Figure 2

Like S. cerevisiae, S. boulardii cells undergo programmed cell death in ethanol . S. Boulardii cells were cultured in rich YPD media overnight and resuspended in fresh media and allowed to reach exponential phase. They were then resuspended either in water or fresh media or in water or fresh media containing either 15% or 22% ethanol, and allowed to grow at 30°C for the indicated times. (A) Mitochondrial fragmentation was detected in cells cultured in 15% ethanol using 10 nM Mitotracker Green. (B) Intracellular ROS accumulation was detected in cells cultured in 22% ethanol with 5 μg/ml of dihydrorhodamine 123. (C) Activated caspase-like enzymatic activity was detected in S. boulardii cells cultured in 22% ethanol using a FLICA apoptosis detection kit according to the manufacturer’s specifications. At least three independent cultures were tested and compared. The differences in staining patterns were deemed statistically significant by the Student’s t-test (p<0.05)