title: Collecting Fecal Samples for Microbiome Analyses in Epidemiology Studies.
abstract: BACKGROUND
  The need to develop valid methods for sampling and analyzing fecal specimens for microbiome studies is increasingly important, especially for large population studies.
  
  
  METHODS
  Some of the most important attributes of any sampling method are reproducibility, stability, and accuracy. We compared seven fecal sampling methods [no additive, RNAlater, 70% ethanol, EDTA, dry swab, and pre/post development fecal occult blood test (FOBT)] using 16S rRNA microbiome profiling in two laboratories. We evaluated nine commonly used microbiome metrics: abundance of three phyla, two alpha-diversities, and four beta-diversities. We determined the technical reproducibility, stability at ambient temperature, and accuracy.
  
  
  RESULTS
  Although microbiome profiles showed systematic biases according to sample method and time at ambient temperature, the highest source of variation was between individuals. All collection methods showed high reproducibility. FOBT and RNAlater resulted in the highest stability without freezing for 4 days. In comparison with no-additive samples, swab, FOBT, and 70% ethanol exhibited the greatest accuracy when immediately frozen.
  
  
  CONCLUSIONS
  Overall, optimal stability and reproducibility were achieved using FOBT, making this a reasonable sample collection method for 16S analysis.
  
  
  CONCLUSIONS
  Having standardized method of collecting and storing stable fecal samples will allow future investigations into the role of gut microbiota in chronic disease etiology in large population studies.
authors.affiliation: Nutritional Epidemiology Branch, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, Maryland. sinhar@exchange.nih.gov chia.nicholas@mayo.edu.
authors.forename: Rashmi
authors.initials: R
authors.lastname: Sinha
authors.affiliation: Microbiome Program, Center for Individualized Medicine, Mayo Clinic, Rochester, Minnesota. Health Sciences Research, Mayo Clinic, Rochester, Minnesota.
authors.forename: Jun
authors.initials: J
authors.lastname: Chen
authors.affiliation: Department of Pediatrics, University of California San Diego, San Diego, California.
authors.forename: Amnon
authors.initials: A
authors.lastname: Amir
authors.affiliation: Nutritional Epidemiology Branch, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, Maryland. Cancer Prevention Fellowship Program, Division of Cancer Prevention, National Cancer Institute, National Institutes of Health, Bethesda, Maryland.
authors.forename: Emily
authors.initials: E
authors.lastname: Vogtmann
authors.affiliation: Biostatistics Branch, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, Maryland.
authors.forename: Jianxin
authors.initials: J
authors.lastname: Shi
authors.affiliation: Department of Cancer Biology, Mayo Clinic, Jacksonville, Florida.
authors.forename: Kristin S
authors.initials: KS
authors.lastname: Inman
authors.affiliation: Nutritional Science Research Group, Division of Cancer Prevention, National Cancer Institute, National Institutes of Health, Bethesda, Maryland.
authors.forename: Roberto
authors.initials: R
authors.lastname: Flores
authors.affiliation: Biostatistics Branch, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, Maryland.
authors.forename: Joshua
authors.initials: J
authors.lastname: Sampson
authors.affiliation: Department of Pediatrics, University of California San Diego, San Diego, California. Department of Computer Science and Engineering, University of California San Diego, San Diego, California.
authors.forename: Rob
authors.initials: R
authors.lastname: Knight
authors.affiliation: Microbiome Program, Center for Individualized Medicine, Mayo Clinic, Rochester, Minnesota. Health Sciences Research, Mayo Clinic, Rochester, Minnesota. Department of Surgery, Mayo Clinic, Rochester, Minnesota. Biomedical Engineering and Physiology, Mayo College, Rochester, Minnesota. sinhar@exchange.nih.gov chia.nicholas@mayo.edu.
authors.forename: Nicholas
authors.initials: N
authors.lastname: Chia
mesh_terms: D000328:Adult; D016021:Epidemiologic Studies; D005243:Feces / Q000382:microbiology*; D005260:Female; D006801:Humans; D008297:Male; D064307:Microbiota; D008875:Middle Aged; D055815:Young Adult