title: Duodenal and faecal microbiota of celiac children: molecular, phenotype and metabolome characterization.
abstract: BACKGROUND
  Epidemiology of celiac disease (CD) is increasing. CD mainly presents in early childhood with small intestinal villous atrophy and signs of malabsorption. Compared to healthy individuals, CD patients seemed to be characterized by higher numbers of Gram-negative bacteria and lower numbers Gram-positive bacteria.
  
  
  RESULTS
  This study aimed at investigating the microbiota and metabolome of 19 celiac disease children under gluten-free diet (treated celiac disease, T-CD) and 15 non-celiac children (HC). PCR-denaturing gradient gel electrophoresis (DGGE) analyses by universal and group-specific primers were carried out in duodenal biopsies and faecal samples. Based on the number of PCR-DGGE bands, the diversity of Eubacteria was the higher in duodenal biopsies of T-CD than HC children. Bifidobacteria were only found in faecal samples. With a few exceptions, PCR-DGGE profiles of faecal samples for Lactobacillus and Bifidobacteria differed between T-CD and HC. As shown by culture-dependent methods, the levels of Lactobacillus, Enterococcus and Bifidobacteria were confirmed to be significantly higher (P = 0.028; P = 0.019; and P = 0.023, respectively) in fecal samples of HC than in T-CD children. On the contrary, cell counts (CFU/ml) of presumptive Bacteroides, Staphylococcus, Salmonella, Shighella and Klebsiella were significantly higher (P = 0.014) in T-CD compared to HC children. Enterococcus faecium and Lactobacillus plantarum were the species most diffusely identified. This latter species was also found in all duodenal biopsies of T-CD and HC children. Other bacterial species were identified only in T-CD or HC faecal samples. As shown by Randomly Amplified Polymorphic DNA-PCR analysis, the percentage of strains identified as lactobacilli significantly (P = 0.011) differed between T-CD (ca. 26.5%) and HC (ca. 34.6%) groups. The metabolome of T-CD and HC children was studied using faecal and urine samples which were analyzed by gas-chromatography mass spectrometry-solid-phase microextraction and 1H-Nuclear Magnetic Resonance. As shown by Canonical Discriminant Analysis of Principal Coordinates, the levels of volatile organic compounds and free amino acids in faecal and/or urine samples were markedly affected by CD.
  
  
  CONCLUSIONS
  As shown by the parallel microbiology and metabolome approach, the gluten-free diet lasting at least two years did not completely restore the microbiota and, consequently, the metabolome of CD children. Some molecules (e.g., ethyl-acetate and octyl-acetate, some short chain fatty acids and free amino acids, and glutamine) seems to be metabolic signatures of CD.
authors.affiliation: Department of Biologia e Chimica Agro-Forestale ed Ambientale, University of Bari Aldo Moro, via Amendola 165/A, Bari, 70126 Italy.
authors.forename: Raffaella
authors.initials: R
authors.lastname: Di Cagno
authors.forename: Maria
authors.initials: M
authors.lastname: De Angelis
authors.forename: Ilaria
authors.initials: I
authors.lastname: De Pasquale
authors.forename: Maurice
authors.initials: M
authors.lastname: Ndagijimana
authors.forename: Pamela
authors.initials: P
authors.lastname: Vernocchi
authors.forename: Patrizia
authors.initials: P
authors.lastname: Ricciuti
authors.forename: Francesca
authors.initials: F
authors.lastname: Gagliardi
authors.forename: Luca
authors.initials: L
authors.lastname: Laghi
authors.forename: Carmine
authors.initials: C
authors.lastname: Crecchio
authors.forename: Maria Elisabetta
authors.initials: ME
authors.lastname: Guerzoni
authors.forename: Marco
authors.initials: M
authors.lastname: Gobbetti
authors.forename: Ruggiero
authors.initials: R
authors.lastname: Francavilla
mesh_terms: D001419:Bacteria / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism; D016022:Case-Control Studies; D002446:Celiac Disease / Q000378:metabolism* / Q000382:microbiology*; D002648:Child; D002675:Child, Preschool; D004386:Duodenum / Q000382:microbiology*; D005243:Feces / Q000737:chemistry / Q000382:microbiology*; D005260:Female; D006801:Humans; D008297:Male; D055442:Metabolome; D054892:Metagenome; D010641:Phenotype; D010802:Phylogeny; D014556:Urine / Q000737:chemistry; D055549:Volatile Organic Compounds / Q000032:analysis / Q000378:metabolism
chemical_list: D055549:Volatile Organic Compounds