Richard Puleston has received funding from Sanofi Pasteur to attend two influenza-related meetings. He will also be providing UNPAID consultancy advice to Novartis. Richard Puleston, Jonathan Nguyen Van Tam, Iain Stephenson and Karl Nicholson have received funding from GlaxoSmithKline to support virus isolation studies as a subsidiary (ongoing) component of this study. Joanne Enstone has received a one-off consultancy fee from GlaxoSmithKline and funding to attend an educational meeting from Sanofi Pasteur. Iain Stephenson has received funding to attend international scientific meetings (Novartis Vaccines), lecture and consultancy fees (Novartis, GSK and Baxter) and research funding from several pharmaceutical companies including Novartis Vaccines, GSK, Baxter Vaccines and Hoffman-La Roche. Maria Zambon and Katja Hoschler have been investigators of clinical trials sponsored by Novartis, Baxter, Sanofi Pasteur, and CSL Australia Ltd. Katja Hoschler has been sponsored by Sanofi Pasteur to take part and speak at one international meeting. Karl Nicholson has been an ad hoc consultant to GlaxoSmithKline, Merck, Sanofi Pasteur, and Novartis. He has received funding to speak at meetings organised by Novartis, Baxter, Berna Biotech, Esteves, and the European Scientific Working Group on Influenza, and H5 vaccines from Novartis to support an MRC-funded research project, and H1N1 vaccines from Baxter AG and GlaxoSmithKline to support an National Institute for Health Research-funded research project. Jonathan Nguyen Van Tam has received funding to attend influenza-related meetings, ad-hoc lecture and consultancy fees and research funding from several influenza antiviral drug and vaccine manufacturers (including both GlaxoSmithKline and Baxter AG), and is a former employee of SmithKline Beecham PLC (now GlaxoSmithKline), Roche Products Ltd, and Sanofi Pasteur MSD. The other authors have for themselves declared that no competing interests exist. This does not alter the authors' adherence to all the PLOS ONE policies on sharing data and materials.
Conceived and designed the experiments: RP JT JVT IS KN MZ PM. Performed the experiments: RP GB JT YD KH MZ JK. Analyzed the data: RP KN. Wrote the paper: RP JVT KN PM GA MZ KH JT JE.
Illness and death from influenza increase during pregnancy. In the United Kingdom pregnant women were targeted in a national programme for vaccination during the H1N1 2009–10 pandemic.
In this study, pregnant women were recruited in labour from November 9, 2009 to March 10, 2010. Pandemic vaccination status was determined. Venous cord blood collected at delivery was evaluated for transplacental transfer of antibodies by measurement of haemagglutination inhibition and microneutralization titres.
Samples were collected from 77 vaccinated and 27 unvaccinated women. Seroprotection (HI titre ≥1∶40) was detected in 58 (75.3%, 95% CI 64.2–84.4) cord blood samples from vaccinated women and 5 (18.5%, 95% CI 6.3–38.1) from unvaccinated women (P<0.0001). There was evidence of transplacental seroprotection 8 days after maternal immunization (77.9%, 95 CI 66.2–87.1), maintained in most cases for at least 16 weeks.
Immunization of pregnant women with AS03A-adjuvanted vaccine is followed by transplacental transfer of passive immunity at titres consistent with clinical protection in three-quarters of new-born infants. The findings support national and international pandemic H1N1 2009 recommendations for immunization during pregnancy.
Influenza infection during pregnancy is associated with increased rates of severe illness, hospitalization and death. The risks to mother and child increase as pregnancy progresses, both during seasonal and pandemic influenza, including the recent H1N1 2009 pandemic.
Infants under 6 months of age experience high rates of influenza and have the highest rates of hospitalization of any age group (other than the over 65 s),
The H1N1 2009 pandemic was the first time when nationally and internationally an adjuvanted monovalent influenza vaccine was advocated during pregnancy. Our aim was to evaluate whether immunization during pregnancy with one 3.75 µg dose of AS03A-adjuvanted split-virion inactivated influenza A/California/7/2009 H1N1 vaccine raised transplacental antibody titres to levels consistent with protection. Our study was undertaken during the second wave of the H1N1 2009 pandemic in the UK, so we were able to compare cord blood samples from vaccinated and unvaccinated women.
From November 2009 to March 2010, we undertook an observational study at three sites in the UK (Queen's Medical Centre, Nottingham; City Hospital, Nottingham; and Leicester Royal Infirmary, Leicester) investigating the transfer of immunity to babies born to women immunized/unimmunized with monovalent AS03A adjuvanted H1N1 2009 vaccine (Pandemrix®: GSK Biologicals.) as part of the UK national pandemic vaccine program. Participants were not vaccinated as part of the study.
Pregnant women normally resident in the East Midlands who presented for delivery beyond the first trimester were eligible for participation. Women were recruited regardless of age, social class, ethnicity, previous pregnancy and childbirth status, past and current medical history (including current medications), ethnicity, mode of delivery and outcome of the pregnancy. The main exclusions were uncertain vaccination status (H1N1 2009 vaccine), being held in legal custody, participation in another clinical study, non-residence in the East Midlands (UK) and clinical situations requiring cord blood. We included vaccinated women regardless of the interval between immunization and delivery.
All participants provided informed consent. The Leicester, Northamptonshire and Rutland Ethics Committee and participating hospitals approved the study.
Potential participants were approached for consent during the admission for delivery. We recorded participants' vaccination status against H1N1 2009 virus and collected basic demographic data, medical, pregnancy and childbirth history and the method of delivery and outcome of the pregnancy. The date of immunization and vaccine batch number were obtained from primary care records. Subjects who received vaccine before the date of delivery were considered ‘vaccinated’ and those who were vaccinated on the day of delivery or afterwards (or never vaccinated) were classified as ‘unvaccinated’.
We took venous cord blood samples at childbirth for antibody titration against NIBRG-121 virus (generated from A/California/7/2009 and A/PR/8/34 strains by reverse genetics). Sera were separated and stored at −20°C until transfer to the Respiratory Virus Unit, Centre for Infections (Health Protection Agency, UK) for serological analysis. Antibodies were titrated by haemagglutination-inhibition (HI) assay with standard methods, as reported previously.
The primary endpoint was the proportion of participants with haemagglutination inhibition titres of 1∶40 or greater. This corresponds with a 50% or greater reduction in the risk of contracting an influenza illness in a susceptible adult population,
All statistical analyses were performed in Stata (StataCorp Inc., version 11). Baseline characteristics of vaccinated and unvaccinated mothers were summarized using un-paired t- and rank sum tests for normally and non-normally distributed continuous data respectively and χ2 test for categorical data. The GMTs of each sample's duplicate HI and MN tests were transformed into binary immune/non-immune status using cut-off titres of 1∶40 or greater for HI and 1∶60 or greater for MN. GMTs of HI and MN antibodies were compared using two group mean comparison t-test of log10-transformed titres. We compared proportions of participants that achieved ‘seroprotection’ by χ2 test. Exact (Clopper–Pearson) CIs are reported for all proportional endpoints. No formal adjustments for multiple testing were done. We assessed the role of possible confounding covariates on immunity, including the duration of ‘exposure’ (defined as the interval between the first UK case of pandemic H1N1 2009 infection and date of vaccination plus 10 days (notional time to seroconversion) for vaccinees, or the date of childbirth for non-vaccinees), gestational age and birth-weight, by multivariable logistic regression.
Investigators in the maternity units could not be blinded to the vaccination status of participants. Cord blood samples were labelled with each participant's unique code and laboratory staff titrated the specimens without knowledge of vaccination status.
From Nov 18 2009 to Mar 20 2010, 117 eligible women were approached and 104 were enrolled: 77 participants were vaccinated before the date of delivery with one 3.75 µg dose of AS03A-adjuvanted split-virion inactivated influenza A/California/7/2009 H1N1 vaccine and 27 were unvaccinated. Cord blood samples were collected at birth from all participants. The gestational age of the babies ranged from 34 to 42 weeks. The date of vaccination was available for 74 of 77 vaccinees; vaccine was administered a median of 42 days (range, 1–108 days) before delivery. Infants whose mothers were vaccinated were delivered from Nov 25 2009 to Mar 12 2010 and those whose mothers were unvaccinated were delivered from Nov 18 2009 to Jan 26 2010. Baseline demographic characteristics of the two groups were similar (
Vaccine recipients (n = 77) | Unvaccinated (n = 27)) | p | |
|
31 (29–32) | 29 (26–31) | 0.1413 |
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White | 67 (89) | 25 (93) | 0.6253 |
Other | 8 (11) | 2 (7) | |
|
|||
1 | 35 (46) | 10 (37) | 0.4172 |
2 or more | 41 (54) | 17 (63) | |
|
|||
0 | 45 (59) | 16 (59) | 0.9965 |
1 or more | 31 (41) | 11 (41) | |
|
|||
0 | 67 (87) | 22 (81) | 0.4815 |
1 or more | 10 (13) | 5 (19) | |
|
|||
0 | 50 (65) | 16 (59) | 0.5982 |
1 or more | 27 (35) | 11 (41) | |
|
|||
Normal, assisted, elective or caesarean unspecified | 67 (88) | 22 (88) | 0.9831 |
Emergency caesarean section | 9 (12) | 3 (12) | |
|
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Male | 38 (50) | 16 (59) | 0.4079 |
|
3365 (3223–3507) | 3600 (3400–3799) | 0.0848 |
|
39.3 (38.9–39.7) | 40.1 (39.5–40.7) | 0.0612 |
|
|||
0 | 43 (57) | 17 (63) | 0.6103 |
1 or more | 32 (43) | 10 (37) | |
|
|||
0 | 54 (82) | 16 (70) | 0.2169 |
1 or more | 12 (18) | 7 (30) |
Data are means for: years/birth weight/gestational age (95% confidence interval) or number (%) for all other parameters.
(Titres are expressed as reciprocal of the dilution and are given on a log2 scale.)
(Titres are expressed as reciprocal of the dilution and are given on a log2 scale.).
Duration of antibody titre protection has also been estimated.
(Titres are expressed as reciprocal of the dilution and are given on a log2 scale.).
The relationship between HI and MN titres is displayed in
Examination of potential (
Our results show that antibodies against H1N1 2009 pandemic virus are present in cord-blood samples from substantially more vaccinated than unvaccinated women and at much higher titres. Our findings are particularly relevant given the substantial impact of H1N1 2009 virus on pregnant women
In our study over 90% of cord-bloods from vaccinated mothers had HI titres of ≥1∶8 and MN titres of ≥1∶10. These high sero-prevalence rates (>90%) contrast with levels of 37% for the presence of HI and MN antibodies in cord-bloods from non-vaccinated women. This ‘background rate’ of 37% for both HI and MN antibody in cord blood from non-vaccinated mothers is comparable to sero-incidence rates of 10–40% found in venous blood collected across England from people aged 25- to 44-years during October 2009–February 2010.
Although there are no established correlates of protection for HI or MN in children and infants, evidence is accumulating that antibodies against seasonal influenza viruses, at levels that are normally associated with protection, can be found in a high proportion of cord blood samples
There is evidence that maternal immunization with seasonal influenza vaccine prevents laboratory-confirmed influenza in infants,
A key finding in our study was the appearance of antibody in cord blood shortly after maternal immunization. By day 8, 97.1% and 95.5% of cord bloods respectively contained HI and MN antibody against pandemic H1N1 2009 virus, 77.9% of cord bloods had ‘seroprotective’ HI titres of ≥1∶40 and 86.6% had MN titres of ≥1∶60. The trend-line fitted to the scatter-chart of HI antibody titres in relation to time between vaccination and delivery (
We found no evidence for an effect of other potential confounders on the observed antibody responses. It should be noted that we only studied cord antibody levels in response to AS03A-adjuvanted H1N1 2009 vaccine, so it is unclear whether conventional (i.e., unadjuvanted) split and subunit vaccines, whole virus vaccine, or vaccine with alternative adjuvants trigger H1N1 2009 HI and MN antibody as rapidly in cord blood as in our study, or to the same high titres. (Although two brands of vaccine were available for use in the UK during the pandemic, only the AS03A-adjuvanted H1N1 2009 vaccine was recommended for use in pregnant women).
This was an ‘emergency’ pandemic research study, designed and executed in a very short time frame and as such there are a number of limitations. Comparison of cord sample antibody titres with maternal serum antibody titres would have provided additional detail; however, maternal serum was not obtained. Older school age children in the household/or those attending a child care facility and previous maternal seasonal influenza vaccination could have affected the results however, these data were not collected and seasonal influenza vaccine was not recommended in the UK for pregnant women until the 2010–11 winter season, immediately post-pandemic. In order to check for recruitment bias it would have been useful to know how many eligible subjects refused consent or declined involvement; however for logistical reasons this information was not available. Ideally the mothers and children in the study would have been followed up by taking serial serological samples to assess the duration of antibody persistence. This was considered, but dismissed as being impractical. The babies have been followed up separately to assess the clinical protective effect of the maternally transferred immunity. This separate element of the study was designed to obtain nasal mucous samples (for determination of the presence of influenza virus PCR) from babies in the original cohort if a respiratory illness occurred in the follow up period. This secondary objective study recently completed and will be reported separately.
This study was not powered to test for safety issues relating to the novel vaccination using an AS03A-adjuvanted H1N1 2009 vaccine. Nonetheless no serious adverse events were reported to us during the study.
Our study focused on the presence of HI and MN antibody but did not assess clinical effectiveness. The increased rates of illness and death from pandemic influenza during pregnancy make randomised placebo-controlled trials of the efficacy of immunization during pregnancy with H1N1 2009 vaccine unethical, especially as the WHO and national authorities, including the United Kingdom Departments of Health, recommended vaccination during pregnancy as a priority during 2009–10 (pandemic vaccine) and 2010–11 (seasonal trivalent vaccine). An insight into the protection afforded to mothers and their children by vaccination of pregnant women with H1N1 2009 vaccine could perhaps be obtained from effectiveness studies in countries with large databases that capture all relevant information in primary and secondary healthcare settings. Meanwhile, we consider the decision by the Departments of Health in the UK to target pregnant women for vaccination with an AS03A-adjuvanted vaccine was justified from the perspective of the serological protection conferred both to the mothers and their babies. However, we have no equivalent data for the use of non adjuvanted, trivalent seasonal vaccine in pregnancy and so cannot generalize further.
Our work strengthens the need for a better understanding of the relationship between measurable antibody and protective immunity in infancy. Future comparative studies of different vaccines administered during pregnancy to inform vaccine policy are also justified.
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We thank the women who agreed to take part in this study. We gratefully acknowledge administrative assistance given by: the National Institute of Health Research; the Leicestershire, Northamptonshire & Rutland Research Ethics Committee 1; Division of Epidemiology and Public Health at Nottingham University; Trent Comprehensive Local Research network (CLRN); staff at Nottingham University Hospitals NHS Trust and Leicester University Hospitals NHS Trust; University of Nottingham Research Innovation Services; Mrs Sharon Figgens, University of Nottingham, Division of Epidemiology and Public Health; Health Protection Agency staff at the Centre for Infections, Colindale, London.