Isolation and characterization of Bacillus subtilis spores that are superdormant for germination with dodecylamine or Ca2+ -dipicolinic acid

Abstract

To isolate and characterize spores superdormant (SD) for germination with either Ca(2+)-dipicolinic acid (CaDPA) or dodecylamine. Bacillus subtilis spores were germinated three times with either CaDPA or dodecylamine and germinated spores removed after each germination treatment, yielding 0.9% (CaDPA-SD spores) or 0.4% (dodecylamine-SD spores) of initial dormant spores. Compared to dormant spores, CaDPA-SD spores germinated poorly with CaDPA and better with dodecylamine and nutrient germinants, although release of DPA from individual CaDPA-SD spores was slow during nutrient germination, and this germination was strongly inhibited by TbCl3. The CaDPA-SD spores were sensitive to hypochlorite and had elevated levels of nutrient germinant receptors (GRs) relative to levels in dormant spores. Dodecylamine-SD spores' germination with dodecylamine and nutrients was similar to that of dormant spores, their germination with Ca-DPA was slower than that of dormant spores, and these SD spores' GR levels were lower than in dormant spores. However, dodecylamine-SD spores were not sensitive to hypochlorite, and the nutrient germination of these SD spores was only partially inhibited by TbCl3 . CaDPA-SD spores appear to have a coat defect and accompanying low levels of the cortex-lytic enzyme CwlJ. The defect in dodecylamine-SD spores, however, is not clear. The results suggest that triggering germination by non-GR-dependent germinants is a potential strategy for efficient spore inactivation.