title: Changes in 16s RNA Gene Microbial Community Profiling by Concentration of Prokaryotic DNA.
abstract: Microbial metagenomics are hindered in clinical tissue samples as a result of the large relative amount of human DNA in relation to microbial DNA acting as competitive inhibitors of downstream applications. We evaluated the LOOXSTER® Enrichment Kit to separate eukaryotic and prokaryotic DNA in submucosal intestinal tissue samples having a low microbial biomass and to determine the effects of enrichment on 16s rRNA microbiota sequencing. The enrichment kit reduced the amount of human DNA in the samples 40-70% resulting in a 3.5-fold increase in the number of 16s bacterial gene sequences detected on the Illumina MiSeq platform. This increase was accompanied by the detection of 41 additional bacterial genera and 94 tentative species. The additional bacterial taxa detected accounted for as much as 25% of the total bacterial population that significantly altered the relative prevalence and composition of the intestinal microbiota. The ability to reduce the competitive inhibition created by human DNA and the concentration of bacterial DNA may allow metagenomics to be performed on complex tissues containing a low bacterial biomass.
authors.affiliation: Department of Biological and Physical Sciences, Montana State University-Billings, Billings, Montana, United States.
authors.forename: Angela
authors.initials: A
authors.lastname: Glassing
authors.affiliation: Molecular Research (Mr. DNA), Shallowater, Texas, United States.
authors.forename: Scot E
authors.initials: SE
authors.lastname: Dowd
authors.affiliation: Department of Surgery, University of Louisville, Louisville, Kentucky, United States.
authors.forename: Susan
authors.initials: S
authors.lastname: Galandiuk
authors.affiliation: Department of Surgery, Paul L. Foster School of Medicine, Texas Tech University Health Sciences Center, El Paso, Texas, United States.
authors.forename: Brian
authors.initials: B
authors.lastname: Davis
authors.affiliation: Department of Surgery, University of Louisville, Louisville, Kentucky, United States.
authors.forename: Jeffrey R
authors.initials: JR
authors.lastname: Jorden
authors.affiliation: St. Vincent Healthcare, Sisters of Charity of Leavenworth Health System, Billings, Montana, United States; Department of Biological and Physical Sciences, Montana State University-Billings, Billings, Montana, United States. Electronic address: rodrick.chiodini@msubillings.edu.
authors.forename: Rodrick J
authors.initials: RJ
authors.lastname: Chiodini
mesh_terms: D001419:Bacteria / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D004269:DNA, Bacterial / Q000235:genetics; D006801:Humans; D007082:Ileum / Q000382:microbiology*; D056186:Metagenomics; D064307:Microbiota; D012336:RNA, Ribosomal, 16S / Q000235:genetics*
chemical_list: D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S
keywords: 16s sequencing; Eukaryotic human DNA; Metagenomics; Microbiome; Microbiota; Prokaryotic DNA; Purification; Separation