The animal reservoir from which the SARS-CoV-2 pandemic has emerged has yet to be described. SARS-CoV-2 contains limited adaptions, including the D614G substitution in the spike glycoprotein. Coronaviruses such as SARS-CoV-2 have demonstrated acquisition of substitutions as a result of a proofreading RNA dependent RNA polymerase. Endemic human to human spread is usually contingent on antigenic novelty that condones reinfection in once immune individuals. Deletions in the amino-terminal domain of the S glycoprotein during long-term infections of immunocompromised patients have been reported in recent studies. The authors of this report have described this as an evolutionary pattern that is encompassed by recurrent deletions that can alter defined antibody epitopes. Deletions cannot be corrected using proofreading activity, and may provide an accelerated adaptive advantage to SARS-CoV-2 evolution.
An immunocompromised cancer patient, who could not clear the SARS-CoV-2 infection after 74 days and ultimately succumbed to the virus, has been designated as Pittsburgh long-term infection (PLTI1). Cloned Spike genes were sequenced from clinical materials that were obtained 72 days post COVID-19 diagnosis and two variants with deletions in the amino terminal domain (NTD) were identified. Patient metadata sequences in GISAID were further investigated. Eight patients contained variants that had deletions in the S glycoproteins of the viruses that were collected over an extended period of weeks to months. At earlier points of collection, S sequences were intact while later points demonstrated deletions corresponding with the S gene. Six sequences had deleted mutations that were identical, overlapping, or adjacent to those found within PLTI1. Deletions were also found at an additional site in viruses isolated from two different patients. The parsimonious explanation offered by the authors describes an independence resurgence due to common selective pressure, which in turn yields convergent outcomes.
When conducting further investigations on GISAID sequence database, 1,108 viruses were identified with additional deletions in the S gene. Deletions were observed in mostly replication competent viruses. Under a lack of selective pressure, a preponderance of in-frame deletions with lengths of 3, 6, 9, and 12 were observed. In the deletions found, 93% were in frame and did not produce a stop codon. In the amino-terminal domain, greater than 97% of deletions maintained the open reading frame.
Lower substitution rates have been found in coronaviruses, including SARS-CoV-2 due to an RNA-dependent RNA polymerase with proofreading reactivity. The adaptive evolution of the S glycoprotein is then enhanced by the increased tolerance for deletions, especially with recurrent deletion regions (RDRs). Within the amino terminal domain, the RDRs occupy defined antibody epitopes and deletions confer resistance to a neutralizing antibody at multiple sites. Fitness of the RDR variants is present within the consensus genomes from patients, transmission between individuals, and emergent lineages. Deletions result as a product of replication, therefore occurring at a rate that may exploit replication in healthy populations. When RDRs are selected in vivo, deletion variants are capable of resistance to neutralization by monoclonal antibodies. Viruses cultured in vivo also exhibited similar behavior. As a result, neutralizing responses have increased in regional binding domain and NTD direction.
Additional lineages with altered antigenicity have also arisen globally and have caused concern. In this study, identical and similar recurrent deletions were demonstrated that alter positions 144/145 and 243/244 in the S glycoprotein disrupt binding of antibody 4A8, an antibody which influences immunodominant epitope with the amino-terminal domain. The analysis captured the first representative of what may have become the B.1.1.7 lineage.
Circulating RDR variants may have gone unnoticed, but might be strong indicators for viral evolution. Their potential as intermediates in immune evasion should also be further evaluated. Deletions and substitutions with major amino-terminal domain and RBD epitopes will continue to be an important focus for variant analysis.
