Cohort characteristics
A total of 83 women with twin pregnancies were enrolled from December 2020 to March 2022. Of these, 74 were carrying dichorionic diamniotic (DCDA) and 9 had monochorionic diamniotic (MCDA) twins (Figure 1). Of these women, 49 women were vaccinated before conception, 21 were immunized during pregnancy, and 13 were unvaccinated (Figure 2).
The median maternal age at enrolment was 34.3 years, the median gestational week at enrollment was 12.3 weeks, and the median BMI was 24.9 (all values were centered on the medians). Most of the participants were Jewish, and 37.2% were nulliparous. Conception was spontaneous for all MCDC and 47.3% of the DCDA (p<0.001). The rest were conceived through IVF or other assisted reproductive methods (Table 1, top). Chronic medical complications (diabetes mellitus, hyper-or-hypothyroidism, etc.) were rare, and none had chronic hypertension or cardiovascular diseases. All these features correspond to the general characteristics of our Twin Clinic population [16, 17].
Pregnancy outcomes
During the study period, four miscarried both twins, and ten lost one twin: two spontaneously, and eight after a selective reduction due to genetic or structural defects. Their pregnancy thus continued with one fetus (Figure 1). All MCDC were delivered by Caesarean section (CS) compared to 57.1% of the DCDA. The remainder of the DCDA twins were delivered vaginally (40.5%) or via tool-assisted delivery (Table 1 middle).
A total of 122 babies were born. Of the 6 MCDC pregnancies, 12 babies were born. Of the 60 DCDA deliveries, there were 100 babies (50 pairs) who were born after a full course of twin pregnancy, and 10 babies were born as singletons after spontaneous loss or twin reduction to singleton (Figure 2). There were 47.5% females (71.4% in the MCDC). No significant differences were found between the birthweight of babies born in MCDA and DCDA pregnancies. Of the newborns, 55.6% had a birthweight below 2500 grams, but only 6.6% were below 1500 grams. The majority of the newborns had a normal APGAR score at 1 and 5 minutes, the duration of NICU days was 10 days, and no baby was lost after delivery (Table 1, bottom).
Pregnancy complications
The rate of gestational diabetes melilotus (GDM) was 21.2%, preeclampsia was 5%, and preterm delivery (delivery<37 weeks) was 55.4%, (Table 1, middle) all of which are within the range of known values for twins in Israel [16, 17].
Vaccination and susceptibility to COVID-19
Of the eighty-three women in the sample, 21 were vaccinated during pregnancy, 49 were vaccinated before conception, and 13 were not vaccinated (Figure 2, Table 2). Of the non-vaccinated women, four became PCR positive, one before, and three during pregnancy, and all of the latter had severe COVID-19 symptoms that required hospitalization in the third trimester but they delivered healthy babies. Of the 21 who were vaccinated during pregnancy, only two contracted COVID-19 during pregnancy and both had very mild symptoms. Of the 49 vaccinated before conception, 13 tested positive on a PCR test, five prior to and seven during pregnancy, and all only had mild symptoms.
Serological response
Response as a function of time of vaccination
A. Vaccination before conception
As shown in the violin plot, in women who were vaccinated before conception (Figure 3A), the level of neutralizing Anti-S-IgG doubled from the first to the second trimester (p=0.05), and tripled from the second to the third trimester (p<0.01), reaching 1,475 au/ml [Inetr Quartile Range (IQR)- [392-3,020], reflecting a 7-fold increase during pregnancy (overall, p=0.012). Other studies have reported a third-trimester Anti-S-IgG for this vaccination in singleton pregnancies of 798[424-1,623] [18].
B. Vaccination during pregnancy
Of the women vaccinated during pregnancy, the violin plot shows an increase from 0 in the first trimester to 372 Au/ml in the second trimester, which however rapidly dropped to one-third of this amount in the third trimester to 112[54-357], and was significantly lower (p<0.001) than the third-trimester Anti-S-IgG levels in women vaccinated before conception (Figure 3 A&B). We found no significant differences between the results for the DCDA vs. MCDA (Table 2, lower part). Note that when the women were vaccinated during pregnancy their Anti-S-IgG was one-third of the levels reported elsewhere for women with a singleton who were vaccinated during pregnancy: 380[65.35-1442.5] (p<0.005) [14, 15].
Serological response as a function of the PCR results
The violin plot (Figure 4A) shows that the level of neutralizing Anti-S-IgG in the PCR-negative (healthy) vaccinated patients tripled from the first 102[0-478] to the second trimester 326[73-1,980] (p=0.005), but that the increase in the third trimester, although significant (p=0.05), was small, at 392[96-1,880]. Overall, the increase throughout pregnancy was significant (p<0.001). This is comparable to the level of third trimester neutralizing Anti-S-IgG among singleton vaccinated women who were PCR negative (healthy) as was reported elsewhere [14, 15].
In the women who were PCR positive before conception, there was no significant increase in their levels of neutralizing anti-S IgG throughout their pregnancy, especially when comparing the third trimester at 47[42-98] to the first 82[59-91] or the second at 60[33-123] trimesters. Note that all of these women were vaccinated but if they became PCR-positive before conception, their symptoms at the time of their positive PCR test were very mild.
In the small number of patients who were PCR positive during pregnancy, there was a significant increase in neutralizing Anti-S-IgG levels when positive, compared to the women who were positive for COVID-19 prior to conception, which for 1 patient was in the 1st trimester and for the other six occurred in the third trimester (p=0.005) (Figure 4&B). These levels were very similar to those obtained for the non-pregnant control group women who were vaccinated but became PCR positive (see Figure 5).
Serological response in non-pregnant women
The sample of women aged 32-41 who were vaccinated during the study period were tested in parallel to assess their level of neutralizing Anti-S-IgG before the third booster shot and one and two months afterwards. The violin plot shows that in the non-pregnant women who had negative PCR tests (healthy), the increase in the Anti-S-IgG remained stable at one and three months after vaccination (p< 0.001 for the overall difference between all three groups). Among the few non-pregnant women who were PCR-positive, none were hospitalized, and their level was twice as high as the PCR-negative women.