Germ-free and Gnotobiotic Mouse Models

C57BL/6 mice were re-derived to germ-free status52 (link). Gnotobiotic C57BL/6 mice colonized with ASF or sDMDMm₂ containing 12 defined bacterial strains (called the Oligo-Mouse-Microbiota, see ‘Bacterial strains') were generated and maintained at the clean mouse facility of the University of Bern. SPF mice (on a C57BL/6 background) were purchased from Harlan Laboratories. Monocolonization was performed by intragastric administration of 10¹⁰ colony-forming units (CFU; if not otherwise specified) of E. coli or B. thetaiotaomicron into 8- to 16-week-old mixed-sex germ-free mice. Bicolonization was established in 8- to 16-week-old mixed-sex germ-free mice by orally gavaging with 10¹⁰ CFU of E. coli and 10¹⁰ CFU of B. thetaiotaomicron. Mice were either used germ-free or stably colonized over 5 days with the organism(s) shown, except for trajectory experiments for bacterial replication where time points between 8 and 24 h were used. Control experiments confirmed that the dose of 10¹⁰ CFU was saturating and established a steady state at the outset, whereas doses ≤10⁸ CFU required an initial phase of replication before the biomass reached steady state at 18 h. Germ-free mice were routinely monitored by culture-dependent (Luria-Bertani Broth (LB) agar and Wilkin's Anaerobic media (Oxoid) supplemented with 5% defibrinated sheep's blood) and culture-independent (Gram stain and cell-impermeant nucleic acid stain Sytox green) methods to confirm sterility. All mouse experiments were performed in accordance with the Swiss Federal and Cantonal regulations and were approved by the Ethics Committee for Animal Experimentation of the SAMS and SCNAT.