于是，Vineet D. Menachery教授等利用SARS-CoV的反向遗传系统，从中提取并鉴定了一种嵌合病毒，该病毒在适应小白鼠的SARS-CoV主干中可表达出蝙蝠冠状病毒SHC014的刺突。结果表明，该新型冠状病毒能利用SARS的人类细胞受体——血管紧张素转换酶II（ACE2）的多个同源基因，在人类呼吸道原代细胞中有效复制，并在体外获得与SARS传染性同等的效果。
Ralph Baric, an infectious-disease researcher at the University of North Carolina at Chapel Hill, last week (November 9) published a study on his team’s efforts to engineer a virus with the surface protein of the SHC014 coronavirus, found in horseshoe bats in China, and the backbone of one that causes human-like severe acute respiratory syndrome (SARS) in mice. The hybrid virus could infect human airway cells and caused disease in mice, according to the team’s results, which were published in Nature Medicine.
The results demonstrate the ability of the SHC014 surface protein to bind and infect human cells, validating concerns that this virus—or other coronaviruses found in bat species—may be capable of making the leap to people without first evolving in an intermediate host, Nature reported. They also reignite a debate about whether that information justifies the risk of such work, known as gain-of-function research. “If the [new] virus escaped, nobody could predict the trajectory,” Simon Wain-Hobson, a virologist at the Pasteur Institute in Paris, told Nature.
In October 2013, the US government put a stop to all federal funding for gain-of-function studies, with particular concern rising about influenza, SARS, and Middle East respiratory syndrome (MERS). “NIH [National Institutes of Health] has funded such studies because they help define the fundamental nature of human-pathogen interactions, enable the assessment of the pandemic potential of emerging infectious agents, and inform public health and preparedness efforts,” NIH Director Francis Collins said in a statement at the time. “These studies, however, also entail biosafety and biosecurity risks, which need to be understood better.”
Baric’s study on the SHC014-chimeric coronavirus began before the moratorium was announced, and the NIH allowed it to proceed during a review process, which eventually led to the conclusion that the work did not fall under the new restrictions, Baric told Nature. But some researchers, like Wain-Hobson, disagree with that decision.
The debate comes down to how informative the results are. “The only impact of this work is the creation, in a lab, of a new, non-natural risk,” Richard Ebright, a molecular biologist and biodefence expert at Rutgers University, told Nature.
But Baric and others argued the study’s importance. “[The results] move this virus from a candidate emerging pathogen to a clear and present danger,” Peter Daszak, president of the EcoHealth Alliance, which samples viruses from animals and people in emerging-diseases hotspots across the globe, told Nature.
1、“关于新冠病毒源于人工合成”之所，请贵所认真核实贵所研究员石正丽的论文——A SARS-like cluster of circulating bat coronaviruses shows potential for human emergence发表于2015年11月9日（Published: 09 November 2015 ）在该论文中提到“The results indicate that group 2b viruses encoding the SHC014 spike in a wild-type backbone can efficiently use multiple orthologs of the SARS receptor human angiotensin converting enzyme II (ACE2), replicate efficiently in primary human airway cells and achieve in vitro titers equivalent to epidemic strains of SARS-CoV.
（该论文作者：Vineet D Menachery, Boyd L Yount Jr, Kari Debbink, Sudhakar Agnihothram, Lisa E Gralinski, Jessica A Plante, Rachel L Graham, Trevor Scobey, Xing-Yi Ge, Eric F Donaldson, Scott H Randell, Antonio Lanzavecchia, Wayne A Marasco, Zhengli-Li Shi & Ralph S Baric ”由于我不是专业人士，故只能看出大体意思是“只要把蝙蝠身上的S蛋白里头ACE2这个受体开头只要一调，这个病毒马上就可以传染给人”，请贵所进行公开澄清，这种研究是不是属于人工合成的一部分？在武汉肺炎之前，武汉病毒研究所及其研究员究竟进行了多少这样的研究？
Isolation of a new bat virus in a studypublished in the Journal of Virology on Dec. 30, 2015, titled “Isolation and Characterization of a Novel Bat Coronavirus Closely Related to the Direct Progenitor of Severe Acute Respiratory Syndrome Coronavirus,” found that the virus, named SL-CoV-WIV1, was almost identical to Rs3367 with 99.9% genome sequence identity. The researchers identified that WIV1 can use human ACE2 as an entry receptor and has the potential to infect human cells in this study. Subsequently, the same research group isolated another bat virus that can use ACE2 and infect human cell lines in the lab in 2015.
In addition, Dr. Shi’s group conducted another study in 2018 to address the question of whether some bat viruses can infect humans via using human ACE2, without the need of an intermediate host. But to the date of their study, “no direct transmission of SARS-Like CoVs from bats to people has been reported”.
Zhengli Shi’s group at the Institute of Virology at Wuhan was successful in isolating two infectious clones of bat SARS-Like CoV: SL-CoV-WIV1 and WIV16 from bats. In their further studies, they found out that these SL-CoV Spike protein (S protein) “[were] unable to use any of the three ACE2 molecules as its receptor; Second, the SL-CoV failed to enter cells expressing the bat ACE2; Third, the chimeric S covering the previously defined receptor-binding domain gained its ability to enter cells via human ACE2, albeit with different efficiencies for different constructs; Fourth, a minimal insert region ( Amino acids 310 to 518 ) was found to be sufficient to convert the SL-CoV S from non-ACE2 binding to human ACE2 binding.”
Therefore, Shi’s group found in a study published in the Journal of Virology in February 2008 that the natural bat coronavirus cannot use the human ACE2 receptor to infect humans. However, when inserted with some amino acids from position 310 to 518 for the bat CoV S protein sequence, the chimeric bat CoV can use the human ACE2 receptor.
Meanwhile, another research group led by Dr. Li published their finding in 2013 that 5 amino acid sites on CoV spike proteins are crucial in making the binding to human ACE2 on SARS virus (those positions are Y442, L472, N479, D480, T487). These 5 sites just lie in the region that the Shi group noted to be important above.
Later, Li and Shi jointly conducted a gain-of-function study published in the Journal of Virology in September 2015 on the MERS virus and a bat virus (strain HKU4) in 2015. Since MERS virus can enter human cells but HKU4 can not, they introduced 2 single mutations in the HKU4 spike protein and found that the new mutant S protein can enable HKU4 to enter human cells. If they mutated 2 sites in MERS spike, the resulting MERS pseudovirus (experimental virus) cannot enter human cells anymore.
To their surprise, the chimeric virus (SHC014-MA15) can use SHC014 spike to bind to human ACE2 receptor and enter human cells. SHC014-MA15 can also cause disease in mice and cause death as well. Existing vaccines to SARS cannot protect animals from SHC014-MA15 infection. Therefore, these chimeric virus studies can lead to the generation of more pathogenic, more deadly CoV strains in mammalian models.
Due to the U.S. government-mandated pause on the gain-of-function (GOF) studies, this international research did not proceed further at that time. However, there is no evidence that Shi’s group in China stopped any further study on the track of introducing GOF mutations on the CoV. And it is clear that Shi’s group already mastered the reverse-engineering technology that is sufficient to introduce mutation in current SARS-CoV or SARS-Like CoV to create mutant infectious coronavirus.
t has been two months since the outbreak of the coronavirus in Wuhan and its spread has shown no signs of slowing down in China. More than 35 Chinese cities have been put on lockdown by Chinese authorities in an attempt to isolate confirmed and suspected cases. The lives of millions of people are in danger as the virus shows signs of spreading further in China as well as internationally.
