Study design and setting
Viral genomes were sequenced from combined nose and throat swab samples taken from patients with SARS-CoV-2 infection collected from Nov 9, 2020, for patients acutely admitted to a ward at either University College London Hospitals (UCLH) or North Middlesex University Hospital (NMUH) on or before Dec 20, 2020, for any clinical reason. The study dates were selected because the first hospitalised patient with the B.1.1.7 variant was admitted on Nov 9, 2020, and the B.1.1.7 variant became dominant in both hospitals by Dec 20, with this date coinciding with a surge in hospitalisations that stretched the capacity of the health services. All hospitalised patients with a positive PCR test during this time period were eligible and included in the study.
Concerns have been raised around the emergence of variants of concern in long-shedding, immunocompromised or treated patients, especially when treatment modalities and prophylaxis target the spike protein (eg, convalescent plasma, monoclonal antibodies, and vaccination). Therefore, as part of the virological dataset, two pre-existing UCLH cohorts were analysed separately to investigate the prevalence of B.1.1.7 variant of concern (VOC)-defining mutations: 123 samples from a longitudinal study of 34 long-shedding patients, including immunocompromised patients who had remained PCR positive for more than 21 days and up to 196 days (median 33 days [IQR 26–64]), and 64 samples from a remdesivir-treated cohort of 32 patients (32 samples obtained before and 32 samples obtained after day 1 of treatment; samples were obtained a median of 5 days [IQR 3–10] before treatment and 13 days [6–19] after treatment).
To explore differences in the clinical severity associated with the B.1.1.7 and other lineages, we did a cohort study across our two centres. Inclusion criteria for this hospitalised cohort were individuals aged at least 18 years whose first PCR-positive SARS-CoV-2 result date and admission date met study criteria.
The clinical information and SARS-CoV-2 PCR samples were collected as part of routine clinical care. Data were extracted and analysed using permission granted by the National Health Service London Westminster Research Ethics Committee (IRAS 284088 ; REC 20/HRA/2505 ).
Viral detection for SARS-CoV-2
An array of SARS-CoV-2 RNA assays (Hologic Aptima TMA assay run on a Panther system [Hologic, San Diego, CA, USA], a laboratory-developed PCR run using the open access functionality of the Panther Fusion System [Hologic], a laboratory-developed extraction-free PCR assay, and the Cepheid Xpert Xpress [Cepheid, Sunnyvale, CA, USA]) were used in the diagnostic laboratory, including non-PCR assays such as transcription-mediated amplification assay, which does not allow for Ct reporting (ie, not inferring on quantitation). Therefore, as Ct values were not always available, samples for sequencing were not pre-selected according to Ct.
Next-generation sequencing (NGS) and genomic data analysis
using the V3 version of the ARTIC primer set from Integrated DNA Technologies (Coralville, IA, USA) to create tiled amplicons across the SARS-CoV-2 genome. Libraries were prepared using Nextera Flex and sequenced using Illumina MiSeq 500v2 kits (Nextera DNA Flex library preparation kit and MiSeq reagent cartridge V2 [Illumina, San Diego, CA, USA]).
and aligned to a selection of publicly available SARS-CoV-2 genomes
A read depth cutoff of ten was applied after assembly; genomes with less than 75% alignment coverage were removed from subsequent analysis. Phylogenetic trees were generated from multiple sequence alignments using IQ-Tree
and FigTree, with lineages assigned (including VOC calls) using pangolin and confirmed by manual inspection of alignments using Aliview.
The COG-UK Mutation Explorer was used to identify potential mutations of concern.