Prebiotic galacto-oligosaccharides mitigate the adverse effects of iron fortification on the gut microbiome: a randomised controlled study in Kenyan infants.

OBJECTIVE: Iron-containing micronutrient powders (MNPs) reduce anaemia in African infants, but the current high iron dose (12.5 mg/day) may decrease gut Bifidobacteriaceae and Lactobacillaceae, and increase enteropathogens, diarrhoea and respiratory tract infections (RTIs). We evaluated the efficacy and safety of a new MNP formula with prebiotic galacto-oligosaccharides (GOS) combined with a low dose (5 mg/day) of highly bioavailable iron. DESIGN: In a 4-month, controlled, double-blind trial, we randomised Kenyan infants aged 6.5-9.5 months (n=155) to receive daily (1) a MNP without iron (control); (2) the identical MNP but with 5 mg iron (2.5 mg as sodium iron ethylenediaminetetraacetate and 2.5 mg as ferrous fumarate) (Fe group); or (3) the identical MNP as the Fe group but with 7.5 g GOS (FeGOS group). RESULTS: Anaemia decreased by ≈50% in the Fe and FeGOS groups (p<0.001). Compared with the control or FeGOS group, in the Fe group there were (1) lower abundances of Bifidobacterium and Lactobacillus and higher abundances of Clostridiales (p<0.01); (2) higher abundances of virulence and toxin genes (VTGs) of pathogens (p<0.01); (3) higher plasma intestinal fatty acid-binding protein (a biomarker of enterocyte damage) (p<0.05); and (4) a higher incidence of treated RTIs (p<0.05). In contrast, there were no significant differences in these variables comparing the control and FeGOS groups, with the exception that the abundance of VTGs of all pathogens was significantly lower in the FeGOS group compared with the control and Fe groups (p<0.01). CONCLUSION: A MNP containing a low dose of highly bioavailable iron reduces anaemia, and the addition of GOS mitigates most of the adverse effects of iron on the gut microbiome and morbidity in African infants. TRIAL REGISTRATION NUMBER: NCT02118402.

Low dietary iron intake restrains the intestinal inflammatory response and pathology of enteric infection by food-borne bacterial pathogens.

Orally administrated iron is suspected to increase susceptibility to enteric infections among children in infection endemic regions. Here we investigated the effect of dietary iron on the pathology and local immune responses in intestinal infection models. Mice were held on iron-deficient, normal iron, or high iron diets and after 2 weeks they were orally challenged with the pathogen Citrobacter rodentium. Microbiome analysis by pyrosequencing revealed profound iron- and infection-induced shifts in microbiota composition. Fecal levels of the innate defensive molecules and markers of inflammation lipocalin-2 and calprotectin were not influenced by dietary iron intervention alone, but were markedly lower in mice on the iron-deficient diet after infection. Next, mice on the iron-deficient diet tended to gain more weight and to have a lower grade of colon pathology. Furthermore, survival of the nematode Caenorhabditis elegans infected with Salmonella enterica serovar Typhimurium was prolonged after iron deprivation. Together, these data show that iron limitation restricts disease pathology upon bacterial infection. However, our data also showed decreased intestinal inflammatory responses of mice fed on high iron diets. Thus additionally, our study indicates that the effects of iron on processes at the intestinal host-pathogen interface may highly depend on host iron status, immune status, and gut microbiota composition.

Iron fortification adversely affects the gut microbiome, increases pathogen abundance and induces intestinal inflammation in Kenyan infants.

BACKGROUND: In-home iron fortification for infants in developing countries is recommended for control of anaemia, but low absorption typically results in >80% of the iron passing into the colon. Iron is essential for growth and virulence of many pathogenic enterobacteria. We determined the effect of high and low dose in-home iron fortification on the infant gut microbiome and intestinal inflammation. METHODS: We performed two double-blind randomised controlled trials in 6-month-old Kenyan infants (n=115) consuming home-fortified maize porridge daily for 4 months. In the first, infants received a micronutrient powder (MNP) containing 2.5 mg iron as NaFeEDTA or the MNP without iron. In the second, they received a different MNP containing 12.5 mg iron as ferrous fumarate or the MNP without the iron. The primary outcome was gut microbiome composition analysed by 16S pyrosequencing and targeted real-time PCR (qPCR). Secondary outcomes included faecal calprotectin (marker of intestinal inflammation) and incidence of diarrhoea. We analysed the trials separately and combined. RESULTS: At baseline, 63% of the total microbial 16S rRNA could be assigned to Bifidobacteriaceae but there were high prevalences of pathogens, including Salmonella Clostridium difficile, Clostridium perfringens, and pathogenic Escherichia coli. Using pyrosequencing, +FeMNPs increased enterobacteria, particularly Escherichia/Shigella (p=0.048), the enterobacteria/bifidobacteria ratio (p=0.020), and Clostridium (p=0.030). Most of these effects were confirmed using qPCR; for example, +FeMNPs increased pathogenic E. coli strains (p=0.029). +FeMNPs also increased faecal calprotectin (p=0.002). During the trial, 27.3% of infants in +12.5 mgFeMNP required treatment for diarrhoea versus 8.3% in -12.5 mgFeMNP (p=0.092). There were no study-related serious adverse events in either group. CONCLUSIONS: In this setting, provision of iron-containing MNPs to weaning infants adversely affects the gut microbiome, increasing pathogen abundance and causing intestinal inflammation. TRIAL REGISTRATION NUMBER: NCT01111864.

Characterization of Romboutsia ilealis gen. nov., sp. nov., isolated from the gastro-intestinal tract of a rat, and proposal for the reclassification of five closely related members of the genus Clostridium into the genera Romboutsia gen. nov., Intestinibacter gen. nov., Terrisporobacter gen. nov. and Asaccharospora gen. nov.

A Gram-positive staining, rod-shaped, non-motile, spore-forming obligately anaerobic bacterium, designated CRIBT, was isolated from the gastro-intestinal tract of a rat and characterized. The major cellular fatty acids of strain CRIBT were saturated and unsaturated straight-chain C12-C19 fatty acids, with C16:0 being the predominant fatty acid. The polar lipid profile comprised six glycolipids, four phospholipids and one lipid that did not stain with any of the specific spray reagents used. The only quinone was MK-6. The predominating cell-wall sugars were glucose and galactose. The peptidoglycan type of strain CRIBT was A1σ lanthionine-direct. The genomic DNA G+C content of strain CRIBT was 28.1 mol%. On the basis of 16S rRNA gene sequence similarity, strain CRIBT was most closely related to a number of species of the genus Clostridium, including Clostridium lituseburense (97.2%), Clostridium glycolicum (96.2%), Clostridium mayombei (96.2%), Clostridium bartlettii (96.0%) and Clostridium irregulare (95.5%). All these species show very low 16S rRNA gene sequence similarity (<85%) to the type strain of Clostridium butyricum, the type species of the genus Clostridium. DNA-DNA hybridization with closely related reference strains indicated reassociation values below 32%. On the basis of phenotypic and genetic studies, a novel genus, Romboutsia gen. nov., is proposed. The novel isolate CRIBT (=DSM 25109T=NIZO 4048T) is proposed as the type strain of the type species, Romboutsia ilealis gen. nov., sp. nov., of the proposed novel genus. It is proposed that C. lituseburense is transferred to this genus as Romboutsia lituseburensis comb. nov. Furthermore, the reclassification into novel genera is proposed for C. bartlettii, as Intestinibacter bartlettii gen. nov., comb. nov. (type species of the genus), C. glycolicum, as Terrisporobacter glycolicus gen. nov., comb. nov. (type species of the genus), C. mayombei, as Terrisporobacter mayombei gen. nov., comb. nov., and C. irregulare, as Asaccharospora irregularis gen. nov., comb. nov. (type species of the genus), on the basis of additional data collected in this study. In addition, an emendation of the species Peptostreptococcus anaerobius and the order Eubacteriales is provided.

Mothers' Breast Milk Composition and Their Respective Infant's Gut Microbiota Differ between Five Distinct Rural and Urban Regions in Vietnam.

Microbiota colonization and development in early life is impacted by various host intrinsic (genetic) factors, but also diet, lifestyle, as well as environmental and residential factors upon and after birth. To characterize the impact of maternal nutrition and environmental factors on vaginally born infant gut microbiota composition, we performed an observational study in five distinct geographical areas in Vietnam. Fecal samples of infants (around 39 days old) and fecal and breast milk samples of their mothers (around 28 years) were collected. The microbiota composition of all samples was analyzed by 16S rRNA gene Illumina sequencing and a bioinformatics workflow based on QIIME. In addition, various breast milk components were determined. Strong associations between the geographically determined maternal diet and breast milk composition as well as infant fecal microbiota were revealed. Most notable was the association of urban Ha Noi with relatively high abundances of taxa considered pathobionts, such as Klebsiella and Citrobacter, at the expense of Bifidobacterium. Breast milk composition was most distinct in rural Ha Long Bay, characterized by higher concentrations of, e.g., docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), selenium, and vitamin B12, while it was characterized by, e.g., iron, zinc, and α-linolenic acid (ALA) in Ha Noi. Breast milk iron levels were positively associated with infant fecal Klebsiella and negatively with Bifidobacterium, while the EPA and DHA levels were positively associated with Bifidobacterium. In conclusion, differences between five regions in Vietnam with respect to both maternal breast milk and infant gut microbiota composition were revealed, most likely in part due to maternal nutrition. Thus, there could be opportunities to beneficially steer infant microbiota development in a more desired (rural instead of urban) direction through the mother's diet.

Impact of Sleeve Gastrectomy on Fecal Microbiota in Individuals with Morbid Obesity.

BACKGROUND: The intestinal microbiota plays an important role in the etiology of obesity. Sleeve gastrectomy (SG) is a frequently performed and effective therapy for morbid obesity. OBJECTIVE: To investigate the effect of sleeve gastrectomy on the fecal microbiota of individuals with morbid obesity and to examine whether shifts in microbiota composition are associated with markers of inflammation and intestinal barrier function. METHODS: Fecal and blood samples of healthy individuals (n = 27) and morbidly obese individuals pre-SG (n = 24), and at 2 months (n = 13) and 6 months post-SG (n = 9) were collected. The 16SrRNA gene was sequenced to assess microbiota composition. Fecal calprotectin, plasma inflammatory markers and intestinal permeability markers (multi-sugar test) were determined. RESULTS: Fecal microbiota composition between morbidly obese and lean individuals was significantly different. The fecal microbiota composition changed significantly 2 and 6 months post-SG (p = 0.008) compared to pre-SG but not towards a more lean profile. The post-SG microbiota profile was characterized by an increase in facultative anaerobic bacteria, characteristic for the upper gastrointestinal tract. No correlations were found between inflammatory markers, intestinal permeability and microbial profile changes. CONCLUSIONS: Fecal microbiota composition in morbidly obese individuals changed significantly following SG. This change might be explained by functional changes induced by the SG procedure.

Correlation between protection against sepsis by probiotic therapy and stimulation of a novel bacterial phylotype.

Prophylactic probiotic therapy has shown beneficial effects in an experimental rat model for acute pancreatitis on the health status of the animals. Mechanisms by which probiotic therapy interferes with severity of acute pancreatitis and associated sepsis, however, are poorly understood. The aims of this study were to identify the probiotic-induced changes in the gut microbiota and to correlate these changes to disease outcome. Duodenum and ileum samples were obtained from healthy and diseased rats subjected to pancreatitis for 7 days and prophylactically treated with either a multispecies probiotic mixture or a placebo. Intestinal microbiota was characterized by terminal-restriction fragment length polymorphism (T-RFLP) analyses of PCR-amplified 16S rRNA gene fragments. These analyses showed that during acute pancreatitis the host-specific ileal microbiota was replaced by an "acute pancreatitis-associated microbiota." This replacement was not reversed by administration of the probiotic mixture. An increase, however, was observed in the relative abundance of a novel bacterial phylotype most closely related to Clostridium lituseburense and referred to as commensal rat ileum bacterium (CRIB). Specific primers targeting the CRIB 16S rRNA gene sequence were developed to detect this phylotype by quantitative PCR. An ileal abundance of CRIB 16S rRNA genes of more than 7.5% of the total bacterial 16S rRNA gene pool was correlated with reduced duodenal bacterial overgrowth, reduced bacterial translocation to remote organs, improved pancreas pathology, and reduced proinflammatory cytokine levels in plasma. Our current findings and future studies involving this uncharacterized bacterial phylotype will contribute to unraveling one of the potential mechanisms of probiotic therapy.

Intestinal barrier dysfunction in a randomized trial of a specific probiotic composition in acute pancreatitis.

OBJECTIVES: To determine the relation between intestinal barrier dysfunction, bacterial translocation, and clinical outcome in patients with predicted severe acute pancreatitis and the influence of probiotics on these processes. SUMMARY OF BACKGROUND DATA: Randomized, placebo-controlled, multicenter trial on probiotic prophylaxis (Ecologic 641) in patients with predicted severe acute pancreatitis (PROPATRIA). METHODS: Excretion of intestinal fatty acid binding protein (IFABP, a parameter for enterocyte damage), recovery of polyethylene glycols (PEGs, a parameter for intestinal permeability), and excretion of nitric oxide (NOx, a parameter for bacterial translocation) were assessed in urine of 141 patients collected 24 to 48 h after start of probiotic or placebo treatment and 7 days thereafter. RESULTS: IFABP concentrations in the first 72 hours were higher in patients who developed bacteremia (P = 0.03), infected necrosis (P = 0.01), and organ failure (P = 0.008). PEG recovery was higher in patients who developed bacteremia (PEG 4000, P = 0.001), organ failure (PEG 4000, P < 0.0001), or died (PEG 4000, P = 0.009). Probiotic prophylaxis was associated with an increase in IFABP (median 362 vs. 199 pg/mL; P = 0.02), most evidently in patients with organ failure (P = 0.001), and did not influence intestinal permeability. Overall, probiotics decreased NOx (P = 0.05) but, in patients with organ failure, increased NOx (P = 0.001). CONCLUSIONS: Bacteremia, infected necrosis, organ failure, and mortality were all associated with intestinal barrier dysfunction early in the course of acute pancreatitis. Overall, prophylaxis with this specific combination of probiotic strains reduced bacterial translocation, but was associated with increased bacterial translocation and enterocyte damage in patients with organ failure.

Probiotic prophylaxis in predicted severe acute pancreatitis: a randomised, double-blind, placebo-controlled trial.

BACKGROUND: Infectious complications and associated mortality are a major concern in acute pancreatitis. Enteral administration of probiotics could prevent infectious complications, but convincing evidence is scarce. Our aim was to assess the effects of probiotic prophylaxis in patients with predicted severe acute pancreatitis. METHODS: In this multicentre randomised, double-blind, placebo-controlled trial, 298 patients with predicted severe acute pancreatitis (Acute Physiology and Chronic Health Evaluation [APACHE II] score > or =8, Imrie score > or =3, or C-reactive protein >150 mg/L) were randomly assigned within 72 h of onset of symptoms to receive a multispecies probiotic preparation (n=153) or placebo (n=145), administered enterally twice daily for 28 days. The primary endpoint was the composite of infectious complications--ie, infected pancreatic necrosis, bacteraemia, pneumonia, urosepsis, or infected ascites--during admission and 90-day follow-up. Analyses were by intention to treat. This study is registered, number ISRCTN38327949. FINDINGS: One person in each group was excluded from analyses because of incorrect diagnoses of pancreatitis; thus, 152 individuals in the probiotics group and 144 in the placebo group were analysed. Groups were much the same at baseline in terms of patients' characteristics and disease severity. Infectious complications occurred in 46 (30%) patients in the probiotics group and 41 (28%) of those in the placebo group (relative risk 1.06, 95% CI 0.75-1.51). 24 (16%) patients in the probiotics group died, compared with nine (6%) in the placebo group (relative risk 2.53, 95% CI 1.22-5.25). Nine patients in the probiotics group developed bowel ischaemia (eight with fatal outcome), compared with none in the placebo group (p=0.004). INTERPRETATION: In patients with predicted severe acute pancreatitis, probiotic prophylaxis with this combination of probiotic strains did not reduce the risk of infectious complications and was associated with an increased risk of mortality. Probiotic prophylaxis should therefore not be administered in this category of patients.

Haemophilus is overrepresented in the nasopharynx of infants hospitalized with RSV infection and associated with increased viral load and enhanced mucosal CXCL8 responses.

BACKGROUND: While almost all infants are infected with respiratory syncytial virus (RSV) before the age of 2 years, only a small percentage develops severe disease. Previous studies suggest that the nasopharyngeal microbiome affects disease development. We therefore studied the effect of the nasopharyngeal microbiome on viral load and mucosal cytokine responses, two important factors influencing the pathophysiology of RSV disease. To determine the relation between (i) the microbiome of the upper respiratory tract, (ii) viral load, and (iii) host mucosal inflammation during an RSV infection, nasopharyngeal microbiota profiles of RSV infected infants (< 6 months) with different levels of disease severity and age-matched healthy controls were determined by 16S rRNA marker gene sequencing. The viral load was measured using qPCR. Nasopharyngeal CCL5, CXCL10, MMP9, IL6, and CXCL8 levels were determined with ELISA. RESULTS: Viral load in nasopharyngeal aspirates of patients associates significantly to total nasopharyngeal microbiota composition. Healthy infants (n = 21) and RSV patients (n = 54) display very distinct microbial patterns, primarily characterized by a loss in commensals like Veillonella and overrepresentation of opportunistic organisms like Haemophilus and Achromobacter in RSV-infected individuals. Furthermore, nasopharyngeal microbiota profiles are significantly different based on CXCL8 levels. CXCL8 is a chemokine that was previously found to be indicative for disease severity and for which we find Haemophilus abundance as the strongest predictor for CXCL8 levels. CONCLUSIONS: The nasopharyngeal microbiota in young infants with RSV infection is marked by an overrepresentation of the genus Haemophilus. We present that this bacterium is associated with viral load and mucosal CXCL8 responses, both which are involved in RSV disease pathogenesis.

Probiotic and prebiotic influence beyond the intestinal tract.

Probiotics and prebiotics have long been appreciated for their positive influences on gut health. Research on the mechanisms and effects of these agents shows that their impact reaches beyond the intestine. Effects on the microecology and pathology of the oral cavity, stomach, and vaginal tract have been observed. Likely mediated through immune influences, systemic effects such as reduced severity of colds or other respiratory conditions, impact on allergy incidence and symptoms, and reduced absences from work or daycare have also been noted. These observations, among others, suggest a broader spectrum of influence than commonly considered for these unique substances.

Modification of intestinal flora with multispecies probiotics reduces bacterial translocation and improves clinical course in a rat model of acute pancreatitis.

BACKGROUND: Infection of pancreatic necrosis by gut bacteria is a major cause of morbidity and mortality in patients with severe acute pancreatitis. Use of prophylactic antibiotics remains controversial. The aim of this experiment was assess if modification of intestinal flora with specifically designed multispecies probiotics reduces bacterial translocation or improves outcome in a rat model of acute pancreatitis. METHODS: Male Sprague-Dawley rats were allocated into 3 groups: (1) controls (sham-operated, no treatment), (2) pancreatitis and placebo, and (3) pancreatitis and probiotics. Acute pancreatitis was induced by intraductal glycodeoxycholate and intravenous cerulein infusion. Daily probiotics or placebo was administered intragastrically from 5 days prior until 7 days after induction of pancreatitis. Tissue and fluid samples were collected for microbiologic and quantitative real-time PCR analysis of bacterial translocation. RESULTS: Probiotics reduced duodenal bacterial overgrowth of potential pathogens (Log(10) colony-forming units [CFU]/g 5.0 +/- 0.7 [placebo] vs 3.5 +/- 0.3 CFU/g [probiotics], P < .05), resulting in reduced bacterial translocation to extraintestinal sites, including the pancreas (5.38 +/- 1.0 CFU/g [placebo] vs 3.1 +/- 0.5 CFU/g [probiotics], P < .05). Accordingly, health scores were better and late phase mortality was reduced: 27% (4/15, placebo) versus 0% (0/13, probiotics), respectively, P < .05. CONCLUSIONS: This experiment supports the hypothesis that modification of intestinal flora with multispecies probiotics results in reduced bacterial translocation, morbidity, and mortality in the course of experimental acute pancreatitis.

Design of a multispecies probiotic mixture to prevent infectious complications in critically ill patients.

BACKGROUND & AIMS: Although the potential for probiotics is investigated in an increasing variety of diseases, there is little or no consensus regarding the desired probiotic properties for a particular disease in question, nor about the final design of the probiotic. Specific strain selection procedures were undertaken to design a disease-specific multispecies probiotic. METHODS: From a strain collection of 69 different lactic acid bacteria a primary selection was made of 14 strains belonging to different species showing superior survival in a simulated gastrointestinal environment. Functional tests like antimicrobial activity against a range of clinical isolates and cytokine inducing capacity in cultured human peripheral blood mononuclear cells were used to further identify potential strains. RESULTS: Specific strains inhibited growth of clinical isolates whereas others superiorly induced the anti-inflammatory cytokine IL-10. Based on functional tests and general criteria regarding probiotic design and safety, a selection of the following six strains was made (Ecologic 641); Bifidobacterium bifidum, Bifidobacterium infantis, Lactobacillus acidophilus, Lactobacillus casei, Lactobacillus salivarius and Lactococcus lactis. Combination of these strains resulted in a wider antimicrobial spectrum, superior induction of IL-10 and silencing of pro-inflammatory cytokines as compared to the individual components. CONCLUSIONS: Application of strict criteria during the design of a disease-specific probiotic prior to implementation in clinical trials may provide a rational basis for use of probiotics.

Intestinal colonisation patterns in breastfed and formula-fed infants during the first 12 weeks of life reveal sequential microbiota signatures.

The establishment of the infant gut microbiota is a highly dynamic process dependent on extrinsic and intrinsic factors. We characterized the faecal microbiota of 4 breastfed infants and 4 formula-fed infants at 17 consecutive time points during the first 12 weeks of life. Microbiota composition was analysed by a combination of 16S rRNA gene sequencing and quantitative PCR (qPCR). In this dataset, individuality was a major driver of microbiota composition (P = 0.002) and was more pronounced in breastfed infants. A developmental signature could be distinguished, characterized by sequential colonisation of i) intrauterine/vaginal birth associated taxa, ii) skin derived taxa and other typical early colonisers such as Streptococcus and Enterobacteriaceae, iii) domination of Bifidobacteriaceae, and iv) the appearance of adultlike taxa, particularly species associated with Blautia, Eggerthella, and the potential pathobiont Clostridium difficile. Low abundance of potential pathogens was detected by 16S profiling and confirmed by qPCR. Incidence and dominance of skin and breast milk associated microbes were increased in the gut microbiome of breastfed infants compared to formula-fed infants. The approaches in this study indicate that microbiota development of breastfed and formula-fed infants proceeds according to similar developmental stages with microbiota signatures that include stage-specific species.

Gut microbiome in ADHD and its relation to neural reward anticipation.

BACKGROUND: Microorganisms in the human intestine (i.e. the gut microbiome) have an increasingly recognized impact on human health, including brain functioning. Attention-deficit/hyperactivity disorder (ADHD) is a neurodevelopmental disorder associated with abnormalities in dopamine neurotransmission and deficits in reward processing and its underlying neuro-circuitry including the ventral striatum. The microbiome might contribute to ADHD etiology via the gut-brain axis. In this pilot study, we investigated potential differences in the microbiome between ADHD cases and undiagnosed controls, as well as its relation to neural reward processing. METHODS: We used 16S rRNA marker gene sequencing (16S) to identify bacterial taxa and their predicted gene functions in 19 ADHD and 77 control participants. Using functional magnetic resonance imaging (fMRI), we interrogated the effect of observed microbiome differences in neural reward responses in a subset of 28 participants, independent of diagnosis. RESULTS: For the first time, we describe gut microbial makeup of adolescents and adults diagnosed with ADHD. We found that the relative abundance of several bacterial taxa differed between cases and controls, albeit marginally significant. A nominal increase in the Bifidobacterium genus was observed in ADHD cases. In a hypothesis-driven approach, we found that the observed increase was linked to significantly enhanced 16S-based predicted bacterial gene functionality encoding cyclohexadienyl dehydratase in cases relative to controls. This enzyme is involved in the synthesis of phenylalanine, a precursor of dopamine. Increased relative abundance of this functionality was significantly associated with decreased ventral striatal fMRI responses during reward anticipation, independent of ADHD diagnosis and age. CONCLUSIONS: Our results show increases in gut microbiome predicted function of dopamine precursor synthesis between ADHD cases and controls. This increase in microbiome function relates to decreased neural responses to reward anticipation. Decreased neural reward anticipation constitutes one of the hallmarks of ADHD.

Maternal exposure to a Western-style diet causes differences in intestinal microbiota composition and gene expression of suckling mouse pups.

SCOPE: The long-lasting consequences of nutritional programming during the early phase of life have become increasingly evident. The effects of maternal nutrition on the developing intestine are still underexplored. METHODS AND RESULTS: In this study, we observed (1) altered microbiota composition of the colonic luminal content, and (2) differential gene expression in the intestinal wall in 2-week-old mouse pups born from dams exposed to a Western-style (WS) diet during the perinatal period. A sexually dimorphic effect was found for the differentially expressed genes in the offspring of WS diet-exposed dams but no differences between male and female pups were found for the microbiota composition. Integrative analysis of the microbiota and gene expression data revealed that the maternal WS diet independently affected gene expression and microbiota composition. However, the abundance of bacterial families not affected by the WS diet (Bacteroidaceae, Porphyromonadaceae, and Lachnospiraceae) correlated with the expression of genes playing a key role in intestinal development and functioning (e.g. Pitx2 and Ace2). CONCLUSION: Our data reveal that maternal consumption of a WS diet during the perinatal period alters both gene expression and microbiota composition in the intestinal tract of 2-week-old offspring.

Probiotic prophylaxis in patients with predicted severe acute pancreatitis (PROPATRIA): design and rationale of a double-blind, placebo-controlled randomised multicenter trial [ISRCTN38327949].

BACKGROUND: Infectious complications are the major cause of death in acute pancreatitis. Small bowel bacterial overgrowth and subsequent bacterial translocation are held responsible for the vast majority of these infections. Goal of this study is to determine whether selected probiotics are capable of preventing infectious complications without the disadvantages of antibiotic prophylaxis; antibiotic resistance and fungal overgrowth. METHODS/DESIGN: PROPATRIA is a double-blind, placebo-controlled randomised multicenter trial in which 200 patients will be randomly allocated to a multispecies probiotic preparation (Ecologic 641) or placebo. The study is performed in all 8 Dutch University Hospitals and 7 non-University hospitals. The study-product is administered twice daily through a nasojejunal tube for 28 days or until discharge. Patients eligible for randomisation are adult patients with a first onset of predicted severe acute pancreatitis: Imrie criteria 3 or more, CRP 150 mg/L or more, APACHE II score 8 or more. Exclusion criteria are post-ERCP pancreatitis, malignancy, infection/sepsis caused by a second disease, intra-operative diagnosis of pancreatitis and use of probiotics during the study. Administration of the study product is started within 72 hours after onset of abdominal pain. The primary endpoint is the total number of infectious complications. Secondary endpoints are mortality, necrosectomy, antibiotic resistance, hospital stay and adverse events. To demonstrate that probiotic prophylaxis reduces the proportion of patients with infectious complications from 50% to 30%, with alpha 0,05 and power 80%, a total sample size of 200 patients was calculated. CONCLUSION: The PROPATRIA study is aimed to show a reduction in infectious complications due to early enteral use of multispecies probiotics in severe acute pancreatitis.

Sexually dimorphic characteristics of the small intestine and colon of prepubescent C57BL/6 mice.

BACKGROUND: There is increasing appreciation for sexually dimorphic effects, but the molecular mechanisms underlying these effects are only partially understood. In the present study, we explored transcriptomics and epigenetic differences in the small intestine and colon of prepubescent male and female mice. In addition, the microbiota composition of the colonic luminal content has been examined. METHODS: At postnatal day 14, male and female C57BL/6 mice were sacrificed and the small intestine, colon and content of luminal colon were isolated. Gene expression of both segments of the intestine was analysed by microarray analysis. DNA methylation of the promoter regions of selected sexually dimorphic genes was examined by pyrosequencing. Composition of the microbiota was explored by deep sequencing. RESULTS: Sexually dimorphic genes were observed in both segments of the intestine of 2-week-old mouse pups, with a stronger effect in the small intestine. Amongst the total of 349 genes displaying a sexually dimorphic effect in the small intestine and/or colon, several candidates exhibited a previously established function in the intestine (i.e. Nts, Nucb2, Alox5ap and Retnlγ). In addition, differential expression of genes linked to intestinal bowel disease (i.e. Ccr3, Ccl11 and Tnfr) and colorectal cancer development (i.e. Wt1 and Mmp25) was observed between males and females. Amongst the genes displaying significant sexually dimorphic expression, nine genes were histone-modifying enzymes, suggesting that epigenetic mechanisms might be a potential underlying regulatory mechanism. However, our results reveal no significant changes in DNA methylation of analysed CpGs within the selected differentially expressed genes. With respect to the bacterial community composition in the colon, a dominant effect of litter origin was found but no significant sex effect was detected. However, a sex effect on the dominance of specific taxa was observed. CONCLUSIONS: This study reveals molecular dissimilarities between males and females in the small intestine and colon of prepubescent mice, which might underlie differences in physiological functioning and in disease predisposition in the two sexes.

T lymphocytes control microbial composition by regulating the abundance of Vibrio in the zebrafish gut.

Dysbiosis of the intestinal microbial community is considered a risk factor for development of chronic intestinal inflammation as well as other diseases such as diabetes, obesity and even cancer. Study of the innate and adaptive immune pathways controlling bacterial colonization has however proven difficult in rodents, considering the extensive cross-talk between bacteria and innate and adaptive immunity. Here, we used the zebrafish to study innate and adaptive immune processes controlling the microbial community. Zebrafish lack a functional adaptive immune system in the first weeks of life, enabling study of the innate immune system in the absence of adaptive immunity. We show that in wild type zebrafish, the initial lack of adaptive immunity associates with overgrowth of Vibrio species (a group encompassing fish and human pathogens), which is overcome upon adaptive immune development. In Rag1-deficient zebrafish (lacking adaptive immunity) Vibrio abundance remains high, suggesting that adaptive immune processes indeed control Vibrio species. Using cell transfer experiments, we confirm that adoptive transfer of T lymphocytes, but not B lymphocytes into Rag1-deficient recipients suppresses outgrowth of Vibrio. In addition, ex vivo exposure of intestinal T lymphocytes to Rag1-deficient microbiota results in increased interferon-gamma expression by these T lymphocytes, compared to exposure to wild type microbiota. In conclusion, we show that T lymphocytes control microbial composition by effectively suppressing the outgrowth of Vibrio species in the zebrafish intestine.