The emergence of divergent SARS-CoV-2 lineages has raised concern that novel variants eliciting immune escape or the ability to displace circulating lineages could emerge within individual hosts. Though growing evidence suggests that novel variants arise during prolonged infections, most infections are acute. Understanding how efficiently variants emerge and transmit among acutely-infected hosts is therefore critical for predicting the pace of long-term SARS-CoV-2 evolution. To characterize how within-host diversity is generated and propagated, we combine extensive laboratory and bioinformatic controls with metrics of within- and between-host diversity to 133 SARS-CoV-2 genomes from acutely-infected individuals. We find that within-host diversity is low and transmission bottlenecks are narrow, with very few viruses founding most infections. Within-host variants are rarely transmitted, even among individuals within the same household, and are rarely detected along phylogenetically linked infections in the broader community. These findings suggest that most variation generated within-host is lost during transmission.

The rapid spread of SARS-CoV-2 has gravely impacted societies around the world. Outbreaks in different parts of the globe have been shaped by repeated introductions of new viral lineages and subsequent local transmission of those lineages. Here, we sequenced 3940 SARS-CoV-2 viral genomes from Washington State to characterize how the spread of SARS-CoV-2 in Washington State (USA) in early 2020 was shaped by differences in timing of mitigation strategies across counties, as well as by repeated introductions of viral lineages into the state. Additionally, we show that the increase in frequency of a potentially more transmissible viral variant (614G) over time can potentially be explained by regional mobility differences and multiple introductions of 614G, but not the other variant (614D) into the state. At an individual level, we observed evidence of higher viral loads in patients infected with the 614G variant. However, using clinical records data, we did not find any evidence that the 614G variant impacts clinical severity or patient outcomes. Overall, this suggests that with regards to D614G, the behavior of individuals has been more important in shaping the course of the pandemic in Washington State than this variant of the virus.

Real‐time epidemiological tracking of variants of concern (VOCs) can help limit the spread of more contagious forms of severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2), such as those containing the N501Y mutation. Typically, genetic sequencing is required to be able to track VOCs in real‐time. However, sequencing can take time and may not be accessible in all laboratories. Genotyping by RT‐ddPCR offers an alternative to rapidly detect VOCs through discrimination of specific alleles such as N501Y, which is associated with increased transmissibility and virulence. Here we describe the first cases of the B.1.1.7 lineage of SARS‐CoV‐2 detected in Washington State by using a combination of reverse‐transcription polymerase chain reaction (RT‐PCR), RT‐ddPCR, and next‐generation sequencing. We initially screened 1035 samples positive for SARS‐CoV‐2 by our CDC‐based laboratory‐developed assay using ThermoFisher's multiplex RT‐PCR COVID‐19 assay over four weeks from late December 2020 to early January 2021. S gene target failures (SGTF) were subsequently assayed by RT‐ddPCR to confirm four mutations within the S gene associated with the B.1.1.7 lineage: a deletion at amino acid (AA) 69‐70 (ACATGT), deletion at AA 145, (TTA), N501Y mutation (TAT), and S982A mutation (GCA). All four targets were detected in two specimens; follow‐up sequencing revealed a total of 9 mutations in the S gene and phylogenetic clustering within the B.1.1.7 lineage. Next, we continued screening samples for SGTF detecting 23 additional B.1.1.7 variants by RT‐ddPCR and confirmed by sequencing. As VOCs become increasingly prevalent, molecular diagnostic tools like RT‐ddPCR can be utilized to quickly, accurately, and sensitively distinguish more contagious lineages of SARS‐CoV‐2.

The rapid spread of SARS-CoV-2 has gravely impacted societies around the world. Outbreaks in different parts of the globe are shaped by repeated introductions of new lineages and subsequent local transmission of those lineages. Here, we sequenced 3940 SARS-CoV-2 viral genomes from Washington State to characterize how the spread of SARS-CoV-2 in Washington State (USA) was shaped by differences in timing of mitigation strategies across counties, as well as by repeated introductions of viral lineages into the state. Additionally, we show that the increase in frequency of a potentially more transmissible viral variant (614G) over time can potentially be explained by regional mobility differences and multiple introductions of 614G, but not the other variant (614D) into the state. At an individual level, we see evidence of higher viral loads in patients infected with the 614G variant. However, using clinical records data, we do not find any evidence that the 614G variant impacts clinical severity or patient outcomes. Overall, this suggests that at least to date, the behavior of individuals has been more important in shaping the course of the pandemic than changes in the virus.

The recent emergence of divergent SARS-CoV-2 lineages has raised concerns about the role of selection within individual hosts in propagating novel variants. Of particular concern are variants associated with immune escape and/or enhanced transmissibility. Though growing evidence suggests that novel variants can arise during prolonged infections, most infections are acute. Understanding the extent to which variants emerge and transmit among acutely infected hosts is therefore critical for predicting the pace at which variants resistant to vaccines or conferring increased transmissibility might emerge in the majority of SARS-CoV-2 infections. To characterize how within-host diversity is generated and propagated, we combine extensive laboratory and bioinformatic controls with metrics of within- and between-host diversity to 133 SARS-CoV-2 genomes from acutely infected individuals. We find that within-host diversity during acute infection is low and transmission bottlenecks are narrow, with very few viruses founding most infections. Within-host variants are rarely transmitted, even among individuals within the same household. Accordingly, we also find that within-host variants are rarely detected along phylogenetically linked infections in the broader community. Together, these findings suggest that efficient selection and transmission of novel SARS-CoV-2 variants is unlikely during typical, acute infection.

Our understanding of archaeal virus diversity and structure is just beginning to emerge. Here we describe a new archaeal virus, tentatively named turreted icosahedral virus (MTIV), that was isolated from an acidic hot spring in Yellowstone National Park, USA. Two strains of the virus were identified and found to replicate in an archaeal host species closely related to Each strain encodes for a 9.8-9.9 kb, linear dsDNA genome with large inverted terminal repeats. Each genome encodes for 21 ORFs. Between the strains the ORFs display high homology, but they are quite distinct from other known viral genes. The 70-nm diameter virion is built upon on a T=28 icosahedral lattice. Both single particle cryo-electron microscopy and cryo-tomography reconstructions reveal an unusual structure that has 42 turret-like projections: 12 from each of the 5-fold axes and 30 hexameric units positioned on icosahedral 2-fold axes. Both the virion structural properties and genome content support MTIV as the founding member of a new family of archaeal viruses. Many archaeal viruses are quite different than viruses infecting bacteria and eukaryotes. Initial characterization of MTIV reveals a virus distinct from other known bacterial, eukaryotic, and archaeal viruses; this finding suggests that viruses infecting Archaea are still an understudied group of viruses. As the first known virus infecting the , MTIV provides a new system for exploring archaeal virology by examining host-virus interactions and the unique features of MTIV structure-function relationships. These studies will likely expand our understanding of virus ecology and evolution.

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Real-time epidemiological tracking of variants of interest can help limit the spread of more contagious forms of SARS-CoV-2, such as those containing the N501Y mutation. Typically, genetic sequencing is required to be able to track variants of interest in real-time. However, sequencing can take time and may not be accessible in all laboratories. Genotyping by RT-ddPCR offers an alternative to sequencing to rapidly detect variants of concern through discrimination of specific mutations such as N501Y that is associated with increased transmissibility. Here we describe the first cases of the B.1.1.7 lineage of SARS-CoV-2 detected in Washington State by using a combination of RT-PCR, RT-ddPCR, and next-generation sequencing. We screened 1,035 samples positive for SARS-CoV-2 by our CDC-based laboratory developed assay using ThermoFisher’s multiplex RT-PCR COVID-19 assay over four weeks from late December 2020 to early January 2021. S gene dropout candidates were subsequently assayed by RT-ddPCR to confirm four mutations within the S gene associated with the B.1.1.7 lineage: a deletion at amino acid (AA) 69-70 (ACATGT), deletion at AA 145, (TTA), N501Y mutation (TAT), and S982A mutation (GCA). All four targets were detected in two specimens, and follow-up sequencing revealed a total of 10 mutations in the S gene and phylogenetic clustering within the B.1.1.7 lineage. As variants of concern become increasingly prevalent, molecular diagnostic tools like RT-ddPCR can be utilized to quickly, accurately, and sensitively distinguish more contagious lineages of SARS-CoV-2.