title: Glycans affect DNA extraction and induce substantial differences in gut metagenomic studies.
abstract: Exopolysaccharides produced by bacterial species and present in feces are extremely inhibitory to DNA restriction and can cause discrepancies in metagenomic studies. We determined the effects of different DNA extraction methods on the apparent composition of the gut microbiota using Illumina MiSeq deep sequencing technology. DNA was extracted from the stool from an obese female using 10 different methods and the choice of DNA extraction method affected the proportional abundance at the phylum level, species richness (Chao index, 227 to 2,714) and diversity (non parametric Shannon, 1.37 to 4.4). Moreover DNA was extracted from stools obtained from 83 different individuals by the fastest extraction assay and by an extraction assay that degradated exopolysaccharides. The fastest extraction method was able to detect 68% to 100% genera and 42% to 95% species whereas the glycan degradation extraction method was able to detect 56% to 93% genera and 25% to 87% species. To allow a good liberation of DNA from exopolysaccharides commonly presented in stools, we recommend the mechanical lysis of stools plus glycan degradation, used here for the first time. Caution must be taken in the interpretation of current metagenomic studies, as the efficiency of DNA extraction varies widely among stool samples.
authors.affiliation: URMITE, UM63, CNRS 7278, IRD 198, Inserm 1095, Aix Marseille Université,13005 Marseille, France.
authors.forename: Emmanouil
authors.initials: E
authors.lastname: Angelakis
authors.affiliation: URMITE, UM63, CNRS 7278, IRD 198, Inserm 1095, Aix Marseille Université,13005 Marseille, France.
authors.forename: Dipankar
authors.initials: D
authors.lastname: Bachar
authors.affiliation: Centre National de la Recherche Scientifique, UMR 7257, Aix-Marseille Université, Marseille, France.
authors.forename: Bernard
authors.initials: B
authors.lastname: Henrissat
authors.affiliation: URMITE, UM63, CNRS 7278, IRD 198, Inserm 1095, Aix Marseille Université,13005 Marseille, France.
authors.forename: Fabrice
authors.initials: F
authors.lastname: Armougom
authors.affiliation: URMITE, UM63, CNRS 7278, IRD 198, Inserm 1095, Aix Marseille Université,13005 Marseille, France.
authors.forename: Gilles
authors.initials: G
authors.lastname: Audoly
authors.affiliation: URMITE, UM63, CNRS 7278, IRD 198, Inserm 1095, Aix Marseille Université,13005 Marseille, France.
authors.forename: Jean-Christophe
authors.initials: JC
authors.lastname: Lagier
authors.affiliation: URMITE, UM63, CNRS 7278, IRD 198, Inserm 1095, Aix Marseille Université,13005 Marseille, France.
authors.forename: Catherine
authors.initials: C
authors.lastname: Robert
authors.affiliation: URMITE, UM63, CNRS 7278, IRD 198, Inserm 1095, Aix Marseille Université,13005 Marseille, France.
authors.forename: Didier
authors.initials: D
authors.lastname: Raoult
mesh_terms: D001419:Bacteria / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification*; D005243:Feces / Q000382:microbiology; D005260:Female; D000069196:Gastrointestinal Microbiome / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D007732:Kwashiorkor / Q000382:microbiology; D056186:Metagenomics; D009765:Obesity / Q000382:microbiology; D011134:Polysaccharides / Q000737:chemistry*; D011135:Polysaccharides, Bacterial / Q000737:chemistry; D011502:Protein-Energy Malnutrition / Q000382:microbiology
chemical_list: D004269:DNA, Bacterial; D011134:Polysaccharides; D011135:Polysaccharides, Bacterial