Background
Acute SARS-CoV-2 infections are mostly cleared within a few days but can become chronic in immunocompromised individuals. Impaired immune clearance is also thought to be a major factor in the evolution of new SARS-CoV-2 variants of concern(VOC). However, it is unclear how intra-host variation accumulates over the course of very long-term chronic SARS-CoV-2 infection, and how this reflects evolving mechanisms of adaptive and innate immune evasion
Methods
A chronic infection (505 days) with SARS-CoV-2 B.1 virus of a 23yo male with an unknown immunodeficiency was followed in this study. Longitudinal samples of nasopharyngeal swabs, and corresponding full-length spike haplotypes sequencing, were obtained together with contemporaneous autologous sera. A live virus was isolated for day 329 of infection. Neutralisation assays were performed using FL-virus and lentiviral particles pseudotyped with spike proteins representative of viral quasispecies haplotypes, to assess evolution of immune evasion from autologous and heterologous sera, and a panel of mAb. Calu3 cells were infected with isolated FL-virus or VOCs for comparative studies on innate immune evasion
Results
We observed that original B.1 virus carrying a D614G mutation in spike gradually accumulated 55 further spike mutations during the course of infection, mostly in key antigenic regions. Interestingly, 23 of those are defining of different VOCs such as Alpha and Omicron. Although this patient was not vaccinated or treated with antibody-based therapies, we demonstrated that accumulating mutations led to a virus universally resistant to wave 1 neutralising responses. We also observed that the virus isolate induced significantly lower proinflammatory gene expression in Calu3 cells than B.1 D614G virus. This correlated with a non-spike adaptation that appeared early in the infection, leading to higher expression of the viral innate antagonist protein orf9b. This same mutation has arisen independently in major VOC such as Alpha and Delta
Conclusions
We characterised a pathway by which a wave 1 SARS-CoV-2 virus gradually evolved mechanisms of evasion from both adaptive and innate immune responses redolent of major VOC during a single chronic infection. This provides one of the strongest pieces of evidence for the involvement of long-term infections in the evolution of new variants that resist antibody-mediated immunity. We also suggest adaptations that resist innate immunity occur early in a chronic infection to promote viral persistence