Spore production and purification

All experiments were carried out with Bacillus subtilis trpC2 strain 168 (DSM 402) originally obtained from the German Collection of Microorganisms and Cell Cultures GmbH (DSMZ, Braunschweig, Germany). Spores were prepared by cultivation in double-strength liquid Schaeffer sporulation medium [38] with vigorous aeration at 37°C for 72 hours. Harvested spores were purified by washing with sterile water repeatedly followed by lysozyme and DNase I treatment for the removal of vegetative cells. Further, a heat inactivation step at 80°C for 10 min was performed to inactivate any remaining vegetative or germinating spores, and the spores were subsequently washed with sterile water and checked for purity by phase-contrast microscopy. Spore preparations were free (>99%) of vegetative cells, germinated spores and cell debris.