On April 19, Li Lanjuan, academician of Chinese Academy of engineering, released a new paper without peer review on the preprint platform medrxiv. The paper points out that there have been mutations that can affect the pathogenicity of the new coronavirus. It is necessary to take these mutations into account in drug and vaccine research and development.
Key conclusions of the paper:
The variation and diversity of new coronavirus strains may be greatly underestimated.
The difference of cytopathic effect and viral load of different mutant strains was 270 times.
A trinucleotide mutation can greatly enhance the replication rate and pathogenicity of the virus. It was found that the patients with this mutation remained nucleic acid positive for 45 days.
The researchers isolated live, replicable viruses from faeces, unlike previous reports.
Between January 22, 2020 and February 4, 2020, researchers isolated the virus from 11 patients with new coronavirus infection admitted to the Affiliated Hospital of Zhejiang University. The patients ranged in age from 4 months to 71 years, 10 of whom had a history of exposure to Wuhan.
Epidemiological information of 11 patients
The researchers sequenced the 11 virus isolates from patients in a super deep way and compared them with 1111 genome sequences from the gisaid database. It was found that there were 33 mutations in the virus isolated from these 11 patients, 19 of which were new mutations found for the first time.
The researchers wrote:
Although in this study, we only analyzed viruses isolated from 11 patients, we observed a rich diversity of mutations.
… This shows that the true diversity of new coronavirus strains is still greatly underestimated.
There have been a lot of mutations in the new coronavirus, and a considerable part of the mutations related to adaptability are enriched in the interface between the virus S protein and human ACE2 receptor.
So here comes the key question:
Do these mutations have a functional effect on the pathogenicity of the virus?
As we all know, the symptoms of new coronavirus infection are very complex, but also with the patient’s age, physical condition and other factors. In order to objectively evaluate the effect of these mutations on the pathogenicity of the virus, the researchers decided to carry out an in vitro infection test.
They chose a cell line called Vero-E6, which has ACE2 receptors very similar to human cells. They infected Vero-E6 cells with viruses from all 11 patient sources, collected cells at regular intervals and tested their viral load.
In the first four hours, the viral load of these cells remained stable, at this time, virus replication rarely occurred.
Eight hours later, the viral load of patients 6, 7, 9, 10 and 11 increased significantly.
24 hours later, all virus loads except No. 2 and No. 7 increased significantly, while No. 10 and No. 11 rose much faster than others.
It is worth noting that the virus load of 10 with the strongest replication ability is nearly 270 times higher than that of 2 with the weakest replication ability within 24 hours.
So, the higher the viral load, does it mean the stronger the killing power to cells?
The researchers tested at 48 and 72 hours and found that the higher the viral load, the higher the pathological effect and mortality of cells.
In the virus from patient 11, there was an unexpected trinucleotide mutation, which showed “strong” in the follow-up experiments, greatly improving the replication rate and pathogenicity of the virus strain. In fact, the patient tested positive for the virus for an amazing 45 days. The functional effect of this trinucleotide mutation is worthy of further study.
Finally, in contrast to the recently reported “no live virus has been isolated from feces”, three virus isolates in this study were from patients’ fecal samples, indicating that the new coronavirus has the ability to replicate in feces.
Finally, the researchers call for the development of new coronavirus drugs and vaccines to consider the impact of these cumulative mutations as soon as possible to avoid potential pitfalls.
Fortunately, by the time of writing, all 11 patients mentioned in this paper had recovered and discharged.