Metabolites formed during anaerobic transformation of toluene and o-xylene and their proposed relationship to the initial steps of toluene mineralization.
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Strain T1 is a facultative bacterium that is capable of anaerobic toluene degradation under denitrifying conditions. While 80% of the carbon from toluene is either oxidized to carbon dioxide or assimilated into cellular carbon, a significant portion of the remainder is transformed into two dead-end metabolites. These metabolites were produced simultaneous to the mineralization of toluene and were identified as benzylsuccinic acid and benzylfumaric acid. Identification was based on comparison of mass spectra of the methyl esters of the metabolites and authentic compounds that were chemically synthesized. Strain T1 is also capable of o-xylene transformation during growth on toluene. o-Xylene does not serve as a source of carbon and is not mineralized. Rather, it is transformed to analogous dead-end metabolites, (2-methylbenzyl)-succinic acid and (2-methylbenzyl)-fumaric acid. o-Xylene transformation also occurred during growth on succinic acid, which suggests that attack of the methyl group by succinyl-coenzyme A is a key reaction in this transformation. We reason that the main pathway for toluene oxidation to carbon dioxide involves a mechanism similar to that for the formation of the metabolites and involves an attack of the methyl group of toluene by acetyl-coenzyme A. (+info)
Chromobacterium violaceum in siblings, Brazil.
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Chromobacterium violaceum, a saprophyte bacterium found commonly in soil and water in tropical and subtropical climates, is a rare cause of severe, often fatal, human disease. We report 1 confirmed and 2 suspected cases of C. violaceum septicemia, with 2 fatalities, in siblings after recreational exposure in northeastern Brazil. (+info)
N-butanoyl-L-homoserine lactone (BHL) deficient Pseudomonas aeruginosa isolates from an intensive care unit.
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Acylated homoserine lactones (AHLs) are self-generated diffusible signal molecules that mediate population density dependent gene expression (quorum sensing) in a variety of Gram-negative bacteria, and several virulence genes of human pathogens are known to be controlled by AHLs. In this study, strains of Pseudomonas aeruginosa, Acinetobacter baumannii, Escherichia coli and Klebsiella pneumoniae, isolated from intensive care patients, were screened for AHL production by using AHL responsive indicator strains of Chromobacterium violaceum CV026 and Agrobacterium tumefaciens NT1. Positive reactions were recorded for all 50 isolates of P. aeruginosa and 10 isolates of Acinetobacter baumannii with Agrobacterium tumefaciens NT1. Surprisingly, most P. aeruginosa isolates gave negative results with C. violaceum CV026 in contrast to previous reports. This suggests that the new isolates of P. aeruginosa either failed to make short chain AHLs or the level of the signal molecule is very low. (+info)
A case of Chromobacterium infection after car accident in Korea.
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Chromobacterium violaceum is a gram negative straight rod, 0.8-1.2 by 2.5 to 6.0 m, which is motile by one polar flagella and one to four lateral flagella. The organism inhabits soil and water and is often found in semitropical and tropical climates. Infections in humans are rare. We report a case of infection caused by strains of C. violaceum. A 38-year-old male patient was admitted to KyungHee University Hospital, Seoul, Korea on July 28th, 2003, after a car accident. The patient had multiple trauma and lacerations. He had an open wound in the left tibial area from which C. violaceum was isolated. The strain was resistant to ampicillin, tobramycin, ampicillin/sulbactam, ceftriaxone and cefepime, but was susceptible to amikacin, gentamicin, ciprofloxacin, levofloxacin, trimethoprim/sulfamethoxazole and piperacillin/tazobactam. The patient was treated successfully by debridement, cephapirin sodium and astromicine sulfate. (+info)
L-Canavanine made by Medicago sativa interferes with quorum sensing in Sinorhizobium meliloti.
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Sinorhizobium meliloti is a gram-negative soil bacterium, capable of establishing a nitrogen-fixing symbiosis with its legume host, alfalfa (Medicago sativa). Quorum sensing plays a crucial role in this symbiosis, where it influences the nodulation process and the synthesis of the symbiotically important exopolysaccharide II (EPS II). S. meliloti has three quorum-sensing systems (Sin, Tra, and Mel) that use N-acyl homoserine lactones as their quorum-sensing signal molecule. Increasing evidence indicates that certain eukaryotic hosts involved in symbiotic or pathogenic relationships with gram-negative bacteria produce quorum-sensing-interfering (QSI) compounds that can cross-communicate with the bacterial quorum-sensing system. Our studies of alfalfa seed exudates suggested the presence of multiple signal molecules capable of interfering with quorum-sensing-regulated gene expression in different bacterial strains. In this work, we choose one of these QSI molecules (SWI) for further characterization. SWI inhibited violacein production, a phenotype that is regulated by quorum sensing in Chromobacterium violaceum. In addition, this signal molecule also inhibits the expression of the S. meliloti exp genes, responsible for the production of EPS II, a quorum-sensing-regulated phenotype. We identified this molecule as l-canavanine, an arginine analog, produced in large quantities by alfalfa and other legumes. (+info)
Cell-cell influences on bacterial community development in aquatic biofilms.
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Dialysis tubing containing spent culture media, when placed in a lake, was colonized by a low diversity of bacteria, whereas abiotic controls had considerable diversity. Changes were seen in the presence and absence of acylated homoserine lactones, suggesting that these molecules and other factors may influence adherent-population composition. (+info)
Production and properties of the native Chromobacterium violaceum fucose-binding lectin (CV-IIL) compared to homologous lectins of Pseudomonas aeruginosa (PA-IIL) and Ralstonia solanacearum (RS-IIL).
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Chromobacterium violaceum is a versatile, violet pigment (violacein)-producing beta-proteobacterium, confined to tropical and subtropical regions, dwelling in soil and water, like Pseudomonas aeruginosa and Ralstonia solanacearum. These three bacteria are saprophytes that occasionally become aggressive opportunistic pathogens virulently attacking animals (the first two) and plants (the third). The recent availability of their genome sequences enabled identification in the C. violaceum genome of an ORF (locus no. 1744) that is similar to those of P. aeruginosa and R. solanacearum lectins, PA-IIL and RS-IIL, respectively. A recombinant protein, CV-IIL, encoded by that ORF exhibited fucose>mannose-specific lectin activity resembling PA-IIL. This paper describes production and properties of the native CV-IIL, which, like PA-IIL and RS-IIL, is probably also a quorum-sensing-driven secondary metabolite, appearing concomitantly with violacein. Its formation is repressed in the CV026 mutant of C. violaceum, which lacks endogenous N-acylhomoserine lactone. The upstream extragenic sequence of its ORF contains a 20 bp sequence (5'-101-120) with partial similarities to the luxI-box and the related P. aeruginosa and R. solanacearum promoter boxes of quorum-sensing-controlled genes. The lectin level is augmented by addition of trehalose to the medium. The subunit size of CV-IIL (around 11.86 kDa) is similar to those of PA-IIL (11.73 kDa) and RS-IIL (11.60 kDa). Like PA-IIL, in the tetrameric form CV-IIL preferentially agglutinates alpha1-2 fucosylated H-positive human erythrocytes (regardless of their A, B or O type), as opposed to the O(h) Bombay type, but differs from it in having no interaction with rabbit erythrocytes and in displaying stronger affinity to l-galactose than to l-fucose. The greater similarity of CV-IIL to PA-IIL than to RS-IIL might be related to the selective adaptation of both C. violaceum and P. aeruginosa to animal tissues versus the preferential homing of R. solanacearum to plants. (+info)
Cloning and expression of Chromobacterium violaceum phenylalanine hydroxylase in Escherichia coli and comparison of amino acid sequence with mammalian aromatic amino acid hydroxylases.
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The complete amino acid sequence (296 amino acids) of Chromobacterium violaceum phenylalanine hydroxylase (PAH) was determined by nucleotide analysis of a DNA clone isolated using both a synthetic oligonucleotide probe based on the NH2-terminal amino acid sequence and an antibody against this enzyme. The ApaL I fragment (approximately 1.9 kilobase pairs) containing the entire PAH gene was subcloned in pBluescript II and induced by isopropyl-beta-D-thiogalactopyranoside. In order to eliminate fusion proteins the XbaI/ClaI fragment which contained the PAH gene from the Bluescript construct was subcloned into pMAC 5-8 containing the TAC promoter. The recombinant protein reacts with antibody raised to authentic C. violaceum PAH and its NH2-terminal 20-amino acid sequence and COOH-terminal amino acid residue were identical with the wild-type protein. Key physical and chemical characteristics of the recombinant protein, i.e. its copper content and Michaelis-Menten parameters, were the same as wild-type. Comparison of amino acid sequences revealed a highly conserved region between C. violaceum PAH and three different mammalian aromatic amino acid hydroxylases. This conserved area may well be a catalytically important domain of these pterin- and metal-requiring aromatic amino acid hydroxylases. The over-expression of C. violaceum PAH in Escherichia coli will facilitate the analysis of the enzyme mechanism by various spectroscopic methods. (+info)