Systematic detection of co-infection and intra-host recombination in more than 2 million global SARS-CoV-2 samples (preprint)

Systematic monitoring of SARS-CoV-2 co-infections between different lineages and assessing the risk of intra-host recombinant emergence are crucial for forecasting viral evolution. Here we present a comprehensive analysis of more than 2 million SARS-CoV-2 raw read datasets submitted to the European COVID-19 Data Portal to identify co-infections and intra-host recombination. Co-infection was observed in 0.35% of the investigated cases. Two independent procedures were implemented to detect intra-host recombination. We show that sensitivity is predominantly determined by the density of lineage-defining mutations along the genome, thus we used an expanded list of mutually exclusive defining mutations of specific variant combinations to increase statistical power. We call attention to multiple challenges rendering recombinant detection difficult and provide guidelines for the reduction of false positives arising from chimeric sequences produced during PCR amplification. Additionally, we identify three recombination hotspots of Delta – Omicron BA.1 intra-host recombinants.

Mobilisation and analyses of publicly available SARS-CoV-2 data for pandemic responses (preprint)

The COVID-19 pandemic has seen large-scale pathogen genomic sequencing efforts, becoming part of the toolbox for surveillance and epidemic research. This resulted in an unprecedented level of data sharing to open repositories, which has actively supported the identification of SARS-CoV-2 structure, molecular interactions, mutations and variants, and facilitated vaccine development and drug reuse studies and design. The European COVID-19 Data Platform was launched to support this data sharing, and has resulted in the deposition of several million SARS-CoV-2 raw reads. In this paper we describe (1) open data sharing, (2) tools for submission, analysis, visualisation and data claiming (e.g. ORCiD), (3) the systematic analysis of these datasets, at scale via the SARS-CoV-2 Data Hubs as well as (4) lessons learned. As a component of the Platform, the SARS-CoV-2 Data Hubs enabled the extension and set up of infrastructure that we intend to use more widely in the future for pathogen surveillance and pandemic preparedness.

High incidence of sotrovimab resistance and viral persistence after treatment of immunocompromised patients infected with the SARS-CoV-2 Omicron variant (preprint)

Background: Sotrovimab is a monoclonal antibody that neutralizes SARS-CoV-2 by binding to a highly conserved epitope in the receptor binding domain. It retains activity against the Omicron BA.1 variant and is used to treat immunocompromised patients as they are at increased risk for a severe outcome of COVID-19. Methods: We studied viral evolution in 47 immunocompromised patients infected with Omicron BA.1 or 2 and treated with sotrovimab. SARS-CoV-2 PCR was performed at baseline and weekly thereafter until Ct-value was [≥] 30. All RNA samples were sequenced to determine the variant and occurrence of mutations, in particular in the Spike protein, after treatment. Results: Twenty-four (51%) of the 47 patients were male and their median age was 63 years. Thirty-one (66%) had undergone a solid organ transplantation and 13 (28%) had received prior B-cell depleting therapy. Despite a history of vaccination, 24 of 30 patients with available data on anti-SARS-CoV-2 IgG Spike antibodies prior to treatment with sotrovimab had very low or no antibodies. Median time to viral clearance (Ct-value [≥] 30) after treatment was 15 days (IQR 7-22). However, viral RNA with low Ct-values was continuously detected for at least 28 days after treatment in four patients infected with BA.1. Mutations in the Spike protein at position 337 or 340 were observed in all four patients. Similar mutations were also found after treatment of two patients with a BA.2 infection but both cleared the virus within two weeks. Thus following treatment with sotrovimab, spike mutations associated with reduced in vitro susceptibility were detected in 6 of 47 (13%) patients. Conclusion: Viral evolution towards resistance against sotrovimab can explain treatment failure in most immunocompromised patients and these patients can remain infectious after treatment. Therefore, documenting viral clearance after treatment is recommended to avoid that these patients unintentionally become a source of new, sotrovimab resistant, variants. Research on direct acting antivirals and possibly combination therapy for the treatment of COVID-19 in immunocompromised patients is needed.

Omicron BA.1 and BA.2 are antigenically distinct SARS-CoV-2 variants (preprint)

The emergence and rapid spread of SARS-CoV-2 variants may impact vaccine efficacy significantly. The Omicron variant termed BA.2, which differs genetically substantially from BA.1, is currently replacing BA.1 in several countries, but its antigenic characteristics have not yet been assessed. Here, we used antigenic cartography to quantify and visualize antigenic differences between SARS-CoV-2 variants using hamster sera obtained after primary infection. Whereas early variants are antigenically similar, clustering relatively close to each other in antigenic space, Omicron BA.1 and BA.2 have evolved as two distinct antigenic outliers. Our data show that BA.1 and BA.2 both escape (vaccine-induced) antibody responses as a result of different antigenic characteristics. Close monitoring of the antigenic changes of SARS-CoV-2 using antigenic cartography can be helpful in the selection of future vaccine strains.

Virological characteristics of SARS-CoV-2 vaccine breakthrough infections in health care workers (preprint)

BackgroundSARS-CoV-2 vaccines are highly effective at preventing COVID-19-related morbidity and mortality. As no vaccine is 100% effective, breakthrough infections are expected to occur. MethodsWe analyzed the virological characteristics of 161 vaccine breakthrough infections in a population of 24,706 vaccinated healthcare workers (HCWs), using RT-PCR and virus culture. ResultsThe delta variant (B.1.617.2) was identified in the majority of cases. Despite similar Ct-values, we demonstrate lower probability of infectious virus detection in respiratory samples of vaccinated HCWs with breakthrough infections compared to unvaccinated HCWs with primary SARS-CoV-2 infections. Nevertheless, infectious virus was found in 68.6% of breakthrough infections and Ct-values decreased throughout the first 3 days of illness. ConclusionsWe conclude that rare vaccine breakthrough infections occur, but infectious virus shedding is reduced in these cases.

SARS-CoV-2 infection in cats and dogs in infected mink farms (preprint)

Animals like mink, cats and dogs are susceptible to SARS-CoV-2 infection. In the Netherlands, 69 out of 127 mink farms were infected with SARS-CoV-2 between April and November 2020 and all mink on infected farms were culled after SARS-CoV-2 infection to prevent further spread of the virus. On some farms, (feral) cats and dogs were present. This study provides insight into the prevalence of SARS-CoV-2 positive cats and dogs in ten infected mink farms and their possible role in transmission of the virus. Throat and rectal swabs of 101 cats (12 domestic and 89 feral cats) and 13 dogs of ten farms were tested for SARS-CoV-2 using PCR. Serological assays were performed on serum samples from 62 adult cats and all 13 dogs. Whole Genome Sequencing was performed on one cat sample. Cat-to-mink transmission parameters were estimated using data from all ten farms. This study shows evidence of SARS-CoV-2 infection in twelve feral cats and two dogs. Eleven cats (19%) and two dogs (15%) tested serologically positive. Three feral cats (3%) and one dog (8%) tested PCR-positive. The sequence generated from the cat throat swab clustered with mink sequences from the same farm. The calculated rate of mink-to-cat transmission showed that cats on average had a chance of 12% (95%CI 10% to 18%) of becoming infected by mink, assuming no cat-to-cat transmission. As only feral cats were infected it is most likely that infections in cats were initiated by mink, not by humans. Whether both dogs were infected by mink or humans remains inconclusive. This study presents one of the first reports of interspecies transmission of SARS-CoV-2 that does not involve humans, namely mink-to-cat transmission, which should also be considered as a potential risk for spread of SARS-CoV-2.

Occupational and environmental exposure to SARS-CoV-2 in and around infected mink farms (preprint)

Unprecedented SARS-CoV-2 infections in farmed minks raised immediate concerns regarding human health which initiated intensive environmental investigations. Air sampling was performed in infected mink farms, at farm premises and at residential sites. A range of other environmental samples were collected from minks housing units including bedding material. Inside the farms, high levels of SARS-CoV-2 RNA were found in airborne dust, on surfaces, and on various other environmental matrices. This warns for occupational exposure which was substantiated by considerable SARS-CoV-2 RNA concentrations in personal air samples. Dispersion of SARS-CoV-2 to outdoor air was found to be limited and SARS-CoV-2 RNA was not detected in air samples collected beyond farm premises, implying a negligible environmental exposure risk for nearby communities. Our occupational and environmental risk assessment is in line with whole genome sequences analyses showing mink-to-human transmission in farm workers, but no indications for direct zoonotic transmission events to nearby communities.