11 June 2021

Dear Pr. Shi Zheng-Li,

These ongoing times are probably putting a lot of pressure on you and I imagine that the situation must be very difficult in the present context. I sincerely hope that you can cope with it well. As shown by the recent Science letter, many people in the scientific community are currently not satisfied with the conclusions of the WHO-China joint report and are asking for a more thorough investigation in order to unravel how SARS-CoV-2 spread into the human population. To try to put an end to the discussions about a possible lab origin of the virus, I think that it would be important to clarify the few elements below. Maybe you could help in this process.

1. In April 2012, after clearing bat guano in an abandoned mine in Mojiang (Yunnan), six men contracted severe pneumonia with COVID-19-like symptoms. All were sent to Kunming hospital where three eventually died. Unspecified samples from these patients were sent to different labs including the Wuhan Institute of Virology (WIV) in 2012. You recently announced that the WIV tested the serum samples again.

• Why were these six miners sent to clean guano in the mine in April 2012? Who hired them and sent them all to the same Kunming hospital?
• Why were these lethal pneumonia cases not mentioned in any scientific research article from the WIV, except the Nature’s Addendum from last November?
• What samples were taken from these six patients and sent to the WIV? Are any of these samples still available for independent analysis?
• Were any SARS-like coronaviruses isolated from the patient samples or were any RdRP or spike sequence obtained by RT-PCR?
• Is it possible to provide to the WHO serum samples from the three surviving miners in order to better understand in which condition these miners fell sick and what their exact pathology was?

2. You stated that Wuhan Institute of Virology virus databases were taken offline during the pandemic. However the key bat virus database was taken offline in September 2019, three months before the official date of outbreak started.

• Can you provide further details on why the database was shut down?
• Can all the databases, in their state as of September 2019, be shared with selected expert scientists?
• Do you have any information concerning the reasons why the scientific paper describing the key database (digital object identifier: 10.11922/csdata.2019.0018.zh) was taken offline from the corresponding Chinese journal website “China Science Data” mid-2020?
• Did you know that even the full website of “China Science Data”, where the database is described, became inaccessible in March-April 2021?

3. A bat coronavirus sampled in the Mojiang mine in 2013 (‘RaTG13’) is a virus most closely-related to SARS-CoV-2. In your interview with Science in July 2020, you said that “Ra4991” was renamed to “RaTG13”, the virus was not isolated, and that there is “no more sample”, so no further sequencing is possible. Unfortunately, based on the raw data provided, it has not been possible to assemble the RaTG13 genome sequence.

• How was the RaTG13 genome sequence assembled and how was the 5’ end sequence determined?
• Was “Ra4991” renamed to “RaTG13” for your article, as the Addendum seems to indicate, or are these 2 viruses divergent in some regions such as the spike protein ?
• When was the RaTG13 sample fully depleted?
• Why are a few of the RaTG13 amplicons dated as of June 2017 and named “7896”, which is the name of another closely related virus collected in the same mine?
• As the row data contains low levels of bacterial sequences, was RaTG13 sampled from bronchoalveolar lavage fluid or from bat feces?
• Given that RaTG13 shows weak binding to bat receptor ACE-2 and binds only to one of the ACE2 orthologs of Rhinolophus affinis, is RaTG13 the true whole genome sequence of the sample BtCoV/4991 collected in 2013 by the WIV?
• Did the WIV or any other laboratory ever attempt to recreate RaTG13 or any other coronaviruses by assembling them from synthetic gene sequences?

4. A striking feature of the SARS-CoV-2 genome is the presence of the furin cleavage site. This site was noted as a “cleavage site” in your January 2020 publication.

• Why was this so-called “furin cleavage site,” clearly an important and novel feature of the SARS-CoV-2 virus, not mentioned in your February 2020 Nature publication?

5. You acknowledged isolating three strains of live SARS-related coronaviruses, but based on the WIV naming convention for their live viruses isolates it appears that the WIV did not disclose two potential isolates, WIV6 (not WIV06) and WIV15, as these names are not mentioned in the literature.

• Do these isolates exist? If not, what is the explanation for why these isolate names were skipped in the series?

6. Chinese authorities have stated that all staff at Wuhan labs tested negative for SARS-CoV-2 antibodies.

• How many people were tested, in which Wuhan labs, on which days, and as part of which teams or services within these labs?
• Were any of these serum samples retained?
• Are independent international investigators able to retest the samples of the lab staff to confirm the results?

7. You previously wrote that: ‘Recently we tested the sera from all staff and students in the lab and nobody is infected by either bat SARSr-CoV or SARS-CoV-2. To date, there is “zero infection” of all staff and students in our institute ’. This statement is statistically unlikely (roughly less than one chance in a billion) given that there are more than 590 staff and students at the WIV and about 4.4% of the Wuhan urban population tested positive at around that time. Even if only 85 people were tested, the chance of no positive test would still be less than 4%.

• How can this unexpected observation be explained?
• Is it possible to have more information on the “one or two” WIV’s workers that fell ill in late 2019, as evoked by a member of the WHO-China joint mission?

8. Recently, you published a preprint presenting the 8 coronaviruses that were mentioned in Nature’s Addendum, in November 2020.

• The preprint indicates that these 8 new coronaviruses, including RaTG15, are constituting a novel lineage of SARSr-CoV from bats: “we report the identification of a novel lineage of SARSr-CoVs, including RaTG15 and seven other viruses, form bats”. When did exactly this identification take place?
• Do you confirm that the lineage called “lineage 4” in a phylogenetic tree (Figure 3.1) in the MS thesis by Yu Ping, “Geographic Evolution of Bat SARS-related Coronaviruses” (June, 2019), is the same lineage as that of the 8 new coronaviruses?
• The preprint mentions that these 8 viruses have been collected in “the same location where we found RaTG13 in 2015”. Why is the 2012 Mojiang mine’s outbreak that occurred at the same location not mentioned, which is important information to understand the context of your research?
• The preprint claims that none of the known viruses in bat SARSr-CoV-2 lineage or the novel lineage discovered so far use human ACE2 efficiently compared to SARSr-CoV-2 from pangolin or some of the SARSr-CoV-1 lineage: “We show that none of the known viruses in bat SARSr-CoV-2 lineage or the novel lineage discovered so far use human ACE2 efficiently compared to SARSr-CoV-2 from pangolin or some of the SARSr-CoV-1 lineage viruses.” To support this statement, is it possible to share the complete list of unpublished viruses that are present in your institute, as well as the raw data and the experimentation records related to them?
• Are these 8 new viruses still kept in your institute, in what condition, and if they are physically available can they be made available to other researchers?
• Can you clarify whether the Bat Ra ACE2 sequence MT394204 used in the pre-print comes from the same Mojiang mine?
• How can you explain that the eight viruses are almost identical? How can you explain that each sequence was found only once along the 7 visits to the mine? Doesn’t it contradict your findings of a “rich gene pool” in the longitudinal study in the caves of Jinning? Did you retest all the 1,322 samples?
• Can you provide the raw sequencing data of the 8 viruses?
• Why do the 8 virus sequences display a few nucleotide changes in their RdRp sequences compared to the previous versions that were submitted to NCBI Genbank for the publication of the Latinne et al. paper? Can you explain the methods employed then and now?
• There is some confusion in the nomenclature of these viruses: in the slide of 2 different presentations from your lab, they were labeled Ra or Rs. Can you confirm the bat species infected with such viruses? Isn’t it surprising that RaTG15 is found in one bat species while the extremely close viruses are from another bat species? Could there be an issue regarding the bat species assignment based on COI sequence?

With my sincere and warmest regards,

Virginie

—

Virginie Courtier-Orgogozo

Institut Jacques Monod – CNRS UMR7592 – Université Paris Diderot Bâtiment Buffon – 4e étage – 420B 15 rue Hélène Brion 75013 Paris France tél : (33) 1 57 27 80 43 fax : (33) 1 57 27 80 87 https://virginiecourtier.wordpress.com

@Biol4Ever

—

Expert advice from scientists to protect yourself, understand, and act to stop the spread of SARS-CoV-2, the coronavirus responsible for COVID-19. https://en.adioscorona.org/