key: cord-0795206-j04itdx1
authors: Bochnia-Bueno, L.; De Almeida, S. M.; Raboni, S. M.; Adamoski, D.; Amadeu, L. L. M.; Carstensen, S.; Nogueira, M. B.
title: SARS-CoV-2 vaccination with CoronaVac: seroconversion rate in healthcare workers after 40 days
date: 2021-05-22
journal: nan
DOI: 10.1101/2021.05.20.21255825
sha: 8d2d924c28a2d2f1034716b2971c45bd20db772c
doc_id: 795206
cord_uid: j04itdx1

Background: This study aimed to calculate the seroconversion rate of the CoronaVac vaccine in healthcare workers (HCWs) 40 days after administration. Methods: Serum samples from 133 HCWs from Southern Brazil were collected one day before (Day 0) and 10, 20, and 40 days after administering the vaccines first dose. Immunoglobulin G (IgG) was quantified using immunoassays for anti-N-protein antibodies (Abbott, Sligo, Ireland) and for anti-S1 (spike) protein antibodies (Euroimmun, Lubeck, Germany). Results: Seroconversion by D 40 (20 days after the second dose) occurred in 129 (97%) HCWs for the S1 protein, and in 69 (51.87%) HCWs for the N protein. An absence of IgG antibodies (by both methodologies), occurred in two (1.5%) HCWs undergoing semiannual rituximab administration, and also in another two (1.5%) HCWs with no apparent reason. Conclusion: This study showed that CoronaVac has a high seroconversion rate when evaluated in an HCW population.

This study aimed to identify the seroconversion rate after vaccination with SARS-CoV-2 (CoronaVac) in healthcare workers (HCWs) 40 days after its application.

In (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The inclusion criteria were as follows: answering the questionnaire, being vaccinated with two doses of CoronaVac, and providing serum samples. Fourteen participants were excluded because they did not complete the questionnaire. Seven participants took another vaccine, one participant did not have the second dose, and 15 participants did not provide a sample on days 0 or +40.

Serum samples of 133 healthcare workers included in this study were collected on days 0 (first dose application), +10, +20 (second dose application), and +40. All samples were stored at −20 ºC until analysis.

The participants were divided into two groups based on day 0 serology according to anti-spike-1 (anti-S1) immunoglobulin G (IgG) 6-8 : reactive (n=16) and non-reactive (n=117). The participants were also sorted according to the presence of comorbidities into two divisions: probably immunosuppressed (n=9) or not (n=124).

Semi-quantitative assays were performed to detect anti-SARS-CoV-2 IgG. For all serum samples, assays used the Chemiluminescent Microparticle Immunoassay (CMIA) Architect-I System for anti-nucleocapsid protein (anti-N) IgG (Abbott, Sligo, Ireland).

Additionally, for serum samples from days 0 and +40, assays used the Enzyme-Linked Immunosorbent Assay (EIA) for IgG anti-S1 spike-protein receptor-binding domain (RBD) (Euroimmun, Lübeck, Germany).

Samples were tested in duplicate, following the manufacturer's instructions. Results with a variation coefficient greater than 15·0% were repeated.

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(which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. According to the distribution of seroconversion at day +40, the category variables were evaluated using Pearson's chi-squared test with Yates' continuity correction. The age variable was evaluated using the Wilcoxon signed rank sum test with continuity correction. Samples paired over time were evaluated using the Friedman ANOVA test (as implemented in the rstatix package), followed by the Wilcoxon signed rank test as a post-hoc pairwise comparison. For samples without multiple observations over time, the Wilcoxon signed rank test was used. All statistical analyses were performed using R (R Core Team).

Robust production of S1-protein IgG was observed by day +40 in 129 (97%) HCW participants ( Figure 1B, D) . Although the reactive and non-reactive groups had different values for S1-protein IgG on day 0 (p < 0·0001), on day +40, the average index between the groups was not significantly different (p = 0·3704).

No significant production of the anti-N-protein IgG was observed in non-reactive group participants 10 days after the first vaccine dose (p = 0·5027; Figure 1A ), and although there was a statistical difference in the sample on day +20 (p < 0·0001), there was no apparent seroconversion at that time. By contrast, there was a marked increase in Nprotein IgG levels in 69 (51·87%) participants on day +40 ( Figure 1A) . A significant difference was also observed in the average index for this antibody between the reactive ( Figure 1C ) and non-reactive groups ( Figure 1A ): day 0 (p < 0·0001) and day +40 (p = 0·0657).

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(which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. In the non-reactive group, better-developed antibody responses were observed for N and S1 proteins (p < 0·0001; Figure 1A , B), while in the reactive group, the antibody response showed a significant difference (p < 0·0001) only for antibodies against S1 protein ( Figure 1D ), increasing the level of circulating humoral response. No significant alteration was observed in IgG anti-N protein analysis for the reactive group at days +10, +20, and +40 (p = 0·2231).

Comorbidities were reported by some HCWs, including Crohn's disease, prior bariatric surgery, HIV+, or diabetes (Table 1 ). In general, the participants with comorbidities responded to the vaccine similarly to participants without any comorbidities. However, two cases in the immunosuppressed group did not undergo seroconversion ( Figure 2 ). Furthermore, two other HCWs (not in the immunosuppressed group) did not seroconvert by day +40; both had no apparent cause. This four HCWs without seroconversion were re-evaluated by a serum sample taken at +60 days. Of these, one participant presented seroconversion of the S1 protein ( Figure 3 ).

In the anti-S1 reactive group on day 0, six (37·50%) participants did not have a previous SARS-CoV-2 diagnosis, possibly due to an asymptomatic infection. Furthermore, in the anti-S1 non-reactive group, 7 (5·98%) participants had symptoms suggestive of SARS-CoV-2, although we did not have information about nasopharyngeal RT-PCR or immunological rapid-test detection. Epidemiologic and clinical participant data are shown in Table 1 .

The seroconversion rate of 97% for the anti-S1 IgG observed in HCWs is important data for the scientific community, since in phase III studies of this and other vaccine candidates for SARS-CoV-2, similar or lower seroconversion rates were observed 9 . All rights reserved. No reuse allowed without permission.

(which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. Such mutations confer the potential for the virus to escape the humoral immune response produced by the body due to the disease or to mRNA vaccines 15 . Thus, studies that evaluate vaccine efficacy against these new strains are valuable 16 . Therefore, seroconversion rates observed for N-protein IgG could be valuable with the emergence of SARS-CoV-2 variants, considering the lower mutation levels in this protein 8 , compared to the high mutation levels in the S1 protein 6 . Therefore, seroconversion of N-protein antibodies may be an alternative for the vaccine industry to produce efficient vaccines for circulating strains, including those that may arise in the future.

Curiously, there was no difference in the analysis for the N-protein IgG in the reactive group, possibly due to the antibody levels present at day 0 in this group, the vaccine has not interfered in the humoral response; the group remained at the same average index.

However, 5·98% of the participants in the group without seroconversion had previously In participants with immunosuppressive treatment, the absence of the antibody response was probably due to rituximab having been administered approximately one month before the vaccine. In this situation, as described by Kado et al. (2016) , B lymphocytes All rights reserved. No reuse allowed without permission.

(which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted May 22, 2021. ; https://doi.org/10.1101/2021.05.20.21255825 doi: medRxiv preprint are completely absent in the body. Consequently, there is no production of antibodies until the B lymphocytes recover in 6 to 24 months. In such cases, the response must be evaluated after the repletion time, and re-vaccination considered with medical and clinical endorsement. Of the other two cases lacking seroconversion at day +40, one participant presented seroconversion on day +60: a late response. The only unexplained case of no seroconversion should be further studied to understand what interfered with the immune response.

The immune response developed by vaccination depends not simply on antibodies, but largely on neutralizing antibodies 19 . Both natural infection and vaccination act on the immune system in complex ways, stimulating the production of non-neutralizing antibodies (with their own specific actions) as well as TCD4 + and TCD8 + T cells, which also act to protect against COVID-19, as shown by Tarke et al. (2021) . That study evaluated the immune response to the SARS-CoV-2 variants, and showed that cellular immunity-unlike the humoral response is little affected by the virus variants. In addition to the specific immune response, innate immunity is another important protection mechanism against infections 19 .

The present study has some limitations: the humoral immunity was studied semiquantitatively, there was no quantification and titration of anti-SARS-CoV-2 antibodies, and no testing for neutralizing antibodies. The total number of participants was small, and immunosuppressed comorbidities were low in number and had diverse etiologies.

More studies are needed to elucidate the vaccine response in these specific groups.

However, this is the first study to evaluate the dynamics of IgG production after CoronaVac immunization in the community.

The results of seroconversion have shown the importance of two doses for this vaccine as, until the second dose was applied, there was no change in the production of N-All rights reserved. No reuse allowed without permission.

(which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. protein IgG, as previously described by Zhang et al. (2021) in phase I/II tests for this vaccine, with the antibody response detectable just 14 days after the second dose. The second vaccine dose is important for several types of vaccines, including mRNA vaccines, as described by Dörschug et al. (2021) , resulting in a significant increase in antibody levels. Therefore, with SARS-CoV-2, there would be no difference at this point.

In conclusion, significant antibody production was observed 40 days after the first CoronaVac dose in the large majority of study participants, independent of comorbidities. The anti-N protein and anti-S1 protein antibody responses of participants without prior SARS-CoV-2 infection were comparable with those of the previously infected group, in which the immune response was maintained or optimized, with no decrease in levels. However, more studies are necessary to draw conclusions. The line connecting the boxes represents the trend of the data. The dotted line represents the days of the vaccine application (two doses). A -N-protein IgG evaluation in S1antibody nonreactive participants at day 0. B -S1-protein IgG evaluation in S1-protein IgG nonreactive participants at day 0. C -N-protein IgG evaluation in S1-protein IgG reactive participants at day 0. D -IgG anti-S1 protein evaluation in anti-S1 protein IgG reactive participants at day 0. Purple and green lines represent the participants with Rituximab treatment. The dotted line represents the days of the vaccine application (two doses). A -N-protein IgG evaluation. B -S1-protein IgG evaluation. Table 1 : Clinical and epidemiological characteristics of participants included in the study for each respective group. Information on the handling of special cases: two immunosuppressed (Rituximab 1400 mg/semiannually), one myasthenia gravis (Pyridostigmine 120 mg/day), one Crohn's disease ostomized twenty-two years ago (Azathioprine 100 mg/day), two participants with prior bariatric surgery (11 and 12 years), and one HIV+ (Tenofovir 300 mg, Lamivudine 300 mg + Dolutegravir 50 mg/day; CD4 + 541/µL). *Comorbidities (probably immunosuppressed) included:

Immunosuppressive drugs use, Crohn's disease, Previously bariatric surgery, HIV and

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The authors would like to thank all participants who agreed to participate in this study, those involved in the collection and storage of samples, and the Immunochemistry Laboratory section of Complexo Hospital de Clínicas, UFPR, and CAPES.

The authors declare that there is no conflict. All rights reserved. No reuse allowed without permission.(which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.The copyright holder for this preprint this version posted May 22, 2021. ; https://doi.org/10.1101/2021.05.20.21255825 doi: medRxiv preprint All rights reserved. No reuse allowed without permission.(which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission.(which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. Diabetes. The patient with Myasthenia gravis is not included here because the treatment used was not immunosuppressive.All rights reserved. No reuse allowed without permission.(which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.The copyright holder for this preprint this version posted May 22, 2021. ; https://doi.org/10.1101/2021.05.20.21255825 doi: medRxiv preprint