Louise Kristine Vigsnæs

Decreased colonization of fecal Clostridium coccoides/Eubacterium rectale species from ulcerative colitis patients in an in vitro dynamic gut model with mucin environment.

The mucus layer in the colon, acting as a barrier to prevent invasion of pathogens, is thinner and discontinuous in patients with ulcerative colitis (UC). A recent developed in vitro dynamic gut model, the M-SHIME, was used to compare long-term colonization of the mucin layer by the microbiota from six healthy volunteers (HV) and six UC patients and thus distinguish the mucin adhered from the luminal microbiota. Although under the same nutritional conditions, short-chain fatty acid production by the luminal communities from UC patients showed a tendency toward a lower butyrate production. A more in-depth community analysis of those microbial groups known to produce butyrate revealed that the diversity of the Clostridium coccoides/Eubacterium rectale and Clostridium leptum group, and counts of Faecalibacterium prausnitzii were lower in the luminal fractions of the UC samples. Counts of Roseburia spp. were lower in the mucosal fractions of the UC samples. qPCR analysis for butyryl-CoA:acetate CoA transferase, responsible for butyrate production, displayed a lower abundance in both the luminal and mucosal fractions of the UC samples. The M-SHIME model revealed depletion in butyrate producing microbial communities not restricted to the luminal but also in the mucosal samples from UC patients compared to HV.

Feruloylated and nonferuloylated arabino-oligosaccharides from sugar beet pectin selectively stimulate the growth of Bifidobacterium spp. in human fecal in vitro fermentations.

The side chains of the rhamnogalacturonan I fraction in sugar beet pectin are particularly rich in arabinan moieties, which may be substituted with feruloyl groups. In this work the arabinan-rich fraction resulting from sugar beet pulp based pectin production was separated by Amberlite XAD hydrophobic interaction and membrane separation into four fractions based on feruloyl substitution and arabino-oligosaccharide chain length: short-chain (DP 2-10) and long-chain (DP 7-14) feruloylated and nonferuloylated arabino-oligosaccharides, respectively. HPAEC, SEC, and MALDI-TOF/TOF analyses of the fractions confirmed the presence of singly and doubly substituted feruloylated arabino-oligosaccharides in the feruloyl-substituted fractions. In vitro microbial fermentation by human fecal samples (n = 6 healthy human volunteers) showed a selective stimulation of bifidobacteria by both the feruloylated and the nonferuloylated long-chain arabino-oligosaccharides to the same extent as the prebiotic fructo-oligosaccharides control. None of the fractions stimulated the growth of the potential pathogen Clostridium difficile in monocultures. This work provides a first report on the separation of potentially bioactive feruloylated arabino-oligosaccharides from sugar beet pulp and an initial indication of the potentially larger bifidogenic effect of relatively long-chain arabino-oligosaccharides as opposed to short-chain arabino-oligosaccharides.

Gram-negative bacteria account for main differences between faecal microbiota from patients with ulcerative colitis and healthy controls

Detailed knowledge about the composition of the intestinal microbiota may be critical to unravel the pathogenesis of ulcerative colitis (UC), a human chronic inflammatory bowel disease, since the intestinal microbes are expected to influence some of the key mechanisms involved in the inflammatory process of the gut mucosa. The aim of this study was to investigate the faecal microbiota in patients either with UC in remission (n=6) or with active disease (n=6), and in healthy controls (n=6). The composition of Gram-negative bacteria and Gram-positive bacteria was examined. Antigenic structures of Gram-negative bacteria such as lipopolysaccharides have been related to the inflammatory responses and pathogenesis of inflammatory bowel disease. Dice cluster analysis and principal component analysis of faecal microbiota profiles obtained by denaturing gradient gel electrophoresis and quantitative PCR, respectively, revealed that the composition of faecal bacteria from UC patients with active disease differed from the healthy controls and that this difference should be ascribed to Gram-negative bacteria. The analysis did not show any clear grouping of UC patients in remission. Even with the relatively low number of subjects in each group, we were able to detect a statistically significant underrepresentation of Lactobacillus spp. and Akkermansia muciniphila in UC patients with clinically active disease compared to the healthy controls. In line with previous communications, we have shown that the microbiota in UC patients with active disease differ from that in healthy controls. Our findings indicate that alterations in the composition of the Gram-negative bacterial population, as well as reduced numbers of lactobacilli and A. muciniphila may play a role in UC.

Second meal effect on appetite and fermentation of wholegrain rye foods.

BACKGROUND Wholegrain rye has been associated with decreased hunger sensations. This may be partly mediated by colonic fermentation. Sustained consumption of fermentable components is known to change the gut microflora and may increase numbers of saccharolytic bacteria. OBJECTIVE To investigate the effect of wholegrain rye consumption on appetite and colonic fermentation after a subsequent meal. METHODS In a randomized, controlled, three-arm cross-over study, twelve healthy male subjects consumed three iso-caloric evening test meals. The test meals were based on white wheat bread (WBB), wholegrain rye kernel bread (RKB), or boiled rye kernels (RK). Breath hydrogen excretion and subjective appetite sensation were measured before and at 30 min intervals for 3 h after a standardized breakfast in the subsequent morning. After the 3 h, an ad libitum lunch meal was served to assess energy intake. In an in vitro study, RKB and RK were subjected to digestion and 24 h-fermentation in order to study SCFA production and growth of selected saccharolytic bacteria. RESULTS The test meals did not differ in their effect on parameters of subjective appetite sensation the following day. Ad libitum energy intake at lunch was, however, reduced by 11% (P < 0.01) after RKB and 7% (P < 0.05) after RK compared with after WWB evening meal. Breath hydrogen excretion was significantly increased following RKB and RK evening meals compared with WWB (P < 0.01 and P < 0.05, respectively). Overall, RKB and RK were readily fermented in vitro and exhibited similar fermentation profiles, although total SCFA production was higher for RK compared with RKB (P < 0.001). In vitro fermentation of RKB and RK both increased the relative quantities of Bifidobacterium and decreased Bacteroides compared with inoculum (P < 0.001). The C. coccoides group was reduced after RKB (P < 0.001). CONCLUSION Consumption of wholegrain rye products reduced subsequent ad libitum energy intake in young healthy men, possibly mediated by mechanisms related to colonic fermentation.

Microbiotas from UC patients display altered metabolism and reduced ability of LAB to colonize mucus

We compared fecal microbial communities derived either from Ulcerative Colitis (UC) patients in remission (n = 4) or in relapse (n = 4), or from healthy subjects (n = 4). These communities were used for inoculation of a dynamic in vitro gut model, which contained integrated mucin-covered microcosms. We found that the microbiota of the ‘mucus’ largely differed from that of the ‘lumen’. This was partly due to decreased mucus-associated populations of lactic acid producing bacterial populations (LAB), as LAB originating from UC patients had a significantly decreased capacity to colonize the mucin-covered microcosms as compared to those originating from healthy subjects. We found significant differences between the metabolomes of UC patients in relapse and remission, respectively, while the metabolome of patients in remission resembled that of healthy subjects. These novel findings constitute an important contribution to the understanding of the complex etiology of UC.

Oral supplementation of healthy adults with 2'-O-fucosyllactose and lacto-N-neotetraose is well tolerated and shifts the intestinal microbiota.

The gut microbiota has been established as an important player influencing many aspects of human physiology. Breast milk, the first diet for an infant, contains human milk oligosaccharides (HMO) that shape the infant’s gut microbiota by selectively stimulating the growth of specific bacteria, especially bifidobacteria. In addition to their bifidogenic activity, the ability of HMO to modulate immune function and the gut barrier makes them prime candidates to restore a beneficial microbiota in dysbiotic adults and provide health benefits. We conducted a parallel, double-blind, randomised, placebo-controlled, HMO-supplementation study in 100 healthy, adult volunteers, consuming chemically produced 2′-O-fucosyllactose (2′FL) and/or lacto-N-neotetraose (LNnT) at various daily doses and mixes or placebo for 2 weeks. All participants completed the study without premature discontinuation. Supplementation of 2′FL and LNnT at daily doses up to 20 g was shown to be safe and well tolerated, as assessed using the gastrointestinal symptoms rating scale. 16S rRNA sequencing analysis showed that HMO supplementation specifically modified the adult gut microbiota with the primary impact being substantial increases in relative abundance of Actinobacteria and Bifidobacterium in particular and a reduction in relative abundance of Firmicutes and Proteobacteria. This study provides the first set of data on safety, tolerance and impact of HMO on the adult gut microbiota. Collectively, the results from this study show that supplementing the diet with HMO is a valuable strategy to shape the human gut microbiota and specifically promote the growth of beneficial bifidobacteria.

Associations between common intestinal parasites and bacteria in humans as revealed by qPCR

Several studies have shown associations between groups of intestinal bacterial or specific ratios between bacterial groups and various disease traits. Meanwhile, little is known about interactions and associations between eukaryotic and prokaryotic microorganisms in the human gut. In this work, we set out to investigate potential associations between common single-celled parasites such as Blastocystis spp. and Dientamoeba fragilis and intestinal bacteria. Stool DNA from patients with intestinal symptoms were selected based on being Blastocystis spp.-positive (B+)/negative (B−) and D. fragilis-positive (D+)/negative (D−), and split into four groups of 21 samples (B+ D+, B+ D−, B− D+, and B− D−). Quantitative PCR targeting the six bacterial taxa Bacteroides, Prevotella, the butyrate-producing clostridial clusters IV and XIVa, the mucin-degrading Akkermansia muciniphila, and the indigenous group of Bifidobacterium was subsequently performed, and the relative abundance of these bacteria across the four groups was compared. The relative abundance of Bacteroides in B– D– samples was significantly higher compared with B+ D− and B+ D+ samples (P < 0.05 and P < 0.01, respectively), and this association was even more significant when comparing all parasite-positive samples with parasite-negative samples (P < 0.001). Additionally, our data revealed that a low abundance of Prevotella and a higher abundance of Clostridial cluster XIVa was associated with parasite-negative samples (P < 0.05 and P < 0.01, respectively). Our data support the theory that Blastocystis alone or combined with D. fragilis is associated with gut microbiota characterized by low relative abundances of Bacteroides and Clostridial cluster XIVa and high levels of Prevotella.