Fig 1.
Study subject enrollment flow chart.
Diagram describing the number of participants screened, randomized into treatment groups, screen-failed, withdrew throughout the study period and included in the final analysis.
Table 1.
Participant demographics at baseline.
Fig 2.
Diagram describing the study design including outcomes measured, timing and duration of the study period.
Fig 3.
Diagram describing work flow for bovine colostrum product (Imucon) production. The dried Imucon product (powdered BCP) was produced from the commercially available liquid Imucon product (liquid BCP) for specific use in the current study.
Table 2.
Composition of liquid bovine colostrum product.
Table 3.
Confidence intervals and p-values for gastrointestinal and behavioral questionnaire data.
Fig 4.
Proportion of normal stools with treatment.
Mean ± SD proportion of total recorded stools that were of normal consistency (4 on Bristol Stool Scale), based on stool log data (n = 8 for each group). Significant differences in means (p<0.05) are denoted by an asterisk. D123, days 1,2 and 3 of the study period (baseline); D835, days 8 through 35 of the study period.
Table 4.
Descriptive statistics for stool consistency from stool log data.
Fig 5.
Change in gastrointestinal symptom frequency with treatment.
Mean ± SD change in parental reported frequency of gastrointestinal symptom after treatment based on Likert-type scale where 0 = symptom never occurs, 1 = symptom rarely occurs, 2 = symptom sometimes occurs, 3 = symptom often occurs or 4 = symptom always occurs (n = 7 for diarrhea, n = 5 for constipation, n = 5 for pain and n = 6 for gas per group). Means that are significantly different from 0 (p<0.05), indicating significant improvement, are denoted by an asterisk.
Fig 6.
Change in ABC score with treatment.
Mean ± SD of change in scores on the Aberrant Behavior Checklist (ABC) based on parental report. Data is presented by subscale as well as total scores (n = 8 per group). Mean differences that are statistically different from 0 (p<0.05), indicating significant improvement, are denoted by an asterisk.
Fig 7.
Changes in fecal microbial composition with treatment.
UniFrac principal coordinate analysis (PCoA) plots of 16S sequencing data for (A) unweighted data by treatment group, (B) unweighted data by participant, (C) weighted data by treatment group and (D) weighted data by participant; numbers (e.g. 204, 212) represent participant IDs.
Fig 8.
Microbial community state analysis.
The bottom half of the figure shows four heatmaps corresponding to each of the four enterotypes identified in the study with the abundances of each bacterial taxon displayed by color (bright blue = more abundant; black = no/low abundance). The top half of the figure shows a chart indicating the enterotype of each participant during the various phases of the study. Arrows show the direction of transitions between enterotypes. The numbers (e.g. 204, 212) represent participant IDs to track transitions of individual participant microbiomes between enterotypes. The treatment period is indicated by a letter (a = pre BCP only treatment, b = post BCP only treatment, c = pre combination treatment, d = post combination treatment). The first arm that the participant was enrolled in is denoted by an asterisk.
Fig 9.
Changes in fecal, urinary and serum metabolomes with treatment.
(A) Principal component analysis (PCA) plot of global fecal, urinary and plasma metabolomes changes by participant where numbers (e.g. 204, 212) represent participant IDs and the final metabolome (4th visit) is denoted by an asterisk, (B) mean ± SD fecal ethanol concentrations (n = 15 per group), (C) mean ± SD fecal methanol concentrations (n = 15). Significant differences in mean denoted by an asterisk (p<0.05).
Fig 10.
Changes in cellular cytokine production with treatment.
Median and interquartile range of percentage of stimulated lymphocytes expressing intracellular cytokines before and after treatment (n = 7 per group). (A) Percentage of stimulated CD4+ cells producing intracellular IL-13, and (B) percentage of stimulated CD8+ cells producing TNF-α, before and after treatment. Significant differences in percentage before and after treatment (p<0.05) are denoted by an asterisk.