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Title: Storage and handling of human faecal samples affect the gut microbiome composition: A feasibility study
Contributor(s): Ezzy, Alan C  (author); Hagstrom, Amanda D  (author)orcid ; George, Chris (author); Hamlin, Adam S  (author)orcid ; Pereg, Lily  (author); Murphy, Aron J  (author); Winter, Gal  (author)orcid 
Publication Date: 2019-09
DOI: 10.1016/j.mimet.2019.105668
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Abstract: Human gut microbiome analysis through faecal sampling typically involves five stages: sample collection, storage, DNA extraction, next generation sequencing and bioinformatics analysis. Of these, the first three are considered irreversible. This feasibility study describes an assessment of methodologies used for faecal DNA extraction and sample handling, using the parameters DNA yield, purity and resultant microbial profile. Six DNA extraction techniques, including commercially available kits and manual protocols were compared on human faecal samples (n = 3). Different extraction techniques produced significant variance in DNA yield (range 2.7-164 ng/mg faeces) and microbial diversity profiles, with considerable variation in phyla dominance (Firmicutes (P < 0.001), Bacteroidetes (P = 0.003), Actinobacteria (P = 0.003), One-way ANOVA). The most effective method, with the highest DNA yield, was a simple and inexpensive extraction technique named MetaHIT. Using this method, DNA was extracted from separate faecal samples (n = 3) and had been aliquoted to seven storage conditions including three stabilizing buffers and three temperature conditions, for a period of 120-h, with storage at -80 °C as a control treatment. DNA yield and purity was not statistically different between the control and remaining treatments. 16S rDNA-based diversity profile was largely comparable across the treatments with only minor differences in genera between samples stored at room temperature in air and - 80 °C control. Overall these results suggest that the choice of DNA extraction method has a greater influence on the resultant microbial diversity profile than the short-term storage method.
Publication Type: Journal Article
Source of Publication: Journal of Microbiological Methods, v.164, p. 1-9
Publisher: Elsevier BV
Place of Publication: Netherlands
ISSN: 1872-8359
Fields of Research (FoR) 2008: 060599 Microbiology not elsewhere classified
Fields of Research (FoR) 2020: 320803 Systems physiology
Socio-Economic Objective (SEO) 2008: 970106 Expanding Knowledge in the Biological Sciences
Socio-Economic Objective (SEO) 2020: 200105 Treatment of human diseases and conditions
Peer Reviewed: Yes
HERDC Category Description: C1 Refereed Article in a Scholarly Journal
Appears in Collections:Journal Article
School of Science and Technology

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