Assessment of Predictors for SARS-CoV-2 Antibodies Decline Rate in Health Care Workers after BNT162b2 Vaccination—Results from a Serological Survey

Abstract

Background:
SARS-CoV-2 is a novel human pathogen causing Coronavirus Disease 2019 that has caused widespread global mortality and morbidity. Since health workers in Israel were among the first to be vaccinated, we had a unique opportunity to investigate the post-vaccination level of IgG anti-S levels antibodies (Abs) and their dynamics by demographic and professional factors.

Methods:
Prospective Serological Survey during December 2020-August 2021 at Barzilai Medical Center among 458 health care workers (HCW) followed for 6 months after the second BNT162b2 vaccine dose.

Results:
Antibody levels before the second dose, and 30, 90 and 180 days after were 57.1 ± 29.2, 223 ± 70.2, 172.8 ± 73.3 and 166.4 ± 100.7 AU/mL, respectively. From GEE analysis, females had higher Abs levels (β = 26.37 AU/mL, p = 0.002). Age was negatively associated with Abs, with a 1.17 AU/mL decrease for each additional year (p < 0.001). Direct contact with patients was associated with lower Abs by 25.02 AU/mL (p = 0.009) compared to working with no such contact. The average decline rate overall for the study period was 3.0 ± 2.9 AU/mL per week without differences by demographic parameters and was faster during the first 3 months after vaccination than in the subsequent 3 months.

Conclusions:
All demographic groups experienced a decline in Abs over time, faster during the first 3 months. Findings of overall Abs lower in males, workers with direct contact with patients, and older workers, should be considered for policy-making about choosing priority populations for additional vaccine doses in hospital settings.

Full text

Vaccines 2022, 10, 1443. https://doi.org/10.3390/vaccines10091443 www.mdpi.com/journal/vaccines
Article
Assessment of Predictors for SARS-CoV-2 Antibodies
Decline Rate in Health Care Workers after BNT162b2
Vaccination—Results from a Serological Survey
Nadav Zacks 1,*,†,‡, Amir Bar-Shai 1,2,†, Hezi Levi 1,3, Anna Breslavsky 2, Shlomo Maayan 3, Evgenia Tsyba 3,
Shlomo Feitelovich 4, Ori Wand 1,2, Moshe Schaffer 1,5, Yaniv Sherer 1,6, Gili Givaty 6, Anat Tzurel Ferber 2,
Tal Michael 1 and Natalya Bilenko 1, 7,*
1 Faculty of Health Sciences, Ben-Gurion University of the Negev, Beer-Sheva 84101, Israel
2 Pulmonary Department, Barzilai University Medical Centre, Ashkelon 7830604, Israel
3 Division of Infectious Diseases, Barzilai University Medical Centre, Ashkelon 7830604, Israel
4 Laboratory Division, Barzilai University Medical Centre, Ashkelon 7830604, Israel
5 Department of Oncology, Barzilai University Medical Centre, Ashkelon 7830604, Israel
6 Medical Administration Department, Barzilai University Medical Centre, Ashkelon 7830604, Israel
7 Regional Medical Office of Ministry of Health, Ashkelon District, Ashkelon 7830604, Israel
* Correspondence: zacksn@post.bgu.ac.il (N.Z.); natalyab1@bmc.gov.il (N.B.)
† These authors contributed equally to this work.
‡ Participated in this study as part of the requirements for graduation from the Goldman Medical School at
the Faculty of Health Sciences, Ben-Gurion University of the Negev, Israel.
Abstract: Background: SARS-CoV-2 is a novel human pathogen causing Coronavirus Disease 2019
that has caused widespread global mortality and morbidity. Since health workers in Israel were
among the first to be vaccinated, we had a unique opportunity to investigate the post-vaccination
level of IgG anti-S levels antibodies (Abs) and their dynamics by demographic and professional
factors. Methods: Prospective Serological Survey during December 2020–August 2021 at Barzilai
Medical Center among 458 health care workers (HCW) followed for 6 months after the second
BNT162b2 vaccine dose. Results: Antibody levels before the second dose, and 30, 90 and 180 days
after were 57.1 ± 29.2, 223 ± 70.2, 172.8 ± 73.3 and 166.4 ± 100.7 AU/mL, respectively. From GEE
analysis, females had higher Abs levels (β = 26.37 AU/mL, p = 0.002). Age was negatively associated
with Abs, with a 1.17 AU/mL decrease for each additional year (p < 0.001). Direct contact with pa-
tients was associated with lower Abs by 25.02 AU/mL (p = 0.009) compared to working with no such
contact. The average decline rate overall for the study period was 3.0 ± 2.9 AU/mL per week without
differences by demographic parameters and was faster during the first 3 months after vaccination
than in the subsequent 3 months. Conclusions: All demographic groups experienced a decline in
Abs over time, faster during the first 3 months. Findings of overall Abs lower in males, workers
with direct contact with patients, and older workers, should be considered for policy-making about
choosing priority populations for additional vaccine doses in hospital settings.
Keywords: SARS-CoV-2 antibodies; Barzilai Medical Center; Health-Care-Works; BNT162b2 vac-
cination; demographic factors; vaccination regimens; waning of antibodies, booster vaccinations;
patiant contect
Citation: Zacks, N.; Bar-Shai, A.;
Levi, H.; Breslavsky, A.; Maayan, S.;
Evgenia,T.; Feitelovich, S.; Wand, O.;
Schaffer, M.; Sherer, Y.; et al.
Assessment of Predictors for SARS-
CoV-2 Antibodies Decline Rate in
Health Care Workers after
BNT162b2 Vaccination—Results
from a Serological Survey. Vaccines
2022, 10, 1443. https://
doi.org/10.3390/vaccines10091443
Academic Editor: Drishya Kurup
Received: 28 July 2022
Accepted: 27 August 2022
Published: 1 September 2022
Publisher’s Note: MDPI stays neu-
tral with regard to jurisdictional
claims in published maps and institu-
tional affiliations.
Copyright: © 2022 by the authors. Li-
censee MDPI, Basel, Switzerland.
This article is an open access article
distributed under the terms and con-
ditions of the Creative Commons At-
tribution (CC BY) license (https://cre-
ativecommons.org/licenses/by/4.0/).

Vaccines 2022, 10, 1443 2 of 11
1. Introduction
SARS-CoV-2 is a novel human pathogen causing Coronavirus Disease 2019 (COVID-
19). COVID-19 is a devastating disease that has caused widespread global mortality and
morbidity [1,2]. COVID-19 was declared a pandemic by the World Health Organization
(WHO) on 11 March 2020 [3]. Severe disease may result in respiratory failure and multi-
organ dysfunction, leading to long-term sequela in recovered patients [4]. Until December
2021, there were over 250 million confirmed cases and over 5 million deaths worldwide.
Israel has over 1.3 million confirmed cases and over 8000 deaths [5,6].
Antibodies, including IgM, IgA, and IgG, can be detected in the blood 5 to 15 days
following symptom onset or a positive Reverse Transcriptase Polymerase Chain Reaction
(RT-PCR) test, with IgM typically appearing first [7]. Specific IgG antibodies to SARS-
CoV-2 antigens can be detected long after the resolution of the infection [8].
The pandemic has prompted the rapid development of vaccines. On December 2020,
the U.S. Food and Drug Administration (FDA) granted an Emergency Use Authorization
for the Pfizer-BioNTech BNT162b2 mRNA vaccine [9]. On 23 August 2021, the FDA ap-
proved the BNT162b2 vaccine for persons aged ≥16 years [10]. The BNT162b2 vaccine pro-
motes the generation of IgG antibodies against viral spike glycoproteins (S1 and S2). The
initially recommended vaccination regimen included two intramuscular injections, given
21 days apart. The FDA has approved a third “booster” shot that may be administrated 5
months later due to concerns of waning immunity over time [11–13].
The phase-3 study of BNT162b2 validated the efficacy and safety of the vaccine: two
doses of BNT162b2 provided 95% protection against COVID-19 in persons 16 years and
older, and the safety was similar to other viral vaccines over an average of two months
[14]. The duration of protection, including the rate at which the anti-SARS-CoV-2 antibod-
ies decrease, is not fully known. Several studies have reported the humoral response in
vaccinated subjects over various periods [15], some of which focused on groups with spe-
cific medical conditions.
Several other studies demonstrated that antibody levels were dynamic after the sec-
ond dose [16] and differed by gender and age [17–20]. 95.5% of participants developed
anti-spike antibodies after 14 days, and antibody titers increased 24.9-fold after the second
dose. As part of our study, we examined additional risk factors for postvaccination im-
munity waning in a multisectoral hospital setting.
In this study, we have investigated the dynamics of post-vaccination antibody levels
over time in health care workers (HCW) of Barzilai Medical Center (BMC) and examined
the effects of sociodemographic and professional factors. On 19 December 2020, Israel be-
gan a national vaccination campaign that prioritized individuals at high risk due to
chronic medical conditions, HCWs, and seniors.
Understanding factors associated with peak antibody levels and the rate of decline
may prove significant in planning future vaccination regimens. They may enable policy-
makers to create a better protection plan for specific population groups. It is still debatable
which dosing regime is optimal and which populations will be most affected by additional
doses. Understanding variables that affect the immune response to vaccination and the
rate of decline may allow custom vaccination plans to be created for individuals requiring
prioritization.
We aimed to assess the humoral response to the BNT162b2 vaccine in HCW over
time and to identify sociodemographic and professional factors influencing the waning of
antibodies after the second dose.

Vaccines 2022, 10, 1443 3 of 11
2. Methods
A prospective serological survey was conducted from December 2020–August 2021
among HCWs of BMC, a tertiary hospital in Ashkelon, Southern Israel, serving approxi-
mately 600,000 people.
2.1. Study Population
Barzilai employs 2700 HCWs, of which 90% (2430 workers) have received at least two
doses of the BNT162b2 vaccine at the time of data collection. Twenty percent (20%) of
HCWs do not have direct professional contact with patients. Inclusion criteria were an age
of 18 or older, signing of an informed consent form, receiving at least two doses of the
BNT162b2 vaccine, and having serological tests performed 0-5 days prior and 30 days
after the second dose of the vaccine. HCWs with a history of infection with SARS-CoV-2,
either before or during the serological survey, were excluded.
Of 2430 HCWs, 600 were eligible for the study according to the study protocol and
were offered to participate in the serological survey. Out of the 600 eligible HCW, 458
(76%) have agreed to participate in this study. Four hundred fifty-two were tested 0-5
days before the second dose of the vaccination (Baseline Test), 355 were tested 28 ± 6 days
after the second vaccine dose (Test #1), 263 were tested 98 ± 7 days after the second dose
(Test #2), and 289 were tested 194 ± 7 days after the second dose (Test #3) (Figure 1). At
Test #1, 343 (96.6%) participants were asked to fill out a demographic and vaccine-related
side effects questionnaire. At the third serological test (Test #3), participants filled out a
questionnaire to determine their physical activity habits, including chronic medication
usage (both prescribed medication and off-the-counter). This information was available
for 189 participants. At each time point, we evaluated the participants for SARS-CoV-2
infection and excluded exposed individuals (36 excluded overall; Baseline Test: 18 partic-
ipants; Test #1: 4 participants; Test #2: 8 participants; Test #3: 6 participants). After that,
181 participants completed all tests and tested negative for SARS-CoV-2 during the sur-
vey. Statistical analysis was performed on the participants who were found negative for
COVID-19 exposure, regardless of how many tests they had participated in (N = 422),
except for the IgG Anti-S decline rate calculation, which was calculated on the participants
who completed all the tests and were found negative for COVID-19 exposure to the (N =
181).
Figure 1. Flow chart of HCW who Participated at Each Stage of The Survey.
Baseline Test
N=452
(18 were found positive for
COVID-19 Exposure)
Test #1
N=355
(4 were found positive for
COVID-19 Exposure)
Test #2
N=263
(8 were found positive for
COVID-19 Exposure)
Test #3
N=289
(6 were found positive for
COVID-19 Exposure)
Second Vaccination
V a cc i n a t e d (O n c e ) B M C W o r k e r s
N=2430
Received first vaccination dose and were scheduled to receive the
second dose
E l i g ib l e f o r B a s e l i n e T e s t
N=600
Where scheduled to receive the second does 0-5 days from the
Baseline Test screening
S t ud y S a m p le
N=458
BMC workers who where eligible have aggreged to participate in
the study
C o mp a r a b l e S am p l e
N=181
Completed all tests and where not exposed to covid-19 at any
stage

Vaccines 2022, 10, 1443 4 of 11
2.2. Study Procedures
Antibody levels against the viral spike antigen (IgG anti-S) were assessed at 4 prede-
termined time points: 0-5 days before the second dose of the vaccination (Baseline Test)
and 30, 60, and 180 days after the second dose (Tests #1, #2, and #3, respectively). The
participants were tested for COVID-19 blood S1/S2 IgG type antibodies (Abs), using Liai-
son chemiluminescent immunoassay kit (DiaSorin, Saluggia, Italy; REF 311450) to assess
immunological status after vaccination.
Infection with SARS-CoV-2 was diagnosed either by a positive RT-PCR in a nasal
swab or, retrospectively, by a positive anti-nucleocapsid antibody test (anti-N). All partic-
ipants underwent two tests for anti-N antibodies on each occasion where IgG anti-S were
measured, using both the Elecsys Anti-SARS-CoV-2 immunoassay (Roche Diagnostics,
Basel, Switzerland) on a Cobas analyzer and the Abbott SARS-CoV-2 IgG nucleocapsid
protein assay (Abbott, Abbott Park, IL) on an Architect analyzer.
2.3. Statistical Analysis
Dependent variables used in this study were (1) geometric means of IgG anti-S anti-
bodies levels at each stage and (2) the rate of Abs levels decline (RD) as the difference
between levels of antibodies between different tests per week. Independent variables used
in this analysis were: age as a continuous variable and age as a dichotomous variable (age
≤ 50 years, age > 50 years), sex (male/female), place of birth (Israel, Asia/Africa, Europe),
profession (direct contact with patient [such as physicians, nurses, radiologists, physio-
therapists, etc.] and no direct contact with patient [administrators, laboratory workers,
housekeepers, IT workers, etc.]), obesity (BMI > 25), self-reported smoking status
(ever/never), self-reported overall physical activity (at work and recreational—yes/no),
night shifts (yes/no), self-reported regular use of medications for chronic conditions
(yes/no). We used ANOVA for comparing Abs between categorical variables and Chi-
square or Fisher’s Exact test when appropriate for comparing proportions. We tested as-
sociations between Abs and independent variables to detect possible confounders. Fi-
nally, multivariate linear regression analysis was built to detect the unique and independ-
ent effect of factors contributing to changes in Abs at each Test. Following this stage, gen-
eralized estimating equations (GEE) models were conducted, accounting for each partici-
pant's repeated measures. The goodness of fit was examined using Bayesian information
criteria (BIC), and the most fitted models were chosen. Betas and 95% confidence interval
(CI) were calculated. Statistical significance was set at p ≤ 0.05. Data analysis and statistical
procedures were conducted using SPSS 25.0 ® (SPSS, Chicago, IL, USA), MATLAB 2021 ®
software, and R statistics (version 4.1.1; A Language and Environment for Statistical Com-
puting, Vienna, Austria)), including the R packages: ‘data.table’, ‘ggplot2’, ‘dplyr’, ‘lubri-
date’ and ‘gee’.
2.4. Ethical Considerations
The study was approved by the Ethics Committee and Institutional Review Board of
Barzilai Medical Centers (No. 0009-21-BRZ). The study was performed in accordance with
the Declaration of Helsinki and Good Clinical Practice guidelines. All participants signed
an informed consent form prior to study enrollment. Results are reported according to
STROBE statement guidelines.
3. Results
The study population included 458 HCWs from BMC belonging to different medical,
paramedical, and management sectors. Our study population generally included rela-
tively young (48.7 ± 10.7 years old), mostly female HCWs (Table 1). Fifty-nine percent of
participants had self-reported BMI above 25. Only one-fifth were Israeli-born. Over half
of studied HCWs have direct contact with patients during work.

Vaccines 2022, 10, 1443 5 of 11
Table 1. Demographic characteristics of the study population
Variable n (%) or Mean ± SD (Median)
Sex N = 458
Male 128 (28%)
Female 330 (72%)
Age [years] 48.7 ± 10.7 (50%)
Place of Birth N = 434
Israel 93 (21.4%)
America/Europe 128 (29.5%)
Asia/Africa 213 (49.1%)
Chronic use of medications N = 186
Yes 74 (40%)
No 112 (60%)
Physical activity N = 189
Yes 110 (58.2%)
No 79 (41.8%)
Direct professional contact with a patient N = 434
Yes 235 (54.1%)
No 199 (45.9%)
BMI 26.0 ± 23.4 (29.4)
Smoker (active or past) N = 435
Yes 292 (67.3%)
No 142 (32.7%)
Night shifts N = 187
Yes 134 (72%)
No 53 (28%)
Some data was not available for all participants.
We examined levels of IgG anti-S Abs after the first and second vaccine doses and
the rate of decline per week over time. At the baseline test, performed 0–5 days prior to
the second vaccine dose, the average antibody levels were 57.1 ± 29.2 AU/mL (median
55.8), with no statistically significant difference between males and females.
Four weeks (28 ± 6 days) after the second dose (Test #1), the antibody levels increased
to an average of 223 ± 70.2 AU/mL (Median 214). Ninety days after the second dose of the
vaccination (Test #2), antibody levels declined to a mean of 172.8 ± 73.3 AU/mL (median
160). At the last measurement performed six months after the second vaccine dose (Test
#3), Abs continued to decrease to an average of 166.4 ± 100.7 AU/mL (Median 137.5).
In the univariate analysis, we found statistically significant associations between IgG
anti-S levels and sex, age, origin, type of contact with a patient, and physical activity, but
not with obesity status, working night shifts and smoking (Appendix A, Table A1).
Beginning from Test #1, female participants had higher levels of IgG anti-S, although
this difference was statistically significant only for Test #1 (p = 0.036, 0.416 and 0.163 for
Tests #1, #2 and #3 respectively) (Figure 2A). At all tests, IgG anti-S Abs levels were higher
for younger individuals. This difference was pronounced and statistically significant
when comparing participants 50 years old or younger with older participants (p = 0.041,
0.088, and 0.557 for Tests #1, #2 and #3, respectively). However, the negative association
of antibody levels with age was less pronounced between tests (Figure 2B). Health care
workers who have direct contact with patients had consistently lower antibody levels than
those without such contact starting from Test #1 but less significantly at Test #3 (p= 0.018,
0.015, and 0.082 for Tests #1, #2, and #3, respectively) (Figure 2C).

Vaccines 2022, 10, 1443 8 of 11
Figure 2. IgG anti-S Levels Over Time. (A) Gender; (B) age (50 or younger, Older than 50); (C) direct patient contact. Circles and crosses represent individual IgG
Anti-S values; means and 95% confidence intervals are only calculated for the comparable sample.

Vaccines 2022, 10, 1443 9 of 11
Further, we examined the rate of Abs decline calculated as the IgG anti-S in AU/mL
difference between tests divided by the number of weeks between them. The average
weekly decline rates were 5.3 ± 4.31 AU/mL/week (median 5.1) between Test #1 and Test
#2 and 0.84 ± 3.1 AU/mL/week (median 1.5) between Test #2 and Test #3. The average
decline rate during the overall study period was 3.0 ± 2.9 AU/mL/week (Median 3.1). We
did not find a significant association between sociodemographic parameters and the rate
of decline.
To identify background variables that could have influenced the humoral response
to vaccination, we developed a GEE model that incorporated variables significantly asso-
ciated with IgG anti-S levels as univariates. We found an independent negative associa-
tion between age in years and IgG anti-S levels with a β of -1.17 for each additional year
(p < 0.001). Since the association was similar for age groups 50 years old or younger and
older than 50, we also assessed age as a categorical variable in our model. We found that
antibody levels are almost 20 AU/mL lower in individuals over 50 years old compared to
younger ones (p = 0.013). Female gender and working without direct patient contact were
found to be associated with higher antibody levels (p = 0.002, 0.009, respectively) (Table 2)
Table 2. Independent effects of sociodemographic and personal background variables on levels of
antibodies from the GEE model.
Characteristic Estimate Standard Error p-value
Test Number −32.28 2.39 <0.001
Age
0.013
≤50 years Ref.
>50 years −19.92 7.10
Sex 0.002
Male Ref.
Female 26.37 8.13
Type of contact with patients 0.009
Direct contact Ref.
Nondirect contact 25.02 7.09
4. Discussion
In this prospective study, we aimed to study how sociodemographic and profes-
sional characteristics influence the titers and decline rates of IgG anti-S antibodies in
HCWs vaccinated with two doses of BNT162b2. Like others [21,22], we found a significant
decrease in antibody levels over time in all demographic groups. We found statistically
significant associations between IgG anti-S levels and sex, age, and type of contact with a
patient, but not with obesity status, physical activity, origin, working night shifts and
smoking. These associations were more significant at the beginning of the survey.
In addition to assessing the humoral response to the BNT162b2 vaccine, our study
also aimed to investigate the influence of sociodemographic factors on the rate of decline
in antibody levels. To our knowledge, this is the first time the decline rate has been meas-
ured and described. We found that absolute Abs levels were negatively associated with
age and male sex of HCW while working in direct contact with patients related to lower
levels. However, we have found that sociodemographic variables do not affect the aver-
age rate of decline over 6 months. Nevertheless, we found that the decline rate in the first
3 months after the second dose was 5–7 times faster than during the following 3 months.
Compared to non-mRNA-based vaccines, antibodies’ decline rates are considerably
slower, with no evidence of significant change in the rate during the first years after the
vaccine [23], suggesting that the humoral response to the BNT162b2 vaccine may be less
durable. One advantage of mRNA vaccines is their short life with an early shut-down of
antigen expression [24]. This aspect of the promising vaccine may be the Achilles’ heel of
the technology.

Vaccines 2022, 10, 1443 10 of 11
We developed a model to assess the individual effects of different variables on IgG
anti-S levels over time. Surprisingly, we identified work environments without exposure
to patients as associated with a lesser decline in antibody levels over time. This observa-
tion is possibly explained by the fact that employees exposed to patients are also more
exposed to various infectious diseases; thus, their immune system is engaged in multiple
processes, making it less available for the short-lived BNT162b2 vaccine.
Uwamino et al. [16] reported similar findings among Japanese participants working
in a medical school and its affiliated hospital in Tokyo, for whom serum samples were
collected before the first dose and three weeks after the second dose of the BNT162b2 vac-
cine. They measured antibody titers against the receptor-binding domain of the spike pro-
tein of SARS-CoV-2. They found that young age (<45 years), female sex, and adverse re-
actions after the second dose were independently related to higher antibody titers after
the second dose. Similar findings were reported in other studies [18–20].
Limitations of the study include a modest sample size and high rates of participants
lost to follow-up or excluded due to SARS-CoV-2 infection over time, which could have
biased the results. Humoral responses are commonly used as surrogate markers for im-
munity after vaccination. Measuring antibody levels is easier to perform and standardize,
less expansive, and commercially available compared to assessing antibodies' cellular im-
mune responses or binding capacities. Yet, the clinical implications of antibody levels are
indeterminate at best. Several publications reported that post-vaccination antibody titers
are predictive of immune protection from COVID-19 [25–27], and real-life studies re-
ported that breakthrough infections correlated with low antibody titers before infection
in HCW [28] and hemodialysis patients [29], yet a protective threshold antibody level for
immunity in currently undefined.
While the initial efficacy of mRNA-based vaccines, including BNT162b2, against con-
tracting SARS-CoV-2, and especially against severe COVID-19, is remarkably high, it is
apparent that this efficacy declines over time. Our results strengthen the indispensability
of additional vaccine doses. Additional vaccine doses, “boosters,” confer improved and
longer-term protection from disease. However, the optimal dosing regime is still debated,
and so are the target populations which will benefit the most from additional doses. A
better understanding of variables that affect the immune response to vaccination may al-
low personally tailored vaccination schedules for individuals requiring prioritization.
Moreover, the fast rate of decline observed in our cohort raises the need to develop a more
efficient and long-lasting COVID-19 vaccine.
5. Conclusions
Female sex, younger age, and no direct contact with patients is associated with
higher IgG anti-S levels in vaccinated HCW. All demographic groups experienced a steep
decline in antibody levels over time. The decline rate of antibody levels is significantly
faster during the first months after the vaccination and slows after that. We believe this
study will be used as grounds for further exploration of the differences in IgG anti-S levels
in HCWs with direct contact with patients from HCWs without direct contact. Consider-
ing the variables that affect absolute antibody levels and the rate of decline may be the
key to deciding when to prioritize the administration of additional vaccine doses in HCW.
This group differs from the general population in terms of age, health status, activity, sex
distribution, and close, continuous contact with in-hospital patients.

Vaccines 2022, 10, 1443 11 of 11
Author Contributions: Conceptualization, A.B.-S., H.L., S.M., M.S., G.G. and N.B.; software, N.Z.,
T.M. and N.B.; formal analysis, N.Z., A.B.-S. and N.B.; data curation, N.Z., A.B., E.T., S.F. and A.T.F.;
writing—original draft preparation, N.Z. and A.B.-S.; writing—review and editing, O.W., H.L., Y.S.
and N.B.; visualization, N.Z.; supervision, N.B. All authors have read and agreed to the published
version of the manuscript.
Funding: This research received no external funding.
Institutional Review Board Statement: The study was conducted according to the guidelines of the
Declaration of Helsinki and approved by the Institutional Ethics Committee of Barzilai Medical
Center. Protocol Number–0009-21-BRZ (3 March 2021).
Informed Consent Statement: Informed consent was obtained from all subjects involved in the
study.
Data Availability Statement: The data presented in this study are available on request from the
corresponding author. The data are not publicly available in accordance with this study’s ethical
protocol.
Conflicts of Interest: The authors declare no conflict of interest.
Appendix A
Table A1. Univariate association between anti-S levels and sociodemographic variables. Data cal-
culated for participants tested at this test were not positive for COVID-19 exposure at the test or any
time before.
Baseline Test (N = 434) Test 1 (N = 343) Test 2 (N = 235) Test 3 (N = 185)
Variable N
x
(Geo) ± μ (Med)
[AU/mL] N
x
(Geo) ± μ (Med)
[AU/mL] N
x
(Geo) ± μ (Med)
[AU/mL] N
x
(Geo) ± μ (Med)
[AU/mL]
Sex
Male 119 47.13 ± 29.49 (51.7)
90 196.75 ± 69.71 (207.5)
61 149.49 ± 74.79 (156) 43 113.17 ± 99.52 (104)
Female 315 49.19 ± 29.05 (56.8)
253 217.47 ± 70.36 (215) 174 161.03 ± 67.93 (159.5)
142 138.9 ± 88.69 (144.5)
p
0.417
0.036
0.416
0.163
Origin
Country
Israel 93 45.38 ± 28.88 (54.3)
74 205.51 ± 64.38 (209.5)
51 153.76 ± 67.58 (142) 37 129.47 ± 75.58 (139)
Europe/America 128 54.46 ± 27.7 (60.8) 101 227.57 ± 71.75 (230) 64 167.55 ± 75.13 (175) 52 132.28 ± 103.65 (151)
Others
213
46.8 ± 29.94 (52.1)
168
205.62 ± 71.5 (209.5)
120
154.82 ± 67.43 (157.5)
96
133.69 ± 90.51 (131.5)
p 0.101 0.046 0.286 0.567
Age Group
age < 40 110 60.71 ± 28.92 (65.6)
82 224.9 ± 65.21 (224.5) 56 178.64 ± 67.61 (180) 40 154.04 ± 96.26 (145)
40 ≤ age < 50
125
51.8 ± 27.76 (56)
97
215.77 ± 69.21 (215)
64
157.49 ± 64.55 (159.5)
47
124.09 ± 83.6 (145)
50 ≤ age < 60 139 47.48 ± 28.41 (54.5)
114 212.64 ± 69.22 (219) 78 154.9 ± 69.85 (155.5) 64 140.58 ± 99.32 (135.5)
age > 60 60 29.95 ± 24.41 (32.5)
50 183.68 ± 79.5 (174) 37 137.31 ± 77.44 (124) 34 108.43 ± 76.04 (109.5)
p <0.001 0.025 0.088 0.137
BMI
Normal 178 52.56 ± 27.21 (58.4)
145 212.29 ± 70.47 (213) 96 167.34 ± 70.13 (172.5)
75 143.65 ± 91.36 (142)
Overweight (BMI >
25) 256 46.05 ± 30.38 (53.1)
198 211.49 ± 70.77 (216.5)
139 151.78 ± 69.06 (155) 110 125.3 ± 91.66 (131)
p 0.146 0.933 0.120 0.331
Smoking
Habit
Not smoking 292 48.28 ± 29.1 (52.75)
226 214.96 ± 69.78 (216.5)
153 156.9 ± 70.58 (156) 123 131.63 ± 85.91 (137)
Past or Active
Smoker 142 49.32 ± 29.36 (57.35)
117 205.91 ± 72.1 (208) 82 159.94 ± 68.47 (163) 62 134.06 ± 102.39 (128)
p 0.592 0.428 0.815 0.675
Occupation
Group
exposed to patients 235 46.46 ± 28.63 (54.3)
189 203.41 ± 69.54 (204) 131 147.21 ± 70.9 (147) 104 120.71 ± 91.76 (124)
No exposure 199 51.29 ± 29.66 (56.6)
154 222.64 ± 70.72 (227.5)
104 172.61 ± 66.47 (171.5)
81 149.18 ± 90.06 (151)
p 0.110 0.180 0.015 0.082
Physical
Activity at
Work
No Physical Activity
at work 110 45.96 ± 27.48 (52.45)
103 210.59 ± 72.75 (214) 103 155 ± 68.15 (154) 97 128.32 ± 86.65 (129)
1—Work Requires
some degree of phys-
ical activity
79 52.59 ± 32.52 (59.7)
72 229.36 ± 67.94 (230) 72 170.49 ± 73.05 (172.5)
68 151.8 ± 100.36 (162.5)
p 0.950 0.111 0.114 0.030
Physical
Activity at
Home
No or Low 45 50.17 ± 32.65 (54.3)
42 222.33 ± 62.45 (219) 42 163.79 ± 63.24 (159.5)
39 138.11 ± 83.43 (138)
Long Duration 144 48.15 ± 28.99 (56.85)
133 216.81 ± 73.87 (219) 133 160.38 ± 72.87 (161) 126 137.33 ± 96.79 (139.5)
p 0.614 0.880 0.917 0.731
Work Style
No shift, or Day Shift
134 46.54 ± 28.67 (55.55)
123 214.37 ± 70.65 (215) 123 157.27 ± 67.68 (156) 114 132.77 ± 88.95 (133.5)

Vaccines 2022, 10, 1443 12 of 11
Mixed or Night
Shifts 53 54.19 ± 32.84 (56.9)
50 223.82 ± 70.95 (222.5)
50 167.36 ± 72.41 (181) 49 144.62 ± 99.61 (152)
p 0.175 0.425 0.287 0.195
Medication
No Medication 112 47.73 ± 28.39 (51.8)
104 223.21 ± 69.13 (221.5)
104 164.95 ± 69.9 (165.5) 95 139.93 ± 90.95 (146)
Taking Medication 74 49.56 ± 28.84 (58.95)
69 211.76 ± 74.72 (218) 69 157.76 ± 71.71 (154) 68 136.46 ± 98.09 (136)
p 0.429 0.379 0.597 0.877
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