SARS-CoV-2 vaccination-infection pattern imprints and diversifies T cell differentiation and neutralizing response against Omicron subvariants

Omicron variant infection was first reported on November 26, 2021, and rapidly became the globally dominant variant by early 2022, almost completely replacing the Delta variant shortly after widespread COVID-19 vaccine deployment. Despite increased breakthrough infections as vaccine-induced protection waned over time and as variants with immune-evasive mutations spread, the mechanisms by which Omicron displaced Delta were not fully understood. The study aims to investigate whether host immunity shaped by different vaccination-infection patterns contributed to the rapid replacement of Delta by Omicron. Specifically, the authors compared humoral and cellular immune responses across groups with breakthrough infections by Delta or Omicron following different vaccination strategies, and examined how such patterns imprint T-cell differentiation and neutralizing breadth. Understanding these dynamics may inform the durability of population immunity and guide future vaccination programs.

Retrospective analyses documented increased SARS-CoV-2 breakthrough infection rates and declining vaccine efficacy over time across variant waves (e.g., in New York State, 1,620,214 breakthrough cases among fully vaccinated individuals by June 27, 2022; estimated protective efficacy decreased from 92.8% against the prototype strain in May 2021 to 80% during Delta predominance in mid-July 2021, and 68.7% after Omicron emergence in December 2021). Omicron’s extensive receptor-binding domain (RBD) mutations confer substantial immune escape alongside high transmissibility (R0 ~9.1), facilitating spread among vaccinated populations. Prior studies indicate that natural infection drives continued maturation and broadening of antibodies over a year, with memory B cells evolving to produce antibodies more capable of neutralizing immune-evasive variants (e.g., Beta, Delta), whereas vaccine-elicited antibodies tend to plateau weeks after the second dose. Individuals with prior infection often develop broader anti-spike IgG and neutralizing antibody responses than vaccine-only recipients. Long-lived memory B cells undergo affinity maturation in lymph nodes and expand rapidly upon mRNA vaccination, yielding high-potency antibodies. Inactivated whole-virus vaccines, widely used globally, include antigens beyond spike/RBD but generally induce lower neutralizing titers against Omicron after primary series and booster compared with mRNA, subunit, or adenoviral vector vaccines. Nonetheless, real-world data suggest strong protection against severe disease and death with three-dose inactivated regimens in older adults (e.g., ~98.1% efficacy against severe outcomes with Omicron, comparable to mRNA at ~98.3%). Given the increasing immune escape of Omicron subvariants (BA.1, BA.2, BA.4/5), breakthrough infections are expected even after inactivated vaccination, warranting investigation of sub-population immunity in these contexts.

The study compared immune responses across cohorts defined by vaccination-infection patterns, focusing on breakthrough infections with Delta or Omicron after different vaccine regimens and on vaccine-only controls. Reported groups included: Delta breakthrough infection after two-dose inactivated vaccine; Omicron breakthrough infection after two-dose mRNA vaccine; Omicron breakthrough infection after two-dose inactivated vaccine; Omicron breakthrough infection after three-dose inactivated vaccine; and a three-dose inactivated vaccine group without breakthrough infection. Sample sizes mentioned included groups with n ranging from 9 to 340 depending on cohort. Humoral responses were evaluated by measuring neutralizing antibody (NAb) titers against wild-type (WT) SARS-CoV-2 and variants of concern (VOCs), including Delta and Omicron. Pseudovirus neutralization assays quantified NT50 against prototype, BA.1, BA.2, and BA.4/5 subvariants. Additionally, levels of two representative monoclonal antibodies (13C2 and 08B3; also referred to as 1362 and 08B3 in text) targeting distinct RBD epitopes were measured as markers of NAb components. Statistical analyses included Mann–Whitney tests and Wilcoxon rank-sum tests, with geometric mean titers (GMTs) and fold changes reported. Cellular immunity was assessed by ex vivo assays quantifying virus-specific CD4+ and CD8+ T-cell responses to peptide pools spanning S, M, N, and E proteins, including phenotypic analysis of differentiation subsets (e.g., CD45RA+CD27− CD8+ T cells). In vitro assays evaluated T-cell proliferation and multifunctionality (polyfunctionality) across groups.

- Omicron (BA.1) breakthrough infection, whether after two-dose mRNA (M-M-o) or two-dose inactivated vaccination, elicited higher cross-neutralizing antibody levels against multiple variants and stronger T-cell responses than Delta breakthrough infection after two-dose inactivated vaccination. - Neutralizing activity: Omicron breakthrough conferred higher NAb GMTs than Delta breakthrough. Examples reported include higher NAbs against WT and Omicron in Omicron breakthrough groups compared with Delta breakthrough (e.g., WT 178 vs 157, p < 0.001; Omicron 51 vs 12). Delta breakthrough induced notably low NAbs against Omicron (~12), whereas Omicron breakthrough induced high NAbs against Delta (~194). - mRNA vaccination followed by Omicron breakthrough (M-M-o) produced markedly elevated NAb levels compared with Delta breakthrough after inactivated vaccine, with reported fold increases of approximately 2.5-fold (WT), 5.7-fold (Delta), and 27.3-fold (Omicron), and high absolute titers for Omicron (e.g., 336 vs 12; p < 0.0001). - Component NAbs: M-M-o induced substantially higher levels of monoclonal antibody signatures 13C2/1362 and 08B3 than other groups (e.g., 11.3-fold and 13.3-fold higher than certain comparison groups; multiple-fold higher than Omicron breakthrough after inactivated vaccination and three-dose inactivated vaccination), indicating broader and stronger neutralization breadth. - Pseudovirus assays showed group-dependent differences in NT50 against prototype, BA.1, BA.2, and BA.4/5, with Omicron breakthrough groups generally maintaining higher titers across Omicron subvariants than vaccine-only controls. - T-cell imprinting: Vaccination-infection patterns shaped virus-specific T-cell differentiation. The M-M-o group exhibited higher frequencies of S/M/N/E-specific CD4+ T cells and a lower proportion of virus-specific CD45RA+CD27− CD8+ T cells in ex vivo assays compared with other patterns. - Functional T-cell responses: Breakthrough infection groups demonstrated higher proliferative and multifunctional capacities in vitro than the three-dose inactivated vaccine-only group. - Collectively, these findings suggest that under high vaccine coverage, Omicron’s higher immunogenicity upon breakthrough may have contributed to strengthening population immunity barriers against Delta, potentially facilitating Delta’s rapid replacement by Omicron.

The data indicate that Omicron breakthrough infections, particularly following mRNA vaccination, generate higher magnitude and broader neutralizing antibody responses and more robust, functionally capable T-cell responses than Delta breakthrough after inactivated vaccination or vaccine-only regimens. The enhanced cross-neutralization observed—high NAb titers against Delta following Omicron breakthrough and conversely low titers against Omicron after Delta breakthrough—suggests asymmetric immunity that favors protection against Delta in Omicron-exposed, vaccinated populations. This, together with Omicron’s intrinsic transmissibility and immune-evasive features, may help explain why Delta failed to co-circulate with Omicron and was rapidly displaced. Furthermore, distinct T-cell differentiation patterns (elevated S/M/N/E-specific CD4+ responses and reduced terminally differentiated CD8+ subsets in M-M-o) imply immune imprinting governed by the sequence and type of antigen exposure (vaccine platform and breakthrough variant). Such imprinting may influence the durability and breadth of protection against current and emerging Omicron subvariants. These insights are relevant for optimizing booster strategies and vaccine platform selection to elicit broad, durable immunity at the population level.

Omicron breakthrough infection in vaccinated individuals, especially after mRNA vaccination, imprints stronger and broader neutralizing antibody responses and more robust, multifunctional T-cell immunity than Delta breakthrough or vaccine-only strategies. These immune advantages may have contributed to the rapid global replacement of Delta by Omicron under widespread vaccination. The work advances understanding of how vaccination-infection sequences shape immune imprinting and suggests that future booster programs should consider variant exposure history and vaccine platform to optimize cross-variant protection and durability. Continued evaluation against evolving Omicron subvariants (e.g., BA.1, BA.2, BA.4/5) and assessment of long-term immunity will inform adaptive vaccination strategies.