A sarbecovirus found in Russian bats uses human ACE2

ABSTRACTSpillover of sarbecoviruses from animals to humans resulted in outbreaks of severe acute respiratory syndrome SARS-CoVs and the ongoing COVID-19 pandemic. Efforts to identify the origins of SARS-CoV-1 and -2 has resulted in the discovery of numerous animal sarbecoviruses – the majority of which are only distantly related to known human pathogens and do not infect human cells. The receptor binding domain (RBD) on sarbecoviruses engages receptor molecules on the host cell and mediates cell invasion. Here, we tested the receptor tropism for RBDs from two sarbecoviruses found in Russian horseshoe bats to screen cell entry. While these two viruses are in a viral lineage distinct from SARS-CoV-1 and -2, one virus was capable of using human ACE2 to facilitate cell entry. Our findings demonstrate that sarbecoviruses circulating in wildlife outside of Asia also exhibit compatibility with human ACE2 and should be taken into consideration for future universal sarbecovirus vaccine candidates.

Patient health records and whole viral genomes from an early SARS-CoV-2 outbreak in a Quebec hospital reveal features associated with favorable outcomes

The first confirmed case of COVID-19 in Quebec, Canada, occurred at Verdun Hospital on February 25, 2020. A month later, a localized outbreak was observed at this hospital. We performed tiled amplicon whole genome nanopore sequencing on nasopharyngeal swabs from all SARS-CoV-2 positive samples from 31 March to 17 April 2020 in 2 local hospitals to assess viral diversity (unknown at the time in Quebec) and potential associations with clinical outcomes. We report 264 viral genomes from 242 individuals–both staff and patients–with associated clinical features and outcomes, as well as longitudinal samples and technical replicates. Viral lineage assessment identified multiple subclades in both hospitals, with a predominant subclade in the Verdun outbreak, indicative of hospital-acquired transmission. Dimensionality reduction identified two subclades with mutations of clinical interest, namely in the Spike protein, that evaded supervised lineage assignment methods–including Pangolin and NextClade supervised lineage assignment tools. We also report that certain symptoms (headache, myalgia and sore throat) are significantly associated with favorable patient outcomes. Our findings demonstrate the strength of unsupervised, data-driven analyses whilst suggesting that caution should be used when employing supervised genomic workflows, particularly during the early stages of a pandemic.

A small number of early introductions seeded widespread transmission of SARS-CoV-2 in Québec, Canada

Background Québec was the Canadian province most impacted by COVID-19, with 401,462 cases as of September 24th, 2021, and 11,347 deaths due mostly to a very severe first pandemic wave. In April 2020, we assembled the Coronavirus Sequencing in Québec (CoVSeQ) consortium to sequence SARS-CoV-2 genomes in Québec to track viral introduction events and transmission within the province. Methods Using genomic epidemiology, we investigated the arrival of SARS-CoV-2 to Québec. We report 2921 high-quality SARS-CoV-2 genomes in the context of > 12,000 publicly available genomes sampled globally over the first pandemic wave (up to June 1st, 2020). By combining phylogenetic and phylodynamic analyses with epidemiological data, we quantify the number of introduction events into Québec, identify their origins, and characterize the spatiotemporal spread of the virus. Results Conservatively, we estimated approximately 600 independent introduction events, the majority of which happened from spring break until 2 weeks after the Canadian border closed for non-essential travel. Subsequent mass repatriations did not generate large transmission lineages (> 50 sequenced cases), likely due to mandatory quarantine measures in place at the time. Consistent with common spring break and “snowbird” destinations, most of the introductions were inferred to have originated from Europe via the Americas. Once introduced into Québec, viral lineage sizes were overdispersed, with a few lineages giving rise to most infections. Consistent with founder effects, the earliest lineages to arrive tended to spread most successfully. Fewer than 100 viral introductions arrived during spring break, of which 7–12 led to the largest transmission lineages of the first wave (accounting for 52–75% of all sequenced infections). These successful transmission lineages dispersed widely across the province. Transmission lineage size was greatly reduced after March 11th, when a quarantine order for returning travellers was enacted. While this suggests the effectiveness of early public health measures, the biggest transmission lineages had already been ignited prior to this order. Conclusions Combined, our results reinforce how, in the absence of tight travel restrictions or quarantine measures, fewer than 100 viral introductions in a week can ensure the establishment of extended transmission chains.

Implementation of serological and molecular tools to inform COVID-19 patient management: protocol for the GENCOV prospective cohort study

IntroductionThere is considerable variability in symptoms and severity of COVID-19 among patients infected by the SARS-CoV-2 virus. Linking host and virus genome sequence information to antibody response and biological information may identify patient or viral characteristics associated with poor and favourable outcomes. This study aims to (1) identify characteristics of the antibody response that result in maintained immune response and better outcomes, (2) determine the impact of genetic differences on infection severity and immune response, (3) determine the impact of viral lineage on antibody response and patient outcomes and (4) evaluate patient-reported outcomes of receiving host genome, antibody and viral lineage results.Methods and analysisA prospective, observational cohort study is being conducted among adult patients with COVID-19 in the Greater Toronto Area. Blood samples are collected at baseline (during infection) and 1, 6 and 12 months after diagnosis. Serial antibody titres, isotype, antigen target and viral neutralisation will be assessed. Clinical data will be collected from chart reviews and patient surveys. Host genomes and T-cell and B-cell receptors will be sequenced. Viral genomes will be sequenced to identify viral lineage. Regression models will be used to test associations between antibody response, physiological response, genetic markers and patient outcomes. Pathogenic genomic variants related to disease severity, or negative outcomes will be identified and genome wide association will be conducted. Immune repertoire diversity during infection will be correlated with severity of COVID-19 symptoms and human leucocyte antigen-type associated with SARS-CoV-2 infection. Participants can learn their genome sequencing, antibody and viral sequencing results; patient-reported outcomes of receiving this information will be assessed through surveys and qualitative interviews.Ethics and disseminationThis study was approved by Clinical Trials Ontario Streamlined Ethics Review System (CTO Project ID: 3302) and the research ethics boards at participating hospitals. Study findings will be disseminated through peer-reviewed publications, conference presentations and end-users.

SARS-CoV-2 B.1.1.7 lineage rapidly spreads and overwhelms R.1 lineage in Japan: serial and stationary observation in a community

Background The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) circulates in the world and acquires mutations during evolution. To identify the new emergent variants, the surveillance of the variants of concern (VOC) and variants of interest (VOI) is ongoing. This study aimed to determine how the transition of viral lineage occurred by stationary genome analysis in Yamanashi, Japan. Methods We performed the whole genome sequencing using SARS-CoV-2 positive samples (n=325) collected from February 2020 to the end of June 2021. The number of analyzed samples accounted for 15.4% of the total 2,109 samples identified in our community. Viral lineage was defined by the Phylogenetic Assignment of Named Global Outbreak (PANGO) lineages. Results We identified 13 types of viral lineages including R.1, P.1, B.1.1.7 (Alpha) and B.1.617.2 (Delta) These virus lineages had distinct periods of expansion and decline. After the emerging of the R.1 lineage harboring E484K variant (designated VOI in Japan), the prevalent B.1.1.214 lineage were no longer identified. The R.1 lineages were temporarily prevalent afterwards, but the influx of B.1.1.7 lineage (designated VOC) led to a decline in R.1. Currently, B.1.1.7 has become dominant after mid-April, 2021. Conclusion We clearly elucidated the transition and replacement of viral lineage by the community-based analysis. The virus completely replaced by more infectious lineages, therefore, it will be necessary to continue to monitor the VOC and VOI.

Molecular genetic analysis of West Nile virus variants circulating in European Russia between 2010 and 2019

Aim. A study of the prevalence of West Nile virus (WNV) genetic lineages and genovariants in the south of European Russia between 2010 and 2019.Materials and methods. The study was carried out on 311 WNV containing biological samples from patients, vectors and reservoirs of infection. WNV typing was carried out using reverse transcription and real-time polymerase chain reaction with designed three pairs of primers and three probes and by the sequencing of the 277 bp WNV genome region corresponding to the 5'-untranslated region and locus of the polyprotein gene encoding the capsid protein C. Sequencing results were analyzed using the Nucleotide BLAST software (NCBI).Results. As a result of typing, out of 311 WNV RNA isolates taken for the study, 15 (4.82%) were assigned to lineage 1 (from Astrakhan and Volgograd regions, Krasnodar and Stavropol Territories, Republic of Tatarstan), 285 (91.64%) to lineage 2 (from Astrakhan, Volgograd, Voronezh, Kursk, Lipetsk, Penza, Rostov and Saratov regions, Krasnodar and Stavropol Territories, Republics of Kalmykia and Crimea), and 11 (3.54%) to lineage 4 (from the Volgograd region, Republics of Kalmykia and Crimea). The predominance of viral lineage 2 was demonstrated. The identified isolates of the viral lineage 1 belonged to the «Astrakhan» variant, isolates of lineage 2 belonged to «Russian» and «European» variants. Previously uncommon WNV variants of lineages 1 and 4 were also found.Conclusion. Lineage 2 of WNV prevailed in the south of European Russia in the last decade. The «Russian» variant is most common and its area is expanding. The circulation of various WNV genetic lineages in Russia indicates the need for further study of their spread and improving diagnostic methods and test systems for identifying and differentiating pathogen strains.

Spike protein cleavage-activation mediated by the SARS-CoV-2 P681R mutation: a case-study from its first appearance in variant of interest (VOI) A.23.1 identified in Uganda

The African continent is currently notable as a source of novel SARS-CoV-2 variants. The A.23 viral lineage, characterized by three spike mutations F157L, V367F and Q613H, was first identified in a Ugandan prison in July 2020, and then spilled into the general population adding additional spike mutations (R102I, L141F, E484K and P681R) to comprise lineage A.23.1 by September 2020, with this virus being designated a variant of interest (VOI) in Africa and with subsequent spread to 26 other countries. The P681R spike mutation of the A.23.1 VOI is of note as it increases the number of basic residues in the sub-optimal SARS-CoV-2 spike protein furin cleavage site; as such, this mutation may affect viral replication, transmissibility or pathogenic properties. Here, we performed assays using fluorogenic peptides mimicking the S1/S2 sequence from A.23.1 and observed significantly increased cleavability with furin, compared to sequences matching Wuhan-Hu1 S1/S2. We performed functional infectivity assays using pseudotyped MLV particles harboring SARS-CoV-2 spike proteins and observed an increase in transduction for A.23.1-pseudotyped particles in Vero-TMPRSS2 and Calu-3 cells, compared to Wuhan-Hu1, and a lowered infection in Vero E6 cells. However, these changes in infectivity were not reproduced in a P681R point mutant of Wuhan-Hu1 spike. Our findings suggest that while A.23.1 has increased furin-mediated cleavage linked to the P681R mutation, which may affect viral infection and transmissibility, this mutation needs to occur on the background of other spike protein changes to enable its functional consequences.

Genomic epidemiology and associated clinical outcomes of a SARS-CoV-2 outbreak in a general adult hospital in Quebec

The first confirmed case of COVID-19 in Quebec, Canada, occurred at Verdun Hospital on February 25, 2020. A month later, a localized outbreak was observed at this hospital. We performed tiled amplicon whole genome nanopore sequencing on nasopharyngeal swabs from all SARS-CoV-2 positive samples from 31 March to 17 April 2020 in 2 local hospitals to assess the viral diversity of the outbreak. We report 264 viral genomes from 242 individuals (both staff and patients) with associated clinical features and outcomes, as well as longitudinal samples, technical replicates and the first publicly disseminated SARS-CoV-2 genomes in Quebec. Viral lineage assessment identified multiple subclades in both hospitals, with a predominant subclade in the Verdun outbreak, indicative of hospital-acquired transmission. Dimensionality reduction identified two subclades that evaded supervised lineage assignment methods, including Pangolin, and identified certain symptoms (headache, myalgia and sore throat) that are significantly associated with favorable patient outcomes. We also address certain limitations of standard SARS-CoV-2 bioinformatics procedures, notably when presented with multiple viral haplotypes.