Outbreak.info genomic reports: scalable and dynamic surveillance of SARS-CoV-2 variants and mutations.

In response to the emergence of SARS-CoV-2 variants of concern, the global scientific community, through unprecedented effort, has sequenced and shared over 11 million genomes through GISAID, as of May 2022. This extraordinarily high sampling rate provides a unique opportunity to track the evolution of the virus in near real-time. Here, we present outbreak.info , a platform that currently tracks over 40 million combinations of Pango lineages and individual mutations, across over 7,000 locations, to provide insights for researchers, public health officials and the general public. We describe the interpretable visualizations available in our web application, the pipelines that enable the scalable ingestion of heterogeneous sources of SARS-CoV-2 variant data and the server infrastructure that enables widespread data dissemination via a high-performance API that can be accessed using an R package. We show how outbreak.info can be used for genomic surveillance and as a hypothesis-generation tool to understand the ongoing pandemic at varying geographic and temporal scales.

Outbreak.info Research Library: a standardized, searchable platform to discover and explore COVID-19 resources.

Outbreak.info Research Library is a standardized, searchable interface of coronavirus disease 2019 (COVID-19) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) publications, clinical trials, datasets, protocols and other resources, built with a reusable framework. We developed a rigorous schema to enforce consistency across different sources and resource types and linked related resources. Researchers can quickly search the latest research across data repositories, regardless of resource type or repository location, via a search interface, public application programming interface (API) and R package.

Regional connectivity drove bidirectional transmission of SARS-CoV-2 in the Middle East during travel restrictions.

Regional connectivity and land travel have been identified as important drivers of SARS-CoV-2 transmission. However, the generalizability of this finding is understudied outside of well-sampled, highly connected regions. In this study, we investigated the relative contributions of regional and intercontinental connectivity to the source-sink dynamics of SARS-CoV-2 for Jordan and the Middle East. By integrating genomic, epidemiological and travel data we show that the source of introductions into Jordan was dynamic across 2020, shifting from intercontinental seeding in the early pandemic to more regional seeding for the travel restrictions period. We show that land travel, particularly freight transport, drove introduction risk during the travel restrictions period. High regional connectivity and land travel also drove Jordan's export risk. Our findings emphasize regional connectedness and land travel as drivers of transmission in the Middle East.

The molecular epidemiology of multiple zoonotic origins of SARS-CoV-2.

Understanding the circumstances that lead to pandemics is important for their prevention. We analyzed the genomic diversity of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) early in the coronavirus disease 2019 (COVID-19) pandemic. We show that SARS-CoV-2 genomic diversity before February 2020 likely comprised only two distinct viral lineages, denoted "A" and "B." Phylodynamic rooting methods, coupled with epidemic simulations, reveal that these lineages were the result of at least two separate cross-species transmission events into humans. The first zoonotic transmission likely involved lineage B viruses around 18 November 2019 (23 October to 8 December), and the separate introduction of lineage A likely occurred within weeks of this event. These findings indicate that it is unlikely that SARS-CoV-2 circulated widely in humans before November 2019 and define the narrow window between when SARS-CoV-2 first jumped into humans and when the first cases of COVID-19 were reported. As with other coronaviruses, SARS-CoV-2 emergence likely resulted from multiple zoonotic events.

Introduction and Establishment of SARS-CoV-2 Gamma Variant in New York City in Early 2021.

BACKGROUND: Monitoring the emergence and spread of SARS-CoV-2 variants is an important public health objective. We investigated how the Gamma variant was established in New York City (NYC) in early 2021 in the presence of travel restrictions that aimed to prevent viral spread from Brazil, the country where the variant was first identified. METHODS: We performed phylogeographic analysis on 15,967 Gamma sequences sampled between March 10th through May 1st, 2021, to identify geographic sources of Gamma lineages introduced into NYC. We identified locally circulating Gamma transmission clusters and inferred the timing of their establishment in NYC. RESULTS: We identified 16 phylogenetically-distinct Gamma clusters established in NYC (cluster sizes ranged 2-108 genomes); most of them were introduced from Florida and Illinois and only one directly from Brazil. By the time the first Gamma case was reported by genomic surveillance in NYC on March 10th, the majority (57%) of circulating Gamma lineages had already been established in the city for at least two weeks. CONCLUSIONS: Although travel from Brazil to the US was restricted from May 2020 through the end of the study period, this restriction did not prevent Gamma from becoming established in NYC as most introductions occurred from domestic locations.

Post COVID Syndrome Cohort Characterization

Background Post COVID Syndrome (PCS) is significant morbidity following COVID-19. This study aims to identify biomarkers that predict PCS in a Gulf Coast cohort known for poor health outcomes. Methods Since March 2020 the study Collection of Serum and Secretions for SARS CoV-2 Countermeasure Development (aka ClinSeqSer) has been enrolling subjects with confirmed acute COVID-19, with initial visit at 1 month and follow up every three months from symptom onset. At follow-up, subjects complete symptom questionnaire, physical examination, nasopharyngeal swab/saliva collection, blood draw. Subjects with >= one symptom new since COVID are PCS, remainder are Non-PCS experienced at initial one month visit and six months or longer. Univariate and bivariate analysis was carried out to study significant associations of currently available dataset (N=60). Figure 1. Post-COVID Symptoms Included if “new since covid”. For 60 subjects consented post-covid with completed questionnaire, results were analyzed. Most common symptoms reported were fatigue/tiredness or exhaustion (52%), muscle aches (38%), difficulty concentrating (33%) and headache (32%) as the most common symptoms during one month prior to their initial follow-up visit. The persistent symptoms experienced for six months or longer were fatigue/tiredness or exhaustion (25%), forgetfulness (22%), muscle aches (18%), and sleep difficulties (18%). Results Cohort is 36 (60%) female, 24 (40%) male, age group of 49 (82%) 18-64 years, 11 (18%) 65+ years, 33 (55%) African American, 27 (45%) Caucasian. Median follow-up time after symptom onset: 290 days. Study cohort reported fatigue (52%), myalgias (38%), difficulty concentrating (33%), headache (32%) as most common symptoms during first month from initial symptom onset. Persistent symptoms ( >=6 months) are fatigue (25%), forgetfulness (22%), myalgias (18%), sleep difficulties (18%). Bivariate analysis shows that gender (female, P=0.04), past stroke/transient ischemic attack (P=0.04), deep venous thrombosis (P=0.02), abnormal kidney function (P=0.01) associate with PCS. Convalescent antibodies (ReSARS N IgG, S-RBD IgG) were measured and percentage inhibition of ACE2 spike interaction was recorded. Plasma inflammatory protein levels were measured using multiplex ELISA and Proximity Extension Assay technology during follow-up visit. Increased antibody ReSARS N IgG (2.91, 0.74-10.93;P=0.02) response and higher convalescent IL-10 (P=0.04) was associated with PCS. Percent inhibition of ACE2: spike interaction was not associated (P=0.79) with PCS. Nasal swab/saliva SARS-COV-2 sequencing has not identified a specific SARS-CoV-2 virus mutation predictive of PCS. Table 1. Demographic and Clinical Characteristics The bivariate analysis results showed that the gender (female, P=0.0354), history of stroke or transient ischemic attack (P=0.0382), chest pain from narrow heart vessels (P=0.0479), deep venous thrombosis (P=0.0241) and abnormal kidney function (P=0.0142) were associated with Post-COVID syndrome. Table 2. Antibodies and ACE2 spike inhibition. The convalescent antibodies, ReSARS N IgG and S-RBD IgG were measured in U/mL and percentage inhibition of ACE2 spike interaction was recorded during follow-up visit for PCS vs Non-PCS subjects. The increased antibody ReSARS N IgG (2.91, 0.74-10.93;P=0.0159) response was associated with Post-COVID syndrome. Percent inhibition of ACE2: spike interaction was not associated (P=0.7932) with PCS. Table 3. Plasma inflammatory protein levels. Plasma inflammatory protein levels were measured using multiplex ELISA (MSD) and Proximity Extension Assay technology (Olink) recorded during follow-up visit for PCS vs Non-PCS subjects, revealing IL-10 (P=0.0379) was associated with development of PCS. Conclusion This study identifies initial clinical and biomarker predictors of PCS in a cohort that is 55% African American. Figure 2. Antibody ReSARS N IgG ReSARS N IgG measured in post-covid patients is significantly associated with post-COVID syndrome(P=0.01 9). X axis: number of months from symptom onset to blood draw. Y axis: N IgG U/mL. Figure 3. Spike amino acid mutations Spike amino acid mutations detected in SARS-CoV-2 from acute-phase respiratory isolates. Nasal swab/saliva samples were collected from subjects with acute COVID-19 at time of enrollment into ClinSeqSer, stored at -80°C followed by RNA isolation and SARS-CoV-2 qRT-PCR. Samples with Ct value of ≤30 were then sequenced using NextSeq (Illumina). All sequences are deposited on GISAID and under BioProject (ID PRJNA681020). X axis: subject ID, with ID number increasing chronologically. Y axis: amino acid position of each mutation moving from N- to C-terminus. Disclosures Robert Garry, PhD, Zalgen Labs (Shareholder)

Emergence of an early SARS-CoV-2 epidemic in the United States.

The emergence of the COVID-19 epidemic in the United States (U.S.) went largely undetected due to inadequate testing. New Orleans experienced one of the earliest and fastest accelerating outbreaks, coinciding with Mardi Gras. To gain insight into the emergence of SARS-CoV-2 in the U.S. and how large-scale events accelerate transmission, we sequenced SARS-CoV-2 genomes during the first wave of the COVID-19 epidemic in Louisiana. We show that SARS-CoV-2 in Louisiana had limited diversity compared to other U.S. states and that one introduction of SARS-CoV-2 led to almost all of the early transmission in Louisiana. By analyzing mobility and genomic data, we show that SARS-CoV-2 was already present in New Orleans before Mardi Gras, and the festival dramatically accelerated transmission. Our study provides an understanding of how superspreading during large-scale events played a key role during the early outbreak in the U.S. and can greatly accelerate epidemics.

Successful Clearance of 300 Day SARS-CoV-2 Infection in a Subject with B-Cell Depletion Associated Prolonged (B-DEAP) COVID by REGEN-COV Anti-Spike Monoclonal Antibody Cocktail.

A 59-year-old male with follicular lymphoma treated by anti-CD20-mediated B-cell depletion and ablative chemotherapy was hospitalized with a COVID-19 infection. Although the patient did not develop specific humoral immunity, he had a mild clinical course overall. The failure of all therapeutic options allowed infection to persist nearly 300 days with active accumulation of SARS-CoV-2 virus mutations. As a rescue therapy, an infusion of REGEN-COV (10933 and 10987) anti-spike monoclonal antibodies was performed 270 days from initial diagnosis. Due to partial clearance after the first dose (2.4 g), a consolidation dose (8 g) was infused six weeks later. Complete virus clearance could then be observed over the following month, after he was vaccinated with the Pfizer-BioNTech anti-COVID-19 vaccination. The successful management of this patient required prolonged enhanced quarantine, monitoring of virus mutations, pioneering clinical decisions based upon close consultation, and the coordination of multidisciplinary experts in virology, immunology, pharmacology, input from REGN, the FDA, the IRB, the health care team, the patient, and the patient's family. Current decisions to take revolve around patient's follicular lymphoma management, and monitoring for virus clearance persistence beyond disappearance of REGEN-COV monoclonal antibodies after anti-SARS-CoV-2 vaccination. Overall, specific guidelines for similar cases should be established.

METTL3 regulates viral m6A RNA modification and host cell innate immune responses during SARS-CoV-2 infection.

It is urgent and important to understand the relationship of the widespread severe acute respiratory syndrome coronavirus clade 2 (SARS-CoV-2) with host immune response and study the underlining molecular mechanism. N6-methylation of adenosine (m6A) in RNA regulates many physiological and disease processes. Here, we investigate m6A modification of the SARS-CoV-2 gene in regulating the host cell innate immune response. Our data show that the SARS-CoV-2 virus has m6A modifications that are enriched in the 3' end of the viral genome. We find that depletion of the host cell m6A methyltransferase METTL3 decreases m6A levels in SARS-CoV-2 and host genes, and m6A reduction in viral RNA increases RIG-I binding and subsequently enhances the downstream innate immune signaling pathway and inflammatory gene expression. METTL3 expression is reduced and inflammatory genes are induced in patients with severe coronavirus disease 2019 (COVID-19). These findings will aid in the understanding of COVID-19 pathogenesis and the design of future studies regulating innate immunity for COVID-19 treatment.

Emergence and rapid transmission of SARS-CoV-2 B.1.1.7 in the United States.

The highly transmissible B.1.1.7 variant of SARS-CoV-2, first identified in the United Kingdom, has gained a foothold across the world. Using S gene target failure (SGTF) and SARS-CoV-2 genomic sequencing, we investigated the prevalence and dynamics of this variant in the United States (US), tracking it back to its early emergence. We found that, while the fraction of B.1.1.7 varied by state, the variant increased at a logistic rate with a roughly weekly doubling rate and an increased transmission of 40%-50%. We revealed several independent introductions of B.1.1.7 into the US as early as late November 2020, with community transmission spreading it to most states within months. We show that the US is on a similar trajectory as other countries where B.1.1.7 became dominant, requiring immediate and decisive action to minimize COVID-19 morbidity and mortality.

Genomic surveillance reveals multiple introductions of SARS-CoV-2 into Northern California.

The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread globally, with >365,000 cases in California as of 17 July 2020. We investigated the genomic epidemiology of SARS-CoV-2 in Northern California from late January to mid-March 2020, using samples from 36 patients spanning nine counties and the Grand Princess cruise ship. Phylogenetic analyses revealed the cryptic introduction of at least seven different SARS-CoV-2 lineages into California, including epidemic WA1 strains associated with Washington state, with lack of a predominant lineage and limited transmission among communities. Lineages associated with outbreak clusters in two counties were defined by a single base substitution in the viral genome. These findings support contact tracing, social distancing, and travel restrictions to contain the spread of SARS-CoV-2 in California and other states.