Gut Microbiota and Metabolism Alteration Following Dietary Isoflavone-Photoperiod Interaction

<b>Figure 1S.</b> Schematic diagram of (A) experimental timeline illustrating fecal boli collections and procedures, and (B) microbiome analysis. OFA, Open Field Apparatus. NIF, No Isoflavones. <b>Figure 2 S.</b> Alpha diversity measurements of mouse stool samples. Observed ASVs (A), Shannon index (B), and Inverse Simpson index were calculated for each mouse stool sample in the study. Each plot is split into the two diet groups, isoflavone free (IF) and regular (R) diets. Within each diet group, the photoperiod groups were individually computed at each timepoint (1-4). The equal light/day (12L:12D) groups are shown in black, the long light short dark (16L:8D) groups are shown in yellow. For each box plot, the center line represents the median and the bottom and top hinges of the box represent the 25th and 75th percentiles. The whiskers of the boxplot extend to the largest value no further than 1.5 times the interquartile range. The points represent each mouse’s alpha diversity value. <b>Figure 3S.</b> Top 10 most predictive ASVs for distinguishing RD from NIF mice. A random forest model was trained on 75% of the mice from timepoints 2, 3 and 4. The model was subsequently tested on the remaining 25%. Each plot shows the relative abundance of each predictive ASV for mice within the isoflavone free (NIF, red) and regular diet (RD, blue) groups at time points 2, 3, and 4. The taxonomic information for each ASV is shown along the bottom as Phylum:Order:Family:Genus. Na = no taxonomic information available. <b>Figure 4S.</b> Relative abundance of the top 16 most abundant genera over time. Each bar represents the mean relative abundance and are colored according to diet and photoperiod groups. The x-axis depicts the sample collection time point (1, 2, 3, or 4). Each plot shows the genus name along the top and are ordered from most (top left) to least abundant (bottom right). Error bars depict the standard error of the mean. <b>Figure 5S.</b> Mean relative abundance of the top 30 most abundant genera in the study. Mice were grouped by diet (RD or NIF) and photoperiod (12L:12D, 16L:8D) and the genus relative abundance was determined over time (sample points 1-4). The more yellow the color, the more abundant the genus. A color key is provided. <b>Figure 6S.</b> Relative abundance of the top 30 most abundant genera in the study by mouse. Mice identification numbers are along the x-axis. Each samples time point (1-4) and photoperiod group (12L:12D, 16L:8D) is shown along the top. The data is split so that the regular diet (RD) mice are listed along the top and the isoflavone free diet (NIF) mice are on the bottom. The more yellow the color, the more abundant the genus. A color key is provided. <b>Figure 7S.</b> Top 10 most predictive ASVs for distinguishing photoperiod groups (12L:12D and 16L:8D). Random forest models were trained on 75% of the mice from timepoints 3 and 4 within the regular diet (A) and the isoflavone free diet groups (B). The model was subsequently tested on the remaining 25%. Each plot shows the relative abundance of each predictive ASV for mice within the 12L:12D (red) and 16L:8D (blue) light groups at time points 3 and 4. The taxonomic information for each ASV is shown along the bottom as Phylum:Order:Family:Genus. Na = no taxonomic iformation available. <b>Figure 8S.</b> Relative abundance of the top phyla by mouse. Each mouse’s fecal microbiota phylum composition is shown over time (sample points 1-4). The mouse identification number is shown along the top. The data is split so that the regular diet group (RD) is on the left and the isoflavone free diet group (NIF) is on the right. <b>Figure 9S.</b> Open Field Apparatus measurements. (A, B, C) first OFA, (D, E, F) second OFA, (G, H, I) third OFA. In each OFA (A, D, G) number of fecal boli, (B, E, H) total distance traveled, and (C, F, I) immobile time is shown. Date are expressed as mean +/- standard error of the mean. Adjusted p values are shown. Regular (RD) and isoflavone free (NIF) diet groups. m= meters, s= seconds<b>Figure 10S.</b> Metabolic transcript levels of (A) Peroxisome proliferator activated receptor gamma (Pparg) and (B) nuclear factor erythroid 2 p45-related factor 2 (Nrf2) were determined via qRT-PCR. Date are expressed as mean +/- standard error of the mean. Adjusted p values are shown. Regular (RD) and isoflavone NIF) diet groups. <b>Supplementary Table 1.</b> Microbiome sequencing results for each mouse fecal sample. Each amplicon sequence variant (ASV) is provided along with the taxonmic information for each ASV (Kingdom, Phylum, Class, Order, Famly, Genus). The mouse subject number is given along with the timepoints, photoperiod group, and diet group. The reads column corresponds to the number of ASV reads for that mouse fecal sample.