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ABSTRACT: N-Acylhomoserine lactones (AHLs) are used as quorum-sensing (QS) signals by gram-negative bacteria. We have reported that the cyclic oligosaccharides known as cyclodextrins (CDs) form inclusion complexes with AHLs and disrupt QS signaling. In this study, a series of CD derivatives were designed and synthesized to improve the QS inhibitory activity over that of native CDs. The production of the red pigment prodigiosin by Serratia marcescens AS-1, which is regulated by AHL-mediated QS, was drastically decreased by adding 10 mg/ml 6-alkylacylamino-β-CD with an alkyl chain ranging from C7 to C12. An improvement in the QS inhibitory activity was also observed for 6-alkylamino-α- or γ-CDs and 2-alkylamino-CDs. Furthermore, 6,6'-dioctylamino-β-CD, which contains two octylamino groups, exhibited greater inhibitory activity than 6-monooctylamino-β-CD. The synthesized CD derivatives also had strong inhibitory effects on QS by other gram-negative bacteria, including Chromobacterium violaceum and Pseudomonas aeruginosa. The synthetic alkylamine-modified CD derivatives had higher equilibrium binding constants for binding with AHL than the native CDs did, consistent with the improved QS inhibition. 1H NMR measurements suggested that the alkyl side chains of 6-alkylacylamino-β-CDs with alkyl chains up to 6 carbon atoms long could form self-inclusion complexes with the CD unit.
Journal of Bioscience and Bioengineering 02/2013; · 1.79 Impact Factor
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ABSTRACT: N-Acylhomoserine lactones (AHLs) are used as quorum-sensing (QS) signal molecules by many Gram-negative bacteria. We have reported that Chryseobacterium sp. strain StRB126, which was isolated from the root surface of potato, has AHL-degrading activity. In this study, we cloned and characterized the aidC gene from the genomic library of StRB126. AidC has AHL-degrading activity and shows homology to several metallo-β-lactamase proteins from Bacteroidetes, although not to any known AHL-degrading enzymes. Purified AidC, as a maltose-binding fusion protein, showed high degrading activity against all tested AHLs, whether short- or long-chain forms, with or without substitution at carbon 3. High-performance liquid chromatography (HPLC) analysis revealed that AidC functions as an AHL lactonase catalyzing AHL ring opening by hydrolyzing lactones. An assay to determine the effects of covalent and ionic bonding showed that Zn(2+) is important to AidC activity both in vitro and in vivo. In addition, the aidC gene could also be PCR amplified from several other Chryseobacterium strains. In conclusion, this study indicated that the aidC gene, encoding a novel AHL lactonase, may be widespread throughout the genus Chryseobacterium. Our results extend the diversity and known bacterial hosts of AHL-degrading enzymes.
Applied and environmental microbiology 08/2012; 78(22):7985-92. · 3.69 Impact Factor
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Journal of the American Academy of Dermatology 08/2012; 67(2):e74-5. · 3.99 Impact Factor
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ABSTRACT: N-Acylhomoserine lactone (AHL)-degrading enzyme, AiiM, was identified from the potato leaf-associated Microbacterium testaceum StLB037. In this study, we cloned eight aiiM gene homologues from other AHL-degrading Microbacterium strains. The similarity of the chromosomal locus of the aiiM gene is associated with the phylogenetic classification based on 16S rRNA. Degenerate PCR revealed that the aiiM gene was only conserved in AHL-degrading Microbacterium strains, but not in fifteen Microbacterium type strains or two Microbacterium isolates from other plants. These results suggested that the high level of AHL-degrading activity in Microbacterium strains was caused by the aiiM gene encoded on their chromosome.
Microbes and Environments 03/2012; 27(3):330-3. · 1.91 Impact Factor
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ABSTRACT: N-Acylhomoserine lactones (AHLs) are used as quorum-sensing signal molecules by many gram-negative bacteria. We have reported that Solibacillus silvestris, which was isolated from the potato leaf, has AHL-degrading activity. To identify the AHL-degrading gene, whole genome sequencing of S. silvestris StLB046 was performed by using pyrosequencing technology. As the result of the BLAST search, one predicted ORF (ahlS) showed slight similarity to AiiA-like AHL lactonase from Bacillus cereus group. Escherichia coli harboring the ahlS-expressing plasmid showed high AHL-degrading activity. The ahlS-cording region was also amplified by PCR from the other potato leaf-associated and AHL-degrading S. silvestris strains. Purified AhlS as a maltose binding fusion protein showed high AHL-degrading activity and catalyzes AHL ring opening by hydrolyzing lactones. In addition, expression of ahlS in plant pathogen Pectobacterium carotovorum subsp. carotovorum attenuated maceration of the potato slices. Our results suggest that AHL-degrading activity of ahlS might perform useful functions such as useful biocontrol agents.
Journal of Bioscience and Bioengineering 01/2012; 113(1):20-5. · 1.79 Impact Factor
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ABSTRACT: A total of 828 isolates of fluorescent pseudomonads (FPs) were obtained from the leaves (305 isolates) and roots (523 isolates) of potato plants grown in different geographical locations in Japan, and 16S rRNA gene sequences of 776 isolates were successfully determined by direct PCR sequencing. Clustering analysis (≥99% identity) identified 13 and 26 operational taxonomic units (OTUs) for leaf- and root-associated FPs, respectively, and 29 OTUs were identified in the phytosphere of potato plants. Among them, 7 and 9 OTUs showed a significantly biased distribution to the leaves and roots, respectively. Phylogenetic analysis revealed that 3 dominant OTUs for leaf-associated FPs were grouped in a cluster of leaf-associated pathogens, such as Pseudomonas cichorii and Pseudomonas viridiflava. In contrast, 4 OTUs were located in a cluster of saprophytic pseudomonads. Among them, 3 OTUs showed high similarity to Pseudomonas koreensis and Pseudomonas vancouverensis, both of which have been reported to be beneficial for biological control or plant growth promotion. These data provide key information for efficient surveying and utilization of beneficial FPs in agricultural practices.
Microbes and Environments 01/2012; 27(2):122-6. · 1.91 Impact Factor
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ABSTRACT: Pantoea ananatis SK-1 produces N-acylhomoserine lactones (AHLs) and regulates expression of some virulence factors through AHL-mediated quorum sensing. In this study, we discovered that the strain SK-02I, which has a disrupted AHL-synthetic gene, showed constitutive cell aggregation. SK-1 has the ability to aggregate, and cell aggregation inhibitory factors are expressed under control of AHL-mediated quorum sensing. One of the transposon mutants, SK-33M, constitutively aggregates without defective AHL production. A homology search revealed that the transposon integration site was located in the adhesin-like yeeJ gene. Based on RT-PCR analysis, transcription of yeeJ is regulated by AHL-mediated quorum sensing. However, because both the wild-type and SK-33M strains induced necrotic symptoms in onion leaves, we conclude that the yeeJ gene is not involved in the pathogenicity of SK-1.
Journal of Bioscience and Bioengineering 09/2011; 112(6):566-9. · 1.79 Impact Factor
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ABSTRACT: Members of the genus Nepenthes are carnivorous plants that use the pitfall method of insect capture as a supplementary nutritional source. We extracted metagenomic DNA from the microbial community found in the pitcher fluid of Nepenthes and constructed a plasmid-based metagenomic library. An activity-based screening method enabled the isolation of two lipase genes, lip1 and lip2. Both Lip1 and Lip2 belong to a novel family or subfamily of lipases and show lipase activities in acidic conditions, such as those found in pitcher fluid. This study was conducted under the assumption that the secreted Lip1 and Lip2 were capable of enzymatic activity in the acidic pitcher fluid.
Journal of Bioscience and Bioengineering 07/2011; 112(4):315-20. · 1.79 Impact Factor
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ABSTRACT: N-Acylhomoserine lactones (AHLs) are used as quorum-sensing signaling molecules by many Gram-negative bacteria. Here, 413 bacterial strains were obtained from the roots of potato plants and screened for AHL-degrading bacteria using Chromobacterium violaceum reporter strains. Sixty one isolates degraded N-hexanoyl-L-homoserine lactone (C6-HSL) within 24 h. Based on their 16S rRNA gene sequences, most of these isolates were assigned to the genus Chryseobacterium and divided into eight groups. Most of the strains degraded N-decanoyl-L-homoserine lactone (C10-HSL) and were active against 3-oxo-substituted AHLs. Groups V and VI were more effective at inactivating AHLs. One group V strain, StRB402, showed putative AHL-lactonase activity. This is the first report of AHL-degrading Chryseobacterium strains.
Microbes and Environments 06/2011; 26(2):144-8. · 1.91 Impact Factor
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ABSTRACT: Pantoea ananatis SK-1 produced EPS by AHL-mediated quorum sensing on an LB agar plate containing glucose, fructose, and sucrose. rcsA and rcsB mutants did not produce EPS with or without AHLs and with or without sugars, but they induced necrotic symptoms in onion leaves. These results indicate that EPS production does not relate to the pathogenicity of SK-1.
Bioscience Biotechnology and Biochemistry 05/2011; 75(5):997-9. · 1.28 Impact Factor
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ABSTRACT: The impact of quorum sensing (QS) in in vivo models of infection has been widely investigated, but there are no descriptions for ischemic wound infection. To explore the role of QS in Pseudomonas aeruginosa in the establishment of ischemic wound infection, we challenged a pressure ulcer model in rats with the PAO-1, PAO-1 derivatives ΔlasIΔrhlI and ΔlasRΔrhlR strains, which cannot induce the virulence factor under QS control, thus the reduced tissue destruction was expended in these mutant strains. However unexpectedly, on postwounding day 3, the inflammatory responses in the three groups were similarly severe and the numbers of bacteria in tissue samples did not differ among the three strains. Biofilm formation was immature in QS-deficient strains, defined by the absence of dense bacterial aggregates and extracellular polymeric substance, which was confirmed by scanning electron microscopy. The Pseudomonas aeruginosa QS signal, acylated homoserine lactone, was only quantified from wound samples in the PAO-1 group. The swimming and twitching motilities were significantly enhanced in the ΔlasRΔrhlR group compared with the PAO-1 group in vitro. A significantly larger wound area was correlated with the bacterial motility. The inflammation in the early phase of bacterial challenge to wounds with immature biofilm formation in the QS-deficient strains indicated that the role of QS was more crucial for the chronic phase than for the acute phase of infection. The present findings indicate a difference in the importance of QS in ischemic wound infections compared with other infection models.
Wound Repair and Regeneration 03/2011; 19(2):214-22. · 2.91 Impact Factor
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ABSTRACT: Microbacterium testaceum is an endophytic Gram-positive bacterium that resides within plant hosts. M. testaceum StLB037 was isolated from potato leaves and shows N-acylhomoserine lactone-degrading activity. Here, we present the 3.98-Mb complete genome sequence of StLB037, with an average GC content of 70.28%.
Journal of bacteriology 02/2011; 193(8):2072-3. · 3.94 Impact Factor
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ABSTRACT: Quorum sensing is a cell density-dependent gene regulation system in bacteria. N-(3-oxododecanoyl) homoserine lactone (3-oxo-C12-HSL) is used in the las quorum-sensing system in Pseudomonas aeruginosa, which is an opportunistic pathogen that causes many human diseases. Although many studies have investigated the sole effects of quorum sensing on several types of mammalian cells, including lung cells, little is known about the effects of quorum sensing on the cells associated with wound healing. To better understand the mechanism of bacterial wound infection, we investigated the effects of 3-oxo-C12-HSL on cells using a rat full-thickness wound-healing model. We found that the wound contraction was significantly increased at 24 h after the administration of 3-oxo-C12-HSL to the surface of granulation tissue. Differentiation of fibroblasts to myofibroblasts was induced in the in vivo wound-healing model and was confirmed in vitro using the rat fibroblastic cell line Rat-1. Cyclooxygenase (Cox)-2 expression was also induced in Rat-1 cells by 3-oxo-C12-HSL. This finding suggested that Cox-2 upregulation may be related to the inflammatory findings in the histological examinations, in which infiltrating polymorphonuclear neutrophils were observed at the wound site. Taken together, these results imply that mammals have a potential defense system against invading pathogens by responding to the presence of 3-oxo-C12-HSL and inducing the differentiation of fibroblasts to myofibroblasts as well as inflammation for accelerating wound healing.
FEMS Immunology & Medical Microbiology 02/2011; 62(2):157-63. · 2.44 Impact Factor
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ABSTRACT: A total of 100 isolates of chitinolytic bacteria were obtained from the rhizospheres of various agronomic plants, and the 16S rRNA gene sequences of these isolates were determined. Phylogenetic analyses revealed that 81 isolates belonged to the classes Betaproteobacteria (39 isolates) and Gammaproteobacteria (42 isolates). Of the remaining 19 isolates, 16 belonged to the phylum Firmicutes. Clustering analysis identified 6 and 3 operational taxonomic units (OTUs) in Gammaproteobacteria and Betaproteobacteria, respectively, at the genus level. The majority of chitinolytic bacteria in Gammaproteobacteria belonged to the genera Serratia, Stenotrophomonas, and Lysobacter (14, 15, and 7 isolates, respectively) while those in Betaproteobacteria belonged to the genus Mitsuaria (37 isolates). The 16 isolates placed in Firmicutes belonged to 2 genera, Paenibacillus and Bacillus (8 isolates each). The isolates in the remaining OTUs belonged to the genera Erwinia, Aeromonas, Pseudomonas, Achromobacter, Flavobacterium, and Microbacterium, in less abundance. These results showed a wide distribution of culturable chitinolytic bacteria in the rhizospheres of various agronomic plants. Considering the potential antagonistic activity of chitinolytic enzymes against phytopathogenic fungi, which is exhibited by fungal cell wall degradation, the above-mentioned native chitinolytic bacteria in rhizospheres could potentially be utilized for the biological control of soil-borne phytopathogenic fungi.
Microbes and Environments 01/2011; 26(1):7-14. · 1.91 Impact Factor
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ABSTRACT: In terms of a piecewise affine system representation, which is a kind of hybrid system model, this article discusses a series of approaches to modelling, analysing and synthesizing a biological network such as a gene-regulatory network. First, the input assignment problem, the controllable state set problem (CSP) and the input trajectory generation problem are emphasized as control problems to be addressed for biological networks. Subsequently, after the modelling issue on biological networks developed in the systems and control community is briefly explained, the CSP is described in detail with reference to control of the quorum-sensing system in the pathogen Pseudomonas aeruginosa. Finally, an optimal control design method to the quorum-sensing system is proposed as a solution to the input trajectory generation problem.
Philosophical Transactions of The Royal Society A Mathematical Physical and Engineering Sciences 11/2010; 368(1930):4977-93. · 2.77 Impact Factor
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ABSTRACT: Chromobacterium violaceum produces the purple pigment violacein by quorum-sensing regulation. 20-bp of the lux box-like sequence was found upstream of vioA in C. violaceum ATCC 12472. CviR received C10-HSL and C6-HSL and activated the transcription of vioA in Escherichia coli. However, in strain ATCC 12472, C6-HSL inhibited both C10-HSL-mediated violacein production and the transcription of vioA.
Bioscience Biotechnology and Biochemistry 10/2010; 74(10):2116-9. · 1.28 Impact Factor
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ABSTRACT: N-Acylhomoserine lactones (AHLs) are used as quorum-sensing signal molecules by many Gram-negative bacteria. We have reported that Microbacterium testaceum StLB037, which was isolated from the leaf surface of potato, has AHL-degrading activity. In this study, we cloned the aiiM gene from the genomic library of StLB037, which has AHL-degrading activity and shows high homology with the alpha/beta hydrolase fold family from Actinobacteria. Purified AiiM as a maltose binding fusion protein showed high degrading activity of AHLs with both short- and long-chain AHLs with or without substitution at carbon 3. High-performance liquid chromatography analysis revealed that AiiM works as an AHL lactonase that catalyzes AHL ring opening by hydrolyzing lactones. In addition, expression of AiiM in the plant pathogen Pectobacterium carotovorum subsp. carotovorum reduced pectinase activity markedly and attenuated soft rot symptoms on potato slices. In conclusion, this study indicated that AiiM might be effective in quenching quorum sensing of P. carotovorum subsp. carotovorum.
Applied and environmental microbiology 02/2010; 76(8):2524-30. · 3.69 Impact Factor
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ABSTRACT: Edwardsiella tarda is a gram-negative bacterial pathogen of fish and animals. A number of gram-negative bacteria have quorum-sensing systems and produce N-acylhomoserine lactone (AHL) that they use as a quorum-sensing signaling molecule. We have already reported that E. tarda NUF251 produces AHLs and has the AHL-synthase gene, edwI. Inactivation of NUF251 edwI induces expression of an approximately 45 kDa extracellular protein, identified as a flagellin encoded by FliC. Mutation of edwI also changes the motility pattern of NUF251 from a radial expansion pattern to concentric rings. The addition of exogenous AHL was capable of restoring normal motility to NUF251 edwI mutants. These results demonstrate that quorum sensing negatively regulates motility and expression of the FliC protein.
Journal of Bioscience and Bioengineering 11/2009; 108(4):314-8. · 1.79 Impact Factor
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ABSTRACT: We isolated and identified AHL-degrading bacteria from the leaf surface of Solanum tuberosum. The 16 isolates inactivated both short- and long-chain AHLs. Two of these isolates, identified as Microbacterium testaceum, showed putative AHL-lactonase activity. These two strains interrupted quorum-sensing dependent bacterial infection by plant pathogen Pectobacterium carotovorum subsp. carotovorum. M. testaceum strains, which were isolated in this study, might be useful in the biocontrol of plant diseases.
Bioscience Biotechnology and Biochemistry 10/2009; 73(9):2124-7. · 1.28 Impact Factor
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ABSTRACT: Temperature-induced phase separation (TIPS) technique was applied to create cyclodextrin (CD) immobilized 2-hydroxypropyl
cellulose (HPC) gel sheets. A heterogeneous polymer network could be fixed when the reaction temperature was stepwise increased
above a lower critical solution temperature (LCST) of the HPC polymer at approximately 40°C during the cross-linking reaction.
Dynamics of polymer network formation was characterized by measuring the viscosity change of the pre-gel solution with or
without the TIPS process. A temperature increase is responsible for drastic increase of the viscosity of which change was
translated as the acceleration of the cross-linking reaction rates. The immobilized α-CD or hydroxypropyl-β-CD (HP-β-CD) onto
the TIPS-generated polymer network effectively decreased the production of red pigment prodigiosin that was one of the second
metabolites through the cell-population-density dependent quorum sensing (QS) system in Serratia marcescens AS-1. Since virulence expression in some opportunistic pathogens was regulated by diffusible acylhomoserine lactone (AHL)
mediated QS system, trapping AHLs onto the host matrices could make the CD-immobilized gels interrupt the hierarchical QS
system from outside of cells. The AHL-mediated prodigiosin production could be drastically decreased to approximately 10%
using the CD-AHL inclusion complex formation when the TIPS process was applied to HPC/α-CD gel synthesis with optimized condition
of the phase separation.
KeywordsCell-to-cell communication–Quorum sensing–Cyclodextrin–Gene expression control–
N-Acylhomoserine lactone
08/2009: pages 155-162;
