Aims: To determine the mechanisms of Bacillus subtilis spore killing by and resistance to the general biological decontamination agents, Decon and Oxone(TM). Methods and Results: Spores of B. subtilis treated with Decon or Oxone(TM) did not accumulate DNA damage and were not mutagenized. Spore killing by these agents was increased if spores were decoated. Spores prepared at higher temperatures were more resistant to these agents, consistent with a major role for spore coats in this resistance. Neither Decon nor Oxone(TM) released the spore core's depot of dipicolinic acid (DPA), but Decon- and Oxone(TM)-treated spores more readily released DPA upon a subsequent normally sublethal heat treatment. Decon- and Oxone(TM)-killed spores initiated germination with dodecylamine more rapidly than untreated spores, but could not complete germination triggered by nutrients or Ca2+-DPA and did not degrade their peptidoglycan cortex. However, lysozyme treatment did not recover these spores. Conclusions: Decon and Oxone(TM) do not kill B. subtilis spores by DNA damage, and a major factor in spore resistance to these agents is the spore coat. Spore killing by both agents renders spores defective in germination, possibly because of damage to the inner membrane of spore. Significance and impact of study: These results provide information on the mechanisms of the killing of bacterial spores by Decon and Oxone(TM).