SARS-CoV-2 variants with NSP12 P323L/G671S mutations display enhanced virus replication in ferret upper airways and higher transmissibility

With the emergence of multiple predominant SARS-CoV-2 variants, it becomes important to have a comprehensive assessment of their viral fitness and transmissibility. Here, we demonstrate that natural temperature differences between the upper (33 & DEG;C) and lower (37 & DEG;C) respiratory tract have profound effects on SARS-CoV2 replication and transmissibility. Specifically, SARS-CoV-2 variants containing the NSP12 mutations P323L or P323L/G671S exhibit enhanced RNA-dependent RNA polymerase (RdRp) activity at 33 & DEG;C compared with 37 & DEG;C and high transmissibility. Molecular dynamics simulations and microscale thermophoresis demonstrate that the NSP12 P323L and P323L/G671S mutations stabilize the NSP12-NSP7-NSP8 complex through hydrophobic effects, leading to increased viral RdRp activity. Furthermore, competitive transmissibility assay reveals that reverse genetic (RG)-P323L or RG-P323L/G671S NSP12 outcompetes RG-WT (wild-type) NSP12 for replication in the upper respiratory tract, allowing markedly rapid transmissibility. This suggests that NSP12 P323L or P323L/G671S mutation of SARS-CoV-2 is associated with increased RdRp complex stability and enzymatic activity, promoting efficient transmissibility.

Automated summary

This study demonstrates that natural temperature differences between upper and lower respiratory tracts impact the replication and transmission of SARS-CoV-2. Mutations P323L or P323L/G671S enhance the activity of RNA-dependent RNA polymerase (RdRp) at lower temperatures and increase transmissibility. The stability of the NSP12-NSP7-NSP8 complex is increased by these mutations, leading to efficient viral replication and transmission.