Project description:BACKGROUND:Campylobacter jejuni contains a homologue of the luxS gene shown to be responsible for the production of the signalling molecule autoinducer-2 (AI-2) in Vibrio harveyi and Vibrio cholerae. The aim of this study was to determine whether AI-2 acted as a diffusible quorum sensing signal controlling C. jejuni gene expression when it is produced at high levels during mid exponential growth phase. RESULTS:AI-2 activity was produced by the parental strain NCTC 11168 when grown in rich Mueller-Hinton broth (MHB) as expected, but interestingly was not present in defined Modified Eagles Medium (MEM-alpha). Consistent with previous studies, the luxS mutant showed comparable growth rates to the parental strain and exhibited decreased motility halos in both MEM-alpha and MHB. Microarray analysis of genes differentially expressed in wild type and luxS mutant strains showed that many effects on mRNA transcript abundance were dependent on the growth medium and linked to metabolic functions including methionine metabolism. Addition of exogenously produced AI-2 to the wild type and the luxS mutant, growing exponentially in either MHB or MEM-alpha did not induce any transcriptional changes as analysed by microarray. CONCLUSION:Taken together these results led us to conclude that there is no evidence for the role of AI-2 in cell-to-cell communication in C. jejuni strain NCTC 11168 under the growth conditions used, and that the effects of the luxS mutation on the transcriptome are related to the consequential loss of function in the activated methyl cycle.
Project description:This SuperSeries is composed of the following subset Series: GSE18427: Addition of Exogenous AI-2 to C. jejuni 11168 Wild Type and LuxS01. GSE18451: Wild Type 11168 versus LuxS01 Mutant Cultured in MHB and MEM- ? media Refer to individual Series
Project description:Campylobacter jejuni is a major bacterial cause of human diarrheal diseases worldwide. Despite its sensitivity to environmental stresses, C. jejuni ubiquitously distributes throughout poultry production chains. Biofilm formation mediated by quorum sensing is suggested to be critical to the survival of C. jejuni in agroecosystem. C. jejuni possesses LuxS, the enzyme involved in the production of autoinducer-2 (AI-2) signaling molecules. In this study, two fatty acids, namely decanoic acid and lauric acid, were identified to be effective in inhibiting AI-2 activity of C. jejuni. Both decanoic acid and lauric acid at 100 ppm inhibited ∼90% AI-2 activity (P < 0.05) of C. jejuni without bacterial inactivation. The biofilm biomass of two C. jejuni strains was reduced by 10-50% (P < 0.05) after treatment by both fatty acids, while increased biofilm formation was observed for one C. jejuni strain. In addition, both fatty acids effectively reduced the motility of all tested C. jejuni strains. These findings can aid in developing alternative C. jejuni control strategies in agri-food and clinical settings.
Project description:Campylobacter jejuni is one of the most prevalent causes of bacterial gastroenteritis worldwide, and it is largely associated with consumption of contaminated poultry. Current Campylobacter control measures at the poultry production level remain insufficient, and hence there is the need for alternative control strategies. We evaluated the potential of the monoterpene (-)-α-pinene for control of C. jejuni in poultry. The antibacterial and resistance-modulatory activities of (-)-α-pinene were also determined against 57 C. jejuni strains. In addition, the anti-quorum-sensing activity of (-)-α-pinene against C. jejuni NCTC 11168 was determined for three subinhibitory concentrations (125, 62.5, 31.25 mg/L) over three incubation times using an autoinducer-2 bioassay based on Vibrio harveyi BB170 bioluminescence measurements. The effects of a subinhibitory concentration of (-)-α-pinene (250 mg/L) on survival of C. jejuni, and in combination with enrofloxacin on fluoroquinolone resistance development in C. jejuni, were determined in a broiler chicken model, by addition of (-)-α-pinene to the broiler water supply. The reduction of C. jejuni numbers by (-)-α-pinene was further determined in broiler chickens that were colonized with either fluoroquinolone-susceptible or -resistant strains, by direct gavage treatment. We observed weak in vitro antimicrobial activity for (-)-α-pinene alone (MIC >500 mg/L), but strong potentiating effects on antibiotics erythromycin and ciprofloxacin against different Campylobacter strains (>512 fold change). After 24 h of treatment of C. jejuni with (-)-α-pinene, its quorum-sensing signaling was reduced by >80% compared to the untreated control. When given in the drinking water, (-)-α-pinene did not show any significant inhibitory effects on the level of C. jejuni in the colonized chickens, and did not reduce fluoroquinolone resistance development in combination with enrofloxacin. Conversely, when (-)-α-pinene was administered by direct gavage, it significantly reduced the number of fluoroquinolone susceptible C. jejuni in the colonized broiler chickens. These results demonstrate that (-)-α-pinene modulates quorum-sensing in Campylobacter, potentiates antibiotics against different Campylobacter strains, and reduces Campylobacter colonization in broiler chickens.
Project description:Campylobacter jejuni has been reported as a major cause of bacterial food-borne enteritides in developed countries during the last decade. Despite its fastidious growth requirements, including low level of oxygen and high level of CO2, this pathogen is able to persist in the environment without permanent loss of its viability and virulence. As C. jejuni is not able to multiply outside a host, the cells spend significant amount of time in stationary phase of growth. The entry into the stationary phase is often correlated to resistance to various stresses in bacteria. The switching between exponential and stationary phases is frequently mediated by the regulator sigma S (RpoS). However, this factor is absent in C. jejuni and molecular mechanisms responsible for transition of cells to the stationary phase remain elusive. In this work, proteomic profiles of cells from exponential and stationary phases were compared using 2-D electrophoresis (2DE) fingerprinting combined with mass spectrometry analysis and qRT-PCR. The identified proteins, whose expression differed between the two phases, are mostly involved in protein biosynthesis, carbon metabolism, stress response and motility. Altered expression was observed also in the pleiotropic regulator CosR that was over-expressed during stationary phase. A shift between transcript and protein level evolution of CosR throughout the growth of C. jejuni was observed using qRT-PCR and (2DE). From these data, we hypothesized that CosR could undergo a negative autoregulation in stationary phase. A consensus sequence resulting from promoter sequence alignment of genes potentially regulated by CosR, including its own upstream region, among C. jejuni strains is proposed. To verify experimentally the potential autoregulation of CosR at the DNA level, electrophoretic mobility shift assay was performed with DNA fragments of CosR promoter region and rCosR. Different migration pattern of the promoter fragments indicates the binding capacity of CosR, suggesting its auto-regulation potential.
Project description:Campylobacter jejuni is a major foodborne pathogen, and the LuxS-mediated quorum-sensing (QS) system influences its motility, biofilm formation, invasion, host colonization, and virulence. QS therefore represents a target for the control of C. jejuni. The aim of this study was to investigate the correlation of QS inhibition with changes in C. jejuni motility, adhesion to polystyrene surfaces, and adhesion to and invasion of INT407 cells. This was achieved by studying (i) the luxS-deficient mutant and (ii) treatment of C. jejuni with 20 natural extracts as six essential oils, 11 ethanolic extracts, and three pure compounds. Compared to the wild-type, the ΔluxS mutant showed decreased motility, adhesion to polystyrene surfaces, and invasion of INT407 cells. The anti-QS effects of the treatments (n = 15/20) were assayed using Vibrio harveyi BB170 bioluminescence. Moderate positive correlation was shown between C. jejuni QS reduction and reduced motility (τ = 0.492, p = 0.024), adhesion to polystyrene surfaces (τ = 0.419, p = 0.008), and invasion (r = 0.394, p = 0.068). The best overall effect was achieved with a Sedum rosea (roseroot) extract, with 96% QS reduction, a 1.41 log (96%) decrease in adhesion to polystyrene surfaces, and an 82% decrease in invasion. We show that natural extracts can reduce motility, adhesion to polystyrene surfaces, and invasion of INT407 cells by C. jejuni through modulation of the LuxS (QS) system.
Project description:Multivalency is a common principle in the recognition of cellular receptors, and multivalent agonists and antagonists have played a major role in understanding mammalian cell receptor biology. The study of bacterial cell receptors using similar approaches, however, has lagged behind. Herein we describe our efforts toward the development of a dendrimer-based multivalent probe for studying AI-2 quorum-sensing receptors. From these studies, we have discovered a chemical probe specific for Lsr-type AI-2 quorum-sensing receptors with the potential for enabling the identification of new bacterial species that utilize AI-2 as a quorum-sensing signaling molecule.
Project description:The marine bacterium Phaeobacter inhibens produces tropodithietic acid (TDA), a broad-spectrum antibiotic and anticancer agent. TDA allows P. inhibens to antagonize other bacteria, including several pathogens, and eukaryotes. Since recently antibiotics are also discussed to function as intermicrobial signals. Here we show that ~10% of the genes of P. inhibens are strongly influenced by N-acyl-homoserine lactone (AHL) mediated quorum sensing (QS), switching the bacteriumâs life style from attached to free-living. In an AHL negative mutant of P. inhibens subinhibitory concentrations of TDA caused the same regulatory effect as the AHL. This demonstrates that bacteria can produce antibiotic compounds not only as weapons, but also to substitute their endogenous AHL molecule in QS. The dual function of TDA probably supports the QS system to accelerate regulatory processes and points to a so far neglected role of antibiotics at subinhibitory concentrations in the environment and in microbial interactions. Comparison of whole transcriptomes of wildytype, quorum sensing mutants (pgaI and pgaR) and pgaI grown supplemented with subinhibitory concentration of the antibiotic TDA. RNA isolated in the late exponential growth phase. 4 biological replicates investigated for each strain.