Pure Singlet Oxygen Cytotoxicity for Bacteria

We have modified the separated‐surface‐sensitizer singlet oxygen generating system previously described (Midden and Wang, 1983) for the efficient exposure to pure singlet oxygen of bacteria collected on membrane filters. Physical separation of the photosensitizer and the bacteria eliminates the possibility of direct interaction between bacteria and photoexcited sensitizers that could lead to Type I (non‐singlet oxygen) photooxidation processes. This system was used to examine the bacterial cytotoxicity of singlet oxygen. The role of singlet oxygen was confirmed by measuring the decrease in cytotoxicity as the distance between the singlet oxygen source and the bacteria was increased. The gas phase half‐life of the intermediate responsible for cell killing, determined from this distance dependence analysis (24 ± 6 ms), is the same as that calculated from literature data for the gas phase half‐life of singlet oxygen (53 ± 37 ms). Killing of various strains of Salmonella lyphimurium and Escherichia coli was compared at the same dose of singlet oxygen. Bacteria were killed by singlet oxygen at levels several orders of magnitude lower than those effective in killing by H 2 O 2 . Altered DNA repair capacities (uvrB, recA, xth, nth , pKM101) did not affect survival. Incomplete cell wall lipopolysaccharide formation decreased survival following singlet oxygen exposure. Overproduction of the singlet oxygen quencher histidine increased survival, as did accumulation of the dipeptide carnosine (β‐alanyl‐L‐histidine). No evidence for mutagenicity of exogenous singlet oxygen exposure was obtained in a variety of S. typhimurium strains killed to 35% survival.