COVID-19: zoonosis or leak ?

16 december 2021,

Bonjour,

J'espère que vous allez bien.

A mon avis, il est grand temps de faire preuve d'imagination pour sortir de la crise pandémique.

C'est dans ce but que j'ai soumis à "La Libre" une "Opinion" pour leur rubrique Débats.

Comme le journal refuse de le publier, je vous prie de trouver mon texte ci-dessous.

N'hésitez pas à me contacter pour en parler:  angoffinet@gmail.com

Hello,

I hope you are doing fine.

In my opinion, it is high time to use imagination to get out of the pandemic crisis.

It is with this in mind that I submitted an "Opinion" to "La Libre" for their Debates section.

As the newspaper refuses to publish it, please find my text below.

Do not hesitate to contact me to talk about it: angoffinet@gmail.com

French version

Décréter la fin administrative de la pandémie au printemps prochain. Pourquoi pas ?

Le COVID-19 est la pire pandémie depuis la grippe de 1918 et le SIDA. Prises au dépourvu, les autorités se sont vues contraintes de faire face aux événements de manière réactive, coup par coup: confinement total, partiel, par secteurs, masques, distance … Maintenant, la disponibilité de vaccins de qualité et l’arrivée de nouveaux traitements nous permettent d’envisager une stratégie plus active.

La pandémie entre bientôt dans sa troisième année. On sait que le virus SARS-CoV2 mute facilement. Comme celui de la grippe, il est très probablement installé pour longtemps. Dès lors, il est temps de poser la question qui dérange. Poursuit-on la ligne actuelle en minimisant les risques et les prises de responsabilités,  avec tous les inconvénients connus. Ou prenons-nous une direction radicalement différente en assumant les risques.

Sans critiquer personne (« aurions-nous mieux fait à leur place ? »), reconnaissons franchement que l’approche actuelle est un paquet de mesures parfois infantilisantes et à l’efficacité souvent douteuse, comprenant des dictats qui parfois frisent l’arbitraire et le liberticide: quarantaine, tests, obligation de porter le masque, de vacciner certains, interdiction de se rassembler…  n’allongeons pas la litanie.  La population est mal à l’aise : elle comprend qu’il faut lutter contre la pandémie, mais elle déteste aussi qu’on lui dicte son mode de vie, et elle a bien raison.

Au vu des connaissances épidémiologiques, scientifiques, médicales et sociétales acquises depuis le printemps 2020, ne serait-il pas plus simple et efficace de prévoir une levée de toutes les mesures sanitaires au printemps 2022 ?

En 2020 et 2021, la pandémie s’est fortement atténuée au printemps. Parions que ce sera le cas en 2022. Dès maintenant, une décision proactive peut être prise : profiter du creux de la vague pour abroger toutes les mesures de prévention au printemps. Il n’est pas interdit de sourire et on pourrait choisir le 22 mai 2022, où l’on fête Ste Rita, patronne des causes perdues. Rien ne nous empêche de décréter la fin administrative du cauchemar.

Retour à la situation pré-pandemique, ni plus, ni moins. Plus aucune obligation de masque ou de distance nulle part, aucune restriction dans l’enseignement, l’horéca, les activités culturelles, sportives ou les fiestas nocturnes... Plus aucun contrôle, aucune obligation de tests, plus de quarantaine, aucun CST, rien, nada. Ceux qui le souhaitent restent libres de porter un masque ou d’appliquer toute mesure qu’ils jugent utile, mais personne n’y est contraint.

Quelles seraient les conséquences prévisibles d’une décision aussi drastique?

En positif, coup de fouet au moral de tous, retour à un fonctionnement normal de la société, gain de temps et d’énergie considérable, et bénéfices économiques certains.

L’impact négatif évident, que ne manqueront pas de souligner les Cassandre, est une recrudescence inévitable mais temporaire des cas de COVID, avec répercussions sur le système de santé.  Je pense que cet obstacle n’est pas insurmontable, mais au contraire gérable si la stratégie est bien cadrée au cours des quelques mois à venir.

Premièrement,  il faut poursuivre la campagne de vaccination, convaincre les indécis et insister pour que l’industrie procure des vaccins actifs contre les variants du virus. D’ici mai 2022, toutes les personnes non encore vaccinées pourront l’être, sans obligation. Les non vaccinés seront placés devant la responsabilité de se protéger eux-mêmes. Un autre facteur bénéfique à prendre en compte est la mise à disposition de nouveaux traitements. Des anticorps monoclonaux sont déjà disponibles, mais réservés à des cas exceptionnels. Ils ont deux désavantages : efficacité diminuée lors de  mutations du virus, et coût très élevé. Les inhibiteurs de protéase virale sont très prometteurs et déjà disponibles dans certains pays.

Deuxièmement, il faut préparer le système hospitalier à une recrudescence des cas suite à la levée des mesures sanitaires. Il est impératif de soigner tout le monde au mieux. Notre système de santé est magnifique mais le personnel soignant est las et découragé. Une bonne manière de le motiver est de proposer des incitants financiers. Pas une aumône comme fin 2021. Les coûts de cette opération seraient atténués, voire largement compensés par la reprise de l’activité économique. La fin des mesures en mai doit être planifiée avec une large concertation entre les responsables des soins de santé et les autorités politiques et administratives. Quand on veut, on peut.

Enfin, il existe toujours une possibilité que le virus ne recule pas, voire devienne encore plus grave d’ici le printemps 2022. Ce risque est faible. Dans cette circonstance imprévisible, une levée des mesures en mai devrait être annulée. Ce ne serait pas le premier changement de cap.

Ce que nous vivons tous actuellement me semble ubuesque. Nous ne demandons qu’une chose : en sortir. C’est dans ce seul but que je me permets d’ouvrir le débat.

Namur, 16 décembre 2021

English version:

Declare an administrative end to the pandemic next spring. Why not?

COVID-19 is the worst pandemic since the 1918 flu and AIDS. Caught off guard, the authorities were forced to deal with the events in a reactive way, piecemeal: total or partial containment, by sectors, masks, distance ... Now, the availability of quality vaccines and the arrival of new treatments allow us to consider a more active strategy. 

The pandemic will soon enter its third year. We know that the SARS-CoV2 virus mutates easily. Like influenza, it is very likely to be around for a long time. It is therefore time to ask the uncomfortable question. Do we continue with the current line, minimizing the risks and responsibilities, with all the known drawbacks. Or do we take a radically different direction by assuming the risks. 

Without criticizing anyone ("would we have done better in their place?"), let's frankly recognize that the current approach is a package of measures that are sometimes infantilizing and often of dubious effectiveness, including dictates that sometimes border on the arbitrary and liberticide: quarantine, tests, obligation to wear masks, to vaccinate some people, prohibition of gathering... let's not lengthen the litany.  The population is uncomfortable: it understands that we must fight against the pandemic, but it also hates being dictated to, and rightly so. 

Given the epidemiological, scientific, medical and societal knowledge acquired since spring 2020, would it not be simpler and more effective to plan for a lifting of all health measures in spring 2022?

In 2020 and 2021, the pandemic has strongly subsided in spring. Let's bet that this will be the case in 2022. A proactive decision can be made now: take advantage of the trough to repeal all prevention measures in the spring. It is not forbidden to smile and we could choose May 22, 2022, when we celebrate St. Rita, patron saint of lost causes. Nothing prevents us from declaring the administrative end of the nightmare. 

Back to the pre-pandemic situation, nothing more, nothing less. No more obligation of mask or distance anywhere, no restriction in the teaching, the horéca, the cultural activities, the sports or the night fiestas... No more control, no obligation of tests, no quarantine, no CST, nothing, nada. Those who wish to do so are free to wear a mask or to apply any measure they deem useful, but no one is forced to do so.

What would be the foreseeable consequences of such a drastic decision? 

On the positive side, it would boost everyone's morale, return society to normal functioning, save a lot of time and energy, and provide definite economic benefits.

The obvious negative impact, which the Cassandras will not fail to point out, is an inevitable but temporary increase in the number of cases of COVID, with repercussions on the health care system.  I believe that this obstacle is not insurmountable, but rather manageable if the strategy is well aligned over the next few months. 

First, we need to continue the vaccination campaign, convince the undecideds, and insist that industry provide active vaccines against the virus variants. By May 2022, all those not yet vaccinated will be able to be vaccinated, without obligation. The unvaccinated will be put in charge of protecting themselves. Another beneficial factor to consider is the availability of new treatments. Monoclonal antibodies are already available, but only for exceptional cases. They have two disadvantages: reduced efficacy when the virus mutates, and very high cost. Viral protease inhibitors are very promising and already available in some countries. 

Secondly, the hospital system must be prepared for a resurgence of cases following the lifting of sanitary measures. It is imperative that we treat everyone as well as possible. Our health care system is wonderful, but the health care personnel are tired and discouraged. A good way to motivate them is to offer financial incentives. Not a handout like in late 2021. The costs of this would be mitigated, if not more than offset by the recovery in economic activity. The end of the measures in May must be planned with broad consultation between health care leaders and political and administrative authorities. Where there is a will, there is a way.

Finally, there is always the possibility that the virus will not recede or will become even more severe by the spring of 2022. This risk is small. In this unpredictable circumstance, a lifting of the measures in May would have to be reversed. This would not be the first change of course.

What we are all living through at the moment seems to me surrealistic. We only ask one thing: to get out of it. It is for this sole purpose that I am taking the liberty of opening this debate.

Namur, december 16th, 2021

Thanks to colleagues and friends who contacted me about this text and offered useful comments. This new version takes this into account.

COVID-19: zoonosis or leak?

André Goffinet (edited 15 April 2020)
New information added 23 April 2020

Note added on May 13
An internal email sent by  Yan-Yi WANG,director of the Wuhan Institute of Virology:

Translation as:

"Subject: [Important remind] A notification about the prohibition of disclosing the information related to Wuhan pneumonia of unknown cause

Hi everyone, 

Recently, Our society has been paying close attention to the progress of "Wuhan unknown pneumonia", and the spread of some inappropriate and untrue information in the early stage has caused a certain degree of public panic.
Our institute is carrying out intensive related work, under the guidance of the department of health. I want to inform all our colleagues about a notification received yesterday from the National Health Commission (NHC) via telephone:
"The NHC clearly requires that all tests, experimental data, results, or conclusions related to the epidemic should not be published on any "we media" or social software, nor disclosed to media (including official media) and partner agencies (including technical service companies).
Please follow this policy."

Note added on April 29
A new outbreak in under way in Harbin, North East China, 200 km from Russian border. That outbreak started about mid march and Harbin has been placed in lockdown about one week ago.
This is very troubling because Harbin hosts the second chinese P4 high security biolab, which is operated by veterinarians and the chinese academy of agronomical sciences. Investigators in that P4 have infected domestic animals such as ferrets, cats, dogs, pigs and chicken with SARS-CoV-2. This was published in Science recently. Could there have been a leak during this flurry of activity in that P4? Wuhan and Harbin, two P4, two outbreaks, two lockdowns, lots of troubling coincidences that must be investigated.

The whole world is now mobilized against COVID-19, a nightmarish situation that nobody would have deemed possible a few months ago. We will suffer a lot, fight and ultimately prevail, at a high cost. Then, time will come to think and take action to minimize future risks.

Understanding the origin of the virus is a key element in the story. I do not know what your opinion is, but personally I am not satisfied with the official story. Information on this is scant and comes only from Chinese official channels. While avoiding conspiracy theories, which I abhor, and China bashing, we can look at facts and infer from facts only.

The official version: Covid-19 is a zoonosis

Let us begin by summarizing the official version. Due to poor hygiene in a wild animals market in Wuhan, coronaviruses originating from bats were passed to intermediate mammals, possibly illegally traded pangolins, and then to humans who handled them. This virus happened to be very contagious and quite dangerous, and spread like fire from Wuhan to the rest of the world. This is by definition a zoonosis.

The COVID-19 epidemic (coronovirus induced disease 2019) began in Wuhan, Hubei province, PR China. This is an agglomeration of about 11 million people located in central China, on the Yangtse River about 800 km west of Shanghai. Chinese authorities initially said that the disease started at a market where wild animals and fish are sold for human consumption. That Huanan Seafood market is located at the city center, and is now closed. See the wiki entry (with no guarantee about sources): https://en.wikipedia.org/wiki/Huanan_Seafood_Wholesale_Market.  More recently, data surfaced that raised serious doubt about the market origin, because about one third initial cases had no contact with that market.

COVID-19 is due to infection by coronavirus SARS-CoV-2, which is related to SARS-CoV-1, the agent of the 2002-2003 SARS ep. idemic (SARS = « severe acute respiratory syndrome »), and to MERS CoV (the agent of “Middle East respiratory syndrome”). For a good clinical account of COVID-19, see the Lancet (Huang et al., 2020).

SARS-CoV-1 and SARS-CoV-2 infect target cells via their S (« spike ») protein that protrudes from the viral surface (envelope). S proteins bind to cells via a receptor binding domain (RBD). The viral RBD binds to a peptidase named ACE2 (angiotensin converting enzyme 2) exposed on the cell surface.

Genome sequence analyses showed that SARS-CoV-2 is not only related to SARS-CoV-1 and MERS, but even closer (96% sequence similarity), to coronaviruses such as Bat-CoV-RaTG13, isolated with several others by teams at the Wuhan Institute of Virology (see below), from bats living in caves in the Chinese Yunnan province, 1700 km from Wuhan (Ge et al., 2017; Zhou et al., 2020). However, the SARS-CoV-2 and Bat-CoV-RaTG13 viruses differ significantly in their RBD sequences. This rules out a direct transmission from bats to humans, and leads to the question of an “intermediate host”.

Bats carry various corona- and other viruses that do not infect humans directly. Some of them occasionally infect intermediate mammalian hosts, who rarely pass them to humans. For example, the intermediate host of SARS-CoV-1 was a civet, and for MERS it is the dromedary camel. MERS does not pass from human to human, whereas SARS-CoV-1 could spread from civet to human and then to other humans during the SARS epidemic in 2002-2003.

The key issue of an intermediate mammalian host of SARS-Cov-2 has not been solved. Initial information pointed the Huanan market but Chinese authorities have become more nuanced about the role of that market in initiating the infection.  This is discussed further, among others, in https://www.sciencemag.org/news/2020/01/wuhan-seafood-market-may-not-be-source-novel-virus-spreading-globally. See also a balanced article in The Guardian: https://www.theguardian.com/world/2020/apr/08/how-did-coronavirus-start-and-where-did-it-come-from-was-it-really-wuhans-animal-market?CMP=share_btn_link

There were also persistent rumors that pangolins sold in Wuhan served as intermediate hosts, but that question is far from settled. Coronaviruses have been isolated from pangolins (Manis javanica, aka Sunda, Malayan or Javan pangolin), and a team in Guangzhou published that a coronavirus isolated from smuggled Malaysian pangolins frozen by customs in 2017-2018 (virus GD/P1L) had high sequence similarity to SARS-CoV-2. That similarity concerned the RBD, a 74 aminoacid stretch important for ACE2 binding. The pangolin virus RBD differs from that of the SARS-CoV-2 by only one residue, providing an argument for pangolins being intermediate hosts (https://www.nature.com/articles/s41586-020-2169-0). However, the whole genome is only 90% similar to SARS-CoV-2, and therefore less related to the human virus than the bat Bat-CoV-RaTG13 virus, which is 96% similar to SARS-CoV-2. This does not fit with pangolins as intermediate hosts, because their virus should then have a similarity to SARS-CoV-2 between 96 (bat) and 100%. For example the SARS-CoV isolated from civets was 99.8% similar to the human SARS-CoV-1. Furthermore, pangolins (a CITES protected species !) are regularly smuggled into China from Africa or South Asian countries (https://en.wikipedia.org/wiki/Pangolin_trade), and the pangolin from which the SARS-like virus was isolated was found in Guangdong, far from Hubei province. Smuggled pangolins have been on the market in several cities, more in South than Central China, and it was said that no pangolins were for sale at the Huanan market (unverifiable). Why then did the epidemic start in Wuhan rather than South China like SARS?

That official version is not foolproof and other data are really troubling

Two laboratories in Wuhan are actively involved in studies of bat coronaviruses

The first is the Wuhan Institute of Virology (WIV), a first class institute of the Chinese Academy of Science (CAS) located in a residential area close to central Wuhan, about 12-15 km from the Huanan market (http://english.whiov.cas.cn/). The second is at the Wuhan Center for Disease Control and Prevention (WHCDC) (https://www.whcdc.org/) and is even closer to central Wuhan, about 300 m away from the market.

Since the SARS epidemics, the WIV studies coronaviruses isolated from wild species, and has become a world leader in the field. A team led by Dr Zheng-Li Shi is well known to isolate and identify coronaviruses from bats captured all over China, especially in Yunnan province, and even in other countries. They try to understand how bat viruses can infect human cells such as airway epithelial cells. Dr Shi is a competent and dynamic scientist, as attested by her publications (https://www.ncbi.nlm.nih.gov/pubmed/?term=zhengli+shi ).  She is known and mediatized in China as « Chinese bat woman » (https://www.scientificamerican.com/article/how-chinas-bat-woman-hunted-down-viruses-from-sars-to-the-new-coronavirus1/). It should ne acknowledged that she, like others, drew attention to potential danger of bat coronaviruses being “poised” to generate epidemics in humans. Biosecurity level of their experiments are not always mentioned in publications. I would assume that experiments were initially performed at the biosecurity level (BSL) 2 (P2), and perhaps later at a P3 level. Since 2017, the WIV has a brand new BSL4 (P4) laboratory ( https://en.wikipedia.org/wiki/Wuhan_Institute_of_Virology). I could not assess whether Dr Shi’s team did use that P4, but it seems to be the case according to a publication by The Washington Post (April 14) mentioning that “State Department cables warned of safety issues at Wuhan lab studying bat coronaviruses”, with presumed lax security in the Wuhan P4. In principle, P3 conditions are suitable for coronavirus work. Leaks are rare at P3, and exceptional in P4 conditions. Dr Shi denies that SARS-CoV-2 could have leaked from her laboratory. 

Her team recently published a Nature paper on the bat origin of SARS-CoV-2, with Peng Zhou as first author (Zhou et al., 2020). Peng Zhou has received prestigious grants for his work at the WIV (such as CAS grant XDB29010104; China Natural Science Foundation for excellent scholars 81822028; Mega-Project for Infectious Disease from Minister of Science and Technology of the People’s Republic of China 2018ZX10305409-004-001); he recruited postdocs for bat coronavirus work at WIV as late november 2019 http://www.whiov.cas.cn/105341/201911/t20191118_5438006.html. However, today, his name is not listed among the personnel of WIV. Note also that recent work showing transmission of SARS-CoV-2 to ferrets and cats (Shi et al., 2020) was carried in BSL4 conditions in Harbin, where a second P4 lab is located.

Work on bat coronaviruses at the WHCDC has been carried out by a researcher named Tian Jun-Hua. He has collected hundreds of viruses, especially coronaviruses from bats all over Hubei province and studies them in BSL2 (P2) conditions at the WHCDC. This is evident from a China Science Communication promotion video from the WHCDC that was highly mediatized in China and is available on YouTube:

https://www.youtube.com/watch?v=gYOetKA1o4U&fbclid=IwAR333npF8RTVol4QkNhsIkEAB5o5K59AnALD1JKGZSz1mpAJrGTdSHuKIM8

The internet link to the WHCDC (https://www.whcdc.org/) worked very poorly for days and has been reactivated around April 10^th.  But I could not find any information on bat coronavirus work, not about Tian Jun-Hua, who is not mentioned among personnel. This is strange because WHCDC investigators Tian Jun-Hua and Yu Bin are both coauthors of a recent Nature paper on the bat origin of SARS-CoV-2 (Wu et al., 2020), and, contrary to Tian Jun-Hua,  Yu Bin is mentioned among personnel on the WHCDC website. It is also worth noting that, from comparing publication records, Dr Tian is scientifically much less productive than Dr Shi.

In sum, at least two laboratories, WIV and WHCDC, are situated close to central Wuhan and are actively involved in studies of bat coronaviruses. Both are potential sources for accidental leaks.

The issue of Gain of Function (GOF) experiments

American laboratories also investigated the SARS-CoV-1 following the 2002-2003 epidemics, some working at a BSL3 (P3) level. They showed, for example, that ACE2 is the SARS virus receptor (Li et al., 2003).  Other teams, particularly the one led by Dr Ralph Baric (UNC Chapell-Hill) studied factors that allow coronaviruses to infect humans (Menachery et al., 2015; Yang et al., 2015; Menachery et al., 2016). Some of those studies were carried out in collaboration with Dr Shi from WIV, who is co-author of some papers. A PNAS paper (Menachery et al., 2016) mentions in its title a virus WIV-1 COV (WIV stands for « Wuhan Institute of Virology »).

In those experiments, coronaviruses are cultured in various cell lines such as Vero E6 (derived from green monkey kidney), Huh7 (human liver), Calu-3 (human lung cancer) and Caco-2 (human colon cancer), and on Human Airway Epithelial (HAE) cultures, which effectively mimic the human bronchial environment (B et al., 2009; Menachery et al., 2020).

The strategy to adapt viruses to poise them to infect new organisms is called « Gain of Function » (GOF) in the virologist community.  Publication of GOF experiments to infect human cells raised eyebrows concerning their benefit/risk ratio: Is the scientific benefit sufficient given the risk of generating new human pathogens?

Following this, in October 2014, the NIH issued a document entitled “U.S. Government Gain-of-Function Deliberative Process and Research Funding Pause on Selected Gain-of-Function Research Involving Influenza, MERS, and SARS Viruses” blocking funding of such GOF experiments.  (http://www.phe.gov/s3/dualuse/Documents/gain-of-function.pdf )

Studies under way, such as those in the Baric lab, were allowed to proceed and some were published.

Whether groups in Wuhan followed this American moratorium is unclear as they were obviously not bound to do so. No publications from WHCDC are easily available. Publications from WIV groups mention culture in Vero and some human cells, but they also invested a lot of effort in growing bat coronaviruses in bat cell lines, and in reconstituting viral genomes bioinformatically, without culturing in vitro. On the other hand, a description of research by Shi’s team available on the WIV website gives stong hints (research theme 4.) http://159.226.126.127:8082/web/55577/home

Arguments for a leak scenario

In February 2020, US conservative journals and websites pointed to a leak of SARS-CoV-2 from a Wuhan laboratory. Papers were aggressive and strongly anti-communist, and not too factual.  They were met with strong skepticism by the scientific community who branded them as conspirational and Chinese-bashing. I, for one, dismissed all this without further notice until I started later, in March, to look at facts.

There was also a well documented note published by Botao Xiao and Xiao Lei who pointed to the possibility of an accidental leak from both laboratories in Wuhan. That short paper was briefly available on ResearchGate and was then deleted.

American colleagues published that SARS-CoV-2 is most likely issued from a spontaneous evolution of a bat virus and that COVID-19 is a zoonosis, supporting the official Chinese version. See for example Trevor Bedford on Twitter (https://twitter.com/trvrb/status/1230634351794089984 ), and a commentary in Nat medicine (Andersen et al., 2020) by experts in viral epidemics.

The main arguments for a the zoonotic origin of the epidemics, namely spontaneous mutation of a bat virus with infection of an intermediate host and then humans, are : i) The SARS-CoV-2 genome is 96% similar to a bat coronavirus referred to as BetaCoV/bat/Yunnan/RaTG13 ; ii) The SARS-CoV-2 RBD aminoacid sequence has 73/74 residues in common with a coronavirus (GD/P1L) recovered from frozen pangolins kept by customs; iii) The SARS-CoV-2 sequence shows no sign or signature of genetic engineering (e.g. novel restriction sites); iv) The SARS-CoV-2 RBD sequence binding ACE2 could not have been predicted, and therefore cannot result from targeted engineering.

Those facts are solid, but I find interpretations biased. Except for the high similarity between the RBD in SARS-CoV-2 and pangolins (ii), an alternative explanation to i), iii) and iv) is straightforward. Since 2002-2003, the Wuhan labs carried out experiments involving culture of multiple bat coronaviruses in different cell lines, including human ones. By selecting viruses able to grow on human cells, those experiments mimic evolution and favor the selection of spontaneous mutations that increase the virulence to humans. This does not require any genetic engineering per se. If handled inappropriately, an adapted virus could humans without passing through an intermediate host.

About the similarity of RBD of the pangolin coronavirus and SARS-CoV-2, this is puzzling, but the data are not strong. They were initially published in bioRxiv (https://www.biorxiv.org/content/10.1101/2020.02.13.945485v1.full.pdf) , and the pdf is accessible as an accelerated article preview in Nature https://www.nature.com/articles/s41586-020-2169-0. The pangolin GD/P1L sequence in GenBank is poorly annotated https://www.ncbi.nlm.nih.gov/nuccore/1811123271 , making critical analysis very difficult.

In sum, it is perfectly plausible that SARS-CoV-2 was generated in a Wuhan lab (WIV or WHCDC) by selection and adaptation to human cells, and escaped unintentionally, due to some security breach. Being adapted to human, it did not need an intermediate host (which has thus far not been identified) and could spread to Wuhan and then to the rest of the world. This is the leak scenario.

Summary arguments for a leak from a Wuhan lab 

1. Culture of bat coronaviruses have been done in WIV and WHCDC, resulting in adaptation of coronaviruses from bat to human cells, including human airway epithelial cells.

2. A virus adapted to grow on human cells can infect humans without passing through an intermediate mammal. Indeed, thus far, no intermediate host has been clearly identified.

3. Safety records of Chinese virology labs are not impeccable, as pointed out in The Washington Post on April 14. There were at least two (some reports mention four) leaks of SARS-CoV1 since 2002, fortunately rapidly controlled.

4. Building of the WIV BSL4 lab has been difficult. It lasted more than 10 years (2005-2017) and was/is plagued with internal politics, ego, personal and career problems.

In light of the two possibilities, zoonosis or leak, what should be done ?

SARS-CoV-2 is a moderately pathogenic virus, yet COVID-19 is a world catastrophe. Where would we be if the virus were more aggressive, like MERS for example (30% lethality of infected people)?

Chinese and others, we are all deeply concerned about this happening again in the future. The situation requires that futile face saving considerations be set aside. We MUST know the truth and be proactive.

1. Laboratories at the “Wuhan Center for Disease Control & Prevention” and the « CAS Wuhan Virology Institute », especially its P4 facility, should be investigated by external reviewers, objectively and without hatred or ideological bias.

2. Those laboratories should be closed pending investigation. Laboratories doing GOF or related experiments and the P4 facility should be moved far from city centers. It makes no sense to perform so high risk research close to a densely populated location.

3. Research dealing with GOF and related experiments should be submitted to strict conditions. An International Body should be competent because those issues reach beyond national borders, as COVID-19 amply demonstrates.

André M. GOFFINET, MD, PhD

Prof. Em. Institute of Neuroscience

University of Louvain, Belgium

Email: angoffinet@gmail.com

Opinions expressed here are solely my own and do not reflect any institutional position by the University of Louvain.

Addendum for those interested in more detailed discussion and molecular biology…

The following discussion is largely summarized on a paper in Nat Rev Immunology (de Wit et al., 2016) and from a website of Vincent Racaniello, a renowned virologist (http://www.virology.ws/2020/02/13/furin-cleavage-site-in-the-sars-cov-2-coronavirus-glycoprotein/.

Primer on coronaviruses

SARS-CoV and MERS-CoV belong to the Coronaviridae family and have large, positive-sense RNA genomes of 27.9 kb and 30.1 kb, respectively. Two-thirds of the viral RNA is translated into two large polyproteins (pp1a and pp1ab), and the remainder of the viral genome is transcribed into nested set (9 to 12) subgenomic mRNAs that encode the four structural proteins spike (S), envelope (E), membrane (M) and nucleocapsid (N), as well as several accessory proteins. The two polyproteins, pp1a and pp1ab, are proteolytically cleaved into 16 non-structural proteins (nsp1–nsp16) including papain-like protease (PLpro), 3C-like protease (3CLpro), RNA-dependent RNA polymerase (RdRp), helicase (Hel) and exonuclease (ExoN). These nsp’s make up the replicase-transcriptase complex. The polyproteins are cleaved by two proteases, papain-like protease (PLpro, nsp3) and a main protease, 3C-like protease (3CLpro; nsp5). The nsp’s rearrange membranes derived from the rough endoplasmic reticulum (RER) into double-membrane vesicles, in which viral replication and transcription occur.

Following entry of the virus into the host cell, the viral RNA is uncoated in the cytoplasm. ORF1a and ORF1ab are translated to produce pp1a and pp1ab, which are cleaved by the proteases that are encoded by ORF1a to yield 16 non structural proteins (nsps) that form the RNA replicase–transcriptase complex. This complex localizes to intracellular membranes derived from the rough endoplasmic reticulum (ER), and it drives the production of negative-sense RNAs ((−)RNAs) through both replication and transcription. During replication, full-length (−)RNA copies of the genome are produced and used as templates for full-length (+)RNA genomes. During transcription, a subset of 7–9 subgenomic RNAs, including those encoding all structural proteins, are produced. These subgenomic (−)RNAs are then transcribed into subgenomic (+)mRNAs. The resulting structural proteins are assembled into the nucleocapsid and viral envelope at the ER–Golgi intermediate compartment, followed by release of the virion from the infected cell.

Mechanism of entry into target cells

Virus entry into cells is mediated by the trimeric transmembrane spike (S) glycoprotein. The S protein contains two functional domains: a receptor binding domain (RBD), and a domain that mediates fusion of the viral and cell membranes. Cell entry requires binding to cell surface receptors and S protein cleavage to liberate and activate the fusiogenic sequence.

Receptor binding. The cellular receptor for SARS-CoV-1 and -2 is the membrane peptidase ACE2. Six aminoacids in the receptor-binding domain (RBD) of the S protein are critical for binding to ACE2. In SARS-CoV-1, they are Y442, L472, N479, D480, T487 and Y4911, corresponding to SARS-CoV-2 residues L455, F486, Q493, S494, N501 and Y505. Intriguingly, these residues are conserved in the pangolin virus sequence. The SARS-CoV-2 RBD binds with high affinity to ACE2 from humans, ferrets, cats and other species with high receptor similarity. The structural interactions between the S protein and ACE2 have been defined (Wrapp et al., 2020; Yuan et al., 2020). The physiological role of ACE2 is to cleave angiotensin 2. Note that ACE2 is related but different than ACE, a target of prominent anti-hypertensive drugs.

The SARS-CoV-2 RBD sequence differs from the predicted optimum based on SARS-CoV-1. The high-affinity binding of the SARS-CoV-2 S protein to human ACE2 therefore provides a binding strategy devised by spontaneous evolution, different than the one modeled based on SARS-CoV-1. Some authors consider this as strong evidence that SARS-CoV-2 is not the product of purposeful manipulation (Andersen et al., 2020); more of that below.

Things go fast nowadays, and the structure of the SARS-CoV-2 S protein bound to ACE2 has already been published (Wrapp et al., 2020; Yuan et al., 2020).

S protein cleavage. Each monomer of trimeric S glycoprotein is about 180kDa) and contains an S1 N-terminal part involved in attachment to the cell via the RBD, and an S2 part that mediates fusion of viral and cellular membranes. Glycoprotein S must be cleaved into S1 (that contains the RBD) and S2 (involved in membrane fusion) by host cell proteases prior to cell entry. Depending on virus strains and cell types, S proteins may be cleaved by one or several host proteases, including trypsin, cathepsins, transmembrane protease serine protease-2 (TMPRSS-2), TMPRSS-4, or human airway trypsin-like protease (HAT). The host proteases that cleave S proteins of SARS-CoV-2 have not been clearly defined.

Cleavage of the S glycoprotein can determine whether the virus can cross species. For example, the S glycoprotein from a bat CoV can bind to human cells but cannot be cleaved and cannot mediate virus entry. However, if trypsin is included during infection, the S glycoprotein is cleaved and virus enters. Thus cleavage of the S glycoprotein is a barrier to zoonotic coronavirus transmission.

Another point of interest is the presence of furin cleavage sites in some S proteins. Those sites could be processed during intracellular traffic of virions, prior to their exit from cells. Furin cleavage primes virions to infect other cells directly, by passing the need for S1/S2 cleavage by host proteases after receptor binding.

Contrary to SARS-CoV-1, the S glycoprotein of SARS-CoV-2 contains a furin cleavage sequence (PRRARS|V). As furins are abundant in the respiratory tract, the SARS-CoV-2 S-glycoprotein could be cleaved upon exit from epithelial cells and directly infect other cells. Furins are known to control infection by avian influenza A viruses, in which cleavage of the HA glycoprotein is needed for cell entry. In low-pathogenic viruses, there is a single basic amino acid at the cleavage site in the HA protein, which is cleaved by proteases restricted to the respiratory tract. In highly pathogenic H5N1 viruses, the presence of a furin cleavage site in the HA leads to replication in multiple tissues and higher pathogenicity, due to the distribution of furins in multiple tissues.

The CoV with the highest nucleotide sequence similarity to SARS-CoV-2 is CoV-RaTG-13, isolated from a bat in Yunnan in 2013. But its S glycoprotein lacks a furin cleavage sequence. The acquisition of a furin cleavage site could be a ‘gain of function’ that enabled a bat CoV to jump into humans. A furin cleavage site could have been acquired by recombination with another virus or could have been engineered in a lab, but in the latter case some signature modifications in the genome should be present, which is not the case. Alternatively, it could have appeared spontaneously when adapting and selecting a virus to grow on human epithelial cells, which has been done in the USA until 2015 and continued in WIV; It has been possible de adapt the MERS virus to mice progressively (Li et al., 2017). Upon introduction into a human, at an outdoor meat market or by leaking from a lab, the modified virus began its epidemic spread.

Further data from a Nat Med commentary that claims SARS-CoV-2 cannot have been engineered nor intentionally modified (Andersen et al., 2020)  (https://www.nature.com/articles/s41591-020-0820-9). Mutations that increase pathogenicity can arise spontaneously (for example in an intermediate host). But they can also result from adaptation and selection of a virus able to grow better and better in human epithelial cell culture, which is exactly what GOF experiments are doing. If this is possible in mice (Yamada and Liu, 2009), why not in humans? Note that, despite US government 2015 moratorium, GOF experiments on MERF CoV have been published as recently as 2020 (Menachery et al., 2020).

Andersen et al acknowledge that, in theory, it is possible that SARS-CoV-2 acquired RBD mutations during adaptation to passage in cell culture, as has been observed in studies of SARS-CoV (Sheahan et al., 2008), and that this should be considered. They then propose arguments against evolution in culture in lab conditions. First, “some SARS-CoV from pangolins have RBD almost identical to that of SARS-CoV-2, providing a stronger and more parsimonious explanation of how SARS-CoV-2 acquired these via recombination or mutation (Cui et al., 2019)”. I don’t understand why this explanation is “stronger and more parsimonious” than selecting virus on human airway cells. Second, “the acquisition of both the polybasic cleavage site and predicted O-linked glycans are also proposed to argue against culture-based scenarios. New polybasic cleavage sites have been observed only after prolonged passage of low-pathogenicity avian influenza virus in vitro or in vivo. Furthermore, a hypothetical generation of SARS-CoV-2 by cell culture or animal passage would have required prior isolation of a progenitor virus with very high genetic similarity, which has not been described”. But this is quite wrong. Bat viruses are quite similar to SARS-CoV-2, except they lack a furin site, which could have been acquired, either spontaneously in culture or by recombination with other coronaviruses in cultured cells. The third argument is that “the generation of predicted O-linked glycans is also unlikely to have occurred due to cell-culture passage, as such features suggest the involvement of an immune system”. If this is true, they might have a point, but it is far from settled (two words underlined).

In sum, arguments by Andersen et al. (2020) can equally be presented in favor rather than against the hypothesis that SARS-CoV-2 is derived from a bat virus, by prolonged passage in human cells or by recombination in cells infected with different viruses.

On April 22, we learn that a new outbreak of SARS-CoV-2 happens in Harbin, a city in north east China close to Siberia. I note that Harbin is the place where the second P4 in China is located, operated by the veterinarian branch of the CAS. They just published in Science that the virus caninfect ferrets better than cats, whereas dogs are resistant. They injected series of animals with the virus. The paper is:
https://www.ncbi.nlm.nih.gov/pubmed/32269068

References (in addition to HTML links in the text)

Andersen KG, Rambaut A, Lipkin WI, Holmes EC, Garry RF (2020) The proximal origin of SARS-CoV-2. Nat Med 26:450-452.

B SB, Orenstein JM, Fox LM, Randell SH, Rowley AH, Baker SC (2009) Human airway epithelial cell culture to identify new respiratory viruses: coronavirus NL63 as a model. J Virol Methods 156:19-26.

Cui J, Li F, Shi ZL (2019) Origin and evolution of pathogenic coronaviruses. Nat Rev Microbiol 17:181-192.

de Wit E, van Doremalen N, Falzarano D, Munster VJ (2016) SARS and MERS: recent insights into emerging coronaviruses. Nat Rev Microbiol 14:523-534.

Ge XY, Yang WH, Zhou JH, Li B, Zhang W, Shi ZL, Zhang YZ (2017) Detection of alpha- and betacoronaviruses in rodents from Yunnan, China. Virol J 14:98.

Huang C et al. (2020) Clinical features of patients infected with 2019 novel coronavirus in Wuhan, China. Lancet 395:497-506.

Li K, Wohlford-Lenane CL, Channappanavar R, Park JE, Earnest JT, Bair TB, Bates AM, Brogden KA, Flaherty HA, Gallagher T, Meyerholz DK, Perlman S, McCray PB, Jr. (2017) Mouse-adapted MERS coronavirus causes lethal lung disease in human DPP4 knockin mice. Proc Natl Acad Sci U S A 114:E3119-E3128.

Li W, Moore MJ, Vasilieva N, Sui J, Wong SK, Berne MA, Somasundaran M, Sullivan JL, Luzuriaga K, Greenough TC, Choe H, Farzan M (2003) Angiotensin-converting enzyme 2 is a functional receptor for the SARS coronavirus. Nature 426:450-454.

Menachery VD, Dinnon KH, 3rd, Yount BL, Jr., McAnarney ET, Gralinski LE, Hale A, Graham RL, Scobey T, Anthony SJ, Wang L, Graham B, Randell SH, Lipkin WI, Baric RS (2020) Trypsin Treatment Unlocks Barrier for Zoonotic Bat Coronavirus Infection. J Virol 94.

Menachery VD, Yount BL, Jr., Debbink K, Agnihothram S, Gralinski LE, Plante JA, Graham RL, Scobey T, Ge XY, Donaldson EF, Randell SH, Lanzavecchia A, Marasco WA, Shi ZL, Baric RS (2015) A SARS-like cluster of circulating bat coronaviruses shows potential for human emergence. Nat Med 21:1508-1513.

Menachery VD, Yount BL, Jr., Sims AC, Debbink K, Agnihothram SS, Gralinski LE, Graham RL, Scobey T, Plante JA, Royal SR, Swanstrom J, Sheahan TP, Pickles RJ, Corti D, Randell SH, Lanzavecchia A, Marasco WA, Baric RS (2016) SARS-like WIV1-CoV poised for human emergence. Proc Natl Acad Sci U S A 113:3048-3053.

Sheahan T, Rockx B, Donaldson E, Sims A, Pickles R, Corti D, Baric R (2008) Mechanisms of zoonotic severe acute respiratory syndrome coronavirus host range expansion in human airway epithelium. J Virol 82:2274-2285.

Shi J et al. (2020) Susceptibility of ferrets, cats, dogs, and other domesticated animals to SARS-coronavirus 2. Science.

Wrapp D, Wang N, Corbett KS, Goldsmith JA, Hsieh CL, Abiona O, Graham BS, McLellan JS (2020) Cryo-EM structure of the 2019-nCoV spike in the prefusion conformation. Science 367:1260-1263.

Wu F, Zhao S, Yu B, Chen YM, Wang W, Song ZG, Hu Y, Tao ZW, Tian JH, Pei YY, Yuan ML, Zhang YL, Dai FH, Liu Y, Wang QM, Zheng JJ, Xu L, Holmes EC, Zhang YZ (2020) A new coronavirus associated with human respiratory disease in China. Nature 579:265-269.

Yamada Y, Liu DX (2009) Proteolytic activation of the spike protein at a novel RRRR/S motif is implicated in furin-dependent entry, syncytium formation, and infectivity of coronavirus infectious bronchitis virus in cultured cells. J Virol 83:8744-8758.

Yang Y, Liu C, Du L, Jiang S, Shi Z, Baric RS, Li F (2015) Two Mutations Were Critical for Bat-to-Human Transmission of Middle East Respiratory Syndrome Coronavirus. J Virol 89:9119-9123.

Yuan Y, Qi J, Peng R, Li C, Lu G, Yan J, Wang Q, Gao GF (2020) Molecular Basis of Binding between Middle East Respiratory Syndrome Coronavirus and CD26 from Seven Bat Species. J Virol 94.

Zhou P et al. (2020) A pneumonia outbreak associated with a new coronavirus of probable bat origin. Nature 579:270-273.