FLASH: a next-generation CRISPR diagnostic for multiplexed detection of antimicrobial resistance sequences

Nucleic Acids Res. 2019 Aug 22;47(14):e83. doi: 10.1093/nar/gkz418.


The growing prevalence of deadly microbes with resistance to previously life-saving drug therapies is a dire threat to human health. Detection of low abundance pathogen sequences remains a challenge for metagenomic Next Generation Sequencing (NGS). We introduce FLASH (Finding Low Abundance Sequences by Hybridization), a next-generation CRISPR/Cas9 diagnostic method that takes advantage of the efficiency, specificity and flexibility of Cas9 to enrich for a programmed set of sequences. FLASH-NGS achieves up to 5 orders of magnitude of enrichment and sub-attomolar gene detection with minimal background. We provide an open-source software tool (FLASHit) for guide RNA design. Here we applied it to detection of antimicrobial resistance genes in respiratory fluid and dried blood spots, but FLASH-NGS is applicable to all areas that rely on multiplex PCR.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anti-Bacterial Agents / pharmacology*
  • Bacteria / classification
  • Bacteria / drug effects
  • Bacteria / genetics
  • Bacterial Infections / diagnosis
  • Bacterial Infections / genetics
  • Bacterial Infections / prevention & control
  • CRISPR-Cas Systems*
  • Computational Biology / methods*
  • Drug Resistance, Bacterial / drug effects*
  • Drug Resistance, Bacterial / genetics
  • High-Throughput Nucleotide Sequencing / methods*
  • Humans
  • Metagenomics / methods
  • Reproducibility of Results
  • Sensitivity and Specificity


  • Anti-Bacterial Agents