The Johnson and Johnson Ad26.COV2.S single dose vaccine represents an attractive option for COVID-19 vaccination in resource limited countries. We examined the effect of prior infection with different SARS-CoV-2 variants on Ad26.COV2.S immunogenicity. We compared participants who were SARS-CoV-2 naïve with those either infected with the ancestral D614G virus or infected in the second wave when Beta predominated. Prior infection significantly boosts spike binding antibodies, antibody-dependent cellular cytotoxicity and neutralizing antibodies against D614G, Beta and Delta, however neutralization cross-reactivity varied by wave. Robust CD4 and CD8 T cell responses are induced after vaccination, regardless of prior infection. T cell recognition of variants is largely preserved, apart from some reduction in CD8 recognition of Delta. Thus, Ad26.COV2.S vaccination following infection may result in enhanced protection against COVID-19. The impact of the infecting variant on neutralization breadth after vaccination has implications for the design of second-generation vaccines based on variants of concern.

SARS-CoV-2 variants of concern (VOCs) exhibit escape from neutralizing antibodies, causing concern about vaccine effectiveness. However, while non-neutralizing cytotoxic functions of antibodies are associated with improved disease outcome and vaccine protection, Fc effector function escape from VOCs is poorly defined. Furthermore, whether VOCs trigger Fc functions with altered specificity, as has been reported for neutralization, is unknown. Here, we demonstrate that the Beta VOC partially evades Fc effector activity in individuals infected with the original (D614G) variant. However, not all functions are equivalently affected, suggesting differential targeting by antibodies mediating distinct Fc functions. Furthermore, Beta and Delta infection trigger responses with significantly improved Fc cross-reactivity against global VOCs compared to D614G-infected or Ad26.COV2.S vaccinated individuals. This suggests that, as for neutralization, the infecting spike sequence impacts Fc effector function. These data have important implications for vaccine strategies that incorporate VOCs, suggesting these may induce broader Fc effector responses.

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The Janssen (Johnson & Johnson) Ad26.COV2.S non-replicating viral vector vaccine has been widely deployed for COVID-19 vaccination programs in resource-limited settings. Here we confirm that neutralizing and binding antibody responses to Ad26.COV2.S vaccination are stable for 6 months post-vaccination, when tested against multiple SARS-CoV-2 variants. Secondly, using longitudinal samples from individuals who experienced clinically mild breakthrough infections 4 to 5 months after vaccination, we show dramatically boosted binding antibodies, Fc effector function and neutralization. These high titer responses are of similar magnitude to humoral immune responses measured in convalescent donors who had been hospitalized with severe illness, and are cross-reactive against diverse SARS-CoV-2 variants, including the neutralization resistant Omicron (B.1.1.529) variant that currently dominates global infections, as well as SARS-CoV-1. These data have implications for population immunity in areas where the Ad26.COV2.S vaccine has been widely deployed, but where ongoing infections continue to occur at high levels.

The Johnson and Johnson Ad26.COV2.S single dose vaccine, designed as an emergency response to the pandemic, represents an attractive option for the scale-up of COVID-19 vaccination in resource-limited countries. We examined the effect of prior infection with ancestral (D614G) or Beta variants on Ad26.COV2.S immunogenicity approximately 28 days post-vaccination. We compared healthcare workers who were SARS-CoV-2 naive (n=20), to those infected during the first wave prior to the emergence of Beta (n=20), and those infected in the second wave (n=20), when Beta was the dominant variant. We demonstrate that a priming exposure from infection significantly increased the magnitude of spike binding antibodies, neutralizing antibodies and antibody-dependent cellular cytotoxicity activity (ADCC) against D614G, Beta and Delta variants. The magnitude of antibody boosting was similar in both waves, despite the longer time interval between wave 1 infection and vaccination (7 months), compared to wave 2 (2 months). ADCC and binding cross-reactivity was similar in both waves. However, neutralization cross-reactivity varied by wave, showing that the antibody repertoire was shaped by the spike sequence of the infecting variant. Robust CD4 and CD8 T cell responses to spike of similar or higher magnitude as those elicited by infection were induced after vaccination. In contrast to antibody responses, prior infection was not required for the generation of high magnitude T cell responses, and T cell recognition of the Beta variant was fully preserved. Therefore, Ad26.COV2.S vaccination following prior infection, even >6 months previously, may result in substantially enhanced protection against COVID-19, of particular relevance in settings of high SARS-CoV-2 seroprevalence. Furthermore, the dominant impact of the infecting variant on neutralization breadth after vaccination has important implications for the design of second-generation vaccines based on variants of concern.

The continual emergence of novel coronavirus (CoV) strains, like SARS-CoV-2, highlights the critical need for broadly reactive therapeutics and vaccines against this family of viruses. From a recovered SARS-CoV donor sample, we identify and characterize a panel of six monoclonal antibodies that cross-react with CoV spike (S) proteins from the highly pathogenic SARS-CoV and SARS-CoV-2, and demonstrate a spectrum of reactivity against other CoV. Epitope mapping reveals that these antibodies recognize multiple epitopes on SARS-CoV-2 S, including the receptor binding domain, N-terminal domain, and S2 subunit. Functional characterization demonstrates that the antibodies mediate phagocytosis - and in some cases trogocytosis - but not neutralization in vitro . When tested in vivo in murine models, two of the antibodies demonstrate a reduction in hemorrhagic pathology in the lungs. The identification of cross-reactive epitopes recognized by functional antibodies expands the repertoire of targets for pan-coronavirus vaccine design strategies.

SARS-CoV-2 variants of interest have been associated with increased transmissibility, neutralization resistance and disease severity. Ongoing SARS-CoV-2 genomic surveillance world-wide has improved our ability to rapidly identify such variants. Here we report the identification of a potential variant of interest assigned to the PANGO lineage C.1.2. This lineage was first identified in May 2021 and evolved from C.1, one of the lineages that dominated the first wave of SARS-CoV-2 infections in South Africa and was last detected in January 2021. C.1.2 has since been detected across the majority of the provinces in South Africa and in seven other countries spanning Africa, Europe, Asia and Oceania. The emergence of C.1.2 was associated with an increased substitution rate, as was previously observed with the emergence of the Alpha, Beta and Gamma variants of concern (VOCs). C.1.2 contains multiple substitutions (R190S, D215G, N484K, N501Y, H655Y and T859N) and deletions (Y144del, L242-A243del) within the spike protein, which have been observed in other VOCs and are associated with increased transmissibility and reduced neutralization sensitivity. Of greater concern is the accumulation of additional mutations (C136F, Y449H and N679K) which are also likely to impact neutralization sensitivity or furin cleavage and therefore replicative fitness. While the phenotypic characteristics and epidemiology of C.1.2 are being defined, it is important to highlight this lineage given its concerning constellations of mutations.

The SARS-CoV-2 Omicron variant largely escapes neutralizing antibodies elicited by vaccines or infection. However, whether Omicron triggers humoral responses that are cross-reactive to other variants of concern (VOCs) remains largely unknown. We use plasma from 20 unvaccinated and seven vaccinated individuals infected during the Omicron wave in South Africa to test binding, antibody-dependent cellular cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP) and neutralization against VOCs. In unvaccinated individuals, Fc effector function and binding antibodies target Omicron and other VOCs at comparable levels. However, Omicron-triggered neutralization is not extensively cross-reactive to VOCs, with 20 to 43-fold reductions in titer. In contrast, vaccination followed by breakthrough Omicron infection improved cross-neutralization of VOCs, with titers exceeding 1:2,900. This has important implications for the vulnerability of unvaccinated Omicron-infected individuals to reinfection by circulating and emerging VOCs. Further, while Omicron-based immunogens may be adequate boosters, they are unlikely to be superior to existing vaccines for priming in SARS-CoV-2 naive individuals.