Free PMC article: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8996265/Background and objectives: To understand how organisms evolve, it is fundamental to study how
mutations emerge and establish. Here, we estimated the rate of mutation accumulation of SARS-CoV-
2 in vitro and investigated the repeatability of its evolution when facing a new cell type but no immune
or drug pressures.
Methodology: We performed experimental evolution with two strains of SARS-CoV-2, one carrying the
originally described spike protein (CoV-2-D) and another carrying the D614G mutation that has spread
worldwide (CoV-2-G). After 15 passages in Vero cells and whole genome sequencing, we characterized
the spectrum and rate of the emerging mutations and looked for evidences of selection across the
genomes of both strains.
Results: From the frequencies of the mutations accumulated, and excluding the genes with signals of
selection, we estimate a spontaneous mutation rate of 1.3 10 6 6 0.2 10 6 per-base per-infection
cycle (mean across both lineages of SARS-CoV-262SEM). We further show that mutation accumulation
is larger in the CoV-2-D lineage and heterogeneous along the genome, consistent with the action
of positive selection on the spike protein, which accumulated five times more mutations than the corresponding genomic average. We also observe the emergence of mutators in the CoV-2-G background, likely linked to mutations in
the RNA-dependent RNA polymerase and/or in the error-correcting exonuclease protein.
Conclusions and implications: These results provide valuable information on how spontaneous mutations emerge in SARS-CoV-2 and
on how selection can shape its genome toward adaptation to new environments.
Lay Summary: Each time a virus replicates inside a cell, errors (mutations) occur. Here, via laboratory propagation in cells originally
isolated from the kidney epithelium of African green monkeys, we estimated the rate at which the SARS-CoV-2 virus mutates—an important
parameter for understanding how it can evolve within and across humans. We also confirm the potential of its Spike protein to
adapt to a new environment and report the emergence of mutators—viral populations where mutations occur at a significantly faster
rate.M.A. was supported by ‘Fundação para a Ciência e Tecnologia’ (FCT), fellowships PD/BD/138735/2018. Research was supported by FCT Project PTDC/BIA-EVL/31528/2017 to I.G. and by funds from Portuguese NIH.info:eu-repo/semantics/publishedVersio