Type and Amount of Dietary Fiber Influence the Hindgut Synthesis of Organic Acids from Fermentable Material of Both Total and Nondietary Origin in a Pig Model of the Adult Human.

BACKGROUND: Organic acid synthesis by the hindgut microbiota is commonly believed to be mainly of fermentable material of dietary origin. OBJECTIVE: This study aimed to determine the hindgut organic acid synthesis from fermentable material of dietary (mainly fiber) or nondietary origin for different types and amounts of dietary fiber in growing pigs used as a model for adult humans. METHOD: Seven fiber-containing diets were formulated: 4 fiber types (cellulose, gum acacia, oligofructose, and pectin) at 6% of the diet and 3 (gum acacia, oligofructose, and pectin) at 3% as the sole fiber source. Ileal cannulated female pigs (n = 14; Landrace/Large white) were fed the fiber-containing diets (n = 6 pigs/diet) for 11 days (fiber phase) followed by 3 days on a fiber-free diet (fiber-free phase), using a replicated Youden square. Ileal digesta for each phase were collected and fermented in vitro with a pooled fecal microbial inoculum prepared from feces collected during the fiber phase to determine the organic acids synthesized from fermentable material of dietary (fiber phase) and nondietary (fiber-free phase) origins. RESULTS: The total amount of each individual organic acid synthesized during in vitro hindgut fermentation differed (P ≤ 0.05) across the types and amounts of dietary fiber intake. For example, the amount of acetate was 3.6-fold higher (P ≤ 0.05) for pigs fed the 6% pectin-containing diet than those fed the 6% oligofructose-containing diet. The nondietary substrate contributed between 36% (hexanoate) and 70% (succinate) to the total hindgut organic acid synthesis. The adaptation to the different fiber-containing diets led to different amounts of some organic acids of nondietary origin. CONCLUSIONS: The total amount of organic acids synthesized in the hindgut by the resident microbes is influenced by the type and amount of dietary fiber consumed. This study quantifies the interaction between both dietary and nondietary fermentable materials in hindgut fermentation.

Dietary Fibre and Organic Acids in Kiwifruit Suppress Glycaemic Response Equally by Delaying Absorption-A Randomised Crossover Human Trial with Parallel Analysis of 13C-Acetate Uptake.

Non-sugar components of kiwifruit reduce the amplitude of the glycaemic response to co-consumed cereal starch. We determined the relative contribution of different non-sugar kiwifruit components to this anti-glycaemic effect. Healthy participants (n = 9) ingested equal carbohydrate meals containing 20 g starch as wheat biscuit (WB, 30 g), and the sugar equivalent of two kiwifruit (KFsug, 20.4 g), either intrinsic or added as glucose, fructose and sucrose (2:2:1). The meals were WB+KFsug (control, no non-sugar kiwifruit components), WB + whole kiwifruit pulp (WB+KF), WB + neutralised kiwifruit pulp (WB+KFneut), WB + low-fibre kiwifruit juice (WB+KFjuice) and WB+KFsug + kiwifruit organic acids (WB+KFsug+OA). All meals were spiked with 100 mg sodium [1-13C] acetate to measure intestinal absorption. Each participant ingested all meals in random order. Blood glucose and breath 13CO2 were measured at ingestion and at 15 min intervals up to 180 min. Compared with WB+KFsug, whole kiwifruit pulp (WB+KF) almost halved glycaemic response amplitude (p < 0.001), reduced incremental area under the blood glucose response curve (iAUC) at 30 min (peak) by 50% (p < 0.001), and averted late postprandial hypoglycaemia. All other treatments suppressed response amplitude half as much as whole kiwifruit and averted acute hypoglycaemia, with little effect on iAUC. Effects on 13CO2 exhalation paralleled effects on blood glucose (R2 = 0.97). Dietary fibre and organic acids contributed equally to the anti-glycaemic effect of kiwifruit by reducing intestinal absorption rate. Kiwifruit flesh effectively attenuates glycaemic response in carbohydrate exchange, as it contains fructose, dietary fibre and organic acids.

Chewing differences in consumers affect the digestion and colonic fermentation outcomes: in vitro studies.

It is important to understand variability in consumer chewing behavior for designing food products that deliver desired functionalities for target consumer segments. In this study, we selected 29 participants, representing the large range of chewing variation we had observed in 142 healthy young adults, and investigated the influence of chewing behavior on gastrointestinal digestion and colonic fermentation, using in vitro models and brown rice as a model food. Chewing behavior measured by video observations and chewing outcome differed widely between participants, resulting in large differences in the digestibility of carbohydrates. Inter-individual differences in chewing behavior and chewing outcome also significantly affected in vitro patterns of microbial composition and the production of organic acid metabolites, resulting from colonic fermentation, which is increasingly recognized to be important for human health. These digestion/fermentation outcomes were largely related with the chewing time per mouthful, proportion of bolus particles bigger than 2 mm and amount of saliva added to the bolus during chewing. No significant relationships were found with other chewing trajectory and oral physiological measures. These results suggest that modification of chewing may be an effective strategy to control blood glucose levels and to shape gut microbiota and their metabolites, without altering diets, and that further in vivo studies are warranted to confirm these in vitro findings.

In Vitro Assessment of Hydrolysed Collagen Fermentation Using Domestic Cat (Felis catus) Faecal Inocula.

The gastrointestinal microbiome has a range of roles in the host, including the production of beneficial fermentation end products such as butyrate, which are typically associated with fermentation of plant fibres. However, domestic cats are obligate carnivores and do not require carbohydrates. It has been hypothesised that in the wild, collagenous parts of prey-the so-called animal-derived fermentable substrates (ADFS) such as tendons and cartilage-may be fermented by the cat's gastrointestinal microbiome. However, little research has been conducted on ADFS in the domestic cat. Faecal inoculum was obtained from domestic cats either consuming a high carbohydrate (protein:fat:carbohydrate ratio of 35:20:28 (% dry matter basis)) or high protein (protein:fat:carbohydrate ratio of 75:19:1 (% dry matter basis)) diet. ADFS (hydrolysed collagen, cat hair, and cartilage) were used in a series of static in vitro digestions and fermentations. Concentrations of organic acids and ammonia were measured after 24 h of fermentation, and the culture community of microbes was characterised. The type of inoculum used affected the fermentation profile produced by the ADFS. Butyrate concentrations were highest when hydrolysed collagen was fermented with high protein inoculum (p < 0.05). In contrast, butyrate was not detectable when hydrolysed collagen was fermented in high carbohydrate inoculum (p < 0.05). The microbiome of the domestic cat may be able to ferment ADFS to provide beneficial concentrations of butyrate.

Complementary Food Ingredients Alter Infant Gut Microbiome Composition and Metabolism In Vitro.

We examined the prebiotic potential of 32 food ingredients on the developing infant microbiome using an in vitro gastroileal digestion and colonic fermentation model. There were significant changes in the concentrations of short-chain fatty-acid metabolites, confirming the potential of the tested ingredients to stimulate bacterial metabolism. The 16S rRNA gene sequencing for a subset of the ingredients revealed significant increases in the relative abundances of the lactate- and acetate-producing Bifidobacteriaceae, Enterococcaceae, and Lactobacillaceae, and lactate- and acetate-utilizing Prevotellaceae, Lachnospiraceae, and Veillonellaceae. Selective changes in specific bacterial groups were observed. Infant whole-milk powder and an oat flour enhanced Bifidobacteriaceae and lactic acid bacteria. A New Zealand-origin spinach powder enhanced Prevotellaceae and Lachnospiraceae, while fruit and vegetable powders increased a mixed consortium of beneficial gut microbiota. All food ingredients demonstrated a consistent decrease in Clostridium perfringens, with this organism being increased in the carbohydrate-free water control. While further studies are required, this study demonstrates that the selected food ingredients can modulate the infant gut microbiome composition and metabolism in vitro. This approach provides an opportunity to design nutrient-rich complementary foods that fulfil infants' growth needs and support the maturation of the infant gut microbiome.

The sugar composition of the fibre in selected plant foods modulates weaning infants' gut microbiome composition and fermentation metabolites in vitro.

Eight plant-based foods: oat flour and pureed apple, blackcurrant, carrot, gold- and green-fleshed kiwifruit, pumpkin, sweetcorn, were pre-digested and fermented with pooled inocula of weaning infants' faecal bacteria in an in vitro hindgut model. Inulin and water were included as controls. The pre-digested foods were analysed for digestion-resistant fibre-derived sugar composition and standardised to the same total fibre concentration prior to fermentation. The food-microbiome interactions were then characterised by measuring microbial acid and gas metabolites, microbial glycosidase activity and determining microbiome structure. At the physiologically relevant time of 10 h of fermentation, the xyloglucan-rich apple and blackcurrant favoured a propiogenic metabolic and microbiome profile with no measurable gas production. Glucose-rich, xyloglucan-poor pumpkin caused the greatest increases in lactate and acetate (indicative of high fermentability) commensurate with increased bifidobacteria. Glucose-rich, xyloglucan-poor oats and sweetcorn, and arabinogalactan-rich carrot also increased lactate and acetate, and were more stimulatory of clostridial families, which are indicative of increased microbial diversity and gut and immune health. Inulin favoured a probiotic-driven consortium, while water supported a proteolytic microbiome. This study shows that the fibre-derived sugar composition of complementary foods may shape infant gut microbiome structure and metabolic activity, at least in vitro.

Goat and cow milk differ in altering microbiota composition and fermentation products in rats with gut dysbiosis induced by amoxicillin.

Antibiotics are effective treatments for bacterial infections, however, their oral administration can have unintended consequences and may alter the gut microbiota composition. In this study, we examined the influence of antibiotics on the induction of gut dysbiosis and then evaluated the potential of cow and goat milk to restore the microbiota composition and metabolism in newly weaned rats. In the first study (gut dysbiosis model), rats were treated with amoxicillin, a mixture of antibiotics (ampicillin, gentamicin and metronidazole) or no antibiotics (control). Antibiotics reduced the rat body weights, food intakes and faecal outputs compared to the control group. Gut length was significantly decreased after the antibiotic intake. The bacterial populations (Bifidobacterium spp., Lactobacillus spp. and total bacteria) and short-chain fatty acids (SCFAs; acetic, butyric and propionic) concentrations in rat caecum, colon and faeces were significantly altered by the antibiotic treatments. In the second study, we examined the effects of cow and goat milk in restoring bacterial populations and metabolism in rats with gut dysbiosis induced by amoxicillin. Goat milk significantly increased the numbers of Bifidobacterium spp. and Lactobacillus spp. and decreased the numbers of Clostridium perfringens in the caecum and colon of rats treated with amoxicillin. Whereas, rats fed cow milk had higher Lactobacillus spp. and lower C. perfringens in the gut. Caecal and colonic SCFAs (acetic, butyric and propionic) concentrations differed significantly between rats fed cow and goat milk diets. Overall, goat and cow milk varied in their effects on the immature gut following antibiotic-induced dysbiosis in a rat model.

Influence of kiwifruit on gastric and duodenal inflammation-related gene expression in aspirin-induced gastric mucosal damage in rats.

Kiwifruit (KF) contains bioactive compounds with potential anti-inflammatory properties. In this study, we investigated the protective effects of KF on gastric and duodenal damage induced by soluble aspirin in healthy rats. Sixty-four male Sprague Dawley rats were allocated to eight experimental treatments (n = 8) and the experimental diets were fed for 14 days ad libitum. The experimental diets were 20% fresh pureed KF (green-fleshed and gold-fleshed) or 10% glucose solution (control diet). A positive anti-inflammatory control treatment (ranitidine) was included. At the end of the 14-day feeding period, the rats were fasted overnight, and the following morning soluble aspirin (400 mg/kg aspirin) or water (control) was administered by oral gavage. Four hours after aspirin administration, the rats were euthanized and samples taken for analysis. We observed no significant ulcer formation or increase in infiltration of the gastric mucosal inflammatory cells in the rats with the aspirin treatment. Despite this, there were significant changes in gene expression, such as in the duodenum of aspirin-treated rats fed green KF where there was increased expression of inflammation-related genes NOS2 and TNF-alpha. We also observed that gold and green KF diets had a number of contrasting effects on genes related to inflammation and gastro-protective effects.

In Vivo Assessment of Resistant Starch Degradation by the Caecal Microbiota of Mice Using RNA-Based Stable Isotope Probing-A Proof-of-Principle Study.

Resistant starch (RS) is the digestion resistant fraction of complex polysaccharide starch. By reaching the large bowel, RS can function as a prebiotic carbohydrate, i.e., it can shape the structure and activity of bowel bacterial communities towards a profile that confers health benefits. However, knowledge about the fate of RS in complex intestinal communities and the microbial members involved in its degradation is limited. In this study, 16S ribosomal RNA (rRNA)-based stable isotope probing (RNA-SIP) was used to identify mouse bowel bacteria involved in the assimilation of RS or its derivatives directly in their natural gut habitat. Stable-isotope [U13C]-labeled native potato starch was administrated to mice, and caecal contents were collected before 0 h and 2 h and 4 h after administration. 'Heavy', isotope-labeled [13C]RNA species, presumably derived from bacteria that have metabolized the labeled starch, were separated from 'light', unlabeled [12C]RNA species by fractionation of isolated total RNA in isopycnic-density gradients. Inspection of different density gradients showed a continuous increase in 'heavy' 16S rRNA in caecal samples over the course of the experiment. Sequencing analyses of unlabeled and labeled 16S amplicons particularly suggested a group of unclassified Clostridiales, Dorea, and a few other taxa (Bacteroides, Turicibacter) to be most actively involved in starch assimilation in vivo. In addition, metabolic product analyses revealed that the predominant 13C-labeled short chain fatty acid (SCFA) in caecal contents produced from the [U13C] starch was butyrate. For the first time, this study provides insights into the metabolic transformation of RS by intestinal bacterial communities directly within a gut ecosystem, which will finally help to better understand its prebiotic potential and possible applications in human health.

Consumption of kiwifruit capsules increases Faecalibacterium prausnitzii abundance in functionally constipated individuals: a randomised controlled human trial.

This study investigated the impact of ACTAZIN™ green (2400 and 600 mg) and Livaux™ (2400 mg) gold kiwifruit supplements on faecal microbial composition and metabolites in healthy and functionally constipated (FC) participants. The participants were recruited into the healthy group (n 20; one of whom did not complete the study) and the FC group (n 9), each of whom consumed all the treatments and a placebo (isomalt) for 4 weeks in a randomised cross-over design interspersed with 2-week washout periods. Modification of faecal microbiota composition and metabolism was determined by 16S rRNA gene sequencing and GC, and colonic pH was calculated using SmartPill® wireless motility capsules. A total of thirty-two taxa were measured at greater than 1 % abundance in at least one sample, ten of which differed significantly between the baseline healthy and FC groups. Specifically, Bacteroidales and Roseburia spp. were significantly more abundant (P < 0·05) in the healthy group and taxa including Ruminococcaceae, Dorea spp. and Akkermansia spp. were significantly more abundant (P < 0·05) in the FC group. In the FC group, Faecalibacterium prausnitzii abundance significantly increased (P = 0·024) from 3·4 to 7·0 % following Livaux™ supplementation, with eight of the nine participants showing a net increase. Lower proportions of F. prausnitzii are often associated with gastrointestinal disorders. The discovery that Livaux™ supplementation increased F. prausnitzii abundance offers a potential strategy for improving gut microbiota composition, as F. prausnitzii is a butyrate producer and has also been shown to exert anti-inflammatory effects in many studies.

Monoglobus pectinilyticus gen. nov., sp. nov., a pectinolytic bacterium isolated from human faeces.

A novel anaerobic pectinolytic bacterium (strain 14T) was isolated from human faeces. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain 14T belonged to the family Ruminococcaceae, but was located separately from known clostridial clusters within the taxon. The closest cultured relative of strain 14T was Acetivibrio cellulolyticus (89.7 % sequence similarity). Strain 14T shared ~99 % sequence similarity with cloned 16S rRNA gene sequences from uncultured bacteria derived from the human gut. Cells were Gram-stain-positive, non-motile cocci approximately 0.6 µm in diameter. Strain 14T fermented pectins from citrus peel, apple, and kiwifruit as well as carbohydrates that are constituents of pectins and hemicellulose, such as galacturonic acid, xylose, and arabinose. TEM images of strain 14T, cultured in association with plant tissues, suggested extracellular fibrolytic activity associated with the bacterial cells, forming zones of degradation in the pectin-rich regions of middle lamella. Phylogenetic and phenotypic analysis supported the differentiation of strain 14T as a novel genus in the family Ruminococcaceae. The name Monoglobus pectinilyticus gen. nov., sp. nov. is proposed; the type strain is 14T (JCM 31914T=DSM 104782T).

The fate of (13)C-labelled and non-labelled inulin predisposed to large bowel fermentation in rats.

The fate of stable-isotope (13)C labelled and non-labelled inulin catabolism by the gut microbiota was assessed in a healthy rat model. Sprague-Dawley male rats were randomly assigned to diets containing either cellulose or inulin, and were fed these diets for 3 days. On day (d) 4, rats allocated to the inulin diet received (13)C-labelled inulin. The rats were then fed the respective non-labelled diets (cellulose or inulin) until sampling (d4, d5, d6, d7, d10 and d11). Post feeding of (13)C-labelled substrate, breath analysis showed that (13)C-inulin cleared from the host within a period of 36 hours. Faecal (13)C demonstrated the clearance of inulin from gut with a (13)C excess reaching maximum at 24 hours (d5) and then declining gradually. There were greater variations in caecal organic acid concentrations from d4 to d6, with higher concentrations of acetic, butyric and propionic acids observed in the rats fed inulin compared to those fed cellulose. Inulin influenced caecal microbial glycosidase activity, increased colon crypt depth, and decreased the faecal output and polysaccharide content compared to the cellulose diet. In summary, the presence of inulin in the diet positively influenced large bowel microbial fermentation.

No difference in fecal levels of bacteria or short chain fatty acids in humans, when consuming fruit juice beverages containing fruit fiber, fruit polyphenols, and their combination.

This study examined the effect of a Boysenberry beverage (750 mg polyphenols), an apple fiber beverage (7.5 g dietary fiber), and a Boysenberry plus apple fiber beverage (750 mg polyphenols plus 7.5 g dietary fiber) on gut health. Twenty-five individuals completed the study. The study was a placebo-controlled crossover study, where every individual consumed 1 of the 4 treatments in turn. Each treatment phase was 4-week long and was followed by a 2-week washout period. The trial beverages were 350 g taken in 2 doses every day (ie, 175 mL taken twice daily). The hypothesis for the study was that the combination of polyphenols and fiber would have a greater benefit on gut health than the placebo product or the fiber or polyphenols on their own. There were no differences in fecal levels of total bacteria, Bacteroides-Prevotella-Porphyromonas group, Bifidobacteriumspecies, Clostridium perfringens, or Lactobacillus species among any of the treatment groups. Fecal short chain fatty acid concentrations did not vary among treatment groups, although prostaglandin E2 concentrations were higher after consumption of the Boysenberry juice beverage. No significant differences were found in quantitative measures of gut health between the Boysenberry juice beverage, the apple fiber beverage, the Boysenberry juice plus apple fiber beverage, and the placebo beverage.

In vitro utilization of gold and green kiwifruit oligosaccharides by human gut microbial populations.

We examined the effects of whole kiwifruit on gut microbiota using an in vitro batch model of gastric-ileal digestion and colonic fermentation. Faecal fermentations of gold and green kiwifruit, inulin and water (control) digests were performed for up to 48 h. As compared to the control, gold and green kiwifruit increased Bifidobacterium spp. by 0.9 and 0.8 log(10) cfu/ml, respectively (P < 0.001), and the Bacteroides-Prevotella-Porphyromonas group by 0.4 and 0.5 log(10) cfu/ml, respectively. Inulin only had a bifidogenic effect (+0.4 log(10) cfu/ml). This was accompanied with increases in microbial glycosidases, especially those with substrate specificities relating to the breakdown of kiwifruit oligosaccharides, and with increased generation of short chain fatty acids. The microbial metabolic activity was sustained for up to 48 h, which we attribute to the complexity of the carbohydrate substrate provided by whole kiwifruit. Kiwifruit fermenta supernatant was also separately shown to affect the in vitro proliferation of Bifidobacterium longum, and its adhesion to Caco-2 intestinal epithelial cells. Collectively, these data suggest that whole kiwifruit may modulate human gut microbial composition and metabolism to produce metabolites conducive to increased bifidobacteria-host association.