C.jejuni WT vs pdxA mutant
Ontology highlight
ABSTRACT:
ORGANISM(S): Campylobacter jejuni
SUBMITTER: Hiroshi Asakura
PROVIDER: E-MEXP-3451 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Campylobacter jejuni has become the predominant cause of sheep abortions in the U.S. However, little is know about the genetic diversity among the isolates collected from different time periods. In this study, the genetic diversity of sheep aborion isolates of C. jejuni was investigated by Array-based CGH Each isolate was compared to IA3902, a dye-swap replicate was applied for each isolate
2014-02-11 | E-GEOD-54859 | biostudies-arrayexpress
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Project description:A highly pathogenic Campylobacter jejuni clone has recently emerged as the major cause of Campylobacter-associated sheep abortion in the U.S. and is also associated with foodborne gastroenteritis in humans. A distinct phenotype of this clone is its ability to induce bacteremia and abortion. To facilitate understanding the pathogenic mechanisms of this hyper virulent clone, the differences in global gene expression patterns between this hyper virulent clone (IA3902) and a non-abortifacient strain (NCTC 11168) were compared by DNA microarray. One-condition experiment, IA3902 vs NCTC11168. Biological replicates: 3 IA3902 , 3 NCTC11168. One replicate per array.
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Project description:Campylobacter jejuni is a major cause of food-borne gastroenteritis. Proteomics by label-based two-dimensional liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) identified proteins associated with growth in 0.1% sodium deoxycholate (DOC, a component of gut bile salts), and system-wide validation was performed by data-independent acquisition (DIA-SWATH-MS). Proteins involved in nutrient transport were altered by DOC and aligned with intracellular changes to their respective carbon sources. DOC increased intracellular levels of sulfur-containing amino acids (cysteine and methionine) and the dipeptide cystine (Cys-Cys). A DOC induced transport protein was Cj0025c, which has sequence similarity to bacterial Cys-Cys transporters. Deletion of cj0025c (Δcj0025c) resulted in proteome changes consistent with sulfur starvation.
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Project description:C. jejuni, a spiral-shaped gram-negative bacterium, is a leading bacterial cause of human foodborne illness. Acute disease is associated with C. jejuni invasion of the intestinal epithelium. Further, maximal host cell invasion requires the secretion of proteins termed Campylobacter invasion antigens (Cia). As bile acids are known to alter the pathogenic behavior of other gastrointestinal pathogens, we hypothesized that the virulence potential of Campylobacter may be triggered by the bile acid deoxycholate (DOC). In support of this hypothesis, culturing C. jejuni with a physiologically relevant concentration of DOC significantly altered the kinetics of cell invasion as evidenced by gentamicin-protection assays. In contrast to C. jejuni harvested from Mueller-Hinton (MH) agar plates, C. jejuni harvested from MH agar plates supplemented with DOC demonstrated Cia secretion as judged by metabolic labeling experiments. DOC was also found to induce the expression of the ciaB gene as judged by B-galactosidase reporter assays and real-time RT-PCR. Microarray analysis revealed that DOC induced the expression of virulence genes (i.e., ciaB, cmeABC, dccR, and tlyA). In summary, we demonstrate that it is possible to enhance the pathogenic behavior of C. jejuni by modifying the culture conditions. These results provide a foundation to identify genes expressed by C. jejuni in response to in vivo-like culture conditions. Keywords: Stress response For the expression profiling arrays, an indirect comparison of gene expression was performed, where the expression profile of the C. jejuni F38011 cultured in the presence and absence of DOC was measured separately on different slides as described previously (26). Briefly, Cy5 labeled reference DNA from the C. jejuni F38011 strain was mixed with Cy3 labeled test cDNA (C. jejuni F38011 cultured in the presence or absence of DOC) and hybridized to the Campylobacter cDNA array (26) on separate slides. DNA microarrays were scanned using an Axon GenePix 4000B microarray laser scanner (Axon Instruments, Union City, CA) and the data for spot and background intensities were processed using the GenePix 4.0 software. To compensate for any effect of the amount of template and uneven Cy-dye incorporation, data normalization was performed as previously described (26). For the comparison of genes differentially expressed in the presence and absence of DOC,six hybridization measurements were generated per biological experiment (three technical replicate arrays and two replicate features per array).
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Project description:Objectives: The aim of the study was to characterize (-)-α-Pinene, which is one of the chemical constituents of Alpinia katsumadai seed essential oil responsible for its resistance modulatory activity in Campylobacter jejuni. Methods: Broth microdilution method was used to evaluate the antimicrobial and resistance modulatory potential of (-)-α-Pinene and ethidium bromide accumulation assay to determine its efflux-inhibitory activity in subinhibitory concentration. The target efflux system was identified using knock-out mutants in several efflux related genes. Furthermore, the influence of subinhibitory concentration of (-)-α-Pinene on C. jejuni NCTC 11168 was investigated using microarray technology in order to elucidate the adaptive mechanism of bacteria to treatment with this phytochemical. Knock-out mutants of key adaptation genes were constructed and their role in adaptation to several stress factors, including (-)-α-Pinene, different osmolites and pH, was investigated using Biolog phenotypical microarrays and CFU counts. Results: (-)-α-Pinene was confirmed as highly efficient Campylobacter jejuni resistance modulator, due to its efflux inhibitory activity, which was significantly higher compared to reference inhibitors CCCP and reserpine. The CmeABC along with newly characterized CmeI (Cj1687) was confirmed as its main target efflux system. The transcriptional analysis indicated that the heat shock regulators HspR and HrcA are the main transcription regulators involved in adaptation to (-)-α-Pinene treatment. Conclusions: (-)-α-Pinene is a novel CmeABC and CmeI efflux inhibitor, which evokes the heat shock response in Campylobacter jejuni. Influence of (-)-α-Pinene on gene expression in C. jejuni NCTC 11168 was determined by expressional analysis and qRT-PCR. Exponential phase culture was adjusted to OD600=0.2 in MHB using spectrophotometer (Smart Spec, Bio-Rad, Hercules, CA, USA). Five ml of culture was treated with 62.5 mg/L of (-)-alpha-pinene dissolved in DMSO. Only DMSO (0.048 %) was added to untreated samples. Cultures were treated for 2 h, incubated microaerobically shaking (160 rpm) at 42°C. Experiments were carried out in 4 biological replicates. RNA Protect Bacteria reagent (Qiagen, Maryland, USA) was added to the culture and total RNA was isolated using RNeasy mini kit (Qiagen) and treated with Ambion® Turbo DNA-freeTM kit (Invitrogen, USA). Microarrays with 4751 probes targeting 1756 genes specific for Campylobacter jejuni subsp. jejuni NCTC 11168 (Mycroarray, Biodiscovery-LLC, MI, USA) were used for gene expression analysis. The cDNA was synthesized with random hexamers, SuperScriptTM III Reverse Transcriptase and amynoallyl dUTP (all supplied by Invitrogen, USA) and labeled with monofunctional NHS-ester dye Amersham C3 or Cy5 (GE Healthcare, Buckinghamshire, UK). Concentration of cDNA and labeling efficiency was determined spectrophotometrically with NanoDrop 1000. Four biological replicates were hybridized to four microarrays according to manufacturerâs protocol, incubated for 24 hours at 42â°C and scanned at 532-nm (Cy3) and 635-nm (Cy5) wavelengths using GenePix 4100A (Molecular Devices, Sunnyvale, CA) following the manufacturer's protocol. Fluorescence intensities of each spot were extracted using GenePix Pro 7.0 (Molecular Devices).
2015-11-19 | E-GEOD-59879 | biostudies-arrayexpress
Project description:RNA-seq analysis of the transcriptome of wild type C.jejuni NCTC11168, and of an rpoN mutant of the same strain, both grown in vitro.
2011-06-14 | E-MTAB-706 | biostudies-arrayexpress
Project description:Temperate bacteriophages (prophages) have recently been demonstrated in Campylobacter jejuni. However, what they do there is largely unknown. In the series of studies that are the subject of these submissions we have investigated the relative expression levels of proteins in C. jejuni isolates that differ in the presence or absence of the CJIE1 prophage. At the time of the initial investigations whole genome sequence data were not available for the isolates used, though DNA microarray data indicated that the isolates were very closely related. The overall project was carried out through four separate experiments. Previous work in the scientific literature indicated that growth on medium lacking blood but containing sodium deoxycholate induced the expression of at least some proteins associated with virulence and provided data thought to be of relevance to the virulence of the bacterium. The second set of experiments (experiment 2) therefore compared protein expression in 4-plex iTRAQ experiments using two isolates. Isolate 00-2425 carried the CJIE1 prophage while the second isolate, 00-2426, did not. Three replicate experiments were done. Each isolate was grown on Mueller Hinton agar base and Mueller Hinton agar containing 0.1% sodium deoxycholate.
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