Nutritional profile of rodent diets impacts experimental reproducibility in microbiome preclinical research.

The lack of reproducibility of animal experimental results between laboratories, particularly in studies investigating the microbiota, has raised concern among the scientific community. Factors such as environment, stress and sex have been identified as contributors, whereas dietary composition has received less attention. This study firstly evaluated the use of commercially available rodent diets across research institutions, with 28 different diets reported by 45 survey respondents. Secondly, highly variable ingredient, FODMAP (Fermentable Oligo-, Di-, Mono-saccharides And Polyols) and gluten content was found between different commercially available rodent diets. Finally, 40 mice were randomized to four groups, each receiving a different commercially available rodent diet, and the dietary impact on cecal microbiota, short- and branched-chain fatty acid profiles was evaluated. The gut microbiota composition differed significantly between diets and sexes, with significantly different clusters in ß-diversity. Total BCFA were highest (p = 0.01) and SCFA were lowest (p = 0.03) in mice fed a diet lower in FODMAPs and gluten. These results suggest that nutritional composition of commercially available rodent diets impact gut microbiota profiles and fermentation patterns, with major implications for the reproducibility of results across laboratories. However, further studies are required to elucidate the specific dietary factors driving these changes.

Gluten-free and low-FODMAP sourdoughs for patients with coeliac disease and irritable bowel syndrome: A clinical perspective.

Wheat- and gluten-containing products are often blamed for triggering a wide range of gastrointestinal symptoms, and this has fueled demand for gluten-free products worldwide. The best studied 'gluten intolerance' is coeliac disease, an auto-immune disease that affects the small intestine. Coeliac disease occurs in 1% of the population and requires strict, life-long avoidance of gluten-containing foods as the only medical treatment. There is a larger group of individuals (around 10-15% of the population) who report a wide-range of gastrointestinal symptoms that respond well to a 'gluten-free diet', but who do not have coeliac disease - so called 'non-coeliac gluten sensitivity (NCGS)'. The team at Monash University has identified other factors in gluten-containing foods that may be responsible for symptoms in this group of individuals with so-called, NCGS. We have evidence that certain poorly absorbed short chain carbohydrates (called FODMAPs) present in many gluten-containing food products, induce symptoms of abdominal pain, bloating, wind and altered bowel habit (associated with irritable bowel syndrome, IBS). Our research has shown that FODMAPs, and not gluten, triggered symptoms in NCGS. Going forward, there are great opportunities for the food industry to develop low FODMAP products for this group, as choice of grain variety and type of food processing technique can greatly reduce the FODMAP levels in foods. The use of sourdough cultures in bread making has been shown to reduce the quantities of FODMAPs (mostly fructan), resulting in bread products that are well tolerated by patients with IBS. Greater interaction between biomedical- and food-scientists will improve understanding about the clinical problems many consumers face, and lead to the development of food products that are better tolerated by this group.

Review article: short chain fatty acids as potential therapeutic agents in human gastrointestinal and inflammatory disorders.

BACKGROUND: Butyrate, propionate and acetate are short chain fatty acids (SCFA), important for maintaining a healthy colon and are considered as protective in colorectal carcinogenesis. However, they may also regulate immune responses and the composition of the intestinal microbiota. Consequently, their importance in a variety of chronic inflammatory diseases is emerging. AIMS: To review the physiology and metabolism of SCFA in humans, cellular and molecular mechanisms by which SCFA may act in health and disease, and approaches for therapeutic delivery of SCFA. METHODS: A PubMed literature search was conducted for clinical and pre-clinical studies using search terms: 'dietary fibre', short-chain fatty acids', 'acetate', 'propionate', 'butyrate', 'inflammation', 'immune', 'gastrointestinal', 'metabolism'. RESULTS: A wide range of pre-clinical evidence supports roles for SCFA as modulators of not only colonic function, but also multiple inflammatory and metabolic processes. SCFA are implicated in many autoimmune, allergic and metabolic diseases. However, translating effects of SCFA from animal studies to human disease is limited by physiological and dietary differences and by the challenge of delivering sufficient amounts of SCFA to the target sites that include the colon and the systemic circulation. Development of novel targeted approaches for colonic delivery, combined with postbiotic supplementation, may represent desirable strategies to achieve adequate targeted SCFA delivery. CONCLUSIONS: There is a large array of potential disease-modulating effects of SCFA. Adequate targeted delivery to the sites of action is the main limitation of such application. The ongoing development and evaluation of novel delivery techniques offer potential for translating promise to therapeutic benefit.

Increasing Symptoms in Irritable Bowel Symptoms With Ingestion of Galacto-Oligosaccharides Are Mitigated by α-Galactosidase Treatment.

OBJECTIVES: Galacto-oligosaccharides (GOS) are dietary FODMAPs (fermentable carbohydrates) associated with triggering gastrointestinal symptoms in patients with irritable bowel syndrome (IBS). This randomized, double-blind, placebo-controlled, cross-over trial aimed to assess whether oral α-galactosidase co-ingestion with foods high in GOS and low in other FODMAPs would reduce symptoms. METHODS: Patients meeting the Rome III criteria for IBS who were hydrogen-producers on breath testing were recruited. Participants were treated with full-dose (300 GALU (galactosidic units) α-galactosidase) and half-dose enzyme (150 GALU α-galactosidase), and placebo (glucose) in a random order with ≤14 days washout between arms. Following a 3-day low FODMAP run-in period, participants consumed provided diets high in GOS for a further 3-days. Gastrointestinal symptoms were measured daily using a 100 mm visual-analogue-scale, and breath samples taken hourly on the second last day with hydrogen content analysed as area-under-the-curve. RESULTS: Thirty-one patients with IBS (20 IBS-D, 4 IBS-C, 7 IBS-M) completed the study. The addition of high GOS foods resulted in a significant increase in overall symptoms with 21 patients exhibiting GOS-sensitivity (>10 mm increase for overall symptoms). Of those, full-dose enzyme reduced overall symptoms (median 24. 5(IQR 17.5-35.8) vs. 5.5(1.5-15.0) mm; P=0.006) and bloating (20.5(9.5-42.0) vs. 6.5(2.0-15.8); P=0.017). Breath hydrogen production was minimal with no differences seen between placebo and full-dose (P=0.597). CONCLUSIONS: Oral α-galactosidase taken with high GOS foods provides a clinically significant reduction in symptoms in GOS-sensitive individuals with IBS. This strategy can be translated into practice to improve tolerance to high GOS foods as an adjunct therapy to the low FODMAP diet.

Inaccuracy of patient-reported descriptions of and satisfaction with bowel actions in irritable bowel syndrome.

BACKGROUND: Perception of diarrhea and constipation differs greatly. This study aimed to correlate subjective and objective assessment of fecal characteristics in irritable bowel syndrome (IBS) patients. METHODS: Data from two interventional dietary trials with varying FODMAP (Fermentable Oligosaccharides, Disaccharides, Monosaccharides And Polyols) or gluten content were interrogated. Subjects rated their dissatisfaction with stool consistency daily using a visual analog scale during the interventions. Subjects collected stools at the end of each intervention. Each stool was scored according to the King's Stool Chart (KSC). Fecal water content (FWC) was measured on pooled feces by freeze drying, with diarrhea defined as ≥78%. KEY RESULTS: Seventy IBS (Rome III) and eight healthy subjects were studied. Each subject's self-rating of stool consistency during the most symptomatic diet was approximately double that of their least. Degree of dissatisfaction with stool consistency correlated poorly with changes in FWC and KSC. IBS subtype related poorly to objective measures of stool consistency. Sixty percent of IBS-D subjects had diarrhea on objective measures. Eighty-five percent with IBS-C had hard and formed stools but three patients met the criteria for diarrhea. One healthy subject had diarrhea on FWC and KSC, and six had hard, formed stools. No differences in FWC was observed when subjects consumed differing amounts of FODMAPs or gluten (all P > .200). CONCLUSIONS AND INFERENCES: There are major disparities between patients' stool descriptions and objective features of constipation and diarrhea. Patient-reported bowel habits require more interrogation for accurate IBS subtyping. Varying FODMAP or gluten content of the diet is not associated with consistent change in FWC.

Reducing the maternal dietary intake of indigestible and slowly absorbed short-chain carbohydrates is associated with improved infantile colic: a proof-of-concept study.

BACKGROUND: To investigate if a low fermentable oligo-, di- and mono-saccharides and polyols (FODMAP) diet consumed by breastfeeding mothers may be associated with reduced symptoms of infantile colic. METHODS: Exclusively breastfeeding mothers and their typically-developing healthy infants who met the Wessel Criteria for infantile colic were recruited from the community, to this single-blind, open-label, interventional study. After a 3-day qualifying period, mothers were provided a low FODMAP 7-day diet. On days 5, 6 and 7 mothers completed a Baby Day Diary. At baseline and at the end of the 7-day dietary intervention, breast milk was analysed for FODMAP content and infant faecal samples for pH. RESULTS: Eighteen breastfeeding mothers (aged 27-40 years) adhered (100%) to the low FODMAP diet. Infants were of gestational age 37-40.3 weeks and aged 2-17 weeks. At entry, crying durations were a mean [95% CI] of 142 [106-61] min and fell by 52 [178-120] min (P = 0.005; ancova). Combined crying-fussing durations fell by 73 [301-223] min (n = 13; P = 0.007), as did crying episodes (P = 0.01) and fussing durations (P = 0.011). Infant sleeping, feeding, or awake-and-content durations did not change. Infant faecal pH did not change. Breast milk lactose content was stable and other known FODMAPs were not detected. At end of study, mothers reported their baby 'is much more content' and 'can be put down without crying'. CONCLUSIONS: Maternal low FODMAP diet may be associated with a reduction in infant colic symptoms. A randomized controlled study is warranted to determine if a maternal low FODMAP diet is effective in reducing symptoms.

Adding glucose to food and solutions to enhance fructose absorption is not effective in preventing fructose-induced functional gastrointestinal symptoms: randomised controlled trials in patients with fructose malabsorption.

BACKGROUND: In healthy individuals, the absorption of fructose in excess of glucose in solution is enhanced by the addition of glucose. The present study aimed to assess the effects of glucose addition to fructose or fructans on absorption patterns and genesis of gastrointestinal symptoms in patients with functional bowel disorders. METHODS: Randomised, blinded, cross-over studies were performed in healthy subjects and functional bowel disorder patients with fructose malabsorption. The area-under-the-curve (AUC) was determined for breath hydrogen and symptom responses to: (i) six sugar solutions (fructose in solution) (glucose; sucrose; fructose; fructose + glucose; fructan; fructan + glucose) and (ii) whole foods (fructose in foods) containing fructose in excess of glucose given with and without additional glucose. Intake of fermentable short chain carbohydrates (FODMAPs; fermentable, oligo-, di-, monosaccharides and polyols) was controlled. RESULTS: For the fructose in solution study, in 26 patients with functional bowel disorders, breath hydrogen was reduced after glucose was added to fructose compared to fructose alone [mean (SD) AUC 92 (107) versus 859 (980) ppm 4 h-1 , respectively; P = 0.034). Glucose had no effect on breath hydrogen response to fructans (P = 1.000). The six healthy controls showed breath hydrogen patterns similar to those with functional bowel disorders. No differences in symptoms were experienced with the addition of glucose, except more nausea when glucose was added to fructose (P = 0.049). In the fructose in foods study, glucose addition to whole foods containing fructose in excess of glucose in nine patients with functional bowel disorders and nine healthy controls had no significant effect on breath hydrogen production or symptom response. CONCLUSIONS: The absence of a favourable response on symptoms does not support the concomitant intake of glucose with foods high in either fructose or fructans in patients with functional bowel disorders.

Randomised clinical trial: the efficacy of gut-directed hypnotherapy is similar to that of the low FODMAP diet for the treatment of irritable bowel syndrome.

BACKGROUND: A low fermentable oligosaccharides, disaccharides, monosaccharides and polyols (FODMAP) diet is effective in treating irritable bowel syndrome (IBS). AIM: To compare the effects of gut-directed hypnotherapy to the low FODMAP diet on gastrointestinal symptoms and psychological indices, and assess additive effects. METHODS: Irritable bowel syndrome patients were randomised (computer-generated list), to receive hypnotherapy, diet or a combination. Primary end-point: change in overall gastrointestinal symptoms across the three groups from baseline to week 6. Secondary end-points: changes in psychological indices, and the durability of effects over 6 months. RESULTS: Of 74 participants, 25 received hypnotherapy, 24 diet and 25 combination. There were no demographic differences at baseline across groups. Improvements in overall symptoms were observed from baseline to week 6 for hypnotherapy [mean difference (95% CI): -33 (-41 to -25)], diet [-30 (-42 to -19)] and combination [-36 (-45 to -27)] with no difference across groups (P = 0.67). This represented ≥20 mm improvement on visual analogue scale in 72%, 71% and 72%, respectively. This improvement relative to baseline symptoms was maintained 6 months post-treatment in 74%, 82% and 54%. Individual gastrointestinal symptoms similarly improved. Hypnotherapy resulted in superior improvements on psychological indices with mean change from baseline to 6 months in State Trait Personality Inventory trait anxiety of -4(95% CI -6 to -2) P < 0.0001; -1(-3 to 0.3) P = ns; and 0.3(-2 to 2) P = ns, and in trait depression of -3(-5 to -0.7) P = 0.011; -0.8(-2 to 0.2) P = ns; and 0.6(-2 to 3) P = ns, respectively. Groups improved similarly for QOL (all p ≤ 0.001). CONCLUSIONS: Durable effects of gut-directed hypnotherapy are similar to those of the low FODMAP diet for relief of gastrointestinal symptoms. Hypnotherapy has superior efficacy to the diet on psychological indices. No additive effects were observed.

Review article: insights into colonic protein fermentation, its modulation and potential health implications.

BACKGROUND: Beneficial effects of carbohydrate fermentation on gastrointestinal health are well established. Conversely, protein fermentation generates harmful metabolites but their relevance to gastrointestinal health is poorly understood. AIM: To review the effects of increased protein fermentation on biomarkers of colonic health, factors influencing fermentative activity and potential for dietary modulation to minimise detrimental effects. METHODS: A literature search was performed in PubMed, Medline, EMBASE and Google scholar for clinical and pre-clinical studies using search terms - 'dietary protein', 'fermentation', 'putrefaction', 'phenols', 'sulphide', 'branched-chain fatty acid', 'carbohydrate fermentation', 'gastrointestinal'. RESULTS: High protein, reduced carbohydrate diets alter the colonic microbiome, favouring a potentially pathogenic and pro-inflammatory microbiota profile, decreased short-chain fatty acid production and increased ammonia, phenols and hydrogen sulphide concentrations. These metabolites largely compromise the colonic epithelium structure, causing mucosal inflammation but may also directly modulate the enteric nervous system and intestinal motility. Increased protein fermentation as a result of a high-protein intake can be attenuated by addition of oligosaccharides, resistant starch and nonstarch polysaccharides and a reduction in total protein or specifically, aromatic and sulphur-containing amino acids. These factors may have clinical importance as novel therapeutic approaches to problems, in which protein fermentation may be implicated, such as malodorous flatus, irritable bowel syndrome, ulcerative colitis and prevention of colorectal cancer. CONCLUSIONS: The direct clinical relevance of excessive protein fermentation in the pathogenesis of irritable bowel syndrome, malodorous flatus and ulcerative colitis are underexplored. Manipulating dietary carbohydrate and protein intake have potential therapeutic applications in such settings and warrant further clinical studies.

Review article: gut-directed hypnotherapy in the management of irritable bowel syndrome and inflammatory bowel disease.

BACKGROUND: Gut-directed hypnotherapy is being increasingly applied to patients with irritable bowel syndrome (IBS) and to a lesser extent, inflammatory bowel disease (IBD). AIM: To review the technique, mechanisms of action and evidence for efficacy, and to identify gaps in the understanding of gut-directed hypnotherapy as a treatment for IBS and IBD. METHODS: A review of published literature and a systematic review of clinical trials in its application to patients with IBS and IBD were performed. RESULTS: Gut-directed hypnotherapy is a clearly described technique. Its potential mechanisms of action on the brain-gut axis are multiple with evidence spanning psychological effects through to physiological gastrointestinal modifications. Six of seven randomised IBS studies reported a significant reduction (all P < 0.05) in overall gastrointestinal symptoms following treatment usually compared to supportive therapy only. Response rates amongst those who received gut-directed hypnotherapy ranged between 24% and 73%. Efficacy was maintained long-term in four of five studies. A therapeutic effect was also observed in the maintenance of clinical remission in patients with ulcerative colitis. Uncontrolled trials supported the efficacy and durability of gut-directed hypnotherapy in IBS. Gaps in understanding included to whom and when it should be applied, the paucity of adequately trained hypnotherapists, and the difficulties in designing well controlled-trials. CONCLUSIONS: Gut-directed hypnotherapy has durable efficacy in patients with IBS and possibly ulcerative colitis. Whether it sits in the therapeutic arsenal as a primary and/or adjunctive therapy cannot be ascertained on the current evidence base. Further research into efficacy, mechanisms of action and predictors of response is required.

Randomised clinical trial: gluten may cause depression in subjects with non-coeliac gluten sensitivity - an exploratory clinical study.

BACKGROUND: Current evidence suggests that many patients with self-reported non-coeliac gluten sensitivity (NCGS) retain gastrointestinal symptoms on a gluten-free diet (GFD) but continue to restrict gluten as they report 'feeling better'. AIM: To investigate the notion that a major effect of gluten in those with NCGS is on mental state and not necessarily on gastrointestinal symptoms. METHODS: Twenty-two subjects (24-62 years, five male) with irritable bowel syndrome who had coeliac disease excluded but were symptomatically controlled on a GFD, undertook a double-blind cross-over study. Participants randomly received one of three dietary challenges for 3 days, followed by a minimum 3-day washout before crossing over to the next diet. Challenge gluten-free food was supplemented with gluten (16 g/day), whey (16 g/day) or not supplemented (placebo). End-points included mental state as assessed by the Spielberger State Trait Personality Inventory (STPI), cortisol secretion and gastrointestinal symptoms. RESULTS: Gluten ingestion was associated with higher overall STPI state depression scores compared to placebo [M = 2.03, 95% CI (0.55-3.51), P = 0.010] but not whey [M = 1.48, 95% CI (-0.14 to 3.10), P = 0.07]. No differences were found for other STPI state indices or for any STPI trait measures. No difference in cortisol secretion was identified between challenges. Gastrointestinal symptoms were induced similarly across all dietary challenges. CONCLUSIONS: Short-term exposure to gluten specifically induced current feelings of depression with no effect on other indices or on emotional disposition. Gluten-specific induction of gastrointestinal symptoms was not identified. Such findings might explain why patients with non-coeliac gluten sensitivity feel better on a gluten-free diet despite the continuation of gastrointestinal symptoms.

Dietary sorbitol and mannitol: food content and distinct absorption patterns between healthy individuals and patients with irritable bowel syndrome.

BACKGROUND: Sorbitol and mannitol are naturally-occurring polyol isomers. Although poor absorption and induction of gastrointestinal symptoms by sorbitol are known, the properties of mannitol are poorly described. We aimed to expand data on food composition of these polyols, and to compare their absorptive capacities and symptom induction in patients with irritable bowel syndrome (IBS) and healthy individuals. METHODS: Food samples were analysed for sorbitol and mannitol content. The degree of absorption measured by breath hydrogen production and gastrointestinal symptoms (visual analogue scales) was evaluated in a randomised, double-blinded, placebo-controlled study in 21 healthy and 20 IBS subjects after challenges with 10 g of sorbitol, mannitol or glucose. RESULTS: Certain fruits and sugar-free gum contained sorbitol, whereas mannitol content was higher in certain vegetables. Similar proportions of patients with IBS (40%) and healthy subjects (33%) completely absorbed sorbitol, although more so with IBS absorbed mannitol (80% versus 43%; P = 0.02). Breath hydrogen production was similar in both groups after lactulose but was reduced in patients with IBS after both polyols. No difference in mean (SEM) hydrogen production was found in healthy controls after sorbitol [area-under-the-curve: 2766 (591) ppm 4 h(-1) ] or mannitol [2062 (468) ppm 4 h(-1) ] but, in patients with IBS, this was greater after sorbitol [1136 (204) ppm 4 h(-1) ] than mannitol [404 (154) ppm 4 h(-1) ; P = 0.002]. Overall gastrointestinal symptoms increased significantly after both polyols in patients with IBS only, although they were independent of malabsorption of either of the polyols. CONCLUSIONS: Increased and discordant absorption of mannitol and sorbitol occurs in patients with IBS compared to that in healthy controls. Polyols induced gastrointestinal symptoms in patients with IBS independently of their absorptive patterns, suggesting that the dietary restriction of polyols may be efficacious.

Functional bowel symptoms and diet.

It is well recognised that ingestion of food is a trigger for functional bowel symptoms, particularly those associated with irritable bowel syndrome (IBS). Patients often use manipulation of diet as a means of controlling symptoms. Despite description of multiple dietary methods, few have scientific backing or quality evidence of efficacy. One approach is to define how specific food components influence the pathophysiology of IBS and then rationally design dietary approaches. For example, short-chain poorly absorbed carbohydrates (fermentable oligo-, di- and mono-saccharides and polyols (FODMAP)) cause luminal distension, which is a major stimulus for the development of symptoms in patients with visceral hypersensitivity. By determining food content of FODMAP, a diet in which foods low in FODMAP are favoured over those high in FODMAP can be designed. Observational, comparative and randomised controlled treatment and rechallenge studies have shown that such an approach is efficacious in the majority of patients with IBS. The low FODMAP diet is now the primary dietary therapy for such patients. Similar approaches can be applied to other food components, including proteins (such as gluten), fats and natural bioactive food chemicals. Such approaches have suggestions of efficacy, but the evidence base remains underdeveloped. An additional and important consideration for any dietary therapy is its nutritional adequacy and potential adverse health effects. Dietary manipulation is now a key management strategy in patients with functional bowel symptoms. Future well-designed interventional studies will lead to refinement of dietary approaches taken and to a better understanding of their long-term safety.

Quantification of fructans, galacto-oligosacharides and other short-chain carbohydrates in processed grains and cereals.

BACKGROUND: Wholegrain grains and cereals contain a wide range of potentially protective factors that are relevant to gastrointestinal health. The prebiotics best studied are fructans [fructooligosaccharides (FOS), inulin] and galactooligosaccharides (GOS). These and other short-chain carbohydrates can also be poorly absorbed in the small intestine (named fermentable oligo-, di- and monosaccharides and polyols; FODMAPs) and may have important implications for the health of the gut. METHODS: In the present study, FODMAPs, including fructose in excess of glucose, FOS (nystose, kestose), GOS (raffinose, stachyose) and sugar polyols (sorbitol, mannitol), were quantified using high-performance liquid chromatography with an evaporative light scattering detector. Total fructan was quantified using an enzymic hydrolysis method. RESULTS: Fifty-five commonly consumed grains, breakfast cereals, breads, pulses and biscuits were analysed. Total fructan were the most common short-chain carbohydrate present in cereal grain products and ranged (g per portion as eaten) from 1.12 g in couscous to 0 g in rice; 0.6 g in dark rye bread to 0.07 g in spelt bread; 0.96 g in wheat-free muesli to 0.11 g in oats; and 0.81 g in muesli fruit bar to 0.05 g in potato chips. Raffinose and stachyose were most common in pulses. CONCLUSIONS: Composition tables including FODMAPs and prebiotics (FOS and GOS) that are naturally present in food will greatly assist research aimed at understanding their physiological role in the gut.