MinION nanopore sequencing identifies the position and structure of a bacterial antibiotic resistance island

Short-read, high-throughput sequencing technology cannot identify the chromosomal position of repetitive insertion sequences that typically flank horizontally acquired genes such as bacterial virulence genes and antibiotic resistance genes. The MinION nanopore sequencer can produce long sequencing reads on a device similar in size to a USB memory stick. Here we apply a MinION sequencer to resolve the structure and chromosomal insertion site of a composite antibiotic resistance island in Salmonella Typhi Haplotype 58. Nanopore sequencing data from a single 18-h run was used to create a scaffold for an assembly generated from short-read Illumina data. Our results demonstrate the potential of the MinION device in clinical laboratories to fully characterize the epidemic spread of bacterial pathogens.

[1]  B. R. Callow A new phage-typing scheme for Salmonella typhi-murium , 1959, Journal of Hygiene.

[2]  P Green,et al.  Base-calling of automated sequencer traces using phred. II. Error probabilities. , 1998, Genome research.

[3]  P. Green,et al.  Base-calling of automated sequencer traces using phred. I. Accuracy assessment. , 1998, Genome research.

[4]  Kim Rutherford,et al.  Complete genome sequence of a multiple drug resistant Salmonella enterica serovar Typhi CT18 , 2001, Nature.

[5]  J. Wain,et al.  The role of prophage-like elements in the diversity of Salmonella enterica serovars. , 2004, Journal of molecular biology.

[6]  A. Pühler,et al.  The 120 592 bp IncF plasmid pRSB107 isolated from a sewage-treatment plant encodes nine different antibiotic-resistance determinants, two iron-acquisition systems and other putative virulence-associated functions. , 2005, Microbiology.

[7]  Mark Achtman,et al.  Evolutionary History of Salmonella Typhi , 2006, Science.

[8]  P. Roumagnac,et al.  Clonal Expansion and Microevolution of Quinolone-Resistant Salmonella enterica Serotype Typhi in Vietnam from 1996 to 2004 , 2007, Journal of Clinical Microbiology.

[9]  S. Nair,et al.  Variation in Salmonella enterica Serovar Typhi IncHI1 Plasmids during the Global Spread of Resistant Typhoid Fever , 2008, Antimicrobial Agents and Chemotherapy.

[10]  Julian Parkhill,et al.  Pseudogene accumulation in the evolutionary histories of Salmonella enterica serovars Paratyphi A and Typhi , 2009, BMC Genomics.

[11]  J. Wain,et al.  High-throughput sequencing provides insights into genome variation and evolution in Salmonella Typhi , 2008, Nature Genetics.

[12]  Paul Horton,et al.  Parameters for accurate genome alignment , 2010, BMC Bioinformatics.

[13]  H. Bayley,et al.  Continuous base identification for single-molecule nanopore DNA sequencing. , 2009, Nature nanotechnology.

[14]  Bartek Wilczynski,et al.  Biopython: freely available Python tools for computational molecular biology and bioinformatics , 2009, Bioinform..

[15]  Y. Teo,et al.  Typhoid in Kenya Is Associated with a Dominant Multidrug-Resistant Salmonellaenterica Serovar Typhi Haplotype That Is Also Widespread in Southeast Asia , 2010, Journal of Clinical Microbiology.

[16]  Andrew C. Adey,et al.  Rapid, low-input, low-bias construction of shotgun fragment libraries by high-density in vitro transposition , 2010, Genome Biology.

[17]  B. H. Manh,et al.  Temporal Fluctuation of Multidrug Resistant Salmonella Typhi Haplotypes in the Mekong River Delta Region of Vietnam , 2011, PLoS neglected tropical diseases.

[18]  S. Nair,et al.  Emergence of a Globally Dominant IncHI1 Plasmid Type Associated with Multiple Drug Resistant Typhoid , 2011, PLoS neglected tropical diseases.

[19]  G. Dougan,et al.  High-Resolution Genotyping of the Endemic Salmonella Typhi Population during a Vi (Typhoid) Vaccination Trial in Kolkata , 2012, PLoS neglected tropical diseases.

[20]  Sergey I. Nikolenko,et al.  SPAdes: A New Genome Assembly Algorithm and Its Applications to Single-Cell Sequencing , 2012, J. Comput. Biol..

[21]  T. Dallman,et al.  Public Health Value of Next-Generation DNA Sequencing of Enterohemorrhagic Escherichia coli Isolates from an Outbreak , 2012, Journal of Clinical Microbiology.

[22]  R. Black,et al.  Typhoid fever and paratyphoid fever: Systematic review to estimate global morbidity and mortality for 2010 , 2012, Journal of global health.

[23]  J. Wain,et al.  Revolutionising Bacteriology to Improve Treatment Outcomes and Antibiotic Stewardship , 2013, Infection & chemotherapy.

[24]  J. Wain,et al.  Next-generation sequencing in clinical microbiology , 2013, Expert review of molecular diagnostics.

[25]  Aaron A. Klammer,et al.  Nonhybrid, finished microbial genome assemblies from long-read SMRT sequencing data , 2013, Nature Methods.

[26]  Alexander S. Mikheyev,et al.  A first look at the Oxford Nanopore MinION sequencer , 2014, Molecular ecology resources.

[27]  Long-read, whole-genome shotgun sequence data for five model organisms , 2014 .

[28]  Aaron R. Quinlan,et al.  A reference bacterial genome dataset generated on the MinION™ portable single-molecule nanopore sequencer , 2014, bioRxiv.

[29]  Aaron R. Quinlan,et al.  Poretools: a toolkit for analyzing nanopore sequence data , 2014, bioRxiv.

[30]  Mick Watson,et al.  poRe: an R package for the visualization and analysis of nanopore sequencing data , 2015, Bioinform..