SARS-CoV-2 Seroprevalence Among Parturient Women

Abstract

Limited data are available for pregnant women affected by SARS-CoV-2. Serological tests are critically important to determine exposure and immunity to SARS-CoV-2 within both individuals and populations. We completed SARS-CoV-2 serological testing of 1,293 parturient women at two centers in Philadelphia from April 4 to June 3, 2020. We tested 834 pre-pandemic samples collected in 2019 and 15 samples from COVID-19 recovered donors to validate our assay, which has a ∼1% false positive rate. We found 80/1,293 (6.2%) of parturient women possessed IgG and/or IgM SARS-CoV-2-specific antibodies. We found race/ethnicity differences in seroprevalence rates, with higher rates in Black/non-Hispanic and Hispanic/Latino women. Of the 72 seropositive women who also received nasopharyngeal polymerase chain reaction testing during pregnancy, 46 (64%) were positive. Continued serologic surveillance among pregnant women may inform perinatal clinical practices and can potentially be used to estimate seroprevalence within the community.

One Sentence Summary
Six percent of pregnant women delivering from April 4 to June 3, 2020 had serological evidence of exposure to SARS-CoV-2 with notable race/ethnicity differences in seroprevalence rates.

Full text

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SARS-CoV-2 Seroprevalence Among Parturient Women
Authors: Dustin D. Flannery1,2,3,13; Sigrid Gouma4,13; Miren B. Dhudasia1,3; Sagori
Mukhopadhyay1,2,3; Madeline R. Pfeifer1; Emily C. Woodford1; Jeffrey S. Gerber2,3,5; Claudia P.
Arevalo4; Marcus J. Bolton4; Madison E. Weirick4; Eileen C. Goodwin4; Elizabeth M.
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Anderson4; Allison R. Greenplate6,7; Justin Kim6,7; Nicholas Han6,7; Ajinkya Pattekar6,8; Jeanette
Dougherty6,7; Oliva Kuthuru6,7; Divij Mathew6,7; Amy E. Baxter6,7; Laura A. Vella5,6,7; JoEllen
Weaver9; Anurag Verma10; Rita Leite11; Jeffrey S. Morris12; Daniel J. Rader9,10; Michal A.
Elovitz6,11; E. John Wherry6,7; Karen M. Puopolo1,2,3*; Scott E. Hensley4,6*
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Affiliations:
1 Division of Neonatology, Children’s Hospital of Philadelphia, Philadelphia, PA.
2 Department of Pediatrics, University of Pennsylvania Perelman School of Medicine,
Philadelphia, PA.
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3 Center for Pediatric Clinical Effectiveness, Children’s Hospital of Philadelphia, Philadelphia,
PA.
4 Department of Microbiology, University of Pennsylvania Perelman School of Medicine,
Philadelphia, PA.
5 Division of Infectious Diseases, Children’s Hospital of Philadelphia, Philadelphia, PA.
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6 Institute for Immunology, University of Pennsylvania Perelman School of Medicine,
Philadelphia, PA.
7 Department of Systems Pharmacology and Translational Therapeutics, University of
Pennsylvania, Philadelphia, PA.
8 Division of Gastroenterology, Department of Medicine, University of Pennsylvania Perelman
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School of Medicine, Philadelphia, PA.
9 Institute for Translational Medicine and Therapeutics, University of Pennsylvania Perelman
School of Medicine, Philadelphia, PA.
10 Departments of Genetics and Medicine, Perelman School of Medicine, University of
Pennsylvania, Philadelphia, PA.
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11Maternal and Child Health Research Center, Department of Obstetrics and Gynecology,
University of Pennsylvania Perelman School of Medicine, Philadelphia, PA.
12 Department of Biostatistics Epidemiology and Informatics, University of Pennsylvania,
Philadelphia, PA.
† These authors contributed equally: D.D. Flannery and S. Gouma.
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*Correspondence to: hensley@pennmedicine.upenn.edu and
karen.puopolo@pennmedicine.upenn.edu
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Abstract:
Limited data are available for pregnant women affected by SARS-CoV-2. Serological tests are
critically important to determine exposure and immunity to SARS-CoV-2 within both
individuals and populations. We completed SARS-CoV-2 serological testing of 1,293 parturient
women at two centers in Philadelphia from April 4 to June 3, 2020. We tested 834 pre-pandemic
5
samples collected in 2019 and 15 samples from COVID-19 recovered donors to validate our
assay, which has a ~1% false positive rate. We found 80/1,293 (6.2%) of parturient women
possessed IgG and/or IgM SARS-CoV-2-specific antibodies. We found race/ethnicity differences
in seroprevalence rates, with higher rates in Black/non-Hispanic and Hispanic/Latino women. Of
the 72 seropositive women who also received nasopharyngeal polymerase chain reaction testing
10
during pregnancy, 46 (64%) were positive. Continued serologic surveillance among pregnant
women may inform perinatal clinical practices and can potentially be used to estimate
seroprevalence within the community.
15
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is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted July 10, 2020. .https://doi.org/10.1101/2020.07.08.20149179doi: medRxiv preprint

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One Sentence Summary: Six percent of pregnant women delivering from April 4 to June 3,
2020 had serological evidence of exposure to SARS-CoV-2 with notable race/ethnicity
differences in seroprevalence rates.
5
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is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted July 10, 2020. .https://doi.org/10.1101/2020.07.08.20149179doi: medRxiv preprint

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Main Text:
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can cause serious
disease in adult populations, particularly in those with underlying health conditions (1). SARS-
CoV-2 serological tests are important for determining immunity within individuals and
populations (2). However, many commercial tests have high false positive rates and therefore
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cannot be used to accurately estimate seroprevalence in populations with relatively low levels of
exposures (3,4). Serological tests are especially important for vulnerable populations such as
pregnant women, because immune status has implications for management of both the pregnant
woman and the newborn. Admission to the hospital for delivery is one of the few instances in
which otherwise healthy individuals are consistently interacting with the medical system, and
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therefore provides an opportunity for population surveillance of SARS-CoV-2 serology.
We performed a prospective cohort study of pregnant women presenting for delivery
from April 4 to June 3, 2020 at two academic birth hospitals in Philadelphia, Pennsylvania. Both
hospitals are active clinical and research centers affiliated with the University of Pennsylvania,
and combined represent 50% of live births in Philadelphia (5). Discarded maternal sera from
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delivery admission were collected, deidentified, and tested by enzyme-linked immunosorbent
assay (ELISA) for SARS-CoV-2 immunoglobulin G (IgG) and immunoglobulin M (IgM)
antibodies to the spike receptor binding domain (RBD) antigen.
Demographics and clinical characteristics of the women are shown in Table 1. Most
serum specimens were derived from women living in areas within or immediately bordering the
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city of Philadelphia (Figure 1). Symptomatic pregnant women and those with known risk factors
underwent SARS-CoV-2 nasopharyngeal (NP) nucleic acid polymerase chain reaction (PCR)
testing from April 4-12, 2020; universal PCR testing was recommended for all pregnant women
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presenting for delivery starting April 13, 2020. Of 1,620 women who delivered during the study
period, 1,293 (80%) had available discarded serum specimens and were included in the analysis.
Our serological assay utilized a SARS-CoV-2 spike RBD antigen and modified ELISA
protocol first described by Amanat et al. (6). We validated this serological assay by testing
serum samples collected prior to the pandemic in 2019 from 834 individuals in the Penn
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Medicine Biobank and 15 individuals who recovered from confirmed coronavirus disease 19
(COVID-19) infections in 2020 (Figure 2A-B). All 15 serum samples from COVID-19
recovered donors contained high, but variable, levels of SARS-CoV-2 IgG (Figure 2A) and 10
of 15 samples contained detectable levels of SARS-CoV-2 IgM (Figure 2B). Conversely, only 5
of 834 samples collected before the pandemic contained SARS-CoV-2 IgG and only 4 of 834
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samples contained SARS-CoV-2 IgM; none contained both IgG and IgM. Together, this
indicates that there is an overall false positive rate ~1% (9/834) in our serological assay.
Consistent with our initial validation experiments, only 1 of 140 samples collected from pregnant
women before the pandemic (from 2009-2012) possessed IgG or IgM SARS-CoV-2 antibodies
(Figure 2C-D).
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We found that 80 of 1,293 (6.2%) pregnant women presenting for delivery from April 4
to June 3, 2020 possessed IgG or IgM SARS-CoV-2 antibodies (Figure 2C-D; p = 0.003
comparing samples from pre-pandemic and pandemic pregnant women). We identified 55
women with both SARS-CoV-2 IgG and IgM, 21 women with only SARS-CoV-2 IgG, and 4
women with only SARS-CoV-2 IgM (Table 2). SARS-CoV-2 antibody levels in samples from
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these women were variable (Figure 2C-D), similar to what we found in samples from
individuals recovering from confirmed SARS-CoV-2 infections (Figure 2A-B). The
seroprevalence rate was not statistically different comparing women living within the city limits
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of Philadelphia (62/986, 6.3%) to those living in surrounding areas in Pennsylvania (12/191,
6.3%), or surrounding areas in New Jersey (5/107, 4.7%). In contrast, we observed significant
race/ethnicity differences in seroprevalence rates with higher rates in Black/non-Hispanic (9.7%)
and Hispanic/Latino (10.4%) women and lower rates in White/non-Hispanic (2.0%) and Asian
(0.9%) women (Table 1).
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NP swabs from 1,109 (85.8%) women were tested by SARS-CoV-2 PCR during the
pregnancy or at the time of delivery. We found that 46 of 72 seropositive women who were NP
tested had a SARS-CoV-2 positive PCR result, whereas only 18 of 1,037 seronegative women
who were NP tested had a SARS-CoV-2 positive PCR result (Table 1; p <0.001). While all
serum samples were collected during the delivery admission, NP samples were collected at
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variable times either during the delivery admission or earlier in the pregnancy, and therefore,
further study will be required to evaluate the temporal relationship between SARS-CoV-2
seropositivity and PCR positivity in pregnant women.
Large-scale serology testing is critical for estimating how many individuals have been
infected during the COVID-19 pandemic. Due to widely-imposed social distancing requirements,
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and to decreases in on-site, discretionary medical care, it is currently difficult to collect serum for
population-wide serological testing. The vast majority of pregnant women, however, continue to
have multiple interactions with the medical system for prenatal care and for delivery during this
pandemic, and therefore represent a unique population to assess SARS-CoV-2 immunity within a
community. Our data suggest that ~5.2% (6.2% minus ~1% false positive rate) of parturient
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women in Philadelphia from April 4 to June 3, 2020 were previously exposed to SARS-CoV-2.
As of June 3, 2020, there were 23,160 confirmed cases of COVID-19 in the city of Philadelphia
(7), which has a population size of nearly 1.6 million people, suggesting an infection rate of
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approximately 1.4%. Serologic studies may provide a more accurate means of assessing
population exposure to SARS-CoV-2 by identifying asymptomatic or minimally symptomatic as
well as symptomatic infections. Further studies are needed to determine how the immune status
of pregnant women compares to that of the general population. For example, parturient women
may not represent individuals of different ages within the general population and women and
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men might mount different antibody responses upon infection with SARS-CoV-2 (8).
Pregnant women of all demographics continue to seek medical care during the COVID-
19 pandemic. Therefore, parturient women represent a unique population to assess differences in
SARS-CoV-2 exposures in diverse populations. Our finding that Black/non-Hispanic and
Hispanic/Latino women have higher SARS-CoV-2 seroprevalence rates relative to women of
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other races suggest that there are race/ethnicity differences in SARS-CoV-2 exposures in
Philadelphia and surrounding areas. Identification of factors that contribute to such differences in
exposure to SARS-CoV-2 may inform public health measures aimed at preventing further
infections.
Prior perinatal COVID-19 studies have primarily focused on virus detection (i.e. nucleic
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acid testing) in pregnant women and have not evaluated immunity (9–17). Two published studies
to date have assessed SARS-COV-2 serology in pregnant women with active disease. A study of
6 parturient women in Wuhan, China with confirmed COVID-19 found all 6 women had
elevated levels of SARS-CoV-2 IgG and IgM (18). A case report from Peru detailed a
symptomatic pregnant woman with positive PCR testing and negative serology at presentation,
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who developed severe respiratory failure necessitating delivery; her IgM and IgG turned positive
4 days after delivery (9 days after symptom onset) (19). Beyond describing individual response
to infection, SARS-CoV-2 serological testing among pregnant women will be increasingly
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important for perinatal disease risk management, as well as for optimizing vaccine strategies
when vaccines become available. Additional studies will be needed to address the impact of
maternal infection on neonatal immunity, and to determine those factors that may contribute to
observed disparities in exposure to SARS-CoV-2.
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Acknowledgments: We thank all members of the Wherry Lab and the Penn COVID-19 Sample
Processing Unit for sample procurement, processing and logistics. We thank the staff of the Penn
Medicine Biobank. We thank Florian Krammer (Mt. Sinai) for sending us the SARS-CoV-2
spike RBD expression plasmids. We thank Dr. Steven Melly (Drexel University) for his
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assistance in geographic analyses.
Funding: This work was supported by institutional funds from the University of Pennsylvania
and an NIH grant AI082630 (to E.J.W.). We thank Jeffrey Lurie and we thank Joel Embiid, Josh
Harris, David Blitzer for philanthropic support. E.J.W. is supported by the Parker Institute for
Cancer Immunotherapy which supports the cancer immunology program at UPenn.
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Author contributions: DDF conceptualized and designed the study, collected data, drafted the
initial manuscript, and revised the manuscript. SG led the serological experiments, collected
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data, and revised the manuscript. MBD designed the data collection instruments, collected data,
carried out the analyses, and revised the manuscript. SM conceptualized and designed the study,
designed the data collection instruments, carried out the analyses, and revised the manuscript.
MRP collected data and revised the manuscript. ECW collected data and revised the manuscript.
JSG conceptualized and designed the study, and revised the manuscript. CPA completed
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serological assays, analyzed data, and revised the manuscript. MJB completed serological assays,
analyzed data, and revised the manuscript. MW completed serological assays, analyzed data, and
revised the manuscript. ECG completed serological assays, analyzed data, and revised the
manuscript. EMA completed serological assays, analyzed data, and revised the manuscript. ARG
obtained and proceeded samples from recovered donors. JK obtained and proceeded samples
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from recovered donors. NH obtained and proceeded samples from recovered donors. AP
obtained and proceeded samples from recovered donors. JD obtained and proceeded samples
from recovered donors. OK designed and established recovered donor cohort. DM processed and
characterized samples from recovered donors. AB oversaw acquisition, processing, and
characterization of samples from recovered donors. LAV designed and established recovered
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donor cohort. JW supervised recruitment of participants in PMBB and identification of samples
for serology testing. AV analyzed demographic data of PMBB participants. RL provided samples
for the pre pandemic pregnant controls. JSM provided statistical advice, performed statistical
analyses, and revised the paper. DJR provided input on the use of PMBB controls and revised the
manuscript. MAE provided input and samples for the pre-pandemic pregnant controls and
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revised the manuscript. EJW designed, established, and oversaw healthy donor cohort studies
and made revisions to the manuscript. KMP conceptualized and designed the study, coordinated
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and supervised data collection, and revised the manuscript. SEH conceptualized and designed the
study, coordinated and supervised serological studies, and revised the manuscript.
Competing interests: SEH has received consultancy fee from Sanofi Pasteur, Lumen, Novavax,
and Merck for work unrelated to this report. EJW is a member of the Parker Institute for Cancer
Immunotherapy. EJW has consulting agreements with and/or is on the scientific advisory board
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for Merck, Roche, Pieris, Elstar, and Surface Oncology. EJW is a founder of Surface Oncology
and Arsenal Biosciences. EJW has a patent licensing agreement on the PD-1 pathway with
Roche/Genentech. All other authors declare no competing interests related to this work.
Data and materials availability: All data are included in the manuscript.
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Supplementary Materials:
Materials and Methods
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Table 1. Demographics and Clinical Characteristics of the Study Cohort
Characteristics
Total
(n = 1,293)
Seropositive1
(n = 80)
Seronegative
(n = 1,213)
p-
value2
Age (in years), median (IQR)
31 (27, 35)
28 (24, 32)
31 (27, 35)
<0.001
Race, n (%)
Black/Non-Hispanic
537 (41.5)
52 (65.0)
485 (40.0)
<0.001
White/Non-Hispanic
447 (34.6)
9 (11.3)
438 (36.1)
<0.001
Hispanic/Latino
125 (9.7)
13 (16.2)
112 (9.2)
0.04
Asian
106 (8.2)
1 (1.3)
105 (8.7)
0.01
Other/Unknown3
78 (6.0)
5 (6.2)
73 (6.0)
0.93
Health insurance4, n (%)
<0.001
Private payor
727 (56.2)
26 (33.3)
701 (57.8)
Public payor
561 (43.4)
52 (66.7)
509 (42.0)
Uninsured
3 (0.2)
0
3 (0.2)
Pre-pregnancy BMI5, n (%)
Overweight (25.0 to <30.0)
345 (26.7)
28 (35.9)
317 (26.4)
0.07
Obese (≥30.0)
337 (26.1)
27 (34.6)
310 (25.8)
0.09
Diabetes6, n (%)
113 (8.7)
10 (12.5)
103 (8.5)
0.22
Hypertension6, n (%)
404 (31.3)
33 (41.3)
371 (30.6)
0.05
Asthma6, n (%)
194 (15.0)
13 (16.3)
181 (14.9)
0.75
Cesarean delivery, n (%)
400 (30.9)
30 (37.5)
370 (30.5)
0.19
Preterm delivery at gestational
age <37 weeks, n (%)
128 (9.9)
11 (13.8)
117 (9.7)
0.23
SARS-CoV-2 PCR tested during
pregnancy7, n (%)
1,109 (85.8)
72 (90.0)
1,037 (85.5)
0.26
Positive, n (% tested)
64/1,109 (5.8)
46/72 (63.9)
18/1,037 (1.7)
<0.001
Duration (in hours) between
serology test and first positive
PCR test8, median (IQR)
0.3
(0, 345.3)
75.4
(0, 659.3)
0
(-0.1, 0.2)
0.007
Live-born infant, n (%)
1,282 (99.2)
79 (98.8)
1,203 (99.2)
0.51
Footnotes: 1Seropositivity was based on either IgG or IgM level >0.48 arbitrary units.
2Difference in maternal age was tested using Mann-Whitney U test, differences in proportion for
5
all other characteristics were tested using χ2 tests or Fisher’s exact tests as appropriate. For
race/ethnicity and pre-pregnancy BMI, difference was tested at each level of the characteristic
(e.g. Black women compared to non-Black women). 3Race/ethnicity was unknown for 2
seropositive and 26 seronegative women; race was abstracted from documentation at time of
admission and in clinical practice is usually self-reported. 4Health insurance was missing for 2
10
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15
seropositive women; health insurance was abstracted from documentation in the medical record
of the insurance used during admission for delivery; ‘Public payor’ includes Medicaid, Medicare
and Veterans Affairs Community Care; ‘Uninsured’ includes self-pay and Charity Care. 5Pre-
pregnancy BMI was missing for 2 seropositive and 12 seronegative women; pre-pregnancy BMI
was abstracted from documentation in the medical record or patient’s self-reported entry in birth
5
registration. 6Diagnoses were based on delivery admission International Classification of
Diseases, 10th revision diagnosis codes for diabetes (O24, E08-E13, Z79.4), hypertension (O10,
O11, O13-O16, I10-I13, I15) and asthma (J45). 7Included tests done anytime during pregnancy
up to discharge from delivery admission. 8Difference calculated by subtracting the date and time
sample for PCR was collected from the date and time sample for serology was collected. A
10
negative number indicates PCR sample collected after serology sample. BMI, body mass index;
IQR, interquartile range; PCR, polymerase chain reaction; SARS-CoV-2, severe acute
respiratory syndrome coronavirus 2.
15
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16
Table 2. Relative levels of SARS-COV-2 IgG and IgM in serum collected from seropositive
pregnant women (n = 80).
Patient
Number
IgG levels
(arbitrary
units)
IgM levels
(arbitrary
units)
1
<0.20
0.57
2
<0.20
0.99
3
0.29
0.75
4
0.31
1.09
5
0.50
0.21
6
0.50
1.04
7
0.50
2.28
8
0.54
0.35
9
0.60
0.65
10
0.63
<0.20
11
0.66
1.01
12
0.75
<0.20
13
0.83
<0.20
14
0.86
<0.20
15
0.97
0.43
16
1.07
0.49
17
1.12
<0.20
18
1.70
0.56
19
1.75
0.69
20
1.85
0.33
21
1.87
0.29
22
1.97
<0.20
23
2.16
0.30
24
2.34
0.36
25
2.35
1.78
26
2.45
1.44
27
2.58
0.98
28
2.73
0.30
29
2.75
2.22
30
2.85
0.91
31
2.94
0.39
32
2.99
2.90
33
3.12
1.10
34
3.20
0.28
35
3.32
0.21
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17
36
3.34
1.11
37
3.38
1.40
38
3.42
0.58
39
3.46
<0.20
40
3.54
<0.20
41
3.55
1.40
42
4.40
<0.20
43
4.75
0.96
44
4.88
0.94
45
5.25
3.37
46
5.43
2.41
47
5.46
1.77
48
5.50
12.25
49
5.59
3.43
50
5.87
6.52
51
5.97
3.28
52
6.91
6.44
53
6.94
<0.20
54
7.40
3.27
55
8.60
3.71
56
8.67
0.98
57
9.56
12.60
58
9.59
1.26
59
9.80
1.32
60
10.23
4.40
61
11.28
25.33
62
11.47
0.81
63
11.52
7.90
64
12.45
4.63
65
13.97
1.45
66
14.44
1.56
67
14.46
8.29
68
15.99
2.64
69
18.10
0.63
70
18.76
3.99
71
19.56
1.82
72
20.47
19.70
73
22.32
3.32
74
22.52
8.49
75
31.26
3.67
76
32.85
22.02
77
44.26
17.59
78
44.46
3.27
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18
79
83.77
8.07
80
99.10
11.87
5
10
15
20
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19
Figure Legends
Figure 1. Geographical distribution of women tested for SARS-CoV-2 antibodies. Most
serum specimens analyzed were from women living in areas within or immediately bordering the
city of Philadelphia. Location of birth hospitals where serum samples were collected are shown
as red crosses.
5
Figure 2. Serum SARS-CoV-2 antibody levels in COVID-19 pandemic and pre-pandemic
individuals. (A-B) Relative levels of SARS-CoV-2 IgG (A) and IgM (A) in serum collected
before the COVID-19 pandemic (n = 834) and serum collected from COVID-19 recovered
donors (n = 15). (C-D) Relative levels of SARS-CoV-2 IgG (C) and IgM (D) in serum collected
10
from pregnant women from 2009-2012 (n = 140) and from April 4-June 3, 2020 (n = 1,293).
Dashed lines indicate 0.48 arbitrary units, which was used to distinguish positive versus negative
samples (see Methods). Serum samples that were below the cutoff for seropositivity were
assigned an antibody level of 0.40 arbitrary units.
15
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Figure 1
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A
Figure 2
B
C D
pregnant women
pre-pandemic COVID-19+
(n=834) (n=15)
pre-pandemic COVID-19+
(n=834) (n=15)
pre-pandemic pandemic
(n=140) (n=1,293)
pregnant women
pre-pandemic pandemic
(n=140) (n=1,293)
0.5
1
2
4
8
16
32
64
IgG levels
(arbitrary units)
SARS-CoV-2 IgG
0.5
1
2
4
8
16
32
64
SARS-CoV-2 IgM
SARS-CoV-2 IgG SARS-CoV-2 IgM
IgM levels
(arbitrary units)
IgG levels
(arbitrary units)
IgM levels
(arbitrary units)
0.5
1
2
4
8
16
32
64
128
0.5
1
2
4
8
16
32
64
128
. CC-BY-NC-ND 4.0 International licenseIt is made available under a
is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted July 10, 2020. .https://doi.org/10.1101/2020.07.08.20149179doi: medRxiv preprint