Spore formation by Bacillus cereus in broth as affected by temperature, nutrient availability, and manganese

A study was done to determine the effect of interacting factors on sporulation of Bacillus cereus in broth. Vegetative cells (1.4 to 2.2 log CFU/ml) of B. cereus strain 038-2 (capable of growing at 12°C) and strain F3812/84 (capable of growing at 8°C) were inoculated into 30 ml of tryptic soy broth (TSB), TSB supplemented with manganese (50 μg/ml), diluted (10%) TSB (dTSB), and dTSB supplemented with manganese (50 μg/ml) and incubated at 8, 12, or 22°C for up to 30, 30, or 10 days, respectively. Unheated and heated (80°C for 10 min) cultures were plated on brain heart infusion agar to determine total cell counts (vegetative cells plus spores) and the number of spores produced, respectively. Both strains of B. cereus survived in TSB and dTSB for 30 days at 8°C but did not sporulate. At 12°C, cells grew in TSB to a population of 6.0 ± 0.8 log CFU/ml, which was maintained for 30 days. Neither strain grew in dTSB at 12°C and survived for at least 30 days. Spores were not produced in any of the test broths at 12°C. At 22°C, cells reached a stationary growth phase between 12 and 24 h in TSB, TSB supplemented with manganese, and dTSB supplemented with manganese, and approximately 1% of the CFU were spores. In dTSB, cell growth and spore formation were retarded at 22°C and a significantly lower number of spores was produced compared with the number of spores produced in TSB, TSB supplemented with manganese, and dTSB supplemented with manganese. The addition of manganese to TSB did not affect cell growth or spore formation, but manganese did enhance sporulation in dTSB. This study provides useful information on spore formation by B. cereus as affected by conditions that may be imposed in liquid milieus on the surface of foods and on food contact surfaces in processing environments.