PLACENTAL TRANSFER OF HEPATITIS A ANTIBODIES IN FULL TERM AND PRETERM INFANTS

Abstract

Despite significant improvement in socioeconomic and hygienic standards in Israel, the incidence of hepatitis A infection continues to rise1 and yet the prevalence of hepatitis A antibodies among young adults has declined.2 Transplacental transfer of hepatitis A antibodies (anti-HAV) provides newly born infants with passive protection against infection; hence hepatitis A infection is a rare event during the first year of life. Nevertheless an increased prevalence has recently been reported in an affluent population located in endemic areas.3 Preterm infants may be considered a particularly vulnerable group because of their small amounts of passively acquired antibody.4 The purpose of this study was to investigate, in an endemic area, the passive protection afforded to newborns against hepatitis A infection and to compare full term and preterm infants. Materials and methods.Subjects. Two hundred fifty healthy, Israeli-born mothers and their newborns, delivered at the Chaim Sheba medical center, Tel Hashomer, Israel, participated in the study. They were divided into 2 groups: the full term group with 147 healthy newborns at 37 weeks of gestation or above (mean ± SD 39 ± 1.7 weeks gestation; birth weight 3160 ± 572 g), and the preterm group (n = 103) with a gestational age of <35 weeks (mean ± SD 32 ± 1.8 weeks; birth weight 1626 ± 352 g). (Infants of 36 weeks gestation were not included in the study.) The mean age of the mothers tended to be higher in the preterm group but the difference did not reach statistical significance (mean ± SD 28.7 ± 0.9 and 31.2 ± 1.2 years in the full term and preterm groups, respectively). Fifty-three percent of the preterm group were primigravidas, but in only 35.2% of the full term group was the infant the mother's firstborn. Maternal and umbilical cord blood samples were taken in all cases. The study was approved by the hospital's ethics committee and informed consent was obtained from each participant. Serology. The competitive enzyme-linked immunosorbent assay for total anti-HAV was carried out as described initially by Duermeyer.5 Immulon 1® microtiter wells (Dynatech Laboratories, Inc., Chantilly, VA) were adsorbed overnight at room temperature with 100 μl of anti-HAV antibodies. After adsorption each well was washed three times with phosphate buffer saline containing 0.5% Tween 20 (PBS-Tween). The wells were then filled with 50 μl of HAV antigen. Clarified lysate from fetal rhesus monkey kidney cells (FRhK-4) infected with HAV cytopathic variant (strain H-175) (Cromeans et al., 1987) (provided by Dr. Sobsey, University of North Carolina, School of Public Health, Chapel Hill, NC) was used as a source for HAV antigen in this test. After incubation at room temperature and three washings with PBS-Tween, 50 μl of serum samples in 10-fold dilutions (starting from 1:20) were added. After an incubation period of 1.5 h at 37°C and a washing with PBS-Tween, each well was filled with 50 μl of anti-HAV conjugated to horseradish peroxidase (Type VI; Sigma, St. Louis, MO). The plates were then incubated for 1.5 h at 37°C and washed five times, followed by the addition of 100 μl of substrate solution (o-phenylenediamine and H2O2). After reincubation for 15 to 30 min at room temperature in the dark, the enzyme reaction was stopped with 50 μl of H2SO4. The wells were read in an enzyme-linked immunosorbent assay reader at an optical density of 492 nm. Anti-HAV titers were initially expressed as the dilution giving a 50% blocking effect calculated by interpolation and then were recalculated and presented in international milliunits (ml) using a WHO reference anti-HAV gamma-globulin preparation. A titer of ≥1:20 mIU/ml was considered protective. Statistical analysis. Statistical analysis was performed using the biomedical computer program statistical software package. After applying log transformations to the titers, means and standard deviations were computed. Differences between groups were compared using one-way analysis of variance and Bonferroni's test for multiple comparisons. The geometric mean titers were derived by calculating 10x of the mean values after the log transformations. A paired Student's t test was used to compare the differences between maternal and cord blood titer transformations according to the log values. The proportions of immunized subjects were compared by the chi square test or Fisher's exact test. Correlation between maternal and cord blood titers was performed by linear regression analysis. Regression lines were compared by analysis of variance of regression coefficients over groups. Results. The incidence of protective hepatitis A antibody titers (≥1:20 mIU/ml) and their GMTs in mothers and both the full term and preterm infants, are presented in Table 1. There were similar incidences of protective titers in both the newborns and their mothers; all infants of mothers who had detectable titers of anti-HAV had similarly protective titers. The geometric mean titers against hepatitis A were somewhat lower in the preterm compared with the full term neonates (373 and 493, respectively), but the difference did not reach statistical significance. For the full term and preterm groups a statistically significant linear correlation was found between the maternal and neonatal antibody titers (r = 0.98, r = 0.99 for full term and preterm, respectively).TABLE 1: Percentage of infants with protective hepatitis A titers Analysis of variance of the regression coefficients over groups showed that the transfer of maternal antibodies to the full term neonate was similar to that in the preterm infant (Fig. 1).Fig. 1: Correlation of maternal and neonatal HAV antibody titers for full term and preterm infants.Discussion. Despite considerable improvements in the socioeconomic and hygienic conditions of the Israeli population, the incidence of hepatitis A continues to rise1; however, the prevalence of hepatitis A antibodies among young adults has declined.2 Our study shows that approximately one-half of the mothers and their infants born in an urban area in Israel, a country endemic for hepatitis A, have protective concentrations of HAV antibody. The subsequent decline in HAV antibody titers is another important issue that is being evaluated. Knowledge of the duration of persistence of maternal antibodies in the infant will have significant implications in developing a hepatitis A vaccination policy. A recent study comparing the pattern of anti-HAV prevalence in new immigrants from the former USSR with that in the Israeli population found that ∼90% of young adults in Israel have hepatitis A antibodies.6 This difference probably reflects the higher socioeconomic status of the population in the present study. Our study reconfirms the passive protection afforded the newly born against hepatitis A infection. As in previous reports7 we were able to demonstrate a positive correlation between maternal and neonatal antibody titers. Since preterm infants are a vulnerable group because of low amounts of passively acquired antibodies,4 it was of particular interest to measure their level of protection against hepatitis A infection. Our results indicate that transplacental transfer of hepatitis A antibodies to preterm infants does not differ from that observed in full term infants. This is in contrast to the situation with poliovirus, for which we have previously shown diminished transplacental transmission of maternal antibodies in preterm newborns.8 At present we cannot offer an explanation for this discrepancy. Acknowledgments. We are grateful to Mrs. Elsa Katz for her support, to Mrs. Pnina Lilius of the department of statistics at Tel-Aviv University for performing the data analysis and to Mrs. Nina Golubev of the Central Virology Laboratory for excellent technical assistance. Nehama Linder, M.D. Yuori Karetnyi, M.D., Ph.D. Yariv Gidony, M.D. Gonen Ohel, M.D. Eilona Levin, M.D. Jacov Kuint, M.D. Nadav Davidovich, M.D. Ifat Gidony, M.D. Ella Mendelson, Ph.D. Asher Barzilai, M.D. Department of Neonatology (NL, YG, GO, EL, JK, ND, IG, AB) and Central Virology Laboratory (YK, EM); Chaim Sheba Medical Center; Tel-Hashomer; Sackler School of Medicine; Tel-Aviv University; Tel-Aviv Department of Obstetrics and; Gynecology (GO); Poriah Hospital; Tiberias; Israel