Mechanisms of killing of spores of Bacillus subtilis by iodine, glutaraldehyde and nitrous acid

Treatment of wild-type spores of Bacillus subtilis with glutaraldehyde or a n iodine-based disinfectant (Betadine) did not cause detectable mutagenesis , and spores (termed alpha(-)beta(-)) lacking the major DNA-protective alph a/beta-type, small, acid-soluble proteins (SASP) exhibited similar sensitiv ity to these agents. A recA mutation did not sensitize wild-type or alpha(- )beta(-) spores to Betadine or glutaraldehyde, nor did spore treatment with these agents result in significant expression of a recA-lacZ fusion when t he treated spores germinated. Spore glutaraldehyde sensitivity was increase d dramatically by removal of much spore coat protein, but this treatment ha d no effect on Betadine sensitivity. In contrast, nitrous acid treatment of wild-type and alpha(-)beta(-) spores caused significant mutagenesis, with alpha(-)beta(-) spores being much more sensitive to this agent. A recA muta tion further sensitized both wild-type and alpha(-)beta(-) spores to nitrou s acid, and there was significant expression of a recA-lacZ fusion when nit rous acid-treated spores germinated. These results indicate that: (a) nitro us acid kills B. subtilis spores at least in part by DNA damage, and alpha/ beta-type SASP protect against this DNA damage; (b) killing of spores by gl utaraldehyde or Betadine is not due to DNA damage; and (c) the spore coat p rotects spores against killing by glutaraldehyde but not Betadine. Further analysis also demonstrated that spores treated with nitrous acid still germ inated normally, while those treated with glutaraldehyde or Betadine did no t.