title: Evaluating the Impact of DNA Extraction Method on the Representation of Human Oral Bacterial and Fungal Communities.
abstract: The application of high-throughput, next-generation sequencing technologies has greatly improved our understanding of the human oral microbiome. While deciphering this diverse microbial community using such approaches is more accurate than traditional culture-based methods, experimental bias introduced during critical steps such as DNA extraction may compromise the results obtained. Here, we systematically evaluate four commonly used microbial DNA extraction methods (MoBio PowerSoil® DNA Isolation Kit, QIAamp® DNA Mini Kit, Zymo Bacterial/Fungal DNA Mini PrepTM, phenol:chloroform-based DNA isolation) based on the following criteria: DNA quality and yield, and microbial community structure based on Illumina amplicon sequencing of the V3-V4 region of the 16S rRNA gene of bacteria and the internal transcribed spacer (ITS) 1 region of fungi. Our results indicate that DNA quality and yield varied significantly with DNA extraction method. Representation of bacterial genera in plaque and saliva samples did not significantly differ across DNA extraction methods and DNA extraction method showed no effect on the recovery of fungal genera from plaque. By contrast, fungal diversity from saliva was affected by DNA extraction method, suggesting that not all protocols are suitable to study the salivary mycobiome.
authors.affiliation: Department of Surgery, The University of Auckland, Auckland, New Zealand.
authors.forename: Anna
authors.identifier: http://orcid.org/0000-0003-1600-5802
authors.initials: A
authors.lastname: Vesty
authors.affiliation: Department of Surgery, The University of Auckland, Auckland, New Zealand.
authors.forename: Kristi
authors.initials: K
authors.lastname: Biswas
authors.affiliation: School of Biological Sciences, The University of Auckland, Auckland, New Zealand.
authors.forename: Michael W
authors.initials: MW
authors.lastname: Taylor
authors.affiliation: Oral Health Unit, Green Lane Clinical Centre, Auckland, New Zealand.
authors.forename: Kim
authors.initials: K
authors.lastname: Gear
authors.affiliation: Department of Surgery, The University of Auckland, Auckland, New Zealand.
authors.forename: Richard G
authors.identifier: http://orcid.org/0000-0002-2900-0384
authors.initials: RG
authors.lastname: Douglas
mesh_terms: D044822:Biodiversity; D005591:Chemical Fractionation / Q000379:methods; D004269:DNA, Bacterial / Q000302:isolation & purification*; D004271:DNA, Fungal / Q000302:isolation & purification*; D006801:Humans; D056186:Metagenomics / Q000379:methods*; D064307:Microbiota; D009055:Mouth / Q000382:microbiology*; D012336:RNA, Ribosomal, 16S; D012463:Saliva / Q000382:microbiology
chemical_list: D004269:DNA, Bacterial; D004271:DNA, Fungal; D012336:RNA, Ribosomal, 16S