Current COVID-19 vaccines significantly reduce overall morbidity and mortality and are vitally important to controlling the pandemic. Individuals who previously recovered from COVID-19 have enhanced immune responses after vaccination (hybrid immunity) compared to their naïve-vaccinated peers; however, the effects of post-vaccination breakthrough infections on humoral immune response remain to be determined. Here, we measure neutralizing antibody responses from 104 vaccinated individuals, including those with breakthrough infections, hybrid immunity, and no infection history. We find that human immune sera following breakthrough infection and vaccination following natural infection, broadly neutralize SARS-CoV-2 variants to a similar degree. While age negatively correlates with antibody response after vaccination alone, no correlation with age was found in breakthrough or hybrid immune groups. Together, our data suggest that the additional antigen exposure from natural infection substantially boosts the quantity, quality, and breadth of humoral immune response regardless of whether it occurs before or after vaccination.

Breakthrough infections after vaccination against SARS-CoV-2 are increasingly reported, possibly due to waning of vaccine-induced antibody levels. 1 Moreover, emerging variants of concern with diminished susceptibility to vaccineinduced antibodies are responsible for most new cases. 2,3 Studies have focused on determining the rate of vaccine breakthrough based on antibody levels after standard vaccination practices. 4,5 We assessed antibody levels and variant cross-neutralization after breakthrough infection.Methods | Fully vaccinated health care workers subsequently diagnosed with SARS-CoV-2 breakthrough infection based on a positive polymerase chain reaction (PCR) test result were sequentially recruited at the Oregon Health & Science University between January 31, 2021, and August 18, 2021. Only those with no history of previous infection whose test results were negative for nucleocapsid antibodies were included. Controls were fully vaccinated individuals without a breakthrough infection matched on sex, age, time between vaccine doses, and time between sample collection and most recent antigen exposure (PCR confirmation for those with breakthrough infection and final vaccine dose for controls). Fulllength viral genomic sequencing was used to determine SARS-CoV-2 variant identity. Enzyme-linked immunosorbent assays were used to determine serum dilution titers with a 50% effective concentration (EC 50 ) of IgG, IgA, and IgM antibodies specific to the SARS-CoV-2 spike receptor-binding domain. Live SARS-CoV-2 neutralizing serum dilution titers were determined by 50% focus reduction neutralization tests (FRNT 50 ) against isolates of the original SARS-CoV-2 strain (WA1) and variants of concern (Alpha, Beta, Gamma, and Delta). Median breakthrough and control sera values were calculated in GraphPad Prism and compared with the Wilcoxon matched-pairs signed rank test with the Holm-Šídák correction. Delta-neutralizing potency was determined by comparing Delta-and WA1-neutralizing titers for sequenceconfirmed Delta variant breakthrough cases, non-Delta breakthrough cases, and controls using the Kruskal-Wallis test with Dunn correction. Statistical significance was defined as 2-tailed P < .05. Additional laboratory methods are provided in the Supplement. The Oregon Health & Science University institutional review board approved this study. Written informed consent was obtained.

SARS-CoV-2 and its variants continue to infect hundreds of thousands every day despite the rollout of effective vaccines. Therefore, it is essential to understand the levels of protection that these vaccines provide in the face of emerging variants. Here, we report two demographically balanced cohorts of BNT162b2 vaccine recipients and COVID-19 patients, from which we evaluate neutralizing antibody titers against SARS-CoV-2 as well as the B.1.1.7 (alpha) and B.1.351 (beta) variants. We show that both B.1.1.7 and B.1.351 are less well neutralized by serum from vaccinated individuals, and that B.1.351, but not B.1.1.7, is less well neutralized by convalescent serum. We also find that the levels of variant-specific anti-spike antibodies are proportional to neutralizing activities. Together, our results demonstrate the escape of the emerging SARS-CoV-2 variants from neutralization by serum antibodies, which may lead to reduced protection from re-infection or increased risk of vaccine breakthrough.

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We tested human sera from large, demographically balanced cohorts of BNT162b2 vaccine recipients (n=51) and COVID-19 patients (n=44) for neutralizing antibodies against SARS-CoV-2 variants B.1.1.7 and B.1.351. Although the effect is more pronounced in the vaccine cohort, both B.1.1.7 and B.1.351 show significantly reduced levels of neutralization by vaccinated and convalescent sera. Age is negatively correlated with neutralization in vaccinee, and levels of variant-specific RBD antibodies are proportional to neutralizing activities.