Figure 1. BugFACS-based analysis of the development of gut mucosal IgA responses in healthy USA twin pairs.

a, Bubble plots for OTUs significantly enriched in either the IgA⁺ or IgA⁻ fraction across twin microbiota samples representing at least three age bins (n=534 fecal samples analyzed; 6.7±1.7 (mean±SD) samples/individual from the 1-,3-,6-,9-,12-,15-,18-,21-, and 24-month age bins). The size of the circle indicates the magnitude of enrichment (mean IgA index value) in either fraction. Color intensity indicates the statistical significance, as judged by a Wilcoxon signed-rank test. Gray indicates that a taxon was not detected in any sample at that time point. Taxa are ordered by their mean IgA indices defined across all time points. The rank order of feature importance for OTUs in the RF-derived models of ‘gut microbiota maturation’ and ‘gut mucosal IgA response maturation’ is indicated. The upper bracket to left of the figure encompasses OTUs identified as consistently IgA-targeted after the first three postnatal months, while the lower bracket highlights OTUs designated as consistently not targeted in multiple age bins after the first 3 postnatal months. b, Pearson correlation coefficients were used to compare the similarity of IgA index profiles of twins and unrelated children within a given age bin (n=9 total age bins). The correlation coefficient between members of a twin pair in a given age bin was compared to the mean of the correlations between that pair and all other age-matched unrelated children in the cohort (mean+SEM). Statistical significance was defined with the paired Wilcoxon test. *, P< 0.05; **, P< 0.01; ***, P< 0.001. c. Samples from mothers of the twin pairs were collected during the peripartum period, at 6 months after birth, and at ≥12 months after birth. Samples (n=106) were subjected to BugFACS and analyzed as above. d. Modeling development of gut IgA responses in young gnotobiotic mice colonized with the fecal microbiota from two twin pairs and fed infant formula and complementary foods. Bubble plots for OTUs shown in panel a that were present in BugFACS fractions prepared from the fecal microbiota of mice harboring transplanted fecal microbiota from 6- and 18-month old co-twins belonging to the two twin pairs (n=4–5 mice per group, total of 143 fecal samples analyzed). Data were pooled from the four indicated time points in the diet oscillation. e, Experimental design for gnotobiotic mouse experiment. f, ‘Age’ of the IgA responses documented in mice as a function of donor microbiota and diet. The RF model of human gut mucosal IgA response maturation was applied to BugFACS datasets generated from mouse samples. Data are averaged within recipients of twin pair microbiota from a given age bin (n=4–5 mice per group, 143 fecal samples analyzed). Mean values ± SEM are plotted. *, P<0.05; **, P<0.01; ***, P<0.001; ****, P<0.0001 (Mann-Whitney U test for the indicated comparisons).