The COVID–19 pandemic continues to be a major global epidemiological challenge with the ongoing emergence of new strain lineages that are more contagious, more virulent, drug resistant and in some cases evade vaccine-induced immunity. In response, the HiFiViral SARS-CoV-2 kit (PacBio; Menlo Park, California) was developed as a scalable solution for the Sequel II and Sequel IIe systems. Unlike amplicon sequencing, the HiFiViral SARS-CoV-2 kit uses tiled probes, resulting in robust genome coverage across a broad range of RNA input quantities and in the presence of new variants. The use of long, accurate HiFi reads enables comprehensive variant detection, including single nucleotide variants, indels, and structural variants, as well as phasing of variants if multiple strains are present in samples. The fully kitted solution contains all reagents needed for viral enrichment and barcoding up to 384 samples. Flexible scaling allows the user to pool 24 – 384 samples into a SMRT bell library to be run on a single SMRT Cell 8M. For high throughput labs, up to 8 SMRT Cells may be loaded on an instrument with no subsequent touch points. SMRT Link analysis reports include variant calling, genome coverage, detection of samples with multiple strains, and a plate performance summary for assay prep QC. File outputs include consensus sequences ready for submission to surveillance databases, reads aligned to the Wuhan reference, and consensus sequences aligned to the reference for advanced users. Here we demonstrate performance across a broad range of sample Ct inputs in control samples and nasopharyngeal samples in samples batches up to 384. HiFiViral for SARS-CoV-2 is a cost effective, convenient, and accurate method for viral sequencing, well-suited for scalable surveillance of a rapidly evolving virus to inform public health decision making.
Organization: PacBio
Year: 2022