Vitamin k1

Vitamin K1, brain-heart-infusion broth (BHI), and agar were purchased from Carl Roth (Karlsruhe, Germany). Wilkins-Chalgren anaerobe agar (WC) was purchased from Oxoid (Basingstoke, England). Hemin was purchased from Sigma-Aldrich (St. Luis, United States) and Anaerocult A was purchased from Merck (Darmstadt, Germany). Metronidazole was purchased from Sigma-Aldrich (St. Luis, United States), and Etests were purchased from bioMérieux (Marcy-l’Étoile, France).

Unless otherwise stated, M. schaedleri ASF 457 MCS was cultured under anaerobic conditions under an N₂ and 8% H₂ atmosphere at 37°C without shaking using anaerobic Mucispirillum medium (AMM), which is based on Trypticase soy agar and contains (per liter) 18 g brain heart broth (Merck), 15 g tryptone soy broth (Oxoid), 5 g yeast extract (Bacto yeast extract), 2.5 g K₂HPO₄ (Carl Roth), 1 mg hemin (Sigma), 0.5 mg vitamin K₁ (Carl Roth), 0.4 g Na₂CO₃ (Carl Roth), 3% fetal calf serum (Sigma), 0.5 mg l-cysteine hydrochloride (Sigma), and 0.5 mg alpha-(d+)-glucose monohydrate (Carl Roth) (87 ). M. schaedleri ASF 457 MCS was analyzed for growth in AMM with or without the presence of the following compounds: hydrogen (8%), formate (0.5, 2.5, 10, or 50 mM), nitrate (2 or 10 mM), and fumarate (10 or 50 mM). Growth was quantified by optical density measured at 600 nm (OD₆₀₀) (M107 high-specification visible spectrophotometer; Spectronic Camspec Ltd., Leeds, United Kingdom). OD₆₀₀ values were normalized by subtracting the background absorbance values of abiotic-medium controls.

Transferrin–Alexa-Fluor-647 (#T23366 used at 25 μg/mL), EGF-Alexa-Fluor-555 (E35350, used at 400 ng/mL), vitamin K1 (3804.1, Carl Roth GmbH), puromycin (Toku-E, P001, used at 1 µg/mL), Geneticin® Selective Antibiotic (G418, 11,811,031, used at 1 mg/mL) all from Thermo Fisher Scientific; DAPI (D9542), phalloidin-Atto 390 (50,556) both from Sigma-Aldrich; EGF (AF-100–15, used at 400 ng/mL), PDGF-BB (100-14B, used at 50 ng/mL) both from PeproTech.