This article has been updated to include information about the two reports published by exiled scientist Li-Meng Yan et al. (Updates at the end of the article.)
Six months on from the emergence of SARS-CoV-2 as a global threat, there is still no clarity about its origins. Those who speculate that the virus may have been manipulated in a laboratory are often dismissed as conspiracy theorists, but there is evidence to support the suggestion that gain-of-function research has made it particularly virulent.

While some scientists still argue that SARS-CoV-2 is a product of natural evolution, others consider an accidental or deliberate leak from a laboratory to be a perfectly valid hypothesis that merits further investigation.

For decades, gain-of-function research, which alters viruses to increase their transmissibility, pathogenicity, virulence or lethality, has been carried out by American and Chinese scientists working in collaboration. There have been numerous ‘leaks’ of viruses from laboratories, including during the severe acute respiratory syndrome (SARS) outbreak that occurred in 2003–2004.

As the numbers of Covid-19 cases – and the global death toll – continue to increase, so does the discord over lockdowns and the wearing of face masks.

The race to find a vaccine against SARS-CoV-2 is meanwhile gaining momentum. The companies producing the main contenders say their results are promising, but critics of the “warp speed” approach to vaccine development say fast tracking is reckless and the dangers of pathogenic priming, or disease enhancement, are very real.

Scientists point to ‘suspicious’ SARS-CoV-2 properties

In an interview published in Minerva in Norway on July 2, the Norwegian virologist Birger Sørensen said it was more than 90% certain that SARS-CoV-2 originated in a laboratory.

Sørensen and his fellow scientists Angus Dalgleish and Andres Susrud have done their research with a view to producing a vaccine.

Reporters Aksel Fridstrøm and Nils August Andresen explain in their article that, in 2008, Sørensen’s work came to international attention when he launched a new immunotherapy for HIV. Dalgleish is the professor at St George’s medical school at the University of London who became world famous in 1984 after he discovered a novel receptor that the HIV virus uses to enter human cells.

Sørensen (pictured left) told Minerva that he and his colleagues discovered that SARS-CoV-2 was exceptionally well adjusted to infect humans, to the degree that it was suspicious.

He said he and his colleagues had discovered properties in SARS-CoV-2 that enable it to use an additional receptor and create a binding to human cells in the upper respiratory tract and the intestines that is strong enough to produce an infection.

“This is what enables the virus to transmit to a greater degree between humans, without the virus having attached itself to the ACE2 receptors in the lower respiratory tract, where it causes deep pneumonia,” Sørensen told Minerva. 

The Angiotensin-converting enzyme 2 (ACE2) is a protein found on the surface of human cells, as well as in soluble form in the blood, that has been identified as the receptor for the SARS-CoV-2 viral entry.

Sørensen says the SARS-CoV-2 spike protein is very stable and this points to it being “a fully developed, almost perfected virus for infecting humans”. This, he says, indicates that the structure of the virus cannot have evolved naturally. Sørensen also points to the inserts in SARS-CoV-2, several of which do not, he says, exist in other coronaviruses.

He told Minerva that it’s possible for a virus to attain these properties in nature, but it’s not likely.

“If the mutations had happened in nature, we would have most likely seen that the virus had attracted other properties through mutations, not just properties that help the virus to attach itself to human cells,” he said.

“What we see is that an area that you could observe in the first SARS coronavirus has been moved, so that the parts of the virus that are particularly well suited to attach to humans have become part of the spike protein that the virus uses to penetrate human cells.”

Sørensen told Minerva that the properties seen in SARS-CoV-2 had yet to be discovered anywhere in nature. If the virus came from nature, he says, there should also be many animals infected with it.

“The only place we are aware of where an equivalent virus to that which causes Covid-19 exists is in a laboratory,” Sørensen told Minerva. “So the simplest and most logical explanation is that it comes from a laboratory. Those who claim otherwise, have the burden of proof.”

In an article published in Independent Science News on June 2, Jonathan Latham and Allison Wilson, who are co-founders of the non-profit Bioscience Resource Project, which is based in Ithaca, New York, write: “Unfortunately, in the US at least, the question of the pandemic’s origin has become a political football; either an opportunity for Sinophobia or a partisan ‘blame game’.

“But the potential of a catastrophic lab release is not a game and systemic problems of competence and opacity are certainly not limited to China (Lipsitch, 2018). The US Department of Homeland Security (DHS) is currently constructing a new and expanded national Bio and Agro-defence facility in Manhattan, Kansas.”

The DHS has put the 50-year risk of a release from its laboratory at 70%, Latham and Wilson say.

Citing the comment of the Australian virologist Nikolai Petrovsky, from Flinders University, that no natural virus matching SARS-CoV-2 has been found in nature despite an intensive search to find its origins, Latham and Wilson said: “The idea of an animal intermediate is speculation. Indeed, no credible viral or animal host intermediaries, either in the form of a confirmed animal host or a plausible virus intermediate, has to date emerged to explain the natural zoonotic transfer of Sars-CoV-2 to humans (e.g. Zhan et al., 2020).”

Petrovsky (pictured left) was the lead author of a preprint paper published on the arXiv server on May 13.

The Flinders University scientists found that SARS-CoV-2 targeted humans more potently than any of the tested animal species.

They said: “The binding energy between SARS-CoV-2 spike protein and ACE2 was highest for humans out of all species tested, suggesting that SARS-CoV-2 spike protein is uniquely evolved to bind and infect cells expressing human ACE2.

“This finding is particularly surprising as, typically, a virus would be expected to have highest affinity for the receptor in its original host species, e.g. bat, with a lower initial binding affinity for the receptor of any new host, e.g. humans.

“However, in this case, the affinity of SARS-CoV-2 is higher for humans than for the putative original host species, bats, or for any potential intermediary host species.”

Latham and Wilson note that, in 2014, just before a ban on gain-of-function research went into effect in the US, the director of the Centre for Infectious Diseases at the Wuhan Institute of Virology (WIV), Shi Zhengli, did work with researchers from Ralph Baric’s laboratory in North Carolina, where gain-of-function research on bat coronaviruses was carried out, and published a paper.

The researchers combined the spike of the bat coronavirus SHC014 in a mouse-adapted SARS-CoV backbone into a single engineered live virus, Latham and Wilson wrote.

“The spike was supplied by the Shi lab. They put this bat/human/mouse virus into cultured human airway cells and also into live mice. The researchers observed ‘notable pathogenesis’ in the infected mice.”

Researchers in Shi Zhengli’s laboratory produced recombinant bat coronaviruses and placed these in human cells and monkey cells, Latham and Wilson note. “All these experiments were conducted in cells containing human or monkey ACE2 receptors,” they said.

US-China collaboration

The United States directly, and through collaboration with the non-profit EcoHealth Alliance, has funded Shi Zhengli’s research since at least 2014 and the EcoHealth Alliance has received further SARS-related funding from the US National Institutes of Health (NIH) dating back to 2008.

Zhengli (pictured left) has been referred to as the ‘bat woman’ because of her extensive work in the field of bat virology. She has publicly denied that the SARS-CoV-2 originated in her laboratory. In a WeChat post on February 2 she wrote: “… I promise with my life that the virus has nothing to do with the lab.”

The NIH funded the EcoHealth Alliance to surveil and understand the risks of the transmission to humans of SARS-related coronaviruses. That funding has now been withdrawn.

The director of the NIH’s National Institute of Allergy and Infectious Diseases (NIAID), Anthony Fauci, said the NIH cut off funding for the collaboration between the EcoHealth Alliance, which is based in New York, and researchers in Wuhan after president Donald Trump told it to.

The EcoHealth Alliance is also sponsored by the United States Agency for International Development (USAID), and NIAID . Its president is Peter Daszak, a British zoologist and an expert on disease ecology and zoonosis in particular. Daszak is on the board of the World Health Organisation (WHO).

While researchers from the EcoHealth Alliance, working with scientists from the Wuhan Institute for Virology, identified a sequence of SARS-related coronaviruses found in bats that was 96.11% similar to SARS-CoV-2 in July 2013, the sequence was not published on the NIH website until March this year.

Andre Watson, who is the founder and CEO of the regenerative medicine and pandemic defence biotechnology company Ligandal, which is based in San Francisco, says that a number of other coronaviruses have more than 60% sequence similarity to SARS-CoV-2 and they include viruses that cause neurological, vascular, gastric, and respiratory symptoms. Some of them have been studied for decades, “since the 1970s and earlier”, Watson says.

SARS from 2003 was only 80% similar to SARS-CoV-2 and MERS from 2012 was only 64.3% similar to SARS-CoV-2.

In 2017, the EcoHealth Alliance collaborated with the Wuhan Institute of Virology and the Duke-NUS Medical School in Singapore on research that was funded by the NIH, NIAID, the USAID Emerging Pandemic Threats programme (under the umbrella of the PREDICT project), and by funding sources in China.

The scientists published a paper about research done in bat caves. Researchers drew blood from the bats, or took nasal swabs or fecal samples. They identified 11 new viruses that they called SARS R (SARS-related viruses) in bats, Watson notes. Of these 11 viruses, three of them had spike proteins that had a high affinity for human ACE2.

The authors said: “In this study, we confirmed the use of human ACE2 as receptor of two novel SARSr-CoVs by using chimeric viruses with the WIV1 backbone replaced with the S gene of the newly identified SARSr-CoVs.”

Watson (pictured left) says this means that the researchers admit to doing recombination work with higher-binding ACE2 spike proteins on various SARS-CoV-2-related viral scaffolds.

“Furthermore, overlapping authors published a study in 2015 in which they admitted to discovering ‘SARSr-CoVs’ that had immunological cross-reactivity with SARS-CoV-1 antibodies,” he said.

The 2015 paper is the one Jonathan Latham and Allison Wilson refer to in their June 2 article. The paper’s authors include Shi Zhengli and Ralph Baric.

Watson said: “They have still not published the sequences of viruses taken from those infected with the virus. While they were able to detect these viruses, they also noted that SARS-CoV monoclonal antibodies have marginal efficacy against SARS-like CoVs.

“The authors directly admit to doing genetic engineering of the viruses, which means that they were taking bits and pieces of one type of virus and mixing it with bits and pieces of other clades and strains of virus.”

Vineet Menachery,  Shi Zhengli, Baric et al. said in their article that both wild-type and chimeric viruses were derived from either the epidemic SARS-CoV Urbani strain or the corresponding mouse-adapted (SARS-CoV MA15) infectious clone.

“Plasmids containing spike sequences for SHC014 were extracted by restriction digest and ligated into the E and F plasmid of the MA15 infectious clone,” they wrote.

Menachery et al. go on to explain that plasmids containing wild-type, chimeric SARS-CoV and SHC014-CoV genome fragments were amplified, excised, ligated, and purified.

“In vitro transcription reactions were then preformed to synthesise full-length genomic RNA, which was transfected into Vero E6 cells … The medium from transfected cells was harvested and served as seed stocks for subsequent experiments.”

Synthetic construction of chimeric mutant and full-length SHC014-CoV was approved by the University of North Carolina Institutional Biosafety Committee and the NIH’s Dual Use Research of Concern Committee, the authors said.

Watson explains that ACE2 is not only a receptor but is also a soluble protein that goes into the blood and floats around. “ACE2 is the main transport and uptake mechanism of SARS-CoV-2,” he said.

Watson has been collaborating with the University of California in San Francisco (UCSF) to demonstrate that soluble ACE2 is able to interfere with neutralising antibody binding to the coronavirus spike protein’s receptor binding domain, the portion of the virus that binds to ACE2. He will soon be publishing a paper about this work.

“ACE2 binds to the spike protein of the SARS-CoV-2 coronavirus extremely tightly, about 15 times more strongly than SARS-CoV-1 did,” Watson said.

“This means antibodies have a much harder time binding to this key site so as to inhibit viral binding to cells and to soluble ACE2.

“Neutralising antibodies would normally push away the ACE2 over time, but here ACE2 is super strongly binding, which decreases the amount of neutralising antibodies produced because the site is tightly ‘buried’.  Antibodies can’t stick and create a cloak and an immune-evasive coating on the virus.”

Watson says there is a combination of statistical thermodynamics and exponential cell division of the most strongly binding B cells during antibody generation.

“When a spot is inaccessible, this can decrease the probability of binding. If the site is accessible but buried, there will be instances of limited or nonexisting neutralising responses.

“We are certainly seeing tremendous variability in neutralising IgG antibody titers with many people not generating antibodies or sufficient antibodies. While other immune clearance techniques exist, this really is not good.”

Explaining further, Watson said: “You have the nucleocapsid and ORF polyproteins of SARS-CoV-2, which are on the inside; this binds the RNA and it holds it together. Then you’ve got the envelope and membrane proteins that interact with that lipid envelope on the virus.

“Finally, you’ve got the spike on the outside. The spike is what sticks to cells, and in this case, also inhibits visibility by neutralising antibodies because of the extremely high binding affinity of the viral spike protein for ACE2 in both soluble and membrane-bound forms.”

The EcoHealth Alliance researchers and their Chinese colleagues took an RNA strand from each of the viruses they identified and converted it to DNA, Watson says. They then put the DNA into a plasmid, where it could replicate.

“When you want to do genetic engineering, you don’t do it on RNA, you do it on DNA,” Watson explained. “A plasmid is stable. You can put it in the fridge or the freezer and put it in some cells and they’ll produce RNA.”

The researchers took spike proteins and combined them with envelope and nucleic acid proteins.

“Basically you’re holding the core of the virus stable, and you’re swapping out the pieces on the outside to get one that binds more strongly to human cells,” Watson said.

“They did not publish any of those sequences, but they admitted that they did that work.

“The nature of studying these viruses in cells is that there are going to be mutation events; evolutions are going to happen – especially if you are optimising the virus for infectivity of ACE2-expressing cells with human ACE2.”

The researchers also admitted to infecting humanised mice that had the human ACE2 receptor, Watson says. “It’s clear that they were doing gain-of-function studies.”

Watson says it is known that SARS-related viruses were infecting people in November 2017, or earlier, in rural China, and that these viruses were subsequently transferred to the Wuhan Institute of Virology.

“The funding trail certainly suggests that the USA has collaborated with China, the WHO, and others and had these projects funded since 2011 or earlier,” he said. “This was a joint, multinational effort.”

Watson added: “In rural China people living near bat caves were catching these ‘SARS-related viruses’ and were getting acute respiratory distress.

“The researchers admitted that they drew the blood of these people, collected the viruses, and studied whether or not those viruses were cross-reactive immunity wise with SARS-Cov-1. They were, but they weren’t SARS-Cov-1.

“Those sequences were never published. So, to this day, they did not show us what the sequences were that were part of that outbreak in 2017.”

The paper trails dating back to November 2017 or earlier show that humans were testing positive for a new SARS-related coronavirus, Watson says.

“The WHO, the United States, and China knew that this had occurred in November 2017, or earlier.

“Circulating bat coronaviruses evolved from SARS-CoV-1 (the 2003 outbreak) were known to have potential for human transmission in 2015, or earlier.”

Watson suspects that, in 2019, a SARS virus escaped from the Wuhan laboratory.

“There’s clear evidence that the first infections were not from people coming from the wet market in Wuhan, but that the virus was brought to the wet market.

“It’s awfully suspicious that Wuhan would be home to the only known biosafety level-4 research facility in China, and they worked on these viruses there.”

In the EcoHealth Alliance grant application for 2017, Daszak said the aim of the project was “to understand what factors increase the risk of the next CoV emerging in people by studying CoV diversity in a critical zoonotic reservoir (bats), at sites of high risk for emergence (wildlife markets) in an emerging disease hotspot (China)”.

He said: “Predictive models of host range (i.e. emergence potential) will be tested experimentally using reverse genetics, pseudovirus and receptor binding assays, and virus infection experiments across a range of cell cultures from different species and humanised mice.”

Writing in Science magazine on April 30, Meredith Wadman and Jon Cohen said that Daszak, with NIH vetting and approval, provided Zhengli with $599,000 out of a total of $3.1 million in grant funding to use lab sequencing techniques to identify bat coronaviruses that were at high risk of jumping to humans.

“The grant has also supported blood testing of people who live near bat caves in southern China to see whether they carried antibodies indicating they had been infected with a bat coronavirus,” Wadman and Cohen reported.

According to Wadman and Cohen, Zhengli and her colleagues collected some 15,000 biological samples in the field from bats. “In January, her team published the genetic sequence of a bat virus that shares 96.2% of its genome with SARS-CoV-2.”

On February 3, Shi Zhengli et al. published an article in Nature entitled ‘A pneumonia outbreak associated with a new coronavirus of probable bat origin’.

The researchers obtained full-length genome sequences from five patients at an early stage of the outbreak in Wuhan.

“The sequences are almost identical and share 79.6% sequence identity to SARS-CoV,” Zhengli et al. said.

The researchers explained their discovery that a short region of RNA-dependent RNA polymerase (RdRp) from a bat coronavirus (BatCoV RaTG13) – which was previously detected in Rhinolophus affinis from Yunnan province – showed high sequence identity to 2019-nCoV.

“Simplot analysis showed that 2019-nCoV was highly similar throughout the genome to RaTG13, with an overall genome sequence identity of 96.2 percent,” they said.

According to The Sunday Times in Britain, Daszak has confirmed that RaTG13 is the coronavirus discovered in an abandoned copper mine in Yunnan province in 2013, which was named RaBtCoV/4991 in a scientific paper published by researchers from the Wuhan Institute of Virology in 2016.

The SARS-CoV-2 genome and its spike glycoprotein show 96.11% and 92.86% identities to the Rhinolophus affinis bat coronavirus, respectively.

The director of the Wuhan Institute of Virology, Wang Yanyi, told the Beijing-based English-language news channel CGTN, in an interview published on May 25: “Many people might misunderstand that since our institute reported the RaTG13’s genomic similarity to SARS-CoV-2, we must have the RaTG-13 virus in our lab.

“In fact, that’s not the case. When we were sequencing the genes of this bat virus sample, we got the genome sequence of the RaTG13 but we didn’t isolate nor obtain the live virus of RaTG13. Thus, there is no possibility of us leaking RaTG13.”

Asked about speculation that SARS-CoV-2 leaked from the Wuhan Institute of Virology, Wang Yanyi said: “This is pure fabrication. Our institute first received the clinical sample of the unknown pneumonia on December 30 last year. After we checked the pathogen within the sample, we found it contained a new coronavirus, which is now called SARS-CoV-2.

“We didn’t have any knowledge before that, nor had we ever encountered, researched or kept the virus. In fact, like everyone else, we didn’t even know the virus existed. How could it have leaked from our lab when we never had it?”

Watson says that, throughout the progression of the SARS-CoV-2 pandemic, there have been consistent efforts by officials and the mainstream media to repress factual information about the severity, transmission dynamics, origins, and physiological effects of SARS-CoV-2 and Covid-19.

“On January 21, President Xi Jinping asked the director-general of the WHO, Dr Tedros Adhanom, to withhold information about person-to-person transmission of the virus, as well as pandemic classification. Likely as a consequence, pandemic classification of the virus was delayed four to six weeks.”

‘Escapes’ from laboratories

Concerns about gain-of-function research erupted in 2011 when a team at the University of Wisconsin in the US and researchers at a laboratory at the Erasmus Medical Centre in Rotterdam in the Netherlands announced that they had modified the H5N1 bird flu virus to enable it to spread between ferrets.

The researchers had planned to publish their findings in Science and Nature, but the National Science Advisory Board for Biosecurity in the US asked the journals to refrain from publishing the methods the scientists had used.

The advisory board issued a statement that was published in Science and Nature saying that a pandemic or the deliberate release of a transmissible highly pathogenic influenza A/H5N1 virus “would be an unimaginable catastrophe for which the world is currently inadequately prepared”.

The board added: “Our concern is that publishing these experiments in detail would provide information to some person, organisation or government that would help them to develop similar mammal-adapted influenza A/H5N1 viruses for harmful purposes.”

During the SARS outbreak in 2003 and 2004, there were two separate ‘escapes’ of viruses from the same laboratory in Beijing, China.

The WHO described one of the Beijing incidents, along with two other incidents (one in Singapore and one in Taiwan), as being attributed to “breaches in laboratory biosafety”.

China

In a report in October 2004, the WHO said that two researchers at the National Institute of Virology (NIV) in Beijing, where experiments using live and inactivated forms of the SARS coronavirus were being carried out, developed SARS in late March and mid-April of that year. The outbreak was reported on April 22 and the institute was closed a day later.

In one of the cases, a nurse who had cared for a researcher at the NIV became ill in April 2004 and was diagnosed as being infected with the SARS virus. The researcher, who had worked at the institute for two weeks in March 2004, developed symptoms on March 25, 2004, and was diagnosed as being infected with the SARS virus. The researcher’s mother also became ill and died on April 19.

Another researcher who also worked at the Beijing virology institute developed SARS symptoms on April 17 and was hospitalised. Health authorities diagnosed him as a suspected SARS case.

The WHO said that many shortcomings in biosecurity were found at the NIV and added: “The specific cause of the outbreak was traced to an inadequately inactivated preparation of SARS virus that was used in general (that is, not biosecure) laboratory areas, including one where the primary cases worked.

“It had not been tested to confirm its safety after inactivation, as it should have been.”

The WHO had said on May 18, 2004, that investigators had serious concerns about biosafety procedures at the Beijing institute – including how and where procedures using the SARS coronavirus were carried out, and how and where SARS coronavirus samples were stored.

The organisation added: “WHO and Chinese authorities view with concern the occurrence of laboratory-associated SARS cases. WHO urges all member states to view this latest outbreak as an opportunity to review the biosafety practices of institutions and laboratories working with SARS coronavirus.”

During and after the SARS outbreak of 2003, a large number of specimens were collected from possible human cases, animals, and the environment, the WHO said.

“These specimens, which may contain live SARS coronavirus, are still kept in various laboratories around the world. Some of them are stored in laboratories at an inappropriate containment level.

“SARS coronavirus has also been propagated in reference and research laboratories, and distributed to other laboratories for research purposes. Research using live and inactivated SARS coronavirus – and other pathogens capable of causing serious illness – is being conducted in many laboratories.”

Singapore

In Singapore, in August 2003, a microbiology student who had been doing research on the West Nile virus at the country’s national university became ill. He tested positive for the SARS virus. The student has also been doing work on West Nile at Singapore’s Environmental Health Institute (EHI) laboratory, where others researchers were studying the SARS virus.

The Center for Infectious Disease Research and Policy (CIDRAP) at the University of Minnesota in the US reported on the incident in September 2003 and said an international committee concluded that the researcher had probably acquired the virus in the EHI laboratory.

“Inappropriate laboratory procedures and a cross-contamination of West Nile virus samples with SARS coronavirus in the laboratory led to the infection of the doctoral student,” the Singapore Ministry of Health said. “No evidence could be found of any other source of infection.”

The ministry said the committee determined that there was no evidence that the researcher, who recovered from his illness, transmitted the virus to anyone else.

CIDRAP reported that the international committee that investigated the incident also examined Singapore’s four laboratories rated as biosafety level 3 ((BSL-3), the second highest of four risk categories.

“The group found structural problems as well as training and record-keeping deficiencies at the environmental health lab and recommended that the lab not reopen until the problems are corrected,” CIDRAP reported. “Lesser problems were found at the Singapore General Hospital laboratory and the National University of Singapore laboratory.”

Taiwan

In December 2003, a scientist from Taiwan who had been doing research on SARS fell ill on a flight when returning from a meeting in Singapore and was diagnosed as positive for SARS infection. His 74 contacts in Singapore were quarantined, and none developed SARS. It was alleged in one press report that the scientist had handled leaking biohazard waste without gloves, a mask, or a gown.

Calls for augmented security

On December 18, 2003, the WHO called for augmented biosafety in laboratories where SARS-CoV specimens and cultures were being handled.

The WHO said it strongly recommended biosafety level 3 as the appropriate containment level for working with live SARS-CoV material.

“The possibility that a SARS outbreak could occur following a laboratory accident is a risk of considerable importance, given the relatively large number of laboratories currently conducting research using the SARS-CoV or retaining specimens from SARS patients,” the WHO said.

“These laboratories currently represent the greatest threat for renewed SARS-CoV transmission through accidental exposure associated with breaches in laboratory biosafety.

“Given the severity of the threat, WHO strongly recommends that national governments maintain a registry of laboratories that are approved to safely and securely hold and work with specimens of suspected or confirmed SARS patients or cultures containing SARS-CoV.”

Appropriate national authorities should provide guidelines for laboratories to catalogue and control the storage of cultures and specimens of SARS-CoV for periodic inspections, the WHO said.

The WHO also said it encouraged the destruction of unwanted or unneeded clinical and animal specimens that were suspected of, or were confirmed to contain, SARS-CoV, and/or of stocks of SARS-CoV that could not be kept under secure conditions.

In July 2018, the director of the biosafety laboratory at the Wuhan Institute of Virology, Yuan Zhiming, and Shi Zhengli published a paper in ScienceDirect entitled ‘Quality management in a high-containment laboratory’.

The Wuhan Institute of Virology was accredited in 2017 by the China National Accreditation Service for Conformity Assessment, the scientists noted.

Zhiming and Zhengli said, however: “Because no international dedicated standard exists for biosafety level 4 (BSL-4) laboratories, this paper explains the desire of the laboratory’s director to set up a quality management system (QMS) to accredit this first level 4 containment laboratory in China …”

They added: “There are two international standards currently available, the choice of which depends on whether it is a laboratory intended for medical diagnoses … or a laboratory for both medical and environmental purposes …”

Both standards are recognised by the WHO, Zhiming and Zhengli noted. “Nonetheless, none of them has specific requirements for biorisks, i.e. biosafety and biosecurity.

“Although the 2017 version of ISO/IEC 17025:2017 takes into account risks in a general sense (clause 8.5), the management is free to use either another standard or any guidance more appropriate for the laboratory’s activity.”

Zhiming and Zhengli said that accreditation of a biocontainment laboratory in accordance with an international standard was a real challenge.

In the 14 countries hosting one or several BSL-4 laboratories, very few such laboratories had been accredited according to an international standard because there was no suitable international standard for accrediting the activities carried out at these laboratories, they added.

Gain-of-function moratorium lifted

On December 19, 2017, federal officials in the US ended a moratorium that had been imposed three years earlier on the funding of gain-of-function research.

The head of the NIH, Francis S. Collins, said the research could only be done if a scientific panel decided that the benefits justified the risks.

Collins (pictured left) said that researchers would have to show that their studies were scientifically sound and that they would be done in a high-security lab.

The pathogen to be modified must pose a serious health threat, he said, and the work must produce knowledge that would benefit humans. In addition, there would have to be no safer way to do the research.

Collins said that the new regulations applied to any pathogen that could potentially cause a pandemic.

In October 2014, all federal funding had been halted for gain-of-function research on the flu virus and those causing Middle East respiratory syndrome (MERS) and SARS.

The NIH said that certain gain-of-function studies with the potential to enhance the pathogenicity or transmissibility of potential pandemic pathogens (PPPs) had raised biosafety and biosecurity concerns, “including the potential dual use risks associated with the misuse of the information or products resulting from such research”.

The White House Office of Science and Technology Policy launched a gain-of-function deliberative process to re-evaluate the potential risks and benefits associated with certain experiments.

During the process the government halted federal funding for gain-of-function studies that were aimed at enhancing the pathogenicity or transmissibility among mammals by respiratory droplets of influenza, MERS, or SARS viruses.

The National Science Advisory Board for Biosecurity (NSABB) finalised its recommendations on May 24, 2016. In January 2017, the US government released policy guidance for the review and oversight of research “anticipated to create, transfer, or use enhanced PPPs”.

A new framework was developed to guide funding decisions about proposed research involving pathogens with enhanced pandemic potential.

Writing in Lancet Infectious Diseases in February 2018 about the rescinding of the moratorium on gain-of-function experiments, Talha Burki referred to the suppression by the NSABB in 2011 of the two studies involving H5N1 viruses that had been modified to allow airborne transmission from ferret to ferret.

“They worried that malign actors could replicate the work to deliberately cause an outbreak in human beings,” Burki wrote.

The Department of Health and Human Services (HHS) issued guidelines for funding decisions on experiments likely to result in highly pathogenic H5N1 viruses transmissible from mammal to mammal via respiratory droplets, Burki reported. The guidelines were later expanded to include H7N9 viruses.

Burki reported that, in 2014, several breaches of protocol at US government laboratories brought matters to a head.

“The news that dozens of workers at the Centers for Disease Control and Prevention (CDC) might have been exposed to anthrax, that vials of smallpox virus had been left lying around in an NIH storeroom, and that the CDC had unwittingly sent out samples of ordinary influenza virus contaminated with H5N1, shook faith in the country’s biosafety procedures,” she wrote.

In July 2014 more than 200 scientists signed the Cambridge Working Group declaration in which they said that, for any experiment, the expected net benefits should outweigh the risks.

“Experiments involving the creation of potential pandemic pathogens should be curtailed until there has been a quantitative, objective and credible assessment of the risks, potential benefits, and opportunities for risk mitigation, as well as comparison against safer experimental approaches,” the scientists said.

“A modern version of the Asilomar process, which engaged scientists in proposing rules to manage research on recombinant DNA, could be a starting point to identify the best approaches to achieve the global public health goals of defeating pandemic disease and assuring the highest level of safety.

“Whenever possible, safer approaches should be pursued in preference to any approach that risks an accidental pandemic.”

The scientists said that “recent incidents involving smallpox, anthrax and bird flu in some of the top US laboratories remind us of the fallibility of even the most secure laboratories, reinforcing the urgent need for a thorough reassessment of biosafety”.

Such incidents had been accelerating, the scientists said and had been occurring on average more than twice a week with regulated pathogens in academic and government labs across the country.

“An accidental infection with any pathogen is concerning. But accident risks with newly created ‘potential pandemic pathogens’ raise grave new concerns.

“Laboratory creation of highly transmissible, novel strains of dangerous viruses, especially but not limited to influenza, poses substantially increased risks. An accidental infection in such a setting could trigger outbreaks that would be difficult or impossible to control.

“Historically, new strains of influenza, once they establish transmission in the human population, have infected a quarter or more of the world’s population within two years.”

HIV assertion

French scientist Luc Montagnier, who was a joint winner of the 2008 Nobel Prize for discovery of the human immunodeficiency virus (HIV), has caused controversy with his assertion that some nucleotide sequences of HIV-1 have been found in the SARS-CoV-2 genome. Other scientists have challenged Montagnier’s assertion, saying that each of these sequences also appears in other viruses.

Montagnier (pictured left) also puts forward the theory that SARS-CoV-2 came from the Wuhan laboratory, escaping in an “industrial accident” when Chinese scientists were attempting to develop a vaccine against HIV.

Montagnier says that the sequences he asserts are HIV inserts must have been added to SARS-CoV-2 and that this could not have happened naturally. It is meticulous, professional work, Montagnier says.

Detractors say that a tiny bit of the SARS-CoV-2 genome is about 85% similar to part of the HIV-1 genome, but that the sequence can be found in other viruses.

A paper by Indian scientists who said there were four unique inserts in the 2019-nCoV (SARS-CoV-2) spike glycoprotein that were not present in any other coronavirus reported to date was retracted by the authors.

The paper had been published as a preprint on bioRxiv on January 31. It was stated on bioRxiv that the authors intended to revise the paper “in response to comments received from the research community on their technical approach and their interpretation of the results”.

Prashant Pradhan et al. had said the inserts were either identical or similar to the motifs in the highly variable (V) regions (V1, V4 and V5) in the envelope glycoprotein or in the Gag protein of some unique HIV-1 strains from three different countries (Thailand, Kenya and India).

They speculated that these motif insertions sharing similarity with HIV-1 proteins could provide an enhanced affinity towards host cell receptors and increase the range of host cells of 2019-nCoV. The study implied that 2019-nCoV might be generated by gaining gene fragments from the HIV-1 genome.

In a paper published on February 14 on PubMed Central (PMC), Chuan Xiao et al. discuss the Indian scientists’ assertions and their own examination of the sequences of 2019-nCoV, other CoV viruses and HIV-1 as well as the GenBank database.

Chuan Xiao et al. say their results demonstrate no evidence that the sequences of the four inserts are HIV-1 specific or that 2019-nCoV obtained these insertions from HIV-1.

“First, the results of blast search of these motifs against GenBank shows that the top 100 identical or highly homologous hits are all from host genes of mammalian, insects, bacterial and others,” Chuan Xiao et al. said.

“There are only a few hits on coronaviruses, but none of them are HIV-1 related.”

Chuan Xiao et al. say the insertion sequences in question exist widely in all kinds of viruses.

“While the 100% match between the insertion 1 and 2 sequences and the HIV sequences were found in 19 entries, the matches between the insertion 3 and 4 sequences and HIV-1 sequences were rather poor (from 42% to 88%).

“Sequences that completely match the insertion 3 and 4 sequences were not found in any HIV-1 sequences. This clearly shows that these insertion sequences are widely present in living organisms including viruses, but not HIV-1 specific.”

Evidence of possible manipulation

Andre Watson explains that there is a way that small sequences of HIV may have been inserted into SARS-CoV-2 that would have made the manipulation less noticeable. There could, he says, have been a forensic cover-up.

Watson says it’s very odd that all four of the inserts in question have been found on the surface of the SARS-CoV-2 spike protein.

“One thing that has made this forensically difficult to investigate is that the genomic sequences don’t match. If you line up the RNA letters in those inserts with HIV, they don’t match.

“However, there is something called sense mutation. If I were trying to put a HIV sequence into something, and my job was to cover it up, I wouldn’t use the same genetic sequence, I would get the same protein sequence. I would change the genetic sequence.”

Three DNA or RNA letters create one amino acid, but there are multiple combinations of three that will create a given amino acid, Watson says. “So, you can scramble the genetic letters around and still get the same protein letters to come out.”

Two of the domains that have been found on the surface of the SARS-CoV-2 spike protein are also present in the HIV-1 gp120 surface domain, which, Watson says, is part of HIV’s “viral fusion machinery with CD4+T cells”. (T cells, also called T lymphocytes, are a type of white blood cell. They are an essential part of the body’s immune system.)

Watson says gp120 is known to interfere with dendritic cell function during HIV infections, and also modulates the activity of CD4+ T cells, which interact with dendritic cells to sense and respond to various pathogens.

“While I have yet to see structural modelling data of these domains interacting with known HIV binding motifs and this isn’t damning in and of itself, it does raise several questions, especially when coupled to the known immune-evasive properties of SARS-CoV-2, which includes T cell exhaustion and antibody avoidance in a number of patients,” Watson said.

Watson says he hasn’t a clue what the gp120 domains are doing, “but they are there”.

It’s also very strange, Watson says, to see a malaria sequence on the surface of the SARS-CoV-2 spike protein.

He says the malaria sequences don’t match the known binding domain of malaria to CD147 (malaria’s entry receptor into red blood cell precursor cells and red blood cells).

“However, there is speculation and computational modelling suggesting that SARS-CoV-2 also binds to CD147 and, in our own experiments, we have preliminarily seen some binding responses in the ~100–300 nanomolar range, which need to be further validated.

“What are the odds, if there was not laboratory manipulation, of having four little inserts that match protein sequences from HIV on solvent-accessible domains on the surface of the spike, along with a malaria sequence?”

Watson says he doesn’t want to make claims that cannot be proven, but the presence of the four inserts, and the malaria sequence, need to be investigated – and “certainly line up with the gain-of-function work that is suggested by the plasmid recombination experiments and insertion of higher binding affinity ACE2-binding spike proteins into infectious clones”.

It is important, Watson says, to remember that American and Chinese researchers were doing genetic engineering work in 2015, and admitted that they were doing it.

When a virus is randomly evolving, there is usually a fixed ratio of change if there is mutation, Watson says.

“In the case of SARS-CoV-2, when compared to the 2013 RaTG13 virus that was 96.11% similar to the current outbreak, there is an entire region where the virus has mutated in a way that is statistically impossible,” he said.

“It is virtually impossible for these particular mutations to have happened naturally. And this has happened in the same region of the virus –  the spike protein – on which American and Chinese researchers have admitted doing recombination work.

If SARS-CoV-2 were mutating accidentally, it would have a constant ratio of sense and missense mutations that is typically 5:1, Watson says.

“However, for nearly 2,100 RNA letters, there are virtually no missense mutations, as would be expected naturally between two viruses that are experiencing some form of genetic drift.

“The RNA letters are changing, but most of the protein letters are not – and it’s just in the part of the virus that was known to be spliced around in the SARS-related viral recombination experiments.”

The SARS-CoV-2 research was done in collaboration between the US and China, as well as other global actors, Watson points out.

“To say that we didn’t know what this was when it appeared in December is to outright lie about biological research that we knew was happening. We knew what SARS-CoV-1 did in 2003.”

In a paper published on the pre-print repository ViXr.org in May, independent researcher Murat Seyran from Vienna says that the host tropism (the infection specificity of certain pathogens to particular hosts and host tissues) and the infection pattern of SARS-CoV-2 have three fundamental differences compared to the previous six human pathogenic coronaviruses.

“The unnatural flat pattern of SARS-CoV-2 S protein NTD [N-terminal domain] is conflicting with the evolutionary host tropism strategy of not only the human CoVs but also many different human pathogenic viruses,” Seyran said.

Why have we not seen any pandemic caused by coronaviruses before? Seyran asks. Why did pandemics not emerge in places where people rely on water sources shared with bats or bats are consumed as bushmeat?

If SARS-CoV-2 is not an engineered Bat CoVs RaTG13, its unnatural host tropism pattern and pandemic potential as compared to other human pathogenic coronaviruses raises those questions, he says.

Seyran also says that, in the case of SARS-CoV-2, the S Protein receptor-binding domain (RBD) is not a high-frequency positive selection site, unlike in other coronaviruses.

The SARS-CoV-2 genome is almost identical to the bat coronavirus, but it is only mutated on the RBD, Seyran says. “Why only the RBD had mutations meanwhile the rest of the genome was almost unaltered?”

It is argued that the presence of a furin cleavage spike in SARS-CoV-2’s spike protein is evidence that the virus did not develop naturally.

The international director of Regeneration International, André Leu, explained in a recent article that a furin cleavage site is a segment of four amino acids that enables a virus to use furin in the human body as an enzyme to dissolve its coating so that it can release its genetic material to infect cells.

Furin cleavage sites tend to be more infectious than cleavage sites that use other enzymes, Leu explained.

“The virus that caused the SARS epidemic, SARS-CoV, in 2002–2003 does not have a furin cleavage spike, so it was not as infectious as SARS-CoV-2,” Leu wrote.

“There are several published papers showing that this four-amino acid sequence furin cleavage site is missing from the closest relatives of SARS-CoV-2 and has been inserted in precisely the best place in the spike protein to give it the ability to become highly infectious.

“It is highly improbable that this furin cleavage site has evolved naturally given that there is no sign of it evolving in any of SARS-CoV-2’s relatives in its clade.”

‘Proximal origin’ article challenged

In a controversial article published on March 17 in Nature Medicine, entitled ‘The proximal origin of SARS-CoV-2’, Kristian G. Andersen et al. say it is improbable that SARS-CoV-2 emerged through laboratory manipulation of a related SARS-CoV-like coronavirus.

“Our analyses clearly show that SARS-CoV-2 is not a laboratory construct or a purposefully manipulated virus,” the researchers said.

“It is improbable that SARS-CoV-2 emerged through laboratory manipulation of a related SARS-CoV-like coronavirus.”

Andersen et al.’s findings have been challenged, however. Stuart Newman, who is a professor of cell biology and anatomy at New York Medical College, is quoted by GMWatch as saying that a key argument used to deny that SARS-CoV-2 could be a genetically engineered strain that escaped from a laboratory actually points to the exact opposite. In other words, he is quoted as saying, it indicates that SARS-CoV-2 could well be genetically engineered and that it could have escaped from a lab.

Andersen et al. say that, if genetic manipulation had been performed, one of the several reverse-genetic systems available for betacoronaviruses would probably have been used.

The genetic data irrefutably show that SARS-CoV-2 is not derived from any previously used virus backbone, they say.

Andersen et al. propose two scenarios they say can plausibly explain the origin of SARS-CoV-2: natural selection in an animal host before zoonotic transfer, and natural selection in humans following zoonotic transfer.

The researchers say the high-affinity binding of the SARS-CoV-2 spike protein to ACE2 “is most likely the result of natural selection on a human or human-like ACE2 that permits another optimal binding solution to arise”.

They say that, as many early cases of Covid-19 were linked to the Huanan market in Wuhan, it is possible that an animal source was present at this location.

“Given the similarity of SARS-CoV-2 to bat SARS-CoV-like coronaviruses, it is likely that bats serve as reservoir hosts for its progenitor,” they stated.

They add that Malayan pangolins (Manis javanica) illegally imported into Guangdong province contain coronaviruses similar to SARS-CoV-2.

Although the RaTG13 bat virus remains the closest to SARS-CoV-2 across the genome, some pangolin coronaviruses exhibit strong similarity to SARS-CoV-2 in the receptor-binding domain, the researchers say. This includes all six key RBD residues, they wrote.

According to Andersen et al., this clearly shows that the SARS-CoV-2 spike protein optimised for binding to human-like ACE2 is the result of natural selection.

It is possible, the researchers say, that a progenitor of SARS-CoV-2 jumped into humans, acquiring the genomic features described in their analysis through adaptation during undetected human-to-human transmission.

The researchers say that, although the evidence shows that SARS-CoV-2 is not a purposefully manipulated virus, it is currently impossible to prove or disprove the other theories of its origin described in their article.

However, they say, since they observed all notable SARS-CoV-2 features in related coronaviruses in nature, they do not believe that any type of laboratory-based scenario is plausible.

In an email interview with GMWatch, Stuart Newman (pictured left) said: “The Nature Medicine paper points to variations in two sites of the spike protein of the new coronavirus that the authors claim must have arisen by natural selection in the wild.

“However, genetic engineering of one of these sites, the ACE2 receptor binding domain, has been proposed since 2005 in order to help generate vaccines against these viruses. It is puzzling that the authors of the Nature Medicine commentary did not cite this paper, which appeared in the prominent journal Science.”

Newman also told GMWatch: “The second site that Andersen et al. assert arose by natural means, a target of enzyme cleavage not usually found in this class of viruses, was in fact introduced by genetic engineering in a similar coronavirus in a paper they do cite. This was done to explore mechanisms of pathogenicity.”

He added that he does not believe that changes were deliberately introduced to increase the pathogenicity of any single strain of SARS-CoV-2, but that the virus may have had genetically engineered components in its history before it was inadvertently introduced into the human population.

GMWatch also quoted the London-based molecular geneticist Michael Antoniou, who has also cast doubt on assertions that SARS-CoV-2 was not genetically engineered. Antoniou said that Andersen et al.’s reasoning was not conclusive because it was based largely on computer modelling, which, Antoniou says, is “not definitive but only predictive”.

Antoniou said that, while Andersen et al. may be correct in how they perceive SARS-CoV-2 to have arisen, the data they present “does not exclude the possibility that this new coronavirus variant could have been created through an in vitro, directed iterative evolutionary selection process”.

He added: “Using this method, a very large library of randomly mutagenised coronavirus spike proteins could be selected for strong binding to the ACE2 receptor and consequently high infectivity of human cells.

“The power of such directed evolution to select for optimal enzymatic and protein-protein interactions was acknowledged by the award of the Nobel Prize in Chemistry in 2018.”

GMWatch states: “Neither Dr Antoniou, nor Prof Newman, nor we ourselves make any suggestion that, in the event that genetic engineering was involved, the intention was to create a bioweapon. Such ‘enhanced infectivity’ research is carried out on viruses all over the world (and not just in China) to investigate their behaviour and to develop vaccines and other therapies, as well as for ‘biodefence’ purposes.

“But the question of whether genetic engineering did play a part in the emergence of SARS-CoV-2 must continue to be investigated so that humanity can place appropriate limits and safeguards on such research.”

The race to produce a vaccine

More than a dozen Covid-19 vaccines are now being tested on humans.

Two of the main vaccines in development are the mRNA-1273 vaccine being produced by the American biotech company Moderna and the one being developed by Oxford University’s Jenner Institute, working with the Oxford Vaccine Group.

Testing of a vaccine against SARS-CoV-2 is set to start in Australia, where more than one hundred people in Melbourne and Brisbane have volunteered to take part in the clinical trials.

The Nucleus Network Clinical Research Organisation is working with the US-based biotechnology company Novavax and is set to begin phase one testing of the NVX-CoV2373 vaccine.

Globally, there are dozens of companies and institutions involved in the race to produce a vaccine against SARS-CoV-2. These include 14 projects in India.

The Oxford vaccine

The Oxford University vaccine, originally named ChAdOx1 nCoV-19 and now known as AZD1222, is being tested in South Africa, including on people with HIV.

There have been protests against the vaccine trials, with demonstrators saying they don’t want to be used as guinea pigs.

The Oxford researchers say that, of the two thousand people being enrolled into the trial in South Africa, 1,950 are HIV-negative and 50 are people living with HIV.

Previously, the developers of the Oxford vaccine had said that no saline placebo would be used in the Phase II and Phase III trials and that participants would receive one or two doses of either the ChAdOx1 nCoV-19 vaccine or the MenACWY meningitis vaccine, Nimenrix, which would be used as a “control”.

They say that, in the trials in South Africa, a saline placebo will be used.

AZD1222 is also being tested in the UK, where more 4,000 participants are already enrolled into the clinical trial and enrolment of an additional 10, 000 participants is planned.

The vaccine is also being tested in Brazil. The trial began on June 20 and involves 5,000 volunteers across the country, in Sao Paulo, Rio de Janeiro, and at a site in the northeast of Brazil.

AZD1222 is made from a virus called ChAdOx1, which is a weakened and non-replicating version of a common cold virus (adenovirus) that has been engineered to express the SARS-CoV-2 spike protein.

The chairman and president of the non-profit organisation ACCESS Health International, William Haseltine, points out that, in the trials of the Oxford vaccine, none of the monkeys who were vaccinated were protected against SARS-CoV-2. He told CNBC: “Yes, they didn’t have as much virus in their lungs, but they still had a nasal infection. A hundred percent of the vaccinated monkeys got infected.”

The Oxford University researchers are working in collaboration with the pharmaceutical company AstraZeneca.

AstraZeneca has already entered into agreements with the Coalition for Epidemic Preparedness Innovations (CEPI), the vaccine alliance Gavi, and the Serum Institute of India (SII) to manufacture, procure, and distribute 300 million doses of the AZD1222 vaccine.

The company said in a statement on June 4 that it had reached a $750 million agreement with CEPI and Gavi to support the manufacturing, procurement, and distribution of 300 million doses of the vaccine, with delivery starting by the end of the year.

“In addition, AstraZeneca reached a licensing agreement with SII to supply one billion doses for low and middle-income countries, with a commitment to provide 400 million before the end of 2020,” the company added.

AstraZeneca says its global supply capacity for the AZD1222 vaccine will exceed two billion doses.

The company has also agreed to supply 400 million doses of AZD1222 to the US and the UK.

It said: “AstraZeneca recognises that the vaccine may not work but is committed to progressing the clinical programme with speed and scaling up manufacturing at risk.”

In its description of the vaccine, AstraZeneca says: “It uses a replication-deficient chimpanzee viral vector based on a weakened version of a common cold (adenovirus) virus that causes infections in chimpanzees and contains the genetic material of SARS-CoV-2 spike protein.”

Moderna

There was massive share dumping by Moderna’s chief medical officer, Tal Zaks, and other top executives in the company before Moderna released a press release about its Covid-19 vaccine on May 18.

Three of the 15 people in the high dose (250 microgrammes) cohort of the mRNA-1273 trial suffered adverse effects within 43 days of receiving Moderna’s vaccine.

Moderna describes the effects as “grade 3 systemic symptoms, only following the second dose”.

The company says the 250-microgramme dose is being eliminated from future studies, “not so much because of the side effects, but because the lower doses appeared to work so well that the high dose is not needed”.

Moderna said its findings were based on results from the first eight people who each received two doses of the mRNA-1273 vaccine, starting in March.

The company said it was now proceeding on an accelerated timetable, with the next phase involving six hundred people to begin soon.

Moderna says that additional tests in mice that were vaccinated and then infected showed that its vaccine could prevent SARS-CoV-2 from replicating in their lungs. The company says the animals had levels of neutralising antibodies comparable to those in the people who had received the vaccine.

Critics of the way Moderna has presented its trial information point out that the company has not released its clinical trial study or raw data.

William Haseltine has criticised Moderna for putting out a press release without adequate data, and affecting its share prices. Moderna’s announcement, he says, was premature as only eight people had been studied. “It was not impressive and it was opaque,” Haseltine told CNBC television.

China

In China, the Covid-19 vaccine Ad5-nCoV has been approved for use for one year by the military without phase three trials being conducted. The same vaccine is being tested in Canada.

The news agency Reuters reports that the vaccine was developed jointly by CanSino Biologics and a research institute at the Academy of Military Science (AMS).

CanSino Biologics said that clinical trials had shown the vaccine to be safe and indicated some efficacy.

Ad5-nCoV is one of eight Covid-19 vaccines being developed in China that have been approved for human trials at home and abroad, Reuters reported. Five of the vaccine projects are already at the stage of human trials.

Beijing-based Sinovac Biotech, which developed one of the vaccines being tested, told the Agence France Presse news agency that it was looking to carry out the final stage of its trial abroad because China does not now have a large enough Covid-19 cluster.

Writing for GlobalData Healthcare on June 23, Reynald Castaneda said that CanSino Biologics’ Ad5-nCoV and Johnson & Johnson’s AdVac platform-based vaccine use a human adenovirus vector, “but a significant chunk of people may already have neutralising antibodies against the vector, decreasing efficacy prospects”.

He added that “Phase I Ad5-nCoV data is also underwhelming, adding credence to the issue of pre-existing antibodies”.

AstraZeneca’s AZD1222 and Rome-based ReiThera’s Covid-19 vaccines are also adenovirus vectored but use nonhuman vectors, Castaneda reported.

“However, AZD1222’s recent animal data also leave questions about its utility to prevent virus spread. A possible way to improve efficacy is to add a booster shot down the line, perhaps using a different adenovirus vector or even a different vaccine technology. Perhaps AZD1222 only carrying SARS-CoV-2’s spike protein may not be enough.”

Concerns about spike protein vaccines

Atomic-level structure of the spike protein of SARS-CoV-2. Credit: McLellan Lab, University of Texas at Austin.

There are concerns that there will, with vaccines against SARS-CoV-2, be pathogenic priming, also known as disease enhancement. This happened with vaccines against SARS.

Research scientist James Lyons-Weiler wrote in an article published in March: “In SARS, a type of ‘pathogenic priming’ of the immune system was observed during animal studies of SARS spike protein-based vaccines.

“The exposure of vaccinated animals to the SARS virus led to increased morbidity and mortality. The problem, highlighted in two studies, only became obvious following post-vaccination challenge with the SARS virus.”

Lyons-Weiler added: “SARS-CoV-2 is the sister taxon of SARS-CoV. If pathogenic priming is to occur in humans given spike-protein based SARS-CoV-2 vaccine, as is expected given the SARS spike protein animal studies, the 20% mortality rate expected in the elderly could raise to 40% – and the rest of the population could be sensitised and we could see mortality rates worldwide of the next coronavirus higher than 20%.”

Andre Watson says that old-fashioned vaccines would be more effective than the ones being developed in the US.

“I have raised flags about this to The Biomedical Advanced Research and Development Authority, the HHS, and the White House to no avail.

“It’s too early to know for certain, but I am extremely worried about the potential for reactivation and six- to 12-month delayed reinfection after neutralising antibody counts reduce, especially as social distancing measures are dismantled and viral community load increases.”

Watson says that neutralising antibody counts will decline over time, leading to a potential for reinfection. “Strongly responsive, retained antibodies may lead to enhanced pathogenicity of repeat infections,” he said

Watson says that antibody-dependent enhancement (ADE), in which a person’s body pumps out antibodies that bind to the virus but don’t neutralise it, and vaccine-associated enhanced respiratory disease (VAERD) are a concern with SARS-Cov-2 and Covid-19.

“With spike protein vaccines, there may be even more drift over time towards antibody-dependent enhancement and off-target antibody generation that can enhance disease.

“This has been observed before with spike protein vaccines and SARS-CoV-1 and MERS-CoV, in the sense of vaccine-associated enhanced respiratory distress.”

There is a real possibility that some or many vaccines, especially spike protein vaccines, may make infection worse, especially if neutralising antibody titers decline while off-target antibodies remain highly produced, Watson says.

“This would be made possible by the burying of the spike protein neutralising antibody binding site by ACE2, which binds extremely strongly and we have demonstrated in our lab is capable of inhibiting antibody binding to this site.”

The main focus for most of the companies currently developing Covid-19 vaccines is working on spike protein vaccines, whether they be administered as recombinant proteins, mRNA, or DNA.

In China, however, some researchers are pursuing development of a live attenuated vaccine, and other companies, such as Novavax, are working on virus-like nanoparticles.

Watson’s company is taking a different approach, developing peptides that mimic just the very tip of the spike protein and are showing early results of being able to inhibit viral binding to the ACE2 receptor while stimulating an immune response.

“Ligandal’s drug is intended to be both a prophylactic and a therapeutic,” Watson said. “We are at the preclinical stage of developing an antidote-vaccine and are studying the immune effects of its use before and after infection.”

In the case of Ad5-nCoV, an adenovirus has been used as a vector to deliver the DNA for the coronavirus protein. In other cases, the adenovirus virus is modified (pseudotyped) to produce the viral spike protein on its own surface.

“In this case, the coronavirus protein is expression based, that is it is produced by a gene,” Watson said. “The idea is to generate protein antigens that will call up antigens to SARS-CoV-2.”

In Watson’s view, it is the “live attenuated” and “virus-like particle” approaches that are most likely to be successful, “or whatever presents the spike protein on the surface facing the right way, just like the virus does”.

In the case of SARS-CoV-2, there are about 100 spikes per virus, Watson says. “The latest mutant has an even higher density. Each of the spikes is pointing in a very specific direction and only the very tip of the spike sticks to the ACE2 receptor and should be bound by neutralising antibodies.

“If you cut off the spike and just throw it into circulation, it will face random directions and you may develop many of the wrong antibodies preferentially. The neutralising ones may decline disproportionately to other epitopic, or immune binding, sites.”

Normally, with pretty much every other virus, you’ll get an antibody that will stick to the outside of the virus, and it will flag the virus for destruction, Watson explains.

“Once your body has those antibodies; the moment the virus is in your blood, those antibodies are going to stick to it and the virus is going to get eliminated from your blood very rapidly.

“If you have antibodies against typical viruses, ninety per cent of the virus would be gone from your blood in about 15 minutes.”

Watson says this is seen with a large number of adeno-associated virus patients who have pre-existing immunity to the most commonly used viruses in gene therapy.

“The problem here is that the virus is actually sticking to ACE2 more strongly than immature antibodies, and more strongly than immature B cells and equally as strongly as even good antibodies are.

“It binds with about 700 picomolar affinity to ACE2. This is a greater binding strength for a receptor than anything I have ever seen.”

Watson says that SARS-CoV-2 has evolved to bind to the ACE2 receptor about 15 times more strongly than SARS-CoV-1 did.

He explains further: “You want an antibody that sticks to the very portion of the tip of the spike that would normally stick to the human protein. If you get antibodies on the side of it, they may help to eliminate it, but they can also make the problem worse through antibody dependent enhancement.

“Even in the absence of that, you can get vaccine-induced worsening of effects if you’re exposed to the actual pathogen.”

Watson says the spike protein vaccines are of concern to him because of the issue of immune shielding (the ability of the virus to shield itself from a person’s immune system).

“All five of the vaccines that the US has put $2 billion into through BARDA, which are the Moderna RNA vaccines and also some viral-based vaccines, they all produce a spike protein; they’re all going to have the same issue. I have a little bit more hope for Novavax because at least their spike protein is presenting the right way.

“They’re all going to lead to strong immune responses, but may not create quite the right immune response.”

Watson is also troubled by the fact that BARDA has suspended its funding both for immunomodulators and therapeutics that “improve the clinical response and/or resolution of symptoms associated with … viral respiratory infections” and for pre- and post-exposure prophylactic approaches.

Watson’s approach to dealing with a virus is to make a smaller portion of the spike protein that’s just the very tip. “The portion that they are making, it’s 1,200 amino acids. I’m only making 70 to 90.”

“It still exhibits the viral epitope for creating antibodies, but it only shows the portion that you want to have a neutralising antibody against, and also actively inhibits ACE2 from binding to the cloaking site.

“It doesn’t show any off-target binding sites for antibodies and unwanted immune responses, and also can programme T cell epitope responses.”

With the approach that’s being taken by the big pharmaceutical companies, Watson says, the strongest antibody responses are likely being generated against the sides of the spike protein, not against the tip, because these spike proteins exhibit the same ACE2 binding and immune-shielding effect.

“There are also questions about whether the large spikes have the same sugar modifications (glycosylations) as fully formed spike proteins from whole viruses, which can also affect which regions of the spike the immune system will respond to,” Watson said.

It’s better, Watson says, to only provide the very tip of the spike protein as a vaccine or antidote.

“The ACE2 receptor would then bind to the antidote protein rather than the virus if you’re already infected, and, if you haven’t been infected, this may saturate all the cloaking sites for ACE2 while also serving as a potential vaccine adjuvant to other spike protein vaccines.

“With our approach, the virus can be seen by your antibodies and your B cells and you can get a neutralising response. You can actually generate that response preferentially as opposed to generating a response against parts of the spike protein you don’t want to be targeted.

“When you get exposed to the actual virus, your body will recognise just the parts it needs to.”

In the case of Ligandal’s antidote-vaccine, “nano-scaffolds” are designed to generate ultra-specific antibody responses against just the antibody-binding motif of the virus, Watson says.

“You should be able to administer these nano-scaffolds before or after infection. If you’ve already been infected, the ‘antidote’ component will kick in, preventing ACE2 cloaking and enhancing your immune system’s ability to sense and build a response to the virus.”

Watson has applied to BARDA for funding for his antidote-vaccine project twice, but his applications were rejected. He is more hopeful about getting funding from the NIH and has support from private investors.

“Everything is being driven by politics and how do you make the most revenue. I’m genuinely concerned that those who have control over this don’t actually want the most efficacious treatments to necessarily be out there. That’s why I don’t give up.”

A spike protein of SARS-CoV-2 in ‘closed’ form (ACE2 shown in red). Photo courtesy of Andre Watson.

A spike protein of SARS-CoV-2 in ‘open’ form, in which it binds to ACE2. (Photo courtesy of Andre Watson.)

Francois Balloux, who is the director of the Genetics Institute at University College London, tweeted on June 30: “Most ‘hight-tech’ vaccines tend to focus on eliciting narrow antibody responses. T-cell response works best against the whole viral diversity. This raises the question whether there should not be more efforts towards ‘traditional’ vaccines (i.e. attenuated/inactivated).”

Restrictions and revenue generation

Watson is also concerned about the social requirements that will come once Covid-19 vaccines are made available.

“Credit scores, housing rental availability, the availability of a driver’s licence, and potentially even the ability to get on public transit could readily be dictated by whether or not you’ve already been infected, and whether or not you’ve received a vaccine,” he said.

“If a vaccine is required to get a driver’s licence and even to be able to return to work, people of a lower socioeconomic status will be unfairly targeted.”

Further power is being ceded during the Covid-19 pandemic, Watson says. “Biometrics will merge with other data we already gather on people.

“Hundreds of billions of dollars in recurring revenue can be generated through the Covid crisis in the USA alone, by implementing half measures to treat and prevent the disease.

“For example, remdesivir will get people in and out of the hospital faster without really saving lives.”

“Even if it does save lives, you have to give it to 28 people to save one.”

Gilead Sciences, which manufactures remdesivir, recently announced how much it will be charging for the drug. Media reports said the cost could amount to $5,720 per patient. Gilead said the price for US private insurance companies for a five-day treatment would be $3,120.

The company’s chairman and CEO, Daniel O’Day, said: “We have set a price for governments of developed countries of $390 per vial. Based on current treatment patterns, the vast majority of patients are expected to receive a five-day treatment course using six vials of remdesivir, which equates to $2,340 per patient.”

He added: “In the US, the same government price of $390 per vial will apply. Because of the way the US system is set up and the discounts that government healthcare programmes expect, the price for US private insurance companies will be $520 per vial.”

The Food and Drug Administration (FDA) in the US has not approved remdesivir as a treatment, but granted the drug emergency use authorisation in May.

The US has bought up virtually all of the global supply of remdesivir for the next three months.

The HHS announced on June 29 that it had secured more than 500,000 treatment courses of remdesivir from Gilead Sciences for US hospitals until September.

The stocks make up 100% of Gilead’s projected production for July and 90% for August and September.

Gilead announced on July 8 that it was starting a clinical trial of a nebulised, inhaled version of remdesivir that the company said may enable “more targeted and accessible administration in non-hospitalised patients and potentially lower systemic exposure to the drug”.

The company’s chief medical officer, Merdad Parsey, said about sixty people, aged 18 to 45, would be enrolled for the study.

“Additional clinical trials evaluating remdesivir in combination with anti-inflammatory medicines, in vulnerable patient populations and in outpatient settings are ongoing or planned to initiate in the near future,” Parsey added.

Waning antibodies

Francois Balloux has tweeted that antibody levels to SARS-CoV-2 wane fairly rapidly, in line with other coronaviruses. “As such, antibody tests can fail to detect prior infection in particular for asymptomatic or mild infections. Serological surveys may underestimate the proportion of people infected,” he wrote.

“T-cell response is distinct from antibodies. Antibody titres against SARS-CoV-2 seem to wane fairly fast. T-cell immunity acts late during infection. As such, it is unlikely to protect from (re-)infection but it is expected to reduce symptom severity.”

Balloux says that, in contrast to antibody-mediated immunity, T-cell response is extremely long-lived. “For example, people infected in 2003 with SARS-1 still mount an immune response to SARS-CoV-2 17 years later.

“While there is still no established case of SARS-CoV-2 reinfection to date, it is likely those will be observed soon due to fairly fast waning antibodies.”

A study from China published on June 8 in Nature Medicine indicates that people infected with Covid-19 lose antibodies after only a few months.

Quan-Xin Long et al. said their research showed that antibodies faded quickly in both asymptomatic and symptomatic Covid-19 patients during convalescence.

The researchers observed that IgG levels and neutralising antibodies in a high proportion of patients who recovered from SARS-CoV-2 infection started to decrease within two to three months after infection.

“In another analysis of the dynamics of neutralising antibody titers in eight convalescent patients with Covid-19, four patients showed decreased neutralising antibodies approximately six–seven weeks after illness onset,” they said.

The researchers said their data might indicate risks in using Covid-19 ‘immunity passports’ and supported the prolongation of public health interventions, including social distancing, hygiene, the isolation of high-risk groups, and widespread testing.

The researchers studied 37 asymptomatic and 37 symptomatic patients in the Wanzhou district. They found that more than 90% of the patients in both groups showed steep declines in levels of SARS-COV-2–specific immunoglobulin G (IgG) antibodies during the early convalescent phase (by 8 weeks after release from the hospital).

The decreases in IgG levels were 93.3% in the asymptomatic patients and 96.8% in those who were symptomatic.

The researchers also found declines in specific neutralising antibodies, but the declines were not nearly as large as with IgG antibodies, which are the longest-term antibodies that bind to pathogens and have “matured” to have higher binding affinity than their IgM precursors.

Decreases in the neutralising antibodies were seen in 81% of the asymptomatic patients and 62% of those who were symptomatic. The median decreases were 8.3% in the asymptomatic group and 11.7% in the symptomatic patients.

The median percentage of decrease in IgG levels was 71.1% in the asymptomatic group and 76.2% in the symptomatic group.

Forty percent of the asymptomatic patients became negative for IgG antibodies and 12.9% of the symptomatic group became seronegative in the early convalescent phase.

“In addition, asymptomatic individuals exhibited lower levels of 18 pro- and anti-inflammatory cytokines. These data suggest that asymptomatic individuals had a weaker immune response to SARS-CoV-2 infection,” the researchers said.

Quan-Xin Long et al. cited a study of 191 Covid-19 patients that indicated that the median duration of viral shedding was twenty days. According to another study, the duration of viral shedding in nasopharyngeal aspirates was prolonged up to at least 24 days after the onset of symptoms in 18 patients infected with SARS-CoV-2 in Singapore.

The researchers said that, in their study, the median duration of viral shedding in 37 patients with mild symptoms was 14 days, which was shorter than reported by other scientists.

In comparison to symptomatic patients, the asymptomatic group had a significantly longer, 19-day duration of viral shedding.

“Several factors might contribute to the variation of duration of viral shedding in different studies, including the severity of disease, definition of duration of viral shedding and frequency of specimen collection,” Quan-Xin Long et al. said.

“Notably, detection of viral RNA does not necessarily mean that infectious virus is present in respiratory specimens, and caution is required when applying virus shedding duration that was calculated based on RT-PCR to assess infection potential.”

There are inaccuracies in results from the serology antibody tests, Watson says.

“Serological tests are not very good at telling if you have the virus. There are 13 to 50% false negatives. The false positive rate is 0.2 to 1.2% for most of them, including the Roche one. The false negative rates are higher due to the varying levels of antibodies and delay in producing antibodies.

“The Abbott test is 50% accurate. It’s like flipping a coin. Half the time that you have the virus it won’t tell you, but, if you don’t have it, it will be very good at telling you that you don’t have it.

“Statistically, in terms of the likelihood of someone having symptoms, it’s about the same as measuring a person’s temperature.”

Watson doesn’t understand why the US authorities delayed the testing process and import of PPE from other countries for two months, only to develop expensive tests that have up to 50% false-negative rates, when China could have provided the country with 500,000 test kits a day that were 87% accurate back in early March and were approved by the European Union and the China Food and Drug Administration (CFDA).

A divided society

Watson is dismayed to see the current division in America over how to deal with Covid-19.

“We basically have party lines, and a lot of baiting on each side. You’re either against abortion, pro-guns, anti-mask, and pro-Trump or you’re seen as a neoliberal, Antifa-supporting, mask-wearing libtard. There is very little middle ground and the media controls these narratives.

“There’s a lot of outrage and misinformation and PSYOPs and government-level intervention in our narrative; in how we think and how we speak and what we’re allowed to say. All this is driving a massive divide.”

1. A person suffering from Capgras delusion believes that a friend, spouse, parent, or other close family member (or pet) has been replaced by an identical impostor.

Update 17/7/2020

In a new article, Birger Sørensen, Angus Dalgleish, and Andres Susrud present detailed reasoning for their argument that the origin of SARS-CoV-2 is not natural.

“SARS-CoV-2 is possessed of dual action capability. In this paper we argue that the likelihood of this being the result of natural processes is very small,” the authors state.

“The spike has six inserts which are unique fingerprints with five salient features indicative of purposive manipulation.”

Sørensen, Dalgleish, and Susrud analyse four research projects which, they suggest, show by deduction “how, where, when, and by whom the SARS-CoV-2 spike acquired its special characteristics”.

They state: “This reconstructed historical aetiology meets the criteria of means, timing, agent and place to produce sufficient confidence to reverse the burden of proof.

“Henceforth, those who would maintain that the Covid-19 pandemic arose from zoonotic transfer need to explain precisely why this more parsimonious account is wrong before asserting that their evidence is persuasive, most especially when, as we also show, there are puzzling errors in their use of evidence.”

Sørensen, Dalgleish, and Susrud challenge the arguments put forward by Andersen et al. in their paper published in March.

The three scientists say the contention of Andersen et al. that it is improbable that Covid-19 emerged through laboratory manipulation of a related SARS-CoV-like coronavirus because the ACE2 binding is not ideal “is weakened because Andersen et al. cite two authorities which actually say the reverse of what they say that they say”.

In the course of their bio-chemical analysis, Sørensen, Dalgleish, and Susrud identified five features of SARS-CoV-2 that individually, they say, seem unlikely to be the result of natural evolution and which, taken together, make natural evolution a less likely explanation than purposive manipulation, “specifically for gain of function”.

Sørensen and his colleagues say that a major part of the SARS-CoV-2 spike protein has human-like domains with matured transmission adaption.

“Blasting the spike protein with a rolling window of six amino acids showed that 78.4% of six amino acid windows are human like,” they wrote.

Nearly 80% of the spike protein has a built-in stealth property by having high human similarity, they add.

“Therefore, it is remarkably well-adapted virus for human co-existence,” the scientists wrote. “Such high human similarity also implies a high risk for the development of severe adverse events/toxicity and even Antibody Dependent Enhancement (ADE) unless specific precautions are taken when using the spike protein in any vaccine candidate.”

Sørensen and his co-authors say their discovery that the SARS-CoV-2 spike displays new amino acid inserts “with condensed cumulative charge, all of which are surface exposed” is particularly significant.

“Being physically located on the surface of the spike protein greatly increases the infectivity and pathogenicity of the virus, enabling these inserts to participate in binding to co-receptors/negatively charged attachment receptors or even, as we have discovered, to the negatively charged phospholipid heads on the cell membrane,” they wrote.

“Such a result is typically the objective of gain of function experiments to create chimeric viruses of high potency. Therefore this is a strong indicator of manipulation.”

Sørensen, Dalgleish, and Susrud point to articles by Chinese and American researchers in which, they say, “those researchers demonstrate and discuss how they have manipulated new chimeraviruses into existence, with SARS-coronavirus as a starting point”.

A comprehensive review of the relevant literature shows that a substantial amount of directly relevant gain-of function research has been undertaken, the three scientists say.

“Four studies are especially noteworthy. They are linked in two ways: scientifically, in that the third and fourth build upon the results of the first and second, and in continuity of the institution and personnel across all four.”

Sørensen and his colleagues note that the Wuhan Institute of Virology was a key collaborator in all the projects and Shi Zhengli was a common thread through all the key research projects.

The first project, in 2008, successfully demonstrated technical capabilities to interchange receptor-binding domains (RBDs) between bat SARS-like and human SARS viruses, Sørensen and his co-authors say.

Sørensen and his colleagues also say that, in 2010 scientists from the ‘Special Viruses’ section of the Wuhan Institute of Virology were engaged in gain-of-function experiments, jointly with international collaborators, to increase SARS-CoV infectiousness for humans.

“They used an HIV pseudovirus to express seven bat ACE2 receptors and compared their binding properties to human ACE2 receptors in order to pick the best for further optimising a SARS-like coronavirus’s ability to bind to human cells”, Sørensen and his co-authors wrote.

“They also found that some bat ACE2 receptors are very close to human ACE2 receptors. This study provided a model system for testing the most infectious of SARS-CoV-like viruses which already had been selected in a vast survey of Chinese bat populations between 2005–2013.”

Sørensen and his co-authors also highlight the gain-of-function research carried out in 2015.

“In 2015 scientists from the ‘Special Viruses’ section of the Wuhan Institute of Virology were engaged in ‘gain of function’ experiments jointly with a majority team from the University of North Carolina Chapel Hill,” they wrote.

“Together, they manipulated bat viruses to create a mouse adapted chimeric virus SHC014-MA15 which binds to and can proliferate on human upper airway cells.”

Detailing the way the SHC014 spike, in a wild type backbone, can “efficiently utilise multiple ACE2 receptor orthologs, replicate efficiently in primary human airway cells, and achieve in vitro titers equivalent to epidemic strains of SARS-CoV”, Sørensen and his colleagues say that what are described are “precisely SARS-CoV-2 properties”.

Sørensen and his colleagues say the 2015 authors “were well aware that the chimeric virus which they had created was very dangerous because they discussed this fact”.

They added: “It is certainly the case that this experiment created a chimeric virus with very high infectivity potential targeted to the human upper respiratory tract.”

Sørensen, Dalgleish, and Susrud say that when the in vivo experiments were carried out at Chapel Hill, and the chimeric virus replicated in mouse lung showed significant pathogenesis, this was the opposite of the result the researchers had expected.

“The creation of chimeric viruses like SHC014-MA15 was not expected to increase ‘pathogenicity’,” Sørensen and his collegues wrote.

In summary, the scientists say the work done in 2010 built upon the research carried out in 2008. “The 2010 work (Hou et al., 2010) perfected the ability to express receptors on human cells.

“On these foundations, the central gain of function work that underpins the functionalities of SARS-CoV-2 took place, carrying the WIV spike and plasmid materials to bond successfully to a UNC Chapel Hill human epithelial cell-line.”

Update 21/7/2020

A team of scientists from the Oxford Vaccine Group and Oxford University’s Jenner Institute have published the results of the first human trials of the vaccine ChAdOx1 nCoV-19, now known as AZD1222.

The researchers say the vaccine is “safe and tolerated, with reduced reactogenicity when paracetamol was used prophylactically for the first 24 hours after vaccination”.

They say reactogenicity was reduced after a second dose and the strongest immune response was seen in the ten participants who received two doses. The booster vaccine was administered 28 days after the first dose.

“Neutralising antibodies were induced in all participants after a second vaccine dose. After two doses, potent cellular and humoral immunogenicity was present in all participants studied,” they said.

AZD1222 is a chimpanzee adenovirus viral vector vaccine that expresses the SARS-CoV-2 spike protein.

It uses a weakened common cold virus (adenovirus) that infects chimpanzees and is genetically modified to code for the spike protein of the human SARS-CoV-2 virus. This means that when the adenovirus enters vaccinated people’s cells it also delivers the spike protein genetic code.

The vaccine provoked a T cell response within 14 days of vaccination and an antibody response within 28 days.

Neutralising antibody responses against SARS-CoV-2 were detected in 32 (91%) of 35 participants after a single dose when measured in a microneutralisation assay (MNA) and in 35 (100%) participants when measured in a plaque reduction neutralisation assay (PRNT).

After a booster dose, all participants had neutralising activity by day 56.

No participants were exposed to SARS-CoV-2 virus after vaccination, so the researchers point out that it is not possible for the study to determine whether AZD1222 effectively protects against SARS-CoV-2 infection.

The Oxford researchers say the trial findings are not easily generalisable, “as this was a first-in-human study of fairly young and healthy volunteers, the majority of whom were white”.

They added: “Further studies are required to assess the vaccine in various population groups including older age groups, those with comorbidities, and in ethnically and geographically diverse populations.”

Trials involving older people with comorbidities, health-care workers, and those with higher risk for SARS-CoV-2 exposure will be carried out in the UK and other countries.

“We will also evaluate the vaccine in children, once sufficient safety data have been accumulated in adult studies,” the researchers said.

“Phase 3 trials are now underway in Brazil, South Africa, and the UK and will evaluate vaccine efficacy in diverse populations.”

The Oxford researchers used the MenACWY meningitis vaccine, Nimenrix, as a “control”, not a saline placebo.

Side effects, which included headaches and fatigue, were greater in the group given the vaccine against SARS-CoV-2 than in the group given the meningitis vaccine. Other side effects included muscle aches, malaise, and chills.

The Oxford group’s vaccine trial included 1,077 healthy adults aged 18-55 years with no history of Covid-19, and took place in five UK hospitals between April 23 and 21 May 21. The data included in the paper in The Lancet covered the first 56 days of the trial.

A total of 543 people were given ChAdOx1 nCoV-19 and 543 were given the meningococcal conjugate vaccine. A total of 113 participants (56 who were given ChAdOx1 nCoV-19, and 57 in the control group) were asked to take paracetamol before and for 24 hours after their vaccination.

In the group given ChAdOx1 nCoV-19 and no paracetamol, 60% of participants suffered muscle aches as compared with 48% of those who took paracetamol.

Malaise was suffered by 61% of those not taking paracetamol and 48% of those who took paracetamol. The statistics for chills were 56% and 27% and fever 51% and 36%.

Eight patients in the group vaccinated with ChAdOx1 nCoV-19, and not given paracetamol had a temperature of at least 39°C.

The researchers say that, in the first two days after vaccination with ChAdOx1 nCoV-19, prophylactic paracetamol reduced pain, fever, chills, muscle ache, headache, and malaise.

Writing in a linked comment about the Oxford vaccine trial and a vaccine trial in China, also reported upon in The Lancet, Naor Bar-Zeev and William J Moss from the Johns Hopkins Bloomberg School of Public Health in the US, who were not involved in the two studies, said the vaccine trials were small so inferential caution was warranted, “but the explorations are laudable”

Bar-Zeev and Moss said: “Ethnic diversity in both these trials was very limited. Both trials used adenovirus vectors to deliver and study the Covid-19 vaccine, an innovative and efficient means of vaccine development in the midst of a pandemic.

“Capable of generating humoral, cellular, and innate responses, adenovirus-vectored vaccines have much potential.”

The scientists added, however: “The platform [adenovirus vectored vaccines] only achieved European Commission regulatory licensure on July 1, 2020, with the Ebola vaccine.”

They said that much remained unknown about these and other Covid-19 vaccines in development, including longevity of response and immunogenicity in older adults or other specific groups, such as those with comorbidities who are often excluded from clinical trials, or ethnic or racial groups more severely affected by Covid-19.

“Hand in hand with the trajectory of vaccine study, pharmacovigilance infrastructure is urgently needed, including surveillance for asymptomatic infection among vaccinated and unvaccinated persons if both absolute and relative risk of adverse vaccine outcomes, such as enhanced disease, are to be determined.”

Comments on Twitter that followed the release of the article in The Lancet included concern that the hype being created over the Oxford vaccine might mislead the public and that, in their excitement about a vaccine, people might become reckless about safety.

Dietitian and nutritional scientist Subhasree Ray in India asks what new the vaccines are offering. “Antibodies are producing naturally. So, these vaccines will only enhance that capacity?,” she tweeted. “T cell mediated immunity in Covid-19 patients is already being reported by multiple studies.”

Andre Watson expressed concern about those who don’t generate neutralising antibodies. “What will happen to them when they’re infected? Will they get ADE [antibody-dependent enhancement ]?” he tweeted.

Virologist Peter Kolchinsky tweeted: :Oxford data consistent w/ past results… uncomfortable & fairly weak neutralizing antibody titers compared to other vaccines (until we get outcomes from large studies, we can’t know if good enough).”

There are currently about 250 candidate vaccines against SARS-CoV-2 in development worldwide, including mRNA vaccines, replicating or non-replicating viral vectored vaccines, DNA vaccines, an autologous dendritic cell-based vaccine¹, and inactive virus vaccines. At least 17 of them are currently under evaluation in clinical trials.

Trial in China

The Lancet also carried an article about the phase 2 trial of a recombinant adenovirus type-5-vectored vaccine  that was conducted in China in April 2020 and involved more than 500 people.

The trial was carried out in a single centre in Wuhan. Partipants were healthy adults aged 18 years or older, who were HIV-negative and free of SARS-CoV-2 infection.

The researchers said that the vaccine was shown to be safe and to have induced an immune response.

“Single-dose immunisation with the vaccine induced rapid onset of immune responses within 14 days and significant humoral and cellular immune responses within 28 days in the majority of the recipients,” Feng-Cai Zhu et al. said.

The testing followed a phase 1 trial, whose results were published in May 2020. The results provided data from a wider group of participants than the phase 1 trial, who included a small sub-group of participants aged 55 years and older.

Feng-Cai Zhu et al. said it was important to stress that no participants were exposed to SARS-CoV-2 virus after vaccination, so was is not possible for the study to determine whether the vaccine effectively protects against SARS-CoV-2 infection.

The vaccine uses a weakened human common cold virus to deliver genetic material that codes for the SARS-CoV-2 spike protein to the cells. These cells then produce the spike protein, and travel to the lymph nodes where the immune system creates antibodies that will recognize that spike protein and fight off the coronavirus, the researchers say.

A total 508 participants took part in the trial of the new vaccine. Of these, 253 received a high dose, 129 received a low dose, and 126 received placebo. Sixty-one percent of participants were aged 18–44 years, 26% were aged 45–54 years, and 13% were aged 55 years or older.

The trial found that 95% of participants in the high dose group and 91% of those in the low dose group showed either T cell or antibody immune responses at day 28 post-vaccination.

The vaccine induced a neutralising antibody response in 59% and 47% of participants, and a binding antibody response in 96% and 97% of participants, in the high and low dose groups respectively by day 28.

T cell responses were also found in 90% and 88% of participants receiving the vaccine at high and low dose respectively.

The proportions of participants who had adverse reactions such as fever, fatigue, and injection-site pain were significantly higher in vaccine recipients than those in placebo recipients (72% in the high dose group, 74% in the low dose group, and 37% in the placebo group).

Feng-Cai Zhu et al. say most adverse reactions were mild or moderate and resolved within a short period of time (no more than 48 hours).

They added, however, that, within 28 days, 9% of participants in the high dose group had severe (grade 3) adverse reactions, which was significantly higher than among those receiving the low dose or placebo (one person in the low dose group, and two people in the placebo group). The most common severe reaction was fever.

The researchers note that pre-existing immunity to the human adenovirus that was used as the vector, and increasing age, could partially hamper the specific immune responses to vaccination, particularly in the case of antibody responses.

Among the 508 participants, 52% showed a high pre-existing immunity to the Ad5 vector, while 48% had low pre-existing immunity to the vector.

Those with a higher pre-existing anti-Ad5 immunity showed an inferior immune response (the binding and neutralising antibody levels were about two times greater in people with low pre-existing anti-Ad5 immunity than among those with high pre-existing immunity).

Compared with the younger population, older participants generally had significantly lower immune responses and higher tolerability to the vaccine.

Professor Wei Chen from the Beijing Institute of Biotechnology said it was possible that an additional vaccine dose might be needed in order to induce a stronger immune response in the elderly population. Further research was underway to evaluate this, he said.

Feng-Cai Zhu et al. note that the trial was conducted in Wuhan, and the baseline immunity is representative of Chinese adults at that time, but other countries may have different rates of immunity that should be considered.

“Anti-Ad5 immunity in adults varies from place to place globally, with reported proportions of  adults with neutralising antibodies titres for Ad5 of more than 1:200 of about 80% in India, 78% in Kenya, 67% in Thailand, 64% in Uganda, around 60% in South Africa, 45% in Sierra Leone, and less than 30% in the US,” the researchers said.

“We might expect the candidate Ad5-vectored vaccine to have a superior immunogenicity in the population with a lower pre-existing anti-Ad5 immunity, but an inferior immunogenicity in people with a higher pre-existing anti-Ad5 immunity than observed in this phase 2 trial.”

The researchers added that the trial only followed participants for 28 days and no data about the durability of the vaccine-induced immunity was available from the study.

“Importantly, no participants were exposed to the SARS-CoV-2 virus after vaccination, so it is not possible for this study to determine the efficacy of the candidate vaccine or any immunological risk associated with antibody induced by vaccination when having a virus exposure,” they said.

Update 17/8/2020

Scientists from the University of Waterloo, McMaster University and the University of British Columbia in Canada, the University of Nottingham in England, and the Okinawa Institute of Science and Technology Graduate University in Japan have published a peer-reviewed report in the European Respiratory Journal in which they challenge findings about how SARS-CoV-2 enters cells and affects the lungs.

Previously, it has been accepted that the virus only enters cells through the ACE2 receptor on the cell surface.

Jennifer A. Aguiar et al. found that other receptors could also perhaps facilitate SARS-CoV-2 binding and entry into the cells. These include CD147, GRP78, ADAM17, and cathepsin L.

The researchers found that the ACE2 receptor exists at very low levels in human lung tissue.

One of the lead authors, Andrew Doxey, who is a professor of biology at the University of Waterloo, told the Thailand Medical News. “Finding such low levels of ACE2 in lung tissue has important implications for how we think about this virus. ACE2 is not the full story and may be more relevant in other tissues such as the vascular system.”

According to the Thailand Medical News, the new study could have massive ramifications as the identified possible receptors have different cellular mechanisms to ACE2.

The researchers say that, collectively, their data suggest the presence of a mechanism dynamically regulating ACE2 expression in human lungs, perhaps in periods of SARS-CoV-2 infection, “and also suggest that alternate receptors for SARS-CoV-2 exist to facilitate initial host cell infection”.

They write: “Our data demonstrate rare ACE2 protein expression in human airway epithelial cells in vitro and in situ, consistent with low ACE2 promoter activity and ACE2 gene expression in bronchial epithelial cells.

“We present confirmatory evidence for the presence of TMPRSS2, CD147, and GRP78 protein in vitro in airway epithelial cells and confirm broad in situ protein expression of CD147 and GRP78 in the respiratory mucosa.

“Due to the overwhelming evidence that SARS virus interacts with ACE2, there are likely to be alternate mechanisms regulating ACE2 in the respiratory mucosa in the context of SARS-CoV-2 infection, and/or, perhaps other co-receptors, beyond what is expressed under basal conditions at the protein level.”

The researchers analysed gene expression datasets from the airway epithelial cells of 515 healthy people. They conducted gene promoter activity analysis, single cell RNA sequencing of ten healthy people, immunoblots on multiple airway epithelial cell types, and immunohistochemistry on 98 human lung samples.

1. Autologous dendritic cells are immune cells generated from patients’ own white blood cells.

Update 15/9/2020

Exiled scientist Li-Meng Yan and three of her colleagues at the Rule of Law Society and Rule of Law Foundation in New York in the US have published a report in which they lay out evidence that suggests that SARS-CoV-2 originated in a laboratory in China.

Yan, who specialised in virology and immunology at the Hong Kong School of Public Health, but fled to the US in April, says that the Chinese government and the WHO knew about person-to-person transmission of Covid-19 much earlier than was reported.

She says that her supervisors ignored research that she conducted at the beginning of the pandemic that she believes could have saved lives.

In the new paper, published on the preprint website Zenodo, Yan, Shu Kang, Jie Guan, and Shanchang Hu say their report shows that genetic evidence within the spike gene of the SARS-CoV-2 genome exists that suggests that the genome is a product of genetic manipulation.

“Furthermore, the proven concepts, well-established techniques, and knowledge and expertise are all in place for the convenient creation of this novel coronavirus in a short period of time,” Yan et al. state.

The scientists say that the genome sequence of SARS-CoV-2 has likely undergone genetic engineering, through which the virus has gained the ability to target humans with enhanced virulence and infectivity.

“The characteristics and pathogenic effects of SARS-CoV-2 are unprecedented. The virus is highly transmissible, onset-hidden, multi-organ targeting, sequelae-unclear, lethal, and associated with various symptoms and complications,” they state.

The four scientists say that the theory that SARS-Cov-2 has a natural origin, although widely accepted, lacks substantial support.

“The alternative theory that the virus may have come from a research laboratory is, however, strictly censored on peer-reviewed scientific journals. Nonetheless, SARS-CoV-2 shows biological characteristics that are inconsistent with a naturally occurring, zoonotic virus.

“In this report, we describe the genomic, structural, medical, and literature evidence, which, when considered together, strongly contradicts the natural origin theory.”

Yan et al. say there is evidence indicating that SARS-CoV- 2 is a laboratory product created by using bat coronaviruses ZC45 and/or ZXC21 as a template and/or backbone.

“Evidently, the possibility that SARS-CoV-2 could have been created through gain-of-function
manipulations at the WIV [Wuhan Institute for Virology] is significant and should be investigated thoroughly and independently,” they write.

The four researchers present three main arguments to support their contention that SARS-CoV-2 was manipulated in a laboratory.

They say that the genomic sequence of SARS-CoV-2 is “suspiciously similar” to that of a bat coronavirus discovered in military laboratories in the Third Military Medical University in Chongqing, China, and the Research Institute for Medicine of Nanjing Command.

ZC45 and ZXC21 were discovered (between July 2015 and February 2017), isolated, and characterised by researchers at the military laboratories in the Third Military Medical University and the Research Institute for Medicine of Nanjing Command, Yan et al. say. The data and associated work were published in 2018.

The researchers also say that the receptor-binding motif (RBM) within the spike protein of SARS-CoV-2 cannot be born from nature.  They say that the RBM of the spike resembles that of SARS-CoV from the 2003 epidemic “in a suspicious manner”. Genomic evidence suggests that the RBM has been genetically manipulated, they say.

“The way that SARS-CoV-2 RBM resembles SARS-CoV RBM and the overall sequence conservation pattern between SARS-CoV-2 and ZC45/ZXC21 are highly unusual,” Yan et al. write. “Collectively, this suggests that portions of the SARS-CoV-2 genome have not been derived from natural quasi-species viral particle evolution.”

The scientists further state that SARS-CoV-2 contains a unique furin cleavage site in its spike protein, “which is known to greatly enhance viral infectivity and cell tropism”.

This cleavage site is completely absent in this particular class of coronaviruses found in nature, the scientists say.

“Within the lineage B of β coronaviruses and with the exception of SARSCoV-2, no viruses contain a furin cleavage site at the S1/S2 junction [of the spike protein],” they write.

“In addition, rare codons associated with this additional sequence suggest the strong possibility that this furin cleavage site is not the product of natural evolution and could have been inserted into the SARS-CoV-2 genome artificially by techniques other than simple serial passage or multi-strain recombination events inside co-infected tissue cultures or animals.”

A codon is a trinucleotide sequence of DNA or RNA that corresponds to a specific amino acid.

Yan et al. say that, in a discovery that is consistent with the RBM engineering theory, they have identified two unique restriction sites, EcoRI and BstEII, at either end of the RBM of the SARS-CoV-2 genome.

“These two sites, which are popular choices of everyday molecular cloning, do not exist in the rest of this spike gene,” the scientist say.

“Such EcoRI and BstEII sites do not exist in the spike genes of other β coronaviruses, which strongly indicates that they were unnatural and were specifically introduced into this spike gene of SARS-CoV-2 for the convenience of manipulating the critical RBM.”

A restriction site is a sequence of approximately six–eight base pairs of DNA that binds to a given restriction enzyme. A restriction enzyme is a protein that recognises a specific, short nucleotide sequence and cuts the DNA only at a specific restriction site. The natural function of restriction enzymes is to inactivate invading viruses by cleaving the viral DNA.

Yan et al. say that once restriction sites are successfully introduced, the RBM segment can be swapped conveniently using routine restriction enzyme digestion and ligation.

The feasibility of this RBM-swap strategy has been proven, the researchers add. In 2008, they say, Shi Zhengli’s group swapped a SARS RBM into the spike proteins of several SARS-like bat coronaviruses after introducing a restriction site into a codon-optimised spike gene. They then validated the binding of the resultant chimeric spike proteins with the human ACE2 receptor (hACE2).

Yan et al. refer in their report to the work of Zhengli’s collaborator, Fang Li. “Dr Li was the first person in the world to have structurally elucidated the binding between SARS-CoV RBD and hACE238 and has been the leading expert in the structural understanding of spike-ACE2 interactions,” the researchers said.

“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.

“Rather, it is the smoking gun proving that the RBM/Spike of SARS-CoV-2 is a product of genetic manipulation.”

Although it may be convenient to copy the exact sequence of SARS RBM, it would be too clear a sign of artificial design and manipulation, Yan et al. say.

“The more deceiving approach would be to change a few non- essential residues, while preserving the ones critical for binding … Importantly, changes might have been made intentionally at non-essential sites, making it less like a ‘copy and paste’ of the SARS RBM.”

Yan et al. say that SARS-CoV-2 could have been created in a laboratory in a period of about six months.

The four scientists question the existence in nature of the much-mentioned RaTG13 virus, whose genomic sequence is reportedly 96% identical to that of SARS-CoV-2.

“While suggesting a natural origin of SARS-CoV-2, the RaTG13 virus also diverted the attention of both the scientific field and the general public away from ZC45/ZXC21,” Yan et al. say.

They say that researchers from a Chinese BSL-3 lab (the Shanghai Public Health Clinical Centre), published an article in Nature in which they reported a conflicting close phylogenetic relationship between SARSCoV-2 and ZC45/ZXC21 rather than with RaTG13, but the article “was quickly shut down for ‘rectification’”.

They add: “It is believed that the researchers of that laboratory were being punished for having disclosed the SARS-CoV-2–ZC45/ZXC21 connection.

“On the other hand, substantial evidence has accumulated, pointing to severe problems associated with the reported sequence of RaTG13 as well as questioning the actual existence of this bat virus in nature.”

Yan et al. say that a very recent publication indicates that the RBD of the RaTG13’s spike protein could not bind ACE2 of two different types of horseshoe bats.

They say that their finding further substantiates the suspicion that the reported sequence of RaTG13 could have been fabricated “as the spike protein encoded by this sequence does not seem to carry the claimed function”.

They add: “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.”

Yan et al. say genomic sequence analysis reveals that bat coronavirus ZC45 is the closest match to SARS-CoV-2.

“When SARS-CoV-2 and ZC45/ZXC21 are compared on the amino acid level, a high sequence identity is observed for most of the proteins,” they say.

“The nucleocapsid protein is 94% identical. The membrane protein is 98.6% identical. The S2 portion (2nd half) of the spike protein is 95% identical. Importantly, the Orf8 protein is 94.2% identical and the E protein is 100% identical.”

Yan et al. say sequence blast analysis indicates that, with the exception of SARS-CoV-2, no known coronaviruses share 100% amino acid sequence identity on the E protein with ZC45/ZXC21.

“Although 100% identity on the E protein has been observed between SARS-CoV and certain SARS-related bat coronaviruses, none of those pairs simultaneously share over 83% identity on the Orf8 protein.”

Yan et al. say that the 94.2% identity on the Orf8 protein, 100% identity on the E protein, and the overall genomic/amino acid-level resemblance between SARS-CoV-2 and ZC45/ZXC21 are therefore highly unusual.

“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,” they write.

They add that the high sequence identity between SARS-CoV-2 and ZC45/ZXC21 on various proteins (94-100% identity) do not support the scenario of an ancient recombination event followed by convergent evolution and clearly indicates that SARS-CoV-2 carrying such an RBM cannot come from a ZC45/ZXC21-like bat coronavirus through this convergent evolutionary route.

Yan et al. say that,  if a natural recombination event is responsible for the appearance of SARSCoV-2, then the ZC45/ZXC21-like virus and a coronavirus containing a SARS-like RBM would have to recombine in the same cell by swapping the S1/RBM, which is a rare form of recombination.

“Furthermore, since SARS has occurred only once in human history, it would be at least equally rare for nature to produce a virus that resembles SARS in such an intelligent manner – having an RBM that differs from the SARS RBM only at a few non-essential sites.”

The possibility that this unique SARS-like coronavirus would reside in the same cell with the ZC45/ZXC21-like ancestor virus and the two viruses would recombine in the “RBM-swapping” fashion is extremely low, the researchers say. Also, such a recombination event would have to happen to produce a spike as seen in SARS-CoV-2.

Yan et al. say that, judging from the evidence that they and others have gathered, there should be an independent audit of the WIV P4 laboratories and the laboratories of close collaborators of the WIV researchers.

“Such an investigation should have taken place long ago and should not be delayed any further,” Yan et al. say in their report.

“We also note that in the publication of the chimeric virus SHC015-MA15 in 2015, the attribution of funding of Zhengli Shi by the NIAID was initially left out. It was reinstated in the publication in 2016 in a corrigendum, perhaps after the meeting in January 2016 to reinstate NIH funding for gain-of-function research on viruses.”

This, the four scientists say, is unusual scientific behaviour, which needs explaining.

The researchers also say a critical look should be taken into certain recently published data, “which, albeit problematic, was used to support and claim a natural origin of SARS-CoV-2”.

Update 17/9/2020

Alina Chan, who is a postdoctoral fellow at the Broad Institute of MIT and Harvard in the US, has provided a lengthy critique on Twitter of Yan et al.’s report. Chan is herself the focus of a major article published earlier this month in Boston magazine entitled ‘Could COVID-19 Have Escaped from a Lab?’

Chan, who describes herself as a “human frontier science programme fellow with 12+ years of research training in medical genetics, biochemistry, synthetic biology, and vector engineering” describes the claims that RaTG13 is fabricated and that SARS-CoV-2 is derived from the Zhoushan viruses (published in 2018) as “gut speculations by Yan, who clearly believes that SARS2 is a product of gain of function research by the Chinese government”.

She tweeted: “My position on this: I don’t know. Someone should be looking into the origins of RaTG13 and GD pangolin CoV. What was the sampling+sequencing process? Are there still ways to verify these samples (the former has disintegrated)? Were similar viruses discovered but not reported?

“What I do know: claiming SARS2 was derived from the Zhoushan viruses that are >3000 mutations different — this has destroyed the report’s credibility, and, more importantly, diverted attention away from RaTG13, miners, and the missing WIV virus database …”

Referring to Claim 2 made by Yan et al. that the receptor-binding motif of the SARS-CoV-2  spike was genetically manipulated, Chan says that this claim relies on Claim 1 (that SARS-VoV-2 is similar to the Zhoushan viruses isolated and studied in Chinese military labs) being true. This, Chan says, is a serious weakness.

“Instead of just going with ‘RBD was copied from another virus’, Yan et al. perform enzymatic gymnastics to figure out how it could have gotten into the Zhoushan virus,” Chan tweeted.

Chan says that what Claim 2 “kind of gets correct” is that laboratories, including the WIV, have been codon optimising spikes and swapping in RBMs to study receptor binding for more than a decade.

She says that “this fixation on cloning sites is irrelevant to determining whether SARS2 was ever manipulated in a lab”.

Chan tweeted: “Scientists have been able to clone coronavirus genomes seamlessly for years. They introduce cloning sites to show you that a genome has been manipulated. Another reason to retain a cloning site could be to monitor a feature, e.g. FCS [furin cleavage site], that tends to be lost during cell passage.”

Referring to the claim by Yan et al. about the S1/S2 FCS being artificially inserted, Chan tweeted that “the fixation on whether there is a cloning site surrounding the FCS is unhelpful”.

Chan says that Yan et al. refused to incorporate any of the newly published SARS2-like viruses in their analysis.

“This is a major weakness of the report that has been pointed out by numerous experts. I would argue that at least the 4991 RdRp fragment should be considered,” Chan tweeted.

In a preliminary abstract published on September 17 in Frontiers in Public Health, Indian researchers Monali C. Rahalkar and Rahul A. Bahulikar say that RaTG13/CoV4991 was collected from the Tongguan mineshaft in Mojiang, Yunnan, in 2013.

“Surprisingly, the same mineshaft was also associated with a severe pneumonia-like illness in miners in 2012 killing three of the six miners,” Rahalkar and Bahulikar write.

“A Master’s thesis (in the Chinese language) was found on the cnki.net website which described in detail the severe illness in miners. The thesis concluded that a SARS-like CoV originating from Chinese horseshoe bats (Rhinolophus) was the predicted causative agent.”

Rahalkar and Bahulikar explain that the cases were remotely monitored by a prominent pulmonologist in China.

“The retrospective analysis of the pneumonia cases shows striking similarities with Covid-19. Bilateral pneumonia, vascular complications like pulmonary thromboembolism, and secondary infections are the main similarities,” Rahalkar and Bahulikar write.

‘The treatment regimes were similar to the currently given treatment for Covid-19. We propose that the Mojiang mineshaft and the miners’ illness cases could provide important clues to the investigation of the origin of SARS-CoV-2.”

Update 9/10/2020

Li-Meng Yan, Shu Kang, Jie Guan, and Shanchang Hu have published a second report in which they say SARS-CoV-2 is an “unrestricted bioweapon” and there has been “large-scale, organised scientific fraud”.

They allege that records indicate that “the unleashing of this weaponised pathogen should have been intentional rather than accidental”.

The current pandemic, they say, is “a result of unrestricted biowarfare”.

In the new paper, published on the preprint website Zenodo, Yan, Kang, Guan, and Hu write: “The fact that data fabrications were used to cover up the true origin of SARS-CoV-2 further implicates that the laboratory modification here is beyond simple gain-of-function research.

“The scale and the coordinated nature of this scientific fraud signifies the degree of corruption in the fields of academic research and public health.

“As a result of such corruption, damages have been made both to the reputation of the scientific community and to the well-being of the global community.”

Yan et al. say that, while SARS-CoV-2 meets the criteria of a bioweapon specified by the People’s Liberation Army (PLA) in China, “its impact is well beyond what is conceived for a typical bioweapon”.

In their new paper, Yan et al. write about the novel animal coronaviruses they say were reported by researchers in laboratories in China after the start of the outbreak of SARS-CoV-2.

“While no coronaviruses reported prior to 2020 share more than 90% sequence identity with SARS-CoV-2, these recently published, novel animal coronaviruses (the RaTG13 bat coronavirus, a series of pangolin coronaviruses, and the RmYN02 bat coronavirus) all share over 90% sequence identities with SARS-CoV-2,” they write.

“As a result, these SARS-CoV-2-like viruses have filled an evolutionary gap and served as the founding evidence for the theory that SARS-CoV-2 has a natural origin. In this report, we provide genetic and other analyses, which, when combined with recent findings, prove that these novel animal coronaviruses do not exist in nature and their genomic sequences are results of fabrication.”

Yan et al. say the RaTG13 virus has served as the founding evidence for the theory that SARS-CoV-2 must have a natural origin, but no live virus or intact genome of RaTG13 has ever been isolated or recovered.

“Therefore, the only proof for the ‘existence’ of RaTG13 in nature is its genomic sequence published on GenBank,” they write.

Yan et al. refer to an article by Yong-Zhen Zhang et al., published in Nature magazine on February 3, in which they make no mention of RaTG13 and show that, evolutionarily, SARS-CoV-2 is closest to two bat coronaviruses, ZC45 and ZXC21.

Both of these viruses “were discovered and characterised by military research laboratories under the control of the CCP government,” Yan et al. say.

“Immediately after the publication of this article, Dr Zhang’s laboratory was shut down by the CCP government with no explanations offered,” they write.

In order to have the sequence of a viral genome successfully uploaded onto GenBank, submitters have to provide both the assembled genomic sequence (text only) and raw sequencing reads.

“However, due to the huge amount of work involved in assembling raw reads into complete genomes, no sufficient curation is in place to ensure the correctness or truthfulness of the uploaded viral genomes,” Yan et al. write.

Therefore, Yan et al. say, an entry on GenBank is not a definitive proof that the viral genome is correct or real.

“Clearly, a viral genomic sequence and its GenBank entry can be fabricated if well-planned.”

The RaTG13 virus and its published sequence are suspicious and show signs of fabrication, Yan et al. say.

“The evidence presented both here and from recent literature collectively prove that RaTG13 does not exist in nature and its sequence has been fabricated,” they add.

“If the RaBtCov/4991 virus is equivalent to RaTG13, then RaBtCoV/4991 must be fraudulent as well.”

Yan et al. suggest that the fabrication of RaTG13 was planned and executed in coordination with the laboratory creation of SARS-CoV-2.

The four researchers say in their new report that the complete genomic sequence of RaTG13 was first submitted to GenBank on January 27, 2020, and the raw sequencing reads were made available on February 13.

“However, the sequencing data for gap filling, which is indispensable in assembling a complete genome, was only made available on May 19th, 2020 … The timing and the reversed order of events here are strange and suspicious.”

According to Yan et al., the raw sequencing reads of RaTG13 have multiple abnormal features.

“No independent verification of the RaTG13 sequence seems possible because, according to Dr  Zhengli Shi, the raw sample has been exhausted and no live virus was ever isolated or recovered,” they add.

“However, judging from Shi’s published protocol, exhaustion of the fecal swap sample is highly
unlikely.”

Yan et al. write: “Intriguingly, despite the pivotal role of RaTG13 in revealing the origin of SARS-CoV-2, the information provided for its discovery was surprisingly scarce with key points missing (location and date of sample collection, previous knowledge and publication of this virus, etc).”

They added: “Only in the source section of the NCBI entry for RaTG13 (GenBank accession code: MN996532.1), one could find that the original sample was a ‘fecal swab’ collected on ‘July 24th, 2013’.”

According to Yan et al., there are discrepancies between what Shi Zhengli suggested in the article published in Nature in January 2020 (that the sequencing of the full genome of RaBtCoV/4991 was not done until 2020) and what Zhengli said later in emailed replies to Science magazine.

Yan et al. say that, in June 2020, file names of the raw sequencing reads for RaTG13 uploaded were found, which indicated that the sequencing experiments were done in 2017 and 2018.

“Likely responding to this revelation, in her email interview with Science, Dr Shi contradicted her own description in the Nature publication and admitted that the sequencing of the full genome of RaTG13 was done in 2018.”

The researchers say no follow-up work on RaTG13 has been reported by Zhengli and her colleagues.

“Upon obtaining the genomic sequence of a SARS-like bat coronavirus, the Shi group routinely investigate whether or not the virus is capable of infecting human cells. This pattern of research activities has been shown repeatedly.

“However, such a pattern is not seen here despite that RaTG13 has an interesting RBM and is allegedly the closest match evolutionarily to SARS-CoV-2.”

There were, Yan et al. say, deviations from normal research activities and logical thinking that are difficult to reconcile or explain.

Yan et al. say investigations should be carried out “on the suspected government and individuals and the responsible ones be held accountable for this brutal attack on the global community”.

Yan has told television host in the US Tucker Carlson that her 63-year-old mother has been detained in China.

Alina Chan says that the second report by Yan et al. is “littered with errors”. She tweeted: “… I do wonder why there hasn’t been international impetus to investigate the source of these SARS2-like viruses. Why not go to the Yunnan mine to look for more RaTG13s? Why not investigate the miners – what actually happened in 2012?”

Chan added: “I’m glad Yan is shining the spotlight on these research integrity issues. But I worry that framing the verifiable misdirection surrounding RaTG13/pangolins within a specious article may actually hurt the legitimacy of research integrity inquiries regarding SARS2-like viruses.”

She tweeted: “Yan et al. claim because a cover-up was ‘orchestrated by the CCP’ … ‘the unleashing of the virus must be a planned execution rather than an accident’. I disagree. Just because something is accidental, doesn’t mean it has no consequences & will be admitted openly.”

A new article about the SARS-CoV-2 vaccine race can be found here.

DONATE TO CHANGING TIMES VIA SIMPLE PAYMENTS

1= 5 euro, x 2 = 10 euro, X 3 =15 euro, etc.

€5.00