Cellobiose

Roseburia intestinalis L1-82 (DSMZ14610) was purchased from Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH (Braunschweig, Germany). Bacteroides fragilis (ATCC 25285) and Bacteroides vulgatus were obtained from Ningbo Mingzhou Biotechnology Co., Ltd. (Zhejiang, China). The above bacteria were cultured under anaerobic conditions at 37°C. An anaerobic culture environment was created using Atmosphere Generation Systems (AnaeroJar^™ ASSEMBLY and AnaeroGen^™, OXOID, Thermo Fisher Scientific). The culture medium (Miquel et al., 2015 (link)) was brain heart infusion (BHI) containing 0.5% yeast extract (OXOID), 1 mg/ml cellobiose (Macklin, China), 1 mg/ml maltose (Solarbio, China), 0.5 mg/ml cellobiose (Macklin, China), and 0.5 mg/ml cysteine (Solarbio, China). We established a standard curve to convert the absorbance values at 600 nm (OD600) to CFU values (Supplementary Figure S1). Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 25923, Salmonella typhimurium CMCC 50115, Bacillus subtilis DSM 1088, and Bacillus cereus ATCC 11778 were used as controls. These strains were cultured in nutrient broth at 37°C in an aerobic environment.

The growth profiling was performed in triplicate in 9 cm Petri dishes on MM with 1.5% Ultra-pure agarose (Invitrogen, Carlsbad, CA, USA) with one of the following carbon sources: 25 mM D-glucose (Cat. no. G6172, Macklin, Shanghai, China), D-fructose (Cat. no. D809612, Macklin), D-xylose (Cat. no. XBO998, Sangon Biotech, Shanghai, China), L-arabinose (Cat. no. L824031, Macklin), cellobiose (Cat. no. C6182, Macklin), 1% w/v microcrystalline cellulose from cotton linters (Cat. no. 435236, Sigma, Shanghai, China), or 1% w/v xylan (from corn cob) (Cat. no. X823251, Macklin), or without an added carbon source. For growth profiling plate cultures, the plates were inoculated with a ~4 mm² agar plug from the growing edge of the T. harzianum colony from a PDA plate, and then incubated for at least four days at 28 °C with 12 h light and 12 h dark. After 48 h, the colony diameters were measured, and after both 48 h and 96 h, photos of the colonies were taken. Table S3 contains the colony diameter measurements from the two repeat experiments. Two independent deletion strains were tested for growth on the carbon sources to confirm the reliability of attributing the observed phenotypes to the deleted gene. We selected one of the deletion strains to use in further studies.