In situ changes in native fluorescence from Bacillus subtilis during endospore formation

Native fluorescence emission and excitation spectra were used to monitor changes in Bacillus subtilis (Bs) and Staphylococcus aureus (Sa) subjected to starvation conditions. Initially, the fluorescence spectra from the Bs and Sa was dominated by tryptophan emission. After the second day, a fluorescence band with an emission peak at 410 nm and an excitation peak at 345 nm appeared in the Bs. This emission is from dipicolinic acid, a major constituent of bacterial endospores. The dipicolinic acid intensity increased steadily during the next 2 to 4 days as the number of Bs forming spores increased. No dipicolinic acid signal was observed in the (non-spore forming) Sa. The addition of β-hydroxybutyric acid to either the Bs or Sa resulted in the emergence of a third band with very strong fluorescence emission maximum at 460 nm and with excitation maxima at 250, 270 and 400 nm. This 460 nm emission was quenched with the addition of Fe²⁺, indicating that the source of this emission is a siderophore produced by the bacteria.