Prevalence of saliva immunoglobulin A antibodies reactive with severe acute respiratory syndrome coronavirus 2 among Japanese people unexposed to the virus

  1. Keiichi Tsukinoki
  2. Tetsuro Yamamoto
  3. Jiro Saito
  4. Wakako Sakaguchi
  5. Keiichiro Iguchi
  6. Yoshinori Inoue
  7. Shigeru Ishii
  8. Chikatoshi Sato
  9. Mina Yokoyama
  10. Yuki Shiraishi
  11. Noriaki Kato
  12. Hiroyasu Shimada
  13. Akio Makabe
  14. Akihiro Saito
  15. Masanori Tanji
  16. Isao Nagaoka
  17. Juri Saruta
  18. Tetsutaro Yamaguchi
  19. Shigenari Kimoto
  20. Hideyo Yamaguchi

Reviewed by

Read the full article See related articles

Listed in

Log in to save this article

Abstract

While the COVID‐19 pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) poses a threat to public health as the number of cases and COVID‐19‐related deaths are increasing worldwide, the incidence of the virus infection is extremely low in Japan compared with many other countries. To explain this uncommon phenomenon, we investigated the prevalence of naturally occurring (“natural”) antibodies, focusing on those of the secretory immunoglobulin A (sIgA) form, reactive with SARS‐CoV‐2 among Japanese people. One hundred and eighty healthy Japanese volunteers of a wide range of age who had been considered to be unexposed to SARS‐CoV‐2 participated in this study. Saliva samples and blood samples were collected from all of the 180 participants and 139 adults (aged ≥ 20 years) included therein, respectively. The determination of saliva IgA antibodies, mostly comprising sIgA antibodies, as well as serum IgA and immunoglobulin G antibodies, reactive with the receptor binding domain of the SARS‐CoV‐2 spike‐1 subunit proteins was conducted using an enzyme‐linked immunosorbent assay. The major findings were that 52.78% (95% confidence interval, 45.21%–60.25%) of the individuals who had not been exposed to SARS‐CoV‐2 were positive for saliva IgA antibodies with a wide range of levels between 0.002 and 3.272 ng/mL, and that there may be a negative trend in positivity for the antibodies according to age. As we had expected, a frequent occurrence of assumable “natural” sIgA antibodies reactive with SARS‐CoV‐2 among the studied Japanese participant population was observed.

Article activity feed

  1. Version published to 10.1111/1348-0421.13011
  2. Version published to 10.1101/2022.01.09.22268986v3 on medRxiv
  3. ScreenIT

    SciScore for 10.1101/2022.01.09.22268986: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    EthicsIRB: Study approval and ethics: Approval to undertake the study was obtained from the Kanagawa Dental University Research Ethics Review Board (approval number: 792) on April 6, 2021.
    Consent: Written informed consent was obtained from all participants themselves or informed assent from their legal representatives, mostly parents (in the case of children or adolescent participants aged below 20 years), prior to the study onset.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Detection of anti-SARS-CoV-2 antibodies: Enzyme-linked immunosorbent assays (ELISAs) to measure the binding of IgA in saliva, as well as IgA and IgG in serum, to SARS-CoV-2 spike-1 were performed using an assay system modifying the human IgA ELISA quantitation set (#88-102; Bethyl Laboratories, Montgomery, Texas, USA) that had been reported by Yamamoto et al (61).
    anti-SARS-CoV-2
    suggested: None
    However, since that antigen reacts with IgG, spike-1-His recombinant protein (#40591-V08B1; Sino Biological, Beijin, China) was used to measure the cross-antibodies in serum.
    spike-1-His recombinant protein (#40591-V08B1; Sino Biological, Beijin, China)
    suggested: None
    To the wells thus coated with diluted saliva or serum samples for measurement of SARS-CoV-2-reactive IgA or IgG antibodies, 100 μl per well of biotin-labeled antigen at a concentration of 1 μg/ml was added, and incubated for 1 hour at 25 °C.
    IgG
    suggested: (Bethyl Cat# A300-652A, RRID:AB_519340)
    As positive control, commercially available two antibody products, i.e. spike-neutralizing IgG antibody (cat#40592-R001, Sino Biological; from 0 to 20 μg/ml) and spike-neutralizing IgA antibody (cat#E-AB-V1027, Elabscience, Houston, Texas, USA; from 0 to 2 μg/ml) for detecting SARS-CoV-2-reacting IgG and IgA, respectively.
    spike-neutralizing IgG
    suggested: None
    spike-neutralizing IgA
    suggested: None
    cat#E-AB-V1027
    suggested: (PhosphoSolutions Cat# CoV2-RBT, RRID:AB_2868515)
    IgA
    suggested: None
    Software and Algorithms
    SentencesResources
    ODs were measured at 450 nm in a microplate absorbance reader (Bio-Rad Laboratories, Hercules, CA, USA).
    Bio-Rad Laboratories
    suggested: (Bio-Rad Laboratories, RRID:SCR_008426)
    Analyses were performed using IBM SPSS Statistics version 27 (IBM, USA).
    SPSS
    suggested: (SPSS, RRID:SCR_002865)

    Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

    Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:
    The present study has both strengths and limitations. By implementing this immunological survey in a community-based cohort study with a wide age range of healthy Japanese people who had been unexposed to SARS-CoV-2, we were able to collect preliminary but unprecedented epidemiological data regarding the prevalence and levels of assumable polyreactive natural salivary IgA autoantibodies that exhibit reactivity with the virus. To the best of our knowledge, our survey, along with the preceding one (38), is the only available study which has provided data useful for considering a putative role mucosal natural IgA antibodies in protecting the human host from SARS-CoV-2 infection. Supportably, the presence of natural polyreactive sIgA autoantibodies acting as the frontline of mucosal defense against various infections has been demonstrated (59). Our study has several limitations. Firstly, our sample size was not large with limiting the robustness of our findings in saliva, as well as in serum. If such samples could be increasingly available, statical power for the analyses presented here will be increased. Secondly, we do not know whether and what levels of the salivary IgA detected in the presented study are protective through neutralization against SARS-CoV-2 infection. Thirdly, because of the lack of follow-up data, it was also unable to directly correlate negativity for or low levels of SARS-CoV-2-reactive salivary IgA with the feasibility to have the virus infection. Neverthe...

    Results from TrialIdentifier: No clinical trial numbers were referenced.

    Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

    Results from JetFighter: We did not find any issues relating to colormaps.

    Results from rtransparent:
    • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
    • Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
    • Thank you for including a protocol registration statement.

    Results from scite Reference Check: We found no unreliable references.

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

  4. Version published to 10.1101/2022.01.09.22268986v2 on medRxiv
  5. Version published to 10.1101/2022.01.09.22268986v1 on medRxiv