Identifying the mediators of intracellular E. coli inactivation under UVA light: The (photo) Fenton process and singlet oxygen

Solar disinfection (SODIS) was probed for its underlying mechanism. When Escherichia coli was exposed to UVA irradiation, the dominant solar fraction acting in SODIS process, cells exhibited a shoulder before death ensued. This profile resembles cell killing by hydrogen peroxide (H2O2). Indeed, the use of specialized strains revealed that UVA exposure triggers intracellular H2O2 formation. The resultant H2O2 stress was especially impactful because UVA also inactivated the processes that degrade H2O2-peroxidases through the suppression of metabolism, and catalases through direct enzyme damage. Cell killing was enhanced when water was replaced with D2O, suggesting that singlet oxygen plays a role, possibly as a precursor to H2O2 and/or as the mediator of catalase damage. UVA was especially toxic to mutants lacking miniferritin (dps) or recombinational DNA repair (recA) enzymes, indicating that reactions between ferrous iron and UVA-generated H2O2 lead to lethal DNA damage. Importantly, experiments showed that the intracellular accumulation of H2O2 alone is insufficient to kill cells; therefore, UVA must do something more to enable death. A possibility is that UVA stimulates the reduction of intracellular ferric iron to its ferrous form, either by stimulating O-2(center dot-) formation or by generating photoexcited electron donors. These observations and methods open the door to follow-up experiments that can probe the mechanisms of H2O2 formation, catalase inactivation, and iron reduction. Of immediate utility, the data highlight the intracellular pathways formed under UVA light during SODIS, and that the presence of micromolar iron accelerates the rate at which radiation disinfects water.