Sulbactam

Female C57BL/6j mice were bred under specific pathogen-free (SPF) conditions at the Forschungseinrichtungen für Experimentelle Medizin (Charité – University Medicine, Berlin, Germany). The murine gut microbiota was depleted as described previously [27 (link)]. In brief, 8-weeks-old mice were transferred into sterile cages and subjected to a broad-spectrum antibiotic treatment for 8 to 10 weeks by adding ampicillin plus sulbactam (1 g/L; Ratiopharm, Germany), vancomycin (500 mg/L; Cell Pharm, Germany), ciprofloxacin (200 mg/L; Bayer Vital, Germany), imipenem (250 mg/L; MSD, Germany), and metronidazole (1 g/L; Fresenius, Germany) to the drinking water (ad libitum) resulting in secondary abiotic mice. Three days before infecton, the antibiotic cocktail was replaced by autoclaved tap water to assure antibiotic washout in mice.

Female and male IL10^–/– mice and IL10^–/– mice lacking TLR4 (TLR4^–/– × IL10^–/–) (all in C57BL/10ScSn background) were bred and housed within the same specific pathogen-free unit of the Forschungseinrichtungen für Experimentelle Medizin (FEM, Charité – University Medicine Berlin). Immediately after weaning (i.e., at the age of 3 weeks), sex-matched mice of either genotype were subjected to a broad-spectrum antibiotic treatment as described earlier [13 (link), 20 (link)]. In brief, mice were transferred to sterile cages and treated with a quintuple antibiotic cocktail consisting of ampicillin plus sulbactam (1 g/l; Ratiopharm, Ulm, Germany), vancomycin (500 mg/l; Cell Pharm, Hannover, Germany), ciprofloxacin (200 mg/l; Bayer Vital, Leverkusen, Germany), imipenem (250 mg/l; MSD, Haar, Germany), and metronidazole (1 g/l; Fresenius, Bad Homburg, Germany) via the drinking water ad libitum for 4 months. Cultural and culture-independent (i.e., 16S rRNA based molecular) quality control measures revealed virtual absence of bacteria in fecal samples as described previously [13 (link), 21 (link)].