Antibody Response and Variant Cross-Neutralization After SARS-CoV-2 Breakthrough Infection | Infectious Diseases | JAMA | JAMA Network
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Figure 1.  SARS-CoV-2 Spike Receptor-Binding Domain (RBD)–Specific Antibody Levels After Vaccination and Breakthrough Infection
SARS-CoV-2 Spike Receptor-Binding Domain (RBD)–Specific Antibody Levels After Vaccination and Breakthrough Infection

Enzyme-linked immunosorbent assay measurement of serum dilution titers with a 50% effective concentration (EC50) of SARS-CoV-2 spike RBD-binding antibodies. The dotted lines indicate the assay limits of detection. Two-tailed P values were determined using the Wilcoxon matched-pairs signed rank test with the Holm-Šídák multiple comparison correction. Box plots were generated using the Tukey method. The large box displays the median and IQR. The error bars indicate 1.5 times the IQR or the furthest outlier, whichever is closer to the median. All individual data points are displayed as filled circles.

Figure 2.  Live SARS-CoV-2 Variants Neutralization After Vaccination and Breakthrough Infection
Live SARS-CoV-2 Variants Neutralization After Vaccination and Breakthrough Infection

Live SARS-CoV-2 neutralization by focus-forming assay. A, The dotted line indicates the assay limit of detection. Two-tailed P values were determined using the Wilcoxon matched-pairs signed rank test with the Holm-Šídák multiple comparison correction. B, Participants with inconclusive sequencing information were excluded from this analysis. Two-tailed P values were determined using the Kruskal-Wallis test with the Dunn multiple comparison correction. Box plots were generated using the Tukey method. The large box displays the median and IQR. The error bars indicate 1.5 times the IQR or the furthest outlier, whichever is closer to the median. All individual data points are displayed as filled circles.

1.
Levin  EG, Lustig  Y, Cohen  C,  et al.  Waning immune humoral response to BNT162b2 Covid-19 vaccine over 6 months.   N Engl J Med. 2021. doi:10.1056/NEJMoa2114583PubMedGoogle Scholar
2.
Chen  RE, Zhang  X, Case  JB,  et al.  Resistance of SARS-CoV-2 variants to neutralization by monoclonal and serum-derived polyclonal antibodies.   Nat Med. 2021;27(4):717-726. doi:10.1038/s41591-021-01294-wPubMedGoogle ScholarCrossref
3.
Lopez Bernal  J, Andrews  N, Gower  C,  et al.  Effectiveness of Covid-19 vaccines against the B.1.617.2 (Delta) variant.   N Engl J Med. 2021;385(7):585-594. doi:10.1056/NEJMoa2108891PubMedGoogle ScholarCrossref
4.
Bergwerk  M, Gonen  T, Lustig  Y,  et al.  Covid-19 breakthrough infections in vaccinated health care workers.   N Engl J Med. 2021;385(16):1474-1484. doi:10.1056/NEJMoa2109072PubMedGoogle ScholarCrossref
5.
Chau  NVV, Ngoc  NM, Nguyet  LA,  et al.  An observational study of breakthrough SARS-CoV-2 Delta variant infections among vaccinated healthcare workers in Vietnam.   EClinicalMedicine. 2021;41:101143. doi:10.1016/j.eclinm.2021.101143PubMedGoogle Scholar
6.
Falsey  AR, Frenck  RW  Jr, Walsh  EE,  et al.  SARS-CoV-2 neutralization with BNT162b2 vaccine dose 3.   N Engl J Med. 2021;385(17):1627-1629. doi:10.1056/NEJMc2113468PubMedGoogle ScholarCrossref
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    1 Comment for this article
    Breakthrough Infection and the Need for Variant Boosters
    Bradley Banko, MD, MS | Primary Care
    Your study shows that breakthrough infections broaden immunity in the vaccinated. We should focus on the immunity that results from past and breakthrough infections to see if booster vaccinations are even necessary in those patients, especially those at low risk.
    CONFLICT OF INTEREST: None Reported
    Views 90,251
    Citations 0
    Research Letter
    December 16, 2021

    Antibody Response and Variant Cross-Neutralization After SARS-CoV-2 Breakthrough Infection

    Author Affiliations
    • 1Department of Molecular Microbiology & Immunology, Oregon Health & Science University, Portland
    • 2Division of Infectious Diseases, Oregon Health & Science University, Portland
    • 3Vaccine and Gene Therapy Institute, Oregon Health & Science University, Portland
    JAMA. Published online December 16, 2021. doi:10.1001/jama.2021.22898

    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 vaccine-induced 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). Full-length 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 (EC50) 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 (FRNT50) 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 sequence-confirmed 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.

    Results

    Twenty-six participants with breakthrough infections (mean age, 38 years; 20 [77%] women; 24 [92%] were vaccinated with BNT162b2, sampled a median 28 days after PCR date and 213.5 days after final vaccination; 21 [81%] with mild symptoms) were matched to 26 controls (mean age, 39 years; 21 [81%] women; 26 [100%] were vaccinated with BNT162b2, sampled a median 28 days after final vaccination). Total receptor-binding domain–specific immunoglobulin increased in participants with breakthrough infection with a median EC50 of 2152 (95% CI, 961-3596) compared with 668 (95% CI, 473-892) in controls (322% increase; P < .001) (Figure 1A). Median serum dilutions increased for both IgG and IgA. For example, the median IgA EC50 after breakthrough infection was 120 (95% CI, 44-246), compared with 24 (95% CI, 24-24) for controls (502% increase; P < .001). IgM levels were not significantly different between groups (Figure 1B).

    Among sequence-confirmed breakthrough cases, 10 were Delta and 9 were non-Delta infections. Among breakthrough cases, the median FRNT50 against WA1 was 4646 (95% CI, 2283-7053) vs 489 (95% CI, 272-822) for controls (950% increase; P < .001). FRNT50 results for Alpha, Beta, and Gamma variants are shown in Figure 2A. In breakthrough cases, median FRNT50 against the Delta variant was 2482 (95% CI, 1072-4923), compared with 243 (95%CI, 118-336) for controls (1021% increase; P < .001) (Figure 2A). Sera from Delta breakthrough cases displayed improved potency against the Delta variant at 99% (95% CI, 73-151) of WA1 neutralization for each participant, compared with 36% (95% CI, 33-52) for non-Delta cases and 41% (95% CI, 24-56) for controls (Figure 2B).

    Discussion

    Results of this study showed substantial boosting of humoral immunity after breakthrough infection, despite predominantly mild disease. Boosting was most notable for IgA, possibly due to the differences in route of exposure between vaccination and natural infection. In addition, breakthrough sera demonstrated improved variant cross-neutralization, and Delta breakthrough infections in particular exhibited improved potency against Delta vs WA1, suggesting that the protective immune response may be broadened through development of variant boosters with antigenic inserts matching the emerging SARS-CoV-2 variants. Limitations of this study include the small number of samples and the difference in time from initial vaccination to serum collection between the breakthrough and control groups, which emerging evidence suggests may contribute to the development of variant cross-neutralizing antibody responses.6

    Section Editors: Jody W. Zylke, MD, Deputy Editor; Kristin Walter, MD, Associate Editor.
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    Article Information

    Corresponding Author: Fikadu G. Tafesse, PhD, Department of Molecular Microbiology & Immunology, Oregon Health & Science University, 3181 SW Sam Jackson Park Rd, Portland, OR 97239 (tafesse@ohsu.edu).

    Accepted for Publication: December 3, 2021.

    Published Online: December 16, 2021. doi:10.1001/jama.2021.22898

    Author Contributions: Dr Tafesse had full access to all of the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis.

    Concept and design: Bates, McBride, Winders, Curlin, Tafesse.

    Acquisition, analysis, or interpretation of data: All authors.

    Drafting of the manuscript: Bates.

    Critical revision of the manuscript for important intellectual content: All authors.

    Statistical analysis: Bates, McBride, Schoen.

    Obtained funding: Curlin, Tafesse.

    Administrative, technical, or material support: Bates, McBride, Schoen, Curlin, Tafesse.

    Supervision: Curlin, Tafesse.

    Conflict of Interest Disclosures: None reported.

    Funding/Support: This study was funded in part by an unrestricted grant from the M.J. Murdock Charitable Trust, an unrestricted grant from the Oregon Health & Science University (OHSU) Foundation, National Institutes of Health training grant T32HL083808 on Multidisciplinary Research Training in Pulmonary Medicine, and OHSU Innovative IDEA grant 1018784.

    Role of the Funder/Sponsor: The funders had no role in the design and conduct of the study; collection, management, analysis, and interpretation of the data; preparation, review, or approval of the manuscript; and decision to submit the manuscript for publication.

    Additional Contributions: We acknowledge the efforts of the OHSU COVID-19 serology research team for their assistance with sample acquisition and data collection. We also want to thank the Oregon SARS-CoV-2 Genome Sequencing Center at OHSU for their help in sequencing our samples, including Xuan Qin, PhD, Brian O’Roak, PhD, Andrew Adey, PhD, and Benjamin Bimber, PhD. None of the above individuals were compensated for their contributions. We are grateful for the Oregon Health & Science University faculty, staff, and patients who contributed to this study.

    References
    1.
    Levin  EG, Lustig  Y, Cohen  C,  et al.  Waning immune humoral response to BNT162b2 Covid-19 vaccine over 6 months.   N Engl J Med. 2021. doi:10.1056/NEJMoa2114583PubMedGoogle Scholar
    2.
    Chen  RE, Zhang  X, Case  JB,  et al.  Resistance of SARS-CoV-2 variants to neutralization by monoclonal and serum-derived polyclonal antibodies.   Nat Med. 2021;27(4):717-726. doi:10.1038/s41591-021-01294-wPubMedGoogle ScholarCrossref
    3.
    Lopez Bernal  J, Andrews  N, Gower  C,  et al.  Effectiveness of Covid-19 vaccines against the B.1.617.2 (Delta) variant.   N Engl J Med. 2021;385(7):585-594. doi:10.1056/NEJMoa2108891PubMedGoogle ScholarCrossref
    4.
    Bergwerk  M, Gonen  T, Lustig  Y,  et al.  Covid-19 breakthrough infections in vaccinated health care workers.   N Engl J Med. 2021;385(16):1474-1484. doi:10.1056/NEJMoa2109072PubMedGoogle ScholarCrossref
    5.
    Chau  NVV, Ngoc  NM, Nguyet  LA,  et al.  An observational study of breakthrough SARS-CoV-2 Delta variant infections among vaccinated healthcare workers in Vietnam.   EClinicalMedicine. 2021;41:101143. doi:10.1016/j.eclinm.2021.101143PubMedGoogle Scholar
    6.
    Falsey  AR, Frenck  RW  Jr, Walsh  EE,  et al.  SARS-CoV-2 neutralization with BNT162b2 vaccine dose 3.   N Engl J Med. 2021;385(17):1627-1629. doi:10.1056/NEJMc2113468PubMedGoogle ScholarCrossref
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