The clue behind the emergence of SARS-CoV-2 remains a huge debate among the virologist, policymakers, and general public, while the gain-of-function hypothesizes mostly focused on the furin cleavage site and receptor-binding domain (RBD) of the spike (S) protein. Here, we report that the SARS-CoV-2 gained the novel human-specific spike protein S1-N-Terminal Domain (S1-NTD) (Type-I) which is present only in SARS-CoV-2. Interestingly, SARS-CoV-2-rB-CoV showed unique bat-specific Type-II and Type-III-S1-NTD, which is not present in other SARS coronaviruses, including SARS-CoV-2 variants. We also found widespread recombination and selection pressure in the S1-NTDs of the bat viruses. In addition, the Pangolin/GX/2018 and Pangolin/Guangdong/2019 lineages showed close identity (73-79%) with the Type-I-S1-NTD and Type-II-S1-NTD respectively. This justifies that the pangolin is the mixing vessel (intermediate host) to exchange the bat-specific Type-II-S1-NTD in the SARS-CoV-2-rB-CoV into Type-I-like-S1-NTD in pangolin through recombination. Furthermore, the pangolin virus with Type-I-like-S1-NTD jumped into humans which then transformed into SARS-CoV-2 with Type-I-S1-NTD by host selection pressure. Remarkably, we characterized the bat/Cambodia virus as a recombinant SARS-CoV-2/RatG13 with the S1-NTD of bat-SARS-like viruses;while only bat/RatG13 with Type-I-S1-NTD established a huge pandemic outbreak. Additionally, recent SARS-CoV-2 S1-NTD specific neutralizing antibody-based studies support the role of S1-NTD in the post-attachment of the virus;fusion, virus dissemination, and cell-cell fusion thereby prevent the onset of infection;and most of the SARS-CoV-2 variants with increased transmissibility were linked to the S1-NTD mutations. Collectively, our results strongly suggest that the gain of Type-I-S1-NTD in the SARS-CoV-2 is the reason for the pandemic outbreak.
SARS-CoV-2 gained the unique spike protein S1-N-Terminal Domain from the promising pangolins mixing vessel: A novel clue for the pandemic outbreak