Fecal samples were donated by two healthy individuals (male, age 33 and 32), who did not receive antibiotic or probiotic supplementation for at least 3 months before donation. Fecal samples were collected in a sterile 50 mL Falcon tube in an airtight container together with one Anaerogen sachet (Oxoid) to obtain anaerobic conditions until transfer into an anaerobic chamber (10% CO2, 5% H2 and 85% N2) within 3 h (Coy Laboratories, Ann Arbor, MI, USA). Fecal bacteria were immobilized in 1–2 mm gel beads consisting of gellan gum (2.5%, w/v), xanthan (0.25%, w/v) and sodium citrate (0.2%, w/v) under anaerobic conditions as previously described in detail30 (link),32 (link). Sixty mL of freshly produced fecal beads were transferred in the IR bioreactor containing 140 mL of nutritive medium. For bead colonization, three consecutive fed-batch fermentations were carried out by replacing 100 mL fresh nutritive medium every 8–12 h. Bacteria, growing close to the bead surface are continuously released into the growth medium due to active cell growth in the high-biomass-density peripheral layer29 (link),60 (link).
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