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Serum and breastmilk SARS-CoV-2 specific antibodies following BNT162b2 vaccine: prolonged protection from SARS-CoV-2 in newborns and older children

Open AccessPublished:July 05, 2022DOI:https://doi.org/10.1016/j.ijid.2022.06.055

      Highlights

      • Vaccination is the best strategy against SARS-CoV-2 infection.
      • COVID-19 in newborns may be more serious than in older children.
      • The immune response of lactating women after receiving the BNT162b2 vaccine was measured.
      • Serum and breastmilk contain specific antibodies 6 months after vaccination.

      Abstract

      Objectives

      Vaccination is the best strategy against COVID-19. We aimed to determine antibodies against SARS-CoV-2 in breastmilk and serum of mothers vaccinated with the mRNA vaccine.

      Methods

      This prospective study included 18 lactating women vaccinated with the BNT162b2 vaccine. Serum and breastmilk were collected before the first dose (T0), at the second dose (T1), 3 weeks after the second dose (T2), and 6 months after the first dose (T3). Serum anti-SARS-CoV-2 Spike (S) Immunoglobulin G (IgG) and Immunoglobulin A (IgA) were measured using a semi-quantitative enzyme-linked immunosorbent assay (ELISA) and secretory antibody (s) IgG and IgA in breastmilk using quantitative analysis.

      Results

      We detected serum anti-S IgG and IgA in all women after vaccination. Specific IgG and IgA were higher at T1, T2, and T3 compared with T0 (P <0.0001). Higher antibody levels were observed at T2 and lower values at T3 versus T2 (P = 0.007). After 6 months, all patients had serum IgG, but three of 18 (16%) had serum IgA. In breastmilk, sIgA was present at T1 and T2 and decreased after 6 months at T3 (P = 0.002). Breastmilk sIgG levels increased at T1 and T2 and peaked at T3 (P = 0.008).

      Conclusion

      Secretory antibodies were transmitted through breastmilk until 6 months after anti-COVID-19 mRNA vaccination. Protection of the newborn through breastfeeding needs to be addressed.

      Keywords

      Introduction

      Since December 2019, a novel SARS-CoV-2 causing COVID-19 has been spreading worldwide (
      Coronaviridae Study Group of the International Committee on Taxonomy of Viruses
      The species severe acute respiratory syndrome-related coronavirus: classifying 2019-nCoV and naming it SARS-CoV-2.
      ). The large number of patients requiring hospitalization and the high lethality rates of COVID-19 caused excessive stress on healthcare facilities. Because of the urgent need for preventive strategies, several vaccines have been speeding through the experimental phases. In December 2020, new mRNA vaccines were approved (
      • Polack FP
      • Thomas SJ
      • Kitchin N
      • et al.
      Safety and efficacy of the BNT162b2 mRNA Covid-19 vaccine.
      ;
      • Walter EB
      • Talaat KR
      • Sabharwal C
      • et al.
      Evaluation of the BNT162b2 Covid-19 vaccine in children 5 to 11 years of age.
      ;
      • Woodworth KR
      • Moulia D
      • Collins JP
      • et al.
      The Advisory Committee on Immunization Practices' interim recommendation for use of Pfizer-BioNTech COVID-19 vaccine in children aged 5-11 years - United States, November 2021.
      ). The BNT162b2 vaccine was the first mRNA vaccine distributed worldwide and was administered to high-risk persons first (
      • Polack FP
      • Thomas SJ
      • Kitchin N
      • et al.
      Safety and efficacy of the BNT162b2 mRNA Covid-19 vaccine.
      ).
      Breastfeeding women were not included in phase II/III studies. However, hypothetic damage of liposomal RNA in human breastmilk was not demonstrated. No evidence supporting harmful effects concerning lactation was observed during animal studies (
      • Golan Y
      • Prahl M
      • Cassidy AG
      • et al.
      COVID-19 mRNA vaccination in lactation: assessment of adverse events and vaccine related antibodies in mother-infant dyads.
      ). Very soon, many international and national societies have recommended discussing the risks and benefits of the vaccination for breastfeeding women at higher risk of exposure to the virus as healthcare professionals and offer them vaccination (
      • Bartick MC
      • Valdés V
      • Giusti A
      • et al.
      Maternal and infant outcomes associated with maternity practices related to COVID-19: the COVID mothers study.
      ;

      European Medicines Agency. Comirnaty. https://www.ema.europa.eu/en/news/ema-recommends-first-covid-19-vaccine-authorisation-eu https://www.ema.europa.eu/en/medicines/human/summaries-opinion/comirnaty, 2020 (accessed on 29 december 2020).

      ;

      Joint Committee on Vaccination and Immunisation. COVID-19 vaccination: a guide for women of childbearing age, pregnant or breastfeeding, Updated 2021. https://www.gov.uk/government/publications/covid-19-vaccination-women-of-childbearing-age-currently-pregnant-planning-a-pregnancy-or-breastfeeding, 2021 (accesed on january 2022).

      ;

      Hare H, Womersley K. Healthcare workers who breastfeed should be offered the Covid-19 vaccine. https://blogs.bmj.com/bmj/2020/12/21/healthcare-workers-who-breastfeed-should-be-offered-the-covid-19-vaccine, 2020 (accesed on 15 january 2021).

      ). In addition, preliminary data indicated that after SARS-Cov-2 infection, antibodies against SARS-CoV-2 might be present in breastmilk (
      • Cervia C
      • Nilsson J
      • Zurbuchen Y
      • et al.
      Systemic and mucosal antibody responses specific to SARS-CoV-2 during mild versus severe COVID-19.
      ;
      • Fox A
      • Marino J
      • Amanat F
      • et al.
      Robust and specific secretory IgA against SARS-CoV-2 detected in human milk.
      ), and 90% of those are secretory IgA with neutralizing activity, suggesting the same finding in women receiving a vaccination. Following these recommendations, at San Matteo University Hospital in Pavia, we decided to offer vaccination with the BNT162b2 mRNA vaccine to breastfeeding healthcare workers and collect breastmilk and serum to demonstrate specific antibody secretion in breastmilk after vaccination and to compare serum and secretory immune response.

      Methods

      Subjects enrolled

      Between January 13, 2021, and March 1, 2021, at Fondazione IRCCS Policlinic San Matteo, 18 breastfeeding healthcare workers (median age 34 years, range 29-41) at different months postpartum (median 11 months; range 1-36) receiving the first dose of SARS-CoV-2 vaccine (BNT162b2) were enrolled. Serum and breastmilk samples were collected before the vaccination (T0), at the second dose (T1), at 3 weeks after the second dose (T2), and 6 months after the first dose (T3) for evaluation of SARS-CoV-2 antibody-specific response. Women with previous SARS-CoV-2 infection, confirmed through RT-PCR from nasopharyngeal swabs or serum anti-Nucleocapsid Protein (N) IgG, were excluded from the study. Ethical approval was obtained from the Medical Ethics Committee of the Policlinico San Matteo University Hospital, and written informed consent was obtained from all participants. Participants were asked to collect about 3 ml of milk. After centrifugation at 3000 rpm for 10 minutes, sera and milk were stored at −80°C until analyses were performed.

      Antibody response

      Anti-SARS-CoV-2 Spike (S) serum IgG and IgA antibodies were measured by a semi-quantitative enzyme-linked immunosorbent assay (ELISA) (Euroimmun, Luebeck, Germany) at different time points, according to the manufacturer's instructions. Results were expressed as a ratio concerning an internal calibrator (RU/ml).
      Secretory antibody (sIgG and sIgA) quantification in breastmilk samples was performed with the same kit (Euroimmun). We used a standard curve obtained from a breastmilk pool derived from women previously infected with SARS-CoV-2. We defined that 100 AU/ml of specific sIgA (IgA-AU/ml) and 100 AU/ml of specific sIgG (IgG-AU/ml) were contained in this pool.

      Statistical analysis

      Patient characteristics and COVID-19 symptoms were expressed as mean with SD or median with interquartile range depending on their distribution. Comparisons of antibody levels at T1, T2, and T3 with respect to T0 were performed fitting generalized equation models with first order autocorrelation to take into account the clustered nature of the data.

      Results

      The demographic and clinical characteristics of 18 lactating women and their newborns are listed in Table 1.
      Table 1Demographic and clinical characteristics of 18 lactating women and their newborns
      Age median ± SD at delivery34,0 (± 4,5)
       (minimum, maximum) years29-45
      Caucasian n (%)18/18 (100%)
      Co-morbidities n (%)3/18 (16%)
      Thyroid disorders3/18 (16%)
      Delivery data
       Gestational age at delivery median ± SD (Range)39,5 (1,4)
      Delivery at term18/18
       Cesarean delivery4/18 (22%)
       Vaginal delivery14/18 (78%)
      Infant and breastfeeding data
      Median newborn age at 1st vaccination11,5 (SD 8,5 1-36)
       SGA1/18
      Newborn's side effects after vaccinationNone
      Mother's side effects post-vaccination n (%)
      Minor side effects (n. person)5/8
       Myalgia3/18
       Headache3/18
       Local pain1/18
       Arm lymph node swelling1/18
       Fever1/18
       Skin rash1/18
       Tiredness-1/18
       Arthralgia1/18
       Arm numbness1/18
      Major side effectsNone

      Antibody response elicited by BNT162b2 vaccine

      Serum

      As listed in Table 2, the evaluation of serum anti-S antibodies showed that serum IgG was significantly higher at T1, T2, and T3 compared with T0 (median 5.9 RU/ml, range 0.3-8.2; median 7.6 RU/ml, range 6.3-8.3 and median 5.0 RU/ml, range 2.8-5.8 respectively; P <0.0001 for all). Higher levels were observed at T2 compared with T1 (P <0.02), whereas T3 values were significantly lower than T2 values (P <0.001). At T1, one subject (5.8%) showed specific IgG levels under the cut-off that became positive after the second dose administration. A more homogeneous distribution of antibody values was observed at T2 and T3 compared with T1 (Figure 1).
      Table 2Antibody response elicited by BNT162b2 vaccine in serum and breastmilk over time
      Time from vaccine somministrationT0(1st dose)T1(2nd dose)T2(3 weeks after 2nd dose)T3(6 months after 1st dose)
      Serum anti-S IgG (Median RU/ml, range)<0,8 RU/ml5.9 (0.3-8.2)7.6 (6.3-8.3)5.0 (2.8-5.8)
      Serum anti-S IgA (Median RU/ml, range)<0,8 RU/ml3.3 (0.2-7.5);5.0 (1.6-7.4)3.1 (0.6-8.0)
      Breastmilk anti-S sIgG (Median AU/ml, range)<15 IgG-AU/ml15 (15-95)28 (15-117)114 (82-156)
      Breastmilk anti-S sIgA (Median AU/ml, range)<15 IgG-AU/ml422 (8-1500)565 (47-1500)80 (8-468)
      AU = arbitrary units ; IgA = Immunoglobulin A; IgG = Immunoglobulin G; RU = relative units.
      Figure 1
      Figure 1Serum IgA and IgG, secretory breastmilk IgA and IgG
      AU = arbitrary units ; IgA = Immunoglobulin A; IgG = Immunoglobulin G; RU – relative units
      Serum anti-S IgA values were significantly higher at T1, T2, and T3 compared with T0 (median 3.3 RU/ml, range 0.2-7.5; median 5.0 RU/ml, range 1.6-7.4 and median 3.1 RU/ml, range 0.6-8.0, respectively; P <0.0001 for all). As observed for specific IgG, serum-specific IgA higher levels were observed at T2 compared with T1 (P <0.001). In contrast, significantly lower values were observed at T3 compared with T2 (P = 0.007). For serum IgA, scattered values at each time point were observed (Figure 1). At T3, three samples (17.6%) showed serum anti-S IgA levels under the cut-off value. The same subjects showed low antibody levels even at T1 and T2 (Table 3).
      Table 3Serum IgA levels in the three patients with negative levels at T3
      Subject codeT1 (RU/ml)T2 (RU/ml)T3 (RU/ml)
      #771.401.600.60
      #1061.905.100.90
      #1450.702.200.90

      Breastmilk

      Breastmilk sIgG levels increased at T1 (median 15 IgG-AU/ml, range 15-95), T2 (median 28 AU/ml, range 15-117), and T3 (median 114 AU/ml, range 82-156) (Figure 1, Figure 1 Supplement) compared with T0 (all values <15 IgG-AU/ml), reaching significantly higher levels at T3 (P = 0.008). sIgA levels were significantly higher at T1 (median 422 IgA-AU/ml, range 8-1500) and T2 (median 565 IgA-AU/ml, range 47-1500) compared with T0 (median 80 IgA-AU/ml, range 8-468) (P <0.0001, for all), whereas at T3 (median 155 IgA-AU/ml, range 44.5-350), levels decreased compared with T0. As well as for SARS-CoV-2 IgG and IgA serum response, breastmilk sIgA was induced after the first vaccine dose, reaching higher levels 3 weeks after complete vaccination. However, 6 months after the second dose, we observed a significant decrease (P = 0.002) (Figure 1, Figure 1 Supplement).

      Discussion

      SARS-CoV-2 infections in a newborn may be acquired in two different ways: less common but potentially serious by vertical transmission, as shown by many authors that have gathered evidence of SARS-CoV-2 RNA on the placenta, amniotic fluid, and cord blood (
      • Kotlyar AM
      • Grechukhina O
      • Chen A
      • et al.
      Vertical transmission of Coronavirus disease 2019: a systematic review and meta-analysis.
      ;
      • Patanè L
      • Morotti D
      • Giunta MR
      • et al.
      Vertical transmission of coronavirus disease 2019: severe acute respiratory syndrome coronavirus 2 RNA on the fetal side of the placenta in pregnancies with coronavirus disease 2019-positive mothers and neonates at birth.
      ); and more frequently after delivery through large respiratory droplets containing the SARS-CoV-2 virus from affected mothers (
      • Liguoro I
      • Pilotto C
      • Bonanni M
      • et al.
      Correction to: SARS-COV-2 infection in children and newborns: a systematic review.
      ).
      Maternal transmission of SARS-CoV-2 through breastfeeding was not demonstrated (
      • Centeno-Tablante E
      • Medina-Rivera M
      • Finkelstein JL
      • et al.
      Transmission of SARS-CoV-2 through breast milk and breastfeeding: a living systematic review.
      ). When SARS-CoV-2 was found in the breastmilk of affected women, it was not possible to culture it (
      • Krogstad P
      • Contreras D
      • Ng H
      • et al.
      No infectious SARS-CoV-2 in breast milk from a cohort of 110 lactating women.
      ).
      Conversely, much evidence exists on the benefit of human breastmilk in protecting feeding babies from infections such as influenza and respiratory viruses (
      WHO Collaborative Study Team on the role of breastfeeding on the prevention of infant mortality
      Effect of breastfeeding on infant and child mortality due to infectious diseases in less developed countries: a pooled analysis.
      ;
      • Goldman AS.
      The immune system of human milk: antimicrobial, anti-inflammatory and immunomodulating properties.
      ).
      It is well known that secretory IgA represents the primary protective component of human breastmilk, blocking pathogen entry on the mucosal surface of lactating infants (
      WHO Collaborative Study Team on the role of breastfeeding on the prevention of infant mortality
      Effect of breastfeeding on infant and child mortality due to infectious diseases in less developed countries: a pooled analysis.
      ;
      • Goldman AS.
      The immune system of human milk: antimicrobial, anti-inflammatory and immunomodulating properties.
      ). Anti-SARS-CoV-2 immunity was demonstrated mainly in serum and breastmilk of women recovered from SARS-CoV-2 infection (
      • Fox A
      • Marino J
      • Amanat F
      • et al.
      Robust and specific secretory IgA against SARS-CoV-2 detected in human milk.
      ) and, as observed in our study, in women vaccinated with anti-COVID-19 mRNA vaccines (
      • Baird JK
      • Jensen SM
      • Urba WJ
      • et al.
      SARS-CoV-2 antibodies detected in mother's milk post-vaccination.
      ;
      • Golan Y
      • Prahl M
      • Cassidy AG
      • et al.
      COVID-19 mRNA vaccination in lactation: assessment of adverse events and vaccine related antibodies in mother-infant dyads.
      ;
      • Gonçalves J
      • Juliano AM
      • Charepe N
      • et al.
      Secretory IgA and T cells targeting SARS-CoV-2 spike protein are transferred to the breastmilk upon mRNA vaccination.
      ;
      • Perl SH
      • Uzan-Yulzari A
      • Klainer H
      • et al.
      SARS-CoV-2-specific antibodies in breast milk after COVID-19 vaccination of breastfeeding women.
      ). A previous study reported data where vaccinated breastfeeding women had an immune response characterized by a significant increase in sIgA and sIgG after the second dose (
      • Perl SH
      • Uzan-Yulzari A
      • Klainer H
      • et al.
      SARS-CoV-2-specific antibodies in breast milk after COVID-19 vaccination of breastfeeding women.
      ). However, our results are in accordance with sIgA but not for sIgG.
      For sIgA, this is proof of the presence of a booster effect for mucosal immunity, and this effect may theoretically be replicated with additional doses. The confirmation that sIgA represents the first response of the immune system to SARS-CoV-2 (
      • Sterlin D
      • Mathian A
      • Miyara M
      • et al.
      IgA dominates the early neutralizing antibody response to SARS-CoV-2.
      ), followed by IgG, and that breastfeeding women express robust immune response as reported for non-lactating women (
      • Cassaniti I
      • Bergami F
      • Percivalle E
      • et al.
      Humoral and cell-mediated response against SARS-CoV-2 variants elicited by mRNA vaccine BNT162b2 in healthcare workers: a longitudinal observational study.
      ).
      Our study has shown that after 6 months, sIgA antibody levels significantly decrease. However, this deflection appears to strongly affect women with the weakest initial sIgA response. A possible explanation for the sIgA decline is that the intramuscular route of administering the vaccine cannot trigger a strong mucosal response that lasts over time. Moreover, in this case, we cannot exclude a selective IgA deficiency (
      • Sterlin D
      • Mathian A
      • Miyara M
      • et al.
      IgA dominates the early neutralizing antibody response to SARS-CoV-2.
      ).
      Therefore, no correlation between serum and breastmilk IgA was observed, probably because of the two sources of IgA in breastmilk (serum monomeric or mucosal polymeric from breast mucose-associated lymphoid tissue MALT) (
      • Gonçalves J
      • Juliano AM
      • Charepe N
      • et al.
      Secretory IgA and T cells targeting SARS-CoV-2 spike protein are transferred to the breastmilk upon mRNA vaccination.
      ;
      • Gray KJ
      • Bordt EA
      • Atyeo C
      • et al.
      Coronavirus disease 2019 vaccine response in pregnant and lactating women: a cohort study.
      ;
      • Perl SH
      • Uzan-Yulzari A
      • Klainer H
      • et al.
      SARS-CoV-2-specific antibodies in breast milk after COVID-19 vaccination of breastfeeding women.
      ). Despite a low level of sIgA in breastmilk over time, Gonçalves et al. have shown that repeated breastfeeding probably results in the accumulation of sIgA that retains a neutralization effect on SARS-CoV-2 (
      • Gonçalves J
      • Juliano AM
      • Charepe N
      • et al.
      Secretory IgA and T cells targeting SARS-CoV-2 spike protein are transferred to the breastmilk upon mRNA vaccination.
      ). The same authors have observed a similar neutralizing effect but to a lesser extent for breastmilk sIgG. Interestingly, our group observed a weak increase in breastmilk sIgG levels at 6 months but a decrease in sIgA levels simultaneously. We cannot compare quantification outcomes of breastmilk sIgG and sIgA because a semi-quantitative analysis for serum IgG and IgA evaluation was used. We observed the trend over time (Figure 1 Supplement).
      The explanation for this finding may be that most studies reported a peak of IgG after the second dose. However, as in our study, the observation period is not long enough to observe the real expression of the curve (
      • Charepe N
      • Gonçalves J
      • Juliano AM
      • et al.
      COVID-19 mRNA vaccine and antibody response in lactating women: a prospective cohort study.
      ;
      • Golan Y
      • Prahl M
      • Cassidy AG
      • et al.
      COVID-19 mRNA vaccination in lactation: assessment of adverse events and vaccine related antibodies in mother-infant dyads.
      ;
      • Gonçalves J
      • Juliano AM
      • Charepe N
      • et al.
      Secretory IgA and T cells targeting SARS-CoV-2 spike protein are transferred to the breastmilk upon mRNA vaccination.
      ). Moreover, other studies have observed that other factors may impact the sIgG levels: breastfeeding for longer than 6 months may increase the sIgG level over time, and this is a possible contribution to what we observed (
      • Abuidhail J
      • Al-Shudiefat AA
      • Darwish M.
      Alterations of immunoglobulin G and immunoglobulin M levels in the breast milk of mothers with exclusive breastfeeding compared to mothers with non-exclusive breastfeeding during 6 months postpartum: the Jordanian cohort study.
      ;
      • Charepe N
      • Gonçalves J
      • Juliano AM
      • et al.
      COVID-19 mRNA vaccine and antibody response in lactating women: a prospective cohort study.
      ;
      • Czosnykowska-Łukacka M
      • Lis-Kuberka J
      • Królak-Olejnik B
      • Orczyk-Pawiłowicz M.
      Changes in human milk immunoglobulin profile during prolonged lactation.
      ). Therefore, the small number of patients and the lack of observation after 6 months may limit the interpretation of this finding.
      Notably, increasing evidence in other respiratory infections supports a more important role in neonatal immune response for IgG in breastmilk (
      • Demers-Mathieu V
      • Huston RK
      • Markell AM
      • et al.
      Impact of pertussis-specific IgA, IgM, and IgG antibodies in mother's own breast milk and donor breast milk during preterm infant digestion.
      ;
      • Mazur NI
      • Horsley NM
      • Englund JA
      • et al.
      Breast milk prefusion F Immunoglobulin G as a correlate of protection against respiratory syncytial virus acute respiratory illness.
      ). We suggest a similar role in vaccine-induced breastmilk immunity. The beneficial effects could last for months after anti-COVID-19 vaccination with mRNA formulations.
      Newborns from SARS-CoV-2 infected mothers showed an increased admission rate to a neonatal unit (
      • Allotey J
      • Stallings E
      • Bonet M
      • et al.
      Clinical manifestations, risk factors, and maternal and perinatal outcomes of coronavirus disease 2019 in pregnancy: living systematic review and meta-analysis.
      ). Several cases of severe disease were reported in infants younger than 6 months (
      • de Siqueira Alves Lopes A
      • Fontes Vieira SC
      • Lima Santos Porto R
      • et al.
      Coronavirus disease-19 deaths among children and adolescents in an area of Northeast, Brazil: why so many?.
      ). In this scenario, immune response persistence after 6 months may be particularly important in planning vaccination strategy in puerperium and pregnancy. Numerous studies have observed that certain vaccines induce changes in breastmilk composition, mainly if offered in late pregnancy (
      • Hahn-Zoric M
      • Carlsson B
      • Jeansson S
      • et al.
      Anti-idiotypic antibodies to poliovirus antibodies in commercial immunoglobulin preparations, human serum, and milk.
      ;
      • Schlaudecker EP
      • Steinhoff MC
      • Omer SB
      • et al.
      IgA and neutralizing antibodies to influenza A virus in human milk: a randomized trial of antenatal influenza immunization.
      ;
      • Shahid NS
      • Steinhoff MC
      • Roy E
      • et al.
      Placental and breast transfer of antibodies after maternal immunization with polysaccharide meningococcal vaccine: a randomized, controlled evaluation.
      ;
      • Su F
      • Patel GB
      • Hu S
      • et al.
      Induction of mucosal immunity through systemic immunization: phantom or reality?.
      ). Therefore, many authors have already demonstrated IgG antibodies passage through the placenta in SARS-CoV-2 infected women during pregnancy or after mRNA vaccination (
      • Beharier O
      • Mayo RP
      • Raz T
      • et al.
      Efficient maternal to neonatal transfer of antibodies against SARS-CoV-2 and BNT162b2 mRNA COVID-19 vaccine.
      ;
      • Cassaniti I
      • Percivalle E
      • Zelini P
      • et al.
      Both SARS-CoV-2 infection and vaccination in pregnancy elicited neutralizing antibodies in pregnant women and newborns.
      ). The favorable effect on newborns must be addressed. However, we believe vaccination during lactation or late pregnancy could confer protection thanks to the specific IgG transfer through the placenta, as already demonstrated and used for pertussis and influenza (
      • Su F
      • Patel GB
      • Hu S
      • et al.
      Induction of mucosal immunity through systemic immunization: phantom or reality?.
      ), but also by adding mucosal protection through breastmilk sIgA and sIgG. If established with other studies, breastmilk antibodies may be beneficial in preventing SARS-CoV2 in newborns and infants not included in the vaccination program considering that additional vaccine doses could confer continuous protection.
      We are aware that we have used a commercially available ELISA to detect SARS-CoV-2 specific serum IgA (sample dilution 1:100). We tested it also for sIgA quantification in saliva and breastmilk (sample dilution 1:5). We found it specific and reproducible; however, we will perform more investigations to standardize the assay for secretory antibodies.
      Therefore, the restricted number of cases could represent a limitation of our study. However, the data clearly indicate that passive immunization in this setting is safe and possibly worthwhile.

      Funding source

      This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

      Ethical approval

      This study is a prospective analysis approved by the Medical Ethics Committee of the Fondazione IRCCS Policlinico San Matteo, Pavia, Italy.

      Author contributions

      Alessandra Ricciardi Conceived and designed the analysis; Collected the data; Contributed data or analysis tools; Wrote the paper.Paola Zelini Contributed data or analysis tools; Performed the analysis; Wrote the paper.Irene Cassaniti Conceived and designed the analysis; Collected the data; Contributed data or analysis tools; Marta Colaneri: Contributed data or analysis tools; Performed the analysis; Raffaele Bruno Wrote the paper.Fausto Baldanti Wrote the paper.Maria Antonietta Avanzini Contributed data or analysis tools; Wrote the paper.Annalisa De Silvestri: Contributed data or analysis tools; Performed the analysis

      Conflict of interests

      The authors have no competing interests to declare.

      Acknowledgments

      We are pleased to thank, Alessia Artesani, Alessandra Balestra, Francesca Blaco, Maria Concetta Bonomo, Laura Buono, Costanza Caccia Dominioni, Veronica Di Gregorio, Jessica Esposito, Caterina Fontana, Tomasia Grilli, Silvia Grignaschi, Noemi Manca, Claudia Omes, Anna Pagani, Laura Piccolo, Eleonora Ragni, Rossana Totaro, and Paola Travaglino, for their support and intense cooperation for this study in these hard times and also as women involved in professional care for the others in their public and private life.

      Appendix. Supplementary materials

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