Exposure to two antigenically distinct SARS-CoV-2 variants broadens neutralization patterns

Previous exposure to one severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant influences neutralizing antibody responses induced by subsequent exposures. Consecutive exposures predominantly back-boost pre-existing immunity and expand cross-neutralizing antibodies while de novo variant-specific responses are poorly induced. In this study, we analyzed neutralizing antibodies against a panel of variants in plasma samples from individuals after exactly two exposures: twice pre-Omicron variant (either two dose vaccinated or infected and one dose vaccinated), pre-Omicron followed by early Omicron variant, or twice early Omicron variant. We found that exposure to two antigenically distinct variants, either pre-Omicron followed by Omicron or two different Omicron variants, increased the neutralization breadth. However, no significant cross-neutralization against the genetically closely related human coronavirus SARS-CoV was induced. Using depletion experiments, we showed that the first exposed variant strongly influences the specificity of antibodies. The second exposure primarily expanded cross-reactive antibodies rather than inducing a variant-specific response against the later variant, highlighting the phenomenon of immune imprinting in the context of SARS-CoV-2. Overall, our results indicate that multiple exposures to SARS-CoV-2 improve cross-neutralization against a variety of variants, but also underscore the lack of de novo antibody production against the more recently encountered variant.