Persistence of a SARS-CoV-2 variant with a frameshifting deletion for the duration of a major outbreak

Abstract

Australia experienced widespread COVID-19 outbreaks from infection with the SARS-CoV-2 Delta variant between June 2021 and February 2022. Whole-genome sequencing of virus from an early case revealed a sub-consensus level of sequencing reads supporting a 17-nucleotide frameshift-inducing deletion in ORF7a that truncated the peptide sequence. The variant rapidly became represented at the consensus level (Delta-ORF7aΔ17del) in most of the outbreak cases in Australia. Retrospective analysis of ORF7a deletions in all GISAID clade GK Delta genomes showed that of 4,018,216 genomes, 134,751 (~3.35%) possessed a deletion in ORF7a, with the ORF7aΔ17del mutation by far the most common. Approximately 99.05% of Delta-ORF7aΔ17del genomes on GISAID originated from the Australian Delta outbreak, and comprised 87% of genomes in the outbreak. In vitro comparison of lineages in cell culture showed a significantly greater proportion of cells were infected with Delta-ORF7aΔ17del than with a contemporaneous Delta variant without ORF7aΔ17del (Delta-ORF7aintact), and the proportion was also measurably higher than an early SARS-CoV-2 strain (A.2.2). These results showed that Delta-ORF7aΔ17del potentially has a slight growth advantage compared to Delta-ORF7aintact. Delta-ORF7aΔ17del viruses still produced ORF7a protein, but significantly less than A.2.2, in a different cellular distribution with a more diffuse expression throughout the cytoplasm of infected cells. These data suggest that the proliferation of Delta-ORF7aΔ17del genomes during the Australian Delta outbreak was likely not a result of an intrinsic benefit of the ORF7aΔ17del mutation, but rather a chance founder effect. Nonetheless, the abundance of different ORF7a deletions in genomes worldwide suggests these have some benefit to virus transmission. IMPORTANCE Deletions in the ORF7a region of SARS-CoV-2 have been noted since early in the COVID-19 pandemic, but are generally reported as transient mutations that are quickly lost in the population. Consequently, ORF7a deletions are considered disadvantageous to the virus through possible loss-of-function effects. In constrast to these earlier reports, we present the first report of a SARS-CoV-2 variant with an ORF7a deletion that dominated for the entirety of a protracted outbreak, and found no associated fitness disadvantage or advantage in cell culture. The relatively common rise and fall of ORF7a deletion variants over time likely represent chance founder events followed by proliferation until a more fit variant(s) is introduced to the population. Our global clade-level survey of ORF7a deletions will be a useful resource for future studies into this gene region.