Investigating the influence of spore maturation and sporulation conditions on MalS expression during Bacillus subtilis spore germination

Spore forming bacteria of the orders Bacillales and Clostridiales play a major role in food spoilage and food-borne diseases. The spores remain in a dormant state for extended periods due to their highly resistant features. When environmental conditions become favourable, they can germinate as the germinant receptors located on the spore’s inner membrane get activated via germinant binding. This leads to the formation of vegetative cells via germination and subsequent outgrowth. The present study focuses on the synthesis of protein MalS during B. subtilis spore germination by investigating the dynamics of the fluorescence level of a MalS-GFP fusion protein using time-lapse fluorescence microscopy. Our results show an initial increase within the first 15 minutes of germination, followed by a drop and stabilization of the fluorescence throughout the spore ripening period. Western blot analyses, however, indicate no increase in the levels of the MalS-GFP fusion protein during the first 15 minutes after the addition of the germinants. Thus the instantaneous increase in fluorescence of MalS-GFP may likely due to a change in the physical environment as the spore germination is triggered. Our findings also show that the different sporulation conditions and the maturation time of spores affect the expression of MalS-GFP and the germination behaviour of the spores.