Mapping the evolutionary trajectory of SARS-CoV-2 inside the host

In a recent study published on bioRxiv* preprint server, researchers in the United States have mapped the trajectory of intra-host course of acute severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection.

Study: Intra-host evolutionary dynamics and tissue compartmentalization during acute SARS-CoV-2 infection. Image Credit: ktsdesign/Shutterstock

Background

Large-scale global whole genome sequencing efforts and phylogenetic analyzes of clinical samples during the coronavirus disease 2019 (COVID-19) pandemic have captured the global evolutionary dynamics of SARS-CoV-2. However, there is a lack of understanding of the evolutionary dynamics of SARS-CoV-2 inside the host.

Although a few studies have already captured the intra-host SARS-CoV-2 dynamics, but have focused only on immunocompetent hosts. These studies showed low within-host diversity, with most samples containing 15 or fewer within-host single nucleotide variants (iSNVs). Together, the data from these studies demonstrated that selection-driven emergence of high-frequency iSNVs during acute infection is likely rare. Overall, a high-resolution profile of the within-host evolutionary dynamics of SARS-CoV-2 is lacking.

Moreover, it is poorly understood how pre-existing immunity, obtained by vaccination or prior infection, influences the intra-host evolution of SARS-CoV-2. More importantly, there is a need to characterize the potential for emergence of immune-evading variants in immunocompetent individuals with different vaccination status.

About the study

In the present study, researchers recruited 32 students, faculty, and staff from the University of Illinois in the United States (USA) for longitudinal sampling that resulted in detection of the SARS-CoV-variant. 2 with high confidence to uncover evolutionary dynamics overlooked by less-frequent sampling strategies. Of these 32 study participants, 20 were treatment-naive individuals and 12 had pre-existing immunity acquired through vaccination or natural infection with SARS-CoV-2. From each individual, the team collected daily nasal swabs and saliva from the middle turbinate (MT) of naïve and immune individuals for repeated measurements of iSNV frequencies during the early phase of infection. In this way, they generated high-resolution profiles of iSNV dynamics between tissue compartments and over time.

Study results

Indications of strong positive selection were rare in the study cohort. However, the researchers noted a number of non-synonymous substitutions, including N:P67S, S:Q677H, and ORF1ab:P5402H from the lower detection limit to the high frequency. The substitution at S:Q677 has arisen independently in several SARS-CoV-2 sublines worldwide, showing that mutations at this site may have an evolutionary advantage. The S:Q677H frequency was 56.5% when the associated study participant had a detectable infectious disease. viral load in a nasal swab, indicating the potential for forward transmission of this iSNV.

Furthermore, the authors observed competition between S:Q677H and S:P681H substitutions within the same individual, with S:Q677H appearing at a high frequency for a period of time on a day when the initially fixed frequency of S:P681H decreased. . However, the observed reversion to an S:P681H genotype only after day 7 showed that the selective advantage conferred by S:P681H was greater than that of S:Q677H. The widespread proliferation of SARS-CoV-2 lines containing S:P681H compared to S:Q677H further supports the fitness advantage conferred by this mutation.

Moreover, genome mapping revealed the accumulation of multiple non-synonymous mutation hotspots differing between naïve and immune individuals. The observed enrichment of amino acid substitutions immediately upstream of the SARS-CoV-2 tip subunit 1 (S1)/S2 cleavage site indicated that this region may be subject to intra-host selection. stronger in humans. Therefore, in naïve individuals, they identified hotspots at residues 402-457 in ORF1ab and 655-681 in S, directly adjacent to the S1/S2 cleavage site. S1/S2 cleavage site substitutions are features of the Omicron, Delta, and Alpha SARS-CoV-2 lineages. A recent study by Y. Liu et al. demonstrated that substitutions at the S1/S2 cleavage site were responsible for the increased relative fitness of the Delta variant compared to Alpha.

A high density of nucleocapsid (N) gene substitutions in immune study participants further validated previous data suggesting the importance of the N gene during human adaptation. As a result, the researchers observed a hotspot of mutation accumulation in N:199-204.

Comparison of intra-host single nucleotide variant (iSNV) diversity between samples and individuals. (A) Total number of iSNVs for each sample from each unvaccinated participant. The light gray boxes indicate the total number of iSNVs for all samples and the horizontal black lines indicate the number of shared iSNVs for each participant. (B) iSNV counts for immune participants. (C) iSNV counts for individual samples with Ct <25 from naïve participants as a function of the number of days after enrollment (squared-adjusted R = 0.05007, p = 0.02255). The line represents linear regression. (D) iSNV counts for individual samples with Ct <25 from immune participants as a function of the number of days after enrollment (squared adjusted R = 0.2857, p = 0.006359). The line represents linear regression.

The researchers also observed several shared mutations in the untranslated regions of the SARS-CoV-2 genome, such as the three major untranslated regions (3′ UTRs). The most frequent was a t29760c substitution in the 3′ UTR, shared by nine naïve individuals. Future studies should determine whether the recurrent UTR mutations observed in the current study affect the fitness of intra-host SARS-CoV-2.

Additionally, several study participants exhibited extreme fluctuations at or near iSNVs. They abruptly fell below the detection limit and returned to high frequencies a few days later. This was likely due to spatial patterning, as described for influenza virus by Amato et al. 2021. Spatial patterning promotes drift-induced fluctuations in sampled iSNVs due to bottleneck effects potentially resulting from poor quality sampling of the viral population. This finding further underscores the advantages of longitudinal sampling.

Finally, researchers observed significant tissue compartmentalization between the oral and nasal environments throughout SARS-CoV-2 infection in some study participants. This explains why sampling a single tissue site may not provide a full picture of SARS-CoV-2 diversity within a host.

conclusion

Data from the study provided a high-resolution profile of the evolutionary dynamics of intra-host SARS-CoV-2. The results of the study showed that the evolution of SARS-CoV-2 within the host in naïve and immune individuals appeared to be mainly driven by stochastic forces during acute infections. In addition, the researchers identify mutational hotspots in the SARS-CoV-2 genome, consistent with the selection pressure that favors the emergence of iSNVs capable of onward transmission.

Additionally, they also detected significant tissue compartmentalization of SARS-CoV-2 between nasal swabs and saliva samples in many individuals. Furthermore, the recurrent detection of successful and less successful iSNVs globally indicated areas of alliance and mismatch between intra-host and inter-host selective pressures. These data shed light on the forces shaping global patterns of SARS-CoV-2 evolution.

*Important Notice

bioRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be considered conclusive, guide clinical practice/health-related behaviors, or treated as established information.