Critical Analysis of the New Preprint that Claims Proof for Bioengineered SARS-CoV-2


Taking a hard look at their arguments and data, instead of attacking their credibility.* Illustration: Nadine D

Dr. Li-Meng Yan, a former virologist at the University of Hong Kong School of Public Health, fled the country in April to the U.S. She said she “will be disappeared and killed” if she tries to speak up in China. In the U.S., she has been making headlines that humans created SARS-CoV-2 (that causes Covid-19), which was then released on purpose. And she has ‘proof’ for that, which she released as a preprint this week, titled “Unusual Features of the SARS-CoV-2 Genome Suggesting Sophisticated Laboratory Modification Rather Than Natural Evolution and Delineation of Its Probable Synthetic Route.”

Many have discredited the paper — calling it a ‘poppycock’ — because it is not peer-reviewed and affiliated with the Rule of Law Society, a group with no history of scientific publications.

Dr. Yan, however, has co-authored two Covid-19 papers published in top-tier journals: “Pathogenesis and transmission of SARS-CoV-2 in golden hamsters” in Nature and “Viral dynamics in mild and severe cases of COVID-19” in The Lancet. Her solid background in corona-virology piqued my interest in what her preprint has to say.

On the preprint

The preprint has two sections. Section one presents the theory that SARS-CoV-2 “is a product of laboratory modification.” And section two outlines the “probable pathway for the laboratory creation of SARS-CoV-2.”

As section two is only valuable if section one is reasonable, this article will focus on just section one. Section one is further divided into three points that argue the case for bioengineered SARS-CoV-2. This article will summarize each point Dr. Yan presented, followed by an evaluation of its validity.* Screenshot excerpt from Yan et al. (2020). Unusual Features of the SARS-CoV-2 Genome Suggesting Sophisticated Laboratory Modification Rather Than Natural Evolution and Delineation of Its Probable Synthetic Route

(i) SARS-CoV-2 is created from a ZC45/ZXC21 backbone.

“Searching the NCBI sequence database reveals that, among all known coronaviruses, there were two related bat coronaviruses, ZC45 and ZXC21, that share the highest sequence identity with SARS-CoV-2 (each bat coronavirus is ~89% identical to SARS-CoV-2 on the nucleotide level),” Dr. Yan writes. She further argued that ZC45/ZXC21 and SARS-CoV-2 share two highly identical proteins, a pattern not seen in any other coronaviruses.

(ZC45/ZXC21 are bat coronaviruses the China military research labs isolated from bats in Zhoushan in China, of which the genomic data was published in 2018.)

“Therefore, …the overall genomic/amino acid-level resemblance between SARS-CoV-2 and ZC45/ZXC21 are highly unusual,” the preprint states. “Such evidence, when considered together, is consistent with a hypothesis that the SARS-CoV-2 genome has an origin based on the use of ZC45/ZXC21 as a backbone and/or template for genetic gain-of-function modifications.”

Dr. Yan also admitted that the closest relative of SARS-CoV-2 is the RaTG13 bat sarbecovirus with a 96% sequence identity. But she cited preprints “questioning the actual existence of this [RaTG13] bat virus in nature.” And she closed part (i) with, “The fact that a virus has been fabricated to shift the attention away from ZC45/ZXC21 speaks for an actual role of ZC45/ZXC21 in the creation of SARS-CoV-2.”

Are the arguments of (i) valid?

Her entire case is based on the fact that ZC45/ZXC21 and SARS-CoV-2 are about 89% genetically identical. But evidence of sequence identity does not mean evidence of genetic manipulation. Instead, sequence identity is a standard metric how scientists measure evolutionarily relatedness.

SARS-CoV-2, a coronavirus, is also expected to have genes similar to other coronaviruses, Dr. Angela Rasmussen, a virologist at the Center of Infection and Immunity at Columbia University School of Public Health, told Vox news in response to Dr. Yan’s preprint.

The same applies to the protein sequence identity, where Dr. Yan noted that ZC45/ZXC21 have two proteins resembling SARS-CoV-2. But the genome of SARS-CoV-2 codes for at least 29 known proteins, with an additional 23 previously unknown proteins recently reported in September in Nature. So, she picked only two out of currently known 52 proteins of SARS-CoV-2 to make her case.

Further, the evidence she cited on the real-life actuality of the RaTG13 bat sarbecovirus all are preprints and, thus, remains unconfirmed. But does it really matter if RaTG13 does not exist in the wild? Dr. Yan’s main assertion is that SARS-CoV-2 is bioengineered from ZC45/ZXC21, not from RaTG13.

(ii) The receptor-binding motif (RBM) of SARS-CoV-2 cannot arise from nature and was bioengineered.

SARS-CoV-2 has a spike protein on its surface. The tip of this spike protein — called receptor-binding motif (RBM) — binds to the ACE2 receptor to initiate cell infection. And this RBM of SARS-CoV-2 is genetically similar to the RBM of SARS-1 that caused the 2003 SARS outbreak.

If SARS-CoV-2 arose from nature, its RBM would be highly different from SARS-1, Dr. Yan said. This is because one mechanism of natural evolution is random mutations. So the SARS-CoV-2 RBM would have accumulated many mutations as it evolves naturally.

Another way natural evolution works is by genetic recombination where SARS-CoV-2 gets its RBM from another virus that already has an RBM well adapted to humans. This cannot happen in bats, however, as their ACE2 receptor is very different from humans’. Older data suggested that genetic recombination occurred in pangolins as their ACE2 receptor is similar to humans, but more recent evidence does not support this theory.

All in all, it is doubtful that SARS-CoV-2 RBM arose naturally, from random mutations or genetic recombinations, Dr. Yan argued.

To reinforce her point, Dr. Yan cited a published study in 2008 by the Wuhan Institute of Virology that swapped the RBM of bat coronaviruses with that of SARS-1 — to see if the ‘recombined’ coronaviruses could bind to the human ACE2 receptor. And they did, which confirms that the RBM determines if a coronavirus can infect human cells. Regardless, the purpose of this citation is to show that artificial genetic recombination is possible.

Dr. Yan further identified the presence of restriction sites in the SARS-CoV-2 genome, around its RBM. Restriction sites are genetic areas that tell restriction enzymes where to cut. The cut out genetic sequence can then be swapped with another sequence — creating a recombinant virus.

“The striking finding of EcoRI and BstEII restriction sites at either end of the SARS-CoV-2 RBM, respectively, and the fact that the same RBM region has been swapped both by Dr. Shi and by her long-term collaborator, respectively, using restriction enzyme digestion methods are unlikely a coincidence,” Dr. Yan writes. “Rather, it is the smoking gun proving that the RBM/Spike of SARS-CoV-2 is a product of genetic manipulation.”

Are the arguments of (ii) valid?

Her argument is founded by the observation that SARS-CoV-2 has an RBM similar to SARS-1. Again, as mentioned, sequence identity or similarity indicates evolutionary relatedness. Plus, SARS-1 is a product of natural evolution. If its RBM arose naturally, there is no reason why the RBM of SARS-CoV-2 cannot.

Further, a highly valuable virus segment like the RBM should be ‘conserved’ and protected from too many mutations in a process called negative selection. If the RBM undergoes a mutation that impairs its function, the virus’s infectivity will suffer. Most mutations are indeed not beneficial. There are at least 353,341 mutational events in the SARS-CoV-2 genome, yet only one mutation — D614G — is advantageous for the virus’s survival.

Dr. Yan is correct regarding the improbability of an intermediate host that houses the genetic recombination. Current evidence suggests that the RaTG13 bat sarbecovirus is the ancestor of SARS-CoV-2, which means that SARS-CoV-2 came directly from bats without any intermediate host.

Assuming the SARS-CoV-2 RBM did not originate by nature, Dr. Yan stressed that the restriction sites surrounding the SARS-CoV-2 RBM are a ‘smoking gun’ for its bioengineering. Restriction sites are based on their respective restriction enzymes. There are over 400 known restriction enzymes from bacteria, and each enzyme has a specific restriction site it targets. Importantly, restriction enzymes can be designed artificially, so virtually anywhere in the SARS-CoV-2 genome can serve as restriction sites.

(iii) The furin-cleavage site of SARS-CoV-2 cannot arise from nature and was bioengineered.

Within the RBM of SARS-CoV-2, there is also the furin-cleavage site. This site needs to be cleaved by furin protease — an enzyme ubiquitous in many cell types — to complete virus entry into cells.

According to a published report in May, the furin-cleavage site of SARS-CoV-2 is unique to its lineage and not found in any other related coronaviruses. Although other coronavirus lineages have a furin-cleavage site, they are too distinct from SARS-CoV-2 to share a common ancestor. “Therefore, the furin-cleavage site within the SARS-CoV-2 Spike protein is unlikely to be of natural origin and instead should be a result of laboratory modification,” Dr. Yan writes.

Next, she highlighted a restriction site called FauI near the SARS-CoV-2 furin-cleavage site. This restriction site has a genetic sequence made out of two consecutive rare codons that are not found anywhere else in the SARS-CoV-2 genome. (Codons code for proteins; rare codons are the least used codons by a biological entity).

Put it simply, two rare codons back-to-back that code for a restriction site near the furin-cleavage site of SARS-CoV-2 is too much of a coincidence. Hence, the rare codons should have been inserted by genetic manipulation.

Are the arguments of (iii) valid?

This point is based on the fact that the furin-cleavage site of SARS-CoV-2 is unique to its lineage, which discredits its natural origin.

But the May study she cited also stated that the furin-cleavage site of SARS-CoV-2 is similar to MERS and common cold human coronaviruses. Thus, despite the different lineages, SARS-CoV-2 might get its furin-cleavage site via genetic recombination with other human coronaviruses. Coronaviruses are infamous for their high recombination rates. And Dr. Rasmussen agrees that the furin-cleavage site is present in many coronaviruses, not just SARS-CoV-2.

Point (iii) also pointed out that the two consecutive rare codons coding for a restriction site near the furin-cleavage site, she writes, “should be a result of genetic engineering.”

It is noteworthy, however, that rare codons are just codons that are used the least often. As follows, every biological entity has its own sets of rare codons. Hence, the rare codons Dr. Yan stressed do not prove genetic insertion.

Let’s assume the two consecutive rare codons are inserted artificially. Dr. Yan also said that these rare codons are not found anywhere else in the SARS-CoV-2 genome, supporting its artificial insertion. But it remains unknown if SARS-CoV-2 can read and translate those codons into the FauI restriction site, to begin with. In fact, rare codons are not easily translatable into proteins than common codons; they are called ‘rare’ for a reason.* An image of Dr. Li-Meng Yan by 葉 正道 Ben(busy) is licensed under CC BY-SA 2.0

Closing thoughts

The preprint of Dr. Yan, a virologist who co-authored two Covid-19 papers in top-ranking journals, claims that SARS-CoV-2 is bioengineered. She presented three major points for her assertion. However, none of them are sufficient to prove her case, as this article analyzed in-depth and showed. While many have defamed her preprint because it is not peer-reviewed and affiliated with a group with no history of scientific publications, this article dismissed the preprint due to its flawed arguments.

Lastly, this article is consistent with two other critical reviews that concluded Dr. Yan’s preprint as misleading.

This article was originally published in Microbial Instincts with minor modifications.

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