Looking for a needle in a haystack. SARS-CoV-2 variant characterization in sewage

Abstract

SARS-CoV-2 variants are emerging worldwide, and monitoring them is key in providing early warnings. Here, we summarize the different analytical approaches currently used to study the dissemination of SARS-CoV-2 variants in wastewater and discuss their advantages and disadvantages. We also provide preliminary results of two sensitive and cost-effective approaches: variant-specific reverse transcription-nested PCR assays and a nonvariant-specific amplicon deep sequencing strategy that targets three key regions of the viral spike protein. Next-generation sequencing approaches enable the simultaneous detection of signature mutations of different variants of concern in a single assay and may be the best option to explore the real picture at a particular time. Targeted PCR approaches focused on specific signature mutations will need continuous updating but are sensitive and cost-effective.

Keywords: Next-generation sequencing (NGS); SARS-CoV-2; Signature mutations; Variants of concern (VOCs); Variants of interest (VOIs); Wastewater-based epidemiology (WBE).

Graphical abstract

Figure 1

Spike protein mutations that can affect both tropism (receptor binding) and immune evasion and are therefore the focus of surveillance. All mutations indicated are related to the reference sequence (NC_045512). Variants of concern correspond to: α, β, γ, and δ. To date (15 July 2021), the rest are variants of interest. Orange ticks indicate deletions and yellow ticks amino acid mutations.