Quantification of Listeria monocytogenes Carriage in Mice

Seven- to 11-week-old male mice (C57BL/6 mEcad E16P KI^{77 (link)}) were infected intravenously in the tail vein as previously described^{77 (link)}. Fraternities were kept together in a cage during the whole experiment, except when separated to exclude that carriage was due to coprophagy. To quantify carriage at 30-days post-inoculation, faeces were collected from each individual mouse and weighted before being homogenised in 2 ml of PBS. CFU count was performed by serial dilution of homogenised faeces on ALOA plates (Biomérieux) as described in ref. ⁸ . Separate faecal pellets were collected pre-inoculation and/or 30-days post-inoculation and stored at −20 °C for DNA extraction for 16S rRNA gene sequencing. DNA was extracted with DNeasy PowerSoil Pro Kit (Qiagen).
For experiments with antibiotic treatment, 5-week-old female mice (C57BL/6 mEcad E16P KI^{77 (link)}) received orally every 12 h during 4 days PBS (as control) or the following antibiotics (similarly to^{57 (link)}): ampicillin (100 mg/kg, Sigma-Aldrich A9618), vancomycin (50 mg/kg, Sigma-Aldrich 75423), metronidazole (100 mg/kg, Abcam ab141218), neomycin (100 mg/kg, Gibco 11811-031), amphotericin B (1 mg/kg, ApexBio Technology B1885). The mice were inoculated, as described above, 4 weeks after the antibiotic treatment.