Chloramine Mutagenesis in
            <i>Bacillus subtilis</i>

Shih, Katherine Lu; Lederberg, Joshua

doi:10.1126/science.818709

Cited by 68 publications

(23 citation statements)

References 12 publications

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“…Alternative proposals for killing mechanisms have invoked various consequences of sulfhydryl oxidation (13,26,44), cell envelope alteration (4,33,39), or nucleotide dysfunction (45,46 (47)(48)(49) unless incubated with the microbes for several hours (13). Because chloramines derived from endogenous amines show this same behavior (8,13), their role in physiological disinfection processes is probably minimal.…”

Section: Discussionmentioning

confidence: 99%

Biological reactivity of hypochlorous acid: implications for microbicidal mechanisms of leukocyte myeloperoxidase.

Albrich¹,

McCarthy²,

Hurst³

1981

Proc. Natl. Acad. Sci. U.S.A.

408

241

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Oxidative degradation of biological substrates by hypochlorous acid has been examined under reaction conditions similar to those found in active phagosomes. Iron sulfur proteins are bleached extremely rapidly, followed in decreasing order by f3carotene, nucleotides, porphyrins, and heme proteins. Enzymes containing essential cysteine molecules are inactivated with an effectiveness that roughly parallels the nucleophilic reactivities of their sulfhydryl groups. Other compounds, including glucosamines, qumiones, riboflavin, and, except for N-chlorination, phospholipids, are unreactive. Rapid irreversible oxidation of cytochromes, adenine nucleotides, and carotene pigments occurs when bacterial cells are exposed to exogenous hypochlorous acid; with Escherichia coli, titrimetric oxidation of cytochrome was found to coincide with loss of aerobic respiration. The occurrence of these cellular reactions implicates hypochlorous acid as a primary microbicide in myelo.roxidase-containing leukocytes; the reactivity patterns observed are consistent with the view that bactericidal action results primarily from loss of energy-linked respiration due to destruction of cellular electron transport chains and the adenine nucleotide pool.Bacteria commonly lose their ability to divide within minutes of encountering phagocytosing leukocytes (1). Loss ofcell viability often occurs well before the onset ofcellular digestion as determined by physiological changes (2), macromolecular degradation (3, 4), or loss of macromolecular biosynthesis (5). The specific reactions giving rise to cellular death have not yet been identified, at least in part because leukocytes are capable ofinitiating a diverse set of processes which are potentially lethal (6, 7). Given the above observations, however, the microbicidal reactions must be among the first that attend interaction with the leukocyte.Reactions catalyzed by myeloperoxidase (MPOase) appear to make major contributions to the microbicidal-action of polymorphonuclear leukocytes (PMNs) (6, 7). The cell-free MPOase-H202-Cl-system is 'potentially microbicidal; chlorination of bacteria by the cell-free system (8, 9) and of macromolecular fractions during PMN digestion of bacteria (9) electrophile-nucleophile interactions involving association of electropositive chlorine with electron-rich centers on the substrate; reaction pathways are-correspondingly highly dependent upon medium-conditions, particularly H' and Cl-concentrations (14). With the exception ofamines and amino acids (11,12,15), the reactions of biological compounds with HOC1 are not understood (16).We report 'here the results of a survey of simple biological compounds which can be taken as prototypic ofvarious components of bacterial cells. :The data demonstrate that HOCI is strongly selective -toward nucleotides and compounds that are models for certain components ofrespiratory redox chains. This selectivity is shownito extend to bacterial cells. Table 1 were -determined from loss of their characteristic absorption bands when ...

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Section: Discussionmentioning

confidence: 99%

Biological reactivity of hypochlorous acid: implications for microbicidal mechanisms of leukocyte myeloperoxidase.

Albrich¹,

McCarthy²,

Hurst³

1981

Proc. Natl. Acad. Sci. U.S.A.

408

241

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“…Mutagenesis upon HOCl exposure has been investigated in studies using various tester strains of Salmonella typhimurium (51,55), with contradictory results. Chloramines were reported to be mutagenic in S. typhimurium and in Bacillus subtilis (45,51). Epidemiological studies concluded that there is a relationship between an increased risk of bladder cancer (40) and colorectal cancer (18) and drinking chlorinated water.…”

Section: Discussionmentioning

confidence: 99%

“…There is some evidence that HOCl can attack DNA (55). A few reports have shown that chloramine, which can be generated in vivo by HOCl (10,49), causes DNA damage (10,44,45,51), and genotoxicity of free chlorine and chloramines in drinking water was detected with amphibians (27). Recently, Candeias et al (12) have shown that in vitro, hypochlorous acid can also directly generate hydroxyl radicals via a Fenton-type reaction (24) (9).…”

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confidence: 99%

Hypochlorous acid stress in Escherichia coli: resistance, DNA damage, and comparison with hydrogen peroxide stress

1996

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We have investigated the mechanisms of killing of Escherichia coli by HOCl by identifying protective functions. HOCl challenges were performed on cultures arrested in stationary phase and in exponential phase. Resistance to HOCl in both cases was largely mediated by genes involved in resistance to hydrogen peroxide (H 2 O 2 ). In stationary phase, a mutation in rpoS, which controls the expression of starvation genes including those which protect against oxidative stress, renders the cells hypersensitive to killing by HOCl. RpoSregulated genes responsible for this sensitivity were dps, which encodes a DNA-binding protein, and, to a lesser extent, katE and katG, encoding catalases; all three are involved in resistance to H 2 O 2 . In exponential phase, induction of the oxyR regulon, an adaptive response to H 2 O 2 , protected against HOCl exposure, and the oxyR2 constitutive mutant is more resistant than the wild-type strain. The genes involved in this oxyR-dependent resistance have not yet been identified, but they differ from those primarily involved in resistance to H 2 O 2 , including katG, ahp, and dps. Pretreatment with HOCl conferred resistance to H 2 O 2 in an OxyR-independent manner, suggesting a specific adaptive response to HOCl. fur mutants, which have an intracellular iron overload, were more sensitive to HOCl, supporting the generation of hydroxyl radicals upon HOCl exposure via a Fenton-type reaction. Mutations in recombinational repair genes (recA or recB) increased sensitivity to HOCl, indicative of DNA strand breaks. Sensitivity was visible in the wild type only at concentrations above 0.6 mg/liter, but it was observed at much lower concentrations in dps recA mutants.

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“…In particular, HOCl reacts with NH 4 ϩ and organic amines to form highly toxic chloramines, which also are strong oxidizing and chlorinating compounds and could be the actual killing agents. These chloramines are very diffusible species that can enter cells through membranes and react with intracellular components, including DNA (10,32,33,35).In studies of plant-bacterium interactions, it is sometime necessary to use monoxenic models in which a surface-sterilized plant is associated with a bacterial strain. Oxidative agents like household bleach, which is composed of sodium hypochlorite, are the agents most commonly used for sterilization.…”

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confidence: 99%

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Effects of Rice Seed Surface Sterilization with Hypochlorite on Inoculated Burkholderia vietnamiensis

Miché

Balandreau

2001

Appl Environ Microbiol

116

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When a combination of hydrogen peroxide and hypochlorite was used to surface sterilize rice seeds, a 10 2 -to 10 4 -fold decrease in CFU was observed during the first 15 h after inoculation of the rice rhizosphere organism Burkholderia vietnamiensis TVV75. This artifact could not be eliminated simply by rinsing the seeds, even thoroughly, with sterile distilled water. When growth resumed, a significant increase in the frequency of rifampin-and nalidixic acid-resistant mutants in the population was observed compared to the control without seeds. This phenomenon was a specific effect of hypochlorite; it was not observed with hydrogen peroxide alone. It was also not observed when the effect of hypochlorite was counteracted by sodium thiosulfate. We hypothesized that the hypochlorite used for disinfection reacted with the rice seed surface, forming a chlorine cover which was not removed by rinsing and generated mutagenic chloramines. We studied a set of rifampin-and nalidixic acid-resistant mutants obtained after seed surface sterilization. The corresponding rpoB and gyrA genes were amplified and sequenced to characterize the induced mutations. The mutations in five of seven nalidixic acid-resistant mutants and all of the rifampin-resistant mutants studied were found to correspond to single amino acid substitutions. Hypochlorite surface sterilization can thus be a source of artifacts when the initial bacterial colonization of a plant is studied.The use of hypochlorite salts for disinfection dates back to the mid-18th century. Since that time, chlorination has been the most widely used bactericidal treatment for conventional disinfection of municipal drinking water for prevention of epidemic diseases such as cholera and typhoid, and it is still the most widely used method for disinfecting water (18). Hypochlorite is also routinely used as a sanitizer for domestic uses, as well as in food-processing plants to remove surface contaminants which can alter food quality or lead to food-borne diseases (2,3,23,29).Hypochlorite is known to be a very effective to killer of bacteria; even micromolar concentrations are enough to reduce bacterial populations significantly (27). However, little is known about the exact mechanisms of bacterial killing by this sanitizer. When diluted in water, the hypochlorite salts used [NaOCl, Ca(OCl) 2 , LiOCl, and KOCl] lead to formation of HOCl, whose concentration is correlated with bactericidal activity (27). Bacterial killing by HOCl may be due at least in part to lethal DNA damage (13, 42). However, HOCl itself is so reactive that it is unlikely to penetrate cells and reach the DNA; rather, it seems that the bactericidal activity is due to formation of secondary products, as hypochlorous acid reacts avidly with a wide variety of subcellular compounds (membranes, proteins, etc.) (10, 18). In particular, HOCl reacts with NH 4 ϩ and organic amines to form highly toxic chloramines, which also are strong oxidizing and chlorinating compounds and could be the actual killing agents. These chloramin...

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Chloramine Mutagenesis in Bacillus subtilis

Cited by 68 publications

References 12 publications

Biological reactivity of hypochlorous acid: implications for microbicidal mechanisms of leukocyte myeloperoxidase.

Biological reactivity of hypochlorous acid: implications for microbicidal mechanisms of leukocyte myeloperoxidase.

Hypochlorous acid stress in Escherichia coli: resistance, DNA damage, and comparison with hydrogen peroxide stress

Effects of Rice Seed Surface Sterilization with Hypochlorite on Inoculated Burkholderia vietnamiensis

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