title: Bacterial and viral identification and differentiation by amplicon sequencing on the MinION nanopore sequencer.
abstract: BACKGROUND
  The MinION™ nanopore sequencer was recently released to a community of alpha-testers for evaluation using a variety of sequencing applications. Recent reports have tested the ability of the MinION™ to act as a whole genome sequencer and have demonstrated that nanopore sequencing has tremendous potential utility. However, the current nanopore technology still has limitations with respect to error-rate, and this is problematic when attempting to assemble whole genomes without secondary rounds of sequencing to correct errors. In this study, we tested the ability of the MinION™ nanopore sequencer to accurately identify and differentiate bacterial and viral samples via directed sequencing of characteristic genes shared broadly across a target clade.
  
  
  RESULTS
  Using a 6 hour sequencing run time, sufficient data were generated to identify an E. coli sample down to the species level from 16S rDNA amplicons. Three poxviruses (cowpox, vaccinia-MVA, and vaccinia-Lister) were identified and differentiated down to the strain level, despite over 98% identity between the vaccinia strains. The ability to differentiate strains by amplicon sequencing on the MinION™ was accomplished despite an observed per-base error rate of approximately 30%.
  
  
  CONCLUSIONS
  While nanopore sequencing, using the MinION™ platform from Oxford Nanopore in particular, continues to mature into a commercially available technology, practical uses are sought for the current versions of the technology. This study offers evidence of the utility of amplicon sequencing by demonstrating that the current versions of MinION™ technology can accurately identify and differentiate both viral and bacterial species present within biological samples via amplicon sequencing.
authors: Edgewood Chemical Biological Center, 5183 Black Hawk Rd Bldg E3150 Rm 324, Aberdeen Proving Ground, MD 21010 USA.
authors: Andy
authors: A
authors: Kilianski
authors: Signature Science, LLC, 8329 N. MoPac Expressway, Austin, TX 78759 USA.
authors: Jamie L
authors: JL
authors: Haas
authors: Edgewood Chemical Biological Center, 5183 Black Hawk Rd Bldg E3150 Rm 324, Aberdeen Proving Ground, MD 21010 USA.
authors: Elizabeth J
authors: EJ
authors: Corriveau
authors: Edgewood Chemical Biological Center, 5183 Black Hawk Rd Bldg E3150 Rm 324, Aberdeen Proving Ground, MD 21010 USA.
authors: Alvin T
authors: AT
authors: Liem
authors: Edgewood Chemical Biological Center, 5183 Black Hawk Rd Bldg E3150 Rm 324, Aberdeen Proving Ground, MD 21010 USA ; Defense Threat Reduction Agency, 8725 John J Kingman Rd Stop 6201, Ft. Belvoir, VA 22060-6201 USA.
authors: Kristen L
authors: KL
authors: Willis
authors: Signature Science, LLC, 8329 N. MoPac Expressway, Austin, TX 78759 USA.
authors: Dana R
authors: DR
authors: Kadavy
authors: Edgewood Chemical Biological Center, 5183 Black Hawk Rd Bldg E3150 Rm 324, Aberdeen Proving Ground, MD 21010 USA.
authors: C Nicole
authors: CN
authors: Rosenzweig
authors: Signature Science, LLC, 8329 N. MoPac Expressway, Austin, TX 78759 USA.
authors: Samuel S
authors: SS
authors: Minot
mesh_terms: D001419:Bacteria / Q000235:genetics*; D002965:Classification / Q000379:methods; D058608:Nanopores; D017422:Sequence Analysis, DNA / Q000379:methods*; D014780:Viruses / Q000235:genetics*
keywords: Amplicon; Escherichia coli; Long-read sequencing; MinION™; NGS; Nanopore; Oxford Nanopore; Pathogen detection; Poxvirus; Whole genome sequencing