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Wednesday, September 16, 2020 (Updated Sunday, Sep 20, 2020)

A discussion of Li-Meng Yan's paper on SARS-CoV-2

Three dissident virologists claim to have proof that SARS-CoV-2 was artificially created. What is their evidence? (Updated Sep 20 2020)

O n September 14 2020, Li-Meng Yan, a dissident Chinese virologist, along with three colleagues, posted an article at zenodo.org [1] claiming to prove that SARS-CoV-2 was artificially created. Twitter canceled the author's account two days later and political activists vociferously attacked the paper on political grounds. It is clear that many people don't want to hear her evidence. But the issue is extremely important. If the virus was produced in an experiment and accidentally released, as this paper claims, it means the Wuhan Institute of Virology is in dire need of upgrades to their virus-handling procedures to prevent it from happening again.

My background is 30 years as a researcher in protein biochemistry using biophysical techniques and molecular biology, including design, cloning, and expression of recombinant proteins, to study protein function and structure. Here's a brief summary of the argument put forward in the paper along with my comments.

Update, Sep 20 2020: I've expanded the comments to describe the evidence they would need to make their case more convincing.

Claim 1: “[the sequences of] RaTG13, RmYN02, and several pangolin viruses recently published are highly suspicious and likely fraudulent.” “The RaTG13 virus is excluded from this analysis given the strong evidence suggesting that its sequence may have been fabricated and the virus does not exist in nature.”

Comment: RaTG13 was published by Shi Zhengli at the same time as the SARS-CoV-2 sequence. In 2013 Shi also published a partial gene segment of RaBtCoV/4991, which is identical to RaTG13, but it is incomplete. The authors say that the WIV took this partial sequence and fraudulently added the remainder to exculpate themselves, trying to make it appear that RaTG13 was a pre-existing virus.

Evidence in favor of this claim is that Shi et al. made no mention of RaBtCoV/4991 in their 2020 paper. But unless someone wants to claim that 4991 was also fabricated, RaBtCoV/4991 had to come from somewhere, which is to say it is a real virus. So Claim 1 doesn't make sense.

What is needed: To prove this the authors would have to find another virus that is 100% identical with the published partial RaBtCoV/4991 sequence but which differs from RaTG13.

Claim 2: The WIV could have used transgenic hACE2 transgenic mice to improve the virus's ability to bind to its receptor. “This animal model has been established during the study of SARS-CoV and has been available in the Jackson Laboratory [a commercial supplier of transgenic mice] for many years.” However, hACE2 mice are “not a good model to reflect the virus' transmissibility and associated clinical symptoms in humans.” If they had used golden Syrian hamster instead, the authors say, “the highly contagious nature of SARS-CoV-2 would be extremely evident” and they could have released accurate information.

Comment: This is a good point. If true, it would tend to exculpate the Chinese researchers. But we already know many techniques they could have used. No one was claiming that making the virus was impossible.

What is needed: Sending live animals to another country requires tons of paperwork. They would need Jax's sales records or government export records (which might be available via a FOIA request).

Claim 3: The paper provides a flow chart for how the virus could have been constructed. “Two restriction sites are present at either end of the RBM [receptor-binding motif] of SARS-CoV-2, providing convenience for replacing the RBM within the spike gene.”

Comment: The spike protein consists of two parts: S1, which binds to the receptor, and S2, which fuses the viral and cellular membranes. The point where the subunits are joined is called the S1/S2 site. Proteolysis of this site is essential for infection. The RBM is the part of S1 that binds to the receptor. A common technique is to take functional domains out of one protein and put them into another. The easiest way to do this is by finding (or creating) restriction sites in the DNA that allow you to cut the DNA in a specific place.

The flow chart is quite reasonable, but again no one doubted that artificially creating the virus is possible. So describing how someone could have done it doesn't tell us much; any competent molecular biologist could come up with a similar scheme.

The article makes a big deal about the presence of restriction sites on the S1/S2 sequence, including an EcoRI site and a BstEII site. But this does not indicate engineering. Many restriction sites occur naturally by chance. Their presence is not unusual, but they are a major inconvenience for those of us who try to clone DNA—many restriction sites we might wish to use are ruled out by the presence of another one in an inconvenient location. The probability of finding one of over a thousand known restriction sites at an arbitrary location in a 3800 base sequence by chance is very high.

What is needed: To prove this convincingly, the authors would need to find an authentic copy of the original virus sequence without the restriction sites to show what the virus was like before the supposed change was made.

Claim 4: The furin cleavage site contains rare codons, an unusual sequence not shared by other lineage B betacoronaviruses, and a FauI restriction site. It could not have been introduced by evolution, say the authors, and the probability of successful homologous recombination ever occurring among the ancestors of these viruses is low. “A FauI restriction site is formulated by the codon choices here, suggesting the possibility that the restriction fragment length polymorphism, a technique that a WIV lab is proficient at, could have been involved.” The authors suggest that the FauI site was added to monitor the presence of the furin-cleavage site.

Comment: The furin cleavage site is known to increase the tropism of the virus, and it is now known that the virus even infects the brain, but there is little evidence so far that it increases pathogenicity. There is as yet no good explanation for the presence of the furin cleavage site.

It might make sense for a virus creator to put a restriction site in the middle of the furin cleavage site to make sure it didn't disappear. RFLP analysis is a simple technique and any molecular biology lab could handle it easily. However, as mentioned, restriction sites pop up randomly all the time, so finding one proves little.

What is needed: The FauI restriction site is a red herring. To prove that the furin cleavage site was added, the authors would have to find a trail of evidence showing step by step how the sequence was manipulated from whatever virus the WIV started from. It would also be helpful to have data showing that furin cleavage increases pathogenicity. This is plausible but it needs to be shown experimentally.

Claim 5: WIV most likely used ZC45 and ZXC21, not RaTG13, to create SARS-CoV-2 because RaTG13 is not real. The reasoning behind this claim is that the S1 sequence of SARS-CoV-2 is quite similar to these other viruses except for the receptor-binding motif and furin-cleavage sites. Also, two other proteins on these viruses, the Orf8 protein and E protein, are 94.2% and 100% identical to SARS-CoV-2.

Comment: This is indeed strange but so far circumstantial.

What is needed: The authors need to prove that RaTG13 is not real. They claim to have evidence, and I look forward to seeing it, but to be convincing it would have to be more than the discovery of mysterious unexplained features in the DNA sequence.

The authors say they plan to prove that RaTG13 is fabricated in a follow-up report. If they can do this, it would be solid evidence of consciousness of guilt on the part of WIV researchers. However, the current paper doesn't say very much more than what other bloggers have already revealed.

It would be premature to draw any conclusion about the origins of the virus from this paper. The authors have set for themselves a difficult and possibly impossible task: finding a signature of intel­ligent design in a virus is much like what the creationists have been doing without success for years.

But what is not premature is that social media giants deciding for us what is true and what is false may turn out to be as big a threat as the virus. It is unfair and unscientific to dismiss ideas that one doesn't want to hear as “conspiracy theories” as many people are doing.

1. Yan LM, Kang S, Guan J, Hu S (2020). Unusual Features of the SARS-CoV-2 Genome Suggesting Sophisticated Laboratory Modification Rather Than Natural Evolution and Delineation of Its Probable Synthetic Route. https://zenodo.org/record/4028830

Comment added, Sep 30 2020: While searching the DNA for clues may be valuable, meta-analysis—that is, looking at records and lab notebooks—is more likely to find any evidence on whether the virus was artificially created or modified. Here is an example from a popular vitamin website.

sep 16 2020, 6:52 pm (Updated Sep 20 2020)

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