Bactericidal Effects Of Hydrogen Peroxide Research Articles

Arginine Metabolism Powers Salmonella Resistance to Oxidative Stress.

Salmonella invades host cells and replicates inside acidified, remodeled vacuoles that are exposed to reactive oxygen species (ROS) generated by the innate immune response. Oxidative products of the phagocyte NADPH oxidase mediate antimicrobial activity, in part, by collapsing the ΔpH of intracellular Salmonella. Given the role of arginine in bacterial resistance to acidic pH, we screened a library of 54 single-gene mutants in Salmonella that are each involved in, but do not entirely block, arginine metabolism. We identified several mutants that affected Salmonella virulence in mice. The triple mutant ΔargCBH, which is deficient in arginine biosynthesis, was attenuated in immunocompetent mice, but recovered virulence in phagocyte NADPH oxidase deficient Cybb-/- mice. Furthermore, ΔargCBH Salmonella was profoundly susceptible to the bacteriostatic and bactericidal effects of hydrogen peroxide. Peroxide stress led to a larger collapse of the ΔpH in ΔargCBH mutants than occurred in wild-type Salmonella. The addition of exogenous arginine rescued ΔargCBH Salmonella from peroxide-induced ΔpH collapse and killing. Combined, these observations suggest that arginine metabolism is a hitherto unknown determinant of virulence that contributes to the antioxidant defenses of Salmonella by preserving pH homeostasis. In the absence of phagocyte NADPH oxidase-produced ROS, host cell-derived l-arginine appears to satisfy the needs of intracellular Salmonella. However, under oxidative stress, Salmonella must additionally rely on de novo biosynthesis to maintain full virulence.

Efficacy of Vaporized Hydrogen Peroxide Combined with Silver Ions against Multidrug-Resistant Gram-Negative and Gram-Positive Clinical Isolates.

Antimicrobial resistance (AMR) is a serious public health problem that results in high morbidity and mortality rates. In particular, multidrug-resistant (MDR) strains circulating in hospital settings pose a major threat as they are associated with serious nosocomial infections. Therefore, regular cleaning and disinfection procedures, usually using chemical disinfectants, must be implemented in these facilities. Hydrogen peroxide (HP)-based disinfectants have proven high microbicidal activity and several comparative advantages over conventional disinfectants. We assessed the in vitro biocidal activity of an 8% HP solution combined with 30 mg/L silver ions (HP + Ag) against MDR clinical isolates of Klebsiella pneumoniae (MDRKp) and Pseudomonas aeruginosa (MDRPa), and methicillin-resistant Staphylococcus aureus (MRSA). Accordingly, the in vitro antibacterial activity was determined using the macrodilution method, and the efficacy was determined for 30 min in terms of (1) activity on bacteria in suspension and (2) activity on surfaces using vaporized HP + Ag on a 20 cm2 stainless steel surface. A strong bactericidal effect of HP + Ag was observed against MDRKp, MDRPa, and MRSA strains, with minimum inhibitory concentrations and minimum bactericidal concentrations between 362.5 and 5800 mg/L. A strong effect was observed during the 30 min of HP + Ag exposure to the resistant clinical isolates, with over 4-Log10 reduction in CFUs. Regarding the efficacy of the disinfectant on surfaces, bacterial load reductions of >99% were observed. These results suggest that HP + Ag is potentially useful as an effective disinfectant for decontaminating surfaces in hospital settings suspected of contamination with MDR bacteria.

Эффективность применения дезинфицирующего средства с использованием генератора горячего тумана

The sporicidal and bactericidal effect of hydrogen peroxide using a hot fog generator on the viability and ultrastructure of spores of the vaccine strain Bacillus anthracis STI-1 was studied using a bacteriological method and electron microscopy. The high efficiency of disinfection treatment of various surfaces has been established, which consists in the fact that within 10-20 minutes there is a complete death of test microorganism that contaminate the surfaces treated with hot fog. It has been experimentally proven that in the process of generating hot fog, the concentration of working disinfectant solution increases, which is based on the partial evaporation of moisture from solution of disinfectant during the formation of hot fog and its transfer by a stream of hot gas. Based on the results obtained, it is advisable to use hot fog generators to achieve a pronounced biocidal effect when carrying out disinfection measures in medical organizations, as well as when performing scientific research. Keywords: microorganisms, spores, bacteria, hydrogen peroxide, hot fog generator.

Antimicrobial Effects of Novel H2O2-Ag+ Complex on Membrane Damage to Staphylococcus aureus, Escherichia coli O157:H7, and Salmonella Typhimurium

Diseases caused by harmful microorganisms pose a serious threat to human health. Safe and environmentally friendly disinfectants are, therefore, essential in preventing and controlling such pathogens. This study aimed to investigate the antimicrobial activity and mechanism of a novel hydrogen peroxide and silver (H2O2-Ag+) complex (HSC) in combatting Staphylococcus aureus ATCC 29213, Escherichia coli O157:H7 NCTC 12900, and Salmonella Typhimurium SL 1344. The MICs and MBCs against S. aureus were found to be 0.014% H2O2-3.125 mg/L Ag+, and for both E. coli O157:H7 and Salmonella Typhimurium they were 0.028% H2O2-6.25 mg/L Ag+. Results of the time-kill trial suggest that HSC could inhibit the growth of the tested bacteria, because 99.9% of viable cells were killed following treatment at 1 MIC for 3 h. The mechanism of antibacterial action of HSC was found to include the disruption of the bacterial cell membrane, followed by reduction of intracellular ATP concentration and inhibition of the activity of antioxidases, superoxide dismutase, and catalase. The enhanced bactericidal effect of hydrogen peroxide combined with silver indicates a potential for its application in environmental disinfection, particularly in the food industry.

Escherichia coli survival in plasma‐treated water and in a gas–liquid plasma reactor

AbstractThe effects of post‐plasma exposure to deionized water treated by argon–water plasma and to direct plasma treatment of bacteria in a liquid solution of a strain of Escherichia coli, which was independently evolved to resist the bactericidal effects of hydrogen peroxide, were determined. The hydrogen peroxide resistant phenotype of the evolved strain was proportional to the initial cell density. The catalase activity was significantly elevated in the evolved strain as compared with the wild‐type strain, and the evolved strain resisted the bactericidal effects of plasma‐treated water (post‐plasma exposure) as compared with the wild‐type strain. The evolved strain remained viable after flowing through the plasma reactor and lysed less when exposed to (nonplasma control) hydrogen peroxide solutions as compared with the wild‐type cells.

In vitro antibacterial phenolic extracts from “sugarbag” pot-honeys of Australian stingless bees (Tetragonula carbonaria)

Australian stingless bee honeys have been shown to exert antioxidant and in vitro antimicrobial properties; however their bioactive factors remained unidentified. This study investigated the antibacterial properties of phenolic extracts from Tetragonula carbonaria honeys. Honeys were harvested from beehives in three sites of South East Australia. Liquid-liquid extractions yielded the phenolic concentrates, for analyses by liquid and gas chromatography mass spectrometry. Antibacterial assays were conducted against Staphylococcus aureus and Klebsiella pneumoniae by in vitro agar diffusion and broth dilution assays. The phenolic extracts averaged to 5.87 mg/100 g of raw honeys, and constituents were 3-phenyllactic acid, lumichrome, diglycosyl flavonoids, norisoprenoids. The honeys did not contain methylglyoxal, dihydroxyacetone or phenolics characteristic of Leptospermum nectars. Hydrogen peroxide content amounted up to 155.8 μM in honeys. Beside the bactericidal effects of hydrogen peroxide at 760 μM, other antibacterial factors were the phenolic extracts of sugarbag honeys that were active at minimum bactericidal concentrations of 1.2-1.8 mg/mL. © 2014 American Chemical Society.

In vitro antibacterial phenolic extracts from "sugarbag" pot-honeys of Australian stingless bees (Tetragonula carbonaria).

Australian stingless bee honeys have been shown to exert antioxidant and in vitro antimicrobial properties; however their bioactive factors remained unidentified. This study investigated the antibacterial properties of phenolic extracts from Tetragonula carbonaria honeys. Honeys were harvested from beehives in three sites of South East Australia. Liquid-liquid extractions yielded the phenolic concentrates, for analyses by liquid and gas chromatography mass spectrometry. Antibacterial assays were conducted against Staphylococcus aureus and Klebsiella pneumoniae by in vitro agar diffusion and broth dilution assays. The phenolic extracts averaged to 5.87 mg/100 g of raw honeys, and constituents were 3-phenyllactic acid, lumichrome, diglycosylflavonoids, norisoprenoids. The honeys did not contain methylglyoxal, dihydroxyacetone or phenolics characteristic of Leptospermum nectars. Hydrogen peroxide content amounted up to 155.8 μM in honeys. Beside the bactericidal effects of hydrogen peroxide at 760 μM, other antibacterial factors were the phenolic extracts of "sugarbag" honeys that were active at minimum bactericidal concentrations of 1.2-1.8 mg/mL.

Bactericidal effect of hydroxyl radicals generated from a low concentration hydrogen peroxide with ultrasound in endodontic treatment.

One approach to enhance the disinfection of root canals in endodontic treatment is ultrasonic irrigation with sodium hypochlorite. Reactive oxygen species, such as hydroxyl radical, are generated by biological defense systems to kill invading bacteria. Ultrasonic irrigation with hydrogen peroxide may be a promising option to increase hydroxyl radical generation. We examined the bactericidal effects of hydroxyl radical generated from low concentration hydrogen peroxide with ultrasound in vitro. An ultrasonic tip was submerged in 0.5 or 1.0 M hydrogen peroxide in a microfuge tube. hydrogen peroxide was irradiated with the ultrasound, the tip of which was maintained centered in the tube to mimic ultrasonic irrigation. Hydroxyl radical generation was assessed by electron spin resonance spectroscopy. Subsequently, Enterococcus faecalis suspension in hydrogen peroxide was prepared and irradiated as described above. Bactericidal effects were assessed by viable counting. Electron spin resonance measurements showed that hydroxyl radical generation increased significantly in a time- and dose-dependent manner (two-way analysis of variance and Tukey’s test, p<0.05). Moreover, the bactericidal effects of hydrogen peroxide against Enterococcus faecalis were enhanced by ultrasonic irradiation in a time- and dose-dependent manner. These results suggest that ultrasonic irrigation in the presence of low concentration hydrogen peroxide can serve as a disinfection strategy in endodontic treatment.

Alternative disinfection techniques to extend the shelf life of minimally processed iceberg lettuce

Chlorination continues to be widely used by the fresh-cut industry for washing and disinfecting minimally processed fruits and vegetables. However, as chlorination has been reported to produce unhealthy by-products such as, chloramines and trihalomethanes, many alternative methods have been reported in the literature. This study examined the use of silver and hydrogen peroxide as possible alternative to chlorination. The results revealed an obvious bactericidal effect of hydrogen peroxide, silver and their combination on spoilage organisms. Combination of electrochemically generated silver (5 ppm) and hydrogen peroxide (0.4 ppm) caused significant ( P < 0.05) reduction in the total plate count (0.87 log), Pseudomonas (2.66 logs), Enterobactericeae (1.61 logs) and yeast and mould (1.60 logs) immediately after washing in comparison to water washed shredded lettuce. However, washing with chlorinated water (5 ppm) under same conditions revealed insignificant reduction in TPC (0.17 log), Pseudomonas (0.60 log), Enterobactericeae (0.15 log) or yeast and mould (0.81 log) counts. Results revealed also that both sources of silver (electrochemical silver and silver nitrate) have similar disinfecting effects, however, electrochemical silver maintained the quality of washed lettuce.

A SitABCD homologue from an avian pathogenic Escherichia coli strain mediates transport of iron and manganese and resistance to hydrogen peroxide

An operon encoding a member of the family of ATP-binding cassette (ABC) divalent metal ion transporters, homologous to Salmonella enterica SitABCD, has been identified in the avian pathogenic Escherichia coli (APEC) strain chi7122. The sitABCD genes were located on the virulence plasmid pAPEC-1, and were highly similar at the nucleotide level to the chromosomally encoded sitABCD genes present in Shigella spp. A cloned copy of sitABCD conferred increased growth upon a siderophore-deficient E. coli strain grown in nutrient broth supplemented with the chelator 2,2'-dipyridyl. Ion rescue demonstrated that Sit-mediated growth promotion of this strain was due to the transport of iron. SitABCD mediated increased transport of both iron and manganese as demonstrated by uptake of 55Fe, 59Fe or 54Mn in E. coli K-12 strains deficient for the transport of iron (aroB feoB) and manganese (mntH) respectively. Isotope uptake and transport inhibition studies showed that in the iron transport deficient strain, SitABCD demonstrated a greater affinity for iron than for manganese, and SitABCD-mediated transport was higher for ferrous iron, whereas in the manganese transport deficient strain, SitABCD demonstrated greater affinity for manganese than for iron. Introduction of the APEC sitABCD genes into an E. coli K-12 mntH mutant also conferred increased resistance to the bactericidal effects of hydrogen peroxide. APEC strain chi7122 derivatives lacking either a functional SitABCD or a functional MntH transport system were as resistant to hydrogen peroxide as the wild-type strain, whereas a Deltasit DeltamntH double mutant was more sensitive to hydrogen peroxide. Overall, the results demonstrate that in E. coli SitABCD represents a manganese and iron transporter that, in combination with other ion transport systems, may contribute to acquisition of iron and manganese, and resistance to oxidative stress.

The haemin storage (Hms+) phenotype of Yersinia pestis is not essential for the pathogenesis of bubonic plague in mammals.

The haemin storage (Hms+) phenotype of Yersinia pestis enables this bacillus to form greenish/brown or red colonies on haemin or Congo Red agar plates, respectively, at 26 but not 37 degrees C. Escherichia coli strains that contain mutations in genes essential for siderophore biosynthesis, porphyrin generation and/or haemin transport remain unable to utilize exogenous haemin as a nutritional iron or porphyrin source when transformed with the cloned Y. pestis hmsHFRS locus. Further physiological analysis of the Hms+ phenotype of Y. pestis strain KIM6+ suggests that the haemin and inorganic iron stored by the Hms system was not used nutritionally under subsequent iron-deficient conditions. In vitro analysis of the bactericidal effects of hydrogen peroxide, superoxide and nitric oxide showed that Hms- Y. pestis cells, in certain cases, were more susceptible than the Hms+ parent cells to these reactive oxygen species at 26 and/or 37 degrees C. In adherence assays, a higher percentage of Hms+ cells were associated with HeLa cells and normal human neutrophils, compared to Hms- cells. However, the Hms+ phenotype did not provide any additional protection against the killing effects of neutrophils. Finally, LD50 analysis in subcutaneously infected mice showed that an Hms- strain was slightly more virulent than Hms+, indicating that the Hms phenotype is not essential for the pathogenesis of bubonic plague in mammals.

Photocatalytic bactericidal effect of TiO 2 thin films: dynamic view of the active oxygen species responsible for the effect

The role of active oxygen species in the photocatalytic bactericidal effect was investigated using a thin transparent titanium dioxide (TiO 2) film. The viable number of Escherichia coli ( E. coli) significantly decreased on the illuminated TiO 2 film, and the bactericidal effect was observed even when E. coli was separated from the TiO 2 surface with a 50 μm porous membrane. Mannitol, a hydroxyl radical scavenger, inhibited the effect only in the absence of the membrane. In contrast, catalase inhibited the effect in all cases. On the basis of these results, the long-range bactericidal effect of hydrogen peroxide was proposed, together with a cooperative effect due to other oxygen species. © 1997 Elsevier Science S.A.

Susceptibility of Mycobacterium leprae to the bactericidal activity of mouse peritoneal macrophages and to hydrogen peroxide.

Macrophages from athymic nude mice were infected in vitro with Mycobacterium leprae to study the intracellular fate of this organism. Using the proportional bactericidal test, we have shown that the viability of M. leprae declines rapidly within these macrophages, although results of clearance experiments demonstrate that live and killed organisms are cleared at comparable rates. We have also shown that M. leprae is susceptible to the bactericidal effects of hydrogen peroxide and we suggest that hydrogen peroxide generated by macrophages is responsible for the killing of intracellular M. leprae.

Resistance of Actinobacillus actinomycetemcomitans and differential susceptibility of oral Haemophilus species to the bactericidal effects of hydrogen peroxide.

We compared the sensitivities of oral and nonoral isolates of Actinobacillus actinomycetemcomitans, Haemophilus segnis, H. aphrophilus, and H. paraphrophilus to the bactericidal action of reagent hydrogen peroxide (H2O2). Susceptibility to a range of H2O2 concentrations (10(-6) to 10(-3) M) was assessed by incubating bacterial suspensions for 1 h at 37 degrees C in the presence of H2O2 and plating on chocolate agar to determine the concentration of H2O2 that would produce a 50% reduction in CFU (50% lethal dose). As a group, A. actinomycetemcomitans was more resistant to H2O2 than the oral haemophili, and H. aphrophilus was much more sensitive than all other organisms tested. The range of 50% lethal dose values for A. actinomycetemcomitans was between 8.5 X 10(-5) and 10(-3) M H2O2 or above. In contrast, H. aphrophilus exhibited 50% lethal dose values from below 1 X 10(-6) to 3.4 X 10(-4) M H2O2. The resistance of A. actinomycetemcomitans to H2O2 may be sufficient to protect these organisms from direct H2O2-mediated killing by host phagocytes.

Bactericidal and cytotoxic effects of hypothiocyanite-hydrogen peroxide mixtures.

Lactoperoxidase catalyzes the oxidation of thiocyanate by hydrogen peroxide into hypothiocyanite, a reaction which can protect bacterial and mammalian cells from killing by hydrogen peroxide. The present study demonstrates, however, that lactoperoxidase in the presence of thiocyanate can actually potentiate the bactericidal and cytotoxic effects of hydrogen peroxide under specific conditions, such as when hydrogen peroxide is present in the reaction mixtures in excess of thiocyanate. The toxic agent was also formed in the absence of lactoperoxidase in a reaction between hypothiocyanite and hydrogen peroxide. Sulfate, sulfite, cyanate, carbonate, and ammonia, which have been postulated to be formed in the chemical oxidation of hypothiocyanite by hydrogen peroxide, were not bactericidal and did not potentiate the bactericidal effect of hydrogen peroxide. Cyanosulfurous acid, the only other postulated product of the chemical oxidation of hypothiocyanite by hydrogen peroxide, may be the killing agent.

Potentiation by L-cysteine of the bactericidal effect of hydrogen peroxide in Escherichia coli.

Under anaerobic conditions an exponentially growing culture of Escherichia coli K-12 was exposed to hydrogen peroxide in the presence of various compounds. Hydrogen peroxide (0.1 mM) together with 0.1 mM L-cysteine or L-cystine killed the organisms more rapidly than 10 mM hydrogen peroxide alone. The exposure of E. coli to hydrogen peroxide in the presence of L-cysteine inhibited some of the catalase. This inhibition, however, could not fully explain the 100-fold increase in hydrogen peroxide sensitivity of the organism in the presence of L-cysteine. Of other compounds tested only some thiols potentiated the bactericidal effect of hydrogen peroxide. These thiols were effective, however, only at concentrations significantly higher than 0.1 mM. The effect of L-cysteine and L-cystine could be annihilated by the metal ion chelating agent 2,2'-bipyridyl. DNA breakage in E. coli K-12 was demonstrated under conditions where the organisms were killed by hydrogen peroxide.

Lactoperoxidase and thiocyanate protect bacteria from hydrogen peroxide.

Lactoperoxidase and thiocyanate were shown to protect Escherichia coli and three oral streptococcal species from the bactericidal effect of hydrogen peroxide under aerobic conditions. Lactoperoxidase in the absence of thiocyanate was also protective for two of the bacterial species in a dilution medium but potentiated hydrogen peroxide toxicity for the other two under the same conditions. The products of the reaction between hydrogen peroxide and thiocyanate in the presence of lactoperoxidase were not bactericidal except in the case of E. coli, and then only under special conditions. The results demonstrate the effectiveness of lactoperoxidase and thiocyanate in protecting living cells from hydrogen peroxide toxicity. Although the effect on human cells was not examined in this study, extrapolation of these results to the cells of the oral mucosa would suggest an important protective role of lactoperoxidase and thiocyanate against the toxic effects of hydrogen peroxide in the oral cavity.

Metabolic inhibition of Peptostreptococcus anaerobius decreases the bactericidal effect of hydrogen peroxide.

Peptostreptococcus anaerobius VPI 4330-1 was tested under various conditions for examination of the bactericidal effect of hydrogen peroxide. The cells were most rapidly killed by hydrogen peroxide when they were in the exponential-growth phase. Cooling or starving the cells decreased the bactericidal effect of hydrogen peroxide. The ionophore nigericin and the metal ion chelating agent 2,2'-bipyridine stopped macromolecular syntheses and greatly decreased the bactericidal effect of hydrogen peroxide. The ionophore valinomycin and the glycolytic inhibitor iodoacetate also stopped the syntheses of the macromolecules but only slightly decreased the bactericidal effect of hydrogen peroxide. Novobiocin, an inhibitor of deoxyribonucleic acid gyrase, and chloramphenicol, an inhibitor of protein synthesis, were not able to decrease the bactericidal effect of hydrogen peroxide. These findings implicate metal ions and an active metabolism of the organism in the bactericidal effect of hydrogen peroxide.

Bactericidal effect of hydrogen peroxide is prevented by the lactoperoxidase-thiocyanate system under anaerobic conditions.

Streptococcus sanguis and Peptostreptococcus anaerobius were exposed to various combinations of the components of the lactoperoxidase-thiocyanate-hydrogen peroxide system. The bactericidal effect of hydrogen peroxide was prevented under anaerobic conditions by lactoperoxidase together with thiocyanate, but not by lactoperoxidase or thiocyanate alone. Thiocyanate was effective already at a molar ratio to hydrogen peroxide of 1:100.

Bactericidal effect of hydrogen peroxide on spacecraft isolates.

Solutions of 3, 10, and 15% hydrogen peroxide were found to have pronounced bactericidal effects, as a function of time of exposure, on sporeformers and non-sporeformers isolated from spacecraft.