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Microarray Analysis and Functional Characterization of the Nitrosative Stress Response in Nonmucoid and Mucoid Pseudomonas aeruginosa. Aaron M. Firoved, 2004.The type strain of Pseudomonas aeruginosa, PAO1, showed great upregulation of many nitrosative defense genes upon treatment with S-nitrosoglutathione, while the mucoid strain PAO578II showed no further upregulation above its constitutive upregulation of nor and fhp . NO· consumption however, showed that both strains mount functional, protein synthesis-dependent NO·-consumptive responses . Heterologous Expression of the Enterococcal vanA Operon in Methicillin-Resistant Staphylococcus aureus. Bruno Périchon, 2004.Two methicillin- and vancomycin-resistant Staphylococcus aureus strains, MI-VRSA and PA-VRSA, and Enterococcus faecalis DMC83006B, considered to be the potential donor of glycopeptide resistance to MI-VRSA, were studied . MI-VRSA is highly resistant to both glycopeptides, whereas PA-VRSA displays low-level resistance to vancomycin and reduced susceptibility to teicoplanin . We have analyzed the expression of the vanA operon in the three clinical isolates . Determination of the relative amounts of late peptidoglycan precursors and quantification of the D,D-peptidase activities, in the absence or after induction by glycopeptides, revealed that the resistance genes were expressed at similarly high levels in the three strains . Glycopeptide resistance stability in the three strains was studied by replica plating . Resistance was lost at high frequency, ca . 50%, after overnight growth of PA-VRSA in the absence of antibiotics, whereas it was fully stable in MI-VRSA and E . faecalis DMC83006B . Induction of resistance by vancomycin was significantly delayed in PA-VRSA relative to MI-VRSA . Low-level glycopeptide resistance of S . aureus PA-VRSA is thus likely due to instability of the genetic element, plasmid or transposon, carrying the vanA operon associated with a longer lag phase before growth resumes after induction by vancomycin . Altering the Substrate Specificity of Organophosphorus Hydrolase for Enhanced Hydrolysis of Chlorpyrifos. Catherine Mee-Hie Cho, 2004.Chlorpyrifos is one of the most popular pesticides used for agriculture crop protection, and widespread contamination is a potential concern . However, chlorpyrifos is hydrolyzed almost 1,000-fold slower than the preferred substrate, paraoxon, by organophosphorus hydrolase (OPH), an enzyme that can degrade a broad range of organophosphate pesticides . We have recently demonstrated that directed evolution can be used to generate OPH variants with up to 25-fold improvement in hydrolysis of methyl parathion . The obvious question and challenge are whether similar success could be achieved with this poorly hydrolyzed substrate, chlorpyrifos . For this study, five improved variants were selected from two rounds of directed evolution based on the formation of clear haloes on Luria-Bertani plates overlaid with chlorpyrifos . One variant, B3561, exhibited a 725-fold increase in the kcat/Km value for chlorpyrifos hydrolysis as well as enhanced hydrolysis rates for several other OP compounds tested . Considering that wild-type OPH hydrolyzes paraoxon at a rate close to the diffusion control limit, the 39-fold improvement in hydrolysis of paraoxon by B3561 suggests that this variant is one of the most efficient enzymes available to attack a wide spectrum of organophosphate nerve agents . The ClpXP ATP-Dependent Protease Regulates Flagellum Synthesis in Salmonella enterica Serovar Typhimurium. Toshifumi Tomoyasu, 2002.The ClpXP protease is a member of the ATP-dependent protease family and plays a dynamic role in the control of availability of regulatory proteins and the breakdown of abnormal and misfolded proteins . The proteolytic activity is rendered by the ClpP component, while the substrate specificity is determined by the ClpX component that has ATPase activity . We describe here a new role of the ClpXP protease in Salmonella enterica serovar Typhimurium in which ClpXP is involved in the regulation of flagellum synthesis . Cells deleted for ClpXP show "hyperflagellate phenotype," exhibit overproduction of the flagellar protein, and show a fourfold increase in the rate of transcription of the fliC encoding flagellar filament . The assay for promoter activity of the genes responsible for expression of the fliC showed that the depletion of ClpXP results in dramatic enhancement of the expression of the fliA encoding sigma factor  28, leaving the expression level of the flhD master operon lying at the top of the transcription hierarchy of flagellar regulon almost normal . These results suggest that the ClpXP may be responsible for repressing the expression of flagellar regulon through the control of the FlhD/FlhC master regulators at the posttranscriptional and/or posttranslational levels . Proteome analysis of proteins secreted from the mutant cells deficient for flhDC and clpXP genes demonstrated that the
 flhD mutation abolished the enhanced effect by
clpXP mutation on the production of flagellar proteins, suggesting that the ClpXP possibly defines a regulatory pathway affecting the expression of flagellar regulon that is dependent on FlhD/FlhC master regulators .
Application of Host-Specific Bacteriophages to the Surface of Chicken Skin Leads to a Reduction in Recovery of Campylobacter jejuni. Robert J. Atterbury, 2003.Retail poultry products are widely purported as the major infection vehicle for human campylobacteriosis . Numerous intervention strategies have sought to reduce Campylobacter contamination on broiler carcasses in the abattoir . This study reports the efficacy of bacteriophage in reducing the number of recoverable Campylobacter jejuni cells on artificially contaminated chicken skin .
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