Multiple detection and spread of novel strains of the SARS-CoV-2 B.1.177 (B.1.177.75) lineage that test negative by a commercially available nucleocapsid gene real-time RT-PCR

Abstract

Several lineages of SARS-CoV-2 are currently circulating worldwide. During SARS-CoV-2 diagnostic activities performed in Abruzzo region (central Italy) several strains belonging to the B.1.177.75 lineage tested negative for the N gene but positive for the ORF1ab and S genes (+/+/- pattern) by the TaqPath COVID-19 CE-IVD RT-PCR Kit manufactured by Thermofisher. By sequencing, a unique mutation, synonymous 28948C > T, was found in the N-negative B.1.177.75 strains. Although we do not have any knowledge upon the nucleotide sequences of the primers and probe adopted by this kit, it is likely that N gene dropout only occurs when 28948C > T is coupled with 28932C > T, this latter present, in turn, in all B.1.177.75 sequences available on public databases. Furthermore, epidemiological analysis was also performed. The majority of the N-negative B.1.177.75 cases belonged to two clusters apparently unrelated to each other and both clusters involved young people. However, the phylogeny for sequences containing the +/+/- pattern strongly supports a genetic connection and one common source for both clusters. Though, genetic comparison suggests a connection rather than indicating the independent emergence of the same mutation in two apparently unrelated clusters. This study highlights once more the importance of sharing genomic data to link apparently unrelated epidemiological clusters and to, remarkably, update molecular tests.