Water and Small Molecule Permeation of Dormant Bacillus subtilis Spores


We use a suspended microchannel resonator to characterize the water- and small molecule permeability of Bacillus subtilis spores based on spores' buoyant mass in different solutions. Consistent with previous results, we find that the spore coat is not a significant barrier to small molecules, and the extent to which small molecules may enter the spore is size dependent. We have developed a method to directly observe the exchange kinetics of intra-spore water with deuterium oxide, and we apply this method to wild-type spores and a panel of congenic mutants with deficiencies in the assembly or structure of the coat. Compared to wild-type spores that exchange in approximately 1 sec, several coat-mutant spores are found to have relatively high water permeability with exchange times below the ∼200 msec temporal resolution of our assay. In addition, we find that the water permeability of the spore correlates with the ability of spores to germinate with dodecylamine and with the ability of TbCl3 to inhibit germination with L-valine. These results suggest that the structure of the coat may be necessary for maintaining low water permeability. Importance Spores of Bacillus species cause food spoilage and disease, and are extremely resistant to standard decontamination methods. This hardiness is partly due to spores' extremely low permeability to chemicals, including water. We present a method to directly monitor the uptake of molecules into B. subtilis spores by weighing spores in fluid. The results demonstrate the exchange of core water with sub-second resolution and show a correlation between water permeability and the rate at which small molecules can initiate or inhibit germination in coat-damaged spores. The ability to directly measure the uptake of molecules in the context of spores with known structural or genetic deficiencies is expected to provide insight into the determinants of spores' extreme resistance.

J. Bacteriol.