Objective: To investigate reciprocal regulation between Fur and two RyhB homologs in Methods: Regulatory relationships were assessed by a combination of colony morphology assay, primer extension, electrophoretic mobility shift assay and DNase I footprinting. Results: Fur bound to the promoter-proximal DNA regions of Conclusion Fur and the two RyhB homologs exert negative reciprocal regulation, and RyhB homologs have a positive regulatory effect on biofilm formation in
Bacterial Proteins/metabolism* ; Biofilms ; Gene Expression Regulation, Bacterial/physiology* ; Yersinia pestis/physiology*

Bacterial Proteins ; genetics ; metabolism ; Biofilms ; DNA-Binding Proteins ; genetics ; metabolism ; Flagella ; genetics ; metabolism ; Gene Expression Regulation, Bacterial ; Transcription Factors ; genetics ; metabolism ; Vibrio parahaemolyticus ; cytology ; genetics ; growth & development ; physiology

We used a proteomic approach to identify IbpA in Cronobacter sakazakii (C. sakazaki), which is related to heat tolerance in this strain. The abundance of IbpA in C. sakazakii strains strongly increased after heat shock. C. sakazakii CMCC 45402 ibpA deletion mutants were successfully constructed. The C. sakazakii CMCC 45402 ΔibpA and wild-type strains could not be distinguished based on colony morphology on LB agar plates or biochemical assays. The growth of the C. sakazakii CMCC 45402 ΔibpA mutant in heat shock conditions was indistinguishable from that of the isogenic wild-type, but showed greater heat resistance than E. coli O157:H7 strain CMCC 44828. This study suggests that the absence of a single ibpA gene has no obvious effect on the phenotype or heat resistance of the strain C. sakazakii CMCC 45402.
Bacterial Proteins ; genetics ; metabolism ; Cronobacter sakazakii ; genetics ; physiology ; Gene Expression Regulation, Bacterial ; physiology ; Genotype ; Heat-Shock Proteins ; genetics ; metabolism ; Hot Temperature ; Stress, Physiological

OBJECTIVETo characterize carbapenem (CPM)-non-susceptible Klebsiella pneumoniae (K. pneumoniae) and carbape-nemase produced by these strains isolated from Beijing Children's Hospital based on a five-year surveillance.

METHODSThe Minimal Inhibition Concentration values for 15 antibiotics were assessed using the Phonix100 compact system. PCR amplification and DNA sequencing were used to detect genes encoding carbapenemases. WHONET 5.6 was finally used for resistance analysis.

RESULTSIn total, 179 strains of CPM-non-susceptible K. pneumoniae were isolated from January, 2010 to December, 2014. The rates of non-susceptible to imipenem and meropenem were 95.0% and 95.6%, respectively. In the 179 strains, 95 (53.1%) strains carried the blaIMP gene, and IMP-4 and IMP-8 were detected in 92 (96.8%) and 3 (3.2%) IMP-producing isolates, respectively. 65 (36.3%) strains carried the blaNDM-1 gene. 6 (3.4%) strains carried the blaKPC gene, and KPC-2 were detected in 6 KPC-producing isolates. In addition, New Delhi-Metallo-1 (NDM-1) producing isolates increased from 7.1% to 63.0% in five years and IMP-4 producing isolates decreased from 75.0% to 28.3%.

CONCLUSIONHigh frequencies of multiple resistances to antibiotics were observed in the CPM-non-susceptible K. pneumoniae strains isolated from Beijing Children's Hospital. The production of IMP-4 and NDM-1 metallo-β-lactamases appears to be an important mechanism for CPM-non- susceptible in K. pneumoniae.

Anti-Bacterial Agents ; pharmacology ; Bacterial Proteins ; genetics ; metabolism ; Child ; China ; epidemiology ; Drug Resistance ; Gene Expression Regulation, Bacterial ; physiology ; Gene Expression Regulation, Enzymologic ; physiology ; Hospitals, Pediatric ; Humans ; Klebsiella Infections ; epidemiology ; microbiology ; Klebsiella pneumoniae ; drug effects ; enzymology ; genetics ; Microbial Sensitivity Tests ; Population Surveillance ; Time Factors ; beta-Lactamases ; genetics ; metabolism

Bacterial Proteins ; chemistry ; genetics ; metabolism ; DNA, Bacterial ; chemistry ; genetics ; metabolism ; Gene Expression Regulation, Bacterial ; physiology ; Glutathione ; metabolism ; Listeria monocytogenes ; chemistry ; genetics ; metabolism ; Peptide Termination Factors ; chemistry ; genetics ; metabolism

PURPOSE: In the gastric mucosa of Helicobacter pylori (H. pylori)-infected patients with gastritis or adenocarcinoma, proliferation of gastric epithelial cells is increased. Hyperproliferation is related to induction of oncogenes, such as β-catenin and c-myc. Even though transcription factors NF-κB and AP-1 are activated in H. pylori-infected cells, whether NF-κB or AP-1 regulates the expression of β-catenein or c-myc in H. pylori-infected cells has not been clarified. The present study was undertaken to investigate whether H. pylori-induced activation of NF-κB and AP-1 mediates the expression of oncogenes and hyperproliferation of gastric epithelial cells. MATERIALS AND METHODS: Gastric epithelial AGS cells were transiently transfected with mutant genes for IκBα (MAD3) and c-Jun (TAM67) or treated with a specific NF-κB inhibitor caffeic acid phenethyl ester (CAPE) or a selective AP-1 inhibitor SR-11302 to suppress activation of NF-κB or AP-1, respecively. As reference cells, the control vector pcDNA was transfected to the cells. Wild-type cells or transfected cells were cultured with or without H. pylori. RESULTS: H. pylori induced activation of NF-κB and AP-1, cell proliferation, and expression of oncogenes (β-catenein, c-myc) in AGS cells, which was inhibited by transfection of MAD3 and TAM67. Wild-type cells and the cells transfected with pcDNA showed similar activities of NF-κB and AP-1, proliferation, and oncogene expression regardless of treatment with H. pylori. Both CAPE and SR-11302 inhibited cell proliferation and expression of oncogenes in H. pylori-infected cells. CONCLUSION: H. pylori-induced activation of NF-κB and AP-1 regulates transcription of oncogenes and mediates hyperproliferation in gastric epithelial cells.
Blotting, Western ; Caffeic Acids ; Cell Line, Tumor ; Cell Proliferation ; DNA, Bacterial/analysis/genetics ; DNA-Binding Proteins/*metabolism ; Epithelial Cells/*metabolism ; Gastric Mucosa/*metabolism/pathology ; Gastritis/pathology ; Gene Expression Regulation, Bacterial ; Helicobacter Infections/metabolism/pathology/physiopathology ; Helicobacter pylori/pathogenicity/physiology ; Humans ; NF-kappa B/antagonists & inhibitors/*biosynthesis/metabolism ; Peptide Fragments ; Phenylethyl Alcohol/analogs & derivatives ; Proto-Oncogene Proteins c-jun ; Repressor Proteins ; Transcription Factor AP-1/*biosynthesis ; Transcription Factors/*metabolism ; beta Catenin/*metabolism

Helicobacter pylori (H. pylori) induces the activation of nuclear factor-kB (NF-kappaB) and cytokine expression in gastric epithelial cells. The Janus kinase/signal transducers and activators of transcription (Jak/Stat) cascade is the inflammatory signaling in various cells. The purpose of the present study is to determine whether H. pylori-induced activation of NF-kappaB and the expression of interleukin-8 (IL-8) are mediated by the activation of Jak1/Stat3 in gastric epithelial (AGS) cells. Thus, gastric epithelial AGS cells were infected with H. pylori in Korean isolates (HP99) at bacterium/cell ratio of 300:1, and the level of IL-8 in the medium was determined by enzyme-linked immonosorbent assay. Phospho-specific and total forms of Jak1/Stat3 and IkappaBalpha were assessed by Western blot analysis, and NF-kappaB activation was determined by electrophoretic mobility shift assay. The results showed that H. pylori induced the activation of Jak1/Stat3 and IL-8 production, which was inhibited by a Jak/Stat3 specific inhibitor AG490 in AGS cells in a dose-dependent manner. H. pylori-induced activation of NF-kappaB, determined by phosphorylation of IkappaBalpha and NF-kappaB-DNA binding activity, were inhibited by AG490. In conclusion, Jak1/Stat3 activation may mediate the activation of NF-kappaB and the expression of IL-8 in H. pylori-infected AGS cells. Inhibition of Jak1/Stat3 may be beneficial for the treatment of H. pylori-induced gastric inflammation, since the activation of NF-kappaB is inhibited and inflammatory cytokine expression is suppressed.
Blotting, Western ; DNA, Bacterial/analysis/genetics ; Epithelial Cells/metabolism ; Gastric Mucosa/drug effects/*immunology/microbiology ; Gene Expression Regulation/drug effects/*immunology ; Gene Expression Regulation, Bacterial ; Helicobacter Infections/immunology/*metabolism ; Helicobacter pylori/genetics/pathogenicity/*physiology ; Humans ; Interleukin-8/genetics/*metabolism ; Janus Kinase 1 ; NF-kappa B/biosynthesis/*metabolism ; Phosphorylation ; RNA, Messenger/metabolism ; STAT3 Transcription Factor ; Signal Transduction/genetics

Helicobacter pylori (H. pylori) induces the activation of nuclear factor-kB (NF-kappaB) and cytokine expression in gastric epithelial cells. The Janus kinase/signal transducers and activators of transcription (Jak/Stat) cascade is the inflammatory signaling in various cells. The purpose of the present study is to determine whether H. pylori-induced activation of NF-kappaB and the expression of interleukin-8 (IL-8) are mediated by the activation of Jak1/Stat3 in gastric epithelial (AGS) cells. Thus, gastric epithelial AGS cells were infected with H. pylori in Korean isolates (HP99) at bacterium/cell ratio of 300:1, and the level of IL-8 in the medium was determined by enzyme-linked immonosorbent assay. Phospho-specific and total forms of Jak1/Stat3 and IkappaBalpha were assessed by Western blot analysis, and NF-kappaB activation was determined by electrophoretic mobility shift assay. The results showed that H. pylori induced the activation of Jak1/Stat3 and IL-8 production, which was inhibited by a Jak/Stat3 specific inhibitor AG490 in AGS cells in a dose-dependent manner. H. pylori-induced activation of NF-kappaB, determined by phosphorylation of IkappaBalpha and NF-kappaB-DNA binding activity, were inhibited by AG490. In conclusion, Jak1/Stat3 activation may mediate the activation of NF-kappaB and the expression of IL-8 in H. pylori-infected AGS cells. Inhibition of Jak1/Stat3 may be beneficial for the treatment of H. pylori-induced gastric inflammation, since the activation of NF-kappaB is inhibited and inflammatory cytokine expression is suppressed.
Blotting, Western ; DNA, Bacterial/analysis/genetics ; Epithelial Cells/metabolism ; Gastric Mucosa/drug effects/*immunology/microbiology ; Gene Expression Regulation/drug effects/*immunology ; Gene Expression Regulation, Bacterial ; Helicobacter Infections/immunology/*metabolism ; Helicobacter pylori/genetics/pathogenicity/*physiology ; Humans ; Interleukin-8/genetics/*metabolism ; Janus Kinase 1 ; NF-kappa B/biosynthesis/*metabolism ; Phosphorylation ; RNA, Messenger/metabolism ; STAT3 Transcription Factor ; Signal Transduction/genetics

OBJECTIVETo purify a low-temperature hydroxylamine oxidase (HAO) from a heterotrophic nitrifying bacterium Acinetobacter sp. Y16 and investigate the enzyme property.

METHODSA HAO was purified by an anion-exchange and gel-filtration chromatography from strain Y16. The purity and molecular mass were determined by RP-HPLC and SDS-PAGE. The HAO activity was detected by monitoring the reduction of potassium ferricyanide using hydroxylamine as substrate and ferricyanide as electron acceptor. The partial amino acid sequence was determined by mass spectrometry.

RESULTSThe low-temperature HAO with a molecular mass of 61 kDa was purified from strain Y16 by an anion-exchange and gel-filtration chromatography. The enzyme exhibited an ability to oxidize hydroxylamine in wide temperature range (4-40 °C) in vitro using hydroxylamine as substrate and ferricyanide as electron acceptor. It was stable in the temperature range of 4 to 15 °C and pH range of 6.0 to 8.5 with less than 30% change in its activity. The optimal temperature and pH were 15 °C and 7.5, respectively. Three peptides were determined by mass spectrometry which were shown to be not identical to other reported HAOs.

CONCLUSIONThis is the first study to purify a low-temperature HAO from a heterotrophic nitrifier Acinetobacter sp. It differs from other reported HAOs in molecular mass and enzyme properties. The findings of the present study have suggested that the strain Y16 passes through a hydroxylamine-oxidizing process catalyzed by a low-temperature HAO for ammonium removal.

Acinetobacter ; enzymology ; genetics ; metabolism ; Amino Acid Sequence ; Cold Temperature ; Gene Expression Regulation, Bacterial ; physiology ; Gene Expression Regulation, Enzymologic ; physiology ; Hydrogen-Ion Concentration ; Oxidoreductases ; genetics ; metabolism ; Substrate Specificity

OBJECTIVETo investigate the protein expression profiles of the major food-borne pathogen Campylobacter jejuni NCTC11168.

METHODSMembrane and soluble cellular proteins were extracted from the genome-sequenced C. jejuni strain NCTC11168. Protein expression profiles were determined using two-dimensional gel electrophoresis (2-DE). All the detected spots on the 2-DE map were subjected to matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/TOF) analysis.

RESULTSA total of 537 and 333 spots were detected from the whole cell and membrane-associated proteins of C. jejuni NCTC11168 cultured on Columbia agar medium at 42 °C by 2-DE and Coomassie Brilliant Blue staining, respectively. Analyses of whole cell and membrane-associated proteins included 399 and 133 spots, respectively, which included 182 and 53 functional proteins identified by MALDI-TOF/TOF analysis.

CONCLUSIONThe comprehensive expression protein profiles of C. jeuni NCTC11168 obtained in this study will be useful for elucidating the roles of these proteins in further pathogenesis investigation.

Bacterial Proteins ; genetics ; metabolism ; Campylobacter jejuni ; classification ; genetics ; metabolism ; Electrophoresis, Gel, Two-Dimensional ; methods ; Gene Expression Regulation, Bacterial ; physiology ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; methods ; Transcriptome