Identifying and Characterizing New Ingredients in vitro for Prebiotic and Synbiotic use

The endogenous microbiota, constituting the microbes that live inside and on humans, is estimated to outnumber human cells by a factor of ten. This commensal microbial population has an important role in many physiological functions, with the densest microbiota population found in the colon. The colonic microbiota is a highly complex and diverse bacterial ecosystem, and a delicate balance exists between the gut microbiota and its host. An imbalance in the microbial ecosystem may lead to severe symptoms in and also beyond the gastrointestinal tract. Due to the important role of the gut microbiota in human health, means of its modification have been introduced in the dietary concepts of pro-, pre- and synbiotics. Prebiotics, which are usually carbohydrates, strive to selectively influence beneficial microbes resident in the colon with the aim of modifying the composition and functionality of the commensal microbial population towards a purportedly healthier one. The study of prebiotic effects on colonic micro-organisms is typically done by using human faecal material, though this provides relatively little information on bacterial populations and metabolic events in different parts of the colon. For this reason, several in vitro models have been developed to investigate the gut microbiota. The aim of this doctoral thesis was to screen through some of the promising prebiotic candidates, characterize their effects on the microbiota through the use of two in vitro methods (pure microbial cultures and a colon simulator model) and to evaluate their potential as emerging prebiotics or synbiotics when combined with the probiotic Bifidobacterium lactis . As a result of the screening work and subsequent colon simulation studies, several compounds with promising features were identified. Xylo-oligosaccharides (XOS), which have previously already shown promise as prebiotic compounds, were well fermented by several probiotic Bifidobacterium lactis strains in pure culture studies and in the following simulation studies utilizing the complex microbiota by endogenous B. lactis Another promising compound was panose, a trisaccharide belonging to isomalto-oligosaccharides (IMO) that also was also able to modify the microbiota in vitro by increasing the number of beneficial microbes investigated. Panose has not been widely studied previously and therefore, this thesis work provided the first data on panose fermentation in mixed colonic microbiota. Galacto-oligosaccharide (GOS) is an established prebiotic, and it was studied here in conjunction with another potential polygosaccharide polydextrose (PDX) and probiotic B. lactis Bi-07. In this final study, the synbiotics including GOS were more effective than the constituting pro- or prebiotics alone in modulating the microbiota composition, thus indicating a synergy resulting from the combination. The results obtained in this in vitro work can be, and have already been, utilized in product development aimed at the nutritional modification of the human colonic microbiota. Some of the compounds have entered the human clinical intervention phase to nvestigate in more detail the prebiotic and synbiotic properties seen in these in vitro studies.Siirretty Doriast

Sikiökautinen ympäristö säätelee elintapasairauksien riskiä

•Aikuisiän sairauksien riskin ohjelmoitumisella tarkoitetaan ympäristön vaikutusta kehittyvään yksilöön ­sikiöajan ja varhaislapsuuden herkkyyskausien aikana. •Ohjelmoitumisessa sikiö sopeutuu elinympäristöön muuttamalla solujen ja elinten aineenvaihduntaa. Yksi ohjelmoitumisen mekanismeista on geenien aktiivisuutta muokkaava epigeneettinen säätely. •Varhainen ravitsemusympäristö on keskeinen ohjelmoitumiseen vaikuttava tekijä. Häiriöt, kuten äidin vajaa­ravitsemus tai ylipaino sekä raskausdiabetes, voivat ohjelmoitumisen välityksellä lisätä lapsen elintapa­sairauksien riskiä aikuisiässä. •Ravinnon koostumus vaikuttaa sekä ohjelmoitumiselle altistaviin tekijöihin, kuten ylipainon tai matala-asteisen tulehduksen syntyyn, että epigeneettiseen säätelyyn.</div

The effect of selected synbiotics on microbial composition and short-chain Fatty Acid production in a model system of the human colon.

Prebiotics, probiotics and synbiotics can be used to modulate both the composition and activity of the gut microbiota and thereby potentially affecting host health beneficially. The aim of this study was to investigate the effects of eight synbiotic combinations on the composition and activity of human fecal microbiota using a four-stage semicontinuous model system of the human colon.Carbohydrates were selected by their ability to enhance growth of the probiotic bacteria Lactobacillus acidophilus NCFM (NCFM) and Bifidobacterium animalis subsp. lactis Bl-04 (Bl-04) under laboratory conditions. The most effective carbohydrates for each probiotic were further investigated, using the colonic model, for the ability to support growth of the probiotic bacteria, influence the composition of the microbiota and stimulate formation of short-chain fatty acids (SCFA).The following combinations were studied: NCFM with isomaltulose, cellobiose, raffinose and an oat β-glucan hydrolysate (OBGH) and Bl-04 with melibiose, xylobiose, raffinose and maltotriose. All carbohydrates showed capable of increasing levels of NCFM and Bl-04 during fermentations in the colonic model by 10(3)-10(4) fold and 10-10(2) fold, respectively. Also the synbiotic combinations decreased the modified ratio of Bacteroidetes/Firmicutes (calculated using qPCR results for Bacteroides-Prevotella-Porphyromonas group, Clostridium perfringens cluster I, Clostridium coccoides - Eubacterium rectale group and Clostridial cluster XIV) as well as significantly increasing SCFA levels, especially acetic and butyric acid, by three to eight fold, as compared to the controls. The decreases in the modified ratio of Bacteroidetes/Firmicutes were found to be correlated to increases in acetic and butyric acid (p=0.04 and p=0.03, respectively).The results of this study show that all synbiotic combinations investigated are able to shift the predominant bacteria and the production of SCFA of fecal microbiota in a model system of the human colon, thereby potentially being able to manipulate the microbiota in a way connected to human health

Bifidobacterium lactis 420 and fish oil enhance intestinal epithelial integrity in Caco-2 cells

Increased intestinal permeability is a predisposing factor for low grade inflammation-associated conditions, including obesity and type 2 diabetes. Dietary components may influence intestinal barrier integrity. We hypothesized that the dietary supplements Bifidobacterium lactis 420, Lactobacillus rhamnosus HN001, and fish oil have beneficial impacts on intestinal barrier integrity. In addition, we hypothesized that the co-administration of these components results in synergistic benefits to the integrity of the intestinal barrier. To study this, we investigated the impact of cell free culture supernatant from dietary supplements B. lactis 420 and L.rhamnosus HN001, and fish oil; separately and in combination, on intestinal permeability in a CaCo-2 cell model. Administered separately, both B. lactis 420 supernatant and fish oil significantly increased the integrity of the intestinal epithelial barrier, as determined by an increase in transepithelial electrical resistance (TEER), whereas L. rhamnosus did not. The TEER increase with B. lactis 420 was dose dependent. Interestingly, a combination of B. lactis 420 supernatant and fish oil negated the increase in TEER of the single components. mRNA expression of tight junction proteins, measured by real time qPCR, was not altered, but the mRNA expression of myosin light chain kinase (MLCK) increased after fish oil treatment. To conclude, single dietary components, namely B. lactis 420 and fish oil induced beneficial effects on intestinal barrier integrity in vitro, whilst a combination of two beneficial test compounds, resulted in a null effect.</p

The effect of polydextrose and probiotic lactobacilli in a Clostridium difficile–infected human colonic model

Background: Clostridium difficile is a natural resident of the intestinal microbiota; however, it becomes harmful when the normal intestinal microbiota is disrupted, and overgrowth and toxin production occurs. The toxins can cause bloating and diarrhoea, which may cause severe disease and have the potential to cause outbreaks in hospitals and other healthcare settings. Normally, antibiotic agents are used for treatment, although for some of the patients, these treatments provide only a temporary relief with a recurrence of C. difficile–associated diarrhoea. Objective: The effects of polydextrose (PDX), Lactobacillus acidophilus NCFM, and L. paracasei Lpc-37 on the growth of C. difficile were investigated in an in vitro model of infected human large intestine. Design: The semi-continuous colonic model is composed of four connected vessels inoculated with human faecal microbes and spiked with pathogenic C. difficile (DSM 1296). PDX in two concentrations (2 and 4%), NCFM, and Lpc-37 were fed to the system during the 2-day simulation, and the growth of C. difficile and several other microbial groups were monitored using quantitative polymerase chain reaction (qPCR) and 16S rDNA sequencing. Results: The microbial community structure of the simulation samples was closely grouped according to treatment, and the largest shifts in the microbial composition were seen with PDX. The microbial diversity decreased significantly with 4% PDX, and the OTU containing C. difficile was significantly (p<0.01) decreased when compared to control and lactobacilli treatments. The mean numbers of C. difficile also decreased as detected by qPCR, although the reduction did not reach statistical significance. Conclusions: The treatments influenced the colonic microbiota, and a trend for reduced numbers of C. difficile as well as alterations of several microbial groups could be detected. This suggests that PDX may be able to modulate the composition and/or function of the colonic microbiota in such manner that it affects the pathogenic C. difficile

Gut Microbiota Richness and Composition and Dietary Intake of Overweight Pregnant Women Are Related to Serum Zonulin Concentration, a Marker for Intestinal Permeability

Background: Increased intestinal permeability may precede adverse metabolic conditions. The extent to which the composition of the gut microbiota and diet contribute to intestinal permeability during pregnancy is unknown.Objective: The aim was to investigate whether the gut microbiota and diet differ according to serum zonulin concentration, a marker of intestinal permeability, in overweight pregnant women.Methods: This cross-sectional study included 100 overweight women [mean age: 29 y; median body mass index (in kg/m(2)): 30] in early pregnancy (= 46.4 ng/mL) serum zonulin groups on the basis of the median concentration of zonulin (46.4 ng/mL). The richness of the gut microbiota (Chao 1, observed species and phylogenetic diversity) was higher in the low zonulin group than in the high zonulin group (P = 0.01). The abundances of Bacteroidaceae and Veillonellaceae, Bacteroides and Blautia, and Blautia sp. were lower and of Faecalibacterium and Faecalibacterium prausnitzii higher (P < 0.05) in the low zonulin group than in the high zonulin group. Dietary quantitative intakes of n-3 (omega-3) polyunsaturated fatty acids (PUFAs), fiber, and a range of vitamins and minerals were higher (P < 0.05) in women in the low zonulin group than those in the high zonulin group.Conclusions: The richness and composition of the gut microbiota and the intake of n-3 PUFAs, fiber, and a range of vitamins and minerals in overweight pregnant women are associated with serum zonulin concentration. Modification of the gut microbiota and diet may beneficially affect intestinal permeability, leading to improved metabolic health of both the mother and fetus. This trial was registered at clinicaltrials.gov as NCT01922791

Ratios of <i>Bacteroidetes/Firmicutes</i> as determined by qPCR.

<p>Ratios of <i>Bacteroidetes/Firmicutes</i> (<i>Bacteroides-Prevotella-Porphyromonas</i> group/<i>Clostridium perfringens</i> cluster I, <i>Clostridium coccoides - Eubacterium rectale</i> group and Clostridial cluster XIV) for fermentations with <i>Lactobacillus acidophilus</i> NCFM (A) in combination with isomaltulose (•) (n = 2), cellobiose (▴)(n = 2), raffinose (♦)(n = 2) and OBGH (▪)(n = 2); <i>Bifidobacterium animalis</i> subsp. <i>lactis</i> Bl-04 (B) in combination with melibiose (<b>○</b>)(n = 2), xylobiose (Δ) (n = 2), raffinose (◊)(n = 2) and maltotriose (□)(n = 3). Control fermentations (n = 3) are denoted by crosses and dotted lines and results are shown as mean values for each vessel, V1–V4, ± standard error of mean. *p<0.05.</p