Alum-induced chemokine production with increasing age in infancy.

<p>Whole blood samples from PNG infants aged 1–3 months (n = 10, <i>white bars</i>), 4–6 months (n = 9, <i>light grey bars</i>), 7–12 months (n = 10, <i>dark grey bars</i>) or 13–18 months (n = 9, <i>black bars</i>) were stimulated with Alum. Presented are the geometric means and 95% confidence intervals (pg/mL) for each age group for background-adjusted chemokine responses. Significance level is indicated where p<0.10.</p

Pneumococcal serotype-specific memory B-cell responses.

<p>Serotype-specific Antibody Forming Cells (AFCs) per 1x10⁶ cultured PBMCs were measured at 10 months of age (1 month after PPV23 vaccination) and pre-challenge at 3–5 years of age in a subset of children vaccinated with PCV7 and PPV23 (white bars) or PPV23-only (grey bars), and pre-challenge at 3–5 years of age in a subset of children not vaccinated with any pneumococcal vaccines (striped pattern). Serotypes with an asterisk (*) are included in PPV23 only, while other serotypes are included in both PCV7 and PPV23. Data are presented as means and 95% confidence intervals. No significant differences were found between groups at 10 months or 3–5 years of age (tested using Mann-Whitney U test). Differences in responses at 10 months versus 3–5 years of age within a group were tested using Wilcoxon Signed Rank Test, and where significant differences were found this has been indicated in the graph.</p

Infant characteristics.

<p>Infant characteristics.</p

Pneumococcal serotype-specific IgG antibody titers before and after challenge.

<p>Serum antibody titers were assessed in 3-5-year-old children who had received PCV7 and PPV23 as infants (white bars; n = 100 /n = 86), who had received only PPV23 as infants (grey bars; n = 32 /n = 28), or who had not received pneumococcal vaccines (striped bars; n = 121 pre-challenge/ n = 98 post-challenge) (A) before and (B) after challenge with a low dose of PPV23. Data are presented as geometric mean titers and 95% confidence intervals. Serotype-specific geometric mean titers before or after challenge were compared between each group using Mann-Whitney U test. * p < 0.05.</p

Generalised estimating equations studying associations between change in serotype-specific antibody titers in response to challenge and pneumococcal vaccine history and pre-challenge antibody titers.

<p>Generalised estimating equations studying associations between change in serotype-specific antibody titers in response to challenge and pneumococcal vaccine history and pre-challenge antibody titers.</p

Proportion of 3-5-year-old children with pneumococcal serotype IgG antibody titers ≥ 0.35 μg/ml and ≥ 1 μg/ml before challenge.

<p>Proportion of 3-5-year-old children with pneumococcal serotype IgG antibody titers ≥ 0.35 μg/ml and ≥ 1 μg/ml before challenge.</p

Individual pneumococcal serotype-specific IgG antibody titers following PPV23 vaccination at 9 months versus low-dose challenge at 3–5 years of age.

<p>Graphs present scatterplots of pneumococcal serotype-specific IgG antibody titers measured in individual children 1 month after PPV23 vaccination (vaccinated at 9 months of age) (x axis) and 1 month after challenge with a low dose of PPV23 at 3–5 years of age (y axis), in those who had received 3 doses of PCV7 before the PPV23 vaccine (primed; grey circles), or only received PPV23 (unprimed; black diamonds). Serotypes with an asterisk (*) are included in PPV23 only, while the other serotypes are included in both PCV7 and PPV23. The p-values included in the graphs correspond with non-parametric analysis of paired responses within the treatment groups (Wilcoxon Signed Rank Test).</p

Flowchart for children in the study.

<p>A flowchart of the completed preceding neonatal PCV7 trial is published elsewhere [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0185877#pone.0185877.ref008" target="_blank">8</a>]. A total of 132 of 259 children who participated in the previous PCV7 study and who were vaccinated with PPV23 at 9 months of age were included in this study. One hundred and twenty seven children could not be enrolled for reasons summarized in Fig 1, including death; being too old according to pre-defined inclusion criteria; illness; migration to an area outside the study’s reach capacities; not located; and refusal. A total of 121 of 136 potential community controls who were assented to participate in the study were included in the final analysis: of the 15 not included, three children were over age; three did not consent; three could not be relocated and blood collection pre-challenge was not successful for six children. Post-challenge data were not available for 41 children for reasons summarized. <i>N</i> is Neonatal PCV7 group; <i>I</i> is Infant PCV7 group; <i>C</i> is control group (no PCV7).</p

Maturation of innate immune function in PNG infants.

<p>Whole blood samples from PNG infants aged 1–3 months (n = 18, <i>white bars</i>), 4–6 months (n = 18, <i>light grey bars</i>), 7–12 months (n = 21, <i>dark grey bars</i>) or 13–18 months (n = 10, <i>black bars</i>) were stimulated with TLR (LTA; PolyIC; LPS; Gardiquimod) and NLR (iE-DAP; MDP) ligands, and Alum alone or with LPS co-stimulation (denoted by ♦). Presented are the geometric means and 95% confidence intervals (pg/mL) for each age group for background-adjusted cytokine responses. Significance level is indicated where p<0.05.</p

Innate TNF-α responses in relation to increasing age in infancy.

<p>Presented are the geometric means and 95% confidence intervals (pg/mL) for background-adjusted TNF-α responses. Mann-Whitney U tests for significant differences in log-transformed TNF-α levels compared to the “1–3 months” age group; and Spearman rho tests for significant correlations between log-transformed TNF-α levels and ordered age groups were conducted. Significance level is indicated where p<0.05 only (<b>*</b> p = 0.049).</p>♦<p>denotes LPS co-stimulation used.</p