Detection of SARS-CoV-2 intra-host recombination during superinfection with Alpha and Epsilon variants in New York City

Abstract

Recombination is an evolutionary process by which many pathogens generate diversity and acquire novel functions. Although a common occurrence during coronavirus replication, detection of recombination is only feasible when genetically distinct viruses contemporaneously infect the same host. Here, we identify an instance of SARS-CoV-2 superinfection, whereby an individual was infected with two distinct viral variants: Alpha (B.1.1.7) and Epsilon (B.1.429). This superinfection was first noted when an Alpha genome sequence failed to exhibit the classic S gene target failure behavior used to track this variant. Full genome sequencing from four independent extracts reveals that Alpha variant alleles comprise around 75% of the genomes, whereas the Epsilon variant alleles comprise around 20% of the sample. Further investigation reveals the presence of numerous recombinant haplotypes spanning the genome, specifically in the spike, nucleocapsid, and ORF 8 coding regions. These findings support the potential for recombination to reshape SARS-CoV-2 genetic diversity.

Fig. 1. The distribution of allelic frequencies in the index case (NYCPHL-002130) and named partner with suspected superinfection (NYCPHL-002461).

Frequencies of individual alleles shown as ticks, a smoothed kernel density plot is used to highlight clustering patterns, and colors represent allele types.

Fig. 2. Phylogenetic consistency of major and minor variants.

A Phylogeny of Alpha variant immediate relatives. B Root-to-tip regression for Alpha variant. C Phylogeny of Epsilon variant immediate relatives. D Root-to-tip regression for Epsilon variant. NY-NYCPHL-002461 is the genome deposited in GISAID from the case of putative superinfection. NY-NYCPHL-002130 is the genome from the index case.

Fig. 3. Major, minor, and mixed haplotypes in the 947 nucleotide S (spike) gene cloned sequences.

Each row represents a sequenced clone (n = 104). Colored markings denote mutations from the reference genome. Major strain mutations are those found in the Alpha variant. Minor strain mutations are those found in Epsilon variant. Other mutations are found at intermediate or low frequencies. Shared mutations are those shared by B.1 viruses.

Fig. 4. Major, minor, and mixed haplotypes in the 657 nucleotide S (spike) gene cloned sequences.

Each row represents a sequenced clone (n = 93). Colored markings denote mutations from the reference genome. Major strain mutations are those found in the Alpha variant. Minor strain mutations are those found in Epsilon variant. Other mutations are found at intermediate or low frequencies.

Fig. 5. Major, minor, and mixed haplotypes in the 476 nucleotide ORF8 cloned sequences.

Each row represents a sequenced clone (n = 36). Colored markings denote mutations from the reference genome. Major strain mutations are those found in the Alpha variant. Minor strain mutations are those found in Epsilon variant. Other mutations are found at intermediate or low frequencies.

Fig. 6. Putative Alpha/Iota variant recombinant and the nucleotide variation present in the major, minor, and reference strains.

The distribution of the nucleotide variation found in the major, minor, Iota (B.1.526; EPI_ISL_1635735), and single putative recombinant (EPI_ISL_2965250) strains relative to the reference genome (Wuhan Hu-1; bottom gray sequence).