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\begin{document}

\title{Omicron breakthrough infections in wild-type SARS-CoV-2 vaccinees elicit
high levels of neutralizing antibodies against pangolin coronavirus
GX\_P2V}



\author[1]{Lihua Song}%
\author[1]{Shanshan Lu}%
\author[2]{Shengdong Luo}%
\author[2]{Bingke Bai}%
\author[2]{Zhenping Fan}%
\author[1]{Bixia Hong}%
\author[2]{Wen Xu}%
\author[2]{Hongbin Ma}%
\author[2]{Wei-Wei Chen}%
\author[1]{Huahao Fan}%
\author[1]{Yigang Tong}%
\affil[1]{Beijing University of Chemical Technology College of Life Science and Technology}%
\affil[2]{5th Medical Center of Chinese PLA General Hospital}%


\vspace{-1em}



  
  \date{May 20, 2024}


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\selectlanguage{english}
\begin{abstract}
Omicron BF.7 became the predominant SARS-CoV-2 variant in Beijing after
the adjustment of COVID-19 response strategies in December 2022. The
ability of antibodies elicited by BF.7 infection to cross-react with
SARS-CoV-2-like viruses is unknown. This study aimed to investigate the
cross-reactive neutralizing antibodies against SARS-CoV-2-related
pangolin coronavirus GX\_P2V in sera from vaccinated and/or SARS-CoV-2
infected individuals. All vaccinated individuals who recovered from
Omicron BF.7 breakthrough infections exhibited substantially higher
levels of neutralizing antibodies against GX\_P2V among collected
subject populations (geometric mean titer {[}GMT{]} = 362). Uninfected
individuals who received four-mixed-dose vaccines also demonstrated
higher levels of neutralizing antibodies (GMT = 44) against GX\_P2V than
those who received two- or three-dose vaccines and those who recovered
from wild type SARS-CoV-2. This finding highlights the significance of
prior and hybrid booster vaccinations with wild-type SARS-CoV-2 vaccines
in generating potent cross-protective immunity against future spillovers
of SARS-CoV-2-like viruses.%
\end{abstract}%



\sloppy


\textbf{Omicron breakthrough infections in wild-type SARS-CoV-2
vaccinees elicit high levels of neutralizing antibodies against pangolin
coronavirus GX\_P2V}

Shanshan Lu\textsuperscript{1}, Shengdong Luo\textsuperscript{2}, Bingke
Bai\textsuperscript{2}, Zhenping Fan\textsuperscript{2}, Bixia
Hong\textsuperscript{1}, Wen Xu\textsuperscript{2}, Hongbin
Ma\textsuperscript{2}, Weiwei Chen\textsuperscript{2}, Huahao
Fan\textsuperscript{1}, Yigang Tong\textsuperscript{1}, Lihua
Song\textsuperscript{1}

\textsuperscript{1}Beijing Advanced Innovation Center for Soft Matter
Science and Engineering, College of Life Science and Technology, Beijing
University of Chemical Technology, Beijing,
China;\textsuperscript{2}Department of Infectious Diseases, Fifth
Medical Center of Chinese PLA General Hospital, Beijing, China

\textbf{Correspondence}

Lihua Song, Yigang Tong and Huahao Fan, Beijing Advanced Innovation
Center for Soft Matter Science and Engineering, College of Life Science
and Technology, Beijing University of Chemical Technology, Beijing,
China.

Email: Lihua Song, songlihua@gmail.com, Yigang Tong,
tong.yigang@gmail.com and fanhuahao@mail.buct.edu.cn

Shanshan Lu and Shengdong Luo are joint first authors.

\textbf{Funding information}

NSFC-MFST project (China--Mongolia) {[}grant number 32161143027{]},
Inner Mongolia Key Research and Development Program {[}grant number
2019ZD006{]}, Military Biosecurity Research Program {[}grant number
BWS21J025{]}, National Key R\&D Program of China (2021YFC2301804) and
the Biosafety Special Program (No. 19SWAQ 13).



\textbf{Abstract}

Omicron BF.7 became the predominant SARS-CoV-2 variant in Beijing after
the adjustment of COVID-19 response strategies in December 2022. The
ability of antibodies elicited by BF.7 infection to cross-react with
SARS-CoV-2-like viruses is unknown. This study aimed to investigate the
cross-reactive neutralizing antibodies against SARS-CoV-2-related
pangolin coronavirus GX\_P2V in sera from vaccinated and/or SARS-CoV-2
infected individuals. All vaccinated individuals who recovered from
Omicron BF.7 breakthrough infections exhibited substantially higher
levels of neutralizing antibodies against GX\_P2V among collected
subject populations (geometric mean titer {[}GMT{]} = 362). Uninfected
individuals who received four-mixed-dose vaccines also demonstrated
higher levels of neutralizing antibodies (GMT = 44) against GX\_P2V than
those who received two- or three-dose vaccines and those who recovered
from wild type SARS-CoV-2. This finding highlights the significance of
prior and hybrid booster vaccinations with wild-type SARS-CoV-2 vaccines
in generating potent cross-protective immunity against future spillovers
of SARS-CoV-2-like viruses.

\textbf{KEYWORDS}

COVID-19, Omicron BF.7, pangolin coronavirus GX\_P2V, cross-protective
immunity

\textbf{1 INTRODUCTION}

The coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute
respiratory syndrome coronavirus (SARS-CoV-2), has persisted for more
than three years\textsuperscript{1}, and numerous variants have emerged
globally. In early December 2022, China lifted its COVID-zero policy,
followed shortly by the quick spread of the SARS-CoV-2 Omicron variant
across most parts of the country.\textsuperscript{2} Although
vaccination with wild-type SARS-CoV-2 vaccines did not appear to provide
significant protection against Omicron in Beijing, most people only
experienced influenza-like symptoms. By February 2023, only sporadic
COVID cases were reported each day, indicating that herd immunity
against Omicron variants was achieved in a relatively short period in
China. However, concerns remain regarding the effectiveness of this
immunity against future spillovers of SARS-CoV-2-like viruses.

To address this issue, we analyzed the neutralizing antibodies in sera
from wild-type SARS-CoV-2 vaccine recipients who recovered from Omicron
BF.7 variant infection by performing viral neutralizing assays with a
SARS-CoV-2-related pangolin coronavirus GX\_P2V\textsuperscript{3}. The
differences between SARS-CoV-2 and GX\_P2V primarily lie in the spike
glycoprotein (S), a primary target for neutralizing antibody (Figure
S1). However, compared to SARS-CoV-2, the spike protein of pangolin-CoV
GX\_P2V has a comparable binding affinity to human ACE2 but has a
distinct receptor binding domain (RBD)\textsuperscript{4} (Figure 1A),
making it a practical model for testing neutralizing antibodies against
unpredictable SARS-CoV-2 related viruses.

\textbf{2 MATERIALS AND METHODS}

\textbf{2.1 Cell culture}

BGM cells purchased from the American Type Culture Collection (ATCC,
USA) were maintained in Roswell Park Memorial Institute (RPMI) 1640
medium (Gibco, USA) supplemented with 10\% fetal bovine serum (FBS,
Gibco, USA) and 1\% penicillin/streptomycin (Gibco, USA) at 37\selectlanguage{ngerman}°C in a
5\% CO2 incubator. Mycoplasma testing was performed, and cell cultures
were confirmed to be negative.

\textbf{2.2 Patients and sample collection}

Serum samples were collected from the subsequent cohorts: healthy
individuals uninfected prior to the global COVID pandemic; convalescents
of wild-type SARS-CoV-2 with qPCR-negative results 2-8 weeks
post-diagnosis in 2020; recipients of two-dose inactivated vaccines (2-3
weeks post-second vaccination), three-dose inactivated vaccines (3-5
months post-third vaccination), and four-mixed-dose vaccines (three-dose
inactivated plus one-dose of alternative vaccine types, 2-4 weeks
post-fourth vaccination). These vaccinated participants had no history
of SARS-CoV-2 infection, as confirmed by regular qPCR screening tests.
Serum samples from both vaccinated and unvaccinated individuals infected
with the Omicron BF.7 variant were obtained when the subjects were
tested qPCR-negative 2-9 weeks and 3-5 weeks post-diagnosis,
respectively. Sera from golden hamsters infected twice with GX\_P2V
served as positive controls.

\textbf{2.3 Neutralizing antibody assays}

Serum samples from participants were initially 10-fold diluted and
subsequently 3-fold serially diluted. The diluted serum samples were
then combined with an equal volume of GX\_P2V (100
TCID\textsubscript{50}, GenBank accession number MW532698) incubated at
37\selectlanguage{english} in a 5\% CO\textsubscript{2} incubator for 2 hours. Following
incubation, 100 \selectlanguage{greek}µ\selectlanguage{english}L of the virus-serum mixture was transferred into
96-well plates and combined with an equal volume of BGM cells
(3\selectlanguage{ngerman}×10\textsuperscript{4} cells/well). The inoculated plates were
incubated continuously at 37\selectlanguage{english} in a 5\% CO\textsubscript{2} incubator for
an additional 5 days, after which the cytopathic effect (CPE) of the
virus were observed using a light microscope. GX\_P2V neutralization
titers were calculated using the Reed-Muench equation and expressed as
the sample dilution at which a 50\% neutralization titer
(NT\textsubscript{50}) was achieved.

\textbf{2.4 Statistical analysis}

Statistical analyses were performed using GraphPad Prism 9 software. The
differences between two groups were analyzed by Student's t-test (paired
two-tailed). P-values less than 0.05 (P\textless{} 0.05) were considered
statistically significant.

\textbf{3 RESULTS AND DISCUSSION}

A total of 52 infected patients and 32 uninfected population
participated in this study. Descriptive characteristics for the study
population are presented in Table 1 and Table 2. We assayed the
neutralizing activity against pangolin-CoV GX\_P2V in serum samples
obtained from 27 participants who had been vaccinated and boosted but
recovered from SARS-CoV-2 BF.7 variant. Sera from patients who did not
receive any vaccination but recovered from wild-type SARS-CoV-2 strain
or Omicron infections, and sera from individuals who received two, three
and four mixed vaccinations but had no history of SARS-CoV-2 infection,
were used as controls. Sera from GX\_P2V-infected golden hamsters were
used as positive controls.

The geometric mean titers (GMTs) of neutralizing antibodies against
GX\_P2V in each group are presented in Table S1 and Figure 1B. Notably,
regardless of the vaccine types, all vaccinated individuals who
recovered from Omicron BF.7 infections had substantially higher GMTs of
neutralizing antibodies against GX\_P2V compared to cohorts of wild-type
SARS-CoV-2 convalescents, uninfected two-dose vaccinees, uninfected
three-dose vaccinees, uninfected four-mixed-dose vaccinees, and
unvaccinated Omicron convalescents by factors of 45.3, 27.8, 60.3, 8.2,
and 12.9, respectively (Figure 1B). Thus, Omicron breakthrough
infections in wild-type SARS-CoV-2 vaccinees elicited superior
cross-neutralizing antibodies against GX\_P2V, a SARS-CoV-2 related
virus.

Moreover, compared with unvaccinated individuals recovered from the
Omicron BF.7 variant infection, the GMT was lower by a factor of 3.5,
2.2, 4.7 in wild-type SARS-CoV-2 convalescents, uninfected recipients of
two-dose and three-dose inactivated vaccines, respectively. The GMT of
uninfected participants but with four-mixed-dose vaccines was also
higher by a factor of 5.5, 3.4, 7.3 than in wild-type SARS-CoV-2
convalescents, uninfected recipients of two-dose and three-dose
inactivated vaccines, respectively. The results showed the effectiveness
of mixed booster vaccinations and hybrid immunity protection against
SARS-CoV-2-like virus infection.

In conclusion, our finding highlights the significance of prior
vaccinations with wild-type SARS-CoV-2 vaccines in generating potent
cross-protective immunity against breakthrough infections and supports
reports of pan-SARS-CoV-2 neutralizing epitopes in the wild-type
SARS-CoV-2 RBD.\textsuperscript{5,6} Additionally, our results further
support GX\_P2V as a promising live vaccine candidate against
SARS-CoV-2.\textsuperscript{7} Overall, our study suggests that the
general population in China has likely developed superior immunity to
prevent future breaks of SARS-CoV-2-like viruses after the Omicron
epidemic. Further research is necessary to corroborate these findings
and to evaluate the long-term efficacy of such immunity in the
population.

\textbf{ACKNOWLEDGEMENTS}

We thank the reviewers for critical review of the manuscript. The
funding sources had no role in the study design, data collection,
analysis, interpretation, or writing of the report.

\textbf{AUTHOR CONTRIBUTIONS}

\emph{Study concept and design:} Lihua Song, Yigang Tong; \emph{Data
acquisition} : Shanshan Lu, Shengdong Luo, Wen Xu, Bixia Hong, Wen Xu,
Hongbin Ma; \emph{Analysis and interpretation of data} : Huahao Fan,
Bingke Bai, Zhenping Fan, Hongbin Ma, Weiwei Chen; \selectlanguage{english}\emph{Manuscript
write-up} : Lihua Song, Shanshan Lu. Revision and approval of the final
draft by all authors

\textbf{CONFLICT OF INTERESTS}

No potential conflict of interest was reported by the authors.

\textbf{Ethics statement}

This study was approved by the ethics committee (EC) of the Fifth
Medical Center, General Hospital of Chinese PLA (approval number:
2020027D).

\textbf{DATA AVAILABILITY STATEMENT}

The data in this study are available from the corresponding author upon
reasonable request.

\textbf{REFERENCES}

1. Zhu N, Zhang D, Wang W, et al. A Novel Coronavirus from Patients with
Pneumonia in China, 2019. N Engl J Med. 2020;382(8):727-733.

2. Pan Y, Wang L, Feng Z, et al. Characterisation of SARS-CoV-2 variants
in Beijing during 2022: an epidemiological and phylogenetic analysis.
Lancet. 2023;401(10377):664-672.

3. Lam TT, Jia N, Zhang YW, et al. Identifying SARS-CoV-2-related
coronaviruses in Malayan pangolins. Nature. 2020;583(7815):282-285.

4. Zhang S, Qiao S, Yu J, et al. Bat and pangolin coronavirus spike
glycoprotein structures provide insights into SARS-CoV-2 evolution. Nat
Commun. 2021;12(1):1607.

5. Liu Z, Zhou J, Wang X, et al. A pan-sarbecovirus vaccine based on RBD
of SARS-CoV-2 original strain elicits potent neutralizing antibodies
against XBB in non-human primates. Proc Natl Acad Sci U S A.
2023;120(11):e2221713120.

6. Qi H, Liu B, Wang X, Zhang L. The humoral response and antibodies
against SARS-CoV-2 infection. Nat Immunol. 2022;23(7):1008-1020.

7. Lu S, Luo S, Liu C, et al. Induction of significant neutralizing
antibodies against SARS-CoV-2 by a highly attenuated pangolin
coronavirus variant with a 104nt deletion at the 3'-UTR. Emerg Microbes
Infect. 2023;12(1):2151383.\selectlanguage{english}
\begin{figure}[H]
\begin{center}
\includegraphics[width=0.70\columnwidth]{figures/image1/image1}

\end{center}
\end{figure}

\textbf{FIGURE 1} Neutralizing antibody responses to pangolin
coronavirus GX\_P2V.

(A) Amino acids sequences of the GX\_P2V and SARS-CoV-2 variants were
aligned based on the whole spike protein, S1, S2, and RBD. The percent
identities of the aligned sequences were calculated, and a heatmap was
generated using GraphPad Prism 9 with SARS-CoV-2 as the reference
sequence. (B) Neutralizing antibody responses against pangolin
coronavirus GX\_P2V were assessed and expressed as neutralizing titer 50
(NT\textsubscript{50}). The numbers over the different colors of dots in
each group represent geometric mean titers (GMTs). GMTs for each group
were displayed as horizontal black lines. Ratios between GMTs of two
different groups are indicated. Individual samples of various groups are
represented by different colors of dots.

\textbf{TABLE 1} Characteristics of the infected population\selectlanguage{english}
\begin{longtable}[]{@{}llll@{}}
\toprule
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\textbf{Variable}\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\textbf{Wild-type SARS-CoV-2 convalescents}\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\textbf{Unvaccinated Omicron convalescents}\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\textbf{Vaccinated Omicron convalescents}\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
Number of participants\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
20\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
5\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
27\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\textbf{Age} (years), median (range)\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
53 (17-79)\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
40 (34-43)\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
29 (25-54)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\textbf{Sex at birth}, female\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
10 (50\%)\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
3 (60\%)\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
19 (70\%)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\textbf{COVID vaccination history}\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
Two-dose inactivated vaccines\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
N/A\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
1 (3.7\%)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
Three-dose inactivated vaccines\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
N/A\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
18 (66.7\%)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.24\columnwidth}\raggedright\strut
Two-dose Ad5-nCoV (adenovirus vector vaccine)\strut
\end{minipage} & \begin{minipage}[t]{0.24\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.24\columnwidth}\raggedright\strut
N/A\strut
\end{minipage} & \begin{minipage}[t]{0.24\columnwidth}\raggedright\strut
6 (22.2\%)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.24\columnwidth}\raggedright\strut
Three-dose Recombinant Fusion Protein vaccines\strut
\end{minipage} & \begin{minipage}[t]{0.24\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.24\columnwidth}\raggedright\strut
N/A\strut
\end{minipage} & \begin{minipage}[t]{0.24\columnwidth}\raggedright\strut
1 (3.7\%)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.24\columnwidth}\raggedright\strut
Two-dose inactivated vaccines plus one-dose Ad5-nCoV\strut
\end{minipage} & \begin{minipage}[t]{0.24\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.24\columnwidth}\raggedright\strut
N/A\strut
\end{minipage} & \begin{minipage}[t]{0.24\columnwidth}\raggedright\strut
1 (3.7\%)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\textbf{COVID disease severity}\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
Asymptomatic\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
1 (20\%)\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
0\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
Mild\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
4 (20\%)\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
4 (80\%)\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
19 (70\%)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
Moderate\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
9 (45\%)\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
8 (30\%)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
Severe\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
4 (20\%)\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
0\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
Critical\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
3 (15\%)\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
0\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\textbf{Time between last vaccination and confirmed infection}\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
N/A\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
N/A\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
6-22 months\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\textbf{Time between infection and serum sampling when SARS-CoV-2
negative}\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
2-8 weeks\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
3-5 weeks\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
2-9 weeks\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\textbf{Method of SARS-CoV-2 testing}\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
Quantitative PCR\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
20 (100\%)\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
1 (20\%)\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
5 (19\%)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
Rapid antigen detection\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
4 (80\%)\strut
\end{minipage} & \begin{minipage}[t]{0.22\columnwidth}\raggedright\strut
22 (81\%)\strut
\end{minipage}\tabularnewline
\bottomrule
\end{longtable}

\emph{Note:} Data are median (range), or n (\%).

\textbf{TABLE 2} Characteristics of the uninfected population\selectlanguage{english}
\begin{longtable}[]{@{}lllll@{}}
\toprule
\begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\textbf{Variable}\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\textbf{Healthy individuals}\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\textbf{Uninfected two-dose vaccinees}\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\textbf{Uninfected three-dose vaccinees}\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\textbf{Uninfected four-mixed-dose vaccinees}\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
Number of participants\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
10\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
8\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
10\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
4\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\textbf{Age} (years), median (range)\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
41 (33-63)\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
35 (32-50)\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
/\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
31 (28-51)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\textbf{Sex at birth}, female\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
7 (70\%)\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
7 (88\%)\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
/\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
2 (50\%)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\textbf{COVID vaccination history}\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
Inactivated vaccines\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
8 (100\%)\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
10 (100\%)\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
0\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
Ad5-nCOV (adenovirus vector vaccine)\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
0\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
dNS1-RBD\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
0\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
Recombinant Fusion Protein\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
0\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Three-dose inactivated vaccines plus one-dose dNS1-RBD\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
1 (25\%)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Three-dose inactivated vaccines plus one-dose Ad5-nCOV\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
2 (50\%)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Three-dose inactivated vaccines plus one-dose Recombinant Fusion
Protein\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
0\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
1 (25\%)\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\textbf{Time between last vaccination and serum sampling}\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
N/A\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
2-3 weeks\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
3-5 months\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
2-4 weeks\strut
\end{minipage}\tabularnewline
\bottomrule
\end{longtable}

\emph{Note:} Data are median (range), or n (\%).

\selectlanguage{english}
\FloatBarrier
\end{document}

