SARS-CoV-2 genomic diversity and the implications for qRT-PCR diagnostics and transmission

Abstract

The COVID-19 pandemic has sparked an urgent need to uncover the underlying biology of this devastating disease. Though RNA viruses mutate more rapidly than DNA viruses, there are a relatively small number of single nucleotide polymorphisms (SNPs) that differentiate the main SARS-CoV-2 lineages that have spread throughout the world. In this study, we investigated 129 RNA-seq data sets and 6928 consensus genomes to contrast the intra-host and inter-host diversity of SARS-CoV-2. Our analyses yielded three major observations. First, the mutational profile of SARS-CoV-2 highlights intra-host single nucleotide variant (iSNV) and SNP similarity, albeit with differences in C > U changes. Second, iSNV and SNP patterns in SARS-CoV-2 are more similar to MERS-CoV than SARS-CoV-1. Third, a significant fraction of insertions and deletions contribute to the genetic diversity of SARS-CoV-2. Altogether, our findings provide insight into SARS-CoV-2 genomic diversity, inform the design of detection tests, and highlight the potential of iSNVs for tracking the transmission of SARS-CoV-2.

Figure 1.

Overview of general diversity of SARS-CoV-2. From outer to inner layers: Annotation of SARS-CoV-2 genome (gray), PCR primer designs (dark red), transcription-regulating sequences (TRS) (orange), intra-host variant density including iSNVs (blue), deletions start sites (red), duplication start sites (yellow), and inversion start sites (green) along the entire genome. For SNPs + iSNVs + SVs, we plotted the density scaled by their allele frequency across the population over 100-bp windows.

Figure 2.

Mutational frequencies of iSNVs and SNPs. (A) Distribution of iSNV AF. We note that the distribution of AF is strictly <50% as iSNVs are below consensus-level by definition. (B) Mutational spectrum of SARS-CoV-2. (C) Mutational spectra of SARS-CoV-1, SARS-CoV-2, and MERS. (D) Mutational spectrum of SARS-CoV-2 by orf/nsp.

Figure 3.

Shared SNPs and SNVs across data sets. (A) Illustration differentiating what we define as an intra-host SNV (iSNV) and an inter-host consensus-level SNP. (B) UpSet plot captures the shared single nucleotide variants between iSNVs and consensus-level SNPs. The horizontal bars on the left show the total number of variants in the given category. Vertical bars indicate the size of the intersection between highlighted (with black circles) sets. Every variant contributes to exactly one intersection size to avoid double counting.

Figure 4.

iSNV and SNP presence on widely used primers and probes. This figure shows the locations on WHO probes and primers that contain SNPs (left) and iSNVs (right). Columns correspond to base pair positions within the probe, and the sequences are 5′-aligned. Rows correspond to the oligonucleotide sequences, and squares are highlighted based on how many samples/genomes contain a variant in that position.

Figure 5.

In-depth analysis of shared iSNVs. (A) Paired samples from patient COVSUBJ 9 in NYC. (B) Paired samples from patient COVSUBJ 0639 in NYC. (C) The distribution of the number of genomic pairs and their shared variants. (D) The number of pairs with variants at given nucleotide positions. Red color represents positions that were shown to be highly homoplasic and more likely to be affected by error (De Maio et al. 2020).