It is Difficult to Test Novel Coronavirus

Novel Coronavirus is one of the major themes of our daily life in 2020. Coronavirus is a type of virus. There are many different types, and some can cause disease. For example, COVID-19 is caused by SARS-CoV-2. A virus surface spike protein mediates SARS-CoV-2 entry into cells. To fulfill its function, SARS-CoV-2 spike binds to its receptor human ACE2 (hACE2) through its receptor-binding domain (RBD) and is proteolytically activated by human proteases. That is the way how the sars-cov-2 virus enters cells.

Compared with the SARS virus in 2003, the novel coronavirus (hereinafter referred to as the novel coronavirus) is relatively mild, and the infected person has a slower onset and milder symptoms. However, it is this superficial mildness that has brought about more serious disasters. Like the ten fragrances in Jin Yong's novels, they are colorless, tasteless, and shadowless, but they can kill people invisible when people are most alert. However, for the novel coronavirus, we are not completely untraceable. Compared with 2003, we have more advanced science and technology, in a very short time, obtained the whole genome sequence of the novel coronavirus, and designed specific primers and probes, which allows us to from the nucleic acid level Detect the presence of novel coronavirus.

Principle of nucleic acid molecule detection of novel coronavirus

Nucleic acids include DNA and RNA. Each organism has its own unique nucleic acid sequence. According to this principle, Chinese scientists completed the analysis of the whole genome sequence of the novel coronavirus in a very short time, and found the specific nucleic acid sequence only in the novel coronavirus by comparing with the genome sequences of other species. We only need to detect whether the patient's samples contain these specific sequences from the novel coronavirus, we can know whether the patient has been infected with the novel coronavirus.

The method for detecting the specific sequence of the novel coronavirus is fluorescence quantitative PCR (RT-qPCR). First, the specific sequence of the novel coronavirus is amplified using specific primers (the method of DNA amplification in vitro is called PCR), and then it is checked whether the amplified product can fully match the fluorescent probe. The fluorescent probe is also a unique nucleic acid sequence of the novel coronavirus. If the sequence of the amplified product and the fluorescent probe are identical, the fluorescent probe will emit fluorescence; if it is not completely identical, the fluorescent probe will not emit fluorescence. And if there are more viruses in the patient, the earlier the fluorescence signal appears, that is, the smaller the Ct value. Finally, according to the size of the Ct value to determine whether the patient is infected with novel coronavirus. Generally, if the Ct value is less than 37, it is considered to be positive. The Ct value is between 37 and 40 and needs to be retested; while the Ct value is greater than 40, it means negative.

Novel Coronavirus detection process

The nucleic acid detection of the novel coronavirus begins with pharyngeal swab sampling, which is to dip the throat secretion of the patient with a cotton swab. After obtaining the patient sample, the sample is cryopreserved, packaged, and sent to a special testing agency for further testing. Because patients and patient samples may have a large amount of novel coronavirus, each step in the process may cause infection to the operator.

After receiving the sample, the testing institution needs to pre-process the sample and then extract the nucleic acid in it. Since the novel coronavirus is an RNA virus, its nucleic acid component is RNA. However, RNA is extremely unstable and easily degraded, so this process is also more difficult. In addition, RNA cannot be directly detected by PCR, and it is necessary to reverse transcribe RNA into DNA before testing. Fortunately, most of the current nucleic acid detection kits are one-step methods, that is, the two-step synthesis of reverse transcription and PCR detection is completed in one step, which greatly reduces the workload of the staff. The PCR amplification is done in a special fluorescence quantitative PCR instrument. Through fluorescence quantitative PCR, we can obtain the Ct value of the sample. Through the size of the Ct value, we can determine whether the patient sample contains novel coronavirus. However, it should be noted that patients generally need at least two nucleic acid tests to confirm the diagnosis, and the interval between the two tests needs more than one day. After the test is completed, all test samples and experimental wastes need special and strict treatment to prevent virus environmental pollution.

Difficulties in detection of novel coronavirus

In the detection process of novel coronavirus, the three most critical elements are personnel, laboratory and kit. Unfortunately, as far as the current situation is concerned, these three elements are very scarce. First of all, the entire inspection process is complicated and has a high risk factor. Each operator must be experienced and very careful. Moreover, the number of samples for each test cannot be too large. Therefore, even if our test personnel are working in three shifts, the test throughput is still not up. Secondly, although many laboratories have fluorescence quantitative PCR instruments, they can complete fluorescence quantitative PCR experiments. However, in order to prevent the spread of the virus, laboratories that detect novel coronavirus must meet Class 2 (P2) biosafety standards. Regardless of the layout, instruments, airflow, and operating specifications of the laboratory, there are special requirements. many. The last is the COVID-19 Detection Kit. Although many companies have developed nucleic acid detection kits that can detect novel coronaviruses in a short period of time, it is limited by the fact that the current production personnel cannot fully resume work, the lack of raw materials for production, and the difficulty of full cold chain transportation Factors such as hospitals and testing institutions are still facing a shortage of testing kits. Therefore, the diagnosis of novel coronavirus still faces a grim situation!