Antagonistic pleiotropy plays an important role in governing the evolution and genetic diversity of SARS-CoV-2 (preprint)

Analyses of the genomic diversity of SARS-CoV-2 found that some sites across the genome appear to have mutated independently multiple times with frequency significantly higher than four-fold sites, which can be either due to mutational bias, i.e., elevated mutation rate in some sites of the genome, or selection of the variants due to antagonistic pleiotropy, a condition where mutations increase some components of fitness at a cost to others. To examine how different forces shaped evolution of SARS-CoV-2 in 2020-2021, we analyzed a large set of genome sequences (~ 2 million). Here we show that while evolution of SARS-CoV-2 during the pandemic was largely mutation-driven, a group of nonsynonymous changes is probably maintained by antagonistic pleiotropy. To test this hypothesis, we studied the function of one such mutation, spike M1237I. Spike I1237 increases viral assembly and secretion, but decreases efficiency of transmission in vitro. Therefore, while the frequency of spike M1237I may increase within hosts, viruses carrying this mutation would be outcompeted at the population level. We also discuss how the antagonistic pleiotropy might facilitate positive epistasis to promote virus adaptation and reconcile discordant estimates of SARS-CoV-2 transmission bottleneck sizes in previous studies.

Spatial and temporal origin of the third SARS-CoV-2 Outbreak in Taiwan (preprint)

Since the first report of SARS-CoV-2 in December 2019, Taiwan had gone through three local outbreaks. Unlike the first two, the spatial and temporal origin of the third outbreak (April 20 to November 5, 2021) is still unclear. We sequenced and reconstructed the phylogeny of SARS-CoV-2 genomes and find that the third outbreak was caused by a single virus lineage (T-III), which carries four genetic fingerprints, including spike M1237I (S-M1237I), and three silent changes. The T-III is closest to sequences derived from Turkey on February 8, 2021. The estimated date of divergence from the most recent common ancestor (TMRCA) of T-III is March 23, 2021 (95% HPD February 24 - April 13, 2021), almost one month before the first three confirmed cases on April 20, 2021. The effective population size of the T-III showed approximately 20-fold increase after the onset of the outbreak and reached a plateau in early June. Consequently, the lineage leading to the third outbreak most likely originated from Europe, perhaps Turkey, in February 2021. In addition, the T-III could have circulated in Taiwan in mid-March 2021. The virus was unnoticed while spreading within the community.

Signatures of adaptive evolution during human to mink SARS CoV2 cross species transmission inform estimates of the COVID19 pandemic timing (preprint)

One of the unique features of SARS-CoV-2 is that it mainly evolved neutrally or under purifying selection during the early pandemic. This contrasts with the preceding epidemics of the closely related SARS-CoV and MERS-CoV, both of which evolved adaptively. It is possible that the SARS-CoV-2 exhibits a unique or adaptive feature which deviates from other coronaviruses. Alternatively, the virus may have been cryptically circulating in humans for a sufficient time to have acquired adaptive changes for efficient transmission before the onset of the current pandemic. In order to test the above scenarios, we analyzed the SARS-CoV-2 sequences from minks (Neovision vision) and parenteral human strains. In the early phase of the mink epidemic (April to May 2020), nonsynonymous to synonymous mutation ratios per site within the spike protein was 2.93, indicating a selection process favoring adaptive amino acid changes. In addition, mutations within this protein concentrated within its receptor binding domain and receptor binding motif. Positive selection also left a trace on linked neutral variation. An excess of high frequency derived variants produced by genetic hitchhiking was found during middle (June to July 2020) and early late (August to September 2020) phases of the mink epidemic, but quickly diminished in October and November 2020. Strong positive selection found in SARS-CoV-2 from minks implies that the virus may be not unique in super-adapting to a wide range of new hosts. The mink study suggests that SARS-CoV-2 already went through adaptive evolution in humans, and likely been circulating in humans at least six months before the first case found in Wuhan, China. We also discuss circumstances under which the virus can be well-adapted to its host but fail to induce an outbreak.

One viral sequence for each host? - The neglected within-host diversity as the main stage of SARS-CoV-2 evolution (preprint)

The standard practice of presenting one viral sequence for each infected individual implicitly assumes low within-host genetic diversity. It places the emphasis on the viral evolution between, rather than within, hosts. To determine this diversity, we collect SARS-CoV-2 samples from the same patient multiple times. Our own data in conjunction with previous reports show that two viral samples collected from the same individual are often very different due to the substantial within-host diversity. Each sample captures only a small part of the total diversity that is transiently and locally released from infected cells. Hence, the global SARS-CoV-2 population is a meta-population consisting of the viruses in all the infected hosts, each of which harboring a genetically diverse sub-population. Advantageous mutations must be present first as the within-host diversity before they are revealed as between-host polymorphism. The early detection of such diversity in multiple hosts could be an alarm for potentially dangerous mutations. In conclusion, the main forces of viral evolution, i.e., mutation, drift, recombination and selection, all operate within hosts and should be studied accordingly. Several significant implications are discussed.

D614G Substitution of SARS-CoV-2 Spike Protein Increases Syncytium Formation and Viral Transmission via Enhanced Furin-mediated Spike Cleavage (preprint)

ABSTRACT Since the D614G substitution in the spike (S) of SARS-CoV-2 emerged, the variant strain underwent rapid expansion to become the most abundant strain worldwide. Therefore, this substitution may provide an advantage of viral spreading. To explore the mechanism, we analyzed 18 viral isolates containing S proteins with either G614 or D614. Both the virus titer and syncytial phenotype were significantly increased in S-G614 than in S-D614 isolates. We further showed increased cleavage of S at the furin substrate site, a key event that promotes syncytium, in S-G614 isolates. These functions of the D614G substitution were validated in cells expressing S protein. The effect on syncytium was abolished by furin inhibitor treatment and mutation of the furin-cleavage site, suggesting its dependence on cleavage by furin. Our study provides a mechanistic explanation for the increased transmissibility of S-G614 containing SARS-CoV-2 through enhanced furin-mediated S cleavage, which increases membrane fusion and virus infectivity.

The origin and underlying driving forces of the SARS-CoV-2 outbreak (preprint)

The spread of SARS-CoV-2 since December 2019 has become a pandemic and impacted many aspects of human society. Here, we analyzed genetic variation of SARS-CoV-2 and its related coronavirus and found the evidence of intergenomic recombination. After correction for mutational bias, analysis of 137 SARS-CoV-2 genomes as of 2/23/2020 revealed the excess of low frequency mutations on both synonymous and nonsynonymous sites which is consistent with recent origin of the virus. In contrast to adaptive evolution previously reported for SARS-CoV in its brief epidemic in 2003, our analysis of SARS-CoV-2 genomes shows signs of relaxation of selection. The sequence similarity of the spike receptor binding domain between SARS-CoV-2 and a sequence from pangolin is probably due to an ancient intergenomic introgression. Therefore, SARS-CoV-2 might have cryptically circulated within humans for years before being recently noticed. Data from the early outbreak and hospital archives are needed to trace its evolutionary path and reveal critical steps required for effective spreading. Two mutations, 84S in orf8 protein and 251V in orf3 protein, occurred coincidentally with human intervention. The 84S first appeared on 1/5/2020 and reached a plateau around 1/23/2020, the lockdown of Wuhan. 251V emerged on 1/21/2020 and rapidly increased its frequency. Thus, the roles of these mutations on infectivity need to be elucidated. Genetic diversity of SARS-CoV-2 collected from China was two time higher than those derived from the rest of the world. In addition, in network analysis, haplotypes collected from Wuhan city were at interior and have more mutational connections, both of which are consistent with the observation that the outbreak of cov-19 was originated from China. SUMMARYIn contrast to adaptive evolution previously reported for SARS-CoV in its brief epidemic, our analysis of SARS-CoV-2 genomes shows signs of relaxation of selection. The sequence similarity of the spike receptor binding domain between SARS-CoV-2 and a sequence from pangolin is probably due to an ancient intergenomic introgression. Therefore, SARS-CoV-2 might have cryptically circulated within humans for years before being recently noticed. Data from the early outbreak and hospital archives are needed to trace its evolutionary path and reveal critical steps required for effective spreading. Two mutations, 84S in orf8 protein and 251V in orf3 protein, occurred coincidentally with human intervention. The 84S first appeared on 1/5/2020 and reached a plateau around 1/23/2020, the lockdown of Wuhan. 251V emerged on 1/21/2020 and rapidly increased its frequency. Thus, the roles of these mutations on infectivity need to be elucidated.