Genome-wide tiled detection of circulating Mycobacterium tuberculosis cell-free DNA using Cas13

Nat Commun. 2023 Mar 31;14(1):1803. doi: 10.1038/s41467-023-37183-8.

Abstract

Detection of microbial cell-free DNA (cfDNA) circulating in the bloodstream has emerged as a promising new approach for diagnosing infection. Microbial diagnostics based on cfDNA require assays that can detect rare and highly fragmented pathogen nucleic acids. We now report WATSON (Whole-genome Assay using Tiled Surveillance Of Nucleic acids), a method to detect low amounts of pathogen cfDNA that couples pooled amplification of genomic targets tiled across the genome with pooled CRISPR/Cas13-based detection of these targets. We demonstrate that this strategy of tiling improves cfDNA detection compared to amplification and detection of a single targeted locus. WATSON can detect cfDNA from Mycobacterium tuberculosis in plasma of patients with active pulmonary tuberculosis, a disease that urgently needs accurate, minimally-invasive, field-deployable diagnostics. We thus demonstrate the potential for translating WATSON to a lateral flow platform. WATSON demonstrates the ability to capitalize on the strengths of targeting microbial cfDNA to address the need for point-of-care diagnostic tests for infectious diseases.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • CRISPR-Cas Systems
  • Cell-Free System
  • DNA, Bacterial / analysis
  • DNA, Bacterial / genetics
  • Genome, Bacterial
  • Humans
  • Mycobacterium tuberculosis* / genetics
  • Tuberculosis / diagnosis
  • Tuberculosis / microbiology

Substances

  • DNA, Bacterial