Culture-independent analysis of liver abscess using nanopore sequencing

PLoS One. 2018 Jan 9;13(1):e0190853. doi: 10.1371/journal.pone.0190853. eCollection 2018.

Abstract

The identification of microbial species has depended predominantly upon culture-based techniques. However, the difficulty with which types of organisms are cultured implies that the grown species may be overrepresented by both cultivation and plate counts. In recent years, culture-independent analysis using high-throughput sequencing has been advocated for use as a point-of-care diagnostic tool. Although it offers a rapid and unbiased survey to characterize the pathogens in clinical specimens, its accuracy is reduced by the high level of contamination of human DNA. In this paper, we propose using a culture-independent analysis for a Klebsiella pneumoniae clinical strain within a liver abscess using nanopore sequencing. Owing to the highly-contaminated cell population within a liver abscess, we managed to reduce the confounding effects of human DNA through the use of DNase and differential centrifugation. Genomic DNA was sequenced through the use of Nanopore MinION sequencer and analyzed using a suite of bioinformatics approaches. K. pneumoniae was successfully identified along with antibiotic-resistant genes. Our results indicate that, by integrating real-time nanopore sequencing and bioinformatics software, real-time pathogen identification in a liver abscess can be achieved.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • High-Throughput Nucleotide Sequencing
  • Humans
  • Klebsiella pneumoniae / drug effects
  • Klebsiella pneumoniae / genetics
  • Klebsiella pneumoniae / isolation & purification
  • Liver Abscess / diagnostic imaging
  • Liver Abscess / microbiology*
  • Microbial Sensitivity Tests
  • Nanopores*

Grants and funding

This work was supported in part by the Ministry of Science and Technology (MOST) with grant numbers 103-2923-E-194-001-MY3 and 104-2221-E-194-048-MY2 to YTH and this was supported by grants from Taichung Veterans General Hospital in Taiwan (TCVGH-1073901B and TCVGH-NCHU1067611) to PYL. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.