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Putida
Kinds of Putida Terms modified by Putida Selected AbstractsGas Diffusion Electrodes for Use in an Amperometric Enzyme BiosensorELECTROANALYSIS, Issue 21 2008Martin Hämmerle Abstract The preparation of gas diffusion electrodes and their use in an amperometric enzyme biosensor for the direct detection of a gaseous analyte is described. The gas diffusion electrodes are prepared by covering a PTFE membrane (thickness 250,,m, pore size 2,,m, porosity 35%) with gold, platinum, or a graphite/PTFE mixture. Gold and platinum are deposited by e-beam sputtering, whereas the graphite/PTFE layer is prepared by vacuum filtration of a respective aqueous suspension. These gas diffusion electrodes are exemplarily implemented as working electrodes in an amperometric biosensor for gaseous formaldehyde containing NAD-dependent formaldehyde dehydrogenase from P. putida [EC. 1.2.1.46] as enzyme and 1,2-naphthoquinone-4-sulfonic acid as electrochemical mediator. The resulting sensors are compared with regard to background current, signal noise, linear range, sensitivity, and detection limit. In this respect, sensors with gold or graphite/PTFE covered membranes outclass ones with platinum for this particular analyte and sensor configuration. [source] Dynamics of Benzene and Toluene Degradation in Pseudomonas putida F1 in the Presence of the Alternative Substrate SuccinateENGINEERING IN LIFE SCIENCES (ELECTRONIC), Issue 4 2007I. Rüegg Abstract In batch and continuous culture, the regulation of benzene and toluene degradation by Pseudomonas putida,F1 was investigated in the presence of the alternative carbon and energy source succinate. In batch culture, benzene and toluene were used simultaneously, whereas succinate suppressed benzene consumption under carbon excess conditions resulting in diauxic growth. In carbon-limited continuous culture mixed substrate growth was observed. Since in nature low substrate concentrations and ever changing conditions prevail, this paper focuses on the dynamics of benzene/toluene degradation, biomass synthesis, and the regulation of benzene/toluene-degrading enzymes in cultures growing continuously at a dilution rate of 0.1,h,1, when shifting the supply of the carbon and energy source from succinate to various mixtures of succinate and benzene/toluene, or to benzene only. When the succinate concentration was kept constant (1.25,mM) and the medium was supplemented with benzene (2,mM), growth with benzene began already two hours after the shift. In contrast, replacing succinate with benzene only led to a wash out of biomass for more then ten hours, before biomass production from benzene started. A striking and reproducible transition pattern was observed for all shifts where the succinate concentration was reduced or succinate was omitted. After an initial period of biomass production from benzene, the culture collapsed and a wash-out of biomass was observed. However, this wash-out was not accompanied by an increase in benzene in the cultivation liquid, indicating a benzene uptake without conversion into biomass. Another possibility is that in phases of low biomass concentrations, cells were only able to use the low amounts of benzene/toluene dissolved in the cultivation liquid yielding low biomass concentrations whereas in phases of high biomass concentrations, they were able to rapidly utilize the aromatic solvents so that additional benzene from the gas phase diffused into the cultivation liquid resulting in more biomass production. In most cases, growth resumed again after 10 to 80,h. Currently, the reasons for the decrease in biomass after the first rise are unknown. However, several indications rule out intoxication of the cells by either the solvents benzene or toluene themselves, or by toxic degradation intermediates, or by-products. [source] DNA sequence-based analysis of the Pseudomonas speciesENVIRONMENTAL MICROBIOLOGY, Issue 6 2010Magdalena Mulet Summary Partial sequences of four core ,housekeeping' genes (16S rRNA, gyrB, rpoB and rpoD) of the type strains of 107 Pseudomonas species were analysed in order to obtain a comprehensive view regarding the phylogenetic relationships within the Pseudomonas genus. Gene trees allowed the discrimination of two lineages or intrageneric groups (IG), called IG P. aeruginosa and IG P. fluorescens. The first IG P. aeruginosa, was divided into three main groups, represented by the species P. aeruginosa, P. stutzeri and P. oleovorans. The second IG was divided into six groups, represented by the species P. fluorescens, P. syringae, P. lutea, P. putida, P. anguilliseptica and P. straminea. The P. fluorescens group was the most complex and included nine subgroups, represented by the species P. fluorescens, P. gessardi, P. fragi, P. mandelii, P. jesseni, P. koreensis, P. corrugata, P. chlororaphis and P. asplenii. Pseudomonas rhizospherae was affiliated with the P. fluorescens IG in the phylogenetic analysis but was independent of any group. Some species were located on phylogenetic branches that were distant from defined clusters, such as those represented by the P. oryzihabitans group and the type strains P. pachastrellae, P. pertucinogena and P. luteola. Additionally, 17 strains of P. aeruginosa, ,P. entomophila', P. fluorescens, P. putida, P. syringae and P. stutzeri, for which genome sequences have been determined, have been included to compare the results obtained in the analysis of four housekeeping genes with those obtained from whole genome analyses. [source] Lack of CbrB in Pseudomonas putida affects not only amino acids metabolism but also different stress responses and biofilm developmentENVIRONMENTAL MICROBIOLOGY, Issue 6 2010Cristina I. Amador Summary The CbrAB two-component system has been described in certain species of Pseudomonads as a global regulatory system required for the assimilation of several amino acids (e.g. histidine, proline or arginine) as carbon or carbon and nitrogen sources. In this work, we used global gene expression and phenotypic analyses to characterize the roles of the CbrAB system in Pseudomonas putida. Our results show that CbrB is involved in coordination with the nitrogen control system activator, NtrC, in the uptake and assimilation of several amino acids. In addition, CbrB affects other carbon utilization pathways and a number of apparently unrelated functions, such as chemotaxis, stress tolerance and biofilm development. Based on these new findings, we propose that CbrB is a high-ranked element in the regulatory hierarchy of P. putida that directly or indirectly controls a variety of metabolic and behavioural traits required for adaptation to changing environmental conditions. [source] New alk genes detected in Antarctic marine sedimentsENVIRONMENTAL MICROBIOLOGY, Issue 3 2009Emanuele Kuhn Summary Alkane monooxygenases (Alk) are the key enzymes for alkane degradation. In order to understand the dispersion and diversity of alk genes in Antarctic marine environments, this study analysed by clone libraries the presence and diversity of alk genes (alkB and alkM) in sediments from Admiralty Bay, King George Island, Peninsula Antarctica. The results show a differential distribution of alk genes between the sites, and the predominant presence of new alk genes, mainly in the pristine site. Sequences presented 53.10,69.60% nucleotide identity and 50.90,73.40% amino acid identity to alkB genes described in Silicibacter pomeroyi, Gordonia sp., Prauserella rugosa, Nocardioides sp., Rhodococcus sp., Nocardia farcinica, Pseudomonas putida, Acidisphaera sp., Alcanivorax borkumensis, and alkM described in Acinetobacter sp. This is the first time that the gene alkM was detected and described in Antarctic marine environments. The presence of a range of previously undescribed alk genes indicates the need for further studies in this environment. [source] Stable implantation of orthogonal sensor circuits in Gram-negative bacteria for environmental releaseENVIRONMENTAL MICROBIOLOGY, Issue 12 2008Aitor De Las Heras Summary A broad host range, orthogonal genetic platform has been developed to format sensor circuits in the chromosome of Gram-negative microorganisms destined for environmental release as bioindicators of toxic or perilous compounds (e.g. explosives) in soil. The genetic scheme includes the generation of a genomic landing pad for the sensor module with a Tn5 -mini-transposon bearing an optimal attTn7 sequence and a choice of reporter systems with optical and enzymatic outputs. The array of functional elements thereby inserted in the chromosome match that of a cognate plasmid vector which delivers the transcription factors and the promoters to a frame that places the regulatory parts in front of the reporters. Site-specific recombination sites allow the deletion of antibiotic resistances and enables reporter output prior to deliberate release. The system thus allows the production and maintenance of cells in a pre-release state and its intentional conversion in deliverable strains that fulfil all safety, stability and performance criteria. The combination of such a genetic platform with a variant of the transcriptional regulator XylR of Pseudomonas putida that responds to 2,4-dinitrotoluene has been the basis for the production of strains that emit light upon exposure to residues of explosives in a soil microcosm. [source] Characterizing the regulation of the Pu promoter in Acinetobacter baylyi ADP1ENVIRONMENTAL MICROBIOLOGY, Issue 7 2008Wei E. Huang Summary Effective gene trapping and screening requires sensory and regulatory compatibility of both host and exogenous systems. The naturally competent bacterium Acinetobacter baylyi ADP1 is able to efficiently take up and integrate exogenous DNA into the chromosome, making it an attractive host system for a wide range of metagenomic applications. To test the ability of A. baylyi ADP1 to express the XylR-regulated Pu promoter from Pseudomonas putida mt-2, we have constructed and examined an A. baylyi ADP1 strain, ADPWH- Pu-lux-xylR. The Pu promoter in ADPWH- Pu-lux-xylR was specifically induced by toluene, m -, p - and o- xylene. The substrate-induced Pu promoter was highly dependent on the growth medium: it was repressed in rich media until stationary phase, but was immediately induced in minimal medium with glucose as the sole carbon source (MMG). However, the Pu promoter was repressed in MMG when it was supplemented with 5 g l,1 yeast extract. Further investigation showed that the Pu promoter in MMG was repressed by 0.5 g l,1 aspartic acid or asparagine, but not repressed by glutamine. Changing the carbon/nitrogen ratios by addition of ammonia did not significantly affect the Pu promoter activity but addition of nitrate did. These results show that A. baylyi ADP1 reproduced characteristics of the XylR-regulated Pu promoter observed in its original host. It demonstrates that A. baylyi could provide an excellent genetic host for a wide range of functional metagenomic applications. [source] Temperature and pyoverdine-mediated iron acquisition control surface motility of Pseudomonas putidaENVIRONMENTAL MICROBIOLOGY, Issue 7 2007Miguel A. Matilla Summary Pseudomonas putida KT2440 is unable to swarm at its common temperature of growth in the laboratory (30°C) but exhibits surface motility similar to swarming patterns in other Pseudomonas between 18°C and 28°C. These motile cells show differentiation, consisting on elongation and the presence of surface appendages. Analysis of a collection of mutants to define the molecular determinants of this type of surface movement in KT2440 shows that while type IV pili and lipopolysaccharide O-antigen are requisites flagella are not. Although surface motility of flagellar mutants was macroscopically undistinguishable from that of the wild type, microscopy analysis revealed that these mutants move using a distinct mechanism to that of the wild-type strain. Mutants either in the siderophore pyoverdine (ppsD) or in the FpvA siderophore receptor were also unable to spread on surfaces. Motility in the ppsD strain was totally restored with pyoverdine and partially with the wild-type ppsD allele. Phenotype of the fpvA strain was not complemented by this siderophore. We discuss that iron influences surface motility and that it can be an environmental cue for swarming-like movement in P. putida. This study constitutes the first report assigning an important role to pyoverdine iron acquisition in en masse bacterial surface movement. [source] The ttgGHI solvent efflux pump operon of Pseudomonas putida DOT-T1E is located on a large self-transmissible plasmidENVIRONMENTAL MICROBIOLOGY, Issue 6 2007José J. Rodríguez-Herva Summary Pseudomonas putida DOT-T1E is a solvent-tolerant strain able to grow in the presence of > 1% (v/v) toluene in the culture medium. A set of multidrug efflux pumps have been found to play a major role in the tolerance of this bacterium to organic solvents (Rojas et al., J Bacteriol 183: 3967,3973). In the course of studies of the mechanisms underlying solvent tolerance in DOT-T1E, we isolated a spontaneous solvent-sensitive mutant derivative which had lost the genes encoding the TtgGHI efflux pump, the most important extrusion element in quantitative terms. Genomic comparisons between the mutant and its parental strain by microarray analysis revealed that in addition to the ttgVW-ttgGHI gene cluster, another group of genes, highly similar to those found in the Tn4653A and ISPpu12 transposable elements of the TOL plasmid pWW0 from P. putida mt-2, were also absent from this strain. Further analysis demonstrated that strain DOT-T1E harboured a large plasmid (named pGRT1) that was lost from the solvent-sensitive mutant. Mapping analysis revealed that the ttgVW-ttgGHI genes and the Tn4653A -like transposon are borne by the pGRT1 plasmid. Plasmid pGRT1 is highly stable and its frequency of loss is below 10,8 per cell per generation under a variety of growth conditions, including nutritional and physical stresses. The pGRT1 plasmid is self-transmissible, and its acquisition by the toluene-sensitive P. putida KT2440 and Pseudomonas aeruginosa PAO1 increased the recipient's tolerance to toluene up to levels similar to those exhibited by P. putida DOT-T1E. We discuss the importance and potential benefits of this plasmid for the development of bacteria with enhanced solvent tolerance, and its potential impact for bioremediation and whole-cell biotransformations. [source] Biofilm formation and cellulose expression among diverse environmental Pseudomonas isolatesENVIRONMENTAL MICROBIOLOGY, Issue 11 2006Susanne Ude Summary The ability to form biofilms is seen as an increasingly important colonization strategy among both pathogenic and environmental bacteria. A survey of 185 plant-associated, phytopathogenic, soil and river Pseudomonas isolates resulted in 76% producing biofilms at the air,liquid (A,L) interface after selection in static microcosms. Considerable variation in biofilm phenotype was observed, including waxy aggregations, viscous and floccular masses, and physically cohesive biofilms with continuously varying strengths over 1500-fold. Calcofluor epifluorescent microscopy identified cellulose as the matrix component in biofilms produced by Pseudomonas asplenii, Pseudomonas corrugata, Pseudomonas fluorescens, Pseudomonas marginalis, Pseudomonas putida, Pseudomonas savastanoi and Pseudomonas syringae isolates. Cellulose expression and biofilm formation could be induced by the constitutively active WspR19 mutant of the cyclic-di-GMP-associated, GGDEF domain-containing response regulator involved in the P. fluorescens SBW25 wrinkly spreader phenotype and cellular aggregation in Pseudomonas aeruginosa PA01. WspR19 could also induce P. putida KT2440, which otherwise did not produce a biofilm or express cellulose, as well as Escherichia coli K12 and Salmonella typhimurium LT2, both of which express cellulose yet lack WspR homologues. Statistical analysis of biofilm parameters suggest that biofilm development is a more complex process than that simply described by the production of attachment and matrix components and bacterial growth. This complexity was also seen in multivariate analysis as a species-ecological habitat effect, underscoring the fact that in vitro biofilms are abstractions of those surface and volume colonization processes used by bacteria in their natural environments. [source] Pseudomonas putida: a cosmopolitan opportunist par excellenceENVIRONMENTAL MICROBIOLOGY, Issue 12 2002Kenneth N. Timmis No abstract is available for this article. [source] Complete sequence of the IncP-9 TOL plasmid pWW0 from Pseudomonas putidaENVIRONMENTAL MICROBIOLOGY, Issue 12 2002Alicia Greated Summary The TOL plasmid pWW0 (117 kb) is the best studied catabolic plasmid and the archetype of the IncP-9 plasmid incompatibility group from Pseudomonas. It carries the degradative (xyl) genes for toluenes and xylenes within catabolic transposons Tn4651 and Tn4653. Analysis of the complete pWW0 nucleotide sequence revealed 148 putative open reading frames. Of these, 77 showed similarity to published sequences in the available databases predicting functions for: plasmid replication, stable maintenance and transfer; phenotypic determinants; gene regulation and expression; and transposition. All identifiable transposition functions lay within the boundaries of the 70 kb transposon Tn4653, leaving a 46 kb sector containing all the IncP-9 core functions. The replicon and stable inheritance region was very similar to the mini-replicon from IncP-9 antibiotic resistance plasmid pM3, with their Rep proteins forming a novel group of initiation proteins. pWW0 transfer functions exist as two blocks encoding putative DNA processing and mating pair formation genes, with organizational and sequence similarity to IncW plasmids. In addition to the known Tn4651 and IS1246 elements, two additional transposable elements were identified as well as several putative transposition functions, which are probably genetic remnants from previous transposition events. Genes likely to be responsible for known resistance to ultraviolet light and free radicals were identified. Other putative phenotypic functions identified included resistance to mercury and other metal ions, as well as to quaternary ammonium compounds. The complexity and size of pWW0 is largely the result of the mosaic organization of the transposable elements that it carries, rather than the backbone functions of IncP-9 plasmids. [source] Genetically engineered Pseudomonas: a factory of new bioplastics with broad applicationsENVIRONMENTAL MICROBIOLOGY, Issue 10 2001Elías R. Olivera Summary New bioplastics containing aromatic or mixtures of aliphatic and aromatic monomers have been obtained using genetically engineered strains of Pseudomonas putida. The mutation (,) or deletion (,) of some of the genes involved in the ,-oxidation pathway (fadA,, fadB,,fadA or ,fad,BA mutants) elicits a strong intracellular accumulation of unusual homo- or co-polymers that dramatically alter the morphology of these bacteria, as more than 90% of the cytoplasm is occupied by these macromolecules. The introduction of a blockade in the ,-oxidation pathway, or in other related catabolic routes, has allowed the synthesis of polymers other than those accumulated in the wild type (with regard to both monomer size and relative percentage), the accumulation of certain intermediates that are rapidly catabolized in the wild type and the accumulation in the culture broths of end catabolites that, as in the case of phenylacetic acid, phenylbutyric acid, trans -cinnamic acid or their derivatives, have important medical or pharmaceutical applications (antitumoral, analgesic, radiopotentiators, chemopreventive or antihelmintic). Furthermore, using one of these polyesters (poly 3-hydroxy-6-phenylhexanoate), we obtained polymeric microspheres that could be used as drug vehicles. [source] Comparison of biodegradation kinetic parameters for naphthalene in batch and sand column systems by pseudomonas putidaENVIRONMENTAL PROGRESS & SUSTAINABLE ENERGY, Issue 2 2001Jeong-Hun Park Kinetic parameters for the degradation of naphthalene by Pseudomonas putida ( ATCC 17484) were estimated in both batch and column assays, in order to evaluate the role of flow and cell attachment on biodegradation rates. Suspended cells and cells attached to Ottawa sand were used under a variety of biomass levels, column flow-rates, and substrate concentrations. In batch systems, degradation followed zero order kinetics across the entire concentration range, while the columns exhibited decreased rates at concentrations less than 100 (,g/L), describable by Michaelis-Menten kinetics. This is reflected in elevated values of the half-saturation constant, Ks, in columns. We offer the explanation that this may have resulted from reactive heterogeneity within the porous media, imposing a distribution of length-scales for transfer of substrate to the cell surfaces. Well-mixed batch systems are expected to have both shorter and more uniform transfer distances. When kinetic parameters obtained in batch system are used for prediction of degradation in columns, at least two factors,exposed reduction of exposed cell surface are a and heterogeneity of cell distribution,will likely reduce overall column degradation rates. [source] Toxicity assessment of mono-substituted benzenes and phenols using a Pseudomonas initial oxygen uptake assayENVIRONMENTAL TOXICOLOGY & CHEMISTRY, Issue 2 2005Ded-Shih Huang Abstract A methodology is presented for assessing the toxicity of chemical substances through their inhibitory action toward the Pseudomonas initial oxygen uptake (PIOU) rate. The current studies reveal that the PIOU assay is rapid, cost-efficient, and easy to perform. The oxygen uptake rate was found to be associated with a putative benzoate transporter and highly dependent on benzoate concentration. The putative benzoate transporter has been shown to follow Michaelis,Menten kinetics. Most phenols were found to be noncompetitive inhibitors of the benzoate transporter. The inhibition constant (Ki) of these noncompetitive inhibitors can be related to the concentration causing 50% oxygen uptake inhibition in Pseudomonas putida. Modeling these data by using the response,surface approach leads to the development of a quantitative structure,activity relationship (QSAR) for the toxicity of phenols ((1/Ki) = ,0.435 (±0.038) lowest-unoccupied-molecular orbital + 0.517 (±0.027)log KOW ,2.340 (±0.068), n = 49, r2 = 0.930, s = 0.107, r2adj = 0.926, F = 303.1). A comparison of QSAR models derived from the Ki data of the PIOU method and the toxicity data of 40-h Tetrahymena pyrifomis growth inhibition assay (Tetratox) indicated that there was a high correlation between the two approaches (r2 = 0.925). [source] Effect of nonaqueous-phase liquids on the availability of polycyclic aromatic hydrocarbons in soil for worm uptake and bacterial genotoxicityENVIRONMENTAL TOXICOLOGY & CHEMISTRY, Issue 11 2003Antonio Quiñones-Rivera Abstract A study was conducted to determine the effect of nonaqueous-phase liquids (NAPLs) on the bioavailability of benzo[a]pyrene (BaP) in soil. Sentry 19 oil and pristane reduced the availability of BaP for assimilation by the earthworm Eisenia fetida and for mutagenicity in a rifampicin-sensitive strain of Pseudomonas putida. As much as 80% of the compound could be rendered unavailable to the worms or for genotoxicity. Tests with five alkanes and an oil showed that the extent of reduction in genotoxicity of BaP varied with the identity, viscosity, and hydrophobicity of the NAPL. The magnitude of the decline in availability for genotoxicity differed in tests of three soils. Because little or no BaP was lost from the soil, the diminished bioavailability was not the result of a diminished total concentration of the compound. These findings show that exposure to hydrophobic toxicants can be appreciably altered in soils containing NAPLs. [source] Oxidation of polychlorinated benzenes by genetically engineered CYP101 (cytochrome P450cam)FEBS JOURNAL, Issue 5 2001Jonathan P. Jones Polychlorinated benzenes are recalcitrant environmental pollutants primarily because they are resistant to attack by dioxygenases commonly used by micro-organisms for the biodegradation of aromatic compounds. We have investigated the oxidation of polychlorinated benzenes by mutants of the haem mono-oxygenase CYP101 (cytochrome P450cam) from Pseudomonas putida with the aim of generating novel systems for their biodegradation. Wild-type CYP101 had low activity for the oxidation of dichlorobenzenes and trichlorobenzenes to the chlorophenols, but no products were detected for the heavily chlorinated benzenes. Increasing the active-site hydrophobicity with the Y96F mutation increased the activity up to 100-fold, and both pentachlorobenzene and hexachlorobenzene were oxidized slowly to pentachlorophenol. Decreasing the space available at the top of the active site with the F87W mutation to force the substrate to be bound closer to the haem resulted in a further 10-fold increase in activity with most substrates. Introducing the F98W mutation, also at the top of the active site, decreased the NADH-turnover rates but increased the coupling efficiencies, and >,90% coupling was observed for 1,3-dichlorobenzene and 1,3,5-trichlorobenzene with the F87W,Y96F,F98W mutant. The V247L mutation generally increased the NADH-turnover rates, and the F87W,Y96F,V247L mutant showed reasonably fast NADH turnover (229 min,1) with the highly insoluble pentachlorobenzene without the need for surfactants or organic cosolvents. As all chlorophenols are degraded by micro-organisms, novel biodegradation systems could be constructed in which CYP101 mutants convert the inert polychlorinated benzenes to the phenols, which are then readily degraded by natural pathways. [source] Riding the sulfur cycle , metabolism of sulfonates and sulfate esters in Gram-negative bacteriaFEMS MICROBIOLOGY REVIEWS, Issue 2 2000Michael A. Kertesz Abstract Sulfonates and sulfate esters are widespread in nature, and make up over 95% of the sulfur content of most aerobic soils. Many microorganisms can use sulfonates and sulfate esters as a source of sulfur for growth, even when they are unable to metabolize the carbon skeleton of the compounds. In these organisms, expression of sulfatases and sulfonatases is repressed in the presence of sulfate, in a process mediated by the LysR-type regulator protein CysB, and the corresponding genes therefore constitute an extension of the cys regulon. Additional regulator proteins required for sulfonate desulfonation have been identified in Escherichia coli (the Cbl protein) and Pseudomonas putida (the AsfR protein). Desulfonation of aromatic and aliphatic sulfonates as sulfur sources by aerobic bacteria is oxygen-dependent, carried out by the ,-ketoglutarate-dependent taurine dioxygenase, or by one of several FMNH2 -dependent monooxygenases. Desulfurization of condensed thiophenes is also FMNH2 -dependent, both in the rhodococci and in two Gram-negative species. Bacterial utilization of aromatic sulfate esters is catalyzed by arylsulfatases, most of which are related to human lysosomal sulfatases and contain an active-site formylglycine group that is generated post-translationally. Sulfate-regulated alkylsulfatases, by contrast, are less well characterized. Our increasing knowledge of the sulfur-regulated metabolism of organosulfur compounds suggests applications in practical fields such as biodesulfurization, bioremediation, and optimization of crop sulfur nutrition. [source] Mutualistic symbiosis between Bursaphelenchus xylophilus and bacteria of the genus PseudomonasFOREST PATHOLOGY, Issue 5 2005B. G. Zhao Summary Interactions between the pine wood nematode (PWN), Bursaphelenchus xylophilus, and bacteria of the genus Pseudomonas were examined by cultivating axenic PWN and bacterial strains using callus of Pinus thunbergii. Ten (Pseudomonas fluorescens, Pseudomonas putida, Pseudomonas cepacia and Pseudomonas spp.) of the 29 bacterial strains tested, significantly increased the reproduction of PWN. The rest of the bacteria (19 strains of 10 species) inhibited the reproduction of PWN completely. The growth of 18 of the 29 bacterial strains tested, including the 10 strains promoting PWN reproduction, was significantly increased by the presence of PWN. It indicated a mutualistic symbiotic relationship between PWN and the 10 bacterial strains in the genus Pseudomonas. The bacterial mutualistic symbionts are organisms, which may have co-evolved with PWN rather than being accidentally associated. The finding provides further evidence for our hypothesis that pine wilt disease is complex, induced by both PWN and associated phytotoxin-producing bacteria. Résumé Les interactions entre le nématode des pins Bursaphelenchus xylophilus (PWN) et des bactéries du genre Pseudomonas ont étéétudiées en cultivant de manière axénique PWN et des souches bactériennes sur des cals de Pinusthunbergii. Dix souches bactériennes (Pseudomonas fluorescens, P. putida, P. cepacia et Pseudomonas spp.) sur les 29 testées ont significativement augmenté la reproduction de PNW. Le reste des bactéries (19 souches de 10 espèces) ont complètement inhibé la reproduction de PNW. La croissance de 18 souches bactériennes sur 29, incluant les 10 favorisant la reproduction de PNW, a été significativement augmentée en présence de PNW. Ceci indique une relation symbiotique mutualiste entre PNW et 10 souches bactériennes du genre Pseudomonas. Les symbiontes bactériens mutualistes pourraient être des organismes ayant coévolué avec PNW plutôt qu'associés de façon fortuite. Ces observations renforcent l'hypothèse selon laquelle le flétrissement des pins est une maladie complexe induite par PWN en association avec des bactéries productrices de phytotoxines. Zusammenfassung Die Interaktionen zwischen dem Kiefernsplintholznematoden (PWN, Bursaphelenchus xylophilus) und Bakterien der Gattung Pseudomonas wurden in axenischen Kulturen des Nematoden mit verschiedenen Bakterienstämmen und Kallus von Pinus thunbergii untersucht. Zehn Bakterienstämme (Pseudomonas fluoreszens, Pseudomonas putida, Pseudomonas cepacia und Pseudomonas spp.) von 29 getesteten Isolaten erhöhten die Reproduktion des PWN signifikant. Die übrigen Isolate (19 Stämme von 10 Arten) hemmten die Vermehrung des Nematoden vollständig. Das Wachstum von 18 der 29 getesteten Bakterienstämme (inkl. der 10 Stämme, welche die Vermehrung des Nematoden förderten), wurde durch die Präsenz des Nematoden signifikant erhöht. Dieser Befund deutet auf eine mutualistische Beziehung zwischen dem PWN und 10 Pseudomonas -Isolaten hin. Die mutualistischen bakteriellen Symbionten dürften sich wahrscheinlich in Coevolution mit dem Nematoden entwickelt haben. Dieser Befund unterstützt die Hypothese, dass die Kiefernwelke eine Komplexkrankheit darstellt, die sowohl durch B. xylophilus als auch die damit assoziierten phytotoxinbildenden Bakterien ausgelöst wird. [source] Effects of some bacteria (Pseudomonas spp. and Erwinia herbicola) on in vitro growth of Piptoporus betulinusFOREST PATHOLOGY, Issue 6 2000K. Przyby Summary Bacteria including Pseudomonas putida, Pseudomonas fluorescens biovar I, Pseudomonas fluorescens biovar V, Pseudomonas aureofaciens and Erwinia herbicola were isolated from discoloured zones in birch trunks. Antagonistic effects of these bacteria to growth of Piptoporus betulinus mycelium were tested in vitro, both in dual culture and using bacterial cell-free culture filtrates. In dual cultures, P. putida was most effective at inhibiting mycelial growth of Piptoporus betulinus. Filtrates of P. putida inhibited growth of P. betulinus mycelium irrespective of filtrate concentration, incubation time of bacteria and timing of recording mycelium growth. The strongest antagonistic effect (inhibition of fungal growth) was observed on a medium containing 80% of sterile filtrate obtained from 15-day-old bacterial cultures. The highest stimulating effect on mycelium growth was noted on medium containing 80% filtrate obtained from 7-day-old E. herbicola cultures. Résumé Des bactéries, Pseudomonas putida, Pseudomonas fluorescens biovar I, Pseudomonas fluorescens biovar V, Pseudomonas aureofaciens et Erwinia herbicola, ont été isolées de zones colorées de troncs de bouleau. Les effets antagonistes de ces bactéries sur la croissance mycélienne de Piptoporus betulinus ont étéévalués in vitro, en cultures doubles et à partir de filtrats bactériens. En cultures doubles, P. putida a été le plus inhibiteur de la croissance du P. betulinus. Les filtrats de P. putida inhibaient la croissance quel que soit la concentration du filtrat, la durée d'incubation de la bactérie, et le délai dans lequel la croissance mycélienne était mesurée. L'effet inhibiteur le plus fort a été observé sur un milieu contenant 80% de filtrat stérile obtenu de cultures bactériennes de 15 jours. L'effet stimulant le plus fort a été noté sur un milieu contenant 80% d'un filtrat obtenu de cultures de 7 jours de E. herbicola. Zusammenfassung Verschiedene Bakterienarten (Pseudomonas putida, Pseudomonas fluorescens Biovar I, Pseudomonas fluorescens Biovar V, Pseudomonas aureofaciens und E. herbicola) wurden aus verfärbtem Holz in Birkenstämmen isoliert. Antagonistische Effekte dieser Bakterien gegenüber Myzel von Piptoporus betulinus wurden in vitroüberprüft (Dualkulturen und bakterienzellfreie Kulturfiltrate). In Dualkulturen zeigte P. putida den stärksten Hemmeffekt auf das Myzelwachstum von P. betulinus. Filtrate von P. putida hemmten das Wachstum von P. betulinus, unabhängig von der Filtratkonzentration, der Inkubationszeit der Bakterien und dem Zeitpunkt der Messung des Myzelwachstums. Der antagonistische Effekt (Hemmung des Myzelwachstums) war am ausgeprägtesten auf einem Medium, das 80% Sterilfiltrat von 15 Tage alten Bakterienkulturen enthielt. Der stärkste Stimulationseffekt auf das Myzelwachstum wurde auf einem Medium beobachtet, welches 80% Filtrat von sieben Tage alten E. herbicola -Kulturen enthielt. [source] Impact of different pasteurization temperatures on the survival of microbial contaminants isolated from pasteurized milkINTERNATIONAL JOURNAL OF DAIRY TECHNOLOGY, Issue 2 2005PHOLISA DUMALISILE The thermal inactivation of selected microbes was studied using the low temperature long time (LTLT), high temperature short time (HTST) and ,pot' pasteurization methods. Survivors were enumerated after heating for up to 40 min for the LTLT and HTST pasteurization methods and after heating for up to 30 min for the ,pot' pasteurization method. With the exception of the Bacillus cereus strain, the selected microbes did not survive the LTLT and HTST pasteurization methods. The results from the ,pot' pasteurizer showed that B. cereus, Chryseobacterium meningosepticum, Pseudomonas putida, Acinetobacter baumannii and Escherichia coli strains survived the pasteurization conditions applied, showing that the ,pot' pasteurizer does not pasteurize effectively. [source] Development of a homologous expression system for rubber oxygenase RoxA from Xanthomonas sp.JOURNAL OF APPLIED MICROBIOLOGY, Issue 3 2010N. Hambsch Abstract Aims:, Natural rubber (poly-[cis -1,4-isoprene]) can be cleaved into 12-oxo-4,8-dimethyltrideca-4,8-diene-1-al by rubber oxygenase A (RoxA) isolated from Xanthomonas sp. RoxA is a novel type of dihaem dioxygenase with unknown cleavage mechanism of the rubber carbon backbone. Analysis of mutant RoxA after mutagenesis could be a way to investigate the function of selected amino acids of RoxA during catalysis. Unfortunately, expression of functional RoxA in recombinant Escherichia coli or in recombinant ,-Proteobacteria such as Pseudomonas putida was not possible in our hands. Therefore, expression of recombinant RoxA in the homologous host, Xanthomonas, was performed. Methods and Results:, A transformation system via electroporation was established, and a conjugation system was optimized for Xanthomonas sp. Inactivation of the chromosomal roxA gene by insertional mutagenesis resulted in inability of Xanthomonas sp. to produce active RoxA and to utilize rubber as a sole source of carbon and energy. When an intact copy of roxA was cloned under control of a rhamnose-inducible promoter in a broad host range vector and was transferred to Xanthomonas sp., high expression levels of functional RoxA in the presence of rhamnose were obtained. Conclusions and Significance and Impact of the Study:, Purification of recombinantly expressed RoxA was simplified because of drastically shortened fermentation times and because separation of RoxA from remaining rubber latex particles was not necessary with rhamnose-induced cultures. About 6 mg purified RoxA were obtained from 1 l of cell-free culture fluid. Purified recombinant RoxA was highly active and revealed comparable spectral properties as RoxA purified from the wild type. The results of our study are the methodical basis for molecular biological manipulation in Xanthomonas sp. and will simplify investigation into the biochemical mechanisms by which rubber can be biodegraded in the environment by this novel extracellular dihaem dioxygenase RoxA. [source] Calorimetric investigations into the starvation response of Pseudomonas putida growing on phenol and glucoseJOURNAL OF APPLIED MICROBIOLOGY, Issue 6 2009Andreas Lißner Abstract Aims:, To investigate the stress response during nutrient deprivation, particularly with regard to the application of phenol as growth substrate of Pseudomonas putida with calorimetric measurements as a new method. Methods and Results:, The online and noninvasive measurement of the thermal power P0 permits the detection of microbial activity during the starvation period. While the results of the investigations with phenol reveal a significant loss of activity as a function of the temporal nutrient dosage, only a small loss of activity was detected by using glucose. Microbiological methods (colony forming units (CFU) and activity of catechol-2,3-dioxygenase) showed a loss of the enzyme activity at a constant CFU. The introduction of a simple decay parameter kD in the kinetic description of the growth process on phenol was sufficient for the successful kinetic modelling. Conclusions:, The combination of calorimetric measurements and the determination of the enzymatic activity proved the loss of activity of Ps. putida during the deprivation of the substrate phenol. Significance and Impact of the study:, The initial heat power (P0) proves to be a suitable parameter for the characterization of the physiological state of the culture and can be used for the regulation of nutrient supply in biotechnological process development. [source] Functional, genetic and chemical characterization of biosurfactants produced by plant growth-promoting Pseudomonas putida 267JOURNAL OF APPLIED MICROBIOLOGY, Issue 2 2009Marco Kruijt Abstract Aims:, Plant growth-promoting Pseudomonas putida strain 267, originally isolated from the rhizosphere of black pepper, produces biosurfactants that cause lysis of zoospores of the oomycete pathogen Phytophthora capsici. The biosurfactants were characterized, the biosynthesis gene(s) partially identified, and their role in control of Phytophthora damping-off of cucumber evaluated. Methods and Results:, The biosurfactants were shown to lyse zoospores of Phy. capsici and inhibit growth of the fungal pathogens Botrytis cinerea and Rhizoctonia solani. In vitro assays further showed that the biosurfactants of strain 267 are essential in swarming motility and biofilm formation. In spite of the zoosporicidal activity, the biosurfactants did not play a significant role in control of Phytophthora damping-off of cucumber, since both wild type strain 267 and its biosurfactant-deficient mutant were equally effective, and addition of the biosurfactants did not provide control. Genetic characterization revealed that surfactant biosynthesis in strain 267 is governed by homologues of PsoA and PsoB, two nonribosomal peptide synthetases involved in production of the cyclic lipopeptides (CLPs) putisolvin I and II. The structural relatedness of the biosurfactants of strain 267 to putisolvins I and II was supported by LC-MS and MS-MS analyses. Conclusions:, The biosurfactants produced by Ps. putida 267 were identified as putisolvin-like CLPs; they are essential in swarming motility and biofilm formation, and have zoosporicidal and antifungal activities. Strain 267 provides excellent biocontrol activity against Phytophthora damping-off of cucumber, but the lipopeptide surfactants are not involved in disease suppression. Significance and Impact of the Study:,Pseudomonas putida 267 suppresses Phy. capsici damping-off of cucumber and provides a potential supplementary strategy to control this economically important oomycete pathogen. The putisolvin-like biosurfactants exhibit zoosporicidal and antifungal activities, yet they do not contribute to biocontrol of Phy. capsici and colonization of cucumber roots by Ps. putida 267. These results suggest that Ps. putida 267 employs other, yet uncharacterized, mechanisms to suppress Phy. capsici. [source] Degradation of naphthenic acids by sediment micro-organismsJOURNAL OF APPLIED MICROBIOLOGY, Issue 5 2006L.F. Del Rio Abstract Aims:, Naphthenic acids (NAs) are naturally occurring, linear and cyclic carboxylic surfactants associated with the acidic fraction of petroleum. NAs account for most of the acute aquatic toxicity of oil sands process-affected water (OSPW). The toxicity of OSPW can be reduced by microbial degradation. The aim of this research was to determine the extent of NA degradation by sediment microbial communities exposed to varying amounts of OSPW. Methods and Results:, Eleven wetlands, both natural and process-affected, and one tailings settling pond in Northern Alberta were studied. The natural wetlands and process-affected sites fell into two distinct groups based on their water chemistry. The extent of degradation of a 14C-labelled monocyclic NA surrogate [14C-cyclohexane carboxylic acid (CCA)] was relatively uniform in all sediments (approximately 30%) after 14 days. In contrast, degradation of a bicyclic NA surrogate [14C-decahydronaphthoic acid (DHNA)]was significantly lower in non process-affected sediments. Enrichment cultures, obtained from an active tailings settling pond, using commercially available NAs as the sole carbon source, resulted in the isolation of a co-culture containing Pseudomonas putida and Pseudomonas fluorescens. Quantitative GC,MS analysis showed that the co-culture removed >95% of the commercial NAs, and partially degraded the process NAs from OSPW with a resulting NA profile similar to that from ,aged wetlands'. Conclusions:, Exposure to NAs induced and/or selected micro-organisms capable of more effectively degrading bicyclic NAs. Native Pseudomonas spp. extensively degraded fresh, commercial NA. The recalcitrant NAs resembled those found in process-affected wetlands. Significance and Impact of the Study:, These results suggest that it may be possible to manipulate the existing environmental conditions to select for a microbial community exhibiting higher rates of NA degradation. This will have significant impact on the design of artificial wetlands for water treatment. [source] Succession of microbial communities during a biostimulation process as evaluated by DGGE and clone library analysesJOURNAL OF APPLIED MICROBIOLOGY, Issue 4 2001A. Ogino Aims:,The objective of this study was to investigate the changes in the indigenous bacterial community structure for assessing the impact of biostimulation on spilled oil. Methods and Results:,Changes in the bacterial community structure were monitored by denaturing gradient gel electrophoresis (DGGE) and clone library methods based on 16S rRNA gene (rDNA) sequences. The results of DGGE, coupled with the use of the Shannon index and principal component analysis (PCA) and clone library analyses, were consistent. In the treated (fertilized) area, one operational taxonomic unit (OTU) became dominant during the fertilization period, and it was most closely related to Pseudomonas putida. Conclusions:,The bacterial community structure in the treated area was markedly different from that in the control (non-fertilized) area during the fertilization period, but in the two areas it became similar at 14 weeks after the end of fertilization. Significance and Impact of the Study:,The results suggest that the bacterial community structure was disrupted by the biostimulation treatment, but that it recovered immediately after the end of fertilization. [source] Evolution of a degradative bacterial consortium during the enrichment of naphtha solventJOURNAL OF APPLIED MICROBIOLOGY, Issue 6 2000L. Cavalca A microbial mixed culture able to degrade naphtha solvent, a model of hydrocarbon aromatic mixture, was isolated from a hydrocarbon-polluted soil. Composition of the population was monitored by phenotypic and molecular methods applied on soil DNA, on whole enrichment culture DNA, and on 85 isolated strains. Strains were characterized for their 16S rDNA restriction profiles and for their random amplified polymorphic DNA profiles. Catabolic capabilities were monitored by phenotypic traits and by PCR assays for the presence of the catabolic genes methyl mono-oxygenase ( xylA,M), catechol 2,3 dioxygenase (xylE) and toluene dioxygenase (todC1) of TOL and TOD pathways. Different haplotypes belonging to Pseudomonas putida, Ps. aureofaciens and Ps. aeruginosa were found to degrade aromatic compounds and naphtha solvent. The intrinsic catabolic activity of the microbial population of the polluted site was detected by PCR amplification of the xylE gene directly from soil DNA. [source] Biodesulfurization of dibenzothiophene using recombinant Pseudomonas strainJOURNAL OF CHEMICAL TECHNOLOGY & BIOTECHNOLOGY, Issue 3 2008Lavanya Meesala Abstract BACKGROUND: The sulfur content in crude oil available from various sources ranges from 0.03 to values as high as 8.0 wt%. These high quantities of sulfur must be removed before the crude oil is processed because combustion of this oil would result in severe environmental pollution, such as acid rain. Due to high utility and operating costs, the conventional hydrodesulfurization process (HDS) is considered to be uneconomic. The biotechnological option, biodesulfurization (BDS) seems an attractive low cost, environmentally benign technology. RESULTS: This paper reports the development of a recombinant strain of bacteria designed by introducing desulfurizing, dsz genes containing plasmid pSAD 225-32, which was isolated from Rhodococcus erythropolis IGTS8 into a gram negative solvent-tolerant bacterium, Pseudomonas putida (MTCC 1194). This recombinant bacterium can desulfurize the dibenzothiophene (DBT) in the sulfur selective 4S-pathway. It has been observed that for the same concentration of DBT, the recombinant strain's growth rate is greater than that of the parent strain. Increasing the concentration of DBT resulted in an increase of lag phase as well as decreased growth rate, which shows that the bacteria is following substrate inhibition type kinetics. This genetically modified bacterium can desulfurize 73.1% of 1.2 mmol L,1 DBT (dissolved in ethanol) in 67 h of cultivation time using growing cells. CONCLUSIONS: It is concluded that further research in this area of biodesulfurization using genetically modified organisms may remove the bottlenecks presently in the way of commercialization of the BDS process. Copyright © 2007 Society of Chemical Industry [source] Continuous bioremediation of phenol-polluted air in an external loop airlift bioreactor with a packed bed,JOURNAL OF CHEMICAL TECHNOLOGY & BIOTECHNOLOGY, Issue 6 2006Hossein Nikakhtari Abstract An external loop airlift bioreactor with a small amount (99% porosity) of stainless steel mesh packing inserted in the riser section was used for bioremediation of a phenol-polluted air stream. The packing enhanced volatile organic chemical and oxygen mass transfer rates and provided a large surface area for cell immobilization. Using a pure strain of Pseudomonas putida, fed-batch and continuous runs at three different dilution rates were completed with phenol in the polluted air as the only source of growth substrate. 100% phenol removal was achieved at phenol loading rates up to 33 120 mg h,1 m,3 using only one-third of the column, superior to any previously reported biodegradation rates of phenol-polluted air with 100% efficiency. A mathematical model has been developed and is shown to accurately predict the transient and steady-state data. Copyright © 2006 Society of Chemical Industry [source] MICROBIOLOGICAL AND PATHOGENIC CONTAMINANTS OF SEAFOOD IN GREECEJOURNAL OF FOOD QUALITY, Issue 1 2007C. PAPADOPOULOU ABSTRACT A total of 360 samples, including 105 marine fish, 25 prawns, 50 squid, 50 octopus, 30 mussels and 100 freshwater fish were examined microbiologically for the presence of microorganisms potentially pathogenic. All samples were examined following standard microbiological procedures. The isolated microorganisms were: Aeromonas hydrophilia (38,93%), Klebsiella ozonae (1,40%), Escherichia coli (14,87%), Yersinia enterocolitica (0,40%), Hafnia alvei (0,36.6%), Enterobacter agglomerans (0,42%), Citrobacter freundii (0,46%), Proteus vulgaris (15,80%), Proteus mirabilis (7,82%), Morganella morganii (0,30%), Pseudomonas fluorescens (0,34%), Pseudomonas putida (0,6%), Plesiomonas shigelloides (0,4%), Listeria innocua (1,3.3%), Vibrio parahaemolyticus (0,2%), Clostridium perfringens (0,1%), Staphylococcus aureus (0,80%) and Candida quillermondi (0,1%), Candida albicans (0,1%), Penicillium oxalicum (0,1%) and Penicillium italicum (0,12%). [source] | |||||||||||||||||||||||||