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

Journal name:
Nature Biotechnology
Volume:
33,
Pages:
296–300
Year published:
DOI:
doi:10.1038/nbt.3103
Received
Accepted
Published online

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.

At a glance

Figures

  1. A box-and-whiskers plot of the z-score for deleted k-mers of 3-6 bp in length, grouped by the proportion of GC content (0 = 0% GC content; 1 = 100% GC content).
    Figure 1: A box-and-whiskers plot of the z-score for deleted k-mers of 3–6 bp in length, grouped by the proportion of GC content (0 = 0% GC content; 1 = 100% GC content).
  2. Genetic organization of the S. Typhi chromosomal resistance island.
    Figure 2: Genetic organization of the S. Typhi chromosomal resistance island.

    Gene names were assigned using BLAST analysis and manual annotation.

  3. Comparison of the S. Typhi chromosomal resistance island with two closely related plasmids.
    Figure 3: Comparison of the S. Typhi chromosomal resistance island with two closely related plasmids.

    The S. Typhi chromosomal resistance island was compared to the S. Typhi multi-drug resistance plasmid pHCM1 and the plasmid with the highest BLASTN similarity to the island, pRSB107, from an uncultured organism in sewage. Top, pHCM1; middle, genomic island; bottom, pRSB107. (a) Overview to show full-length sequence of pHCM1 and pRSB107. (b) Zoomed-in view to show detail of regions of similarity. Red indicates 100% similarity and blue indicates 99% similarity; the cut-off minimum was set at 800.

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Primary accessions

NCBI Reference Sequence

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Author information

  1. These authors contributed equally to this work.

    • Philip M Ashton &
    • Satheesh Nair

Affiliations

  1. Gastrointestinal Bacteria Reference Unit, Public Health England, Colindale, London, UK.

    • Philip M Ashton,
    • Satheesh Nair &
    • Tim Dallman
  2. Department of Biomedical Sciences, University of Sassari, Sassari, Italy.

    • Salvatore Rubino
  3. Department of Infection and Immunity, King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia.

    • Salvatore Rubino
  4. National Reference Centre for Salmonellae and other Enterics, Robert Koch Institute, Wernigerode, Germany.

    • Wolfgang Rabsch
  5. Norwich Medical School, University of East Anglia, Norwich, UK.

    • Solomon Mwaigwisya,
    • John Wain &
    • Justin O'Grady

Contributions

P.M.A., S.N., T.D., J.W. and J.O'G. conceived the study, performed the analysis and wrote the first draft of the manuscript. J.O'G. and S.M. performed the MinION sequencing. P.M.A. and T.D. performed the bioinformatics analysis. S.N. performed the PCR analysis and coordinated the Illumina sequencing. P.M.A., T.D., S.R., W.R., J.W. and J.O'G. analyzed the resistance island structure and insertion site and devised the figures. All authors contributed to editing and data analysis of the final manuscript.

Competing financial interests

J.O'G. is a participant of Oxford Nanopore's MinION Access Programme (MAP) and received the MinION device and flowcells used for this study free of charge.

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