Background: Breakthrough infections with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants are increasingly observed in vaccinated individuals. Immune responses towards SARS-CoV-2 variants, particularly Omicron-BA.5, are poorly understood. We investigated the humoral and cellular immune responses of hospitalized COVID-19 patients during Delta and Omicron infection waves.

Methods: The corresponding SARS-CoV-2 variant of the respective patients were identified by whole genome sequencing. Humoral immune responses were analyzed by ELISA and a cell culture-based neutralization assay against SARS-CoV-2 D614G isolate (wildtype), Alpha, Delta (AY.43) and Omicron (BA.1 and BA.5). Cellular immunity was evaluated with an IFN-γ ELISpot assay.

Results: On a cellular level, patients showed a minor IFN-γ response after stimulating PBMCs with mutated regions of SARS-CoV-2 variants. Neutralizing antibody titers against Omicron-BA.1 and especially BA.5 were strongly reduced. Double-vaccinated patients with Delta breakthrough infection showed a significantly increased neutralizing antibody response against Delta compared to double-vaccinated uninfected controls (median complete neutralization titer (NT₁₀₀) 640 versus 80, p<0.05). Omicron-BA.1 infection increased neutralization titers against BA.1 in double-vaccinated patients (median NT₁₀₀ of 160 in patients versus 20 in controls, p=0.07) and patients that received booster vaccination (median NT₁₀₀ of 50 in patients versus 20 in controls, p=0.68). For boosted patients with BA.5 breakthrough infection, we found no enhancing effect on humoral immunity against SARS-CoV-2 variants.

Conclusion: Neutralizing antibody titers against Omicron-BA.1 and especially BA.5 were strongly reduced in SARS-CoV-2 breakthrough infections. Delta and Omicron-BA.1 but not Omicron-BA.5 infections boosted the humoral immunity in double-vaccinated patients and patients with booster vaccination. Despite BA.5 breakthrough infection, those patients may still be vulnerable for reinfections with BA.5 or other newly emerging variants of concern.