How to detect the SARS-CoV-2 virus?
Figure from: Cryo-EM image of
IVDC, China CDC
On January 21, 2020, the U.S. Centers for Disease Control and Prevention (CDC) reported the first case in the United States of the new coronavirus. The arrival of new SARS-like coronavirus in U.S. heightens concerns about global spread after the infection cases reported in Thailand, Japan, South Korea and Taiwan. Meanwhile, a panel of Chinese health experts confirmed that the new virus is able to spread between people, which indicates it could be much harder to control.
Via next generation sequencing (NGS) of cultured virus or samples from several pneumonia patients, the etiologic agent responsible for the pneumonia cases has been identified as a novel betacoronavirus (in the same family as SARS-CoV and MERS-CoV). Electron microscopy revealed the virus is a coronavirus with a characteristic crown morphology.
The genome sequence of this betacoronavirus is crucial to develop specific diagnostic tests and to identify potential intervention options. Full genome sequence data from the viruses have been released and are available on NCBI (https://www.ncbi.nlm.nih.gov/nuccore/MN908947 ). Working directly from sequence information, a few laboratories developed several genetic amplification (PCR) assays to detect the novel coronaviruses. A few diagnostic kits developed by commercial companies, using the real-time PCR (RT-PCR) assay have been put to use rapidly. The timely emergence of detection methods is also one reason for the sharp rise in the number of confirmed cases.
Principle of novel coronavirus detection
The virus detection method can be basically illustrated as below.
Isolate the novel coronavirus (2019-nCoV) from the patients and sequence its genome.
Compare the genome sequence of 2019-nCoV with human genome to find out the specific sequence in the virus genome.
Design PCR amplification primers and fluorescent probe primers for detecting the specific sequences identified in step 2.
Extract RNA from suspected individual’s serum, and convert the RNA into cDNA. The cDNA is then used as template and mixed with the PCR primers and probes for amplification. If the fluorescence signal increases rapidly and Ct value is less than 37, it can be determined as positive; If there’s no fluorescence detected, or the fluorescence signal grows slowly and Ct value finally ends up above 40, it can be determined as negative.