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Query: UMLS:C0023241 (Legionella)
 6,990 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An Escherichia coli K-12 strain deleted for sodA and sodB (manganese and iron superoxide dismutases) was constructed and characterized by Southern blotting, enzyme assays, and physiological analyses. The sod deletion strain was used to clone the iron superoxide dismutase gene of Legionella pneumophila by complementation to paraquat resistance.
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PMID:Construction of an Escherichia coli K-12 strain deleted for manganese and iron superoxide dismutase genes and its use in cloning the iron superoxide dismutase gene of Legionella pneumophila. 158 12

Legionella pneumophila (Lp) grow in cultures in human, guinea pig, and mouse macrophages from A/J strain mice. Because exudate macrophages from this strain of mice have been reported deficient in responsiveness to lymphokines, we thought it of interest to document the extent of responsiveness to interferon-gamma in the context of growth restriction of Lp. Peritoneal exudate macrophages were obtained from A/J mice and cultured in either the presence or absence of recombinant interferon-gamma. These cultures were then infected with Lp and the extent of bacterial growth estimated 48 hr later by means of a colony-forming unit (CFU) assay and electron microscopy. Interferon-gamma treatment significantly restricted the number of CFUs in the culture at concentrations as low as 20 U/ml, but did not affect the uptake of bacteria by macrophages. Furthermore, treatment with interferon induced morphological changes consistent with activated macrophages. The involvement of oxygen-dependent mechanisms in phagocyte killing and growth restriction was examined by the use of inhibitors such as superoxide dismutase (SOD) and catalase. Neither one of these inhibitors of toxic oxygen metabolites affected the interferon-gamma-induced suppression of Lp growth. These results suggest that although thioglycolate-induced exudate macrophages from A/J mice support the growth of Lp, these cells readily respond to the activating influence of interferon-gamma. Furthermore, lymphokine treatment does not inhibit Lp uptake by macrophages and apparently restricts the growth of bacteria by mechanisms independent of the activity of toxic oxygen metabolites.
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PMID:Interferon-gamma induced resistance to Legionella pneumophila in susceptible A/J mouse macrophages. 189 14

Four pairs of virulent/avirulent strains of Legionella pneumophila were examined for their adherence/uptake and activation of human monocytes. Oxidative metabolic responses of monocytes were quantitated by measuring intracellular hydrogen peroxide generation using flow cytometry and by assessment of superoxide dismutase-inhibitable superoxide anion generation. All L. pneumophila strains induced less of a response than did Escherichia coli. Within each pair of isolates, virulent strains of L. pneumophila stimulated the oxidative response of monocytes less than avirulent variants. To determine effects of complement fixation by each strain on their adherence to monocytes, a phagocytic index (PI) was determined under various conditions. In autologous donor serum (AS), all L. pneumophila strains had a PI in the range of 2.1-3.1 bacteria per monocyte, with E. coli having a PI of 9.1. No significant differences were observed between virulent L. pneumophila strains and their avirulent variants. In the presence of heat-inactivated AS, all PI fell to 0.13-0.20 for the L. pneumophila strains, and to 2.16 for E. coli. Using heat-inactivated AS reconstituted with exogenous human complement as a source of opsonization, levels of PI were indistinguishable from their respective levels in AS. This suggests that complement fixation plays an important role in the adherence of virulent and avirulent L. pneumophila to human monocytes.
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PMID:Interactions of virulent and avirulent Legionella pneumophila with human monocytes. 215 38

Keto acids and reduced-oxygen-scavenging enzymes were examined for their roles in supporting the growth of Legionella species and for their potential reactions between the chemical components of the media. When grown in an experimental ACES (2-[(2-amino-2-oxoethyl)-amino] ethanesulfonic acid)-buffered chemically defined (ABCD) broth, the presence of keto acids shortened the lag periods, increased the rates of growth, and gave maximum cell yields. In addition, keto acids affected the specific activities of reduced-oxygen-scavenging enzymes determined during growth. The specific activities of superoxide dismutase of Legionella pneumophila (Knoxville) and L. dumoffii (TEX-KL) were increased three- to eightfold, while that of L. bozemanii (WIGA) was not affected. All strains appeared to be equally sensitive to the effects of superoxide anion (O2-) generated by light-activated riboflavin, and all were equally protected by the presence of keto acids in the ABCD broth. Production of trace amounts of acetate and succinate in pyruvate- and alpha-ketoglutarate-containing media exposed to light suggested that hydrogen peroxide was formed. Pyruvate and alpha-ketoglutarate were products of growth on amino acids, and there was no quantitative evidence that these keto acids were metabolized when they were added to the medium. The rate of cysteine oxidation in ABCD broth was increased by the presence of ferric ion or by exposure to light or by both, and keto acids reduced the rate of this oxidation. ACES buffer was a substrate for the production of O2- in the presence of light, and the combined addition of Fe2+ ions, cysteine, and either keto acid to the medium strongly inhibited the production of O2-. Thus, keto acids inhibited the rate of cysteine oxidation, they stimulated rapid growth by an unknown process, and, in combination with added Fe2+ ions and cysteine, they reversed the toxic effects of light by inhibiting O2- production.
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PMID:Role of keto acids and reduced-oxygen-scavenging enzymes in the growth of Legionella species. 300 29

The difficulties associated with the growth of Legionella species in common laboratory media may be due to the sensitivity of these organisms to low levels of hydrogen peroxide and superoxide radicals. Exposure of yeast extract (YE) broth to fluorescent light generated superoxide radicals (3 microM/h) and hydrogen peroxide (16 microM/h). Autoclaved YE medium was more prone to photochemical oxidation than YE medium sterilized by filtration. Activated charcoals and, to a lesser extent, graphite, but not starch, prevented photochemical oxidation of YE medium, decomposed hydrogen peroxide and superoxide radicals, and prevented light-accelerated autooxidation of cysteine. Also, suspensions of charcoal in phosphate buffer and in charcoal yeast extract medium readily decomposed exogenous peroxide (17 and 23 nmol/ml per min, respectively). Combinations of bovine superoxide dismutase and catalase also decreased the rate of photooxidation of YE medium. Medium protected from light did not accumulate appreciable levels of hydrogen peroxide, and autoclaved YE medium protected from light supported good growth of Legionella micdadei. Various species of Legionella (10(4) cells per ml) exhibited sensitivity to relatively low levels of hydrogen peroxide (26.5 microM) in challenge experiments. The level of hydrogen peroxide that accumulated in YE medium over a period of several hours (greater than 50 microM) was in excess of the level tolerated by Legionella pneumophila, which contained no measurable catalase activity. Strains of L. micdadei, Legionella dumoffi, and Legionella bozmanii contained this enzyme, but the presence of catalase did not appear to confer appreciable tolerance to exogenously generated hydrogen peroxide.
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PMID:Production of superoxide and hydrogen peroxide in medium used to culture Legionella pneumophila: catalytic decomposition by charcoal. 630 19

We studied the interaction between Legionella pneumophila, which is principally a pulmonary pathogen, with primate alveolar macrophages (AM), which are the primary pulmonary cellular defense mechanism. For these studies we used L. pneumophila, type I, which were grown in albumin-yeast extract broth, were greater than 80% viable, and were comparable in virulence for guinea pigs to organisms from guinea pig spleen homogenates. For comparison, avirulent agar-passed L. pneumophila, type I, and a strain of Escherichia coli were also used. In the absence of detectable antibody, AM phagocytosed similar numbers of virulent and avirulent Legionella and killed the majority of ingested Legionella in 15-30 min, as determined by two different assays. The virulent and avirulent Legionella appeared to be equally susceptible to the cidal systems of the AM and both were killed more readily than were E. coli under both assay conditions. Phagocytosis of Legionella by AM was associated with a localized respiratory burst, as indicated by nitroblue tetrazolium reduction around ingested organisms. Killing of AM-associated Legionella was inhibited by the hydroxyl radical (OH.) scavenger mannitol (but not by an equiosmolar concentration of sodium sulfate), and by a combination of superoxide dismutase and catalase (but not by either enzyme alone). These findings suggest a contribution by OH., one generated by the metal-catalyzed interaction of superoxide and hydrogen peroxide (Haber-Weiss reaction) in the anti-Legionella activity of AM. The virulent Legionella that survived intracellularly increased in number from 4 X 10(4) at 1 h to 6 X 10(6) at 96 h after infection. In contrast, avirulent Legionella replicated more slowly, increasing in number from 4 X 10(4) to 1 X 10(5) over the same period. Replication of virulent Legionella destroyed the AM monolayers by 120 h, whereas monolayers containing avirulent organisms remained intact. Thus, virulence of Legionella appears not to correlate with its ability to survive early killing by AM, but rather with the ability of the small fraction of surviving organisms to replicate within these cells.
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PMID:Interaction of primate alveolar macrophages and Legionella pneumophila. 637 25

We examined 40 strains of Legionella for reduced-oxygen scavenging enzymes. Using a simple reaction chamber with a Swinney filter for the Beers and Sizer assay, we determined the catalase activity of live cells grown on buffered charcoal-yeast extract agar. For 29 strains of Legionella pneumophila, the apparent first-order rate constants for catalase ranged from 0.000 to 0.005. Similarly, low values ranging from 0.001 to 0.005 were observed for Legionella wadsworthii, Legionella oakridgensis, and Legionella gormanii. High catalase activities were found for Legionella jordanis, Legionella longbeachae, Legionella micdadei, and Legionella bozemanii, with first-order rate constant values of 0.010 to 0.035. Cell-free extracts were analyzed for catalase, peroxidase, and superoxide dismutase. Cell-free extracts of all strains had superoxide dismutase levels ranging from 8.2 to 30.5 U per mg of protein. The species could be characterized by their catalase and peroxidase since L. pneumophila and L. gormanii had only peroxidase (relative molecular weight [Mr], 150,000); L. dumoffii had a peroxidase (Mr, 150,000) plus a catalase (Mr, 174,000); and all remaining species had catalase only (Mr, 300,000, 220,000, or 150,000).
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PMID:Determination of catalase, peroxidase, and superoxide dismutase within the genus Legionella. 649 Aug 28

Recent advances in the field of molecular biology have revolutionized our understanding of the functioning of living organisms and facilitated the development of robust tools for both diagnosis and treatment of diseases. With particular reference to the field of critical care medicine, development of molecular biology techniques have aided in the following: (1) rapid and highly specific detection of pathogenic infectious agents (eg, Mycobacterium tuberculosis, Pneumocystis carinii, cytomegalovirus, Legionella); (2) development of assays for measurement of circulating cytokines such as tumor necrosis factor (TNF) and interleukin (IL)-1 that has helped our understanding of the pathogenesis of the sepsis syndrome; (3) administration of antibodies or soluble receptors to attempt to prevent untoward effects of cytokines such as TNF or IL-1; and (4) the administration of recombinant deoxyribonucleic acid (DNA) or proteins to patients in an attempt to alter the course of a disease such as antioxidant enzymes (superoxide dismutase). The rapidity of progress in this field has been staggering, which necessitates frequent updating of our knowledge for clinicians to put these molecular tools to their best use. This brief review attempts to explain the basic principles of commonly used techniques in molecular biology including recombinant DNA, polymerase chain reaction, DNA libraries, gene therapy, and protein biochemistry in a manner that is understandable to those without an in-depth knowledge of the field.
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PMID:Current techniques in cell and molecular biology. 749 50

Thioglycolate-elicited murine macrophages from genetically susceptible A/J mice activated with lipopolysaccharide (LPS) and infected with Legionella pneumophila in vitro evince marked inhibition of intracellular growth of this bacterium. The mechanism of inhibition by LPS-activated macrophages in terms of replication of this intracellular pathogen is unclear. LPS activation of murine macrophages induced a downshift in transferrin receptor (TfR) expression and reduction in cellular iron content, and this was correlated with augmented intracellular growth of Legionella in the cells. When LPS-stimulated macrophages were first saturated with iron, partial reversion of L. pneumophila growth restriction was observed. However, an excess of exogenous L-tryptophan (Trp) did not reverse this growth inhibition, nor did supplementation of the macrophage culture medium with both iron and Trp. The antilegionella activity of the macrophages induced by LPS activation was independent of reactive oxygen intermediates (ROI), since the scavengers catalase, superoxide dismutase, mannitol, and thiourea had no effect on growth restriction. Likewise, notwithstanding the ability of LPS-activated macrophages to synthesize reactive nitrogen intermediates (RNI), which was inhibited by L-arginine analogs NG-monomethyl-L-arginine and L-aminoguanidine), as well as by incubation in arginine-free medium, their ability to inhibit the intracellular replication of L. pneumophila was not affected. Thus, we conclude that LPS-activated macrophages inhibit the intracellular growth of L. pneumophila partially by iron-dependent, Trp-independent, and ROI- and RNI-independent mechanisms. We also suggest that additional unknown mechanisms are involved, since complete reversion was not obtained.
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PMID:LPS inhibits the intracellular growth of Legionella pneumophila in thioglycolate elicited murine peritoneal macrophages by iron-dependent, tryptophan-independent, oxygen-independent, and arginine-independent mechanisms. 782 75

A/J mouse macrophages infected with Legionella pneumophila and treated with gamma interferon (IFN-gamma) in vitro developed potent antimicrobial activity. This antilegionella activity was independent of the macrophage capacity to generate reactive oxygen intermediates, since the oxygen radical scavengers catalase, superoxide dismutase, mannitol, and thiourea had no effect on the antilegionella activity of IFN-gamma-activated macrophages. Likewise, whereas the ability of IFN-gamma-activated macrophages to synthesize reactive nitrogen intermediates was markedly inhibited by the L-arginine (Arg) analogs, NG-monomethyl-L-arginine and L-aminoguanidine, as well as by incubation in L-Arg-free medium, their ability to inhibit the intracellular growth of L. pneumophila remained intact. The intracellular growth of L. pneumophila in A/J macrophages was inhibited by the iron(III) chelator desferrioxamine and reversed by Fe-transferrin as well as by ferric salts. Additionally, IFN-gamma-activated macrophages incorporated 28% less 59Fe(III) compared with nonactivated cells. Nonetheless, only partial blocking of growth restriction was observed when IFN-gamma-stimulated macrophages were saturated with iron(III). Indole-propionic acid, which appears to inhibit the biosynthesis of L-tryptophan (L-Trp), was an L-Trp-reversible growth inhibitor of L. pneumophila in macrophages, implying that the intracellular replication of this pathogen is also L-Trp dependent. However, an excess of exogenous L-Trp did not reverse the growth inhibition due to IFN-gamma, though a small synergistic effect was observed when the culture medium was supplemented with both iron(III) and L-Trp. We conclude that IFN-gamma-activated macrophages inhibit the intracellular proliferation of L. pneumophila by reactive oxygen intermediate- and reactive nitrogen intermediate-independent mechanisms and just partially by nutritionally dependent mechanisms. We also suggest that additional mechanisms, still unclear, may be involved, since complete reversion was never obtained and since at higher concentrations of IFN-gamma, iron(III) did not induce any significant reversion in the L. pneumophila growth inhibition.
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PMID:Inhibition of Legionella pneumophila growth by gamma interferon in permissive A/J mouse macrophages: role of reactive oxygen species, nitric oxide, tryptophan, and iron(III). 803 89


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