After a 7-day adaptation period, mice were randomly divided into 4 groups: NFD (n = 10); NFD plus JSRS (90% NFD plus 10% JSRS, n = 10); HFD (n = 10); HFD plus JSRS (90% HFD plus 10% JSRS, n = 10). Body weight was assessed at weeks 1, 2, 4, and 8. Feces were collected at weeks 2, 4, 8, and stored at −80 °C until further microbiota analysis. After 8 weeks of intervention, all mice were subjected to a 16-h fast and then euthanized and dissected. Blood was collected, and serum was isolated to determine TG as a blood lipid indicator. All fecal samples from week 2 and week 4 were used for high-throughput sequencing of the V3-V4 region of the bacterial 16s rRNA gene [24], and fecal samples from week 8 were processed for deep metagenomic sequencing [25]. The experimental design is presented graphically in Figure 1A.

Experimental design. (A) In stage Ι, mice were randomly divided into four groups: NFD (normal-fat diet, n = 5); NFD plus JSRS (90% normal-fat diet plus 10% jackfruit seed sourced resistant starch, n = 6); HFD (high-fat diet, n = 6); HFD plus JSRS (90% high-fat diet plus 10% jackfruit seed sourced resistant starch, n = 6). The mice were kept for eight weeks. (B) The 4 groups that were treated for 3 weeks in order to make nutritionally obese mice model: NFD (normal-fat diet, n = 10); TR (HFD was chosen for the first 3 weeks for the development of obese mice model, then, 90% NFD plus 10% JSRSplus 8Log CFU B. pseudolongum infusions done later n = 10); HFD (high-fat diet, n = 10); PR (90% HFD plus 10% JSRS plus 8Log B. pseudolongum infusions, n = 10). After 3 weeks, three mice in each group were euthanized to observe abdominal fat accumulation. Then a 3-week intervention was continued on respective dietary treatments until the end of the experiment.

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