title: Development of a standardized approach for environmental microbiota investigations related to asthma development in children.
abstract: Standardized studies examining environmental microbial exposure in populations at risk for asthma are necessary to improve our understanding of the role this factor plays in disease development. Here we describe studies aimed at developing guidelines for high-resolution culture-independent microbiome profiling, using a phylogenetic microarray (PhyloChip), of house dust samples in a cohort collected as part of the NIH-funded Inner City Asthma Consortium (ICAC). We demonstrate that though extracted DNA concentrations varied across dust samples, the majority produced sufficient 16S rRNA to be profiled by the array. Comparison of array and 454-pyrosequencing performed in parallel on a subset of samples, illustrated that increasingly deeper sequencing efforts validated greater numbers of array-detected taxa. Community composition agreement across samples exhibited a hierarchy in concordance, with the highest level of agreement in replicate array profiles followed by samples collected from adjacent 1×1 m(2) sites in the same room, adjacent sites with different sized sampling quadrants (1×1 and 2×2 m(2)), different sites within homes (living and bedroom) to lowest in living room samples collected from different homes. The guidelines for sample collection and processing in this pilot study extend beyond PhyloChip based studies of house-associated microbiota, and bear relevance for other microbiome profiling approaches such as next-generation sequencing.
authors.affiliation: Division of Gastroenterology, University of California, San Francisco, CA 94143, United States.
authors.forename: Kei E
authors.initials: KE
authors.lastname: Fujimura
authors.forename: Marcus
authors.initials: M
authors.lastname: Rauch
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Matsui
authors.forename: Shoko
authors.initials: S
authors.lastname: Iwai
authors.forename: Agustin
authors.initials: A
authors.lastname: Calatroni
authors.forename: Henry
authors.initials: H
authors.lastname: Lynn
authors.forename: Herman
authors.initials: H
authors.lastname: Mitchell
authors.forename: Christine C
authors.initials: CC
authors.lastname: Johnson
authors.forename: James E
authors.initials: JE
authors.lastname: Gern
authors.forename: Alkis
authors.initials: A
authors.lastname: Togias
authors.forename: Homer A
authors.initials: HA
authors.lastname: Boushey
authors.forename: Suzanne
authors.initials: S
authors.lastname: Kennedy
authors.forename: Susan V
authors.initials: SV
authors.lastname: Lynch
mesh_terms: D001249:Asthma / Q000209:etiology*; D058448:Biota; D002648:Child; D004391:Dust; D004783:Environmental Microbiology; D006801:Humans; D054892:Metagenome; D056186:Metagenomics / Q000379:methods* / Q000592:standards*; D046228:Microarray Analysis / Q000379:methods / Q000592:standards; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D017422:Sequence Analysis, DNA
chemical_list: D004391:Dust; D012336:RNA, Ribosomal, 16S