Study population

This study was based on an ongoing prospective biorepository cohort of parturients recruited on admission to the delivery room from May 5, 2021 to October 25, 2021, at the Hadassah Mount Scopus Medical Center in Jerusalem, Israel. Eligibility criteria included an age of 18 to 45 years and a willingness to participate and provide informed consent. Pregnant women with active maternal COVID-19 at delivery were excluded from the study. The institutional review board of the Hadassah Medical Center approved the study (HMO-0389-20, HMO-0274-21).

Because our first aim was to describe the impact of the time elapsed from infection to delivery on antibody levels in parturients with a history of COVID-19, we analyzed all available samples of unvaccinated patients infected at any time during pregnancy. SARS-CoV-2 infection was confirmed by a positive reverse transcriptase polymerase chain reaction test on a nasopharyngeal swab.

Subsequently (aim 2), parturients were assigned to 1 of 3 study groups, selected according to time of exposure, as follows:

• 1.
  

  Gravidae with a history of SARS-CoV-2 infection before or during pregnancy (from 29 weeks before pregnancy up to gestational week 22 ⁺⁰ ) who did not receive a boosting vaccine dose were assigned to the convalescent group;

  
  
• 2.
  

  Gravidae with a history of SARS-CoV-2 infection before or during pregnancy (from 29 weeks before pregnancy up to gestational week 22 ⁺⁰ ) who received a single boosting dose of the BNT162b2 mRNA vaccine during the index pregnancy (from gestational week 4 ⁺⁶ to week, 38 ⁺⁵ ) were assigned to the boosted convalescent group;

  
  
• 3.
  

  Naïve, fully vaccinated controls with no history of SARS-CoV-2 infections who received 2 doses of the BNT162b2 mRNA vaccine and who received the second dose from gestational week 4 ⁺⁶ to week 38 ⁺⁵ were assigned to the vaccinated group.

For the participants, maternal and umbilical cord blood samples were drawn at the time of delivery and during the postpartum period for the mothers (maternal blood drawn, 6–13 weeks postpartum). Demographic, medical, and side-effect profile data were collected for all patients using the medical computerized chart and designated questionnaires (aim 3).

A group of nongravid women of reproductive age was also recruited to characterize and compare the pattern of humoral response over time.

Sample and data collection and handling

Maternal and umbilical cord blood samples were collected immediately following delivery for patients who enrolled after they provided informed consent. Blood samples were centrifuged at 1000×g for 10 minutes at room temperature, and serum samples were aliquoted and stored at −80°C until analysis. Demographic and clinical data were collected at the time of enrollment.

Laboratory technique

Serum anti–SARS-CoV-2 spike receptor binding domain (RBD)–specific antibodies were assessed using the Architect SARS-CoV-2 IgG II Quant assay (Abbott Diagnostics, Chicago, IL).

For a subgroup of randomly selected mother-newborn dyads, neutralizing antibody titers against SARS-CoV-2 were determined using a wild-type SARS-CoV-2 microneutralization assay as previously described with minor changes. Briefly, following serum heat inactivation, samples were serially diluted using 2-fold dilutions (starting from 1:10; diluted in Dulbecco’s Modified Eagle’s Medium in a total volume of 50 μL). Diluted samples were incubated for 1 hour at 37°C in a humidified atmosphere with 5% CO ₂ with an equivalent volume of viral solution, including 100 median tissue culture infectious doses (TCID50) of SARS-CoV-2 isolate USA-WA1/2020 (cat. no. NR-52281; obtained from BEI Resources, Manassas, VA). The serum-virus mixtures (8 replicates for every serum dilution) were then added to a 96-well plate containing a semiconfluent VERO E6 cell monolayer (ATCC CRL-1586; maintained as described previously ). After incubation for 3 days at 37°C in a humidified atmosphere with 5% CO ₂ , the viral cytopathic effect was evaluated for each well. The median neutralization titer (NT50) was defined as the reciprocal of the highest serum dilution that protected 50% of culture wells from the cytopathic effect. Each assay included positive and negative serum controls, a cell control, and viral back-titration control.

Statistical analysis

Statistical analyses were performed using IBM SPSS 27 for Windows (IBM Corp, Armonk, NY), and Prism 5.01 (GraphPad Software, San Diego, CA). Correlations between fetal and maternal antibodies were analyzed by linear regression tests. Comparisons of the antibody concentrations among groups, and continuous parameters (eg, clinical data), were analyzed using Kruskal-Wallis 1-way ANOVA tests followed by a Dunn all-pairwise comparisons test or, alternatively, using Wilcoxon rank sum tests (if only 2 groups were compared). Comparisons between maternal and fetal concentrations within each group were analyzed with Wilcoxon matched-pairs signed-rank test. A Pearson chi-square analysis was used to compare proportional data. All statistical tests were based on 2-tailed hypotheses. Differences were considered significant at a P value <.05.