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Background Irritable bowel syndrome (IBS) is a functional disorder affecting around 11% of the world population, which diagnostic is mainly based on clinical parameters. IBS shares many symptoms with other gastrointestinal disorders such as inflammatory bowel disease (IBD), which makes positive diagnosis a difficult task. Aim This work presents the design of a new test, called RAID‐Dx, which is a mathematical algorithm based on the combination of nine faecal microbial markers capable of diagnosing IBS. Methods A cohort consisting of 165 subjects (52 IBS and 52 IBD patients, and 61 healthy controls) was recruited from the Gastroenterology Department of six hospitals. Each patient provided a stool sample from which DNA was extracted, and microbial markers composing RAID‐Dx were analysed by qPCR. The results obtained were used to define and validate the RAID‐Dx algorithm. Results The abundance of the biomarkers included in the algorithm differed according to the diagnosis. RAID‐Dx shows a high capacity to diagnose IBS with a sensitivity of 82.4% and a specificity of 85.7%. RAID‐Dx also reports higher sensitivity and specificity values than faecal calprotectin for IBS and IBD differentiation. Conclusions RAID‐Dx is a noninvasive tool aimed to diagnose IBS with high sensitivity and specificity. The use of this new tool for IBS diagnosis could significantly improve disease management, minimise its misdiagnosis and increase patients’ quality of life

Inflammatory bowel disease (IBD), including its two main categories (Crohn's disease and ulcerative colitis), has been linked both to gut microbiota and to diet. Bread is a daily food that has a potential capacity as a prebiotic. Our aim was to evaluate different bread-making processes and their effect on fecal colonic microbiota in IBD patients. The microbial composition of several sourdoughs and dough samples was analyzed by high-throughput sequencing of 16S and 18S rRNA genes. Three types of bread, which followed different bread-making processes, were in vitro digested and incubated with feces from IBD patients. Changes in gut microbiota were assessed by a quantitative polymerase chain reaction using specific bacterial sequence targets. Short-chain fatty acid production was also analyzed by gas chromatography. Lactobacillus sanfranciscensis was the dominant lactic acid bacteria species found in sourdough and bread doughs prepared using sourdough, whereas Saccharomyces cerevisiae was the most dominant yeast in all groups, especially in bread doughs before baking. Differences in microbial composition in raw bread doughs were more related to the type of dough and elaboration than to fermentation time lengths. The analysis of in vitro fecal incubations with bread conditions revealed an increase in most bacterial groups analyzed and short-chain fatty acid production, both in Crohn's disease and ulcerative colitis samples. Most remarkable increases in short-chain fatty acid production mirrored higher abundances of Roseburia species. The potential prebiotic properties observed were mainly obtained when using a high quantity of bread, regardless of bread type. Overall, this study highlights the bacterial dynamics within the bread-making process and the potential prebiotic effect in IBD patients.

Background and Aims Crohn’s disease and ulcerative colitis evolve with alternate outbreaks and remissions of variable duration in both cases. Despite the advances, about 10-30% of patients do not respond to the treatment after the induction period. Besides, between 20% to 50% further patients need an optimization of the dose to respond the treatment. Recent studies have pointed gut microbiota can play a role in the anti-TNF treatment response. This study aimed to define a bacterial signature that could be used to predict the response of patients to anti-TNF treatment. Methods There were obtained 38 stool samples from 38 IBD patients before starting anti-TNF treatments: Adalimumab, Golimumab or Infliximab. Patients were differentiated in 2 groups: responders and non-responders to biological treatment. From each sample, DNA was purified and used in a qPCR for the quantification of the 8 microbial markers. Results In this proof of concept, the predictive ability to identify anti-TNF treatment responders was analyzed. An algorithm consisting in the combination of 4 bacterial markers showed a high capacity to discriminate between responders and non- responders. The algorithm proved high sensitivity and specificity reporting values of 93.33% and 100% respectively, with a positive predictive value of 100% and a negative predictive value of 75% for predicting response to biologic treatment. Conclusions A specific bacterial signature could beneficiate patients with inflammatory bowel disease predicting the therapeutic effectiveness of an anti-TNF treatment, leading to a personalized therapy, improving the patients’ quality of life, saving costs and gaining time in patient improvement.

Inflammatory bowel disease (IBD) and irritable bowel syndrome (IBS) patients have different faecal microbiota profiles compared to healthy controls. Prebiotics intake influences intestinal microbiota composition which in turn influence the growth of short-chain fatty acids (SCFA) producing bacteria. This study aimed to evaluate the capacity of Previpect, a new prebiotic obtained from grapes fibre, to balance the dysbiosis found in patients with intestinal disorders. This was achieved through the analysis of specific bacterial markers and SCFA production using an in vitro fermentation system and comparing the obtained results with those obtained with other commercial prebiotics. Fresh faecal samples from patients with IBD (N = 6), IBS (N = 3), and control subjects (N = 6) were used. Previpect showed high fermentative ability enabling the growth of butyrate producing bacteria and increasing SCFA concentration up to 2.5-fold. Previpect is a promising prebiotic which may be used as a therapeutic strategy towards promotion of intestinal microbiota restoration, microbial healing, and as a preventive supplement for healthy individuals This work was supported by the Centro para el Desarrollo Tecnológico Industrial (SNEO-20151529) and Ministry of Science and Innovation (PTQ-17-09413)

Background Inflammatory Bowel Disease (IBD) and Irritable Bowel Syndrome (IBS) are two intestinal disorders with unknown aetiology. IBD presents two main forms which are Crohn’s disease (CD) and ulcerative colitis (UC). It has been reported recently that bacterial communities present in the colon of patients with either IBD or IBS are structurally different compared to those found in healthy individuals. This dysbiosis mainly consists of a decrease of butyrate producing bacteria and, therefore, the recovery of this metabolic group should be at the baseline of any successful treatment. The main goal of this proof-of-concept study was to test the capacity of a new prebiotic of selected dietary fibre obtained from grape to balance the dysbiosis found in patients with different intestinal disorders. Methods In this study, fresh faecal samples from 6 healthy subjects, 3 CD, 3 UC, and 3 IBS were collected at the Hospital Universitari Dr. Josep Trueta. Fresh samples were diluted 1:5 with fermentation buffer and incubated with 200 mg of prebiotic at 37 ºC under agitation for 72 h. Results were compared with those obtained with various commercial prebiotics after simulating in vitro digestion: inulin, grape pectin, and grape seed extract (GSPE). Total DNA was extracted and the abundance of butyrate-producing bacterial markers: F. prausnitzii (Fpra), its two phylogroups (PHGI and PHGII), Roseburia spp. (Ros) and S. variabile (Sva) was analysed by qPCR. Bacterial activity was determined by measuring short chain fatty acids (SCFA) concentration (butyrate, acetate, and propionate) by gas chromatography. Results The abundance of all butyrate-producing bacteria analysed increased after incubation with the new prebiotic. Among them, Ros presented significantly higher abundance values with respect to negative control in healthy, CU, and IBS patients. It also showed a significant increase in all individuals analysed compared to inulin, grape pectin, and GSPE. Fermentation of the tested new prebiotic, produced significantly higher concentrations of SCFA (p < 0.001) when compared to negative control and all other commercial fibres. Acetate concentration was a 241% higher than negative control, followed by butyrate (187%), and propionate (169%). Conclusion The studied prebiotic produces in vitro an increase of the abundance and enhancement of the metabolism of butyrate-producing bacteria, when compared to commercial prebiotics. These results suggest that this new fibre is a promising prebiotic to be used in promotion of intestinal microbiota restoration at mucosa.

Guidelines recommend routine screening for colorectal cancer (CRC) in asymptomatic adults starting at age 50. The most extensively used noninvasive test for CRC screening is the fecal immunochemical test (FIT), which has an overall sensitivity for CRC of approximately 61.0%-91.0%, which drops to 27.0%-67.0% for advanced adenomas. These figures contain a high false-positive rate and a low positive predictive value. This work aimed to develop a new, noninvasive CRC screening tool based on fecal bacterial markers capable of decreasing FIT false-positive rates in a FIT-positive population. We defined a fecal bacterial signature (RAID-CRC Screen) in a proof-of-concept with 172 FIT-positive individuals and validated the obtained results on an external cohort of 327 FIT-positive subjects. All study participants had joined the national CRC screening program. In the clinical validation of RAID-CRC Screen, a sensitivity of 83.9% and a specificity of 16.3% were obtained for the detection of advanced neoplasm lesions (advanced adenomas and/or CRC). FIT 20 μg/g produced 184 false-positive results. Using RAID-CRC Screen, this value was reduced to 154, thus reducing the false-positive rate by 16.3%. The RAID-CRC Screen test could be implemented in CRC screening programs to allow a significant reduction in the number of colonoscopies performed unnecessarily for FIT-positive participants of CRC screening programs JGG, XA, MSP, MS SRP, JA, LO, MM are employees from GoodGut, a company that has received private and public funding: RTC-2016-5017-1 (Spanish Ministry of Economy, Industry and Competitivity (MINECO)); SNEO-20151529 (Neotec)

Background Dysbiosis is a widely used but unspecific term. It has been defined as any change in the composition of resident microbial communities relative to the ones found in healthy individuals. But it is still unclear which are the appropriate communities to define it and the reference values to measure it. Different studies have described dysbiosis in various diseases such as Inflammatory Bowel Disease (IBD), diabetes, rheumatoid arthritis, and autism, among others. Besides, the microbiota can be altered by transient factors such as antibiotics, diet, stress, or infections. Therefore, different degrees of severity can be associated with the term dysbiosis from which pathological and transient dysbiosis can be differentiated. This work aimed at defining more specifically the term intestinal dysbiosis and to differentiate both transient and pathological dysbiosis. Methods Fifteen key microbial markers belonging to the principal families, classes and orders found in the human intestinal microbiota were accurately selected based on its functionality: F. prausnitzii (Fpra), E. coli (Eco), Firmicutes (Fir), Bacteroidetes (Bac), A. muciniphila (Akk), Ruminococcus sp. (Rum), Roseburia sp. (Ros), Gammaproteobacteria (Gam), Clostridia cluster I (Clo), Clostridia cluster XIV (XIV), Enterococcus sp., Lactobacillus sp. (Lac), C. albicans (Can), M. smithii (Msm), and the total bacterial load (Eub). The dysbiosis was defined using stool samples in a cohort of healthy subjects (n=24) and then validated with 9 patients diagnosed with intestinal diseases 4 IBD and 2 Irritable Bowel Syndrome (IBS)). Total DNA was extracted and the abundance of microbial markers was analysed by qPCR. Together with the establishment of the most common range in which each microbial marker was found, an index to define the pathological dysbiosis was calculated. Results Almost all healthy subjects analysed presented one or two slightly altered markers. The affected microbial markers in this “transient dysbiosis” were Fpra, Akk, and Ros, which are indicative of the mucous layer state, Firm as indicative of the lifestyle and fiber intake, Gam as characteristic of the pro-inflammatory state of the gut, and Msm as an altered intestinal habit and gas production. All patients analyzed (IBD and IBS) presented alterations mainly of the bacteria inhabiting the mucosa and an alteration of the opportunistic species related to the disrupted mucous layer and defining “pathological dysbiosis”. Conclusion This study establishes an appropriate abundance range of key microbial markers in the gut, leading to a specific definition of dysbiosis which allows to differentiate the pathological from the transient dysbiosis. These results need further validation in a larger patient cohort.

Background Inflammatory Bowel Disease (IBD) is a clinical condition of the gastrointestinal tract of unknown aetiology. The two main forms of IBD are Crohn’s disease (CD) and Ulcerative colitis (UC). Recently, it has been reported that the bacterial communities present in the colon of patients with IBD are structurally different compared with those in healthy individuals. This particular dysbiosis consists of a decrease of butyrate producing bacteria and an increase of the pro-inflammatory species. The goal of this work was to test a new prebiotic of selected dietary fibre made from grape for its capacity to balance the dysbiosis typically found in patients with intestinal disorders. Methods Faecal samples from 16 healthy subjects and 11 IBD patients (5 CD and 6 UC) were collected by the Hospital Universitari Dr. Josep Trueta. Fresh stool samples were incubated with 200 mg, 600 mg of prebiotic, and 200 mg of apple pectin. A negative control without substrate addition was also performed. The tubes were incubated under continuous stirring for 72 hours at 37 ºC in semi-anaerobic atmosphere. Total DNA was extracted and the abundances of butyrate-producing bacterial markers (Faecalibacterium prausnitzii, and its phylogroups I and II, Roseburia hominis and Subdolinogranulum variabile) were analysed by qPCR. Concentrations of both butyrate and acetate were determined by gas chromatography as an indication of the bacterial metabolic activity. Results A significant increase of butyrate producing species, such as S. variabile, R. hominis and F. prausnitzii was observed in CD and UC samples when incubated with 200 mg of prebiotic compared with the negative control. F. prausnitzii phylogroup II, which is underrepresented in UC patients, also increased. No differences were found when samples were incubated with 600 mg. Regarding apple pectin, the increase on the abundance of butyrate producing bacteria was higher than the prebiotic in UC samples. In samples from healthy subjects, an increase in the butyrate producing species abundances was observed in both concentrations of prebiotic when compared with the negative control. In those samples no differences were observed when comparing apple pectin with the prebiotic. Concerning butyrate and acetate production, these substances increased in all prebiotic-supplemented samples when compared with the negative control. Conclusion The studied prebiotic causes an increase of the abundance and activity of butyrate-producing bacteria in both IBD patients and healthy subjects. These results points to this new dietary fibre as a promising prebiotic to maintain eubiosis and to promote the microbiota restoration of the intestinal mucosa.

Background Colorectal cancer is the second commonest cause of cancer mortality. Some countries are implementing colorectal cancer screening to detect lesions at an early stage using non-invasive tools like the faecal immunochemical test. Despite affordability, this test shows a low sensitivity for precancerous lesions and a low positive predictive value for colorectal cancer, resulting in a high false-positive rate. Aim To develop a new, non-invasive colorectal cancer screening tool based on bacterial faecal biomarkers, which in combination with the faecal immunochemical test, could allow a reduction in the false-positive rate. This tool is called risk assessment of intestinal disease for colorectal cancer (RAID-CRC). Methods We performed both the faecal immunochemical test and the bacterial markers analysis (RAID-CRC test) in stool samples from individuals with normal colonoscopy (167), non-advanced adenomas (88), advanced adenomas (30) and colorectal cancer (48). All the participants showed colorectal cancer-associated symptoms. Results Performance of the faecal immunochemical test for advanced neoplasia (ie advanced adenoma and colorectal cancer) was determined by using the cut-off value established in Catalonia (20 µg haemoglobin/g of faeces) for a population-based screening approach. Sensitivity and specificity values of 83% and 80%, respectively, and positive and negative predictive values of 56% and 94%, respectively, were obtained. When both the immunological and the biological analysis were combined, the corresponding values were 80% and 90% for sensitivity and specificity, respectively, and 70% and 94% for positive and negative predictive values, respectively, resulting in a 50% reduction of the false-positive rate. Conclusions RAID-CRC test allows a substantial reduction in the faecal immunochemical test false-positive results (50%) in a symptomatic population. Further validation is indicated in a colorectal cancer-screening scenario.

Background Inflammatory bowel disease (IBD) is defined as a group of autoimmune diseases that affects the digestive tract by causing chronic inflammatory processes. Crohn’s disease (CD) is one of the major types of IBD. Although there are several efficient drugs for its treatment, almost 80% of patients will require an intestinal resection throughout their lives. Some bacterial species found in faecal samples can be used as indicators to diagnose and monitor activity in CD patients. However, this methodology has not yet been tested in patients that underwent an intestinal resection. The aim of this study was to investigate the changes in the abundance of these species associated with total or partial intestinal resection. Methods In this study, we used 30 faecal samples from patients diagnosed with CD in remission withdrawn from the Biobank of the Hospital Universitari Dr. Josep Trueta of Girona. Fifteen samples corresponded to patients who underwent an intestinal resection. The following markers were quantified by qPCR: Faecalibacterium prausnitzii (Fpra), F. prausnitzii phylogroup I (PHG-I), F. prausnitzii phylogrup II (PHG-II), Escherichia coli (Eco), Akkermansia muciniphila (Akk), Ruminococcus sp. (Rum), Bacteroidetes (Bac) and Methanobrevibacter smithii (Msm). Results The abundances of Fpra, PHG-I, PHG-II, Rum, and Msm were significantly lower (p < 0.05) in resected CD patients, whereas Eco was significantly more abundant in those patients. No significant differences were observed in the abundances of Akk and Bac. Conclusion In resected CD patients, beneficial butyrate-producing bacteria are underrepresented, whereas pro-inflammatory species are overpopulated. Any quantitative relationship of these indicators with the relapse probability would help to stratify these patients for a better orientation of therapeutic strategies.