The Origins of COVID-19 by Dr Sam Bailey
March 22nd 2023 Why I don’t believe there ever was a Covid virus - Dr Mike Yeadon https://www.conservativewoman.co.uk/why-i-dont-believe-there-ever-was-a-covid-virus/
March 11th 2022 Top 20 Origins Of Covid19 Part 3 ft You'll Believe Anything https://nurembergtrials.net/nuremberg-trials-2-0/f/poll-top-20-origins-of-covid-19-ft-youll-believe-anything
September 20th 2021 Covid Origin Revisit: Coal Mine, Fish Market, Wuhan Lab, Phantom? https://nurembergtrials.net/nuremberg-trials-2-0/f/origins-of-covid-19-revisited-coal-mine-fish-market
Two Years of Origins of Covid19: 1) Coal Mine 2) Wet Market 3) 5G 4) China Bio-Lab 5) Bat Bites 6) Chemtrail Covid (Alex Jones) 7) Cats & Dogs (NIH) 8) Pangolins (NIH) 9) Animal Products (WHO) 10) Vampires in Ukraine 11) Ukraine Bio-Lab 12) Fukushima Radiation 13) Aliens 14) Asteroids 15) Bigfoot 16) Popped Out Of No-where (LA Times) 17) The Swamp (Biden, Newsom & Bloomberg) 18) GMO Mosquito's (Bill Gates) 19) All Of The Above 20) I'll Believe Anything
The original report of "isolation" came from a report that they isolated "virus particles" from the anal swab of a monkey... (SNIP) In the same fashion, Koch Postulates to "Isolate" a "Virus" or duplicated "genetic material" for Covid has only been done with Monkeys, with anal swabs, and only found "virus particles". https://ncbi.nlm.nih.gov/pmc/articles/PMC7095368/
https://nurembergtrials.net/nuremberg-trials-2-0/f/virology-debunked-w-cracker-jack-pcr-test---10-points-on-the-box https://nurembergtrials.net/nuremberg-trials-2-0/f/20-fake-covid-19-tests-none-look-for-a-whole-unique-virus https://nurembergtrials.net/nuremberg-trials-2-0/f/666-ways-covid-19-is-a-full-blown-hoax-by-tesla-leaks-2921
March 13th 2022 VIROLOGY ON TRIAL PART 6: FREEDOM POISONED: IS VIROLOGY ROOTED IN NONSENSE & SILENCE? LARGEST CRIMES EVER?! https://www.bitchute.com/video/RckOAckiSmTG/
Bonus: March 11th 2022 MEDIA CONTROL: POPULATION PREPPED FOR NEXT COMMUNISM VARIANTS (BA 2 & DELTACRON) aka Bare Ass Part 2 w/ Covid19 Toilet Trackers https://www.bitchute.com/video/MFterQgBJlsE/
Breaking: March 11th 2022 Republicans Will, Eventually, Pivot to the Position That Zelensky Is a Secret Sex Criminal Who Invented COVID https://slate.com/news-and-politics/2022/03/tucker-carlson-madison-cawthorn-republicans-ukraine.html
March 11th 2022 Pfizer CEO: Virus's are now "smart", "knows how to mutate", "knows how to trust the science". This is 100% proof your a fucking idiot and you'll believe anything!
Meaning of Omi-Cron or Omni-Cron aka a False Messiah or the Devil aka Imhotep aka Fauci: The all knowing Sickle Maker or "Death Personified":
Omni- Is a Latin prefix meaning "all" knowing or "every" thing https://en.wikipedia.org/wiki/Omni
Omi- Portending evil or harm; foreboding; threatening; inauspicious: an ominous bank of dark clouds. https://villains.fandom.com/wiki/Evil_Omi
Cron- An omnipresent being whos knowledge is unquestionable and power is all encompassing. https://en.wikipedia.org/wiki/Cron
September 20th 2021 Origins of Covid19 Part 2: Coal Mine, Fish Market, Wuhan Lab, Phantom? https://nurembergtrials.net/nuremberg-trials-2-0/f/origins-of-covid-19-revisited-coal-mine-fish-market
June 10th 2021 Origins of Covid19 Part 1: Common Cold, Guano, Pangolin or Vaccine Bioweapon! https://nurembergtrials.net/nuremberg-trials-2-0/f/origins-of-coronavirus---the-common-cold-bat-guano-or-pangolin
March 22nd 2021 A pandemic of jokes? The Israeli COVID-19 meme and the construction of a collective response to risk https://www.degruyter.com/document/doi/10.1515/humor-2021-0012/html
May 6th 2020 These are the 35 best COVID-19 memes for the week of May 4 https://www.wearethemighty.com/mighty-survival/covid-19-memes-2645929842/
Covid's Top 10 Origins Revisited: Where's the original Covid "Virus", SARS-1 or SARS-2 Covid-19 or Variants? Since PCR tests are for copying genetic sequences and identifying "markers"; not actual Virus's 1) Bat bites Man in a coal plant in West China 2) Bat Guano found 1000 miles away in a Fish Market in East China 3) The Pangolin 4) The Cold Virus 5) Escaped from the Wuhan Lab 6) A computer generated genetic code uploaded to the public Gene Database by China; from the "original" Pneumonia "cases"; then removed by China from the Gene Database 7) The gene sequence' then used in Vaccine's using Vaccinology as a Bio-weapon 8) Popped out of nowhere as reported by the LA Time's 9) Sprayed out of airplanes as "chemtrails" as reported by Alex Jones, Mike Adams etal 10) Jumped, hopped, skipped or otherwise transmitted into Humans from the feces of a Chimpanzee
-----> It can't be all 10. https://nurembergtrials.net/nuremberg-trials-2-0/f/origins-of-coronavirus---the-common-cold-bat-guano-or-pangolin
Bat Guano - The #1 Competitor to Dow Chemical's Atrazine Herbicide. As a manure, guano is a highly effective fertilizer due to its exceptionally high content of nitrogen, phosphate, and potassium: key nutrients essential for plant growth. Guano was also, to a lesser extent, sought for the production of gunpowder and other explosive materials.
Cold Virus - Coronaviruses comprises of a big class of (cold) viruses that consists of a core of genetic material (single-stranded RNA genome) enveloped by a spherical lipid layer with spikes like protein structure attached to the surface. This structure gives it a crown-like appearance, and in Latin, corona means crown, so this is why, these viruses are labelled as coronaviruses [3, 4]
Pangolin - Although a SARS-CoV-2-like coronavirus was detected in the lungs of these pangolins, a direct association between the clinical signs or pathology and active virus replication is not available as we lack evidence from immunohistochemistry or in situ hybridization experiments. The experimental infection of healthy pangolins with pangolin-CoV would provide more definitive answers; however, as pangolins are protected it is difficult to carry out such experiments. Further studies are needed to confirm the role of pangolins in the transmission of SARS-related coronaviruses. Our findings support the call for stronger enforcement of regulations against the illegal trade in pangolins. Owing to the demand for their meat as a delicacy and their scales for use in traditional medicine in China, the illegal smuggling of pangolins from Southeast Asia to China is widespread18. International co-operation in the implementation of stricter regulations against illegal wildlife trade (and illegal fish market) and consumption of game meat should be encouraged, as this will increase the protection of endangered animals and help to prevent future outbreaks of diseases caused by SARS-related coronaviruses.
BATS & BIRDS & HUMANS OH MY!
First Bats, now Humans?? Recently, on the basis of genomic resemblance with previously reported SARS-CoV, the International Committee on Taxonomy of Viruses (ICTV) coronavirus study group named this virus “SARS-CoV-2” [13]. It has been confirmed that SARS-CoV-2 can spread with human-to-human transmission via respiratory droplets (e.g. through coughing or sneezing) or even by contact with contaminated surfaces [14, 15]. https://translational-medicine.biomedcentral.com/articles/10.1186/s12967-020-02439-0
EVOLUTIONARY THEORY OF MUTATIONS
Why there is no antiviral drugs? Because, none worked?? This could be the case for SARS-CoV-2, in that viral proteins are actively acquiring mutations due to SNPs and thus escape from being targeted by antiviral drugs
IF IT LOOKS LIKE A DUCK, QUACKS LIKE A DUCK, MUST BE A DUCK!
Genome organization of all coronaviruses are similar and contain 5' and 3' untranslated regions (UTR’s) for characteristic genes coding for ORF1ab, spike, envelope, membrane, and nucleocapsid proteins [24]. An #evolutionary-based study was performed to understand the relationships of SARS-CoV-2 genome with other SARS isolates. #Homology modeling for predicting SARS-CoV-2 RdRp structure was performed with a reference template of SARS-CoV RdRp.
VACCINOLOGY
Of 932, 94 total sequences were selected that exhibited > 90% similarity with our input RdRp sequence.
The best antiviral drug was selected with distinct S-score, root-mean-square deviation (RMSD)
Most importantly, clade III—which included the experimental SARS-CoV-2 isolate originating from the Wuhan seafood market
Modeling, predictions and estimates. within favorable regions ... Ramachandran plot shows 94.1% in most favored regions.
An MLE tree constructed for all validated proteins revealed that most of the proteins shared (close association) with each other.
#Similarty, #close relation, #resemblance ... but sufficiently diverge. this virus may show similarity to other viruses based on nucleotide sequences, such as MERS, which could help to find attractive therapeutic options [53, 54, 56, 57]. It has been reported that nucleotide sequence-based analysis of SARS-CoV-2 and SARS-CoV (Bats origin) represent a close relation with each other but sufficiently divergent due to single nucleotide mutations [13, 21, 23, 56, 57]. Most of the viral species examined showed resemblance to each other and clustered together.
First Humans, Now Bats! SARS-CoV-2 whole-genome sequences reported from Hong Kong and USA (MN975262.1, MN997409.1, MN994467.1, MN988713.1) revealed close relation with each other as they originated from same clade and are considered as sister species (Fig. 1). Our results are consistent with the results of Wu et al. [58], who demonstrated that SARS-CoV-2 and bat-origin SARS-CoV clustered together, revealing similarity in their genomes (Fig. 1).
However, the latter group only predicted 10 models, instead of the ~ 100 structures here. In addition, of these possible structures we selected the most suitable among them to predict the 3D structure of the RdRp protein of SARS-CoV. It is suggested that the #close #association between all these RdRps is due to the similarity in their genome organization. Therefore, more research is needed to explore the differentiation between these viral RdRps.
https://translational-medicine.biomedcentral.com/articles/10.1186/s12967-020-02439-0
However, all these approaches have intrinsic #limitations. Bioinformatic prediction tools are trained on sets of previously described epitopes, but since the available epitope repertoire for many human leukocyte antigen (HLA) alleles is limited, its prediction capacity is also limited [20,21]. Inferences based on epitope sequence homology with other coronaviruses are hampered because past studies on SARS-CoV-1 and MERS only included few selected viral proteins.
T cell recognition of epitopes is very sensitive to mismatches and not matching the recall antigen with the autologous virus can lead to missed responses
HIGHLY VARIABLE VIRUS'S "May not warrant inclusion into a single test"
Direct comparisons of these different central sequence approaches have been performed for a highly variable pathogen (human immunodeficiency virus, HIV) and shown that the different designs yielded comparable results when synthetic peptides covering these sequences were used to measure virus-specific T cell responses [42,43]. However, the additional costs in terms of peptide synthesis and cells needed for ex-vivo experiments, may not warrant inclusion of all the different variants into a single test set.
SCIENCE BY CONSENSUS
2.1. Consensus Sequence ORF Generation and Entropy Calculation. A nucleotide consensus sequence was generated by keeping all nucleotides present in at least 25% of the sequences in the alignment. 50 reference sequences (RefSeq) with the lowest E-values resulting from a pBLAST search [51] using the ORF-specific consensus sequences (pan-coronavirus alignment) (ii) homologous proteins from 17 viruses representing the Betacoronavirus taxon (beta-coronavirus alignment) or, (iii) homologous proteins from the 7 full-genome sequenced human coronaviruses (including SARS-CoV, MERS-CoV, and common cold species OC43, NL63, 229E, HKU1, human-coronavirus alignment).
For creation of the CoV-2 Consensus sequence, nucleotide sequences from 1731 SARS-CoV-2 genomes were aligned and a full genome nucleotide consensus was created, 23 open reading frames (ORF) were then located in the alignment using the NC_045512.2 and the Finkel et al. [46] coordinates and translated to amino acids. Of the 23 ORF, 12 were canonical ORF as annotated in NC_045512.2 and 11 in alternative reading frames described by Finkel et al. Covid Variability Mismatches between the sequence of in vitro antigen sets and the autologous virus in an infected individual can lead to missed responses. In the CoV-2-cons 15–11 OLP set, mean OLP normalized entropies were overall low (Range: 0.947–0.758) and comparable between OLP covering the canonical ORF (Range: 0.947–0.879) and OLP matching the alternative frameshift ORF (Range: 0.932–0.758).
APPLES AND ORANGES ARE SIMILAR "They are both round"!
Based on the SARS-CoV-2 alignment used to design the consensus, only nine amino acid positions in the entire SARS-CoV-2 genome showed two amino acids present in at least 25% of the sequences.
(>90% sequence identity) 125 #similar epitopes were identified. The similar epitopes were found to be derived from the following organisms; SARS-CoV: 71, Human coronavirus 229E: 1, Alphacoronavirus 1: 1, Unknown origin: 3, and Homo sapiens:
However, longer test peptides tend to yield fewer responses and imply bigger efforts for subsequent epitope mapping. Canonical and alternative frame ORF were considered in the present CoV-2-consensus sequence design to ensure an as broad as possible screening for all potentially expressed protein sequences. Consensus sequence design is highly dependent on the sequences included in the alignments used to construct them. As noted, coverage of sequence diversity for in-vitro antigen test sets is critical as responses to autologous viral variants may be missed if these #variant sequences are not matched [27]. To achieve this, peptide synthesis uses mixes of amino acids at variable positions, so that the resulting OLP resembles a mini-peptide library that can achieve an a-priori set coverage of circulating viral variants [64]. This would readily allow to cover more sequence diversity beyond the 25% frequency cut-off that was applied in the present study. For the interpretation of T cell responses, it has to be taken into account that some degree of cross-reactivity can exist among human coronavirus [5,65]. This implies that responses to these regions could be associated with previous infections by other human coronaviruses, some of them triggering much milder infections that can pass unnoticed, like those by coronaviruses causing a common cold.
Such a vaccine could allow to prevent infection not only with SARS-CoV-2, but also with other, clinically relevant coronavirus like SARS-CoV-1 and MERS, and even with new coronaviruses jumping the species barrier to humans.
At the same time, the existence of SARS-CoV-2 homologous regions in the human genome, together with the existence of described epitopes in these regions raise some concern that coronaviruses could be involved in a molecular mimicry process triggering autoimmune diseases like the Guillain-Barré syndrome [66,67,68,69].
The present study is currently limited to the design of the CoV-2 consensus sequence, without functional immune analyses of the OLP sets in samples from infected individuals. included ~30 randomly selected sequences per host family or region to mitigate sampling and surveillance intensity bias. phylogenetic reconstructions for a-CoVs in China suggest an evolutionary origin within rhinolophid and vespertilionid bats (Fig. 2a). suggesting that frequent cross-species transmission events have occurred among bats.
At the genus level, we identified 20 highly supported inter-genus host switches for a-CoVs, 17 of them were also highly significant using the random subset (Fig. 5a and Supplementary Table 6). In order to identify the hotspots of CoV phylogenetic diversity in China and evaluate phylogenetic clustering of CoVs, we calculated the mean phylogenetic distance (MPD) and the mean nearest taxon distance (MNTD) statistics50 and their standardized effect size (SES).
https://www.mdpi.com/2076-393X/8/3/444/htm
COVID-19 CAUSATIVE AGENT "the evolution and diversification of these coronaviruses remains poorly understood"
Bats are #presumed reservoirs of diverse coronaviruses (CoVs) including progenitors of Severe Acute Respiratory Syndrome (SARS)-CoV and SARS-CoV-2, the causative agent of COVID-19. However, the evolution and diversification of these coronaviruses remains poorly understood. Here we use a Bayesian #statistical framework and a large sequence data set from bat-CoVs (including 630 novel CoV sequences). Finally, we present a phylogenetic analysis suggesting a likely origin for SARS-CoV-2 in Rhinolophus spp. bats.
COVID-19 ORIGIN Animal to Human Transmission was "thought" to be the "likely origin"
All CoVs known to infect humans are zoonotic, or of animal origin, with many #thought to originate in bat hosts1,2. (the largest non-segmented RNA viral genome). This phenomenon is thought to have led to the emergence of a number of CoVs affecting livestock and human health4,5,6,7,8,9. Subsequent investigations identified horseshoe bats (genus Rhinolophus) as the natural reservoirs of SARS-related CoVs and the likely origin of SARS-CoV13,14,15,16
This emerging human virus is closely related to SARS-CoV, and also appears to have originated in horseshoe bats20,21—with its full genome 96% similar to a viral sequence reported from Rhinolophus affinis20. Closely related sequences were also identified in Malayan pangolins22,23. The emergence of SARS-CoV-2 further underscores the importance of bat-origin CoVs to global health, and understanding their origin and cross-species transmission is a high priority for pandemic preparedness20,29. Host switching of viruses over evolutionary time is an important mechanism driving the evolution of bat-CoVs in nature and appears to vary geographically32,33. However, detailed analyses of host switching have been hampered by incomplete or opportunistic sampling, typically with relatively low numbers of viral sequences from any given region34. However, the macroevolution of CoVs in their bat hosts in China and their cross-species transmission dynamics remain poorly understood. We use a phylogeographic Bayesian statistical framework to reconstruct virus transmission history between different bat host species and virus spatial spread over evolutionary time. We generated 630 partial sequences (440 nt) of the RNA-dependent RNA polymerase (RdRp) gene from bat rectal swabs collected in China and added 608 bat-CoV and 8 pangolin-CoV sequences from China available in GenBank or GISAID to our datasets. #guano
CORRELATION IS NOT CAUSATION
Mantel tests revealed a positive and significant #correlation between CoV genetic differentiation (FST) and geographic distance matrices, both with and without provinces, including fewer than four viral sequences
In our phylogenetic analysis that includes all known bat-CoVs from China, we found that SARS-CoV-2 is likely derived from a clade of viruses originating in horseshoe bats (Rhinolophus spp.). For these reasons, we cannot rule out an origin for the clade of viruses that are progenitors of SARS-CoV-2 that is outside China, and within Myanmar, Lao PDR, Vietnam, or another Southeast Asian country. Finally, while our analysis shows that the RdRp sequences of CoVs from the Malayan pangolin are closely related to SARS-CoV-2 RdRp, analysis of full genomes of these viruses suggest that these terrestrial mammals are less likely to be the origin of SARS-CoV-2 than Rhinolophus spp. bats22,23. This analysis also demonstrates that a significant amount of cross-species transmission has occurred among bat hosts over evolutionary time. The Rhinolophidae comprises a single genus, Rhinolophus, and is the most speciose bat family after the Vespertilionidae in China51, with 20 known species, just under a third of global Rhinolophus diversity, mostly in Southern China35. This family likely originated in Asia52,53, but some studies suggest an African origin54,55. A significant correlation between geographic distance and genetic differentiation of both a- and ß-CoVs has been detected, even if only a relatively small proportion of the variance is explained by geographic distance.
WHAT? THE SPIKE PROTEIN IS USED TO USER IN THE VIRUS INTO THE HOST CELL?
((The ectodomain region of the spike protein (S) is essential for attachment and eventual entry of the viral protein into the host cell.)) Variation in the extent of host jumps between a- and ß-CoVs within the same hosts in the same environment may be due to virus-specific factors, such as differences in receptor usage between a- and ß-CoVs60,61,62., CoVs use a large diversity of receptors, and their entry into host cells is mediated by the spike protein with an ectodomain consisting of a receptor-binding subunit S1 and a membrane-fusion subunit S263. An ectodomain is the domain of a membrane protein that extends into the extracellular space (the space outside a cell). ...
First, only partial RdRp sequences were generated in this study and used in our phylogenetic analysis as the noninvasive samples (rectal swabs/feces) collected in this study prevented us from generating longer sequences in many cases. The RdRp gene is a suitable marker for this kind of study as it reflects vertical ancestry and is less prone to recombination than other regions of the CoV genome such as the spike protein gene16,72. Second, most sequences in this study were obtained by #consensus PCR using primers targeting highly conserved regions. Even if this broadly reactive PCR assay designed to detect widely variant CoVs has proven its ability to detect a large diversity of CoVs in a wide diversity of bats and mammals 30,74,75,76,77, we may not rule out that some bat-CoV variants remained undetected. Using deep sequencing techniques would allow to detect this unknown and highly divergent diversity. Additionally, faster rates of #evolution in the tropics have been described for other RNA viruses that could favor cross-species transmission of RNA viruses in these regions81. Both SARS-CoV and SADS-CoV emerged in this region, and several bat SARSr-CoVs with high zoonotic potential have recently been reported from there, although the dynamics of their circulation in wild bat populations remain poorly understood16,61.
PCR TESTS AT 40 CYCLES CAN IDENTIFY A SINGLE FRAGMENT AND "OPTIMIZED FOR BATS"?
RNA was extracted from 200 µl swab rectal samples or fecal pellets with the High Pure Viral RNA Kit (Roche) following the manufacturer’s instructions. RNA was eluted in 50 µl elution buffer and stored at -80 °C. A one-step hemi-nested reverse transcription-PCR (Invitrogen) was used to detect CoV RNA using a set of primers targeting a 440-nt fragment of the RdRp gene and optimized for bat-CoV detection (CoV-FWD3: GGTTGGGAYTAYCCHAARTGTGA; CoV-RVS3: CCATCATCASWYRAATCATCATA; CoV-FWD4/Bat: GAYTAYCCHAARTGTGAYAGAGC)82. For the first round PCR, the amplification was performed as follows: 50 °C for 30 min, 94 °C for 2 min, followed by 40 cycles consisting of 94 °C for 20 s, 50 °C for 30 s, 68 °C for 30 s, and a final extension step at 68 °C for 5 min. For the second round PCR, the amplification was performed as follows: 94 °C for 2 min, followed by 40 cycles consisting of 94 °C for 20 s, 59 °C for 30 s, 72 °C for 30 s, and a final extension step at 72 °C for 7 min. PCR products were gel purified and sequenced with an ABI Prism 3730 DNA analyzer (Applied Biosystems, USA). PCR products with low concentration or bad sequencing quality were cloned into pGEM-T Easy Vector (Promega) for sequencing. Positive results detected in bat genera that were not known to harbor a specific CoV lineage previously were repeated a second time (PCR + sequencing) as a confirmation. Species identifications from the field were also confirmed and re-confirmed by cytochrome (cytb) DNA barcoding using DNA extracted from the feces or swabs83. Only viral detection and barcoding results confirmed at least twice were included in this study. Our final datasets include 630 sequences generated for this study and 616 sequences from GenBank. Model selection and phylogenetic analysis. Preliminary analysis were run to select the best-fitting combination of substitution models. TEMPEST showed that both datasets did not contain sufficient temporal information to accurately estimate substitution rates or time to the most recent common ancestor. Therefore, we used a fixed substitution rate of 1.0 for all our BEAST analysis. The MPD and the MNTD statistics50 and their SES were calculated for each zoogeographic region, bat family, and genus using the R package picante94. All subsequent BEAST analysis were performed under the best-fitting model
https://www.nature.com/articles/s41467-020-17687-
PANGOLINS USED TO BAN WILDLIFE TRADE AND BATS TO BAN THE PRIVATE FISHING TRADE?
Here we show that a coronavirus, which we name pangolin-CoV, isolated from a Malayan pangolin has 100%, 98.6%, 97.8% and 90.7% amino acid identity with SARS-CoV-2 in the E, M, N and S proteins, respectively. In particular, the receptor-binding domain of the S protein of pangolin-CoV is almost identical to that of SARS-CoV-2, with one difference in a noncritical amino acid. Our comparative genomic analysis suggests that SARS-CoV-2 may have originated in the recombination of a virus similar to pangolin-CoV with one similar to RaTG13. Pangolin-CoV was detected in 17 out of the 25 Malayan pangolins that we analysed. Infected pangolins showed clinical signs and histological changes, and circulating antibodies against pangolin-CoV reacted with the S protein of SARS-CoV-2. This newly identified coronavirus from pangolins—the most-trafficked mammal in the illegal wildlife trade—could represent a future threat to public health if wildlife trade is not effectively controlled. March–August 2019, and analysed them for SARS-related coronaviruses using reverse-transcription polymerase chain reaction (RT–PCR) with primers that target a conservative region of betacoronaviruses. RNA from 17 of the 25 Malayan pangolins generated the expected PCR product, whereas RNA from the Chinese pangolins did not amplify. Plasma samples of four PCR-positive and four PCR-negative Malayan pangolins were used in the detection of IgG and IgM antibodies against SARS-CoV-2 using a double-antigen sandwich enzyme-linked immunosorbent assay (ELISA). One of the PCR-positive sample reacted strongly, showing an optical density at 450 nm (OD450) value of 2.17 (cut-off value = 0.11) (Extended Data Table 2). The plasma remained positive at the dilution of 1:80, which suggests that the pangolin was naturally infected with a virus similar to SARS-CoV-2. The other three PCR-positive pangolins had no detectable antibodies against SARS-CoV-2. They showed typical coronavirus morphology (Fig. 1e). (It looks like a duck, must be a duck). RT–PCR targeting the spike (S) and RdRp genes produced the expected PCR products: these PCR products had approximately 84.5% and 92.2% nucleotide sequence identity, respectively, to the partial S and RdRp genes of SARS-CoV-2. We obtained the completed coronavirus genome (29,825 bp)—which we designated pangolin-CoV—using the assembled contigs, short sequence reads and targeted PCR analysis. The predicted S, E, M and N genes of pangolin-CoV are 3,798, 228, 669 and 1,260 bp, respectively, in length and the proteins they encode share 90.7%, 100%, 98.6% and 97.8% amino acid identity to the equivalent proteins of SARS-CoV-2 (Table 1). In a Simplot analysis of whole-genome sequences, we found that pangolin-CoV was highly similar to SARS-CoV-2 and RaTG13, with sequence identity between 80 and 98% (except for the S gene) (Fig. 2). In particular, the receptor-binding domain (RBD) of the S protein of pangolin-CoV has only one amino acid difference with SARS-CoV-2. The phylogenetic tree was constructed using RAxML with the substitution model GTRGAMMAI and 1,000 bootstrap replicates. Numbers (>70) above or below branches are percentage bootstrap values for the associated nodes. The scale bar represents the number of substitutions per site. As the S proteins of both SARS-CoV and SARS-CoV-2 have previously been shown to specifically recognize angiotensin-converting enzyme 2 (ACE2) during the entry of host cells2,8, we conducted molecular binding simulations of the interaction of the S proteins of the four closely related SARS-related coronaviruses with ACE2 proteins from humans, civets and pangolins. No animals thus far have been implicated as carriers of the virus. In addition, a bat coronavirus (RaTG13) has about 96% sequence identity to SARS-CoV-2 at the whole-genome level2. Therefore, it is reasonable to assume that bats are the native host of SARS-CoV-2, as has previously been suggested for SARS-CoV and MERS-CoV12,13. The SARS-related coronavirus identified in the present study and the metagenomic assemblies of viral sequences from Malayan pangolins14 is genetically related to SARS-CoV-2, but is unlikely to be directly linked to the current outbreak because of its substantial sequence differences from SARS-CoV-2. However, a virus related to pangolin-CoV appears to have donated the RBD to SARS-CoV-2. SARS-related coronavirus sequences have previously been detected in dead Malayan pangolins15. These sequences appear to be from the same virus (pangolin-CoV) that we identified in the present study, as judged from their sequence similarity. Although a SARS-CoV-2-like coronavirus was detected in the lungs of these pangolins, a direct association between the clinical signs or pathology and active virus replication is not available as we lack evidence from immunohistochemistry or in situ hybridization experiments. The experimental infection of healthy pangolins with pangolin-CoV would provide more definitive answers; however, as pangolins are protected it is difficult to carry out such experiments. Further studies are needed to confirm the role of pangolins in the transmission of SARS-related coronaviruses. Our findings support the call for stronger enforcement of regulations against the illegal trade in pangolins. Owing to the demand for their meat as a delicacy and their scales for use in traditional medicine in China, the illegal smuggling of pangolins from Southeast Asia to China is widespread18. International co-operation in the implementation of stricter regulations against illegal wildlife trade and consumption of game meat should be encouraged, as this will increase the protection of endangered animals and help to prevent future outbreaks of diseases caused by SARS-related coronaviruses.
https://www.nature.com/articles/s41586-020-2313-x
The Origins of COVID-19 "How do you investigate something that doesn't exist?" (NurembergTrials.net)