The Mysterious Animal Origin of “SARS-COV-2” Part 1: Bats

The natural origin of “SARS-COV-2,” a “virus” which has never been properly purified/isolated nor proven pathogenic, relies heavily on its zoonotic roots to its closest relative RatG13, a bat “coronavirus.” However, there are serious issues with this theory. The existence of this bat “virus” is highly questionable as its sequence mysteriously appeared around the same time the “SARS-COV-2” sequence was published online. The genome of RatG13 has serious discrepancies/errors and it is seemingly unable to be reproduced in order to be independently verified as the actual source material has been depleted. These are but a few of the issues and a handful of studies showcased below help to break down the mess that is RatG13. I will provide highlights from each one and put together a summary of the pertinent points at the end:

Anomalies in BatCoV/RaTG13 sequencing and provenance

“To this date, the most critical piece of evidence on the purposed “natural origin” theory of SARS-CoV-2, was the sequence known as RaTG13, allegedly collected from a single fecal sample from Rhinolophus Affinis. Understanding the provenance of RaTG13 is critical on the ongoing debate of the Origins of SARS-CoV-2. However, this sample is allegedly “used up” and therefore can no longer be accessed nor sequenced independently [1], and the only available data was the 3 related Genbank accessions: MN996532.1, SRX7724752 and SRX8357956.

We report these datasets possessed multiple significant anomalies, and the provenence of the promised claims of RaTG13 or it’s role in proving a “probable bat origin”[2] of SARS-CoV-2 can not be satisfied nor possibly be confirmed.”

“The raw data of BtCoV/RaTG13 contained multiple anomalies that signifies that the original sample could not have contained enough RNA template for the extraction of a complete viral genome as in MN996532.1.

Furthermore, many of these anomalies points toward the fraudulent use of a mixed DNA library, rather than genuine mRNA, for the sequencing of SRX7724752, evident by the presence of widespread A-T ligation of unrelated dsDNA fragments that can only happen if the same library preparation process have been ran on dsDNA instead of ssRNA. which would constitute Academic fraud.

The Spike glycoprotein of RaTG13 does not resemble that of a wild virus but instead possessed multiple signatures of artificial attenuation in a cell culture when compared to the SARS-CoV-2 Spike and the Spike sequences of other related viruses, indicating that the sequence did not derive from what the Wuhan Institute of Virology claimed to be. Therefore, the sequencing of BtCoV/RaTG13 cannot be considered to be valid or honest as is, and any publications, including [2], and other publications that cites or use RaTG13 as critical pieces of evidence or proof, must be immediately invalidated and retracted.”

https://zenodo.org/record/3969272#.YCKgs-pME0M

It can be seen that there are many discrepancies in the RatG13 dataset and that there is evidence of academic fraud in the methods used for its creation. Many independent researchers have had difficulty verifying the accuracy of RatG13. On top of this, the origins for this bat “Coronavirus” are as mysterious as that of “SARS-COV-2:”

The genetic structure of SARS-CoV-2 does not rule out a laboratory origin

“Zhou et al.[3] from the Wuhan Institute of Virology (WIV) were the first to identify and characterize a new coronavirus (CoV), SARS-CoV-2. The genomic sequences obtained from early cases shared 79% sequence identity to the CoVs that caused severe acute respiratory syndrome (SARS-CoV) in 2002–2003 and 96.2% sequence identity to RaTG13 (MN996532), a CoV sequence detected from a Rhinolophus affinis bat. RaTG13 is currently the closest phylogenetic relative for SARS-CoV-2 found,[4] but its complete genomic sequence was not published before the outbreak of SARS-CoV-2 and the original sample was collected in the Yunnan province (China) by the same group of WIV researchers in 2013. Zhou et al.[3] stated to have found a match between SARS-CoV-2 and a short region of RNA-dependent RNA polymerase (RdRp) of a CoV in their database and then fully sequenced the original sample collected in 2013, which they called RaTG13.

We discovered that the RdRp of RaTG13 has 100% nucleotide identity with the sequence BtCoV/4991 (KP876546), which was identified by Ge et al.[5] in a Rhinolophus affinis bat in the Yunnan province in 2013, same location and year as RaTG13. BtCoV/4991 was collected in a mine colonized by bats near Tongguanzhen, Mojiang, Yunnan. The WIV researchers were invited to investigate the mine after six miners there had contracted severe pneumonia in 2012iii , and three of the miners have died.[6] The miners have been tasked with clearing out bat droppings in the mine, and the severity of their pneumonia correlated with the duration of exposure to the mine.[7] Four miners’ samples subsequently underwent testing at WIV, where Immunoglobulin G (IgG) antibodies against SARS were identified in all samples.[8] Considering that only about 5300 people were infected in mainland China during the SARS outbreak of 2002–2004, most of whom resided in Guandong, the odds of four miners in Yunnan retaining antibodies from the 2002–2004 SARS outbreak are negligible. On the other hand, it is possible that the SARS antibody test administered to the miners cross-reacted with a novel SARS-like bat virus that the miners had acquired at the mine. Ge et al.[5] have identified a number of CoVs in the mine, but based on the phylogenetic analysis, BtCoV/4991 was the only SARS-related strain, clearly separated from all known alpha- and beta-CoVs at that time. BtCoV/4991 was also different from other bat CoVs in the phylogenetic analysis carried out by Wang et al. in 2019.[9] Chen et al.[10identified BtCoV/4991 as the closest sequence to SARS-CoV-2 because RaTG13 had not yet been published at that time. BtCoV/4991 and RaTG13 have been later asserted to be two different coding names of the same strain, as their original authors at WIV registered the two strains as one entry in the Database of Bat-associated Viruses (DBatVir).iv

In late July 2020, Zhengli Shi, the leading CoV researcher from WIV, in an email interview [11] asserted the renaming of the RaTG13 sample and unexpectedly declared that the full sequencing of RaTG13 has been carried out as far back as in 2018 and not after the SARS-CoV-2 outbreak, as stated in Zhou et al.[3] The reversal in WIV’s stance on when exactly RaTG13 was fully sequenced could have been due to the discovery by independent researchers into the origins of SARS-CoV-2 that the filenames of the raw sequencing reads deposited by WIV on May 19, 2020v seem to indicate that sequencing for RaTG13 was done in 2017 and 2018.vi However, no formal erratum about year of sequencing and sample renaming from the authors of Zhou et al. [3] has yet appeared, or as far as is currently known, has been submitted.”

It is important to mention that RaTG13 and the pangolin CoV sequences from smuggled pangolins confiscated in the GD province in March 2019, and to which most of published papers supporting a natural origin of SARS-CoV-2 refer,[2] have recently been questioned as to the accuracy of their assembly dataxii and require further analyses to prove their correctness.[xiii ,xiv ] It should also be noted that in vitro receptor binding studies of reconstituted RaTG13 yielded some peculiar results.[xi] The most surprising observation was that RaTG13, unlike SARS-CoV-2, is unable to bind ACE2 in R. macrotis bats, a close relative of RaTG13’s purported host, R. affinis[59] (whose ACE2 receptor has not yet been tested). At the same time, RaTG13 was observed to bind hACE2[60], but not as well as ACE2 of rats and mice, to which SARS-CoV-2 did not bind at all. Is it possible that just as SARS-MA15 was a mouse-adapted strain of SARS, RaTG13 is actually a mouse-adapted version of a CoV extracted from the Mojiang cave, rather than a strain obtained from a bat fecal swab? Unfortunately, the RaTG13 sample has been exhausted and it is no longer available for external examination,[11] which is unfortunate given a number of inconsistencies in its sequencing raw data. Also, the status and availability of the Mojiang miners’ samples remain as well an open and highly relevant question. Several samples from the miners have been collected[78] and likely stored, and it would be of great value to test them for the presence of SARS-CoV-2-like CoVs.

Another open question is the reason for modification and subsequent deletion of WIV’s own viral database. In May 2020, several media outlets have reported that the change tracking system of WIV’s internal database showed that the database was renamed from “Wildlife-borne viral pathogen database” to “Bat and rodent-borne viral pathogen database,” and its description was edited to replace instances of “wild animal” by “bat and rodent”; in addition, mention of “arthropod vectors” was deleted.xv The database description reported that it contained over 60 Mb of data in structured query language (SQL) format, but at as of early May 2020 the download link no longer worked.xvi Subsequently, the database page was taken down in its entirety but its snapshot is still available on Web Archive.xvii It is possible that other international CoV labs might have downloaded the SQL archive of the WIV database before it was taken down, in which case such groups should make those data publicly available.”

“On the basis of our analysis, an artificial origin of SARS-CoV-2 is not a baseless conspiracy theory that is to be condemned[66] and researchers have the responsibility to consider all possible causes for SARS-CoV-2 emergence.”

https://onlinelibrary.wiley.com/doi/full/10.1002/bies.202000240

According to the above source, RatG13 was apparently not sequenced before the “SARS-COV-2” outbreak…until it was…once they changed the dates in the computer as well as their stories. As mentioned previously, there appears to be much difficulty in independently verifying RatG13 as well as tracking its origins. The samples from which the RatG13 genome came from no longer exist and there are some mysterious changes as well as the eventual deletion of the WIV database.

This last study draws out some further anomalies/errors with the RatG13 genome such as the existence of numerous sources of DNA within the genome which come from animals other than the bat species it supposedly comes from as well as an extremely low makeup of “viral” sequences:

The Anomalous Nature of the Fecal Swab Sample Used for RaTG13 Genome Assembly as Revealed by NGS Data Analysis

“RaTG13 beta coronavirus, which exists in the form of a genome sequence, is the closest relative of SARS-CoV-2 reported till date. The sample from which RaTG13 virus was sequenced was a bat fecal swab collected in 2013 from Tongguan, Mojiang, Yunnan province, China. The genome data for RaTG13, MN996532.1, was deposited on 27th Jan 2020 and the raw data (Illumina reads) was deposited a fortnight later on 13th Feb 2020 https://www.ncbi.nlm.nih.gov/sra/SRX7724752%5Baccn%5D. Comparison of the RNA Seq data of RaTG13 fecal swab sample to the corresponding data from the bat fecal swabs deposited by the same working group indicated that the raw data seemed to be anomalous in several aspects. Thirty percent of the reads did not match with anything. From the rest of the 70%, an abnormal high proportion was contributed by reads derived from eukaryotes (~68%). These matched with the sequences of not one but various bat species (round leaf bats, fruit bats and other bats) and animal species (squirrels, foxes, etc.) as per Krona analysis included with the SRA data. The proportion of the bacterial reads in the swab was exceptionally low, i.e. 0.7%, which is abnormal, compared to the 70-90% bacterial abundance in other bat fecal swabs. Furthermore, we also found another set of raw data associated with RaTG13, amplicon sequencing of the genome (SRX8357956), which was submitted in May 2020. Analysis of the amplicons by BLAST showed that these collectively do not cover the whole genome (MN996532.1). On closer inspection, the dates mentioned in the files of the sequenced amplicons were also found to be older (2017, 2018). Collectively, the anomalies in the raw data of RaTG13 pose an important question about the overall authenticity of the RaTG13 genome sequence.”

The coronavirus sequence (RaTG13) contributes to ~0.003% of the total sequence reads. These raw reads were used to build an almost complete assembly, though the overall coverage is less ~7-8X. Though there were less overlaps in a few regions, there are only 3 gaps as per our analysis using RaTG13 as the reference genome.”

https://www.preprints.org/manuscript/202008.0205/v2

In Summary:

  • The sample used to sequence RatGq3 is allegedly “used up” and therefore can no longer be accessed nor sequenced independently
  • The datasets possessed multiple significant anomalies
  • The original sample could not have contained enough RNA template for the extraction of a complete “viral” genome
  • Many of these anomalies points toward the fraudulent use of a mixed DNA library which would constitute academic fraud
  • The Spike glycoprotein of RaTG13 does not resemble that of a wild “virus” but instead possessed multiple signatures of artificial attenuation in a cell culture
  • The complete genomic sequence was not published before the outbreak of “SARS-CoV-2” and the original sample was collected in the Yunnan province (China) by the same group of WIV researchers in 2013
  • BtCoV/4991 (2013) and RaTG13 (2020) were later asserted to be two different coding names of the same strain, as their original authors at WIV registered the two strains as one entry in the Database of Bat-associated Viruses
  • In an email from July 2020, the WIV asserted the renaming of the RaTG13 sample and unexpectedly declared that the full sequencing of RaTG13 has been carried out as far back as in 2018 and not after the SARS-CoV-2 outbreak, as stated in Zhou et al.
  • No formal erratum about year of sequencing and sample renaming from the authors of Zhou et al. has yet appeared, or as far as is currently known, has been submitted
  • The sequences for both RatG13 and the Pangolin (considered another close relative) have recently been questioned as to the accuracy of their assembly data and require further analyses to prove their correctness
  • Another open question is the reason for modification and subsequent deletion of WIV’s own “viral” database
  • WIV’s internal database showed that the database was renamed from “Wildlife-borne viral pathogen database” to “Bat and rodent-borne viral pathogen database,” and its description was edited to replace instances of “wild animal” by “bat and rodent”
  • As of early May 2020, the download link no longer worked and the database page was taken down in its entirety
  • Analysis showed that an artificial origin of SARS-CoV-2 is not a baseless conspiracy theory that is to be condemned
  • The “virus” RatG13 waa based on has never been isolated or recovered from any sample and the sample itself is no longer available for any further validation
  • The published genome sequence of RaTG13, which was first published at the same time as that of the “SARS-COV-2” genome, is the stand-alone proof of its existence
  • Only 0.7% of the dataset is of bacterial origin, which should actually constitute the bulk (70-90%) of DNA sequences isolated from a bat fecal sample
  • 30% of the data did not match anything, and the rest instead matched with a number of different bats and other animals such as Mexican bats, squirrels, flying foxes, red foxes, etc.
  • The “virus” itself comprised only 0.003% of the total sample

It is clear that there are many problems with the RatG13 genome dataset. There are very legitimate questions as to it’s accuracy. Whether or not the claims are valid would require further independent analysis. Unfortunately, the sample used to sequence RatG13 is conveniently all used up which makes it impossible to verify the accuracy of its genome.

All of this is to say that “SARS-COV-2,” as it was never properly purified/isolated, is only as good as its reference genomes. If the RatG13 reference genome is of questionable origin/existence, then the existence of the “virus” that shares 96.2% sequence identity with it must also be questioned.

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