Evolution of the SARS-CoV-2 Mutational Spectrum.

SARS-CoV-2 evolves rapidly in part because of its high mutation rate. Here, we examine whether this mutational process itself has changed during viral evolution. To do this, we quantify the relative rates of different types of single-nucleotide mutations at 4-fold degenerate sites in the viral genome across millions of human SARS-CoV-2 sequences. We find clear shifts in the relative rates of several types of mutations during SARS-CoV-2 evolution. The most striking trend is a roughly 2-fold decrease in the relative rate of G→T mutations in Omicron versus early clades, as was recently noted by Ruis et al. (2022. Mutational spectra distinguish SARS-CoV-2 replication niches. bioRxiv, doi:10.1101/2022.09.27.509649). There is also a decrease in the relative rate of C→T mutations in Delta, and other subtle changes in the mutation spectrum along the phylogeny. We speculate that these changes in the mutation spectrum could arise from viral mutations that affect genome replication, packaging, and antagonization of host innate-immune factors, although environmental factors could also play a role. Interestingly, the mutation spectrum of Omicron is more similar than that of earlier SARS-CoV-2 clades to the spectrum that shaped the long-term evolution of sarbecoviruses. Overall, our work shows that the mutation process is itself a dynamic variable during SARS-CoV-2 evolution and suggests that human SARS-CoV-2 may be trending toward a mutation spectrum more similar to that of other animal sarbecoviruses.

Potential impact of seasonal forcing on a SARS-CoV-2 pandemic.

A novel coronavirus (SARS-CoV-2) first detected in Wuhan, China, has spread rapidly since December 2019, causing more than 100,000 confirmed infections and 4000 fatalities (as of 10 March 2020). The outbreak has been declared a pandemic by the WHO on Mar 11, 2020. Here, we explore how seasonal variation in transmissibility could modulate a SARS-CoV-2 pandemic. Data from routine diagnostics show a strong and consistent seasonal variation of the four endemic coronaviruses (229E, HKU1, NL63, OC43) and we parameterise our model for SARS-CoV-2 using these data. The model allows for many subpopulations of different size with variable parameters. Simulations of different scenarios show that plausible parameters result in a small peak in early 2020 in temperate regions of the Northern Hemisphere and a larger peak in winter 2020/2021. Variation in transmission and migration rates can result in substantial variation in prevalence between regions. While the uncertainty in parameters is large, the scenarios we explore show that transient reductions in the incidence rate might be due to a combination of seasonal variation and infection control efforts but do not necessarily mean the epidemic is contained. Seasonal forcing on SARS-CoV-2 should thus be taken into account in the further monitoring of the global transmission. The likely aggregated effect of seasonal variation, infection control measures, and transmission rate variation is a prolonged pandemic wave with lower prevalence at any given time, thereby providing a window of opportunity for better preparation of health care systems.

A pseudovirus system enables deep mutational scanning of the full SARS-CoV-2 spike.

A major challenge in understanding SARS-CoV-2 evolution is interpreting the antigenic and functional effects of emerging mutations in the viral spike protein. Here, we describe a deep mutational scanning platform based on non-replicative pseudotyped lentiviruses that directly quantifies how large numbers of spike mutations impact antibody neutralization and pseudovirus infection. We apply this platform to produce libraries of the Omicron BA.1 and Delta spikes. These libraries each contain ∼7,000 distinct amino acid mutations in the context of up to ∼135,000 unique mutation combinations. We use these libraries to map escape mutations from neutralizing antibodies targeting the receptor-binding domain, N-terminal domain, and S2 subunit of spike. Overall, this work establishes a high-throughput and safe approach to measure how ∼105 combinations of mutations affect antibody neutralization and spike-mediated infection. Notably, the platform described here can be extended to the entry proteins of many other viruses.

Swiss public health measures associated with reduced SARS-CoV-2 transmission using genome data.

Genome sequences from evolving infectious pathogens allow quantification of case introductions and local transmission dynamics. We sequenced 11,357 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genomes from Switzerland in 2020 - the sixth largest effort globally. Using a representative subset of these data, we estimated viral introductions to Switzerland and their persistence over the course of 2020. We contrasted these estimates with simple null models representing the absence of certain public health measures. We show that Switzerland's border closures de-coupled case introductions from incidence in neighboring countries. Under a simple model, we estimate an 86-98% reduction in introductions during Switzerland's strictest border closures. Furthermore, the Swiss 2020 partial lockdown roughly halved the time for sampled introductions to die out. Last, we quantified local transmission dynamics once introductions into Switzerland occurred, using a phylodynamic model. We found that transmission slowed 35-63% upon outbreak detection in summer 2020, but not in fall. This finding may indicate successful contact tracing over summer before overburdening in fall. The study highlights the added value of genome sequencing data for understanding transmission dynamics.

Impact of cross-border-associated cases on the SARS-CoV-2 epidemic in Switzerland during summer 2020 and 2021.

During the summers of 2020 and 2021, the number of confirmed cases of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections in Switzerland remained at relatively low levels, but grew steadily over time. It remains unclear to what extent epidemic growth during these periods was a result of the relaxation of local control measures or increased traveling and subsequent importation of cases. A better understanding of the role of cross-border-associated cases (imports) on the local epidemic dynamics will help to inform future surveillance strategies. We analyzed routine surveillance data of confirmed cases of SARS-CoV-2 in Switzerland from 1 June to 30 September 2020 and 2021. We used a stochastic branching process model that accounts for superspreading of SARS-CoV-2 to simulate epidemic trajectories in absence and in presence of imports during summer 2020 and 2021. The Swiss Federal Office of Public Health reported 22,919 and 145,840 confirmed cases of SARS-CoV-2 from 1 June to 30 September 2020 and 2021, respectively. Among cases with known place of exposure, 27% (3,276 of 12,088) and 25% (1,110 of 4,368) reported an exposure abroad in 2020 and 2021, respectively. Without considering the impact of imported cases, the steady growth of confirmed cases during summer periods would be consistent with a value of Re that is significantly above the critical threshold of 1. In contrast, we estimated Re at 0.84 (95% credible interval, CrI: 0.78-0.90) in 2020 and 0.82 (95% CrI: 0.74-0.90) in 2021 when imported cases were taken into account, indicating that the local Re was below the critical threshold of 1 during summer. In Switzerland, cross-border-associated SARS-CoV-2 cases had a considerable impact on the local transmission dynamics and can explain the steady growth of the epidemic during the summers of 2020 and 2021.

Estimation and worldwide monitoring of the effective reproductive number of SARS-CoV-2.

The effective reproductive number Re is a key indicator of the growth of an epidemic. Since the start of the SARS-CoV-2 pandemic, many methods and online dashboards have sprung up to monitor this number through time. However, these methods are not always thoroughly tested, correctly placed in time, or are overly confident during high incidence periods. Here, we present a method for timely estimation of Re, applied to COVID-19 epidemic data from 170 countries. We thoroughly evaluate the method on simulated data, and present an intuitive web interface for interactive data exploration. We show that, in early 2020, in the majority of countries the estimated Re dropped below 1 only after the introduction of major non-pharmaceutical interventions. For Europe the implementation of non-pharmaceutical interventions was broadly associated with reductions in the estimated Re. Globally though, relaxing non-pharmaceutical interventions had more varied effects on subsequent Re estimates. Our framework is useful to inform governments and the general public on the status of epidemics in their country, and is used as the official source of Re estimates for SARS-CoV-2 in Switzerland. It further allows detailed comparison between countries and in relation to covariates such as implemented public health policies, mobility, behaviour, or weather data.

Over the past two and a half years, countries around the globe have struggled to control the transmission of the SARS-CoV-2 virus within their borders. To manage the situation, it is important to have an accurate picture of how fast the virus is spreading. This can be achieved by calculating the effective reproductive number (Re), which describes how many people, on average, someone with COVID-19 is likely to infect. If the Re is greater than one, the virus is infecting increasingly more people, but if it is smaller than one, the number of cases is declining. Scientists use various strategies to estimate the Re, which each have their own strengths and weaknesses. One of the main difficulties is that infections are typically recorded only when people test positive for COVID-19, are hospitalized with the virus, or die. This means that the data provides a delayed representation of when infections are happening. Furthermore, changes in these records occur later than measures that change the infection dynamics. As a result, researchers need to take these delays into account when estimating Re. Here, Huisman, Scire et al. have developed a new method for estimating the Re based on available data records, statistically taking into account the above-mentioned delays. An online dashboard with daily updates was then created so that policy makers and the population could monitor the values over time. For over two years, Huisman, Scire et al. have been applying their tool and dashboard to COVID-19 data from 170 countries. They found that public health interventions, such as mask requirements and lockdowns, did help reduce the Re in Europe. But the effects were not uniform across the globe, likely because of variations in how restrictions were implemented and followed during the pandemic. In early 2020, the Re only dropped below one after countries put lockdowns or other severe measures in place. The Re values added to the dashboard over the last two years have been used pro-actively to inform public health policies in Switzerland and to monitor the spread of SARS-CoV-2 in South Africa. The team has also recently released programming software based on this method that can be used to track future disease outbreaks, and extended the method to estimate the Re using SARS-CoV-2 levels in wastewater.

Evolution, geographic spreading, and demographic distribution of Enterovirus D68.

Worldwide outbreaks of enterovirus D68 (EV-D68) in 2014 and 2016 have caused serious respiratory and neurological disease. We collected samples from several European countries during the 2018 outbreak and determined 53 near full-length genome ('whole genome') sequences. These sequences were combined with 718 whole genome and 1,987 VP1-gene publicly available sequences. In 2018, circulating strains clustered into multiple subgroups in the B3 and A2 subclades, with different phylogenetic origins. Clusters in subclade B3 emerged from strains circulating primarily in the US and Europe in 2016, though some had deeper roots linking to Asian strains, while clusters in A2 traced back to strains detected in East Asia in 2015-2016. In 2018, all sequences from the USA formed a distinct subgroup, containing only three non-US samples. Alongside the varied origins of seasonal strains, we found that diversification of these variants begins up to 18 months prior to the first diagnostic detection during a EV-D68 season. EV-D68 displays strong signs of continuous antigenic evolution and all 2018 A2 strains had novel patterns in the putative neutralizing epitopes in the BC- and DE-loops. The pattern in the BC-loop of the USA B3 subgroup had not been detected on that continent before. Patients with EV-D68 in subclade A2 were significantly older than patients with a B3 subclade virus. In contrast to other subclades, the age distribution of A2 is distinctly bimodal and was found primarily among children and in the elderly. We hypothesize that EV-D68's rapid evolution of surface proteins, extensive diversity, and high rate of geographic mixing could be explained by substantial reinfection of adults. Better understanding of evolution and immunity across diverse viral pathogens, including EV-D68 and SARS-CoV-2, is critical to pandemic preparedness in the future.

Interactions between seasonal human coronaviruses and implications for the SARS-CoV-2 pandemic: A retrospective study in Stockholm, Sweden, 2009-2020.

OBJECTIVES: The four seasonal coronaviruses 229E, NL63, OC43, and HKU1 are frequent causes of respiratory infections and show annual and seasonal variation. Increased understanding about these patterns could be informative about the epidemiology of SARS-CoV-2. METHODS: Results from PCR diagnostics for the seasonal coronaviruses, and other respiratory viruses, were obtained for 55,190 clinical samples analyzed at the Karolinska University Hospital, Stockholm, Sweden, between 14 September 2009 and 2 April 2020. RESULTS: Seasonal coronaviruses were detected in 2130 samples (3.9 %) and constituted 8.1 % of all virus detections. OC43 was most commonly detected (28.4 % of detections), followed by NL63 (24.0 %), HKU1 (17.6 %), and 229E (15.3 %). The overall fraction of positive samples was similar between seasons, but at species level there were distinct biennial alternating peak seasons for the Alphacoronaviruses, 229E and NL63, and the Betacoronaviruses, OC43 and HKU1, respectively. The Betacoronaviruses peaked earlier in the winter season (Dec-Jan) than the Alphacoronaviruses (Feb-Mar). Coronaviruses were detected across all ages, but diagnostics were more frequently requested for paediatric patients than adults and the elderly. OC43 and 229E incidence was relatively constant across age strata, while that of NL63 and HKU1 decreased with age. CONCLUSIONS: Both the Alphacoronaviruses and Betacoronaviruses showed alternating biennial winter incidence peaks, which suggests some type of immune mediated interaction. Symptomatic reinfections in adults and the elderly appear relatively common. Both findings may be of relevance for the epidemiology of SARS-CoV-2.

Spread of a SARS-CoV-2 variant through Europe in the summer of 2020.

Following its emergence in late 2019, the spread of SARS-CoV-21,2 has been tracked by phylogenetic analysis of viral genome sequences in unprecedented detail3-5. Although the virus spread globally in early 2020 before borders closed, intercontinental travel has since been greatly reduced. However, travel within Europe resumed in the summer of 2020. Here we report on a SARS-CoV-2 variant, 20E (EU1), that was identified in Spain in early summer 2020 and subsequently spread across Europe. We find no evidence that this variant has increased transmissibility, but instead demonstrate how rising incidence in Spain, resumption of travel, and lack of effective screening and containment may explain the variant's success. Despite travel restrictions, we estimate that 20E (EU1) was introduced hundreds of times to European countries by summertime travellers, which is likely to have undermined local efforts to minimize infection with SARS-CoV-2. Our results illustrate how a variant can rapidly become dominant even in the absence of a substantial transmission advantage in favourable epidemiological settings. Genomic surveillance is critical for understanding how travel can affect transmission of SARS-CoV-2, and thus for informing future containment strategies as travel resumes.

Novel SARS-CoV-2 variants: the pandemics within the pandemic.

BACKGROUND: Many new variants of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been termed variants of concern/interest (VOC/I) because of the greater risk they pose due to possible enhanced transmissibility and/or severity, immune escape, diagnostic and/or treatment failure, and reduced vaccine efficacy. AIMS: We sought to review the current knowledge of emerging SARS-CoV-2 variants, particularly those deemed VOC/Is: B.1.351, B.1.1.7, and P.1. SOURCES: MEDLINE and BioRxiv databases, as well as the grey literature, were searched for reports of SARS-CoV-2 variants since November 2020. Relevant articles and their references were screened. CONTENT: Mutations on the spike protein in particular may affect both affinity for the SARS-CoV-2 cell receptor ACEII and antibody binding. These VOC/Is often share similar mutation sets. The N501Y mutation is shared by the three main VOCs: B.1.1.7, first identified in the United Kingdom, P.1, originating from Brazil, and B.1.351, first described in South Africa. This mutation likely increases transmissibility by increasing affinity for ACEII. The B.1.351 and P.1 variants also display the E484K mutation which decreases binding of neutralizing antibodies, leading to partial immune escape; this favours reinfections, and decreases the in vitro efficacy of some antibody therapies or vaccines. Those mutations may also have phenotypical repercussions of greater severity. Furthermore, the accumulation of mutations poses a diagnostic risk (lowered when using multiplex assays), as seen for some assays targeting the S gene. With ongoing surveillance, many new VOC/Is have been identified. The emergence of the E484K mutation independently in different parts of the globe may reflect the adaptation of SARS-CoV-2 to humans against a background of increasing immunity. IMPLICATIONS: These VOC/Is are increasing in frequency globally and pose challenges to any herd immunity approach to managing the pandemic. While vaccination is ongoing, vaccine updates may be prudent. The virus continues to adapt to transmission in humans, and further divergence from the initial Wuhan sequences is expected.

Multiplex qPCR discriminates variants of concern to enhance global surveillance of SARS-CoV-2.

With the emergence of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) variants that may increase transmissibility and/or cause escape from immune responses, there is an urgent need for the targeted surveillance of circulating lineages. It was found that the B.1.1.7 (also 501Y.V1) variant, first detected in the United Kingdom, could be serendipitously detected by the Thermo Fisher TaqPath COVID-19 PCR assay because a key deletion in these viruses, spike Δ69-70, would cause a "spike gene target failure" (SGTF) result. However, a SGTF result is not definitive for B.1.1.7, and this assay cannot detect other variants of concern (VOC) that lack spike Δ69-70, such as B.1.351 (also 501Y.V2), detected in South Africa, and P.1 (also 501Y.V3), recently detected in Brazil. We identified a deletion in the ORF1a gene (ORF1a Δ3675-3677) in all 3 variants, which has not yet been widely detected in other SARS-CoV-2 lineages. Using ORF1a Δ3675-3677 as the primary target and spike Δ69-70 to differentiate, we designed and validated an open-source PCR assay to detect SARS-CoV-2 VOC. Our assay can be rapidly deployed in laboratories around the world to enhance surveillance for the local emergence and spread of B.1.1.7, B.1.351, and P.1.

SARS-CoV-2 N501Y Introductions and Transmissions in Switzerland from Beginning of October 2020 to February 2021-Implementation of Swiss-Wide Diagnostic Screening and Whole Genome Sequencing.

The rapid spread of the SARS-CoV-2 lineages B.1.1.7 (N501Y.V1) throughout the UK, B.1.351 (N501Y.V2) in South Africa, and P.1 (B.1.1.28.1; N501Y.V3) in Brazil has led to the definition of variants of concern (VoCs) and recommendations for lineage specific surveillance. In Switzerland, during the last weeks of December 2020, we established a nationwide screening protocol across multiple laboratories, focusing first on epidemiological and microbiological definitions. In January 2021, we validated and implemented an N501Y-specific PCR to rapidly screen for VoCs, which are then confirmed using amplicon sequencing or whole genome sequencing (WGS). A total of 13,387 VoCs have been identified since the detection of the first Swiss case in October 2020, with 4194 being B.1.1.7, 172 B.1.351, and 7 P.1. The remaining 9014 cases of VoCs have been described without further lineage specification. Overall, all diagnostic centers reported a rapid increase of the percentage of detected VOCs, with a range of 6 to 46% between 25 to 31 of January 2021 increasing towards 41 to 82% between 22 to 28 of February. A total of 739 N501Y positive genomes were analysed and show a broad range of introduction events to Switzerland. In this paper, we describe the nationwide coordination and implementation process across laboratories, public health institutions, and researchers, the first results of our N501Y-specific variant screening, and the phylogenetic analysis of all available WGS data in Switzerland, that together identified the early introduction events and subsequent community spreading of the VoCs.

Emergence and spread of a SARS-CoV-2 variant through Europe in the summer of 2020.

Following its emergence in late 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global pandemic resulting in unprecedented efforts to reduce transmission and develop therapies and vaccines (WHO Emergency Committee, 2020; Zhu et al., 2020). Rapidly generated viral genome sequences have allowed the spread of the virus to be tracked via phylogenetic analysis (Worobey et al., 2020; Hadfield et al., 2018; Pybus et al., 2020). While the virus spread globally in early 2020 before borders closed, intercontinental travel has since been greatly reduced, allowing continent-specific variants to emerge. However, within Europe travel resumed in the summer of 2020, and the impact of this travel on the epidemic is not well understood. Here we report on a novel SARS-CoV-2 variant, 20E (EU1), that emerged in Spain in early summer, and subsequently spread to multiple locations in Europe. We find no evidence of increased transmissibility of this variant, but instead demonstrate how rising incidence in Spain, resumption of travel across Europe, and lack of effective screening and containment may explain the variant's success. Despite travel restrictions and quarantine requirements, we estimate 20E (EU1) was introduced hundreds of times to countries across Europe by summertime travellers, likely undermining local efforts to keep SARS-CoV-2 cases low. Our results demonstrate how a variant can rapidly become dominant even in absence of a substantial transmission advantage in favorable epidemiological settings. Genomic surveillance is critical to understanding how travel can impact SARS-CoV-2 transmission, and thus for informing future containment strategies as travel resumes.

Cryptic transmission of SARS-CoV-2 in Washington state.

After its emergence in Wuhan, China, in late November or early December 2019, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus rapidly spread globally. Genome sequencing of SARS-CoV-2 allows the reconstruction of its transmission history, although this is contingent on sampling. We analyzed 453 SARS-CoV-2 genomes collected between 20 February and 15 March 2020 from infected patients in Washington state in the United States. We find that most SARS-CoV-2 infections sampled during this time derive from a single introduction in late January or early February 2020, which subsequently spread locally before active community surveillance was implemented.

Geographical and temporal distribution of SARS-CoV-2 clades in the WHO European Region, January to June 2020.

We show the distribution of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) genetic clades over time and between countries and outline potential genomic surveillance objectives. We applied three genomic nomenclature systems to all sequence data from the World Health Organization European Region available until 10 July 2020. We highlight the importance of real-time sequencing and data dissemination in a pandemic situation, compare the nomenclatures and lay a foundation for future European genomic surveillance of SARS-CoV-2.

Coast-to-Coast Spread of SARS-CoV-2 during the Early Epidemic in the United States.

The novel coronavirus SARS-CoV-2 was first detected in the Pacific Northwest region of the United States in January 2020, with subsequent COVID-19 outbreaks detected in all 50 states by early March. To uncover the sources of SARS-CoV-2 introductions and patterns of spread within the United States, we sequenced nine viral genomes from early reported COVID-19 patients in Connecticut. Our phylogenetic analysis places the majority of these genomes with viruses sequenced from Washington state. By coupling our genomic data with domestic and international travel patterns, we show that early SARS-CoV-2 transmission in Connecticut was likely driven by domestic introductions. Moreover, the risk of domestic importation to Connecticut exceeded that of international importation by mid-March regardless of our estimated effects of federal travel restrictions. This study provides evidence of widespread sustained transmission of SARS-CoV-2 within the United States and highlights the critical need for local surveillance.