Quantitative Profiling of Gut Microbiota of Children With Diarrhea-Predominant Irritable Bowel Syndrome

OBJECTIVES:Human intestinal microbiota has a number of important roles in human health and is also implicated in several gastrointestinal disorders. The goal of this study was to determine the gut microbiota in two groups of pre- and adolescent children: healthy volunteers and children diagnosed with diarrhea-predominant irritable bowel syndrome (IBS-D).METHODS:Phylogenetic Microbiota Array was used to obtain quantitative measurements of bacterial presence and abundance in subjects’ fecal samples. We utilized high-throughput DNA sequencing, quantitative PCR, and fluorescent in situ hybridization to confirm microarray findings.RESULTS:Both sample groups were dominated by the phyla Firmicutes, Bacteroidetes, and Actinobacteria, which cumulatively constituted 91% of overall sample composition on average. A core microbiome shared among analyzed samples encompassed 55 bacterial phylotypes dominated by genus Ruminococcus; members of genera Clostridium, Faecalibacterium, Roseburia, Streptococcus, and Bacteroides were also present. Several genera were found to be differentially abundant in the gut of healthy and IBS groups: levels of Veillonella, Prevotella, Lactobacillus, and Parasporobacterium were increased in children diagnosed with IBS, whereas members of Bifidobacterium and Verrucomicrobium were less abundant in those individuals. By calculating a nonparametric correlation matrix among abundances of different genera in all samples, we also examined potential associations among intestinal microbes. Strong positive correlations were found between abundances of Veillonella and both Haemophilus and Streptococcus, between Anaerovorax and Verrucomicrobium, and between Tannerella and Anaerophaga.CONCLUSIONS:Although at the higher taxonomical level gut microbiota was similar between healthy and IBS-D children, specific differences in the abundances of several bacterial genera were revealed. Core microbiome in children was dominated by Clostridia. Putative relationships identified among microbial genera provide testable hypotheses of cross-species associations among members of human gut microbiota.

[1]  J. Corander,et al.  The fecal microbiota of irritable bowel syndrome patients differs significantly from that of healthy subjects. , 2007, Gastroenterology.

[2]  A. Palva,et al.  Analysis of the Fecal Microbiota of Irritable Bowel Syndrome Patients and Healthy Controls with Real-Time PCR , 2005, The American Journal of Gastroenterology.

[3]  R Amann,et al.  Application of a suite of 16S rRNA-specific oligonucleotide probes designed to investigate bacteria of the phylum cytophaga-flavobacter-bacteroides in the natural environment. , 1996, Microbiology.

[4]  O. Paliy,et al.  Application of phylogenetic microarrays to interrogation of human microbiota. , 2012, FEMS microbiology ecology.

[5]  G. Di Nardo,et al.  A distinctive 'microbial signature' in celiac pediatric patients , 2010, BMC Microbiology.

[6]  T. Næs,et al.  Unveiling an abundant core microbiota in the human adult colon by a phylogroup-independent searching approach , 2011, The ISME Journal.

[7]  A. Gasbarrini,et al.  Prevalence of small intestinal bacterial overgrowth in children with irritable bowel syndrome: a case-control study. , 2009, The Journal of pediatrics.

[8]  H. Flint,et al.  Understanding the effects of diet on bacterial metabolism in the large intestine , 2007, Journal of applied microbiology.

[9]  Anne Salonen,et al.  Gastrointestinal microbiota in irritable bowel syndrome: present state and perspectives. , 2010, Microbiology.

[10]  M. Samsom,et al.  Lower Bifidobacteria counts in both duodenal mucosa-associated and fecal microbiota in irritable bowel syndrome patients. , 2009, World journal of gastroenterology.

[11]  F. Bushman,et al.  Linking Long-Term Dietary Patterns with Gut Microbial Enterotypes , 2011, Science.

[12]  E. Zoetendal,et al.  Microarray Analysis and Barcoded Pyrosequencing Provide Consistent Microbial Profiles Depending on the Source of Human Intestinal Samples , 2011, Applied and Environmental Microbiology.

[13]  I. Wiklund,et al.  An irritable bowel syndrome-specific symptom questionnaire: development and validation. , 2003, Scandinavian journal of gastroenterology.

[14]  R. Joerger,et al.  Composition of microbiota in content and mucus from cecae of broiler chickens as measured by fluorescent in situ hybridization with group-specific, 16S rRNA-targeted oligonucleotide probes. , 2003, Poultry science.

[15]  H. Khamis,et al.  Distal gut microbiota of adolescent children is different from that of adults. , 2011, FEMS microbiology ecology.

[16]  Marcus J Claesson,et al.  An irritable bowel syndrome subtype defined by species-specific alterations in faecal microbiota , 2011, Gut.

[17]  B. Foy,et al.  Optimizing the analysis of human intestinal microbiota with phylogenetic microarray. , 2011, FEMS microbiology ecology.

[18]  F. Guarner,et al.  A Global Perspective on Irritable Bowel Syndrome: A Consensus Statement of the World Gastroenterology Organisation Summit Task Force on Irritable Bowel Syndrome , 2012, Journal of clinical gastroenterology.

[19]  R. Sartor,et al.  Luminal and mucosal-associated intestinal microbiota in patients with diarrhea-predominant irritable bowel syndrome , 2010, Gut pathogens.

[20]  N. Pace,et al.  Disease phenotype and genotype are associated with shifts in intestinal‐associated microbiota in inflammatory bowel diseases , 2011, Inflammatory bowel diseases.

[21]  A. H. Stouthamer,et al.  Oxygen metabolism by the anaerobic bacteriumVeillonella alcalescens , 1978, Archives of Microbiology.

[22]  Oliver Grundmann,et al.  Irritable bowel syndrome: Epidemiology, diagnosis and treatment: An update for health‐care practitioners , 2010, Journal of gastroenterology and hepatology.

[23]  F. Leroy,et al.  Cross-feeding between bifidobacteria and butyrate-producing colon bacteria explains bifdobacterial competitiveness, butyrate production, and gas production. , 2011, International journal of food microbiology.

[24]  S. Fukudo,et al.  Altered profiles of intestinal microbiota and organic acids may be the origin of symptoms in irritable bowel syndrome , 2009, Neurogastroenterology and motility : the official journal of the European Gastrointestinal Motility Society.

[25]  J. Conklin,et al.  The effects of methane and hydrogen gases produced by enteric bacteria on ileal motility and colonic transit time , 2012, Neurogastroenterology and motility : the official journal of the European Gastrointestinal Motility Society.

[26]  J. Hyams,et al.  Abdominal pain and irritable bowel syndrome in adolescents: a community-based study. , 1996, The Journal of pediatrics.

[27]  J. Doré,et al.  Faecalibacterium prausnitzii is an anti-inflammatory commensal bacterium identified by gut microbiota analysis of Crohn disease patients , 2008, Proceedings of the National Academy of Sciences.

[28]  R. Knight,et al.  Fast UniFrac: Facilitating high-throughput phylogenetic analyses of microbial communities including analysis of pyrosequencing and PhyloChip data , 2009, The ISME Journal.

[29]  B. Flourié,et al.  Bacterial populations contaminating the upper gut in patients with small intestinal bacterial overgrowth syndrome , 1999, American Journal of Gastroenterology.

[30]  Aleksandar Milosavljevic,et al.  Gastrointestinal microbiome signatures of pediatric patients with irritable bowel syndrome. , 2011, Gastroenterology.

[31]  R. McCallum,et al.  Results of small intestinal bacterial overgrowth testing in irritable bowel syndrome patients: clinical profiles and effects of antibiotic trial. , 2007, Advances in Medical Sciences.

[32]  R. Knight,et al.  Bacterial Community Variation in Human Body Habitats Across Space and Time , 2009, Science.

[33]  J. Nikkilä,et al.  Diarrhoea-predominant irritable bowel syndrome distinguishable by 16S rRNA gene phylotype quantification. , 2009, World journal of gastroenterology.

[34]  J. Frick,et al.  Microbiota in pediatric inflammatory bowel disease. , 2010, The Journal of pediatrics.

[35]  K. L. Johnston,et al.  Small intestinal bacterial overgrowth. , 1999, The Veterinary clinics of North America. Small animal practice.

[36]  F. Shanahan,et al.  A Molecular Analysis of Fecal and Mucosal Bacterial Communities in Irritable Bowel Syndrome , 2010, Digestive Diseases and Sciences.

[37]  Robert J. Palmer,et al.  Characterization of a Streptococcus sp.-Veillonella sp. Community Micromanipulated from Dental Plaque , 2008, Journal of bacteriology.

[38]  Igor Jurisica,et al.  NAViGaTOR: Network Analysis, Visualization and Graphing Toronto , 2009, Bioinform..

[39]  R. Satokari,et al.  Prevalence and temporal stability of selected clostridial groups in irritable bowel syndrome in relation to predominant faecal bacteria. , 2006, Journal of medical microbiology.

[40]  L. Walker,et al.  Validation of the Pediatric Rome II Criteria for Functional Gastrointestinal Disorders Using the Questionnaire on Pediatric Gastrointestinal Symptoms , 2005, Journal of pediatric gastroenterology and nutrition.

[41]  장재훈,et al.  The effects of methane and hydrogen gases produced by enteric bacteria on ileal motility and colonic transit time , 2012 .

[42]  Mario Vaneechoutte,et al.  Active Crohn's disease and ulcerative colitis can be specifically diagnosed and monitored based on the biostructure of the fecal flora , 2008, Inflammatory bowel diseases.

[43]  M. Icaza-Chávez,et al.  Gut microbiota in health and disease , 2013 .

[44]  Christophe Caron,et al.  Towards the human intestinal microbiota phylogenetic core. , 2009, Environmental microbiology.

[45]  E. Björnsson,et al.  Small intestinal bacterial overgrowth in patients with irritable bowel syndrome , 2006, Gut.

[46]  L. Paulin,et al.  Multilayered epithelium in a rat model and human Barrett's esophagus: Similar expression patterns of transcription factors and differentiation markers , 2008, BMC gastroenterology.

[47]  R. McCallum,et al.  Small intestinal bacterial overgrowth in irritable bowel syndrome: are there any predictors? , 2010, BMC gastroenterology.

[48]  R. Williams,et al.  The acid end-products of glucose metabolism of oral and other haemophili. , 1984, Journal of general microbiology.

[49]  Lynne A. Goodwin,et al.  Complete genome sequence of Veillonella parvula type strain (Te3T) , 2010, Standards in genomic sciences.

[50]  W. D. de Vos,et al.  Intestinal Microbiota in Healthy Adults: Temporal Analysis Reveals Individual and Common Core and Relation to Intestinal Symptoms , 2011, PloS one.

[51]  M. Pop,et al.  Metagenomic Analysis of the Human Distal Gut Microbiome , 2006, Science.

[52]  Henry C. Lin,et al.  Small intestinal bacterial overgrowth: a framework for understanding irritable bowel syndrome. , 2004, JAMA.

[53]  William A. Walters,et al.  QIIME allows analysis of high-throughput community sequencing data , 2010, Nature Methods.

[54]  W. Whitman,et al.  Prokaryotes: the unseen majority. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[55]  Michiel Kleerebezem,et al.  High temporal and inter-individual variation detected in the human ileal microbiota. , 2010, Environmental microbiology.

[56]  A. Palva,et al.  Composition and temporal stability of gastrointestinal microbiota in irritable bowel syndrome--a longitudinal study in IBS and control subjects. , 2005, FEMS immunology and medical microbiology.

[57]  O. Paliy,et al.  High-Throughput Quantitative Analysis of the Human Intestinal Microbiota with a Phylogenetic Microarray , 2009, Applied and Environmental Microbiology.

[58]  A. Caminero,et al.  Differences of small intestinal bacteria populations in adults and children with/without celiac disease: Effect of age, gluten diet, and disease , 2012, Inflammatory bowel diseases.