Oxygen toxicity in Treponema pallidum: deoxyribonucleic acid single-stranded breakage induced by low doses of hydrogen peroxide

Abstract

The effect of hydrogen peroxide on Treponema pallidum was investigated. The in vitro loss of virulence (as measured by rabbit inoculation) of T. pallidum was accelerated by as low as 100 μM hydrogen peroxide in the complex maintenance medium used. Higher doses led to rapidly accelerated death with 500 μM hydrogen peroxide causing sterilization of the medium within 3 to 4 h. Since hydrogen peroxide is known to cause single-stranded breaks in DNA, the effect of hydrogen peroxide on the treponemal genome was examined. Extensive breakage was caused by 100 μM hydrogen peroxide as determined on alkaline sucrose gradients. A limit was reached at 250 μM and above. Single-stranded breaks could be demonstrated as early as 5 – 10 min after exposure to hydrogen peroxide when the treponemes were exposed to 250 μM hydrogen peroxide; accelerated death was evident by 2 h past exposure demonstrating that DNA breakage was preceding death. Treponemal death caused by penicillin did not result in DNA breakage. The repair-proficient bacterium Escherichia coli K-12 was compared with T. pallidum. It required 10 – 100 times more hydrogen peroxide to cause various levels of breakage. Escherichia coli K-12 rapidly repaired DNA breakage once hydrogen peroxide was removed by addition of catalase. Treponema pallidum, in comparison, showed little or no repair in vitro. Addition of catalase or dithiothreitol to the medium protected against all but a low level of breakage; this may reflect on the ability of catalase and reducing agents to protect T. pallidum against oxygen toxicity in vitro.