Comparatively low rates of COVID-19 in women admitted in labor and their newborns prior to routine vaccination of pregnant women: insights from Denmark

Abstract Background: In a country with a high-test frequency, societal lockdown, and pregnancy leave granted from 28 gestational weeks, we investigated SARS-CoV-2 infection in women admitted in labor and their newborn in the pre-vaccine period. Material and methods: A total of 1042 women admitted for delivery in two Danish hospitals agreed to a plasma sample and nasopharyngeal, vaginal, and rectal swabs and to sampling of umbilical cord blood and a nasopharyngeal swab from their newborn at delivery. Plasma samples from women were examined for SARS-CoV-2 antibodies. If antibodies were detected, or the woman had a positive nasopharyngeal swab upon admission or had a household contact with symptoms consistent with COVID-19, SARS-CoV-2 PCR was performed on plasma and swab samples from mother and child. Results: Seventeen women (1.6%) were seropositive. Half the newborn (n = 9 (53%)) of seropositive mothers were also seropositive. None of the seropositive women or newborns had clinical signs of COVID-19 and all had SARS-CoV-2 PCR negative plasma and swab samples. Conclusion: Adherence to specific national guidelines pertaining to testing, self-imposed isolation, and cautious behaviors among pregnant women likely contributed to the exceptionally low prevalence of both prior and current COVID-19 infections detected at the time of childbirth preceding the routine vaccination of pregnant women in Denmark.


Introduction
Significant variations exist in the incidence and severity of COVID-19 in pregnant and laboring women during the pre-vaccine period, even within the Scandinavian countries [1][2][3].These disparities could potentially be attributed to variations in recommendations, rules and regulations related to SARS-CoV-2 testing, availability of tests and isolation, and timing of pregnancy leave.National policies and guidelines are of paramount importance for pandemic protection.The national guidelines issued by the Board of Health emphasized alcohol based hand rub before and after contact, use of mask in public transportation, keeping physical distance, and restrictions on the number of people gathering socially.However, the implementation of public health measures and hospital restrictions during the pandemic left pregnant women more anxious and vulnerable, and differences in self-imposed isolation and other behavioral measures in families and during pregnancy could also play a role [4,5].
At the early phase of the pandemic, there was an increasing focus on protection of the pregnant population, and The Danish Health Authorities defined pregnant women as having an increased risk of a serious course of infection [6].This involved a very extensive test strategy and granted leave from gestational week 28 (usually 32 weeks).
The aim of this study was to examine presence of SARS-CoV-2 infection and pregnancy outcome in an unselected cohort of women admitted in labor prior to the routine vaccination of pregnant women introduced in Denmark in July 2021.Additionally, we also aimed to discuss our results in the lights of published findings in Danish non-pregnant populations of the same age and period and to discuss the possible impact of a high test-frequency, societal lockdown, and the granted early pregnancy leave.

Materials and methods
From April 23 to November 15, 2020, all women admitted in labor at two Danish hospitals were invited; 1042 participated.Upon arrival, a structured interview was conducted to assess the presence of current and previous COVID-19 symptoms among household members.Nasopharyngeal, rectal and vaginal swabs and a blood sample were obtained from the women during delivery.SARS-CoV-2 PCR testing was conducted on nasopharyngeal swabs obtained from all women immediately upon their arrival in labor.Umbilical cord blood and a nasopharyngeal swab were collected from the newborns for subsequent analysis.Plasma samples from all mothers underwent SARS-CoV-2 total antibody analysis.When positive, the plasma sample was also investigated for IgM antibodies while corresponding umbilical cord plasma was examined for both total and IgM antibodies.
For all seropositive women and their newborns, SARS-CoV-2 PCR analysis was performed on plasma and swab samples, irrespective of the newborn's antibody status.
Additionally, if a woman's nasopharyngeal swab tested positive upon admission or if there was a household contact with COVID-19 symptoms, SARS-CoV-2 PCR was performed on women's and newborns' plasma sample.
For details on SARS-CoV-2 PCR assays, see Appendix 1.For details on serology assays, see Appendix 2.

Changes in national guidelines and lock-down
During the study period, there were significant updates to the national COVID-19 guidelines including the implementation of stricter protocols for the use of protective equipment, the adoption of populationbased testing strategies, the enforcement of travel restrictions, and regulations of quarantine and self-isolation were implemented.
Notably, on April 2 nd 2020, three weeks before initiation of our study, pregnancy leave was granted from 28 gestational weeks, as opposed to the previous 32 weeks.Furthermore, starting from April 30, 2020, women in labor were tested as part of the national strategy.Halfway through the study period, Denmark emerged as one of the leading countries globally in terms of the frequency of testing [7].Women were invited to participate in our study regardless of the changes in the national guidelines.
Maternal characteristics, pregnancy complications and condition of the newborn at delivery were retrieved from electronic medical patient records.A structured interview was conducted with all women to assess the presence of current or previous COVID-19 symptoms among all household members.
Characteristics of the women and their newborn by antibody status were shown as relative risks (RR) with 95% confidence intervals (CI).Fisher's exact test for categorical variables and Wilcoxon rank-sum test for two independent samples with two groups were used.Goodness of fit was investigated by Hosmer-Lemeshow's test.Data was analyzed by use of STATA, version 16.

Results
During the seven months study period, 4795 women gave birth (2878 at Aarhus University Hospital and 1917 at Lillebaelt Hospital) of which 1042 women and their newborns participated.Basic characteristics of women, their pregnancy and newborns by SARS-CoV-2 antibody status are presented in Table 1.
Among the 1042 women, 17 (1.6%)were total antibody positive.Three of the total antibody positive were also IgM positive.Only one woman had a SARS-CoV-2 PCR positive nasopharyngeal swab at delivery.She was seronegative and had no COVID-19 symptoms.Nine newborns of the 17 seropositive mothers were total antibody positive (53%).Two were also IgM positive (Figure 1 and Appendix 3).
Pregnancy outcome was similar in seropositive and seronegative women (Table 1).The seropositive newborns were all born at term with normal birthweight and were clinically well (normal Apgar scores and normal umbilical artery pH).
All SARS-CoV-2 PCR tests of seropositive women (plasma, nasopharyngeal, rectal and vaginal swab) and their newborn (plasma, nasopharyngeal swab) were negative and none of the women or newborn had any symptoms of COVID-19.

Discussion
Among the 1042 participating women in our study admitted in labor during the first seven months of the pandemic in Denmark, less than 2% were SARS-CoV-2 total antibody positive and less than 0.5 0% were IgM positive.Half the newborn infants of the seropositive mothers in the study were SARS-CoV-2 antibody positive.Only one woman was SARS-CoV-2 PCR positive.
We found evidence of transplacental transfer of maternal antibodies against SARS-CoV-2 in half the newborn of seropositive mothers in line with current evidence of transfer [8,9].We found no transmission of the virus from mother to fetus, which has also been shown previously in studies with selected groups of seropositive women and newborns at delivery [10,11].The low rate of SARS-CoV-2 antibody-positive participants in our study is compatible with the finding from another Danish study on antibody prevalence at the time of delivery [12].For comparison, Sweden, a neighboring country, found a seroprevalence of about 15% in December 2020 in both a randomly selected group of pregnant women and a group of otherwise healthy blood donors [13].In another Swedish study from the same inclusion period as ours, around 5% of women admitted in labor were nasopharyngeal SARS-CoV-2 PCR positive [14].These differences from our findings could be partially explained by the different national guidelines in Sweden and Denmark, where Sweden had a unique strategy for managing COVID-19 involving no national lockdown [13].
French studies have reported seroprevalence rates in pregnant women of 4.7% [15] and 8% [16] and Flannery et al. found a seroprevalence of 6.2% in pregnant women at delivery in Philadelphia [3].
The seroprevalence in our study group of pregnant women is lower than that reported from the background population.A Danish study including hospital healthcare and administrative personnel during our inclusion period found a seroprevalence of 3.4% in women below the age of 40 years [17].A Danish study of blood donors (17-35 years), also from the same period and uptake area, reported a seroprevalence around 2.5% [18].
Our very low rate of SARS-CoV-2 PCR-positive women giving birth (<0.1% positive) were also different from that of the background population in which 1.3% of Danish women of 20 to 39 years were SARS-CoV-2 PCR positive during the study period [19] (Statens_Seruminstitut_report_positve_SARS_Cov_2_ 230420_151120; Personal communication).
Studies have reported increased risk of anxiety symptoms among pregnant women during the pandemic and a strong relation between mental health and the feelings related to burdens experienced.Kajdy et al. [5] found that satisfaction with government reactions and fear appraisal played an important role in the perception of the efficacy of restrictions.Chivers et al. [20] found that general information on COVID-19 safe behaviors did not meet the pregnant women's particular needs, potentially exacerbating the risk of psychological and psychosocial distress.Feduniw et al. [21] described women's changes in childbirth plans during the pandemic and found that the decision-making process surrounding childbirth was markedly influenced and that some women were more likely to choose a home birth, with or without medical assistance.
We do not have data on women in our inclusion period who may have avoided hospital deliveries by giving birth at home, or data on women who may have postponed pregnancy due to anxiety related to the pandemic.
This study possesses several notable strengths.Firstly, since the inclusion period of our study was from April to November 2020, all included women were pregnant at the onset of the pandemic and subsequently delivered during its initial phase.Secondly, a meticulously structured interview was conducted to also assess the presence of current and previous COVID-19 symptoms among household members.Lastly, longitudinally collected, comprehensive medical data pertaining to delivery and the newborn were obtained.
The use of the Wantai assay for total and IgM antibodies in our serology testing is a major strength as it performed best among 16 serological assays compared [22].Another strength is the consistently performed SARS-CoV-2 PCR analysis on maternal and newborn plasma and swab samples.
A major limitation is the relative low rate of participance (21.6%).This could not be attributed to skewed selection of women, e.g.related to age, BMI, birthweight, or mode of delivery and therefore we consider the likelihood of bias to be small.
The low prevalence of SARS-CoV-2 antibodies indicates that national guidelines emphasizing testing and pregnancy leave starting from the 28th week of gestation had a significant preventive effect on pregnant women.Moreover, it is plausible that the prevailing sense of uncertainty experienced by pregnant women during the COVID-19 pandemic has contributed to a distinct cautious behavior including self-imposed isolation, further reinforcing the need for comprehensive information and adequate support for pregnant women during times of disease outbreaks.

Conclusion
Adherence to specific national guidelines pertaining to testing, self-imposed isolation, and cautious behaviors among pregnant women likely contributed to the exceptionally low prevalence of both prior and current COVID-19 infections detected during the period of childbirth preceding the routine vaccination of pregnant women in Denmark.Notably, the seroprevalence during pregnancy was a mere 1.6% compared to 3.4% for a similar age population of women working in healthcare.Moreover, extensive SARS-CoV-2 PCR testing of plasma, rectal swabs and vaginal swabs yielded uniformly negative results for seropositive women, and none of the women exhibited symptoms of COVID-19.
results from maternal plasma, rectal or vaginal swabs, umbilical cord plasma and nasopharyngeal swabs from the newborn were negative (Figure 1).
One of the two IgM positive newborn had a mother who was total antibody and IgM positive, the other had a mother who was only total antibody positive.The mothers of the remaining seven seropositive newborn were total antibody positive only.
The three mothers with serology indicating current infection (total antibody and IgM positive) gave birth to five newborn of whom one had no antibodies, one was total antibody and IgM positive.Serum was not available for the remaining three.

Table 1 .
Characteristics of women in labor (N ¼ 1042) by SARS-CoV-2 total antibody status.