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

Jenai Quan ; Charles Langelier ; Alison Kuchta ; Joshua Batson ; Noam Teyssier ; Amy Lyden ; Saharai Caldera ; Aaron McGeever ; Boris Dimitrov ; Ryan King ; +18 more... Jordan Wilheim ; Maxwell Murphy ; Lara Pesce Ares ; Katherine A Travisano ; Rene Sit ; Roberto Amato ; Davis R Mumbengegwi ; Jennifer L Smith ; Adam Bennett ; Roly Gosling ORCID logo ; Peter M Mourani ; Carolyn S Calfee ; Norma F Neff ; Eric D Chow ; Peter S Kim ; Bryan Greenhouse ; Joseph L DeRisi ; Emily D Crawford ; (2019) FLASH: a next-generation CRISPR diagnostic for multiplexed detection of antimicrobial resistance sequences. Nucleic acids research, 47 (14). e83-. ISSN 0305-1048 DOI: 10.1093/nar/gkz418
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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.


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