title: Performance comparison of Illumina and ion torrent next-generation sequencing platforms for 16S rRNA-based bacterial community profiling.
abstract: High-throughput sequencing of the taxonomically informative 16S rRNA gene provides a powerful approach for exploring microbial diversity. Here we compare the performances of two common "benchtop" sequencing platforms, Illumina MiSeq and Ion Torrent Personal Genome Machine (PGM), for bacterial community profiling by 16S rRNA (V1-V2) amplicon sequencing. We benchmarked performance by using a 20-organism mock bacterial community and a collection of primary human specimens. We observed comparatively higher error rates with the Ion Torrent platform and report a pattern of premature sequence truncation specific to semiconductor sequencing. Read truncation was dependent on both the directionality of sequencing and the target species, resulting in organism-specific biases in community profiles. We found that these sequencing artifacts could be minimized by using bidirectional amplicon sequencing and an optimized flow order on the Ion Torrent platform. Results of bacterial community profiling performed on the mock community and a collection of 18 human-derived microbiological specimens were generally in good agreement for both platforms; however, in some cases, results differed significantly. Disparities could be attributed to the failure to generate full-length reads for particular organisms on the Ion Torrent platform, organism-dependent differences in sequence error rates affecting classification of certain species, or some combination of these factors. This study demonstrates the potential for differential bias in bacterial community profiles resulting from the choice of sequencing platform alone.
authors.affiliation: Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA stevesal@uw.edu ngh2@uw.edu.
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Salipante
authors.affiliation: Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA.
authors.forename: Toana
authors.initials: T
authors.lastname: Kawashima
authors.affiliation: Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA.
authors.forename: Christopher
authors.initials: C
authors.lastname: Rosenthal
authors.affiliation: Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA.
authors.forename: Daniel R
authors.initials: DR
authors.lastname: Hoogestraat
authors.affiliation: Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA.
authors.forename: Lisa A
authors.initials: LA
authors.lastname: Cummings
authors.affiliation: Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA.
authors.forename: Dhruba J
authors.initials: DJ
authors.lastname: Sengupta
authors.affiliation: Life Technologies, South San Francisco, California, USA.
authors.forename: Timothy T
authors.initials: TT
authors.lastname: Harkins
authors.affiliation: Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA Department of Microbiology, University of Washington, Seattle, Washington, USA.
authors.forename: Brad T
authors.initials: BT
authors.lastname: Cookson
authors.affiliation: Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA stevesal@uw.edu ngh2@uw.edu.
authors.forename: Noah G
authors.initials: NG
authors.lastname: Hoffman
mesh_terms: D001419:Bacteria / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D001424:Bacterial Infections / Q000382:microbiology*; D004269:DNA, Bacterial / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing / Q000295:instrumentation / Q000379:methods*; D006801:Humans; D012336:RNA, Ribosomal, 16S / Q000235:genetics*
chemical_list: D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S