A new coronavirus found in bat samples obtained last year in Yunnan’s Yula County, with 97.2% consistent with the new coronavirus (SARS-CoV-2) in the longest encoded gene region, 1ab, according to a new study. At the same time, the team in this new coronavirus S protein (protogy glycoprotein) S1 and S2 junction also found the insertion of three residual PAA, they concluded that SARS-CoV-2 at the junction of S1 and S2 four amino acid insertion is not an “artificial trace”, a strong proof that it originated in nature.
On May 10, local time, Current Biology, a sub-publication of Cell, published online the “A novel beta beta corona virus ly to SARS-CoV-2 natural sat s1/S2 core of the sea sciences research institute” from Shandong First Medical University, Shandong University of Higher Education, The Institute of The Phoebe Of infectious diseases, the Xishuangbanna Tropical Botanical Garden of the Chinese Academy of Sciences, the Chinese Academy of Sciences,
In the paper, the team reported a new bat-sourced coronavirus, called RmYN02, identified in genomic analysis of 227 bats collected from Yunnan Province, China, between May and October 2019.
Studies show that The homologous nature of RmYN02 and the new coronavirus SARS-CoV-2 in the whole viral genome was 93.3% and 97.2% of the homologous in the 1ab gene closest to SARS-CoV-2. In contrast, the RmYN02 receptor binding domain (RBD) and SARS-CoV-2 exhibit low sequence homologousness (61.3%) and may not bind to angiotensin-converting enzyme 2 (ACE2).
However, similar to SARS-CoV-2, RmYN02 is characterized by multiple amino acid insertions at shear sites in S1 and S2 subunits of the S protein. The team believes this is strong evidence that such insertion events can occur in nature.
These data suggest that SARS-CoV-2 stems from multiple natural recombinations of viruses present in bats and other wild animals.
The authors of this paper are Shi Weifeng, director of the Institute of Pathogen Biology of the School of Basic Medicine of Shandong First Medical University, Director of the Laboratory of Epidemiology of The Cause seology of new infectious diseases in Shandong University, Bi Yuhai, research group of the Institute of Microbiology of the Chinese Academy of Sciences, and Bi Yuhai, project researcher, and Alice Alice Hughes, director of the Landscape Ecology Group of the Xishuangbanna Tropical Botanical Garden Integrated Conservation Center of the Chinese Academy of Sciences. The study was published on March 5, local time, on the preprinted website bioRxiv.
The origin of the new coronavirus is not yet clear, there are still a large number of coronaviruses in wild animals
SARS-CoV-2 has caused an unprecedented pneumonia epidemic in China and elsewhere, which has caused global public health concern. Although bats are considered the most likely natural host of SARS-CoV-2, the origin of the virus remains unclear.
System development analysis shows that SARS-CoV-2 is a new type of coronavirus different from SARS-CoV and MERS-CoV. By far the most closely related virus to SARS-CoV-2 is RaTG13, which was isolated in a chinese chrysanthemum manta ray specimen collected in Yunnan in 2013 by the team of Shi Zhengli of the Wuhan Virus Institute of the Chinese Academy of Sciences. RaTG13 virus strains and neo-coronaviruses have 96.1% nucleotide homomorphism and 92.9% S gene homologousness. These data once again show that bats are important hosts of coronaviruses.
However, it is worth noting that the University of Hong Kong School of Public Health, the National Key Laboratory of Emerging Infectious Diseases Professor Guan Qi, Guangxi Medical University Professor Hu Yanling team, as well as South China Agricultural University, Lingnan Modern Agricultural Science and Technology Guangdong Province Laboratory Professor Shen Yongyi, Professor Xiao Lihua team of two research groups have previously reported the presence of the Malay pangolin SARS-CoV-2-related coronavirus, these pangolins through illegal smuggling into Guangxi and Guangdong.
The team noted that although the coronaviruses detected in these pangolins are far more distant from SARS-CoV-2 than RaTG13 and SARS-CoV-2, they are very similar to SARS-CoV-2 on the receptor binding domain (RBD) of the S protein.
Therefore, although it is not clear whether pangolins are intermediate hosts of SARS-CoV-2 transmission to human processes, they may play an important role in the ecology and evolution of coronaviruses.
They believe that these viruses found in pangolins suggest that there are still large samples of coronaviruses in wild animals, some of which may have been directly involved in the emergence of SARS-CoV-2.
Comparison of SARS-CoV-2 and several representative bat source coronavirus sequences.
A new bat source coronavirus, named RmYN02
Between May and October 2019, the team collected 302 samples from 227 bats in Lula County, Yunnan Province, the paper said. These bats belong to 20 different species, and most of the samples were taken from the Malay chrysanthemum manta ray Rhinolophus Malayanus (n?48, 21.1%), the mid-shoe manta ray Hipposiderostus (n?41, 18.1%) and the small brown chrysanthemum-headed manta ray Rhinolophus stheno (n?39, 17.2%). The samples were derived from a variety of tissues, including wing membranes (219), lungs (2), liver (3), and feces (78).
All but three bats were sampled and released while alive.
Using a new generation of macro genome sequencing technology, the team first locked in two initial consistent sequences. Samples from these sequences were derived from 11 Malay chrysanthemum manta rays from 6 May to 30 July 2019. After a series of validation steps, the team obtained a partial (23395bp) and a complete (29671bp) sequence of bat coronavirus genomes named BetaCoV/Rm/Yunnan/YUNNAN/YN01/2019 (RmYN01) and BetaCoV/Rm/Yunnan/YN02/2019 (RmYN002).
In contrast, RmYN02 was closely related to SARS-CoV-2, showing 93.3% nucleotide sequence consistency, but at the full-length genomic level, RaTG13 and SARS-CoV-2 were more consistent (96.1%). RmYN02 and SARS-CoV-2 are very similar in most genomic regions (e.g. 1ab, 3a, E, 6, 7a, N and 10) (?gt;96% sequence consistency). In particular, RmYN02 was consistent with SARS-CoV-2 at 97.2% in the longest encoded gene region of 1ab (n-21285).
However, the sequence consistency of RmYN02 and SARS-CoV-2 in the S gene (71.8% nucleotides, 72.9% amino acids) was well below the 97.4% between RaTG13 and SARS-CoV-2. It is also worth noting that the amino acid homologousness of RmYN02 and SARS-CoV-2 in RBD was only 62.4%. The amino acid homologousity of pangolin coronavirus and SARS-CoV-2 in RBD from Guangdong is 97.4%, which is also the closest to SARS-CoV-2 in the RBD region.
The results of three-dimensional structural analysis of the homologous model, in vitro experiments and S proteins around SARS-CoV-2 show that ACE2 can also be used as cell receptors, as can SARS-CoV-2. The team also analyzed the RBD of RmYN02, RaTG13 and two pangolin CoVs using homologous models.
The homologous modeling and structural comparison of RmYN02 and the representative coronavirus RBD structure.
It was found that amino acid deficiency in RmYN02 RBD formed two rings shorter than SARS-CoV-2 RBD near the receptor binding site. Importantly, the conservative secondary sulfur key in the external domain (external sub-sub-domain) of SARS-CoV, SARS-CoV-2, RaTG13, Pangolin/MP789/2019, Pangolin/GX/P5L/2017 is missing in RmYN02. The team speculated that these deficiencies could lead to structural changes, thereby reducing the combination of RmYN02 RBD and ACE2, and even lead to non-binding.
Of course, it is possible that the circulatory SARS-related coronaviruses, including RmYN02, ZXC21 and ZC45, use a receptor that we currently do not know about.
It is worth mentioning that previous studies have concluded that the six amino acid residues (L455, F486, Q493, S494, N501 and Y505) in RBD are the main determinants of the binding of SARS-CoV-2 to ACE2 effective receptors. Consistent with homologous modeling, pangolin/MP789/2019 has the same amino acid residue as SARS-CoV-2 in all six locations. In contrast, RaTG13, RmYN02 and RmYN01 and SARS-CoV-2 all had the same amino acid residue in only one location.
The team believes that this evolutionary model is a manifestation of a complex combination of recombination and natural selection.
System Development Tree: A full-length genome; B, S gene; C, RBD ;D, RdRp (RNA-dependent RNA polymerase)
The team also conducted systematic developmental analysis of bat coronaviruses in RmYN02, RaTG13, SARS-CoV-2, and pangolins. Consistent with previous studies, pangolin beta-CoVs form two subtypes. However, it is mentioned in the paper that pangolins are natural reservoirs for these viruses, or whether they are obtained independently from bats or other wild animals, which need further verification.
More notably, in most viral genomes, the kinship between RmYN02 and SARS-CoV-2 is recent, although there is still a long branch distance between the two viruses. The S gene tree showed that SARS-CoV-2 was closer to RaTG13 and farther from RmYN02, indicating that the latter had undergone recombination in the S gene. SARS-CoV-2 and pangolin-CoV/GD are most closely related to the system development tree of RBD, and are far away from bat viruses, again indicating recombination. Finally, systematic developmental analysis of the complete RNA-dependent RNA polymerase (RdRp) gene (often used in the systematic developmental analysis of RNA viruses) shows that RmYN02, RaTG13, and SARS-CoV-2 form a distinct subgroup of pangolin viruses.
SARS-CoV-2 is a natural origin and may be obtained through recombination
Similar to the avian influenza virus (AIVs) hemagglutinin (HA) protein, the S protein of the coronavirus is functionally split into two sub-subsequial S1 and S2. The insertion of multi-base amino acids on shear sites in some AIV subtypes is thought to be associated with increased pathogenicity.
It is worth noting that one of the characteristics of SARS-CoV-2 is the insertion of a four-amino acid at the junction of S1 and S2, which has not been observed in other lineage of the beta-coronavirus. This insertion, known as the Flynn protease shear site, is unique to SARS-CoV-2 and is currently found in all detected SARS-CoV-2 sequences.
The team also found three residual PAAs at the junction of S1 and S2 in RmYN02, which they said was very important. “Although the residual slots (and the resulting nucleotides) inserted by SARS-CoV-2 are different from those in RmYN02, they can be shown to be separate insertion events that strongly indicate that they are naturally occurring in wild animals (bats) and may have been obtained through recombination.” “
Therefore, these data strongly indicate the natural origin of SARS-CoV-2.
In addition, the team again identified RmYN02’s bat host as the Rhinolophus Malayanus, and a sequence obtained in a Malay chrysanthemum manta ray (GenBank accession MK900703) that was 100% consistent.
Malay chrysanthemum bats and Chinese chrysanthemum bats are widely distributed in southwestern China and Southeast Asia. In general, these bats do not migrate long distances, are strong in groups, and are likely to live in the same cave, which may facilitate the exchange and recombination of viruses between them, the paper notes.
Notably, scientists found RaTG13 from swabs and RmYN02 from feces. The feces is therefore a simple but feasible way for bats to spread the virus to other animals, especially those that can take advantage of the cave environment.