title: Species identification and profiling of complex microbial communities using shotgun Illumina sequencing of 16S rRNA amplicon sequences.
abstract: The high throughput and cost-effectiveness afforded by short-read sequencing technologies, in principle, enable researchers to perform 16S rRNA profiling of complex microbial communities at unprecedented depth and resolution. Existing Illumina sequencing protocols are, however, limited by the fraction of the 16S rRNA gene that is interrogated and therefore limit the resolution and quality of the profiling. To address this, we present the design of a novel protocol for shotgun Illumina sequencing of the bacterial 16S rRNA gene, optimized to amplify more than 90% of sequences in the Greengenes database and with the ability to distinguish nearly twice as many species-level OTUs compared to existing protocols. Using several in silico and experimental datasets, we demonstrate that despite the presence of multiple variable and conserved regions, the resulting shotgun sequences can be used to accurately quantify the constituents of complex microbial communities. The reconstruction of a significant fraction of the 16S rRNA gene also enabled high precision (>90%) in species-level identification thereby opening up potential application of this approach for clinical microbial characterization.
authors.affiliation: Genome Institute of Singapore, Genome #02-01, Singapore, Singapore.
authors.forename: Swee Hoe
authors.initials: SH
authors.lastname: Ong
authors.forename: Vinutha Uppoor
authors.initials: VU
authors.lastname: Kukkillaya
authors.forename: Andreas
authors.initials: A
authors.lastname: Wilm
authors.forename: Christophe
authors.initials: C
authors.lastname: Lay
authors.forename: Eliza Xin Pei
authors.initials: EX
authors.lastname: Ho
authors.forename: Louie
authors.initials: L
authors.lastname: Low
authors.forename: Martin Lloyd
authors.initials: ML
authors.lastname: Hibberd
authors.forename: Niranjan
authors.initials: N
authors.lastname: Nagarajan
mesh_terms: D000328:Adult; D001419:Bacteria / Q000145:classification / Q000235:genetics; D002675:Child, Preschool; D019295:Computational Biology / Q000379:methods; D041981:Gastrointestinal Tract / Q000382:microbiology; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D054892:Metagenome; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S
chemical_list: D012336:RNA, Ribosomal, 16S