Effective vaccines against SARS-CoV-2 are now available, but the ongoing emergence of variants that evade natural and vaccine immunity are a concern. SARS-CoV-2 naturally acquires mutations as it replicates within a population, and the current hypothesis is that variants arise during chronic infections, where selective pressure from antibodies and prolonged infection time give rise to intra-host single-nucleotide variants (iSNVs) that are selected and can be transmitted. However, most SARS-CoV-2 infections are acute, and it is not known if variants also arise during acute infections, or if these intra-host variants can be transmitted to others. The existence of iSNVs derived from acute infections would indicate that variants can arise more frequently, increasing the risk for variants of concern and the need for novel vaccines.
To answer both these questions, Katarina Braun, a graduate student in the Friedrich lab at the University of Wisconsin-Madison, along with Dr. Louise Moncla and colleagues from the Bedford group in the Fred Hutch Vaccine and Infectious Disease Division, recently published a paper in PLoS Pathogens in which they used viral sequencing and bioinformatics to study iSNVs. RNA sequencing methods notoriously are prone to multiple sources of technical error, which can overestimate the single-nucleotide mutations that occur biologically. To overcome this technical artifact, the authors employed rigorous biologic and statistical controls to sequence and align viral genomes isolated from 2020 Wisconsin-area nasal swabs to identify true iSNVs present in SARS-CoV-2 strains. Overall, they found that iSNVs that arise within acutely infected hosts were relatively rare, as most infections contained only 0-6 iSNVs within the approximately 30,000 nucleotide viral genome.
Although an iSNV might sometimes arise during acute infection, the authors next interrogated whether intra-host variants could actually transmit out of the host to seed a new infection. If this were to occur, infections that are phylogenetically linked should share iSNVs. However, this analysis revealed only one instance of an iSNV from a host being detected in a downstream consensus strain. In cases of concurrent infections in multiple people within a household, the presence of shared iSNVs was higher than in randomly sampled infections; however, transmission linkage could not be inferred without additional epidemiologic data. Likewise, the found that intra-host iSNVs were not adaptively beneficial, further negating the possibility that acute infection-derived variants go on to leave the host to infect others.
To further investigate whether iSNVs from acute infection leave the host to infect others, the authors assessed viral transmission bottlenecks. In a narrow transmission bottleneck, where relatively few viral particles found an infection, the likelihood that novel variants are lost is high. Conversely, wide bottlenecks contain many viral particles, increasing the chance that a novel variant enters the new host. The authors found that most acute infections sampled were seeded by fewer than 50 viral particles—a low founder number—suggesting a narrow transmission bottleneck and that iSNVs are not productively transmitted from acutely infected people to others.
This study is one of few that have interrogated intra-host SARS-CoV-2 variants, finding that a low frequency of iSNVs result from acute infection. Importantly, this work also demonstrates that the iSNVs that do arise are largely lost upon transmission. Dr. Moncla, the senior author in this study, commented on this finding: “our data shows that novel variants arising in an infected host and then getting transmitted onward is likely a pretty rare event. What's remarkable to me is that we've seen that variants of concern have arisen on a global scale really rapidly, which I think that is a testament to the sheer number of infections of SARS-CoV-2 that have occurred. Even rare evolutionary events can occur given enough opportunities."
These findings demonstrate that chronic, and not acute, SARS-CoV-2 infections are likely the source of novel variants of concern, and that these intra-host analysis techniques could be employed to study iSNVs in prolonged infections. Going forward, further studies are needed to assess the impact of SARS-CoV-2 immunity on intra-host viral selection, as this information will be important to understand SARS-CoV-2 variants and breakthrough transmission among those with vaccine or natural immunity. “I think that seeing how variants arise and are selected within individuals with vaccine or infection-induced immunity will be a really important follow-up. We don't know yet how immune escape variants will be selected, and understanding that process will be important for future vaccine updates,” said Dr. Moncla.
Braun KM, Moreno GK, Wagner C, Accola MA, Rehrauer WM, Baker DA, Koelle K, I’Connor DH, Bedford T, Friedrich TC, Moncla LH. Acute SARS-CoV-2 infections harbor limited within-host diversity and transmit via tight transmission bottlenecks. PLoS Pathogens. 2021 Aug 23;17(8):e1009849. doi: 10.1371/journal.ppat.1009849. eCollection 2021 Aug.
This work was supported by the National Institute for Allergy and Infectious Disease and the National Library of Medicine.