Both the icSARS-CoV and the MA-SARS-CoV have become the most widely used recombinant viruses to study SARS-like viruses and no specific sequences were found in the human SARS-CoV-2Wu. The identity of MA-SARS-CoV compared with the original SARS-CoV-Urbani is 99.97% with only six distinct nucleotides, that cannot be used as markers of this recombinant virus since same mutations are naturally acquired by the WT-SARS-CoV, as demonstrated from the sequences of other isolated SARS-CoV 14. This is expected since the first part of these two genomes show high similarities.Īnother recombinant SARS-CoV was produced in 2007 which derived from fifteen passages of the SARS-CoV-Urbani in BALB/c mouse lungs, therefore it was named Mouse-Adapted (MA)-SARS-CoV 14. All the genomes did not contain these sites (Table S1), in particular, the hSARS-CoV-19-Wuhan and hCoV-19-Italy-Vr.īy analysing the sequences of Bat-CoV-raTG13 and the hCoV-19-Pangolin, we observed that only one sequence from SARS-CoV-Urbani (GCCAGCGTGGT) was found in SARS-CoV-2Wu (Wuhan). We analysed natural sequences isolated from four different SARS-CoV (hCoV-19-Italy-Vr/hSARS-CoV-19-Wuhan/hCoV-19 Pangolin/Bat-Cov-raTG13) to look for Bgl1 RS ‘marker’ (GCCNNNN/NGGC). The newly sequences introduced in the recombinant cDNA of SARS-CoV can be used as markers to follow possible virus laboratory spillage. ![]() Also, in this case, the recombinant virus shows several nucleotide mutations which exclude the manipulations performed using modified primers and unique restriction sites. ( D) A specific area of the alignment performed between the mutant SARS-CoV-Urbani MA15 containing the SHC014 spike with the hCoV-19-Italy-VR and the SARS-CoV-19 Wuhan. Despite some small regions are conserved the chimeric spikes show single bp mutation (substitution, deletion, insertions) which support natural evolutions instead of man-made manipulation. ( C) Multiple sequence alignment performed with ClustalW and visualised with JalView show the poor similarities in the RBD between chimeric Spikes generated in the laboratory (line 2 and 4) compared with other SARS-CoV sequences. In the hCoV-19-Italy-VR sequence most of these markers’ sites are not present, while are similar to the wilt type virus HKU3 and RP3. ( B) This alignment shows in violet specific markers used to build a recombinant spike between the Bat-SCoVs genomes HKU3 and RP3. The red boxes show the different nucleotides present in the wilt type SARS-CoV Urbani. The violet box highlight BglI RSs used to build the recombinant icSARS-CoV. ( A) Alignment between the WT SARS-CoV Urbani and the icSARS-CoV. Although other several coronavirus experts have discredited the hypothesis of a man-made coronavirus 8, 9, 10, 11, 12, here we aim to present a different method based on the analysis of restriction site (RS) sequences in the genome of SARS-Cov-2 to reconstruct its origin and follow the new variants.Īnalyses of several RSs sequences in natural and recombinant viruses. In March 2020, Anderson and colleagues published a detailed analysis showing that SARS-CoV-2 does not derive from a laboratory construct 7. Therefore, one of the major discussions around SARS-CoV-2 has been related to its origin with the assumption that SARS-CoV-2 could have been the result of genetic manipulations or spill-over from laboratories studying these viruses. ![]() The World Health Organization declared a coronavirus disease 2019 (COVID-19) pandemic in March 2020. Phylogenetic analysis demonstrated the highly similarity between human SARS-CoV-2 and the sequence isolated from the Bat-Cov-raTG13 4 (97.2% identity) and the Pangolin-SARS-CoV 5 (80% identity), particularly in the receptor-binding-domain (RBD) of the S protein, important to mediate binding to human-receptor-angiotensin-converting-enzyme-2 (hACE2) 6. Although SARS-CoV-2 belongs to the same lineage of CoVs that causes SARS, it is genetically different and it cluster apart exploiting phylogenetic trees 3. In December 2019, a new coronavirus (SARS-CoV-2) started to cause viral pneumonia bringing to severe and fatal infection. They have been associated with two major disease outbreaks, the severe acute respiratory syndrome (SARS-CoV, 2002) and the Middle East respiratory syndrome (MERS-CoV, 2012) 2. ![]() Coronaviruses (CoVs) goes into the family Coronaviridae causing symptoms primarily in the upper respiratory tracts which range from common cold to severe to fatal illnesses 1.
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