Replacement of the Alpha variant of SARS-CoV-2 by the Delta variant in Lebanon between April and June 2021 (preprint)

Background The COVID-19 pandemic continues to expand globally, with case numbers rising in many areas of the world, including the Eastern Mediterranean Region. Lebanon experienced its largest wave of COVID-19 infections from January to April 2021. Limited genomic surveillance was undertaken, with just twenty six SARS-CoV-2 genomes available for this period, nine of which were from travellers from Lebanon detected by other countries. Additional genome sequencing is thus needed to allow surveillance of variants in circulation. Methods Nine hundred and five SARS-CoV-2 genomes were sequenced using the ARTIC protocol. The genomes were derived from SARS-CoV-2-positive samples, selected retrospectively from the sentinel COVID-19 surveillance network, to capture diversity of location, sampling time, gender, nationality and age. Results Although sixteen PANGO lineages were circulating in Lebanon in January 2021, by February there were just four, with the Alpha variant accounting for 97% of samples. In the following two months, all samples contained the Alpha variant. However, this had changed dramatically by June and July, when all samples belonged to the Delta variant. Discussion This study provides a ten-fold increase in the number of SARS-CoV-2 genomes available from Lebanon. The Alpha variant, first detected in the UK, rapidly swept through Lebanon, causing the country’s largest wave to date, which peaked in January 2021. The Alpha variant was introduced to Lebanon multiple times despite travel restrictions, but the source of these introductions remains uncertain. The Delta variant was detected in Gambia in travellers from Lebanon in mid-May, suggesting community transmission in Lebanon several weeks before this variant was detected in the country. Prospective sequencing in June/July 2021 showed that the Delta variant had completely replaced the Alpha variant in under six weeks.

Large scale sequencing of SARS-CoV-2 genomes from one region allows detailed epidemiology and enables local outbreak management (preprint)

The COVID-19 pandemic has spread rapidly throughout the world. In the UK, the initial peak was in April 2020; in the county of Norfolk (UK) and surrounding areas, which has a stable, low-density population, over 3,200 cases were reported between March and August 2020. As part of the activities of the national COVID-19 Genomics Consortium (COG-UK) we undertook whole genome sequencing of the SARS-CoV-2 genomes present in positive clinical samples from the Norfolk region. These samples were collected by four major hospitals, multiple minor hospitals, care facilities and community organisations within Norfolk and surrounding areas. We combined clinical metadata with the sequencing data from regional SARS-CoV-2 genomes to understand the origins, genetic variation, transmission and expansion (spread) of the virus within the region and provide context nationally. Data were fed back into the national effort for pandemic management, whilst simultaneously being used to assist local outbreak analyses. Overall, 1,565 positive samples (172 per 100,000 population) from 1,376 cases were evaluated; for 140 cases between two and six samples were available providing longitudinal data. This represented 42.6% of all positive samples identified by hospital testing in the region and encompassed those with clinical need, and health and care workers and their families. 1,035 cases had genome sequences of sufficient quality to provide phylogenetic lineages. These genomes belonged to 26 distinct global lineages, indicating that there were multiple separate introductions into the region. Furthermore, 100 genetically-distinct UK lineages were detected demonstrating local evolution, at a rate of ~2 SNPs per month, and multiple co-occurring lineages as the pandemic progressed. Our analysis: identified a sublineage associated with 6 care facilities; found no evidence of reinfection in longitudinal samples; ruled out a nosocomial outbreak; identified 16 lineages in key workers which were not in patients indicating infection control measures were effective; found the D614G spike protein mutation which is linked to increased transmissibility dominates the samples and rapidly confirmed relatedness of cases in an outbreak at a food processing facility. The large-scale genome sequencing of SARS-CoV-2-positive samples has provided valuable additional data for public health epidemiology in the Norfolk region, and will continue to help identify and untangle hidden transmission chains as the pandemic evolves.

CoronaHiT: large scale multiplexing of SARS-CoV-2 genomes using Nanopore sequencing (preprint)

The COVID-19 pandemic has spread to almost every country in the world since it started in China in late 2019. Controlling the pandemic requires a multifaceted approach including whole genome sequencing to support public health interventions at local and national levels. One of the most widely used methods for sequencing is the ARTIC protocol, a tiling PCR approach followed by Oxford Nanopore sequencing (ONT) of up to 24 samples at a time. There is a need for a higher throughput method to reduce cost per genome. Here we present CoronaHiT, a method capable of multiplexing up to 95 small genomes on a single Nanopore flowcell, which uses transposase mediated addition of adapters and PCR based addition of symmetric barcodes. We demonstrate the method using 48 and 94 SARS-CoV-2 genomes per flowcell, amplified using the ARTIC protocol, and compare performance with Illumina and ARTIC ONT sequencing. Results demonstrate that all sequencing methods produce inaccurate genomes when the RNA extract from SARS-CoV-2 positive clinical sample has a cycle threshold (Ct) >= 32. Results from set same set of 23 samples with a broad range of Cts show that the consensus genomes have >90% coverage (GISAID criteria) for 78.2% of samples for CoronaHiT-48, 73.9% for CoronaHiT-94, 78.2% for Illumina and 73.9% for ARTIC ONT, and all have the same clustering on a maximum likelihood tree. In conclusion, we demonstrate that CoronaHiT can multiplex up to 94 SARS-CoV-2 genomes per nanopore flowcell without compromising the quality of the resulting genomes while reducing library preparation complexity and significantly reducing cost. This protocol will aid the rapid expansion of SARS-CoV-2 genome sequencing globally, to help control the pandemic.