Differences And Mechanisms Of VBNC Bacteria Induced By Different Radicals

Disinfection is critical for inactivating pathogens and preventing waterborne diseases.As research on water disinfection technology has intensified,the potential risks that may exist during the disinfection process have received widespread attention.Viable but non-culturable cells(VBNC)are a special form of bacteria formed to resist the adverse external environment,which is mainly characterized by reduced cell size and the ability of bacteria to grow without growth on plates but still have metabolic activity and pathogenicity.In recent years,the persulfate system based on sulfate radical(·SO4-),hydrogen peroxide system based on hydroxyl radical(·OH)and ozone system based on ozone(O3)have become hot topics of research.Although·SO4-,·OH and O3 have strong bactericidal ability,it has been reported that the factors induced by different systems can cause microorganisms to enter the VBNC state when disinfection is not complete,thus posing a "hidden risk".Therefore,it is imperative to investigate the differences and mechanisms of different free radicals inducing cells to enter the VBNC state.In this study,we first clarified the reaction order of ozone,hydroxyl radicals and sulfate radicals in different systems,and then evaluated the differences between disinfection systems based on hydroxyl radicals,sulfate radicals and ozone molecules in inducing Pseudomonas aeruginosa into the viable but non-culturable cells using Fe/PS,Fe/H2O2 and O3 as representatives,while further in-depth analysis of the role of sulfate radicals(-S04-)and hydroxyl radicals(·SO4-)and hydroxyl radicals(·OH)in in the induction of cells into VBNC and the mechanism of bacterial resistance to the two free radical stresses.The main findings are summarized as follows:Part Ⅰ:Quantitative analysis of ozone,hydroxyl radicals and sulfate radicals in sterilized systems.Firstly,the residual ozone,hydrogen peroxide and persulfate concentrations in the solution after disinfection by Fe/PS,Fe/H2O2,O3,UV/PS and UV/H2O2 were determined by the sodium indigo disulfonate(IDS)method,N,Ndiethyl-p-p-phenylenediamine(DPD)/horseradish peroxidase(POD)method and iodometric method.Next,the concentrations of hydroxyl radicals and sulfate radicals in the systems were determined by high performance liquid chromatography and UV spectrophotometry.The selectivity of O3,·OH and·SO4-for Pseudomonas aeruginosa under experimental conditions was clarified.It was found that the selectivity of sulfate radicals and ozone was higher than that of hydroxyl radicals in this experimental system.Part Ⅱ:Study of the disinfection effect and mechanism of Fe/PS,Fe/H2O2 and O3 on Pseudomonas aeruginosa.This study evaluated the actual disinfection effects of Fe/H2O2,Fe/PS and O3 from the perspective of inducing P.aeruginosa into the VBNC state and revealed the subcellular mechanisms that produce the differences.The results show that Fe/PS can produce a strong oxidative stress response,which affects the anabolism of substances.In addition,Fe/PS disinfection interfered more significantly with P.aeruginosa metabolism,mainly in the form of reduced biofilm metabolite content,styrene degradation and disturbed aminyl-tRNA biosynthesis.Part Ⅲ:Differential induction of cells into VBNC state by·SO4-and·OH and the induction mechanism.In this study,the role of·SO4-and·OH in UV/PS disinfection was evaluated from the VBNC perspective,using UV/H2O2 as the reference group and UV/PS as the experimental group.The results of the study showed that·SO4-played a more important role in the UV/PS system.Specifically,·SO4-induced less VBNC state than·OH,and the presence of ·SO4-enhanced the cellular oxidative stress response and increased the effect of cell membrane permeability,resulting in a more sustained bacterial inhibitory effect.The results of this study elucidated the toxic effects of Fe/PS,Fe/H2O2 and O3 on Pseudomonas aeruginosa and their differential and possible mechanisms of induction of VBNC cells,and also analyzed the role played by hydroxyl radicals,sulfate radicals in the induction of P.aeruginosa into the VBNC state.It provides a theoretical basis for the precise assessment of the disinfection effect of H2O2,O3 and PS systems on microorganisms.