Ultra-sensitive and rapid detection of nucleic acids and microorganisms in body fluids using single-molecule tethering.
Ultra-sensitive and rapid detection of nucleic acids and microorganisms in body fluids using single-molecule tethering.
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DOI：
10.1038/s41467-020-18574-7
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发表时间：
2020-09-22
影响因子：
16.6
通讯作者：
Celedon A
中科院分区：
文献类型：
--
作者：
Cheng WC;Horn T;Zayats M;Rizk G;Major S;Zhu H;Russell J;Xu Z;Rothman RE;Celedon A
体液微生物核酸检测已成为许多传染病快速诊断的首选方法。然而，在某些情况下，基于培养的耗时诊断仍然是金标准方法，例如败血症。迫切需要新的无培养方法。在这里，我们描述了单分子系留或SMOLT，这是一种无需扩增和纯化的分子检测方法，可以在不需要培养的情况下以高灵敏度检测体液中的微生物。SMOLT的信号是由由DNA探针连接的微米大小的珠子的位移产生的，这些珠子的长度在1到7微米之间。数千个DNA探针的分子延伸是用一种强大而快速的光学方法以亚微米精度确定的。我们证明，SMOLT可以在亚飞摩尔浓度下直接检测血液、尿液和痰中的核酸，以及在1 CFU mL - 1（每毫升菌落形成单位）下检测血液中的微生物，速度快三倍，具有更高的复用能力，并且比放大方法更直接。SMOLT的临床应用通过开发一种多重检测方法来进一步证明，该方法可以直接在全血中同时检测引起败血症的念珠菌种类。基于培养的微生物检测诊断方法耗时，但仍然是脓毒症等疾病的金标准。在这里，作者提出了一种无需扩增和纯化的方法，以高灵敏度检测体液中的微生物：单分子系留（SMOLT）。
Detection of microbial nucleic acids in body fluids has become the preferred method for rapid diagnosis of many infectious diseases. However, culture-based diagnostics that are time-consuming remain the gold standard approach in certain cases, such as sepsis. New culture-free methods are urgently needed. Here, we describe Single MOLecule Tethering or SMOLT, an amplification-free and purification-free molecular assay that can detect microorganisms in body fluids with high sensitivity without the need of culturing. The signal of SMOLT is generated by the displacement of micron-size beads tethered by DNA probes that are between 1 and 7 microns long. The molecular extension of thousands of DNA probes is determined with sub-micron precision using a robust and rapid optical approach. We demonstrate that SMOLT can detect nucleic acids directly in blood, urine and sputum at sub-femtomolar concentrations, and microorganisms in blood at 1 CFU mL−1 (colony forming unit per milliliter) threefold faster, with higher multiplexing capacity and with a more straight-forward protocol than amplified methodologies. SMOLT’s clinical utility is further demonstrated by developing a multiplex assay for simultaneous detection of sepsis-causing Candida species directly in whole blood. Culture-based diagnostic methods for microorganism detection are time-consuming but still the gold standard for conditions such as sepsis. Here the authors present an amplification and purification-free method to detect microorganisms in bodily fluids with high sensitivity: Single MOLecule Tethering (SMOLT).
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影响因子：
6.1
作者：
Rane TD;Chen L;Zec HC;Wang TH
通讯作者：
Wang TH
影响因子：
4.3
作者：
Sidstedt M;Hedman J;Romsos EL;Waitara L;Wadsö L;Steffen CR;Vallone PM;Rådström P
通讯作者：
Rådström P
影响因子：
48
作者：
Hindson, Christopher M.;Chevillet, John R.;Briggs, Hilary A.;Gallichotte, Emily N.;Ruf, Ingrid K.;Hindson, Benjamin J.;Vessella, Robert L.;Tewari, Muneesh
通讯作者：
Tewari, Muneesh
影响因子：
11.8
作者：
Mylonakis, Eleftherios;Clancy, Cornelius J.;Pappas, Peter G.
通讯作者：
Pappas, Peter G.
影响因子：
16.6
作者：
Lam, Brian;Das, Jagotamoy;Kelley, Shana O.
通讯作者：
Kelley, Shana O.