Investigating the evolutionary dynamics and mutational pattern of SARS-CoV-2 spike gene on selected SARS-CoV-2 variants

The continuous evolution of SARS-CoV-2 has led to the emergence of several vari- ants representing significant challenges for public health. Many studies highlight the relevance of phylogenetic inference or mutational pattern analysis to understand the evolutionary relatedness of viral variants and to estimate the potential effect of new mutations on viral transmission, virulence and antigenicity. Here we describe an evo- lutionary investigation approach combined with mutational analyses of SARS-CoV-2 Spike gene to annotate and potentially track important amino acid site variation of specific functional domain relevant for viral survival. This approach was applied on XBB*, EG* and BA* and their sub-lineages (see materials and methods) available from GISAID. In addition, we considered the major variants of concern (Alpha, Delta, Omicron) and Wuhan-Hu-1 strain as references. Maximum likelihood phyloge- netic tree was constructed from the complete dataset while selection pressure and mutational analyses were conducted on single variants separately. The obtained phylogenetic tree of Spike amino acid gene sequence showed a clear separation of viral variants as well as their expected appearance order. This result supported the significance of selection pressure analyses outcomes combined with amino acid mutational frequencies where in many cases they showed a linear and parallel trend. This allowed also to hypothesize the potential importance of low-frequency mutations in new potential virus variants. This study constitutes an asset of important insights to be considered in regular monitoring programs. In addition, the analysis framework described here introduces a starting point for further standardization, optimization and application on different data types and in large-scale studies.