP EU), we will deeply investigate the diet-microbiome-host metabolic communication to progress in the identification of the bacteria and pathways involved in the metabolism of different nutrients and their health effects. This information will be used for the design of intervention strategies that could modulate the microbiome composition and function and, thereby, reduce the risk of obesity and the associated co-morbidities. To do so, we will develop a multidisciplinary research approach, using functional omics-technologies and systems biology, in well-controlled human trials and humanized mouse models. This information will contribute to progressing in the development of more personalized and efficient dietary strategies to reduce the socioeconomic burden of diet-related diseases in the EU. Keywords: Microbiome, Diet, Energy Balance, Obesity, Metabolic disease

Effect of Ingestion of Yogurt Supplemented with Bifidobacterium Longum on the Gut Microbiota Disturbance Caused by Animal-based Diet Kato K. Morinaga Milk Industry, Japan Introduction: The balance of gut microbiota in healthy adult is basically stable, but can be influenced by lifestyle, psychological stress, aging, use of antibiotics and meal, etc. Animal-based diet, which was composed of meat, meat substitutes and egg, has been reported markedly to affect the intestinal environment as well as the composition of gut microbiota. Ingestion of probiotics has been well demonstrated to have function in maintaining and/or modulating the gut environment, however, its effect on microbiota disturbance caused by animal-based diet has not been investigated. Methods: Objective: In this study, we assessed the effect of ingestion of yogurt supplemented with a probiotic strain on the changes of microbiota induced by animal-based diet. Design: A total of 33 healthy Japanese subjects, aged between 20 and 50 years old, were enrolled in an open, randomized, parallel-group study. After 7 days pre-observation period, all subjects were provided with entire animal-based diet for 5 days followed by balanced diet (boxed lunch delivery) for 14 days. Following the pre-observation period, subjects were allocated into 3 groups. Subjects in the first group ingested 200g daily of Yogurt fermented with Bifidobacterium longum strain BB536 (containing more than 2 × 109 colony forming units per 100 g) together with Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus during both the Animal-based and Balanced diet period (YAB group). Subjects in the second group ingested Yogurt only during the Balanced diet period (YB group). Subjects without ingestion of yogurt throughout the intervention were used as the control (CTR group). Fecal samples were collected at day -7, 0 (before animal-based diet), 5 and 19. The fecal microbiota was analysed based on the V3-V4 region of the bacterial 16S rRNA genes using a by high-throughput sequencing technique. Results: The animal-based diet caused a significant increase in the abundance of Bilophila, Odoribacter, Dorea and Ruminococcus (belonging to Lachnospiraceae) and a significant decrease in the level of Bifidobacterium (in YB and CTR groups). These changes, with the exception of Ruminococcus, were not observed in the YAB group. A significantly higher abundance of genus Lactobacillus was observed after the animal-based diet period in YAB group. No significant effect was found by the post-animal diet yogurt supplementation (YB group). Discussion: These results demonstrated that the balance of the gut microbiota will be disturbed by an entire animalbased diet. The intake of yogurt supplemented with probiotic bifidobacteria played a role in maintaining a normal microbiota composition during the ingestion of a meat-based diet. Keywords: Yogurt, Bifidobacterium longum, Animal-based diet, Bilophila, BB536

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Microbial Properties of GIT and Hatcher Cabinets After in Ovo or Hatcher Spray of Probiotics Hargis B. University of Arkansas, United States Introduction: Early colonization of beneficial bacteria in chicks may improve GIT development and performance, but appropriate application methods need investigation. Options for early application include multiple hatchery processes, and presently in ovo and hatchery application of probiotics were investigated. Methods: In experiment 1, effects of two co-administered egg shell-origin MRS-recoverable bacteria (MRSR) on hatchability and GIT recovery of selected bacterial groups on day of hatch were evaluated. Embryos (d18) were in ovo injected with saline (CON) or MRSR. Immediately after hatch, GIT samples were aseptically collected for microbial recovery on MRS and tryptic soy agar (TSA). For hatcher cabinet application in experiment 2, combinations of either dry (D) or aqueous suspension (W) of Bacillus spores (BS) plus lactic acid bacteria (W; LAB) were sprayed at transfer only, or 3 separate time points (transfer, 20% & 50% hatch). Combinations included single spray of DBS + LAB (W;Trt1), single spray of WBS + LAB (Trt2), or 3X spray WBS + LAB (Trt3). Six (each) TSA, Rogosa (RA), and MacConkey’s (MCA) agar plates were distributed in cabinets at transfer, ~50% pip, ~20%, ~50%, and ~75% hatch to measure aerobic microbial profiles during the hatch period. Results: In experiment 1, hatchability was not affected by in ovo treatment in this study. Foregut MRS recovery of bacteria was 6.05±0.32 Log10 CFU, vs. 3.25±0.63 in CON (P