FIG 3.

Microbial biomarkers and breastmilk feeding in early preterm subjects with high and low IP. (A) Abundance of bacterial groups stratified by postmenstrual age at study days 7 to 10. The results indicate that the Actinobacteria (Bifidobacterium) and Clostridia (Clostridiales) were observed mainly in low-IP subjects but not in high-IP subjects (red). The abundance values of read counts for each ASVs are stacked in order from highest to lowest, separated by a horizontal line. (B) Box plot of Bifidobacterium relative abundance and the cumulative amount of mother’s breastmilk feeding (milliliters per kilogram) during the first 7 to 10 postnatal days in subjects with high or low IP. IP was calculated using the ratio of urine lactulose (La) to rhamnose (Rh), with low or high IP defined by an La/Rh ratio of >0.05 or ≤0.05, respectively. Plotted are interquartile ranges (IQRs) (boxes), medians (lines in boxes), and means (diamonds). Significance values were calculated using a Wilcoxon rank sum test. * denotes the level of significance. NS, nonsignificant. (C and D) Volatility plots demonstrating the fluctuation of microbial community diversity (characterized by the Shannon diversity index) (C) and Bifidobacterium diversity over mother’s own breastmilk (MOM) feeding volumes in high- or low-IP groups (D). The plot was generated in QIIME (October 2019 version) (106). Nonoverlapping vertical error bars at each measuring point were considered significantly different. (E) Temporal characterization of the intestinal microbiota of early preterm infants with profile changes over the first 21 days after birth. The taxonomic profile was generated using 16S rRNA gene sequencing. Community type is shown in heatmap clusters in Fig. S3 in the supplemental material. The dates when IP was measured, MOM, pasteurized human donor’s milk (PHDM), formula feeding day, and antibiotic administration are shown in the plots. Each circle is sized proportionally to the feeding volume. Abbreviations: BW, body weight; F, female; M, male; CS, cesarean section; SVD, spontaneous vaginal delivery; AA, African American; Ap1, Apgar score 1 minute category; Ap5, Apgar score 5 mintues category; GA, gestational age; PMA, postmenstrual age; BW, birthweight.