Molecular Diagnosis of a Novel Coronavirus (2019-nCoV) Causing an Outbreak of Pneumonia
Molecular Diagnosis of a Novel Coronavirus (2019-nCoV) Causing an Outbreak of Pneumonia
复制标题
DOI：
10.1093/clinchem/hvaa029
复制
发表时间：
2020-04-01
影响因子：
9.3
通讯作者：
Poon, Leo L. M.
中科院分区：
文献类型：
--
作者：
Chu, Daniel K. W.;Pan, Yang;Poon, Leo L. M.
背景：最近在急性呼吸道疾病患者中发现了一种新的人畜共患冠状病毒(2019-nCoV)。这种病毒在基因上与SARS冠状病毒和蝙蝠SARS类冠状病毒相似。疫情最初是在中国的主要城市武汉发现的，但随后在中国的其他省份也发现了疫情。其他几个国家也报告了与旅行相关的病例。卫生保健工作者的暴发表明人与人之间的传播。为了早期识别感染者，迫切需要对该病毒进行分子检测。方法：我们建立了两种一步定量实时逆转录聚合酶链式反应检测病毒基因组两个不同区域(ORF1b和N)的方法。设计了与这种新型冠状病毒及其密切相关的病毒如SARS冠状病毒反应的引物和探针组。使用一组阳性对照和阴性对照对这些检测进行了评估。结果：以从SARS冠状病毒感染的细胞中提取的RNA为阳性对照，这些方法的动态范围至少有7个数量级(2×10(-)(4)-2000 TCID50/反应)。以DNA质粒为阳性标准，每一反应的检测限均在10个拷贝以下。所有阴性对照样品的检测结果均为阴性。两例2019-nCoV感染者的检测结果均为阳性。结论：建立的检测方法可以快速检测出人体标本中的2019n-CoV，从而实现对患者的早期识别。
Background: A novel coronavirus of zoonotic origin (2019-nCoV) has recently been identified in patients with acute respiratory disease. This virus is genetically similar to SARS coronavirus and bat SARS-like coronaviruses. The outbreak was initially detected in Wuhan, a major city of China, but has subsequently been detected in other provinces of China. Travel-associated cases have also been reported in a few other countries. Outbreaks in health care workers indicate human-to-human transmission. Molecular tests for rapid detection of this virus are urgently needed for early identification of infected patients.Methods: We developed two 1-step quantitative real time reverse-transcription PCR assays to detect two different regions (ORF1b and N) of the viral genome. The primer and probe sets were designed to react with this novel coronavirus and its closely related viruses, such as SARS coronavirus. These assays were evaluated using a panel of positive and negative controls. In addition, respiratory specimens from two 2019-nCoV-infected patients were tested.Results: Using RNA extracted from cells infected by SARS coronavirus as a positive control, these assays were shown to have a dynamic range o f at least seven orders of magnitude (2 x 10(-)(4)-2000 TCID50/reaction). Using DNA plasmids as positive standards, the detection limits of these assays were found to be below 10 copies per reaction. All negative control samples were negative in the assays. Samples from two 2019-nCoV-infected patients were positive in the tests.Conclusions: The established assays can achieve a rapid detection of 2019n-CoV in human samples, thereby allowing early identification of patients.