Pertussis Maternal Immunization: Narrowing the Knowledge Gaps on the Duration of Transferred Protective Immunity and on Vaccination Frequency

Abstract

Maternal safety through pertussis vaccination and subsequent maternal-fetal-antibody transfer are well documented, but information on infant protection from pertussis by such antibodies and by subsequent vaccinations is scarce. Since mice are used extensively for maternal-vaccination studies, we adopted that model to narrow those gaps in our understanding of maternal pertussis immunization. Accordingly, we vaccinated female mice with commercial acellular pertussis (aP) vaccine and measured offspring protection against Bordetella pertussis challenge and specific-antibody levels with or without revaccination. Maternal immunization protected the offspring against pertussis, with that immune protection transferred to the offspring lasting for several weeks, as evidenced by a reduction (4-5 logs, p < 0.001) in the colony-forming-units recovered from the lungs of 16-week-old offspring. Moreover, maternal-vaccination-acquired immunity from the first pregnancy still conferred protection to offspring up to the fourth pregnancy. Under the conditions of our experimental protocol, protection to offspring from the aP-induced immunity is transferred both transplacentally and through breastfeeding. Adoptive-transfer experiments demonstrated that transferred antibodies were more responsible for the protection detected in offspring than transferred whole spleen cells. In contrast to reported findings, the protection transferred was not lost after the vaccination of infant mice with the same or other vaccine preparations, and conversely, the immunity transferred from mothers did not interfere with the protection conferred by infant vaccination with the same or different vaccines. These results indicated that aP-vaccine immunization of pregnant female mice conferred protective immunity that is transferred both transplacentally and via offspring breastfeeding without compromising the protection boostered by subsequent infant vaccination. These results-though admittedly not necessarily immediately extrapolatable to humans-nevertheless enabled us to test hypotheses under controlled conditions through detailed sampling and data collection. These findings will hopefully refine hypotheses that can then be validated in subsequent human studies.

Keywords: Bordetella pertussis; acellular vaccine; pertussis; pregnancy immunization; protection.

Figure 1

Effect of maternal immunization on protection of offspring against Bordetella pertussis infection. (A) Schematic representation of the maternal vaccination and challenge protocols. Female mice were vaccinated twice with a commercial acellular pertussis (aP) at a 2-week interval; mated with male mice; and when the vaginal plug was detected, a third aP dose was administered. Non-immunized females were mated with male mice at the same time as the immunized females. Mice were born 3–4 weeks after the last immunization. Mice born to aP-immunized or non-immunized females (controls) were challenged with B. pertussis at 4 and 16 weeks after birth. Mothers and pups were bled at different time points as indicated by the dotted vertical arrows. (B) Short- and long-term protection of offspring through maternal immunization. Mice born to aP-immunized (Ipups n = 6, black bars) or non-mmunized (Cpups n = 6, white bars) females were challenged with B. pertussis Tohama I at 4 and 16 weeks after birth. The number of bacteria recovered from mouse lungs, expressed as the log₁₀ of CFUs per lungs, is plotted on the ordinate for the times after birth in weeks indicated on the abscissa, with the data representing the means ± the SD. The dotted horizontal line indicates the lower limit of detection. *p < 0.001 Ipups versus Cpups. (C) The anti-whole-cell-B. pertussis (a-Bp)- and anti-pertussis toxin (a-PTxA)-specific IgG titers were determined in the mother and in the offspring at the indicated time points. The titers are expressed as the geometric mean of the data from each group. p < 0.001 a-PTxA and a-Bp IgG titers in Ipups at 4 weeks after birth versus those detected at 16 weeks after birth. The results from one representative experiment are shown. nd, not detected.

Figure 2

Duration of protective immunity conferred by maternal immunization beyond the first pregnancy. (A) Schematic representation of the maternal vaccination and pup-challenge protocol. Female mice were vaccinated twice with commercial acellular pertussis (aP) at a 2-week interval; mated with male mice; and when the vaginal plug was detected, a third aP dose was administered. Non-immunized females were mated with male mice at the same time as the immunized females. The first birth was 3–4 weeks after the last immunization. Successive births occurred at intervals of 5–8 weeks. Mice born to aP-immunized or non-immunized females (the negative controls) were challenged with Bordetella pertussis at 4 weeks after birth. Mothers and pups were bled 1 day before pup challenge as indicated by the dotted vertical arrows. (B) Protection conferred to the offspring born in later pregnancies by maternal immunity acquired during the first pregnancy. Mice born to aP-immunized (Ipups n = 5) or non-immunized (Cpups n = 5) females were challenged with B. pertussis Tohama I 4 weeks after birth. The protection conferred to the offspring through maternal immunization during the first pregnancy was estimated by determining the number of bacteria recovered from mouse lungs. The bacterial counts expressed as the log₁₀ of CFU per lungs, is plotted on the ordinate as a function of the number of births after the initial delivery indicated on the abscissa. *p < 0.001 Ipups versus Cpups. The anti-pertussis toxin (a-PTxA) IgG titers in the mothers and the offspring determined at the time of challenge are listed below the abscissa. The titers are expressed as the geometric mean of the data from each group (n = 5). p < 0.001 a-PTxA IgG titers in Ipups from the first and second deliveries versus those detected in Ipups from the third through the fifth deliveries. The results from one representative experiment are presented.

Figure 3

Effect of passive immunization on protection through sera collected from Ipups. Pooled sera from either early pregnancy Ipups with high anti-pertussis toxin (a-PTxA) titer or late-pregnancy Ipups with low a-PTxA titer were tested by transfer to naive (Cpup n = 5) female mice. Pooled sera from Cpups transferred to naive female mice were used as negative control of protection. Twenty-four hours after transfer, the mice were infected with Bordetella pertussis and sacrificed 7 days later to determine the CFUs in the lungs. In the figure, the number of bacteria recovered from mouse lungs, expressed as the log₁₀ of CFUs per lungs, is plotted on the ordinate for each of the two serum-donor groups (early- and late-pregnancies) indicated above the bars. *p < 0.05 Ipup serum versus Cpup serum. The IgG a-PTxA titers are listed for each pool below the abscissa (nd, not detected).

Figure 4

Effect of passive immunization through spleen cells collected from Ipups on protection from infection. Whole spleen cells (20–50 × 10⁶) from Ipups or Cpups (controls) were tested as possible vehicles of immune protection by passive transfer to naïve mice. Twenty-four hours after transfer, the mice were infected with Bordetella pertussis and sacrificed 7 days later to determine the CFUs in the lungs. Ipup-recipients were used as a positive control. In the figure, the number of bacteria recovered from the mouse lungs, expressed as the log₁₀(mean CFUs ± SD of per lungs), is plotted on the ordinate for each of the spleen-cell-recipient groups (n = 7) indicated on the abscissa. The dotted horizontal line marks the lower limit of detection. *p < 0.05 Ipups versus both spleen-cell recipient groups.

Figure 5

Effect of passive immunization through lactation on pertussis protection. Pups born from immune mothers (Ipups) were fostered to non-immune mothers (Ipups–Cmother), and pups born from control mothers were fostered to immune mothers (Cpups–Imother). Ipups breast-fed by their immune mother (Ipups–Imother) and Cpups breast-fed by their non-immune mother (Cpups–Cmother) were used as the respective positive and negative controls. All the pups (n = 6 for each group) were suckled for 21 days. (A) Sera from mothers and pups were collected at different time points to analyze the titers of anti-pertussis toxin (a-PTxA) and anti-whole-cell-B. pertussis (a-Bp) IgGs. The titers are expressed as the geometric mean of the data for each group (n = 6). (B) Pups were challenged with a sublethal dose of Bordetella pertussis Tohama phase I to analyze protection. In the figure, the number of bacteria recovered from the mouse lungs, expressed as the log₁₀(mean CFUs ± SD of per lungs), is plotted on the ordinate for each of the dam-suckling groups indicated on the abscissa. The dotted horizontal line marks the lower limit of detection. Two independent experiments were performed. *p < 0.05 for Ipups–Imother and Cpups–Imother versus the other groups and for Ipups–Cmother versus Cpups–Cmother.

Figure 6

Effect of heterologous mother–pup vaccination on protection against Bordetella pertussis challenge. (A) Schematic representation of vaccination and challenge protocols. Female mice were immunized with three doses of commercial aP vaccine on days 0 and 14; mated with male mice; and when the vaginal plug was detected, a third aP dose was administered. Four weeks after birth, the offspring received either no vaccine (n = 8) or one dose of the aP, OMV, or wP vaccine (n = 8 in each group), as indicated below the abscissas of panels (B,C). Two weeks later, the pups were challenged with B. pertussis and the bacterial burden in the lungs measured 7 days after challenge. Pups born to vaccinated or non-vaccinated mothers and receiving no postnatal vaccine served as controls. The pups (n = 5 for each group) were bled and the anti-pertussis toxin (a-PTxA) IgG titers determined 1 day before the challenge as designated by the dotted vertical arrow. (B) The titers, plotted on the ordinate, are expressed as the geometric mean of the data from each group specified on the abscissa. *p < 0.05 pups born to immunized mother versus others groups, and the nd signifies not detected. (C) In the figure, the number of bacteria recovered from the mouse lungs, expressed as the log₁₀(mean CFUs ± SD per lung), is plotted on the ordinate for each group indicated on the abscissa. *p < 0.05 for negative control versus other groups, and aP-immunized pups born to non-immunized mother versus others groups. The dotted horizontal line marks the lower limit of detection. Of the three independent experiments that were performed, the results from a single representative one is presented.