Actinobacteria: A relevant minority for the maintenance of gut homeostasis

Accepted Manuscript Title: Actinobacteria: a relevant minority for the maintenance of gut homeostasis Author: Cecilia Binda Loris Riccardo Lopetuso Gianenrico Rizzatti Giulia Gibiino Vincenzo Cennamo Antonio Gasbarrini PII: DOI: Reference:

S1590-8658(18)30210-X https://doi.org/doi:10.1016/j.dld.2018.02.012 YDLD 3676

To appear in:

Digestive and Liver Disease

Received date: Revised date: Accepted date:

18-10-2017 26-1-2018 19-2-2018

Please cite this article as: Binda C, Lopetuso LR, Rizzatti G, Gibiino G, Cennamo V, Gasbarrini A, Actinobacteria: a relevant minority for the maintenance of gut homeostasis, Digestive and Liver Disease (2018), https://doi.org/10.1016/j.dld.2018.02.012 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

Title page ACTINOBACTERIA: A RELEVANT MINORITY FOR THE MAINTENANCE OF GUT HOMEOSTASIS

Cecilia Binda*, Loris Riccardo Lopetuso*, Gianenrico Rizzatti*, Giulia Gibiino*, Vincenzo

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Cennamo^, Antonio Gasbarrini*

* Department of Internal Medicine, Gastroenterology and Hepatology, Catholic University of

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Sacred Heart of Rome, A. Gemelli Hospital - Italy.

^ Unit of Gastroenterology and Digestive Endoscopy, AUSL Bologna Bellaria-Maggiore Hospital,

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Corresponding Author:

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Electronic word count: 4120

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Bologna - Italy

Prof. Antonio Gasbarrini

Department of Internal Medicine, Gastroenterology Division, Catholic University of Rome, Policlinico “A. Gemelli” Hospital Largo Gemelli, 8

00168 Rome (ITALY)

Phone/fax number: 0039-06-30156018 E-mail: [email protected]

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Abstract Actinobacteria are one the four major phyla of the gut microbiota and, although they represent only a small percentage, are pivotal in the maintenance of gut homeostasis. During the last decade many studies focused the attention on Actinobacteria, especially on their role both in gastrointestinal and systemic diseases and on their possible therapeutic use. In fact, classes of this phylum, especially

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Bifidobacteria, are widely used as probiotic demonstrating beneficial effects in many pathological

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conditions, even if larger in vivo studies are needed to confirm such encouraging results. This

review aims to explore the current knowledge on their physiological functions and to speculate on

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their possible therapeutic role(s) in gastrointestinal and systemic diseases.

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Key words: gut microbiota, Actinobacteria, Bifidobacteria spp, gut homeostasis, dysbiosis,

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probiotic.

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INTRODUCTION The gut microbiota is composed by multiple commensal microbial species including 100 trillion (10^14) bacteria, quadrillion viruses, fungi, parasites, archeas and yeasts, reaching an overall biomass of about 1 kg and more than 3 million of genes [1- 4]. The different gastrointestinal regions are characterized by a different bio-compartmentalization with a distinct and stable microbial

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community. This is influenced by the acid environment, the presence of bile and pancreatic

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secretion and the high peristaltic activity in the stomach and small intestine, which do not allow a stable bacterial colonization, differently from the colon where bacterial colonization is favored by

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the low redox potential and the slow transit [5, 6]. The majority of microbes forming the human microbiota can be assigned to four major phyla: Bacteroidetes, Firmicutes, Proteobacteria and

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Actinobacteria [7, 8]. Firmicutes and Bacteroidetes represent more than 90% of the relative

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abundance of the gut microbiome and their relationship plays a pivotal role in the maintenance of gut homeostasis. Actinobacteria and Proteobacteria represent the remaining 10% [7, 8].

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Mainly oropharyngeal origin aerobic gram-positive bacteria inhabit stomach, duodenum and

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jejunum, whereas coliforms and anaerobic species (such as Bacteroides, Bifidobateria, Clostridia and Lactobacilli) are predominant in the ileum and post ileocecal valve, respectively [9, 10]. Gut microbiota is involved in many useful functions, such as energy production from nutrient biotransformation [11, 12], regulation of lipid metabolism [13], metabolism of vitamins and absorption of calcium, magnesium and iron [12, 14, 15], maintenance of the intestinal barrier function [16, 17], the development of immune system from the first days of life [18- 23]. Many conditions are supposed to be related to quantitative and qualitative changes in gut microbiota composition and function, such as inflammatory bowel diseases, celiac disease, irritable bowel syndrome (IBS), obesity, non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steato-hepatitis (NASH), diabetes, cardiovascular disease, arthritis, psoriasis and psychiatric disorders. This gut microbiota impairment is known as dysbiosis [24- 30].

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Recently, increasing interest has focused on the phyla of Actinobacteria, especially on Bifidobacteria family. The purpose of this review is to explore the crucial role of Actinobacteria in the maintenance of gut homeostasis and the critical importance of their future therapeutic

ACTINOBACTERIA IN THE GASTROINTESTINAL TRACT

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applications.

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Actinobacteria are Gram positive, multiple branching rods, non-motile, non-spore-forming and anaerobic bacteria [31], that include three main anaerobe families (Bifidobacteria, Propionibacteria

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and Corynebacteria) and an aerobe family (Streptomyces). The most represented in the human gut are Bifidobacteria. Interestingly, the complete genome sequences are available for certain

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Bifidobacterial species, like B. adolescents, B. animalis, B. breve, B. bifidum, B. long and B.

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angulatum [31].

Many factors can impact on the presence of Actinobacteria in the intestine. Although the relation

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between phylum diversity and delivery is still unclear, studies demonstrated a higher diversity

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within Actinobacteria phylum among children that underwent vaginal delivery [32-37]. In fact, phylum diversity within Actinobacteria, (i.e., Bifidobacterium) was significantly lower in infants delivered by caesarean section compared to vaginally delivered newborns during the first week of life [33 -35], at the age of 1 month [36], and three months [37]. Other studies did not confirm such differences or showed a lower effect on Bifidobacteria and Bacteroides colonization at the age of 6 and 12 month [38, 39]. The lower abundance of Bifidobacteria and Bacteroides in c-section delivered infants may be explained with a higher mother consumption of antibiotics before, during and after the delivery. Indeed, postnatal use of antibiotic has been associated with decreased numbers of Bifidobacterium and Bacteroides and a higher relative abundance of the Clostridium leptum [40- 42]. Furthermore, a higher abundance of Bifidobacterium genus has been demonstrated in breastfed infants [43]. Interestingly, it has been demonstrated that human milk is rich in

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substances, like human milk oligosaccharides (HMOs), that may act as probiotics and stimulate the growth of Lactobacillus and Bifidobacterium species [44]. Weaning causes a further change in microbial composition and Bifidobacteria are no longer the dominant group, with the adult microbiota established after the second year of life [45]. During the adulthood, the percentage of Actinobacteria remains stable around 8% [46] and constitutes one of

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the prevalent commensal bacterial in the distal small and large intestine [47]. Intriguingly, an Italian

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study on centenarian patients showed a reduction in the total number of anaerobes compared to young adults, with a reduction of Bifidobacteria and Bacteroides [48]. These evidences have opened

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new horizons for the concept of “fragile” microbiota that could deeply affect the delicate health

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ACTINOBACTERIA AND GUT BARRIER

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equilibrium in old patients.

Gut barrier is a multi-layer system able to afford a daily exposure to pathogens and external

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environment. It can be divided into a superficial physical barrier constituted by the epithelial cells,

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the tight junctions and the mucus; and a functional barrier represented by gut-associated mucosa lymphoid tissue (GALT), peristalsis and antimicrobial substances, able to regulate the immunological response to pathogens and tolerance to commensal bacteria [16, 49]. Gut microbiota is pivotal in the maintenance of intestinal barrier functions, increasing tight junctions expression, regulating mucin biosynthesis and catabolism, providing energy for epithelial cells proliferation and stimulating the immune system [17, 49]. In this scenario, Actinobacteria are absolute players in maintaining gut barrier homeostasis.

The production of short chain fatty acids (SCFA), such as acetate, propionate and butyrate, from carbohydrate fermentation is crucial for providing energy to epithelial cells turnover and for their potent antibacterial activity [16, 17]. In this field, Bifidobacteria have beneficial effects in the maintenance of gut barrier thanks to their great production of SCFA [50]. In particular, they produce high concentration of acetate that can protect the host from enteropathogenic infections,

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such as entero-hemorrhagic Escherichia coli and Shigella [51]. Moreover it has been demonstrated that Bifidobacteria have a non-negligible production of lactate, which can be metabolized by a group of bacteria (“lactate utilizer”) to produce butyrate [52]. In this scenario, in vitro studies demonstrated that SCFA, especially butyrate, are correlated with an increased expression of the gene MUC2, a mucin glycoprotein that is one of the major component of the mucous layer and thus

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of intestinal barrier [53-55]. The mechanism responsible for the increased production of MUC

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induced by butyrate is not definitely established. Gaudier et al demonstrated that butyrate could modify MUC gene expression in goblet cells depending on the energy source available with a

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consequent higher expression of MUC2 in conditions of glucose deprivation when the consumption of butyrate by goblet cells is increased [54]. Authors concluded that butyrate metabolism is

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involved not only in mucin synthesis but can play a role also in gene transcription. Another in vitro

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study showed a relationship between SCFA, especially butyrate, the production of prostaglandins (PG) by intestinal myofibroblasts and epithelial mucin expression [53]. In fact, SCFA seemed to

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enhance the production of PGE1 in subepithelial myofibroblasts that in turn was able to stimulate

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epithelial mucin expression, probably through the epithelial differentiation induced by cell-to-cell contact, extracellular matrix and soluble factors such as transforming growth factor β. Other important elements that contribute to the viscoelastic properties of the mucous layer are the Trefoil factors (TFFs). These mucin-associated peptides are mainly secreted by goblet cells, are supposed to be involved in the mucosal maintenance and repair, and seem to reduce the recruitment of inflammatory cells [56, 57]. Moreover, butyrate contributes to the colonic defense barrier by increasing the expression of TFFs [58]. In vitro and in vivo studies and in studies showed a possible effects of butyrate on the expression of heat shot proteins (HSPs), which contribute to the gut barrier homeostasis by suppressing the production of inflammatory modulators [59, 60]. A mechanism for the enhancement of HSPs expression has not been already discerned. So far, only some hypotheses have been proposed. Indeed, SCFA are able to induce cellular acidification

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through a non-ionic diffusion that may induce the production of stress kinases and HSPs. Moreover, the butyrate seems to inhibit the histone deacetylase, a crucial transcriptional regulator [59]. Finally, a possible role of butyrate on intestinal permeability has been assessed in different in vitro studies, which demonstrated a butyrate influence on tight junctions expression. Interestingly, these effects seem to be concentration-dependent, with a concentration up to 2 mM that can decrease the

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intestinal permeability and a concentration higher than 8 mM that, on the contrary, is able to induce

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an increased permeability [56, 61, 62].

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ACTINOBACTERIA AND METABOLISM

Human intestine does not possess many of the enzymes involved in the degradation and

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biotransformation of substances introduced with the diet. Indeed, the majority of enzymes engaged

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in energy production is provided by the commensal bacteria, especially located in the colon and belonging to the genus Bacteroides, Bifidobacterium, Ruminococcus and Roseburia [63]. In

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particular, the pathways involved include the fermentation of large polysaccharides,

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oligosaccarides, unabsorbed sugars and fibers, that release hydrogen, carbon dioxide and SCFAs, the degradation of proteins, the regulation of lipid metabolism through lipoprotein lipase (LPL), and the absorption and biosynthesis of vitamin K, B12, iron, calcium and magnesium [13-15, 64-67]. Bifidobacteria are able to produce large quantities of acetate, but do not produce propionate and butyrate, which are mainly produced by Bacteroides phylium and Clostridium cluster XIVa and IV [68, 69]. However, the acetate released by B. longum NCC2705 represents a co-substrate for butyrate production [70] that constitutes the main energy source for colonocytes [71]. Actinobacteria are also involved in the biodegradation of resistant starch [72 - 74]. Actinobacteria, in particular Bifidobacteria, through the glycosyl hydrolases (GHs) hydrolyze the glycosidic bond between two or more sugars and cooperate to the breakdown of plant-derived carbohydrate starch and polysaccharides, such as FOS, GOS, XOS, inulin or arabinoxilan [75 - 77]. Moreover, Bifidobacteria are supposed to be involved in the transformation of linoleic acid (LA) into

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conjugated linoleic acids (CLA), a group of isomers of LA [78], that have potential healthpromoting properties like anticarcinogenesis, anti-atherosclerosis, anti-diabetes, anti obesity and enhancement of immune functions [79]. Although it has been largely demonstrated that the diet strongly influences gut microbiota composition, the relationship between diet and Actinobacteria is still unclear. In fact, energy

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restrictions, high fiber diet and dietary components such as fructo-oligosaccharides (FOS), galacto-

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oligosaccharides (GOS), xylo-oligosaccharides (XOS) are associated to higher microbial diversity [80]. While some studies showed that Actinobacteria abundance is positively associated with a

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high-fat diet and negatively associated with fiber intake [81, 82], an opposite association is suggested by other studies in which a high concentration of Bifidobacterium spp is positively

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correlated with lean individuals, high consumption of complex carbohydrates, improvement of

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glucose homeostasis, reduction of obesity and inflammation [83 – 86]. A study comparing fecal samples of lean and obese women showed different concentration of Bifidobacteria and Clostridium

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coccoides between the two groups. In particular, there was a negative correlation between

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Bifidobacteria and body fat percentage with an inverse correlation with insulin levels and the homeostasis model assessment (HOMA) index [83]. However, in this study there was no difference in LPS levels between lean and obese subjects and no correlation with body fat percentage, insulin levels or HOMA-index. Nevertheless, a study by Cani et al [85] demonstrated that mice fed with an high fat diet enriched with prebiotic (fermentable dietary fibre, oligofructose (OFS)) could restore their levels of Bifidobacteria, improve glucose tolerance, insulin secretion, and induce an increase of proglucagon mRNA precursor. Authors highlighted that all these changes were correlated with a reduction of endotoxiemia, suggesting a protective role of Bifidobacteria, while no relationship was evident with other bacterial group [85]. Despite the mechanism of action has not been completely understood, authors suggested that the product of OFS degradation, the SCFA, could ameliorate the gut barrier function both through a direct action on colonic cells and by modulating gut microbiota, especially Bifidobacteria [85]. Furthermore, the use of dextrins derived from maize starch is able to

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stimulate the growth of Actinobacteria and Bacteroidetes in both normal-weight and obese children [87], with possible beneficial effect on the production of SCFA. A modulation on gut microbiota, and therefore on Actinobacteria phylum, has been shown with the diet low in fermentable oligosaccharides, disaccharides and polyols, fructo-oligosaccharides and galacto-oligosaccharides (FODMAPs). FODMAPs are small carbohydrate molecules that are

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slowly absorbed in the form of monosaccharides or not absorbed when disaccharides or

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polysaccharides, lasting for a prolonged period into the intestinal lumen [67]. There, FODMAPs exert an osmotic action collecting water into the lumen and are fermented by commensal bacteria

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releasing SCFA, dioxide, hydrogen, methane and carbon [88, 89]. A study by McIntosh et al compared the impact of high and low FODMAPs diet in IBS patients and demonstrated an

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increased richness of Actinobacteria with a decreased number of Bifidobacteria in the low

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FODMAPs diet group [90]. On the contrary, other studies showed that a low FODMAPs diet could lead to a lower abundance in both Bifidobacteria and Actinobacteria because of their FODMAPs

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need for their metabolism [91, 92]. However, not all the patients treated with low FODMAPs

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experienced an improvement of IBS symptoms [90]. This could probably due to different effects of diverse Bifidobacteria strains. Recent studies demonstrated that the use of Bifidobacterium infantis and Bifidobacterium animalis has beneficial effects on IBS symptoms [93 - 95]. However, all the available studies have not examined the microbiome of patients with IBS following the reintroduction phase, but only after the end of the strict low-FODMAP diet, which is usually only recommended for 2 to 6 weeks.

ACTINOBACTERIA AND IMMUNOLOGICAL FUNCTION The gut microbiota plays a pivotal role in the development of immune system and Bifidobacteria, are also critical in this field. The stimulation of intraepithelial lymphocytes, the production of mucosal immunoglobulins and the promotion of a tolerogenic immune response are the main functions exerted by commensal bacteria [19, 20,22, 23, 96].

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A decreased number of Bifidobacteria is associated to an enhancement of gut permeability [97] that leads to the translocation of LPS into the serum. This triggers the immune system activation and sustains chronic inflammatory conditions, such as insulin resistance, diabetes and liver diseases [98]. The administration of Bifidobacterium pseudocatenulatum CECT 7765 along with high fat diet in mice is able to down-regulate the inflammation by reducing the production of inflammatory

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cytokines and chemokines, especially IL-6 and MCP-1, which are usually increased in obesity and

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metabolic disorders [99]. The reduction of these inflammatory markers is concomitant with the improvement of glucose tolerance and with the increasing of IL-4 and IL-13. These cytokines are

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able to stimulate the M2 phenotypic differentiation of macrophages, a subtype of adipose tissue macrophages that secretes the anti-inflammatory cytokine IL-10, and to promote the control of

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inflammation and normal insulin sensitivity [100]. However, the same study showed that after the

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administration of B. pseudocatenulatum CECT 7765 there were increased level of IL-10 only in standard diet fed mice and not in high fat diet-fed mice, indicating that probably parallel regulatory

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mechanisms are involved. Furthermore, B. pseudocatenulatum CECT 7765 can boost an

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appropriate inflammatory response to a bacterial stimulus (LPS) that is usually impaired in high fat diet fed mice. In fact, the administration of this probiotic in these mice stimulates the production of TNF-α by LPS-stimulated macrophages, boosts the oxidative burst and therefore the phagocytosis function in peritoneal macrophages. Finally, it increases the ability of DCs to present antigens and prime a T-lymphocyte proliferative response [99]. In this direction, the reduced phagocytic capacity and oxidative burst of macrophages and the impaired function of DCs are possible reasons for the increased susceptibility to infections in obese subjects [101, 102]. Moreover, Actinobacteria, and mainly Bifidobacteria species, can modulate immune-inflammatory and autoimmune response by inducing regulatory T-cells [103, 104]. Grounding on the evidence that Bifidobacterium longum subspecies infantis (B.infantis) induces regulatory T cells activity in animal models [103, 105 - 108] and increases the relative proportion of the same cells in peripheral blood of healthy humans [109], Groeger et al studied the effect of

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B.infantis inflammatory mediator production in inflammatory disorders in patients with ulcerative colitis (UC), psoriasis and chronic fatigue syndrome (CFS) [110]. They found that 6-8 weeks of treatment with B.infantis significantly reduced plasma C-reactive protein (CRP) levels in all three group of patients, whereas a statistically significant attenuation of TNF-α was observed for psoriasis and CFS. In the UC group they observed a trend of reduction in TNF-α levels that however did not

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reach the statistical significance, probably because of the shorter treatment period (6 weeks of

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treatment vs 8 weeks of treatment of the other two groups). Instead, plasma levels of IL-6 were only marginally decreased after treatment reinforcing the hypothesis of a specific influence on immune

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response for each gut microbiota component [110]. The use of Bifidobacterium species has also been studied in respiratory pathology, alone or in combination with prebiotic or other probiotics like

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Lactobacilllus. In this field, Bifidobacteria seem to be effective to prevent asthma-like symptoms in

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infant with atopic dermatitis and to treat the symptoms of allergic rhinitis [111 – 115]. This is probably due to an immunomodulatory effect of Bifidobacteria on Th balance, regulating Th-2

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eosinophils and IgE production [113].

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immunoresponse that is usually skewed in atopic patients and is responsible of higher levels of

ACTINOBACTERIA AND GUT-BRAIN AXIS Recently, the interest on the impact of gut microflora on gut-brain axis is increasing. This could involve neuroendocrine, immunological and direct neural mechanisms [116 - 118]. Dysbiosis and the consequent increased intestinal permeability are associated to an up-regulation of systemic inflammation that may also involve the central nervous system [119]. Both the gut microbiota production of neurochemicals (i.e., serotonin, SCFA, dopamine and γ-aminobutyric acid) [116, 120] and neurotoxic metabolites (i.e., ammonia) [119] by strengthens the possibility of an implication of the gut-brain axis in depression, anxiety, IBS, IBD, and neurodevelopmental disorders such as autism [116, 121 - 124]. A direct neural communication between the gut and the brain through the

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stimulation of the enteric nervous system and consequently the vagus nerve has also been supposed [125, 126]. Standing on previous studies that showed a reduction of Bifidobacteria and lactobacilli following a stressful experience and emotional stress both in animals and humans [127 - 131], Desbonnet et al conducted a study to assess the potential antidepressant properties of probiotic Bifidobacteria

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infantis in rats [132]. They demonstrated that a two-week treatment with the probiotic

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Bifidobacteria infantis determines an attenuation of pro-inflammatory immune response and an elevation of tryptophan, a precursor of serotonine that represents a target of antidepressant

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treatments. Authors concluded that these results might support the beneficial effect of this probiotic in the treatment of depression, especially when associated to gastrointestinal disorders. Two years

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later, these findings were supported by the evidence that the chronic treatment with Bifidobacteria

life [133].

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infantis was able to attenuate depression in the rats exposed to maternal separation stress in early

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On the other side, a recent study by Lyte at al [134] showed that mice fed with resistant starch (RS)

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had increased levels of Bifidobacteria, according also to a previous study by Tachon et al [135]. RS-fed mice evidenced a significant increase in anxiety-like behavior that the authors considered to be induced by the same diet with a concomitant abundance of Bifidobacterium [134]. Furthermore, a role of Actinobacteria, mainly Bifidobacteria, in the deterioration of cognitive function has been supposed. In fact, preliminary data on the use of the probiotic VSL#3, a mixture of 8 different strains of bacteria including three Bifidobacteria species, showed an improvement in the age-related decrease of long-term potentiation (LTP) in rats [136]. One of the mechanisms supposed to be involved in the regulation of LTP is the anti-inflammatory effect of VSL#3 on the modulation of the hippocampal molecules expression, such as CD 68 and CD 11b. These are markers of microglial activation, and therefore of inflammation. Moreover the authors reported that VSL#3 is able to influence the expression of several genes in the cortex. In particular, it is able to attenuate the age-related changes in three genes, PLA2G3, Nid2 and Alox15, which are associated

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to inflammation. Changes of genes expression in the brain may be linked to the variation of several molecules induced by VSL#3. In fact, this study showed that this probiotic regulates specific mediators involved in synaptic plasticity in the hippocampus of aged rats, especially brain-derived neurotrophic factor (BDNF), synapsin and syntaxin. The authors suggested that the synaptotrophic effect of VSL#3 is mainly due to the BDNF, whose expression was increased in those rats treated

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with the probiotic. The increase of BDNF levels was associated to higher level of synapsin. Of note,

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previous studies recognized its fundamental role in sustaining LTP through the induction of neurogenesis and synaptogenesis [136].

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Lastly, the existence and the importance of gut-brain axis have also been suggested in alcohol dependence. Some alcohol dependent subjects develop a high intestinal permeability that is

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associated with significantly lower levels of Bifidobacteria [137]. These subjects are characterized

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by higher levels of depression, anxiety and craving and have a consistent higher risk of persistence of psychological symptoms after detoxification when compared to alcohol dependent subject with a

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lower intestinal permeability, that on the contrary recover completely after the abstinence period.

BIFIDOBACTERIA AS PROBIOTIC

Bifidobacteria, together with Lactobacilli, represents the cornerstone of probiotic therapeutic approach, but only few Bifidobacteria strains have been studied alone. Thus, in the latter cases the beneficial effects cannot be exclusively related to this probiotic family. For example, the probiotic formulation VSL#3, a mixture of lyophilized four Lactobacilli (L. casei, L. plantarium, L acidophilus, L. bulgaricus), three Bifidobacteria strains (B. longuum, B. breve, B. infantis) and Streptococcus salivaris subspecies termophilus demonstrated to be effective in several conditions. Its efficacy has been mainly assessed in the prevention of pouchitis relapse in patients with UC [138, 139], with promising results in maintaining remission and treating mild-to-moderate UC both in adult and children [140 - 143], in metabolic disorders, in non-alcoholic steato-hepatitis (NASH) [144, 145] and in the prevention of antibiotic-associated diarrhea [146]. An amelioration of

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the pro-inflammatory status in patients affected by UC has been demonstrated with the use of Bifidobacterium infantis alone [110], as mentioned above, and Bifidobacterium breve in association with GOS [147]. The use of Bifidobacteria, alone or in association with other probiotics is also effective in the alleviation of symptoms in patients with IBS [93 - 95, 148 - 150], constipation [151, 152] and in the reduction of antibiotic-associated diarrhea in infants [153]. Moreover, a recent study

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showed that a supplementation of B.breve in association with breast feeding is able to reduce the

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incidence of necrotizing enterocolitis (NEC) in preterm neonates born between 28 and 34 weeks of gestation. However, further studies are needed for neonates < 28 weeks where the reduction of NEC

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didn’t reach a statistical significance [154].

Furthermore, in vitro and murine models studies highlighted how Bifidobacteria, especially B.

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longum and B. lactis strains, seem to be active against rotavirus infection [155] and could protect

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against colorectal cancer (CRC) [156, 157]. Grounding on a previous study where the use of a combination of RS and B. lactis had significantly increased the acute apoptotic response to a

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genotoxic carcinogen (AARGC) in rat colon [158], Le Leu et al confirmed his protective action

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against CRC in rats model, supporting the importance of this synbiotic in cancer prevention. The mechanism(s) of actions that sustain(s) apoptosis with this symbiotic remain(s) unclear, but authors supposed the presence of immunomodulating properties derived by the interaction between B.lactis and RS resulting in butyrate production [156]. A recent randomized, double-blind, placebocontrolled study on humans affected by colorectal polyps (no CRC) or CRC [157] highlighted how the dietary symbiotic, B. lactis, L. Rhamnosus and oligofructose-enriched inulin, was able to reduce colon cell proliferation and cell DNA-damage. However, the two groups responded differently to the synbiotic, maybe because of an increased resistance to qualitative and quantitative changes of cancer patients intestinal microflora [157]. In addition, the effects of Bifidobacteria are not limited to the gastrointestinal tract. In fact, a possible therapeutic role of such probiotic has also been supposed in the treatment of respiratory

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pathologies [112-115], psoriasis, chronic fatigue syndrome [110], depression [132, 133] and in the deterioration of cognitive functions [136].

CONCLUSIONS This review shows how Actinobacteria phylum, despite it represents a minority group of

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commensal bacteria, plays a pivotal role in the development and maintenance of gut homeostasis

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(Figure 1). Its involvement has been supposed in the modulation of gut permeability, immune

system, metabolism and gut-brain axis. An unbalanced abundance has been evidenced in several

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pathological conditions. For this reason, the interest in the use of Actinobacteria, especially of Bifidobacteria family, as probiotic is constantly increasing. This group may represent a significant

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part of the next-generation of probiotics with a potential efficacy both in gut diseases and extra-

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intestinal disorders (Table 1).

However, further studies are needed to deeply understand the interaction between Actinobacteria

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and the host and consequently their therapeutic implications in the prevention and treatment of

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several systemic disorders with the end goal of promoting gut health.

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REFERENCES

1. Mondot S, de Wouters T, Doré J, Lepage P. The human gut microbiome and its dysfunctions. Dig Dis 2013; 31(3-4):278-85 2. Leser TD, Molbach L: Better living through microbial action: the benefits of the mammalian

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gastrointestinal microbiota on the host. Envirol Microbiol 2009; 11:2194-2206

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3. Neish AS. Microbes in gastrointestinal health and disease. Gastroenterology 2009; 136: 65-80. 4. Lopetuso LR, Ianiro G, Scaldaferri F, Cammarota G, Gasbarrini A. Gut Virome and

us

Inflammatory Bowel Disease. Inflamm Bowel Dis 2016;22:1708-12

5. Donaldson GP, Lee SM, Mazmanian SK. Gut biogeography of the bacterial microbiota. Nat Rev

an

Microbiol 2016; 14(1):20-32

M

6. Guarner F, Malagelada JR. Gut flora in health and disease. Lancet 2003; 361:512-9 7. Arumugam M, Raes J, Pelletier E, Le Paslier D, Yamada T, Mende DR et al. Enterotypes of the

d

human gut microbiome. Nature 2011; 473: 174-180.

Ac ce pt e

8. Segata N, Haake SK, Mannon P, Lemon KP, Waldron L, Gevers D et al. Composition of the adult digestive tract bacterial microbiome based on seven mouth surfaces, tonsils, throat and stool samples. Genome Biol 2012; 13:R42. 9. D'Aversa F, Tortora A, Ianiro G, Ponziani FR, Annicchiarico BE, Gasbarrini A.Gut microbiota and metabolic syndrome. Intern Emerg Med 2013; 8 Suppl1:S11-5 10. Gallo A, Passaro G, Gasbarrini A, Landolfi R, Montalto M. Modulation of microbiota as treatment for intestinal inflammatory disorders: An uptodate. World J Gastroenterol 2016; 22:7186-202 11. Round JL, O'Connell RM, Mazmanian SK. Coordination of tolerogenic immuneresponses by the commensal microbiota. J Autoimmun 2010; 34:J220-5 12. Sekirov I, Russel SL, Antunes LC, Finlay BB. Gut microbiota in health and disease. Physiol Rev 2010;90:859-904.

Page 16 of 36

13. Backhed G, Crawford PA. Coordinated regulation of the metabolome and lipidome at the hostmicrobial interface. Biochem Biophys Acta 2010; 1801:240-245 14. Conly JM, Stein K, Worobetz L, Rutledge-Harding S. The contribution of vitaminK2 (menaquinones) produced by the intestinal microflora to human nutritional requirements for vitamin K. Am J Gastroenterol 1994; 89:915-23

ip t

15. Younes H, Coudray C, Bellanger J, Demigné C, Rayssiguier Y, Rémésy C. Effects of two

cr

fermentable carbohydrates (inulin and resistant starch) and their combination on calcium and magnesium balance in rats. Br J Nutr 2001; 86:479-85

us

16. Scaldaferri F, Pizzoferrato M, Gerardi V, Lopetuso L, Gasbarrini A. The gut barrier: new acquisition and therapeutic approaches. J Clin Gastroenterol 2012; 46(Suppl):S12-S17.

an

17. Purchiaroni F, Tortora A, Gabrielli M, Bertucci F, Gigante G, Ianiro G, OjettiV, Scarpellini E,

Pharmacol Sci 2013; 17:323-33

M

Gasbarrini A. The role of intestinal microbiota and the immune system. Eur Rev Med

Ac ce pt e

2008;635:93-101.

d

18. Allen CA, Torres AG. Host-microbe communication within the GI tract. Adv Expt Med Biol

19. Maynard CL, Elson CO, Hatton RD, Weaver CT. Reciprocal interactions of theintestinal microbiota and immune system. Nature 2012; 489:231-41 20. Atarashi K, Tanoue T, Oshima K, Suda W, Nagano Y, Nishikawa H et al. Treg induction bya rationally selected mixture of Clostridia strains from the human microbiota.Nature 2013; 500:232-6

21. Galdean CM, Perdigon G. The probiotic bacterium Lactobacillus casei induces activation of the gut mucosal immune system through innate immunity. Colin Vaccine Immunol 2006;13:219226. 22. Sun M, He C, Cong Y, Liu Z. Regulatory immune cells in regulation of intestinal inflammatory response to microbiota. Mucosal Immunol 2015; 8:969-78

Page 17 of 36

23. McCarville JL, Caminero A, Verdu EF. Novel perspectives on therapeutic modulation of the gut microbiota. Therap Adv Gastroenterol 2016; 9:580-93 24. Sartor RB. Microbial influences in inflammatory bowel disease. Gastroenterology 2008; 134:577-594. 25. De Palma G, Blennerhassett P, Lu J, Deng Y, Park AJ, Green W, et al. Microbiota and host

ip t

determinants of behavioural phenotype in maternally separated mice. Nat Commun 2015;

cr

6:7735

26. Hill DA, Artis D. Intestinal bacteria and the regulation of immune cell homeostasis. Annu Rev

us

Immunol 2010; 28:623-667.

27. Cox A, West N and Cripps A. Obesity, inflammation, and the gut microbiota. Lancet Diabetes

an

Endocrinol 2015; 3:207-215

M

28. Koeth RA, Wang Z, Levison BS, Buffa JA, Org E, Sheehy BT, et al. Intestinal microbiotametabolism of L-carnitine, a nutrient in red meat, promotes atherosclerosis. Nat Med

d

2013; 19:576-85

Ac ce pt e

29. Sun J, Furio L, Mecheri R, van der Does AM, Lundeberg E, Saveanu L, et al. Pancreatic β-Cells Limit Autoimmune Diabetes via an Immunoregulatory Antimicrobial Peptide Expressed under the Influence ofthe Gut Microbiota. Immunity 2015; 43:304-17 30. Scher JU, Ubeda C, Artacho A, Attur M, Isaac S, Reddy SM, et al. Decreasedbacterial diversity characterizes the altered gut microbiota in patients withpsoriatic arthritis, resembling dysbiosis in inflammatory bowel disease. Arthritis Rheumatol 2015; 67:128-39. 31. Belizario JE, Napolitano M. Human microbes and their roles in dysbiosis, common disease, and novel therapeutic approaches. Front Microbiol 2015; 6: 1050. 32. Adlerberth I, Carlsson B, de Man P, Jalil F, Khan SR, Larsson P, Mellander L, Svanborg C, Wold AE, Hanson LA. Intestinal colonization with Enterobacteriaceae in Pakistani and Swedish hospital-delivered infants. Acta Paediatr Scand. 1991;80(6–7):602–10.

Page 18 of 36

33. Hesla HM, Stenius F, Jaderlund L, Nelson R, Engstrand L, Alm J, Dicksved J. Impact of lifestyle on the gut microbiota of healthy infants and their mothers - the ALADDIN birth cohort. FEMS Microbiol Ecol. 2014;90(3):791–801. 34. Mitsou EK, Kirtzalidou E, Oikonomou I, Liosis G, Kyriacou A. Fecal microflora of Greek healthy neonates. Anaerobe. 2008;14(2):94–101.

ip t

35. Dogra S, Sakwinska O, Soh SE, Ngom-Bru C, Bruck WM, Berger B, Brussow H, Lee YS, Yap

cr

F, Chong YS et al.: Dynamics of infant gut microbiota are influenced by delivery mode and gestational duration and are associated with subsequent adiposity. MBio 2015, 6(1).doi:

us

10.1128/mBio.02419-14.

36. Huurre A, Kalliomaki M, Rautava S, Rinne M, Salminen S, Isolauri E. Mode of delivery -

an

Effects on gut microbiota and humoral immunity. Neonatology 2008;93(4):236–40.

M

37. Kabeerdoss J, Ferdous S, Balamurugan R, Mechenro J, Vidya R, Santhanam S, Jana AK, Ramakrishna BS. Development of the gut microbiota in southern Indian infants from birth to 6

d

months: a molecular analysis. J Nutr Sci 2013;2:e18.

Ac ce pt e

38. Gronlund MM, Lehtonen OP, Eerola E, Kero P. Fecal microflora in healthy infants born by different methods of delivery: permanent changes in intestinal flora after cesarean delivery. J Pediatr Gastroenterol Nutr. 1999; 28(1):19–25. 39. Bezirtzoglou E. The intestinal microflora during the first weeks of life. Anaerobe. 1997;3(2– 3):173–7.

40. Yap GC, Chee KK, Hong PY, Lay C, Satria CD, Sumadiono AA, Soenarto Y, Haksari EL, Aw M, Shek LPC, et al. Evaluation of stool microbiota signatures in two cohorts of Asian (Singapore and Indonesia) newborns at risk of atopy. Bmc Microbiol. 2011;11:193. 41. Dethlefsen L, Huse S, Sogin ML, Relman DA. The pervasive effects of an antibiotic on the human gut microbiota, as revealed by deep 16S rRNA sequencing. PLoS Biol. 2008;6(11):e280.

Page 19 of 36

42. Penders J, Thijs C, Vink C, Stelma FF, Snijders B, Kummeling I, van den Brandt PA, Stobberingh EE. Factors influencing the composition of the intestinal microbiota in early infancy. Pediatrics. 2006;118(2):511–21. 43. Bezirtzoglou E, Tsiotsias A, Welling GW. Microbiota profile in feces of breast- and formula-fed newborns by using fluorescence in situ hybridization (FISH). Anaerobe. 2011;17(6):478–82.

ip t

44. Underwood MA, German JB, Lebrilla CB, Mills DA. Bifidobacterium longum subspecies

cr

infantis: champion colonizer of the infant gut. Pediatr Res. 2015; 77(1–2):229–35.

diversity in the human intestine. Cell 2006; 124:837-848.

us

45. Ley RE, Peterson DA, Gordon JI. Ecological and evolutionary forces shaping microbial

46. D’Argenio V, Salvatore F. The role of the gut microbiome in the healthy adult status. Clin Chim

an

Acta 2015; 451(PtA):97-102.

M

47. Eckburg PB, Bik EM, Bernstein CN, Purdom E, Dethlefsen L, Sargent M, Gill SR, Nelson KE, Relman DA. Diversity of the human intestinal microbial flora. Science. 2005; 308:1635-16388.

d

48. Drago L, Toscano M, Rodighiero V, De Vecchi E, Mogna G. Cultivable and pyrosequenced

84.

Ac ce pt e

fecal microflora in centenarians and young subjects. J Clin Gastroenterol 2012;46(suppl):S81-

49. Ashida H, Ogawa M, Kim M, Mimuro H, Sasakawa C. Bacterial and host interactions in the gut epithelial barrier. Nature 2012; 489:231-241. 50. Hardy H, Harris J, Lyon E, Beal J, Foey AD. Probiotics, prebiotics and immunomodulation of gut mucosal defences: homeostasis and immunopathology.Nutrients 2013; 5:1869-912. 51. Fukuda S, Toh H, Taylor TD, Ohno H, Hattori M. Acetate-producing bifidobacteria protect the host from enteropathogenic infection via carbohydrate transporters. Gut Microbes. 2012; 3:44954. 52. Scott KP, Martin JC, Duncan SH, Flint HJ. Prebiotic stimulation of human colonic butyrateproducing bacteria and bifidobacteria, in vitro. FEMS Microbial Ecol 2014;87:30-40

Page 20 of 36

53. Willemsem LE, Koetsier MA, van Deventer SJ et al. Short chain fatty acids stimulate epithelial mucin 2 expression through differential effects on prostaglandin E(1) and E(2) production by intestinal myofibroblasts. Gut 2003;52:1442-47 54. Gaudier E, Jarry A, Blottiere HM et al. Butyrate specifically modulates MUC gene expression in intestinal epithelial goblet cells deprived of glucose. Am J Physiol Gastrointest Liver Physiol

ip t

2004;287:G1168-74

cr

55. Gendler SJ,Spicer AP. Epithelial mucin genes. Annu Rev Physiol 1995; 57:607-34

56. Hamer HM, Jonkers D, Venema K, Vanhoutviny S, Troost FJ.Review article: the role of

us

butyrate on colonic function. Aliment Pharmacy There 2008; 27:104-119.

57. Barrett KE. A new twist on trefoils. Focus on "TFF3 modulates NF-{kappa}B and a novel

an

regulatory molecule of NF-{kappa}B in intestinal epithelial cells via a mechanism distinct from

M

TNF-{alpha}". Am J Physiol Cell Physiol 2005; 289:C1069-71

58. Song M, Xia B, Li J. Effects of topical treatment of sodium butyrate and 5-aminosalicylic acid

d

on expression of trefoil factor 3, interleukin 1beta, and nuclear factor kappaB in trinitrobenzene

Ac ce pt e

sulphonic acid induced colitis in rats. Postgrad Med J 2006; 82:130-5 59. Arvans DL, Vavricka SR, Ren H, et al. Luminal bacterial flora determines physiological expression of intestinal epithelial cytoprotective heat shock proteins 25 and 72. Am J Physiol Gastrointest Liver Physiol 2005; 288: G696–704 60. Ren H, Musch MW, Kojima K, et al. Short-chain fatty acids induce intestinal epithelial heat shock protein 25 expression in rats and IEC 18 cells. Gastroenterology 2001; 121: 631–9 61. Peng L, He Z, Chen W, et al. Effects of butyrate on intestinal barrier function in a caco-2 cell monolayer model of intestinal barrier. Pediatr Res 2007; 61: 37–41 62. Mariadason JM, Kilias D, Catto-Smith A, et al. Effect of butyrate on paracellular permeability in rat distal colonic mucosa ex vivo. J Gastroenterol Hepa- tol 1999; 14: 873–9

Page 21 of 36

63. Davila AM, Blachier F, Gotteland M, Andriamihaja M, Benetti PH, Sanz Y, Tomé D. Intestinal luminal nitrogen metabolism: Role of the gut microbiota and consequences for the host. Pharmacol Res 2012; 68:95-107. 64. Bernalier-Donadille A. Fermentative metabolism by the human gut microbiota. Gastroenterol Clin Biol 2010; 34 Suppl 1:S16-22

ip t

65. CummingsJand MacfarlaneG. The control and consequences of bacterial fermentation in the

cr

human colon. J. Appl Microbiol 1991; 70:443–459

66. Hamer HM, De Preter V, Windey Kand Verbeke K. Functional analysis of colonic bacterial

us

metabolism: relevant to health? Am J Physiol Gastrointest Liver Physiol 2012; 302, G1–9. 67. Ponziani FR, Binda C, Gasbarrini A. How to modulate gut microbiota:diet, pre-probiotic or

an

antibiotics? Minerva Gastroenterol Dietol 2017; 8:1026-1025.

M

68. Bindels LB, Delzenne NM, Cani PD, Walter J. Towards a more comprehensive concept for prebiotics. Nat Rev Gastroenterol Hepatol 2015; 12:303-310.

d

69. O’Callaghan A, van Sinderen D. Bifidobacteria and their role as members of the human gut

Ac ce pt e

microbiota. Front Microbiol 2016; 7:925.

70. Rivière A, Gagnon M, Weckx S, Roy D, De Vuyst L. Mutual cross-feeding interactions between Bifidobacterium longum NCC2705 and eubacterium rectale ATCC336656 explain the bifidogenic and butyrogenic effects of arabinoxylan-oligosaccharides. Appl Environ Microbiol 2015; 80:204-217.

71. Clausen MR, Mortensen PB: Kinetic studies on colonocyte metabolism of short chain fatty acids and glucose in ulcerative colitis. Gut 1995; 37:684-689. 72. Macfarlane GT, Englyst HN. Starch utilization by the human large intestinal microflora. J Appl Bacteriol 1986;60:195–201. 73. Ryan SM, Fitzgerald GF, van Sinderen D. Screening for and identification of starch-, amylopectin-, and pullulan-degrading activities in bifidobacterial strains. Appl Environ Microbiol. 2006;72(8):5289–96.

Page 22 of 36

74. Salyers AA, West SE, Vercellotti JR, Wilkins TD. Fermentation of mucins and plant polysaccharides by anaerobic bacteria from the human colon. Appl Environ Microbiol. 1977;34(5):529–33. 75. Pokusaeva K, Fitzgerald GF and Van Sinderen D. Carbohydrate metabolism in Bifidobacteria. Genes Nutr 2011; 6:285-306.

ip t

76. El Kaoutari A, Armougom F, Gordon JI, Raoult D, Henrissat B. The abundance and variety of

cr

carbohydrate-active enzymes in the human gut microbiota. Nat Rev Microbol 2013; 11:497-504. 77. Milani C, Lugli AG, Duranti S, Turroni F, Mancabelli L, Ferrario C et al. Bifidobacteria exhibit

us

social behaviour through carbohydrate resource sharing in the gut. Sci Rep 2015; 5:15782. 78. Raimondi S, Amaretti A, Leonardi L, Quartieri A, Gozzoli C, Rossi M. Conjugated linoileic

an

acid production by Bifidobacteria: screening, kinetic, and composition. Biomed Res Int 2016;

M

2016:8654317.

79. Kim JH, Kim Y, Kim YJ, Park Y. Conjugated linoleic acids: potential health benefits as a

d

functional food ingredient. Annu Rev Food and Sci Techol 2016; 7:221-244.

Ac ce pt e

80. Cotillard A, Kennedy SP, Kong LC, Prifti E, Pons N, Le Chatelier E et al. Dietary intervention impact on gut microbial gene richness. Nature 2013; 500:585-8 81. TurnbaughPJ, BäckhedF, FultonLand Gordon JI. Diet-induced obesity is linked to marked but reversible alterations in the mouse distal gut microbiome. Cell Host Microbe 2008; 3:213–223 82. Wu GD, Chen J, Hoffmann C, Bittinger K, Chen YY, Keilbaugh SA, Bewtra M, Knights D, Walters WA, Knight R, et al. Linking long-term dietary patterns with gutmicrobial enterotypes. Science 2011; 334:105–108.

83. Teixeira TFS, Grze_skowiak ŁM, Salminen S, Laitinen K, Bressan J, Gouveia Peluzio Mdo C. Faecal levels of Bifidobacterium and Clostridium coccoides but not plasma lipopolysaccharide are inversely related to insulin andHOMA index in women. Clin Nutr 2013; 32:1017–1022.

Page 23 of 36

84. Zimmer J, Lange B, Frick JS, Sauer H, Zimmermann K, Schwiertz A, Rusch K, et al. A vegan or vegetarian diet substantially alters the human colonic faecal microbiota. Eur J Clin Nutr 2012; 66:53–60. 85. Cani PD, Neyrinck AM, Fava F, Knauf C, Burcelin RG, Tuohy KM, Gibson GR, Delzenne NM. Selective increases of bifidobacteria in gut microflora improve high-fat-diet-induced diabetes in

ip t

mice through a mechanism associated with endotoxaemia. Diabetologia 2007; 50:2374–2383.

cr

86. Geurts L, Neyrinck AM, Delzenne NM, Knauf C, Cani PD. Gut microbiota controls adipose tissue expansion, gut barrier and glucose metabolism: novel insights into molecular targets and

us

interventions using prebiotics. Benef Microbes 2014; 5:3–17.

87. Barczynska R, Kapusniak J, Litwin M, Slizewska K, Szalecky M. Dextrins from maize starch as

an

substances activating the growth of Bacteroidetes and Actinobacteria simultaneosly inhibiting

2016; 71:190-196.

M

the growth of Firmicutes, responsible for the occurrence of obesity. Plant Foods Hum Nutr

d

88. Barrett JS, Gearry RB, Muir JG, Irving PM, Rose R, Rosella O, et al. Dietary poorly absorbed,

Ac ce pt e

short-chain carbohydrates increase delivery of water and fermentable substrates to the proximal colon. Aliment Pharmacol Ther. 2010;31:874-82 89. De Giorgio R, Volta U, Gibson PR. Sensitivity to wheat, gluten and FODMAPs in IBS: facts or fiction? Gut. 2016;65:169-78

90. McIntosh K, Reed DE, Schneider T, Dang F, Keshteli AH, De Palma G, Madsen K, Bercik P, Vanner S. FODMAPs alter symptoms and the metabolome of patients with IBS: a randomised controlled trial. Gut. 2017 Jul;66(7):1241-1251. 91. Staudacher HM, Lomer MC, Anderson JL, et al. Fermentable carbohydrate restriction reduces luminal bifidobacteria and gastrointestinal symptoms in patients with irritable bowel syndrome. J Nutr 2012;142:1510–18.

Page 24 of 36

92. Bennet SMP, Böhn L, Störsrud S, Liljebo T, Collin L, Lindfors P, Törnblom H, Öhman L, Simrén M. Multivariate modelling of faecal bacterial profiles of patients with IBS predicts responsiveness to a diet low in FODMAPs. Gut 2017. [Epub ahead of print] 93. O’Mahony L, McCarthy J, Kelly P, et al. Lactobacillus and Bifidobacterium in irritable bowel syndrome: Symptom responses and relationship to cytokine profiles. Gastroenterology

ip t

2005;128:541–51.

cr

94. Whorwell PJ, Altringer L, Morel J, et al. Efficacy of an encapsulated probiotic Bifidobacterium infantis 35624 in women with irritable bowel syndrome. Am J Gastroenterol 2006;101:1581–90.

us

95. Guyonnet D, Chassany O, Ducrotte P, et al. Effect of a fermented milk containing

Bifidobacterium animalis DN-173 010 on the health-related quality of life and symptoms in

an

irritable bowel syndrome in adults in primary care: a multicentre, randomized, double-blind,

M

controlled trial. Aliment Pharmacol Ther 2007;26:475–86.

96. Sivan A, Corrales L, Hubert N, Williams JB, Aquino-Michaels K, Earley ZM, et al. Commensal

Ac ce pt e

2015;350:1084-9.

d

Bifidobacterium promotes antitumor immunity and facilitates anti-PD-L1 efficacy. Science.

97. Duca FA, Sakar Y, Covasa M. The modulatory role of high fat feeding on gastrointestinal signals in obesity. J Nutr Biochem 2013; 24:1663-77. 98. Scarpellini E, Tack J. Obesity and metabolic syndrome: an inflammatory condition. Dig Dis 2012; 30:148-153.

99. Gauffin Cano P, Santacruz A, Trejo MF, Sanz Y. Bifidobacterium CECT 7765 improves metabolic and immunological alterations associated with obesity in high-fat diet-fed mice. Obesity 2013; 21:2310-21. 100.

Cani PD, Possemiers S, Van de Wiele T et al. Changes in gut microbiota control

inflammation in obese mice through a mechanism involving GLP-2 driven improvement of gut permeability. Gut 2009; 58:1091-1103.

Page 25 of 36

101.

Zhou Q, Leeman SE, Amar S. Signaling mechanism involved in altered function of

macrophages from diet-induced obese mice affect immune responces. Proc Natl Acad Sci USA 2009; 106:10740-10745. 102.

Macia L, Declare M, Abboud G et al. Impairment of dendritic cell functionally and steady-

state number in obese mice. J Immunol 2006; 177: 5997-6006. O’Mahony C, Scully P, O’Mahony D, Murphy S, O’Brien F, Lyons A et al. Commensal-

ip t

103.

cr

induced regulatory T cells mediate protection against pathogen-stimulated NF-kappaB activation. PLoS Pathog 2008; 4:e1000112.

Lyons A, O’Mahony D, O’Brien F, MacSharry J, Sheil B, Ceddia M et al. Bacterial strain-

us

104.

specific induction of Foxp3+ T regulatory cells is protective in murine allergy models. Clin Exp

O'Mahony L, Feeney M, O'Halloran S, Murphy L, Kiely B, Fitzgibbon J, Lee G, O'Sullivan

M

105.

an

Allergy 2010; 40:811-19.

G, Shanahan F, Collins JK. Probiotic impact on microbial flora, inflammation and tumour

Konieczna P, Akdis CA, Quigley EM, Shanahan F, O'Mahony L. Portrait of an

Ac ce pt e

106.

d

development in IL-10 knockout mice. Aliment Pharmacol Ther 2001; 15:1219-25.

immunoregulatory Bifidobacterium. Gut Microbes 2012; 3:261-6. 107.

McCarthy J, O'Mahony L, O'Callaghan L, Sheil B, Vaughan EE, Fitzsimons N, Fitzgibbon

J, O'Sullivan GC, Kiely B, Collins JK, Shanahan F. Double blind, placebo controlled trial of two probiotic strains in interleukin 10 knockout mice and mechanistic link with cytokine balance. Gut 2003; 52:975-80. 108.

McKernan DP, Fitzgerald P, Dinan TG, Cryan JF. The probiotic Bifidobacterium infantis

35624 displays visceral antinociceptive effects in the rat. Neurogastroenterol Motil 2010; 22:1029-35, e268. 109.

Konieczna P, Groeger D, Ziegler M, Frei R, Ferstl R, Shanahan F, Quigley EM, Kiely B,

Akdis CA, O'Mahony L. Bifidobacterium infantis 35624 administration induces Foxp3 T

Page 26 of 36

regulatory cells in human peripheral blood: potential role for myeloid and plasmacytoid dendritic cells. Gut 2012; 61:354-66. 110.

Groeger D, O'Mahony L, Murphy EF, Bourke JF, Dinan TG, Kiely B, Shanahan F, Quigley

EM. Bifidobacterium infantis 35624 modulates host inflammatory processes beyond the gut. Gut Microbes 2013; 4:325-39. Tojo R, Suárez A, Clemente MG, de los Reyes-Gavilán CG, Margolles A, Gueimonde M,

ip t

111.

cr

Ruas-Madiedo P. Intestinal microbiota in health and disease: role of bifidobacteria in gut homeostasis. World J Gastroenterol 2014; 20:15163-76.

van der Aa LB, van Aalderen WM, Heymans HS, Henk Sillevis Smitt J, Nauta AJ, Knippels

us

112.

LM, Ben Amor K, Sprikkelman AB; Synbad Study Group. Synbiotics prevent asthma-like

Xiao JZ, Kondo S, Yanagisawa N, Takahashi N, Odamaki T, Iwabuchi N, Miyaji K,

M

113.

an

symptoms in infants with atopic dermatitis. Allergy 2011;66:170-7.

Iwatsuki K, Togashi H, Enomoto K, Enomoto T. Probiotics in the treatment of Japanese cedar

Singh A, Hacini-Rachinel F, Gosoniu ML, Bourdeau T, Holvoet S, Doucet-Ladeveze R,

Ac ce pt e

114.

d

pollinosis: a double-blind placebo-controlled trial. Clin Exp Allergy 2006 Nov;36(11):1425-35.

Beaumont M, Mercenier A, Nutten S. Immune-modulatory effect of probiotic Bifidobacterium lactis NCC2818 in individuals suffering from seasonal allergic rhinitis to grass pollen: an exploratory, randomized, placebo-controlled clinical trial. Eur J Clin Nutr 2013; 67:161-7. 115.

Odamaki T, Xiao JZ, Iwabuchi N, Sakamoto M, Takahashi N, Kondo S, Miyaji K, Iwatsuki

K, Togashi H, Enomoto T, Benno Y. Influence of Bifidobacterium longum BB536 intake on faecal microbiota in individuals with Japanese cedar pollinosis during the pollen season. J Med Microbiol 2007; 56:1301-8. 116.

Mayer EA, Tillisch K, Gupta A. Gut/brain axis and the microbiota. J Clin Invest 2015; 125:

926-938. 117.

Grenham S, Clarke G, Cryan JF, Dinan TG. Brain-gut-microbe communication in health

and disease. Front Physiol 2011; 2: 94.

Page 27 of 36

118.

Rhee SH, Pothoulakis C, Mayer EA. Principles and clinical implications of the brain-gut-

enteric microbiota axis. Nat Rev Gastroenterol Hepatol 2009; 6: 306-314. 119.

Galland L. The gut microbiome and the brain. J Med Food 2014; 17: 1261-1272.

120.

Lyte M. Microbial endocrinology: Host-microbiota neuroendocrine interactions influencing

brain and behavior. Gut Microbes 2014; 5: 381-389. Borre YE, O’Keeffe GW, Clarke G, Stanton C, Dinan TG, Cryan JF. Microbiota and

ip t

121.

cr

neurodevelopmental windows: implications for brain disorders. Trends Mol Med 2014; 20: 509518.

Dinan TG, Cryan JF. Melancholic microbes: a link between gut microbiota and depression?

us

122.

Neurogastroenterol Motil 2013; 25: 713-719.

Bonaz BL, Bernstein CN. Brain-gut interactions in inflammatory bowel disease.

an

123.

124.

M

Gastroenterology 2013; 144: 36-49.

Hsiao EY, McBride SW, Hsien S, Sharon G, Hyde ER, McCue T, Codelli JA, Chow J,

d

Reisman SE, Petrosino JF, Patterson PH, Mazmanian SK. Microbiota modulate behavioral and

Ac ce pt e

physiological abnormalities associated with neurodevelopmental disorders.Cell 2013; 155: 1451-1463. 125.

Forsythe P, Bienenstock J, Kunze WA. Vagal pathways for microbiome-brain-gut axis

communication. Adv Exp Med Biol 2014; 817: 115-133. 126.

Mulak A, Bonaz Bruno. Brain-gut-microbiota axis in Parkinson’s disease. World J

Gastroenterol 2015; 21:10609-10620. 127.

Bailey MT, Coe CL. Maternal separation disrupts the integrity of the intestinal microflora in

infant rhesus monkeys. Dev Psychobiol 1999;35:146-55. 128.

Bailey MT, Lubach GR, Coe CL. Prenatal stress alters bacterial colonization of the gut in

infant monkeys. J Pediatr Gastroenterol Nutr 2004; 38:414–421. 129.

Tannock GW, Savage DC. Influences of dietary and environmental stress on microbial

populations in the murine gastrointestinal tract. Infect Immun 1974; 9:591-8.

Page 28 of 36

130.

Suzuki K, Harasawa R, Yoshitake Y, Mitsuoka T. Effects of crowding and heat stress on

intestinal flora, body weight gain, and feed efficiency of growing rats and chicks. Nippon Juigaku Zasshi 1983; 45:331–338. 131.

Goncharova GI, Liz’ko NN, Liannaia AM, Shilov VM, Spitsa TI. Bifidobacterium flora

status of cosmonauts before and after com- pleting space flights. Kosm Biol Aviakosm Med

Desbonnet L, Garrett L, Clarke G, Bienenstock J, Dinan TG. The probiotic Bifidobacteria

cr

132.

ip t

1981; 15:14–18.

infantis: an assessment of potential antidepressant properties in the rat. J Psych Res 2009;

133.

us

43:164-174.

Desbonnet L, Garrett L, Clarke G, Kiely B, Cryan JF, Dinan TG. Effects of the probiotic

an

bifidobacterium infantis in the maternal separation model of depression. Neuroscience 2010;

134.

M

170:1179-1188.

Lyte M, Chapel A, Lyte JM, Ai Y, Proctor A, Jane JL, Phillips GJ. Resistant starch alters

135.

Ac ce pt e

11:e0146406.

d

the microbiota-gut-brain axis: implications for dietary modulation of behavior. PloS ONE 2016;

Tachon S, Zhou J, Keenan M, Martin R, Marco ML. The intestinal microbiota in aged mice

is modulated by dietary resistant starch and correlated with improvements in host responses. FEMS Microbiol Ecol. 2013; 83:299–309. 136.

Distrutti E, O’Reilly J-A, McDonald C, Cipriani S, Renga B, et al. Modulation of Intestinal

Microbiota by the Probiotic VSL#3 Resets Brain Gene Expression and Ameliorates the AgeRelated Deficit in LTP. PLoS ONE 2014; 9: e106503. 137.

Leclercq S, Matamoros S, Cani PD, Neyrinck AM, Jamar F, Stärkel P, Windey K,Tremaroli

V, Bäckhed F, Verbeke K, de Timary P, Delzenne NM. Intestinalpermeability, gut-bacterial dysbiosis, and behavioral markers of alcohol-dependence severity. Proc Natl Acad Sci U S A 2014;111:e4485-93.

Page 29 of 36

138.

Gionchetti P, Rizzello F, Helwig U, Venturi A, Lammers KM, Brigidi P, et al. Prophylaxis

of pouchitis onset withprobiotic therapy: a double-blind, placebo-controlled trial. Gastroenterology 2003; 124:1202-9. 139.

Mimura T, Rizzello F, Helwig U, Poggioli G, Schreiber S, Talbot IC, et al. Once daily high

dose probiotic therapy(VSL#3) for maintaining remission in recurrent or refractory pouchitis.

Sood A, Midha V, Makharia GK, Ahuja V, Singal D, Goswami P, Tandon RK. The

cr

140.

ip t

Gut 2004; 53:108-14.

probiotic preparation, VSL#3 induces remission in patients with mild-to-moderately active

141.

us

ulcerative colitis. Clin Gastroenterol Hepatol 2009; 7:1202-9.

Mardini HE, Grigorian AY. Probiotic mix VSL#3 is effective adjunctive therapy for mild to

Tursi A, Brandimarte G, Papa A, Giglio A, Elisei W, Giorgetti GM, Forti G, Morini S,

M

142.

an

moderately active ulcerative colitis: a meta-analysis. Inflamm Bowel Dis 2014; 20:1562-7.

Hassan C, Pistoia MA, Modeo ME, Rodino' S, D'Amico T, Sebkova L, Sacca’ N, Di Giulio E,

d

Luzza F, Imeneo M, Larussa T, Di Rosa S, Annese V, Danese S, Gasbarrini A. Treatment of

Ac ce pt e

relapsing mild-to-moderate ulcerative colitis with the probiotic VSL#3 as adjunctive to a standard pharmaceutical treatment: a double-blind, randomized, placebo-controlled study. Am J Gastroenterol 2010; 105:2218-27. 143.

Miele E, Pascarella F, Giannetti E, Quaglietta L, Baldassano RN, Staiano A. Effect of a

probiotic preparation (VSL#3) on induction and maintenance of remission in children with ulcerative colitis. Am J Gastroenterol 2009;104: 437-43. 144.

Abu-Shanab A, Quigley EM. The role of the gut microbiota in nonalcoholic fatty liver

disease. Nat Rev Gastroenterol Hepatol 2010; 7:691-701. 145.

Machado MV, Cortez-Pinto H. Gut microbiota and nonalcoholic fatty liverdisease. Ann

Hepatol 2012; 11:440-9.

Page 30 of 36

146.

Selinger CP, Bell A, Cairns A, Lockett M, Sebastian S, Haslam N. Probiotic VSL#3

prevents antibiotic-associated diarrhoea in a double-blind, randomized, placebo-controlled clinical trial. J Hosp Infect 2013; 84:159-65. 147.

Ishikawa H, Matsumoto S, Ohashi Y, Imaoka A, Setoyama H, Umesaki Y, Tanaka R, Otani

T. Beneficial effects of probiotic bifidobacterium and galacto-oligosaccharide in patients with

Kajander K, Hatakka K, Poussa T, Färkkilä M, Korpela R. A probiotic mixture alleviates

cr

148.

ip t

ulcerative colitis: a randomized controlled study. Digestion. 2011;84:128-33.

symptoms in irritable bowel syndrome patients: a controlled 6-month intervention. Aliment

149.

us

Pharmacol Ther 2005; 22:387-94.

Kajander K, Myllyluoma E, Rajilić-Stojanović M, Kyrönpalo S, Rasmussen M, Järvenpää

an

S, Zoetendal EG, de Vos WM, Vapaatalo H, Korpela R. Clinical trial: multispecies probiotic

M

supplementation alleviates the symptoms of irritable bowel syndrome and stabilizes intestinal microbiota. Aliment Pharmacol Ther 2008; 27:48-57.

Guandalini S, Magazzù G, Chiaro A, La Balestra V, Di Nardo G, Gopalan S, Sibal A,

d

150.

Ac ce pt e

Romano C, Canani RB, Lionetti P, Setty M. VSL#3 improves symptoms in children with irritable bowel syndrome: a multicenter, randomized, placebo-controlled, double-blind, crossover study. J Pediatr Gastroenterol Nutr 2010; 51:24-30. 151.

Kleessen B, Sykura B, Zunft HJ, Blaut M. Effects of inulin and lactose on fecal microflora,

microbial activity, and bowel habit in elderly constipated persons. Am J Clin Nutr 1997; 65:1397-402. 152.

Leahy SC, Higgins DG, Fitzgerald GF, van Sinderen D. Getting better with bifidobacteria. J

Appl Microbiol 2005; 98:1303-15. 153.

Corrêa NB, Péret Filho LA, Penna FJ, Lima FM, Nicoli JR. A randomized formula

controlled trial of Bifidobacterium lactis and Streptococcus thermophilus for prevention of antibiotic-associated diarrhea in infants. J Clin Gastroenterol 2005; 39:385-9.

Page 31 of 36

154.

Patole SK, Rao SC, Keil AD, Nathan EA, Doherty DA, Simmer KN. Benefits of

Bifidobacterium breve M-16V Supplementation in Preterm Neonates - A Retrospective Cohort Study. PLoS One. 2016;11:e0150775. 155.

Chenoll E, Rivero M, Codoñer FM, Martinez-Blanch JF, Ramón D, Genovés S, Moreno

Muñoz JA. Complete Genome Sequence of Bifidobacterium longum subsp. infantis Strain

ip t

CECT 7210, a Probiotic Strain Active against Rotavirus Infections. Genome Announc 2015

156.

cr

;3(2). pii: e00105-15.

Le Leu RK, Hu Y, Brown IL, Woodman RJ, Young GP. Synbiotic intervention of

us

Bifidobacterium lactis and resistant starch protects against colorectal cancer development in rats. Carcinogenesis. 2010; 31:246-51.

Rafter J, Bennett M, Caderni G, Clune Y, Hughes R, Karlsson PC, Klinder A, O'Riordan M,

an

157.

M

O'Sullivan GC, Pool-Zobel B, Rechkemmer G, Roller M, Rowland I, Salvadori M, Thijs H, Van Loo J, Watzl B, Collins JK. Dietary synbiotics reduce cancer risk factors in polypectomized and

Le Leu,R.K. et al. (2005) A synbiotic combination of resistant starch and Bifidobacterium

Ac ce pt e

158.

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colon cancer patients. Am J Clin Nutr 2007;85:488-96.

lactis facilitates apoptotic deletion of carcinogen-damaged cells in rat colon. J. Nutr., 135, 996– 1001

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FIGURE LEGEND Figure 1 Actinobacteria phylum groups four main families: Bifidobacteria, Propionibacteria, Corynebacteria, Stremptomyces. Actinobacteria exert crucial physiological functions in the gut. FOS: fructooligosaccharides, GOS: galacto-oligosaccharides, XOS: xylo-oligosaccharides, BDNF: brain-

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derived neurotrophic factor.

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Table 1 Potential modulatory and therapeutic role of Actinobacteria in intestinal and extra-intestinal diseases.

EXTRA-INTESTINAL DISEASES

UC: prevention of pouchitis relapse, maintenance of remission and treatment of mild-to-moderate UC

NASH

IBS

RESPIRATORY PATHOLOGIES

CONSTIPATION

PSORIASIS

ANTIBIOTIC-ASSOCIATED DIARHEA

CHRONIC FATIGUE SYNDROME

NEC

DEPRESSION

COLORECTAL CANCER

COGNITIVE DETERIORATION

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INTESTINAL DISEASES

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NASH: non-alcoholic steato-hepatitis.

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RCU/ UC: ulcerative colitis, IBS: irritable bowel syndrome, NEC: necrotizing entero-colitis,

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CONFLICT OF INTEREST

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Authors declare no conflict of interest.

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Figure 1

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PHYSIOLOGICAL FUNCTIONS

Metabolism:

-maintenance of gut permeability -down-regulation of inflammation production of IL-4 and IL-13 -induction of regulatory T-cells -regulation of Th-2 immunoresponse

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-production of acetate -production of lactate that is used by “lactate-utilizer” for the production of butyrate -maintenance of gut permeability

Immunological function:

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Gut Barrier:

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-production of acetate -biodegradation of resistant starch with production of FOS, GOS, XOS, inulin, arabinoxilan -formation of conjugated linoleic acids

by

Gut-Brain Axis:

Actinobacteria Bifidobacteria

Propionibacteria

Corynebacteria

Streptomyces

-potential anti-depressant effects with the elevation of tryptophan levels -regulation of mediators involved in synaptic plasticity like BDNF, synapsin, syntaxin -modulation of hippocampal molecules such as CD68 and CD11b

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