The Effect of Delivery Matrix on Bifidobacterium animalis subsp. lactis HN019 Survival through In Vitro Human Digestion.

Bifidobacterium animalis subsp. lactis HN019 is a probiotic with several documented human health benefits. Interest in probiotics has led to the development of new formats that probiotics, including HN019, can be supplemented into. In this study, we looked at common HN019 formats such as frozen culture and freeze-dried powder as well as supplementing it into the following food matrices: yogurts (dairy, soy, and oat based), xanthan gum-based tablets, pulpless orange juice, whey sports drink, and dark chocolate (70% cocoa). In this work, our aim was to investigate whether the food matrix that carried HN019 via simulated human digestion (a dual model system mimicking both upper and lower gastrointestinal digestion) influenced probiotic delivery. To that end, we validated and used a real-time qPCR assay to detect HN019 after simulated digestion. In addition, we also measured the effect on a panel of metabolites. After simulated digestion, we were able to detect HN019 from all the matrices tested, and the observed changes to the metabolite profile were consistent with those expected from the food matrix used. In conclusion, this work suggests that the food matrix supplemented with HN019 did not interfere with delivery to the colon via simulated human digestion.

An in vitro model of the chicken gastrointestinal tract with special emphasis to the cecal microbiota.

An in vitro model of the upper gastrointestinal tract as well as the chicken cecum was developed to have a predictive tool for estimating the production performance of animals by analyzing the feeding value of a certain diet. The upper gastrointestinal tract consists of a batch type model, whereas the cecal model is comprised of 4 semi-continuous connected vessels inoculated with cecal or fecal microbes. The upper gastrointestinal tract and cecal simulations were both run with a corn- and a wheat-based diet to simulate 2 typical feed types. Samples were collected after the 5-h cecal simulations and aliquots were frozen to assess inoculum stability. The microbiota was analyzed by 16S rRNA gene sequencing, whereas short chain fatty acids as microbial metabolites were analyzed by using gas chromatography. As expected, some significant differences in microbial abundance after simulation between the cecal and fecal slurry samples (P = 0.001) were detected, as well between the fresh and frozen status (P = 0.001), hence simulations inoculated with cecal and fresh samples being more diverse. For the measured metabolites, almost all of them increased (P < 0.05) significantly when comparing fresh and frozen inoculum. The present chicken intestinal in vitro model represents a rapid systematic screening system for studying dietary related microbial changes and reducing the need of animal sacrifice for experimentation.

Dynamics of the Lipidome in a Colon Simulator.

Current evidence suggests that gut microbiome-derived lipids play a crucial role in the regulation of host lipid metabolism. However, not much is known about the dynamics of gut microbial lipids within the distinct gut biogeographic. Here we applied targeted and untargeted lipidomics to in vitro-derived feces. Simulated intestinal chyme was collected from in vitro gut vessels (V1-V4), representing proximal to distal parts of the colon after 24 and 48 h with/without polydextrose treatment. In total, 44 simulated chyme samples were collected from the in vitro colon simulator. Factor analysis showed that vessel and time had the strongest impact on the simulated intestinal chyme lipid profiles. We found that levels of phosphatidylcholines, sphingomyelins, triacylglycerols, and endocannabinoids were altered in at least one vessel (V1-V4) during simulation. We also found that concentrations of triacylglycerols, diacylglycerols, and endocannabinoids changed with time (24 vs. 48 h of simulation). Together, we found that the simulated intestinal chyme revealed a wide range of lipids that remained altered in different compartments of the human colon model over time.

In Vitro Screen of Lactobacilli Strains for Gastrointestinal and Vaginal Benefits.

Traditional probiotics comprise mainly lactic acid bacteria that are safe for human use, tolerate acid and bile, and adhere to the epithelial lining and mucosal surfaces. In this study, one hundred commercial and non-commercial strains that were isolated from human feces or vaginal samples were tested with regards to overall growth in culture media, tolerance to acid and bile, hydrogen peroxide (H2O2) production, and adhesion to vaginal epithelial cells (VECs) and to blood group antigens. As a result, various of the tested lactobacilli strains were determined to be suitable for gastrointestinal or vaginal applications. Commercial strains grew better than the newly isolated strains, but tolerance to acid was a common property among all tested strains. Tolerance to bile varied considerably between the strains. Resistance to bile and acid correlated well, as did VEC adhesion and H2O2 production, but H2O2 production was not associated with resistance to bile or acid. Except for L. iners strains, vaginal isolates had better overall VEC adhesion and higher H2O2 production. Species- and strain-specific differences were evident for all parameters. Rank-ordered clustering with nine clusters was used to identify strains that were suitable for gastrointestinal or vaginal health, demonstrating that the categorization of strains for targeted health indications is possible based on the parameters that were measured in this study.

A Healthy Vaginal Microbiota Remains Stable during Oral Probiotic Supplementation: A Randomised Controlled Trial.

The primary objective of this randomised, placebo-controlled, triple-blind study was to assess whether orally consumed Lactobacillus acidophilus La-14 (La-14) and Lacticaseibacillus rhamnosus HN001 (HN001) colonise a healthy human vagina. Furthermore, potential effects on vaginal microbiota and immune markers were explored. Fifty women devoid of vaginal complaints (Nugent score 0-3 and vaginal pH ≤ 4.5) were randomised into a 2-week intervention with either La-14 and HN001 as the verum product or a comparable placebo. Vaginal swab samples were collected at baseline, after one and two weeks of intervention, and after a one-week follow-up, for assessing colonisation of the supplemented lactobacilli, vaginal microbiota, and six specific immune markers. Colonisation of L. acidophilus and L. rhamnosus was not observed above the assay detection limit (5.29 and 5.11 log 10 genomes/swab for L. acidophilus and L. rhamnosus, respectively). Vaginal microbiotas remained stable and predominated by lactobacilli throughout the intervention, and vaginal pH remained optimal (at least 90% of participants in both groups had pH 4.0 or 4.5 throughout the study). Immune markers elafin and human ß-defensin 3 (HBD-3) were significantly decreased in the verum group (p = 0.022 and p = 0.028, respectively) but did not correlate with any microbiota changes. Adverse events raised no safety concerns, and no undesired changes in the vaginal microbiota or immune markers were detected.

Cross-sectional observational study protocol: missing microbes in infants born by caesarean section (MiMIC): antenatal antibiotics and mode of delivery.

INTRODUCTION: The intestinal microbiome in early life plays a major role in infant health and development. Factors like antibiotic exposure, breast/formula feeding and mode of delivery are known to affect the microbiome. The increasing occurrence of caesarean section (C-section) deliveries and antibiotic exposure warrants further insight into the potential missing microbes in those infants. The study objective is to study the effect of maternal antibiotic administration during pregnancy and/or C-section mode of delivery on the development of the infant's intestinal microbiome until the age of 2 years. METHODS AND ANALYSIS: A single site, cross-sectional observational study of C-section and vaginally delivered infants being either exposed to maternal antibiotic treatment or not during the third trimester of pregnancy. Throughout the nine visits, stool, urine, saliva, hair, breast milk and vaginal swabs will be collected from either mother and/or infant for microbiome and metabolomic analysis. ETHICS AND DISSEMINATION: The protocol was approved by the Clinical Research Ethics Committee of the Cork Teaching Hospitals. The trial has been registered at ClinicalTrials.gov.The findings from this study will be disseminated in peer-reviewed journals, during scientific conferences, and directly to the study participants. Sequencing data will be deposited in public databases. TRIAL REGISTRATION NUMBER: NCT04134819.

Bifidobacterium animalis subsp. lactis Bi-07 supports lactose digestion in vitro and in randomized, placebo- and lactase-controlled clinical trials.

BACKGROUND: Probiotics may alleviate lactose maldigestion. OBJECTIVES: The objective was to select a probiotic with high lactase activity and compare it with lactase and placebo in clinical trials. METHODS: Bacterial cultures were screened for lactase activity in a model of the upper gastrointestinal (GI) tract. Bifidobacterium animalis subsp. lactis Bi-07 (Bi-07) counts were adjusted in subsequent experiments to correspond to 4500 Food Chemicals Codex (FCC) units of lactase, the amount in the European Food Safety Authority (EFSA)-approved health claim. Two crossover clinical trials, Booster Alpha and Booster Omega, were performed in participants with lactose intolerance, where 2 × 1012 CFUs Bi-07, 4662 FCC lactase, or placebo was consumed simultaneously with a lactose challenge, with 1-wk washouts between challenges. The trial designs were identical except for the source of lactose. Breath hydrogen concentration (BHC) was measured to assess the effect of the investigational products on lactose digestion, for which incremental area under the curve (iAUC) was the primary outcome. Peak BHC, cumulative BHC, and GI symptoms were secondary outcomes. RESULTS: Bi-07 was superior to placebo in reducing BHC [iAUC, parts per million (ppm) ∙ h] in both trials (Booster Alpha: geometric least square mean ratio: 0.462; 95% CI: 0.249, 0.859; P = 0.016; Booster Omega: 0.227; 95% CI: 0.095, 0.543; P = 0.001). Lactase was superior to placebo in Booster Alpha (0.190; 95% CI: 0.102, 0.365; P < 0.001) but not Booster Omega (0.493; 95% CI: 0.210, 1.156; P = 0.102). Noninferiority of Bi-07 compared with lactase was observed in Booster Omega (0.460; 95% CI: 0.193, 1.096; P = 0.079; CI upper limit < 1.25 noninferiority margin). Odds of abdominal pain (compared with placebo: 0.32, P = 0.036) and flatulence (compared with placebo: 0.25, P = 0.007) were lower with lactase in Booster Alpha. Increased odds of nausea were seen with Bi-07 (compared with placebo: 4.0, P = 0.005) in Booster Omega. CONCLUSIONS: Bi-07 has high lactase activity, and in 2 clinical trials, it supported lactose digestion in individuals with lactose intolerance.These trials were registered at clinicaltrials.gov as NCT03659747 (Booster Alpha) and NCT03814668 (Booster Omega).

Identification and Characterization of a Novel Species of Genus Akkermansia with Metabolic Health Effects in a Diet-Induced Obesity Mouse Model.

Akkermansia muciniphila is a well-known bacterium with the ability to degrade mucin. This metabolic capability is believed to play an important role in the colonization of this bacterium in the gut. In this study, we report the identification and characterization of a novel Akkermansia sp. DSM 33459 isolated from human feces of a healthy donor. Phylogenetic analysis based on the genome-wide average nucleotide identity indicated that the Akkermansia sp. DSM 33459 has only 87.5% similarity with the type strain A. muciniphila ATCC BAA-835. Akkermansia sp. DSM 33459 showed significant differences in its fatty acid profile and carbon utilization as compared to the type strain. The Akkermansia sp. DSM 33459 strain was tested in a preclinical obesity model to determine its effect on metabolic markers. Akkermansia sp. DSM 33459 showed significant improvement in body weight, total fat weight, and resistin and insulin levels. Interestingly, these effects were more pronounced with the live form as compared to a pasteurized form of the strain. The strain showed production of agmatine, suggesting a potential novel mechanism for supporting metabolic and cognitive health. Based on its phenotypic features and phylogenetic position, it is proposed that this isolate represents a novel species in the genus Akkermansia and a promising therapeutic candidate for the management of metabolic diseases.

One Giant Leap from Mouse to Man: The Microbiota-Gut-Brain Axis in Mood Disorders and Translational Challenges Moving towards Human Clinical Trials.

The microbiota-gut-brain axis is a bidirectional communication pathway that enables the gut microbiota to communicate with the brain through direct and indirect signaling pathways to influence brain physiology, function, and even behavior. Research has shown that probiotics can improve several aspects of health by changing the environment within the gut, and several lines of evidence now indicate a beneficial effect of probiotics on mental and brain health. Such evidence has prompted the arrival of a new term to the world of biotics research: psychobiotics, defined as any exogenous influence whose effect on mental health is bacterially mediated. Several taxonomic changes in the gut microbiota have been reported in neurodevelopmental disorders, mood disorders such as anxiety and depression, and neurodegenerative disorders such as Alzheimer's disease. While clinical evidence supporting the role of the gut microbiota in mental and brain health, and indeed demonstrating the beneficial effects of probiotics is rapidly accumulating, most of the evidence to date has emerged from preclinical studies employing different animal models. The purpose of this review is to focus on the role of probiotics and the microbiota-gut-brain axis in relation to mood disorders and to review the current translational challenges from preclinical to clinical research.

Current Perspectives on Gastrointestinal Models to Assess Probiotic-Pathogen Interactions.

There are different models available that mimic the human intestinal epithelium and are thus available for studying probiotic and pathogen interactions in the gastrointestinal tract. Although, in vivo models make it possible to study the overall effects of a probiotic on a living subject, they cannot always be conducted and there is a general commitment to reduce the use of animal models. Hence, in vitro methods provide a more rapid tool for studying the interaction between probiotics and pathogens; as well as being ethically superior, faster, and less expensive. The in vitro models are represented by less complex traditional models, standard 2D models compromised of culture plates as well as Transwell inserts, and newer 3D models like organoids, enteroids, as well as organ-on-a-chip. The optimal model selected depends on the research question. Properly designed in vitro and/or in vivo studies are needed to examine the mechanism(s) of action of probiotics on pathogens to obtain physiologically relevant results.

Effects of Bifidobacterium animalis ssp. lactis 420 on gastrointestinal inflammation induced by a nonsteroidal anti-inflammatory drug: A randomized, placebo-controlled, double-blind clinical trial.

AIMS: Use of nonsteroidal anti-inflammatory drugs (NSAIDs) can cause damage to the gastric and duodenal mucosa. Some probiotics have proven useful in ameliorating the harmful side-effects of NSAIDs. Our aim was to evaluate whether oral administration of Bifidobacterium animalis ssp. lactis 420 (B420) can attenuate the increase of calprotectin excretion into faeces induced by intake of diclofenac sustained-release tablets. METHODS: A double-blind, parallel-group, placebo-controlled and randomized clinical study was performed in 50 healthy male and female volunteers aged 20-40 years, in Finland. Study participation consisted of 4 phases: run-in, intervention with B420 or placebo, B420 or placebo + NSAID treatment, and follow-up. The primary outcome was the concentration of calprotectin in faeces. Secondary outcomes were haemoglobin and microbial DNA in faeces and blood haemoglobin levels. RESULTS: Intake of diclofenac increased the faecal excretion of calprotectin in both groups. The observed increases were 48.19 ± 61.55 µg/g faeces (mean ± standard deviation) in the B420 group and 31.30 ± 39.56 µg/g in the placebo group (difference estimate 16.90; 95% confidence interval: -14.00, 47.77; P = .276). There were no significant differences between the treatment groups in changes of faecal or blood haemoglobin. Faecal B. lactis DNA was much more abundant in the B420 group compared to the placebo group (ANOVA estimate for treatment difference 0.85 × 109 /g faeces; 95% confidence interval: 0.50 × 109 , 1.21 × 109 ; P < .0001). CONCLUSIONS: Short-term administration of the probiotic B420 did not protect the healthy adult study participants from diclofenac-induced gastrointestinal inflammation as determined by analysis of faecal calprotectin levels.

Probiotic triangle of success; strain production, clinical studies and product development.

The successful development of probiotic foods and dietary supplements rests on three pillars; each with their specific challenges and opportunities. First, strain production; this depends on selecting the right strain with promising technological properties and safety profile. Further the manufacturing of the strain in a stable format at sufficiently high yield, following regulatory and customer requirements on culture media ingredients and other processing aids. The second pillar are the preclinical and clinical studies to document that the strain is a probiotic and exerts a health benefit on the host, the consumer. Especially when aiming for a regulator approved health claim, clinical studies need to be thoroughly performed; following appropriate ethical, scientific and regulatory guidelines. Finally, the probiotic will need to be incorporated in a product that can be brought to the consumer; a dietary supplement or a functional food. Because of the live nature of probiotics, specific challenges may need to be dealt with. Although experience from other strains is helpful in the process, the development is strain specific. Commercialisation and marketing of probiotics are strictly but differently regulated in most jurisdictions; defining what can and cannot be claimed.

Fecal Recovery of Probiotics Administered as a Multi-Strain Formulation during Antibiotic Treatment.

The present study aimed to investigate whether probiotic recovery is affected when consumed together with antibiotics. Fecal samples were collected from an earlier antibiotic associated diarrhea, randomized, placebo-controlled study with a product consisting of a combination of Lactobacillus acidophilus NCFM, Lactobacillus paracasei Lpc-37, and Bifidobacterium lactis Bi-07, B. lactis Bl-04 at equal numbers and at a total dose of 1010 CFU. Fecal samples were collected during the screening visit (T0), i.e., at the time of antibiotic prescription, and then on the last day of the antibiotic treatment (T1) as well as seven days after the subject had stopped taking the antibiotic treatment (T2) and at two weeks after completing antibiotic treatment and one week after probiotic/placebo consumption stopped (T3). Samples were analyzed for the presence of the four administered strains. The study was registered at clinicaltrials.gov as NCT01596829. Detection levels of all four strains were significantly increased from T0 to T1 and returned to baseline level from T2 to T3. There were also significantly more subjects with detectable levels of L. paracasei Lpc-37, B. lactis Bi-07, and B. lactis Bl-04 at T1 and T2 compared to T0 and T3, and compared to placebo. Each of the four strains could be detected in the feces of patients apparently unaffected by the simultaneous consumption of antibiotics.

Dose-Response Recovery of Probiotic Strains in Simulated Gastro-Intestinal Passage.

The probiotic definition stipulates "adequate amounts". Here, we investigated the metabolic output and recovery rate of probiotic strains using a simulated upper gastro-intestinal passage and colonic fermentation. Two different doses, 7 × 109 colony forming units (CFU) and 7 × 1010 CFU, of a probiotic mixture (Bifidobacterium lactis Bl-04, Lactobacillus acidophilus La-14, Lactobacillus paracasei Lpc-37, and Lactobacillus plantarum Lp-115) or placebo were tested. The four strains were quantified by qPCR and the metabolites analyzed by gas chromatography. There was a dose-response in the detection of all four strains. There was a slightly larger increase between the two doses for L. paracasei Lpc-37 as compared with the other strains; this may suggest a greater robustness of this strain. Compared with the placebo, the high dose simulations generated more propionic acid and a higher total of short chain fatty acids (SCFA). Higher doses of a species are required to reach measurable increases above the baseline level of this species.

Nasal microbiota clusters associate with inflammatory response, viral load, and symptom severity in experimental rhinovirus challenge.

The role of nasal and fecal microbiota in viral respiratory infections has not been established. We collected nasal swabs and washes, and fecal samples in a clinical study assessing the effect of probiotic Bifidobacterium animalis subsp. lactis Bl-04 on experimental rhinovirus infection. The nasal and fecal microbiota were characterized by 16S rRNA gene sequencing. The resulting data were compared with nasal inflammatory marker concentrations, viral load, and clinical symptoms. By using unsupervised clustering, the nasal microbiota divided into six clusters. The clusters predominant of Staphylococcus, Corynebacterium/Alloiococcus, Moraxella, and Pseudomonadaceae/Mixed had characteristic inflammatory marker and viral load profiles in nasal washes. The nasal microbiota clusters of subjects before the infection associated with the severity of clinical cold symptoms during rhinovirus infection. Rhinovirus infection and probiotic intervention did not significantly alter the composition of nasal or fecal microbiota. Our results suggest that nasal microbiota may influence the virus load, host innate immune response, and clinical symptoms during rhinovirus infection, however, further studies are needed.

Metabolic Fate of 13C-Labeled Polydextrose and Impact on the Gut Microbiome: A Triple-Phase Study in a Colon Simulator.

The present study introduces a novel triple-phase (liquids, solids, and gases) approach, which employed uniformly labeled [U-13C] polydextrose (PDX) for the selective profiling of metabolites generated from dietary fiber fermentation in an in vitro colon simulator using human fecal inocula. Employing 13C NMR spectroscopy, [U-13C] PDX metabolism was observed from colonic digest samples. The major 13C-labeled metabolites generated were acetate, butyrate, propionate, and valerate. In addition to these short-chain fatty acids (SCFAs), 13C-labeled lactate, formate, succinate, and ethanol were detected in the colon simulator samples. Metabolite formation and PDX substrate degradation were examined comprehensively over time (24 and 48 h). Correlation analysis between 13C NMR spectra and gas production confirmed the anaerobic fermentation of PDX to SCFAs. In addition, 16S rRNA gene analysis showed that the level of Erysipelotrichaceae was influenced by PDX supplementation and Erysipelotrichaceae level was statistically correlated with SCFA formation. Overall, our study demonstrates a novel approach to link substrate fermentation and microbial function directly in a simulated colonic environment.

Development of dietary soluble fibres by enzymatic synthesis and assessment of their digestibility in in vitro, animal and randomised clinical trial models.

The aim was to develop novel fibres by enzymatic synthesis, to determine their total dietary fibre by AOAC method 2009.01 and to estimate their potential digestibility and assess their digestibility in vivo using glycaemic and insulinaemic responses as markers in mice and randomised clinical trial models. We found that fibre candidates to which α-(1,2) branching was added were resistant to digestion in the mouse model, depending on the amount of branching. These results show that in vivo models are needed to reliably assess the digestibility of α-glycosidic-linked oligomeric dietary fibre candidates, possibly due to absence of brush border α-glucosidase activity in the current in vitro assessment. α-(1,3)-linked and α-(1,6)-linked glucose oligomers were completely digested in humans and mice. In conclusion, it is possible to develop dietary soluble fibres by enzymatic synthesis. Adding α-(1,2) branching increases their resistance to digestion in vivo and can thus improve their suitability as potential fibre candidates. Clinical Trial Registry: ClinicalTrials.gov, NCT02701270.

Independent and Combined Effects of Lactitol, Polydextrose, and Bacteroides thetaiotaomicron on Postprandial Metabolism and Body Weight in Rats Fed a High-Fat Diet.

Obesity is related to the consumption of energy-dense foods in addition to changes in the microbiome where a higher abundance of gut Bacteroidetes can be found in lean subjects or after weight loss. Lactitol, a sweet-tasting sugar alcohol, is a common sugar-replacement in foods. Polydextrose (PDX), a highly branched glucose polymer, is known to reduce energy intake. Here, we test if the combined effects of lactitol or PDX in combination with Bacteroides species will have a beneficial metabolic response in rats fed a high-fat (HF) diet. A total of 175 male Wistar rats were fed either a LF or HF diet. Bacteroides thetaiotaomicron (10(10) bacteria/animal/day) was orally administered with or without lactitol (1.6-2 g/animal/day) or PDX (2 g/animal/day) for 8 days. Postprandial blood samples, cecal digesta, and feces were collected on the last day. Measurements included: body weight, feed consumption, cecal short-chain fatty acids, fecal dry matter and heat value, blood glucose, insulin, triglyceride, and satiety hormone concentrations. Lactitol and PDX decreased the mean body weight when administered with B. thetaiotaomicron or when lactitol was administered alone. Levels of postprandial plasma triglycerides declined with lactitol and PDX when administered with B. thetaiotaomicron. For intestinal hormone release, lactitol - alone or with B. thetaiotaomicron - increased the release of gastrointestinal peptide tyrosine tyrosine (PYY) as well as the area under the curve (AUC) measured for PYY (0-8 h). In addition, levels of insulin AUC (0-8 h) decreased in the lactitol and PDX-supplemented groups. Lactitol and PDX may both provide additional means to regulate postprandial metabolism and weight management, whereas the addition of B. thetaiotaomicron in the tested doses had only minor effects on the measured parameters.

Probiotic approach to prevent antibiotic resistance.

Probiotics are live microorganisms, mainly belonging to the genera Lactobacillus and Bifidobacterium, although also strain of other species are commercialized, that have a beneficial effect on the host. From the perspective of antibiotic use, probiotics have been observed to reduce the risk of certain infectious disease such as certain types of diarrhea and respiratory tract infection. This may be accompanied with a reduced need of antibiotics for secondary infections. Antibiotics tend to be effective against most common diseases, but increasingly resistance is being observed among pathogens. Probiotics are specifically selected to not contribute to the spread of antibiotic resistance and not carry transferable antibiotic resistance. Concomitant use of probiotics with antibiotics has been observed to reduce the incidence, duration and/or severity of antibiotic-associated diarrhea. This contributes to better adherence to the antibiotic prescription and thereby reduces the evolution of resistance. To what extent probiotics directly reduce the spread of antibiotic resistance is still much under investigation; but maintaining a balanced microbiota during antibiotic use may certainly provide opportunities for reducing the spread of resistances. Key messages Probiotics may reduce the risk for certain infectious diseases and thereby reduce the need for antibiotics. Probiotics may reduce the risk for antibiotic-associated diarrhea Probiotics do not contribute to the spread of antibiotic resistance and may even reduce it.

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.