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

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.

中文翻译:








使用 Cas13 对循环结核分枝杆菌无细胞 DNA 进行全基因组平铺检测


检测血液中循环的微生物游离 DNA (cfDNA) 已成为一种很有前途的感染诊断新方法。基于 cfDNA 的微生物诊断需要能够检测稀有和高度片段化的病原体核酸的检测方法。我们现在报告 WATSON(使用核酸平铺监测的全基因组分析),这是一种检测少量病原体 cfDNA 的方法,该方法将平铺在基因组中的基因组目标的合并扩增与这些目标的基于 CRISPR/Cas13 的合并检测相结合。我们证明,与单个目标基因座的扩增和检测相比,这种平铺策略改进了 cfDNA 检测。WATSON 可以检测结核分枝杆菌的 cfDNA在活动性肺结核患者的血浆中,这种疾病迫切需要准确、微创、可现场部署的诊断。因此,我们展示了将 WATSON 转化为横向流动平台的潜力。WATSON 展示了利用靶向微生物 cfDNA 的优势来满足传染病即时诊断测试需求的能力。








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