title: Resolving the Complexity of Human Skin Metagenomes Using Single-Molecule Sequencing.
abstract: Deep metagenomic shotgun sequencing has emerged as a powerful tool to interrogate composition and function of complex microbial communities. Computational approaches to assemble genome fragments have been demonstrated to be an effective tool for de novo reconstruction of genomes from these communities. However, the resultant "genomes" are typically fragmented and incomplete due to the limited ability of short-read sequence data to assemble complex or low-coverage regions. Here, we use single-molecule, real-time (SMRT) sequencing to reconstruct a high-quality, closed genome of a previously uncharacterized Corynebacterium simulans and its companion bacteriophage from a skin metagenomic sample. Considerable improvement in assembly quality occurs in hybrid approaches incorporating short-read data, with even relatively small amounts of long-read data being sufficient to improve metagenome reconstruction. Using short-read data to evaluate strain variation of this C. simulans in its skin community at single-nucleotide resolution, we observed a dominant C. simulans strain with moderate allelic heterozygosity throughout the population. We demonstrate the utility of SMRT sequencing and hybrid approaches in metagenome quantitation, reconstruction, and annotation.
  
  
  OBJECTIVE
  The species comprising a microbial community are often difficult to deconvolute due to technical limitations inherent to most short-read sequencing technologies. Here, we leverage new advances in sequencing technology, single-molecule sequencing, to significantly improve reconstruction of a complex human skin microbial community. With this long-read technology, we were able to reconstruct and annotate a closed, high-quality genome of a previously uncharacterized skin species. We demonstrate that hybrid approaches with short-read technology are sufficiently powerful to reconstruct even single-nucleotide polymorphism level variation of species in this a community.
authors: Pacific Biosciences, Menlo Park, California, USA.
authors: Yu-Chih
authors: YC
authors: Tsai
authors: Translational and Functional Genomics Branch, National Human Genome Research Institute, Bethesda, Maryland, USA.
authors: Sean
authors: http://orcid.org/0000-0001-6848-3465
authors: S
authors: Conlan
authors: Translational and Functional Genomics Branch, National Human Genome Research Institute, Bethesda, Maryland, USA.
authors: Clayton
authors: C
authors: Deming
authors: Translational and Functional Genomics Branch, National Human Genome Research Institute, Bethesda, Maryland, USA.
authors: Julia A
authors: JA
authors: Segre
authors: Dermatology Branch, Center for Cancer Research, National Cancer Institute, Bethesda, Maryland, USA.
authors: Heidi H
authors: HH
authors: Kong
authors: Pacific Biosciences, Menlo Park, California, USA.
authors: Jonas
authors: J
authors: Korlach
authors: Translational and Functional Genomics Branch, National Human Genome Research Institute, Bethesda, Maryland, USA julia.oh@jax.org.
authors: Julia
authors: J
authors: Oh
mesh_terms: D001435:Bacteriophages / Q000235:genetics* / Q000302:isolation & purification; D003352:Corynebacterium / Q000235:genetics* / Q000302:isolation & purification / Q000821:virology*; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods; D006801:Humans; D056186:Metagenomics / Q000379:methods*; D064307:Microbiota; D008969:Molecular Sequence Data; D017422:Sequence Analysis, DNA / Q000379:methods; D012867:Skin / Q000382:microbiology*