SARS-CoV-2 whole-genome sequencing using reverse complement PCR: For easy, fast and accurate outbreak and variant analysis.

During the course of the SARS-CoV-2 pandemic reports of mutations with effects on spreading and vaccine effectiveness emerged. Large scale mutation analysis using rapid SARS-CoV-2 Whole Genome Sequencing (WGS) is often unavailable but could support public health organizations and hospitals in monitoring transmission and rising levels of mutant strains. Here we report a novel WGS technique for SARS-CoV-2, the EasySeqTM RC-PCR SARS-CoV-2 WGS kit. By applying a reverse complement polymerase chain reaction (RC-PCR), an Illumina library preparation is obtained in a single PCR, thereby saving time, resources and facilitating high-throughput screening. Using this WGS technique, we evaluated SARS-CoV-2 diversity and possible transmission within a group of 173 patients and healthcare workers (HCW) of the Radboud university medical center during 2020. Due to the emergence of variants of concern, we screened SARS-CoV-2 positive samples in 2021 for identification of mutations and lineages. With use of EasySeqTM RC-PCR SARS-CoV-2 WGS kit we were able to obtain reliable results to confirm outbreak clusters and additionally identify new previously unassociated links in a considerably easier workaround compared to current methods. Furthermore, various SARS-CoV-2 variants of interest were detected among samples and validated against an Oxford Nanopore sequencing amplicon strategy which illustrates this technique is suitable for surveillance and monitoring current circulating variants.

Automated summary

This study introduces a novel whole genome sequencing technique for monitoring the transmission and variants of SARS-CoV-2, which proved to be effective in investigating outbreak clusters and identifying new connections amidst the emergence of variants of concern.