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Sample records for putida kt2440 metabolic

  1. Root inoculation with Pseudomonas putida KT2440 induces transcriptional and metabolic changes and systemic resistance in maize plants.

    Science.gov (United States)

    Planchamp, Chantal; Glauser, Gaetan; Mauch-Mani, Brigitte

    2014-01-01

    Pseudomonas putida KT2440 (KT2440) rhizobacteria colonize a wide range of plants. They have been extensively studied for their capacity to adhere to maize seeds, to tolerate toxic secondary metabolites produced by maize roots and to be attracted by maize roots. However, the response of maize plants to KT2440 colonization has not been investigated yet. Maize roots were inoculated with KT2440 and the local (roots) and systemic (leaves) early plant responses were investigated. The colonization behavior of KT2440 following application to maize seedlings was investigated and transcriptional analysis of stress- and defense-related genes as well as metabolite profiling of local and systemic maize tissues of KT2440-inoculated were performed. The local and systemic responses differed and more pronounced changes were observed in roots compared to leaves. Early in the interaction roots responded via jasmonic acid- and abscisic acid-dependent signaling. Interestingly, during later steps, the salicylic acid pathway was suppressed. Metabolite profiling revealed the importance of plant phospholipids in KT2440-maize interactions. An additional important maize secondary metabolite, a form of benzoxazinone, was also found to be differently abundant in roots 3 days after KT2440 inoculation. However, the transcriptional and metabolic changes observed in bacterized plants early during the interaction were minor and became even less pronounced with time, indicating an accommodation state of the plant to the presence of KT2440. Since the maize plants reacted to the presence of KT2440 in the rhizosphere, we also investigated the ability of these bacteria to trigger induced systemic resistance (ISR) against the maize anthracnose fungus Colletotrichum graminicola. The observed resistance was expressed as strongly reduced leaf necrosis and fungal growth in infected bacterized plants compared to non-bacterized controls, showing the potential of KT2440 to act as resistance inducers.

  2. Root inoculation with Pseudomonas putida KT2440 induces transcriptional and metabolic changes and systemic resistance in maize plants

    Directory of Open Access Journals (Sweden)

    Chantal ePlanchamp

    2015-01-01

    Full Text Available Pseudomonas putida KT2440 (KT2440 rhizobacteria colonize a wide range of plants. They have been extensively studied for their capacity to adhere to maize seeds, to tolerate toxic secondary metabolites produced by maize roots and to be attracted by maize roots. However, the response of maize plants to KT2440 colonization has not been investigated yet. Maize roots were inoculated with KT2440 and the local (roots and systemic (leaves early plant responses were investigated. The colonization behavior of KT2440 following application to maize seedlings was investigated and transcriptional analysis of stress- and defense-related genes as well as metabolite profiling of local and systemic maize tissues of KT2440-inoculated were performed. The local and systemic responses differed and more pronounced changes were observed in roots compared to leaves. Early in the interaction roots responded via jasmonic acid- and abscisic acid-dependent signaling. Interestingly, during later steps, the salicylic acid pathway was suppressed. Metabolite profiling revealed the importance of plant phospholipids in KT2440-maize interactions. An additional important maize secondary metabolite, a form of benzoxazinone, was also found to be differently abundant in roots three days after KT2440 inoculation. However, the transcriptional and metabolic changes observed in bacterized plants early during the interaction were minor and became even less pronounced with time, indicating an accommodation state of the plant to the presence of KT2440. Since the maize plants reacted to the presence of KT2440 in the rhizosphere, we also investigated the ability of these bacteria to trigger induced systemic resistance (ISR against the maize anthracnose fungus Colletotrichum graminicola. The observed resistance was expressed as strongly reduced leaf necrosis and fungal development in infected bacterized plants compared to non-bacterized controls, showing the potential of KT2440 to act as

  3. Root inoculation with Pseudomonas putida KT2440 induces transcriptional and metabolic changes and systemic resistance in maize plants

    OpenAIRE

    Chantal ePlanchamp; Gaetan eGlauser; Brigitte eMauch-Mani

    2015-01-01

    Pseudomonas putida KT2440 (KT2440) rhizobacteria colonize a wide range of plants. They have been extensively studied for their capacity to adhere to maize seeds, to tolerate toxic secondary metabolites produced by maize roots and to be attracted by maize roots. However, the response of maize plants to KT2440 colonization has not been investigated yet. Maize roots were inoculated with KT2440 and the local (roots) and systemic (leaves) early plant responses were investigated. The colonization b...

  4. Root inoculation with Pseudomonas putida KT2440 induces transcriptional and metabolic changes and systemic resistance in maize plants

    OpenAIRE

    Chantal ePlanchamp; Gaetan eGlauser; Brigitte eMauch-Mani

    2015-01-01

    Pseudomonas putida KT2440 (KT2440) rhizobacteria colonize a wide range of plants. They have been extensively studied for their capacity to adhere to maize seeds, to tolerate toxic secondary metabolites produced by maize roots and to be attracted by maize roots. However, the response of maize plants to KT2440 colonization has not been investigated yet. Maize roots were inoculated with KT2440 and the local (roots) and systemic (leaves) early plant responses were investigated. The colonization b...

  5. Metabolic engineering of Pseudomonas putida KT2440 for the production of para-hydroxy benzoic acid

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    Shiqin Yu

    2016-11-01

    Full Text Available para-hydroxy benzoic acid (PHBA is the key component for preparing parabens, a common preservatives in food, drugs and personal care products, as well as high performance bioplastics such as liquid crystal polymers (LCP. Pseudomonas putida KT2440 was engineered to produce PHBA from glucose via the shikimate pathway intermediate chorismate. To obtain the PHBA production strain, chorismate lyase UbiC from Escherichia coli and a feedback resistant 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase encoded by gene aroGD146N were overexpressed individually and simultaneously. In addition, genes related to product degradation (pobA or competing for the precursor chorismate (pheA and trpE were deleted from the genome. To further improve PHBA production, the glucose metabolism repressor hexR was knocked out in order to increase erythrose-4- phosphate and NAPH supply. The best strain achieved a maximum titre of 1.73 g L-1 and a carbon yield of 18.1 % (C-mol C-mol-1 in a non-optimized fed-batch fermentation. This is to date the highest PHBA concentration produced by P. putida using a chorismate lyase.

  6. Metabolic Engineering of Pseudomonas putida KT2440 for the Production of para-Hydroxy Benzoic Acid

    Science.gov (United States)

    Yu, Shiqin; Plan, Manuel R.; Winter, Gal; Krömer, Jens O.

    2016-01-01

    para-Hydroxy benzoic acid (PHBA) is the key component for preparing parabens, a common preservatives in food, drugs, and personal care products, as well as high-performance bioplastics such as liquid crystal polymers. Pseudomonas putida KT2440 was engineered to produce PHBA from glucose via the shikimate pathway intermediate chorismate. To obtain the PHBA production strain, chorismate lyase UbiC from Escherichia coli and a feedback resistant 3-deoxy-d-arabino-heptulosonate-7-phosphate synthase encoded by gene aroGD146N were overexpressed individually and simultaneously. In addition, genes related to product degradation (pobA) or competing for the precursor chorismate (pheA and trpE) were deleted from the genome. To further improve PHBA production, the glucose metabolism repressor hexR was knocked out in order to increase erythrose 4-phosphate and NADPH supply. The best strain achieved a maximum titer of 1.73 g L−1 and a carbon yield of 18.1% (C-mol C-mol−1) in a non-optimized fed-batch fermentation. This is to date the highest PHBA concentration produced by P. putida using a chorismate lyase. PMID:27965953

  7. A genome-scale metabolic reconstruction of Pseudomonas putida KT2440: iJN746 as a cell factory

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    Thiele Ines

    2008-09-01

    Full Text Available Abstract Background Pseudomonas putida is the best studied pollutant degradative bacteria and is harnessed by industrial biotechnology to synthesize fine chemicals. Since the publication of P. putida KT2440's genome, some in silico analyses of its metabolic and biotechnology capacities have been published. However, global understanding of the capabilities of P. putida KT2440 requires the construction of a metabolic model that enables the integration of classical experimental data along with genomic and high-throughput data. The constraint-based reconstruction and analysis (COBRA approach has been successfully used to build and analyze in silico genome-scale metabolic reconstructions. Results We present a genome-scale reconstruction of P. putida KT2440's metabolism, iJN746, which was constructed based on genomic, biochemical, and physiological information. This manually-curated reconstruction accounts for 746 genes, 950 reactions, and 911 metabolites. iJN746 captures biotechnologically relevant pathways, including polyhydroxyalkanoate synthesis and catabolic pathways of aromatic compounds (e.g., toluene, benzoate, phenylacetate, nicotinate, not described in other metabolic reconstructions or biochemical databases. The predictive potential of iJN746 was validated using experimental data including growth performance and gene deletion studies. Furthermore, in silico growth on toluene was found to be oxygen-limited, suggesting the existence of oxygen-efficient pathways not yet annotated in P. putida's genome. Moreover, we evaluated the production efficiency of polyhydroxyalkanoates from various carbon sources and found fatty acids as the most prominent candidates, as expected. Conclusion Here we presented the first genome-scale reconstruction of P. putida, a biotechnologically interesting all-surrounder. Taken together, this work illustrates the utility of iJN746 as i a knowledge-base, ii a discovery tool, and iii an engineering platform to explore P

  8. Transcriptional tradeoff between metabolic and stress-response programs in Pseudomonas putida KT2440 cells exposed to toluene.

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    Domínguez-Cuevas, Patricia; González-Pastor, José-Eduardo; Marqués, Silvia; Ramos, Juan-Luis; de Lorenzo, Víctor

    2006-04-28

    When Pseudomonas putida KT2440 cells encounter toluene in the growth medium, they perceive it simultaneously as a potential nutrient to be metabolized, as a membrane-damaging toxic drug to be extruded, and as a macromolecule-disrupting agent from which to protect proteins. Each of these inputs requires a dedicated transcriptional response that involves a large number of genes. We used DNA array technology to decipher the interplay between these responses in P. putida KT2440 subjected to a short challenge (15 min) with toluene. We then compared the results with those in cells exposed to o-xylene (a non-biodegradable toluene counterpart) and 3-methylbenzoate (a specific substrate of the lower TOL pathway of the P. putida pWW0 plasmid). The resulting expression profiles suggest that the bulk of the available transcriptional machinery is reassigned to endure general stress, whereas only a small share of the available machinery is redirected to the degradation of the aromatic compounds. Specifically, both toluene and o-xylene induce the TOL pathways and a dedicated but not always productive metabolic program. Similarly, 3-methylbenzoate induces the expression not only of the lower meta pathway but also of the non-productive and potentially deleterious genes for the metabolism of (nonsubstituted) benzoate. In addition, toluene (and to a lesser extent o-xylene) inhibit motility functions as an unequivocal response to aromatic toxicity. We argue that toluene is sensed by P. putida KT2440 as a stressor rather than as a nutrient and that the inhibition by the aromatic compounds of many functions we tested is the tradeoff for activating stress tolerance genes at a minimal cost in terms of energy.

  9. Genome-scale reconstruction and analysis of the Pseudomonas putida KT2440 metabolic network facilitates applications in biotechnology.

    Directory of Open Access Journals (Sweden)

    Jacek Puchałka

    2008-10-01

    Full Text Available A cornerstone of biotechnology is the use of microorganisms for the efficient production of chemicals and the elimination of harmful waste. Pseudomonas putida is an archetype of such microbes due to its metabolic versatility, stress resistance, amenability to genetic modifications, and vast potential for environmental and industrial applications. To address both the elucidation of the metabolic wiring in P. putida and its uses in biocatalysis, in particular for the production of non-growth-related biochemicals, we developed and present here a genome-scale constraint-based model of the metabolism of P. putida KT2440. Network reconstruction and flux balance analysis (FBA enabled definition of the structure of the metabolic network, identification of knowledge gaps, and pin-pointing of essential metabolic functions, facilitating thereby the refinement of gene annotations. FBA and flux variability analysis were used to analyze the properties, potential, and limits of the model. These analyses allowed identification, under various conditions, of key features of metabolism such as growth yield, resource distribution, network robustness, and gene essentiality. The model was validated with data from continuous cell cultures, high-throughput phenotyping data, (13C-measurement of internal flux distributions, and specifically generated knock-out mutants. Auxotrophy was correctly predicted in 75% of the cases. These systematic analyses revealed that the metabolic network structure is the main factor determining the accuracy of predictions, whereas biomass composition has negligible influence. Finally, we drew on the model to devise metabolic engineering strategies to improve production of polyhydroxyalkanoates, a class of biotechnologically useful compounds whose synthesis is not coupled to cell survival. The solidly validated model yields valuable insights into genotype-phenotype relationships and provides a sound framework to explore this versatile

  10. The Functional Structure of Central Carbon Metabolism in Pseudomonas putida KT2440

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    Sudarsan, Suresh; Dethlefsen, Sarah; Blank, Lars M.; Siemann-Herzberg, Martin

    2014-01-01

    What defines central carbon metabolism? The classic textbook scheme of central metabolism includes the Embden-Meyerhof-Parnas (EMP) pathway of glycolysis, the pentose phosphate pathway, and the citric acid cycle. The prevalence of this definition of central metabolism is, however, equivocal without experimental validation. We address this issue using a general experimental approach that combines the monitoring of transcriptional and metabolic flux changes between steady states on alternative carbon sources. This approach is investigated by using the model bacterium Pseudomonas putida with glucose, fructose, and benzoate as carbon sources. The catabolic reactions involved in the initial uptake and metabolism of these substrates are expected to show a correlated change in gene expressions and metabolic fluxes. However, there was no correlation for the reactions linking the 12 biomass precursor molecules, indicating a regulation mechanism other than mRNA synthesis for central metabolism. This result substantiates evidence for a (re)definition of central carbon metabolism including all reactions that are bound to tight regulation and transcriptional invariance. Contrary to expectations, the canonical Entner-Doudoroff and EMP pathways sensu stricto are not a part of central carbon metabolism in P. putida, as they are not regulated differently from the aromatic degradation pathway. The regulatory analyses presented here provide leads on a qualitative basis to address the use of alternative carbon sources by deregulation and overexpression at the transcriptional level, while rate improvements in central carbon metabolism require careful adjustment of metabolite concentrations, as regulation resides to a large extent in posttranslational and/or metabolic regulation. PMID:24951791

  11. Optimization of Pseudomonas putida KT2440 as host for the production of cis, cis-muconate from benzoate

    NARCIS (Netherlands)

    Duuren, van J.B.J.H.

    2011-01-01

    Optimization of Pseudomonas putida KT2440 as host for the production of cis, cis-muconate from benzoate P. putida KT2440 was used as biocatalyst given its versatile and energetically robust metabolism. Therefore, a mutant was generated and a process developed based on which a life cycle assessment

  12. Influence of putative exopolysaccharide genes on Pseudomonas putida KT2440 biofilm stability

    DEFF Research Database (Denmark)

    Nilsson, Martin; Chiang, Wen-Chi; Fazli, Mustafa;

    2011-01-01

    . putida KT2440 biofilm. The gene clusters alg and bcs, which code for proteins mediating alginate and cellulose biosynthesis, were found to play minor roles in P. putida KT2440 biofilm formation and stability under the conditions tested. A P. putida KT2440 derivative devoid of any identifiable...

  13. An upp-based markerless gene replacement method for genome reduction and metabolic pathway engineering in Pseudomonas mendocina NK-01 and Pseudomonas putida KT2440.

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    Wang, Yuanyuan; Zhang, Chi; Gong, Ting; Zuo, Zhenqiang; Zhao, Fengjie; Fan, Xu; Yang, Chao; Song, Cunjiang

    2015-06-01

    A markerless gene replacement method was adapted by combining a suicide plasmid, pEX18Tc, with a counterselectable marker, the upp gene encoding uracil phosphoribosyltransferase (UPRTase), for the medium-chain length polyhydroxyalkanoates (PHA(MCL))-producing strain Pseudomonas mendocina NK-01. An NK-01 5-fluorouracil (5-FU) resistant background strain was first constructed by deleting the chromosomal upp gene. The suicide plasmid pEX18Tc, carrying a functional allele of the upp gene of P. mendocina NK-01, was used to construct the vectors to delete the algA (encoding mannose-1-phosphate guanylyltransferase) and phaZ (encoding PHA(MCL) depolymerase) genes, and a 30 kb chromosomal fragment in the 5-FU resistant background host. The genes were removed efficiently from the genome of P. mendocina NK-01 and left a markerless chromosomal mutant. In addition, two exogenous genes were inserted into the phaC1 (PHA(MCL) polymerase) loci of Pseudomonas putida KT-∆UPP simultaneously. Thus, we constructed a genetically stable and marker-free P. putida KT2440 mutant with integrated mpd (encoding methyl parathion hydrolase (MPH)) and pytH (encoding a pyrethroid-hydrolyzing carboxylesterase (PytH)) gene on the chromosome. The upp-based counterselection system could be further adapted for P. mendocina NK-01 and P. putida KT2440 and used for genome reduction and metabolic pathway engineering.

  14. Genomotyping of Pseudomonas putida strains using P. putida KT2440-based high-density DNA microarrays: Implications for transcriptomics studies

    NARCIS (Netherlands)

    Ballerstedt, H.; Volkers, R.J.M.; Mars, A.E.; Hallsworth, J.E.; Santos, V.A.M.D.; Puchalka, J.; Duuren, J. van; Eggink, G.; Timmis, K.N.; Bont, J.A.M. de; Wery, J.

    2007-01-01

    Pseudomonas putida KT2440 is the only fully sequenced P. putida strain. Thus, for transcriptomics and proteomics studies with other P. putida strains, the P. putida KT2440 genomic database serves as standard reference. The utility of KT2440 whole-genome, high-density oligonucleotide microarrays for

  15. Generation of a catR deficient mutant of P. putida KT2440 that produces cis, cis-muconate from benzoate at high rate and yield

    NARCIS (Netherlands)

    Duuren, J.B.J.H. van; Wijte, D.; Leprince, A.; Karge, B.; Puchalka, J.; Wery, J.; Dos Santos, V.A.P.M.; Eggink, G.; Mars, A.E.

    2011-01-01

    Pseudomonas putida KT2440-JD1 was derived from P. putida KT2440 after N-methyl-N'-nitro-N-nitrosoguanidine (NTG)-mutagenesis and exposure to 3-fluorobenzoate (3-FB). The mutant was no longer able to grow using benzoate as a sole carbon source, but co-metabolized benzoate to cis, cis-muconate during

  16. Comprehensive proteome analysis of the response of Pseudomonas putida KT2440 to the flavor compound vanillin.

    Science.gov (United States)

    Simon, Oliver; Klaiber, Iris; Huber, Armin; Pfannstiel, Jens

    2014-09-23

    Understanding of the molecular response of bacteria to precursors, products and environmental conditions applied in bioconversions is essential for optimizing whole-cell biocatalysis. To investigate the molecular response of the potential biocatalyst Pseudomonas putida KT2440 to the flavor compound vanillin we applied complementary gel- and LC-MS-based quantitative proteomics approaches. Our comprehensive proteomics survey included cytoplasmic and membrane proteins and led to the identification and quantification of 1614 proteins, corresponding to 30% of the total KT2440 proteome. 662 proteins were altered in abundance during growth on vanillin as sole carbon source as compared to growth on glucose. The proteome response entailed an increased abundance of enzymes involved in vanillin degradation, significant changes in central energy metabolism and an activation of solvent tolerance mechanisms. With respect to vanillin metabolism, particularly enzymes belonging to the β-ketoadipate pathway including a transcriptional regulator and porins specific for vanillin uptake increased in abundance. However, catabolism of vanillin was not dependent on vanillin dehydrogenase (Vdh), as shown by quantitative proteome analysis of a Vdh-deficient KT2440 mutant (GN235). Other aldehyde dehydrogenases that were significantly increased in abundance in response to vanillin may replace Vdh and thus may represent interesting targets for improving vanillin production in P. putida KT2440. The high demand for the flavor compound vanillin by the food and fragrance industry makes natural vanillin from vanilla pods a scarce and expensive resource rendering its biotechnological production economically attractive. Pseudomonas bacteria are metabolically very versatile and accept a broad range of hydrocarbons as carbon source making them suitable candidates for bioconversion processes. This work describes the impact of vanillin on the metabolism of the reference strain P. putida KT2440 on a

  17. Responses of KT2440 Pseudomonas putida to mild water stress

    DEFF Research Database (Denmark)

    Gülez, Gamze

    betydningen af flagellær motilitet og produktion af EPS under matric stess undersøgt ved brug af den såkaldte Porous Surface Model (PSM), som genererer væske-filmens effekter ved kontrol af !m. Det blev vist, at flagellær motilitet var begrænset under matric stress; Pseudomonas putida KT2440 kolonier udviste...... overflade-kolonidannelse under matric stress. Et af hovedformålene med denne afhandling var at forbedre PSM-metoden, således at rækken af matric potentials udvides (dvs. ned til -1.5 MPa, vegetations-visnings-punktet), idet det nuværende spektrum af matric potentials, der kan genereres med PSM-metoden, er...... smalt og nærmest begrænset til 1 m H2O (10 kPa). Med dette formål modificerede vi PSM-metoden, sådan at den ønskede !m kan fastsættes svarende til den negative værdi af det givne pneumatiske tryk. Denne modificerede model har vi kaldt Pressurized Porous Surface Model (PPSM). Gennem validering af...

  18. Proteomic characterization of the outer membrane vesicle of Pseudomonas putida KT2440.

    Science.gov (United States)

    Choi, Chi-Won; Park, Edmond Changkyun; Yun, Sung Ho; Lee, Sang-Yeop; Lee, Yeol Gyun; Hong, Yeonhee; Park, Kyeong Ryang; Kim, Sang-Hyun; Kim, Gun-Hwa; Kim, Seung Il

    2014-10-03

    Outer membrane vesicles (OMVs) are produced by various pathogenic Gram-negative bacteria such as Escherichia coli, Pseudomonas aeruginosa, and Acinetobacter baumannii. In this study, we isolated OMVs from a representative soil bacterium, Pseudomonas putida KT2440, which has a biodegradative activity toward various aromatic compounds. Proteomic analysis identified the outer membrane proteins (OMPs) OprC, OprD, OprE, OprF, OprH, OprG, and OprW as major components of the OMV of P. putida KT2440. The production of OMVs was dependent on the nutrient availability in the culture media, and the up- or down-regulation of specific OMPs was observed according to the culture conditions. In particular, porins (e.g., benzoate-specific porin, BenF-like porin) and enzymes (e.g., catechol 1,2-dioxygenase, benzoate dioxygenase) for benzoate degradation were uniquely found in OMVs prepared from P. putida KT2440 that were cultured in media containing benzoate as the energy source. OMVs of P. putida KT2440 showed low pathological activity toward cultured cells that originated from human lung cells, which suggests their potential as adjuvants or OMV vaccine carriers. Our results suggest that the protein composition of the OMVs of P. putida KT2440 reflects the characteristics of the total proteome of P. putida KT2440.

  19. Growth independent rhamnolipid production from glucose using the non-pathogenic Pseudomonas putida KT2440

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    Wittgens Andreas

    2011-10-01

    Full Text Available Abstract Background Rhamnolipids are potent biosurfactants with high potential for industrial applications. However, rhamnolipids are currently produced with the opportunistic pathogen Pseudomonas aeruginosa during growth on hydrophobic substrates such as plant oils. The heterologous production of rhamnolipids entails two essential advantages: Disconnecting the rhamnolipid biosynthesis from the complex quorum sensing regulation and the opportunity of avoiding pathogenic production strains, in particular P. aeruginosa. In addition, separation of rhamnolipids from fatty acids is difficult and hence costly. Results Here, the metabolic engineering of a rhamnolipid producing Pseudomonas putida KT2440, a strain certified as safety strain using glucose as carbon source to avoid cumbersome product purification, is reported. Notably, P. putida KT2440 features almost no changes in growth rate and lag-phase in the presence of high concentrations of rhamnolipids (> 90 g/L in contrast to the industrially important bacteria Bacillus subtilis, Corynebacterium glutamicum, and Escherichia coli. P. putida KT2440 expressing the rhlAB-genes from P. aeruginosa PAO1 produces mono-rhamnolipids of P. aeruginosa PAO1 type (mainly C10:C10. The metabolic network was optimized in silico for rhamnolipid synthesis from glucose. In addition, a first genetic optimization, the removal of polyhydroxyalkanoate formation as competing pathway, was implemented. The final strain had production rates in the range of P. aeruginosa PAO1 at yields of about 0.15 g/gglucose corresponding to 32% of the theoretical optimum. What's more, rhamnolipid production was independent from biomass formation, a trait that can be exploited for high rhamnolipid production without high biomass formation. Conclusions A functional alternative to the pathogenic rhamnolipid producer P. aeruginosa was constructed and characterized. P. putida KT24C1 pVLT31_rhlAB featured the highest yield and titer reported

  20. TOL Plasmid Carriage Enhances Biofilm Formation and Increases Extracellular DNA Content in Pseudomonas Putida KT2440

    DEFF Research Database (Denmark)

    Smets, Barth F.; D'Alvise, Paul; Yankelovich, T.

    Adherent growth of Pseudomonas putida KT2440 with and without the TOL plasmid (pWWO) at the solid-liquid and air-liquid interface was examined. We compared biofilm formation on glass in flow cells, and assayed pellicle (air-liquid interface biofilm) formation in stagnant liquid cultures by confoc...

  1. Carbon Source Influences Population Heterogeneity In Pseudomonas Putida Kt2440 Biofilms

    DEFF Research Database (Denmark)

    Juel Christensen, Anne-Mette; Sternberg, Claus; Molin, Søren

    2015-01-01

    Introduction: Pseudomonas putida is well known as a potential cell factory for many different biochemicals. Biofilm-based production can be advantageous for possibly toxic products due to increased chemical tolerance and robustness. Biofilm cells frequently differentiate, which challenges...... the benefits of biofilm-based production, and knowledge about factors driving the heterogeneity is therefore of importance.Methods: Biofilm flow chamber systems connected to confocal laser scanning microscopy were used to study biofilm structures of P. putida KT2440 at different carbon conditions. Subsequent...... plating of mature biofilm allowed for variant selection followed by pheno- and genotypic analysis.Results: Structure and cell differentiation in mature P. putida KT2440 biofilms were highly dependent on the type of carbon source utilised. Low glucose concentrations (0.3 mM – 10 mM) did not alter biofilm...

  2. Transcriptome Dynamics of Pseudomonas putida KT2440 under Water Stress

    DEFF Research Database (Denmark)

    Gülez, Gamze; Dechesne, Arnaud; Workman, Christopher;

    2012-01-01

    Water deprivation can be a major stressor to microbial life in surface and subsurface soil. In unsaturated soils, the matric potential (Ψm) is often the main component of the water potential, which measures the thermodynamic availability of water. A low matric potential usually translates...... into water forming thin liquid films in the soil pores. Little is known of how bacteria respond to such conditions, where, in addition to facing water deprivation that might impair their metabolism, they have to adapt their dispersal strategy as swimming motility may be compromised. Using the pressurized...

  3. Molecular characterization of the gallate dioxygenase from Pseudomonas putida KT2440. The prototype of a new subgroup of extradiol dioxygenases.

    Science.gov (United States)

    Nogales, Juan; Canales, Angeles; Jiménez-Barbero, Jesús; García, José Luis; Díaz, Eduardo

    2005-10-21

    In this work we have characterized the galA gene product from Pseudomonas putida KT2440, a ring-cleavage dioxygenase that acts specifically on gallate to produce 4-oxalomesaconate. The protein is a trimer composed by three identical subunits of 47.6 kDa (419 amino acids) that uses Fe2+ as the main cofactor. The gallate dioxygenase showed maximum activity at pH 7.0, and the Km and Vmax values for gallate were 144 microM and 53.2 micromol/min/mg of protein, respectively. A phylogenetic study suggests that the gallate dioxygenase from P. putida KT2440 is the prototype of a new subgroup of type II extradiol dioxygenases that share a common ancestor with protocatechuate 4,5-dioxygenases and whose two-domain architecture might have evolved from the fusion of the large and small subunits of the latter. A three-dimensional model for the N-terminal domain (residues 1-281) and C-terminal domain (residues 294-420) of the gallate dioxygenase from P. putida KT2440 was generated by comparison with the crystal structures of the large (LigB) and small (LigA) subunits of the protocatechuate 4,5-dioxygenase from Sphingomonas paucimobilis SYK-6. The expression of the galA gene was specifically induced when P. putida KT2440 cells grew in the presence of gallate. A P. putida KT2440 galA mutant strain was unable to use gallate as the sole carbon source and it did not show gallate dioxygenase activity, suggesting that the GalA protein is the only dioxygenase involved in gallate cleavage in this bacterium. This work points to the existence of a new pathway that is devoted to the catabolism of gallic acid and that remained unknown in the paradigmatic P. putida KT2440 strain.

  4. The Bistable Behaviour of Pseudomonas putida KT2440 during PHA Depolymerization under Carbon Limitation

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    Stephanie Karmann

    2017-06-01

    Full Text Available Poly(hydroxyalkanoates (PHAs are bacterial polyesters offering a biodegradable alternative to petrochemical plastics. The intracellular formation and degradation of PHAs is a dynamic process that strongly depends on the availability of carbon and other nutrients. Carbon excess and nitrogen limitation are considered to favor PHA accumulation, whereas carbon limitation triggers PHA depolymerization when all other essential nutrients are present in excess. We studied the population dynamics of Pseudomonas putida KT2440 at the single cell level during different physiological conditions, favoring first PHA polymerization during growth on octanoic acid, and then PHA depolymerization during carbon limitation. PHAs accumulate intracellularly in granules, and were proposed to separate preferentially together with nucleic acids, leading to two daughter cells containing approximately equal amounts of PHA. However, we could show that such P. putida KT2440 cells show bistable behavior when exposed to carbon limitation, and separate into two subpopulations: one with high and one with low PHA. This suggests an asymmetric PHA distribution during cell division under carbon limitation, which has a significant influence on our understanding of PHA mobilization.

  5. Analysis of the molecular response of Pseudomonas putida KT2440 to the next-generation biofuel n-butanol.

    Science.gov (United States)

    Simon, Oliver; Klebensberger, Janosch; Mükschel, Björn; Klaiber, Iris; Graf, Nadja; Altenbuchner, Josef; Huber, Armin; Hauer, Bernhard; Pfannstiel, Jens

    2015-06-03

    To increase the efficiency of biocatalysts a thorough understanding of the molecular response of the biocatalyst to precursors, products and environmental conditions applied in bioconversions is essential. Here we performed a comprehensive proteome and phospholipid analysis to characterize the molecular response of the potential biocatalyst Pseudomonas putida KT2440 to the next-generation biofuel n-butanol. Using complementary quantitative proteomics approaches we were able to identify and quantify 1467 proteins, corresponding to 28% of the total KT2440 proteome. 256 proteins were altered in abundance in response to n-butanol. The proteome response entailed an increased abundance of enzymes involved in n-butanol degradation including quinoprotein alcohol dehydrogenases, aldehyde dehydrogenases and enzymes of fatty acid beta oxidation. From these results we were able to construct a pathway for the metabolism of n-butanol in P. putida. The initial oxidation of n-butanol is catalyzed by at least two quinoprotein ethanol dehydrogenases (PedE and PedH). Growth of mutants lacking PedE and PedH on n-butanol was significantly impaired, but not completely inhibited, suggesting that additional alcohol dehydrogenases can at least partially complement their function in KT2440. Furthermore, phospholipid profiling revealed a significantly increased abundance of lyso-phospholipids in response to n-butanol, indicating a rearrangement of the lipid bilayer. n-butanol is an important bulk chemical and a promising alternative to gasoline as a transportation fuel. Due to environmental concerns as well as increasing energy prices there is a growing interest in sustainable and cost-effective biotechnological production processes for the production of bulk chemicals and transportation fuels from renewable resources. n-butanol fermentation is well established in Clostridiae, but the efficiency of n-butanol production is mainly limited by its toxicity. Therefore bacterial strains with

  6. Conversion and assimilation of furfural and 5-(hydroxymethylfurfural by Pseudomonas putida KT2440

    Directory of Open Access Journals (Sweden)

    Michael T. Guarnieri

    2017-06-01

    Full Text Available The sugar dehydration products, furfural and 5-(hydroxymethylfurfural (HMF, are commonly formed during high-temperature processing of lignocellulose, most often in thermochemical pretreatment, liquefaction, or pyrolysis. Typically, these two aldehydes are considered major inhibitors in microbial conversion processes. Many microbes can convert these compounds to their less toxic, dead-end alcohol counterparts, furfuryl alcohol and 5-(hydroxymethylfurfuryl alcohol. Recently, the genes responsible for aerobic catabolism of furfural and HMF were discovered in Cupriavidus basilensis HMF14 to enable complete conversion of these compounds to the TCA cycle intermediate, 2-oxo-glutarate. In this work, we engineer the robust soil microbe, Pseudomonas putida KT2440, to utilize furfural and HMF as sole carbon and energy sources via complete genomic integration of the 12 kB hmf gene cluster previously reported from Burkholderia phytofirmans. The common intermediate, 2-furoic acid, is shown to be a bottleneck for both furfural and HMF metabolism. When cultured on biomass hydrolysate containing representative amounts of furfural and HMF from dilute-acid pretreatment, the engineered strain outperforms the wild type microbe in terms of reduced lag time and enhanced growth rates due to catabolism of furfural and HMF. Overall, this study demonstrates that an approach for biological conversion of furfural and HMF, relative to the typical production of dead-end alcohols, enables both enhanced carbon conversion and substantially improves tolerance to hydrolysate inhibitors. This approach should find general utility both in emerging aerobic processes for the production of fuels and chemicals from biomass-derived sugars and in the biological conversion of high-temperature biomass streams from liquefaction or pyrolysis where furfural and HMF are much more abundant than in biomass hydrolysates from pretreatment.

  7. Conversion and assimilation of furfural and 5-(hydroxymethyl)furfural by Pseudomonas putida KT2440

    Energy Technology Data Exchange (ETDEWEB)

    Guarnieri, Michael T.; Ann Franden, Mary; Johnson, Christopher W.; Beckham, Gregg T.

    2017-02-08

    The sugar dehydration products, furfural and 5-(hydroxymethyl)furfural (HMF), are commonly formed during high-temperature processing of lignocellulose, most often in thermochemical pretreatment, liquefaction, or pyrolysis. Typically, these two aldehydes are considered major inhibitors in microbial conversion processes. Many microbes can convert these compounds to their less toxic, dead-end alcohol counterparts, furfuryl alcohol and 5-(hydroxymethyl)furfuryl alcohol. Recently, the genes responsible for aerobic catabolism of furfural and HMF were discovered in Cupriavidus basilensis HMF14 to enable complete conversion of these compounds to the TCA cycle intermediate, 2-oxo-glutarate. In this work, we engineer the robust soil microbe, Pseudomonas putida KT2440, to utilize furfural and HMF as sole carbon and energy sources via complete genomic integration of the 12 kB hmf gene cluster previously reported from Burkholderia phytofirmans. The common intermediate, 2-furoic acid, is shown to be a bottleneck for both furfural and HMF metabolism. When cultured on biomass hydrolysate containing representative amounts of furfural and HMF from dilute-acid pretreatment, the engineered strain outperforms the wild type microbe in terms of reduced lag time and enhanced growth rates due to catabolism of furfural and HMF. Overall, this study demonstrates that an approach for biological conversion of furfural and HMF, relative to the typical production of dead-end alcohols, enables both enhanced carbon conversion and substantially improves tolerance to hydrolysate inhibitors. This approach should find general utility both in emerging aerobic processes for the production of fuels and chemicals from biomass-derived sugars and in the biological conversion of high-temperature biomass streams from liquefaction or pyrolysis where furfural and HMF are much more abundant than in biomass hydrolysates from pretreatment.

  8. PH-stat fed-batch process to enhance the production of cis, cis-muconate from benzoate by Pseudomonas putida KT2440-JD1

    NARCIS (Netherlands)

    Duuren, J.B.J.H. van; Wijte, D.; Karge, B.; Martins dos Santos, V.A.; Yang, Y.; Mars, A.E.; Eggink, G.

    2012-01-01

    Pseudomonas putida KT2440-JD1 is able to cometabolize benzoate to cis, cis-muconate in the presence of glucose as growth substrate. P. putida KT2440-JD1 was unable to grow in the presence of concentrations above 50 mM benzoate or 600 mM cis, cis-muconate. The inhibitory effects of both compounds wer

  9. pH-stat fed-batch process to enhance the production of cis, cis-muconate from benzoate by Pseudomonas putida KT2440-JD1

    NARCIS (Netherlands)

    Duuren, van J.B.J.H.; Wijte, D.; Karge, B.; Martins Dos Santos, V.A.P.; Yang, Y.; Mars, A.E.; Eggink, G.

    2012-01-01

    Pseudomonas putida KT2440-JD1 is able to cometabolize benzoate to cis, cis-muconate in the presence of glucose as growth substrate. P. putida KT2440-JD1 was unable to grow in the presence of concentrations above 50 mM benzoate or 600 mM cis, cis-muconate. The inhibitory effects of both compounds wer

  10. TOL plasmid carriage enhances biofilm formation and increases extracellular DNA content in Pseudomonas putida KT2440

    DEFF Research Database (Denmark)

    D'Alvise, Paul; Sjoholm, O.R.; Yankelevich, T.;

    2010-01-01

    : TOL carriage leads to more extracellular DNA (eDNA) in pellicles and biofilms. Pellicles were dissolved by DNase I treatment. Enhanced cell lysis due to plasmid carriage was ruled out as the mechanism for eDNA release. We report, for the first time, that carriage of a conjugative plasmid leads......Adherent growth of Pseudomonas putida KT2440 with and without the TOL plasmid (pWWO) at the solid-liquid and air-liquid interface was examined. We compared biofilm formation on glass in flow cells, and assayed pellicle (air-liquid interface biofilm) formation in stagnant liquid cultures by confocal...... laser scanning microscopy. The TOL-carrying strains formed pellicles and thick biofilms, whereas the same strains without the plasmid displayed little adherent growth. Microscopy using fluorescent nucleic acid-specific stains revealed differences in the production of extracellular polymeric substances...

  11. TOL Plasmid Carriage Enhances Biofilm Formation and Increases Extracellular DNA Content in Pseudomonas Putida KT2440

    DEFF Research Database (Denmark)

    Smets, Barth F.; D'Alvise, Paul; Yankelovich, T.;

    of extracellular polymeric substances: TOL carriage leads to more extracellular DNA (eDNA) in pellicles and biofilms. Pellicles were dissolved by DNAse I treatment. eDNA was observed as ominous fibrous structures. Quantitative analysis of live and dead cells in static cultures was performed by flow cytometry......Adherent growth of Pseudomonas putida KT2440 with and without the TOL plasmid (pWWO) at the solid-liquid and air-liquid interface was examined. We compared biofilm formation on glass in flow cells, and assayed pellicle (air-liquid interface biofilm) formation in stagnant liquid cultures by confocal...... combined with specific cytostains; release of cytoplasmic material was assayed by a β-glucosidase assay. Enhanced cell lysis due to plasmid carriage was ruled out as the mechanism for eDNA release. We report, for the first time, that carriage of a conjugative plasmid leads to increased biofilm formation...

  12. Antimicrobial activities of commercial nanoparticles against an environmental soil microbe, Pseudomonas putida KT2440

    Directory of Open Access Journals (Sweden)

    Huang Wenjie

    2009-06-01

    Full Text Available Abstract Background The release of heavy metal-containing nanoparticles (NP into the environment may be harmful to the efficacy of beneficial microbes that function in element cycling, pollutant degradation and plant growth. Nanoparticles of Ag, CuO and ZnO are of interest as antimicrobials against pathogenic bacteria. We demonstrate here their antimicrobial activity against the beneficial soil microbe, Pseudomonas putida KT2440. Results Toxicity was detected in a KT2440 construct possessing a plasmid bearing the luxAB reporter genes. "As manufactured" preparations of nano- Ag, -CuO and -ZnO caused rapid dose-dependent loss of light output in the biosensor. Cell death accompanied loss in Lux activity with treatments by nano-Ag and -CuO, but with -ZnO the treatments were bacteriostatic rather than bactericidal. Bulk equivalents of these products showed no inhibitory activity, indicating that particle size was determinant in activity. Flow Field-Flow Fractionation (FlFFF of an aqueous suspension of the nano-CuO and ZnO revealed a small proportion of 5 nm NP and aggregated particulates with sizes ranging between 70 nm and 300 nm; the majority portion of material was aggregated into particles larger than 300 nm in size. Thus within the commercial preparation there may be microbially active and inactive forms. Conclusion The "as-made" NP of Ag, CuO and ZnO have toxic effects on a beneficial soil microbe, leading to bactericidal or bacteriostatic effects depending on the NP employed. The lack of toxicity from bulk materials suggests that aggregation of the NP into larger particles, possibly by factors present in the environment may reduce their nontarget antimicrobial activity.

  13. Biofilm as a production platform for heterologous production of rhamnolipids by the non‑pathogenic strain Pseudomonas putida KT2440

    DEFF Research Database (Denmark)

    Wigneswaran, Vinoth; Nielsen, Kristian Fog; Sternberg, Claus

    2016-01-01

    C10 fatty acids. Conclusion This study shows a successful application of synthetic promoter library in P. putida KT2440 and a heterologous biosynthesis of rhamnolipids in biofilm encased cells without hampering biofilm capabilities. These findings expands the possibilities of cultivation setups......Background Although a transition toward sustainable production of chemicals is needed, the physiochemical properties of certain biochemicals such as biosurfactants make them challenging to produce in conventional bioreactor systems. Alternative production platforms such as surface-attached biofilm...... as the model compound in biofilm encased Pseudomonas putida KT2440. The rhlAB operon from P. aeruginosa was introduced into P. putida to produce mono-rhamnolipids. A synthetic promoter library was used in order to bypass the normal regulation of rhamnolipid synthesis and to provide varying expression levels...

  14. Functional analysis of aromatic biosynthetic pathways in Pseudomonas putida KT2440.

    Science.gov (United States)

    Molina-Henares, M Antonia; García-Salamanca, Adela; Molina-Henares, A Jesús; de la Torre, Jesús; Herrera, M Carmen; Ramos, Juan L; Duque, Estrella

    2009-01-01

    Pseudomonas putida KT2440 is a non-pathogenic prototrophic bacterium with high potential for biotechnological applications. Despite all that is known about this strain, the biosynthesis of essential chemicals has not been fully analysed and auxotroph mutants are scarce. We carried out massive mini-Tn5 random mutagenesis and screened for auxotrophs that require aromatic amino acids. The biosynthesis of aromatic amino acids was analysed in detail including physical and transcriptional organization of genes, complementation assays and feeding experiments to establish pathway intermediates. There is a single pathway from chorismate leading to the biosynthesis of tryptophan, whereas the biosynthesis of phenylalanine and tyrosine is achieved through multiple convergent pathways. Genes for tryptophan biosynthesis are grouped in unlinked regions with the trpBA and trpGDE genes organized as operons and the trpI, trpE and trpF genes organized as single transcriptional units. The pheA and tyrA gene-encoding multifunctional enzymes for phenylalanine and tyrosine biosynthesis are linked in the chromosome and form an operon with the serC gene involved in serine biosynthesis. The last step in the biosynthesis of these two amino acids requires an amino transferase activity for which multiple tyrB-like genes are present in the host chromosome.

  15. TOL plasmid carriage enhances biofilm formation and increases extracellular DNA content in Pseudomonas putida KT2440.

    Science.gov (United States)

    D'Alvise, Paul W; Sjøholm, Ole R; Yankelevich, Tatiana; Jin, Yujie; Wuertz, Stefan; Smets, Barth F

    2010-11-01

    Adherent growth of Pseudomonas putida KT2440 with and without the TOL plasmid (pWWO) at the solid-liquid and air-liquid interface was examined. We compared biofilm formation on glass in flow cells, and assayed pellicle (air-liquid interface biofilm) formation in stagnant liquid cultures by confocal laser scanning microscopy. The TOL-carrying strains formed pellicles and thick biofilms, whereas the same strains without the plasmid displayed little adherent growth. Microscopy using fluorescent nucleic acid-specific stains revealed differences in the production of extracellular polymeric substances: TOL carriage leads to more extracellular DNA (eDNA) in pellicles and biofilms. Pellicles were dissolved by DNase I treatment. Enhanced cell lysis due to plasmid carriage was ruled out as the mechanism for eDNA release. We report, for the first time, that carriage of a conjugative plasmid leads to increased biofilm formation by production of eDNA. © 2010 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  16. Engineering Pseudomonas putida KT2440 for simultaneous degradation of organophosphates and pyrethroids and its application in bioremediation of soil.

    Science.gov (United States)

    Zuo, Zhenqiang; Gong, Ting; Che, You; Liu, Ruihua; Xu, Ping; Jiang, Hong; Qiao, Chuanling; Song, Cunjiang; Yang, Chao

    2015-06-01

    Agricultural soils are usually co-contaminated with organophosphate (OP) and pyrethroid pesticides. To develop a stable and marker-free Pseudomonas putida for co-expression of two pesticide-degrading enzymes, we constructed a suicide plasmid with expression cassettes containing a constitutive promoter J23119, an OP-degrading gene (mpd), a pyrethroid-hydrolyzing carboxylesterase gene (pytH) that utilizes the upp gene as a counter-selectable marker for upp-deficient P. putida. By introduction of suicide plasmid and two-step homologous recombination, both mpd and pytH genes were integrated into the chromosome of a robust soil bacterium P. putida KT2440 and no selection marker was left on chromosome. Functional expression of mpd and pytH in P. putida KT2440 was demonstrated by Western blot analysis and enzyme activity assays. Degradation experiments with liquid cultures showed that the mixed pesticides including methyl parathion, fenitrothion, chlorpyrifos, permethrin, fenpropathrin, and cypermethrin (0.2 mM each) were degraded completely within 48 h. The inoculation of engineered strain (10(6) cells/g) to soils treated with the above mixed pesticides resulted in a higher degradation rate than in noninoculated soils. All six pesticides could be degraded completely within 15 days in fumigated and nonfumigated soils with inoculation. Theses results highlight the potential of the engineered strain to be used for in situ bioremediation of soils co-contaminated with OP and pyrethroid pesticides.

  17. Systemic defense priming by Pseudomonas putida KT2440 in maize depends on benzoxazinoid exudation from the roots.

    Science.gov (United States)

    Neal, Andrew L; Ton, Jurriaan

    2013-01-01

    Exudation of benzoxazinoid metabolites from roots of young maize seedlings recruits the rhizobacterial strain Pseudomonas putida KT2440 from the soil to the rhizosphere. In this study, we have investigated whether these rhizobacteria are beneficial for maize by eliciting systemic defense priming. Root colonization of the maize hybrid cultivar Delprim by P. putida primed wound- and jasmonic acid (JA)-inducible emission of aromatic and terpenoid volatiles, but not the emission of the green leaf volatile (Z)-3-hexenyl acetate. Furthermore, root colonization by P. putida primed stress-inducible transcription of the JA-dependent gene SerPIN, whereas JA-dependent induction of the MPI gene was unaffected. Systemic priming of SerPIN by P. putida only occurred in benzoxazinoid-producing plants, and was absent in benzoxazinoid-deficient plants. The results from this study suggest that root colonization by P. putida primes a selection of JA-dependent defenses in Maize, which is reliant on benzoxazinoid exudation from the roots.

  18. Gene expression dynamics of pseudomonas putida KT2440 biofilms under water deprivation

    DEFF Research Database (Denmark)

    Gulez, Gamze; Dechesne, Arnaud; Workman, Christopher;

    2010-01-01

    In soil, bacteria can form colonies that are exposed to changing hydration conditions, exerting a stress to which the bacteria should adjust. Some of the phenotypes associated with water deprivation, such as the production extracellular polymeric substance (EPS) and the limitation of motility, have...... previously been observed. However, it is not known how these responses are regulated and temporally expressed. Here, we aimed to investigate the gene level responses by identifying the differentially expressed genes at -0.4 MPa water stress compared to non-stressed condition. We hypothesized that under water...... stress flagellar and EPS genes would be significantly differentially expressed, the former being down- and the latter being up-regulated. The novel Pressurized Porous Surface Model (PPSM) was used to expose KT2440 colonies to -0.4 MPa water stress for 4, 24, and 72 hours. Agilent whole genome 1-color c...

  19. Genetic engineering of Pseudomonas putida KT2440 for rapid and high-yield production of vanillin from ferulic acid.

    Science.gov (United States)

    Graf, Nadja; Altenbuchner, Josef

    2014-01-01

    Vanillin is one of the most important flavoring agents used today. That is why many efforts have been made on biotechnological production from natural abundant substrates. In this work, the nonpathogenic Pseudomonas putida strain KT2440 was genetically optimized to convert ferulic acid to vanillin. Deletion of the vanillin dehydrogenase gene (vdh) was not sufficient to prevent vanillin degradation. Additional inactivation of a molybdate transporter, identified by transposon mutagenesis, led to a strain incapable to grow on vanillin as sole carbon source. The bioconversion was optimized by enhanced chromosomal expression of the structural genes for feruloyl-CoA synthetase (fcs) and enoyl-CoA hydratase/aldolase (ech) by introduction of the strong tac promoter system. Further genetic engineering led to high initial conversion rates and molar vanillin yields up to 86% within just 3 h accompanied with very low by-product levels. To our knowledge, this represents the highest productivity and molar vanillin yield gained with a Pseudomonas strain so far. Together with its high tolerance for ferulic acid, the developed, plasmid-free P. putida strain represents a promising candidate for the biotechnological production of vanillin.

  20. Rhizosphere selection of Pseudomonas putida KT2440 variants with increased fitness associated to changes in gene expression.

    Science.gov (United States)

    Quesada, José Miguel; Fernández, Matilde; Soriano, María Isabel; Barrientos-Moreno, Laura; Llamas, María Antonia; Espinosa-Urgel, Manuel

    2016-08-03

    As the interface between plant roots and soil, the rhizosphere is a complex environment where nutrients released by the plant promote microbial growth. Increasing evidences indicate that the plant also exerts a selective pressure on microbial populations in the rhizosphere, favouring colonization by certain groups. In this work, we have designed an experimental setup to begin analysing the evolution of a specific bacterial population in the rhizosphere, using Pseudomonas putida KT2440 as model organism. After several rounds of selection without passage through laboratory growth conditions, derivatives of this strain with increased fitness in the rhizosphere were isolated. Detailed analysis of one of these clones indicated that this effect is specific for rhizosphere conditions and derives from changes in its transcriptional profile in this environment, with 43 genes being differentially expressed with respect to the parental strain. Several of these genes belong to functional categories which could affect stress adaptation and availability of particular nutrients. By inactivating two genes identified as upregulated in the selected clone (coding for a stress-response protein and a rRNA modifying protein), these functions were shown to contribute to rhizosphere fitness. Our data also suggest the existence of different evolutionary pathways leading to increased rhizosphere fitness.

  1. Production of medium-chain-length polyhydroxyalkanoates by sequential feeding of xylose and octanoic acid in engineered Pseudomonas putida KT2440

    Directory of Open Access Journals (Sweden)

    Le Meur Sylvaine

    2012-08-01

    Full Text Available Abstract Background Pseudomonas putida KT2440 is able to synthesize large amounts of medium-chain-length polyhydroxyalkanoates (mcl-PHAs. To reduce the substrate cost, which represents nearly 50% of the total PHA production cost, xylose, a hemicellulose derivate, was tested as the growth carbon source in an engineered P. putida KT2440 strain. Results The genes encoding xylose isomerase (XylA and xylulokinase (XylB from Escherichia coli W3110 were introduced into P. putida KT2440. The recombinant KT2440 exhibited a XylA activity of 1.47 U and a XylB activity of 0.97 U when grown on a defined medium supplemented with xylose. The cells reached a maximum specific growth rate of 0.24 h-1 and a final cell dry weight (CDW of 2.5 g L-1 with a maximal yield of 0.5 g CDW g-1 xylose. Since no mcl-PHA was accumulated from xylose, mcl-PHA production can be controlled by the addition of fatty acids leading to tailor-made PHA compositions. Sequential feeding strategy was applied using xylose as the growth substrate and octanoic acid as the precursor for mcl-PHA production. In this way, up to 20% w w-1 of mcl-PHA was obtained. A yield of 0.37 g mcl-PHA per g octanoic acid was achieved under the employed conditions. Conclusions Sequential feeding of relatively cheap carbohydrates and expensive fatty acids is a practical way to achieve more cost-effective mcl-PHA production. This study is the first reported attempt to produce mcl-PHA by using xylose as the growth substrate. Further process optimizations to achieve higher cell density and higher productivity of mcl-PHA should be investigated. These scientific exercises will undoubtedly contribute to the economic feasibility of mcl-PHA production from renewable feedstock.

  2. Analysis of Pseudomonas putida KT2440 gene expression in the maize rhizosphere: in vivo [corrected] expression technology capture and identification of root-activated promoters.

    Science.gov (United States)

    Ramos-González, María Isabel; Campos, María Jesús; Ramos, Juan L

    2005-06-01

    Pseudomonas putida KT2440, a paradigm organism in biodegradation and a good competitive colonizer of the maize rhizosphere, was the subject of studies undertaken to establish the genetic determinants important for its rhizospheric lifestyle. By using in vivo expression technology (IVET) to positively select single cell survival, we identified 28 rap genes (root-activated promoters) preferentially expressed in the maize rhizosphere. The IVET system had two components: a mutant affected in aspartate-beta-semialdehyde dehydrogenase (asd), which was unable to survive in the rhizosphere, and plasmid pOR1, which carries a promoter-less asd gene. pOR1-borne transcriptional fusions of the rap promoters to the essential gene asd, which were integrated into the chromosome at the original position of the corresponding rap gene, were active and allowed growth of the asd strain in the rhizosphere. The fact that five of the rap genes identified in the course of this work had been formerly characterized as being related to root colonization reinforced the IVET approach. Up to nine rap genes encoded proteins either of unknown function or that had been assigned an unspecific role based on conservation of the protein family domains. Rhizosphere-induced fusions included genes with probable functions in the cell envelope, chemotaxis and motility, transport, secretion, DNA metabolism and defense mechanism, regulation, energy metabolism, stress, detoxification, and protein synthesis.

  3. Genetic Dissection of the Regulatory Network Associated with High c-di-GMP Levels in Pseudomonas putida KT2440.

    Science.gov (United States)

    Ramos-González, María Isabel; Travieso, María L; Soriano, María I; Matilla, Miguel A; Huertas-Rosales, Óscar; Barrientos-Moreno, Laura; Tagua, Víctor G; Espinosa-Urgel, Manuel

    2016-01-01

    Most bacteria grow in nature forming multicellular structures named biofilms. The bacterial second messenger cyclic diguanosine monophosphate (c-di-GMP) is a key player in the regulation of the transition from planktonic to sessile lifestyles and this regulation is crucial in the development of biofilms. In Pseudomonas putida KT2440, Rup4959, a multidomain response regulator with diguanylate cyclase activity, when overexpressed causes an increment in the intracellular levels of c-di-GMP that gives rise to a pleiotropic phenotype consisting of increased biofilm formation and crinkly colony morphology. In a broad genomic screen we have isolated mutant derivatives that lose the crinkly morphology, designed as cfc (crinkle free colony). A total of 19 different genes have been identified as being related with the emergence of the cfc phenotype either because the expression or functionality of Rup4959 is compromised, or due to a lack of transduction of the c-di-GMP signal to downstream elements involved in the acquisition of the phenotype. Discernment between these possibilities was investigated by using a c-di-GMP biosensor and by HPLC-MS quantification of the second messenger. Interestingly five of the identified genes encode proteins with AAA+ ATPase domain. Among the bacterial determinants found in this screen are the global transcriptional regulators GacA, AlgU and FleQ and two enzymes involved in the arginine biosynthesis pathway. We present evidences that this pathway seems to be an important element to both the availability of the free pool of the second messenger c-di-GMP and to its further transduction as a signal for biosynthesis of biopolimers. In addition we have identified an uncharacterized hybrid sensor histidine kinase whose phosphoaceptor conserved histidine residue has been shown in this work to be required for in vivo activation of the orphan response regulator Rup4959, which suggests these two elements constitute a two-component phosphorelay system.

  4. Genetic Dissection of the Regulatory Network Associated with High C-di-GMP Levels in Pseudomonas putida KT2440

    Directory of Open Access Journals (Sweden)

    María Isabel Ramos-González

    2016-07-01

    Full Text Available Most bacteria grow in nature forming multicellular structures named biofilms. The bacterial second messenger cyclic diguanosine monophosphate (c-di-GMP is a key player in the regulation of the transition from planktonic to sessile lifestyles and this regulation is crucial in the development of biofilms. In Pseudomonas putida KT2440, Rup4959, a multidomain response regulator with diguanylate cyclase activity, when overexpressed causes an increment in the intracellular levels of c-di-GMP that gives rise to a pleiotropic phenotype consisting of increased biofilm formation and crinkly colony morphology. In a broad genomic screen we have isolated mutant derivatives that lose the crinkly morphology, designed as cfc (crinkle free colony. A total of nineteen different genes have been identified as being related with the emergence of the cfc phenotype either because the expression or functionality of Rup4959 is compromised, or due to a lack of transduction of the c-di-GMP signal to downstream elements involved in the acquisition of the phenotype. Discernment between these possibilities was investigated by using a c-di-GMP biosensor and by HPLC-MS quantification of the second messenger. Interestingly five of the identified genes encode proteins with AAA+ ATPase domain. Among the bacterial determinants found in this screen are the global transcriptional regulators GacA, AlgU and FleQ and two enzymes involved in the arginine biosynthesis pathway. We present evidences that this pathway seems to be an important element to both the availability of the free pool of the second messenger c-di-GMP and to its further transduction as a signal for biosynthesis of biopolimers. In addition we have identified an uncharacterized hybrid sensor histidine kinase whose phosphoaceptor conserved histidine residue has been shown in this work to be required for in vivo activation of the orphan response regulator Rup4959, which suggests these two elements constitute a two

  5. Deciphering the genetic determinants for aerobic nicotinic acid degradation: the nic cluster from Pseudomonas putida KT2440.

    Science.gov (United States)

    Jiménez, José I; Canales, Angeles; Jiménez-Barbero, Jesús; Ginalski, Krzysztof; Rychlewski, Leszek; García, José L; Díaz, Eduardo

    2008-08-12

    The aerobic catabolism of nicotinic acid (NA) is considered a model system for degradation of N-heterocyclic aromatic compounds, some of which are major environmental pollutants; however, the complete set of genes as well as the structural-functional relationships of most of the enzymes involved in this process are still unknown. We have characterized a gene cluster (nic genes) from Pseudomonas putida KT2440 responsible for the aerobic NA degradation in this bacterium and when expressed in heterologous hosts. The biochemistry of the NA degradation through the formation of 2,5-dihydroxypyridine and maleamic acid has been revisited, and some gene products become the prototype of new types of enzymes with unprecedented molecular architectures. Thus, the initial hydroxylation of NA is catalyzed by a two-component hydroxylase (NicAB) that constitutes the first member of the xanthine dehydrogenase family whose electron transport chain to molecular oxygen includes a cytochrome c domain. The Fe(2+)-dependent dioxygenase (NicX) converts 2,5-dihydroxypyridine into N-formylmaleamic acid, and it becomes the founding member of a new family of extradiol ring-cleavage dioxygenases. Further conversion of N-formylmaleamic acid to formic and maleamic acid is catalyzed by the NicD protein, the only deformylase described so far whose catalytic triad is similar to that of some members of the alpha/beta-hydrolase fold superfamily. This work allows exploration of the existence of orthologous gene clusters in saprophytic bacteria and some pathogens, where they might stimulate studies on their role in virulence, and it provides a framework to develop new biotechnological processes for detoxification/biotransformation of N-heterocyclic aromatic compounds.

  6. Genetic Dissection of the Regulatory Network Associated with High c-di-GMP Levels in Pseudomonas putida KT2440

    Science.gov (United States)

    Ramos-González, María Isabel; Travieso, María L.; Soriano, María I.; Matilla, Miguel A.; Huertas-Rosales, Óscar; Barrientos-Moreno, Laura; Tagua, Víctor G.; Espinosa-Urgel, Manuel

    2016-01-01

    Most bacteria grow in nature forming multicellular structures named biofilms. The bacterial second messenger cyclic diguanosine monophosphate (c-di-GMP) is a key player in the regulation of the transition from planktonic to sessile lifestyles and this regulation is crucial in the development of biofilms. In Pseudomonas putida KT2440, Rup4959, a multidomain response regulator with diguanylate cyclase activity, when overexpressed causes an increment in the intracellular levels of c-di-GMP that gives rise to a pleiotropic phenotype consisting of increased biofilm formation and crinkly colony morphology. In a broad genomic screen we have isolated mutant derivatives that lose the crinkly morphology, designed as cfc (crinkle free colony). A total of 19 different genes have been identified as being related with the emergence of the cfc phenotype either because the expression or functionality of Rup4959 is compromised, or due to a lack of transduction of the c-di-GMP signal to downstream elements involved in the acquisition of the phenotype. Discernment between these possibilities was investigated by using a c-di-GMP biosensor and by HPLC-MS quantification of the second messenger. Interestingly five of the identified genes encode proteins with AAA+ ATPase domain. Among the bacterial determinants found in this screen are the global transcriptional regulators GacA, AlgU and FleQ and two enzymes involved in the arginine biosynthesis pathway. We present evidences that this pathway seems to be an important element to both the availability of the free pool of the second messenger c-di-GMP and to its further transduction as a signal for biosynthesis of biopolimers. In addition we have identified an uncharacterized hybrid sensor histidine kinase whose phosphoaceptor conserved histidine residue has been shown in this work to be required for in vivo activation of the orphan response regulator Rup4959, which suggests these two elements constitute a two-component phosphorelay system

  7. Siderophore-mediated iron acquisition in the entomopathogenic bacterium Pseudomonas entomophila L48 and its close relative Pseudomonas putida KT2440.

    Science.gov (United States)

    Matthijs, Sandra; Laus, Georges; Meyer, Jean-Marie; Abbaspour-Tehrani, Kourosch; Schäfer, Mathias; Budzikiewicz, Herbert; Cornelis, Pierre

    2009-12-01

    Pseudomonas entomophila L48 is a recently identified entomopathogenic bacterium which, upon ingestion, kills Drosophila melanogaster, and is closely related to P. putida. The complete genome of this species has been sequenced and therefore a genomic, genetic and structural analysis of the siderophore-mediated iron acquisition was undertaken. P. entomophila produces two siderophores, a structurally new and unique pyoverdine and the secondary siderophore pseudomonine, already described in P. fluorescens species. Structural analysis of the pyoverdine produced by the closely related P. putida KT2440 showed that this strain produces an already characterised pyoverdine, but different from P. entomophila, and no evidence was found for the production of a second siderophore. Growth stimulation assays with heterologous pyoverdines demonstrated that P. entomophila is able to utilize a large variety of structurally distinct pyoverdines produced by other Pseudomonas species. In contrast, P. putida KT2440 is able to utilize only its own pyoverdine and the pyoverdine produced by P. syringae LMG 1247. Our data suggest that although closely related, P. entomophila is a more efficient competitor for iron than P. putida.

  8. Catabolismo de la prolina en pseudomonas putida Kt2440 : construcci??n de un sistema de contenci??n biol??gica dependiente de prolina

    OpenAIRE

    V??lchez Tornero, Susana

    2000-01-01

    En esta tesis se ha estudiado el catabolismo de la prolina en P.putida KT2440. El estudio incluye el aislamiento y secuenciaci??n de los genes putA y putP responsables del catabolismo de la prolina en este organismo, as?? como el aislamiento y caracterizaci??n de mutantes deficientes en la lutilizaci??nd e este compuesto. Tras la secuenciaci??n del oper??n se caracteriz?? la expresi??n de los genes implicados en el catabolismo de la prolina en distintas condiciones y fond...

  9. Catabolismo de la prolina en pseudomonas putida Kt2440 : construcción de un sistema de contención biológica dependiente de prolina

    OpenAIRE

    Vílchez Tornero, Susana

    2013-01-01

    En esta tesis se ha estudiado el catabolismo de la prolina en P.putida KT2440. El estudio incluye el aislamiento y secuenciación de los genes putA y putP responsables del catabolismo de la prolina en este organismo, así como el aislamiento y caracterización de mutantes deficientes en la lutilizaciónd e este compuesto. Tras la secuenciación del operón se caracterizó la expresión de los genes implicados en el catabolismo de la prolina en distintas condiciones y fondos genét...

  10. Toxicity of fungal-generated silver nanoparticles to soil-inhabiting Pseudomonas putida KT2440, a rhizospheric bacterium responsible for plant protection and bioremediation

    Energy Technology Data Exchange (ETDEWEB)

    Gupta, Indarchand R. [Nanobiotechnology Laboratory, Department of Biotechnology, S.G.B. Amravati University, Amravati 444602, Maharashtra (India); Department of Biotechnology, Institute of Science, Nipat Niranjan Nagar, Caves Road, Aurangabad 431004, Maharashtra (India); Anderson, Anne J. [Department of Biology, Utah State University, Logan, Utah 84321 (United States); Rai, Mahendra, E-mail: mahendrarai@sgbau.ac.in [Nanobiotechnology Laboratory, Department of Biotechnology, S.G.B. Amravati University, Amravati 444602, Maharashtra (India); Laboratório de Química Biológica, Instituto de Química, UNICAMP, Cidade Universitária “Zefferino Vaz” Barão Geraldo, CEP 13083-970, Caixa Postal 6150, Campinas, SP (Brazil)

    2015-04-09

    Highlights: • This study incorporates the mycosynthesis of AgNPs and their characterisation by various methods. • A first attempt demonstrating the toxicity assessment of AgNPs on beneficial soil microbe. • Use of biosensor in Pseudomonas putida KT2440, gave accurate antimicrobial results. - Abstract: Silver nanoparticles have attracted considerable attention due to their beneficial properties. But toxicity issues associated with them are also rising. The reports in the past suggested health hazards of silver nanoparticles at the cellular, molecular, or whole organismal level in eukaryotes. Whereas, there is also need to examine the exposure effects of silver nanoparticle to the microbes, which are beneficial to humans as well as environment. The available literature suggests the harmful effects of physically and chemically synthesised silver nanoparticles. The toxicity of biogenically synthesized nanoparticles has been less studied than physically and chemically synthesised nanoparticles. Hence, there is a greater need to study the toxic effects of biologically synthesised silver nanoparticles in general and mycosynthesized nanoparticles in particular. In the present study, attempts have been made to assess the risk associated with the exposure of mycosynthesized silver nanoparticles on a beneficial soil microbe Pseudomonas putida. KT2440. The study demonstrates mycosynthesis of silver nanoparticles and their characterisation by UV–vis spectrophotometry, FTIR, X-ray diffraction, nanosight LM20 – a particle size distribution analyzer and TEM. Silver nanoparticles obtained herein were found to exert the hazardous effect at the concentration of 0.4 μg/ml, which warrants further detailed investigations concerning toxicity.

  11. Ethylene Glycol Metabolism by Pseudomonas putida

    OpenAIRE

    Mückschel, Björn; Simon, Oliver; Klebensberger, Janosch; Graf, Nadja; Rosche, Bettina; Altenbuchner, Josef; Pfannstiel, Jens; Huber, Armin; Hauer, Bernhard

    2012-01-01

    In this study, we investigated the metabolism of ethylene glycol in the Pseudomonas putida strains KT2440 and JM37 by employing growth and bioconversion experiments, directed mutagenesis, and proteome analysis. We found that strain JM37 grew rapidly with ethylene glycol as a sole source of carbon and energy, while strain KT2440 did not grow within 2 days of incubation under the same conditions. However, bioconversion experiments revealed metabolism of ethylene glycol by both strains, with the...

  12. Toxicity of fungal-generated silver nanoparticles to soil-inhabiting Pseudomonas putida KT2440, a rhizospheric bacterium responsible for plant protection and bioremediation.

    Science.gov (United States)

    Gupta, Indarchand R; Anderson, Anne J; Rai, Mahendra

    2015-04-09

    Silver nanoparticles have attracted considerable attention due to their beneficial properties. But toxicity issues associated with them are also rising. The reports in the past suggested health hazards of silver nanoparticles at the cellular, molecular, or whole organismal level in eukaryotes. Whereas, there is also need to examine the exposure effects of silver nanoparticle to the microbes, which are beneficial to humans as well as environment. The available literature suggests the harmful effects of physically and chemically synthesised silver nanoparticles. The toxicity of biogenically synthesized nanoparticles has been less studied than physically and chemically synthesised nanoparticles. Hence, there is a greater need to study the toxic effects of biologically synthesised silver nanoparticles in general and mycosynthesized nanoparticles in particular. In the present study, attempts have been made to assess the risk associated with the exposure of mycosynthesized silver nanoparticles on a beneficial soil microbe Pseudomonas putida. KT2440. The study demonstrates mycosynthesis of silver nanoparticles and their characterisation by UV-vis spectrophotometry, FTIR, X-ray diffraction, nanosight LM20--a particle size distribution analyzer and TEM. Silver nanoparticles obtained herein were found to exert the hazardous effect at the concentration of 0.4 μg/ml, which warrants further detailed investigations concerning toxicity.

  13. Ethylene glycol metabolism by Pseudomonas putida.

    Science.gov (United States)

    Mückschel, Björn; Simon, Oliver; Klebensberger, Janosch; Graf, Nadja; Rosche, Bettina; Altenbuchner, Josef; Pfannstiel, Jens; Huber, Armin; Hauer, Bernhard

    2012-12-01

    In this study, we investigated the metabolism of ethylene glycol in the Pseudomonas putida strains KT2440 and JM37 by employing growth and bioconversion experiments, directed mutagenesis, and proteome analysis. We found that strain JM37 grew rapidly with ethylene glycol as a sole source of carbon and energy, while strain KT2440 did not grow within 2 days of incubation under the same conditions. However, bioconversion experiments revealed metabolism of ethylene glycol by both strains, with the temporal accumulation of glycolic acid and glyoxylic acid for strain KT2440. This accumulation was further increased by targeted mutagenesis. The key enzymes and specific differences between the two strains were identified by comparative proteomics. In P. putida JM37, tartronate semialdehyde synthase (Gcl), malate synthase (GlcB), and isocitrate lyase (AceA) were found to be induced in the presence of ethylene glycol or glyoxylic acid. Under the same conditions, strain KT2440 showed induction of AceA only. Despite this difference, the two strains were found to use similar periplasmic dehydrogenases for the initial oxidation step of ethylene glycol, namely, the two redundant pyrroloquinoline quinone (PQQ)-dependent enzymes PedE and PedH. From these results we constructed a new pathway for the metabolism of ethylene glycol in P. putida. Furthermore, we conclude that Pseudomonas putida might serve as a useful platform from which to establish a whole-cell biocatalyst for the production of glyoxylic acid from ethylene glycol.

  14. Colony morphology and transcriptome profiling of Pseudomonas putida KT2440 and its mutants deficient in alginate or all EPS synthesis under controlled matric potentials

    DEFF Research Database (Denmark)

    Gülez, Gamze; Altintas, Ali; Fazli, Mustafa;

    2014-01-01

    Pseudomonas putida is a versatile bacterial species adapted to soil and its fluctuations. Like many other species living in soil, P. putida often faces water limitation. Alginate, an exopolysaccharide (EPS) produced by P. putida, is known to create hydrated environments and alleviate the effect...... of water limitation. In addition to alginate, P. putida is capable of producing cellulose (bcs), putida exopolysaccharide a (pea), and putida exopolysaccharide b (peb). However, unlike alginate, not much is known about their roles under water limitation. Hence, in this study we examined the role...... of different EPS components under mild water limitation. To create environmentally realistic water limited conditions as observed in soil, we used the Pressurized Porous Surface Model. Our main hypothesis was that under water limitation and in the absence of alginate other exopolysaccharides would be more...

  15. Broad host range ProUSER vectors enable fast characterization of inducible promoters and optimization of p-coumaric acid production in Pseudomonas putida KT2440

    DEFF Research Database (Denmark)

    Calero Valdayo, Patricia; Ingemann Jensen, Sheila; Nielsen, Alex Toftgaard

    2016-01-01

    of ProUSER-reporter vectors was further created to characterize different inducible promoters. The PrhaB and Pm promoters were orthogonal and showed titratable, high, and homogeneous expression. To optimize the production of p-coumaric acid, P. putida was engineered to prevent degradation of tyrosine...... and p-coumaric acid. Pm and PrhaB were used to control the expression of a tyrosine ammonia lyase or AroG* and TyrA* involved in tyrosine production, respectively. Pathway expression was optimized by modulating inductions, resulting in small-scale p-coumaric acid production of 1.2 mM, the highest...... achieved in Pseudomonads under comparable conditions. With broad-host-range compatibility, the ProUSER vectors will serve as useful tools for optimizing gene expression in a variety of bacteria....

  16. Functional Characterization of the Mannitol Promoter of Pseudomonas fluorescens DSM 50106 and Its Application for a Mannitol-Inducible Expression System for Pseudomonas putida KT2440.

    Directory of Open Access Journals (Sweden)

    Jana Hoffmann

    Full Text Available A new pBBR1MCS-2-derived vector containing the Pseudomonas fluorescens DSM10506 mannitol promoter PmtlE and mtlR encoding its AraC/XylS type transcriptional activator was constructed and optimized for low basal expression. Mannitol, arabitol, and glucitol-inducible gene expression was demonstrated with Pseudomonas putida and eGFP as reporter gene. The new vector was applied for functional characterization of PmtlE. Identification of the DNA binding site of MtlR was achieved by in vivo eGFP measurement with PmtlE wild type and mutants thereof. Moreover, purified MtlR was applied for detailed in vitro investigations using electrophoretic mobility shift assays and DNaseI footprinting experiments. The obtained data suggest that MtlR binds to PmtlE as a dimer. The proposed DNA binding site of MtlR is AGTGC-N5-AGTAT-N7-AGTGC-N5-AGGAT. The transcription activation mechanism includes two binding sites with different binding affinities, a strong upstream binding site and a weaker downstream binding site. The presence of the weak downstream binding site was shown to be necessary to sustain mannitol-inducibility of PmtlE. Two possible functions of mannitol are discussed; the effector might stabilize binding of the second monomer to the downstream half site or promote transcription activation by inducing a conformational change of the regulator that influences the contact to the RNA polymerase.

  17. Benzoxazinoids in root exudates of maize attract Pseudomonas putida to the rhizosphere.

    Directory of Open Access Journals (Sweden)

    Andrew L Neal

    Full Text Available Benzoxazinoids, such as 2,4-dihydroxy-7-methoxy-2H-1,4-benzoxazin-3(4H-one (DIMBOA, are secondary metabolites in grasses. In addition to their function in plant defence against pests and diseases above-ground, benzoxazinoids (BXs have also been implicated in defence below-ground, where they can exert allelochemical or antimicrobial activities. We have studied the impact of BXs on the interaction between maize and Pseudomonas putida KT2440, a competitive coloniser of the maize rhizosphere with plant-beneficial traits. Chromatographic analyses revealed that DIMBOA is the main BX compound in root exudates of maize. In vitro analysis of DIMBOA stability indicated that KT2440 tolerance of DIMBOA is based on metabolism-dependent breakdown of this BX compound. Transcriptome analysis of DIMBOA-exposed P. putida identified increased transcription of genes controlling benzoate catabolism and chemotaxis. Chemotaxis assays confirmed motility of P. putida towards DIMBOA. Moreover, colonisation essays in soil with Green Fluorescent Protein (GFP-expressing P. putida showed that DIMBOA-producing roots of wild-type maize attract significantly higher numbers of P. putida cells than roots of the DIMBOA-deficient bx1 mutant. Our results demonstrate a central role for DIMBOA as a below-ground semiochemical for recruitment of plant-beneficial rhizobacteria during the relatively young and vulnerable growth stages of maize.

  18. Benzoxazinoids in root exudates of maize attract Pseudomonas putida to the rhizosphere.

    Science.gov (United States)

    Neal, Andrew L; Ahmad, Shakoor; Gordon-Weeks, Ruth; Ton, Jurriaan

    2012-01-01

    Benzoxazinoids, such as 2,4-dihydroxy-7-methoxy-2H-1,4-benzoxazin-3(4H)-one (DIMBOA), are secondary metabolites in grasses. In addition to their function in plant defence against pests and diseases above-ground, benzoxazinoids (BXs) have also been implicated in defence below-ground, where they can exert allelochemical or antimicrobial activities. We have studied the impact of BXs on the interaction between maize and Pseudomonas putida KT2440, a competitive coloniser of the maize rhizosphere with plant-beneficial traits. Chromatographic analyses revealed that DIMBOA is the main BX compound in root exudates of maize. In vitro analysis of DIMBOA stability indicated that KT2440 tolerance of DIMBOA is based on metabolism-dependent breakdown of this BX compound. Transcriptome analysis of DIMBOA-exposed P. putida identified increased transcription of genes controlling benzoate catabolism and chemotaxis. Chemotaxis assays confirmed motility of P. putida towards DIMBOA. Moreover, colonisation essays in soil with Green Fluorescent Protein (GFP)-expressing P. putida showed that DIMBOA-producing roots of wild-type maize attract significantly higher numbers of P. putida cells than roots of the DIMBOA-deficient bx1 mutant. Our results demonstrate a central role for DIMBOA as a below-ground semiochemical for recruitment of plant-beneficial rhizobacteria during the relatively young and vulnerable growth stages of maize.

  19. Benzoxazinoids in Root Exudates of Maize Attract Pseudomonas putida to the Rhizosphere

    Science.gov (United States)

    Neal, Andrew L.; Ahmad, Shakoor; Gordon-Weeks, Ruth; Ton, Jurriaan

    2012-01-01

    Benzoxazinoids, such as 2,4-dihydroxy-7-methoxy-2H-1,4-benzoxazin-3(4H)-one (DIMBOA), are secondary metabolites in grasses. In addition to their function in plant defence against pests and diseases above-ground, benzoxazinoids (BXs) have also been implicated in defence below-ground, where they can exert allelochemical or antimicrobial activities. We have studied the impact of BXs on the interaction between maize and Pseudomonas putida KT2440, a competitive coloniser of the maize rhizosphere with plant-beneficial traits. Chromatographic analyses revealed that DIMBOA is the main BX compound in root exudates of maize. In vitro analysis of DIMBOA stability indicated that KT2440 tolerance of DIMBOA is based on metabolism-dependent breakdown of this BX compound. Transcriptome analysis of DIMBOA-exposed P. putida identified increased transcription of genes controlling benzoate catabolism and chemotaxis. Chemotaxis assays confirmed motility of P. putida towards DIMBOA. Moreover, colonisation essays in soil with Green Fluorescent Protein (GFP)-expressing P. putida showed that DIMBOA-producing roots of wild-type maize attract significantly higher numbers of P. putida cells than roots of the DIMBOA-deficient bx1 mutant. Our results demonstrate a central role for DIMBOA as a below-ground semiochemical for recruitment of plant-beneficial rhizobacteria during the relatively young and vulnerable growth stages of maize. PMID:22545111

  20. Regulatory tasks of the phosphoenolpyruvate-phosphotransferase system of Pseudomonas putida in central carbon metabolism.

    Science.gov (United States)

    Chavarría, Max; Kleijn, Roelco J; Sauer, Uwe; Pflüger-Grau, Katharina; de Lorenzo, Víctor

    2012-01-01

    Two branches of the phosphoenolpyruvate-phosphotransferase system (PTS) operate in the soil bacterium Pseudomonas putida KT2440. One branch encompasses a complete set of enzymes for fructose intake (PTS(Fru)), while the other (N-related PTS, or PTS(Ntr)) controls various cellular functions unrelated to the transport of carbohydrates. The potential of these two systems for regulating central carbon catabolism has been investigated by measuring the metabolic fluxes of isogenic strains bearing nonpolar mutations in PTS(Fru) or PTS(Ntr) genes and grown on either fructose (a PTS substrate) or glucose, the transport of which is not governed by the PTS in this bacterium. The flow of carbon from each sugar was distinctly split between the Entner-Doudoroff, pentose phosphate, and Embden-Meyerhof-Parnas pathways in a ratio that was maintained in each of the PTS mutants examined. However, strains lacking PtsN (EIIA(Ntr)) displayed significantly higher fluxes in the reactions of the pyruvate shunt, which bypasses malate dehydrogenase in the TCA cycle. This was consistent with the increased activity of the malic enzyme and the pyruvate carboxylase found in the corresponding PTS mutants. Genetic evidence suggested that such a metabolic effect of PtsN required the transfer of high-energy phosphate through the system. The EIIA(Ntr) protein of the PTS(Ntr) thus helps adjust central metabolic fluxes to satisfy the anabolic and energetic demands of the overall cell physiology. This study demonstrates that EIIA(Ntr) influences the biochemical reactions that deliver carbon between the upper and lower central metabolic domains for the consumption of sugars by P. putida. These findings indicate that the EIIA(Ntr) protein is a key player for orchestrating the fate of carbon in various physiological destinations in this bacterium. Additionally, these results highlight the importance of the posttranslational regulation of extant enzymatic complexes for increasing the robustness of the

  1. Pseudomonas putida KT2440 as a host for biochemicals production

    DEFF Research Database (Denmark)

    Calero Valdayo, Patricia

    The extensive use of cell factories for production of biochemicals is still facing a number of challenges to become economically competitive. One of the problems that need to be overcome is the toxicity of certain chemical products, which prevents yields high enough for their implementation in in...... for alternative microorganisms as well as identifying mechanisms that can be applied in the design and construction of cell factories will assist in the advance of biosustainability, leading us to a future free of petroleum-based industry....

  2. Transcriptional regulation of pWW0 transfer genes in Pseudomonas putida KT2440

    DEFF Research Database (Denmark)

    Lambertsen, L.M.; Molin, Søren; Kroer, N.;

    2004-01-01

    The conjugative IncP-9 plasmid pWW0 (TOL) carries transfer genes, many of whose functions can be predicted from sequence similarities to the well-studied IncW and IncP-1 plasmids, and that are clustered with the replication and maintenance genes of the plasmid core. In this study we show that the...

  3. A reduction in growth rate of Pseudomonas putida KT2442 counteracts productivity advances in medium-chain-length polyhydroxyalkanoate production from gluconate

    Directory of Open Access Journals (Sweden)

    Zinn Manfred

    2011-04-01

    Full Text Available Abstract Background The substitution of plastics based on fossil raw material by biodegradable plastics produced from renewable resources is of crucial importance in a context of oil scarcity and overflowing plastic landfills. One of the most promising organisms for the manufacturing of medium-chain-length polyhydroxyalkanoates (mcl-PHA is Pseudomonas putida KT2440 which can accumulate large amounts of polymer from cheap substrates such as glucose. Current research focuses on enhancing the strain production capacity and synthesizing polymers with novel material properties. Many of the corresponding protocols for strain engineering rely on the rifampicin-resistant variant, P. putida KT2442. However, it remains unclear whether these two strains can be treated as equivalent in terms of mcl-PHA production, as the underlying antibiotic resistance mechanism involves a modification in the RNA polymerase and thus has ample potential for interfering with global transcription. Results To assess PHA production in P. putida KT2440 and KT2442, we characterized the growth and PHA accumulation on three categories of substrate: PHA-related (octanoate, PHA-unrelated (gluconate and poor PHA substrate (citrate. The strains showed clear differences of growth rate on gluconate and citrate (reduction for KT2442 > 3-fold and > 1.5-fold, respectively but not on octanoate. In addition, P. putida KT2442 PHA-free biomass significantly decreased after nitrogen depletion on gluconate. In an attempt to narrow down the range of possible reasons for this different behavior, the uptake of gluconate and extracellular release of the oxidized product 2-ketogluconate were measured. The results suggested that the reason has to be an inefficient transport or metabolization of 2-ketogluconate while an alteration of gluconate uptake and conversion to 2-ketogluconate could be excluded. Conclusions The study illustrates that the recruitment of a pleiotropic mutation, whose effects might

  4. The sigma(54) regulon (sigmulon) of Pseudomonas putida

    DEFF Research Database (Denmark)

    Cases, I.; Ussery, David; de Lorenzo, V.

    2003-01-01

    , the sigma(54) regulon has been studied both in Escherichia coli, Salmonella typhimurium and several species of the Rhizobiaceae. Here we present the analysis of the sigma(54) regulon (sigmulon) in the complete genome of Pseudomonas putida KT2440. We have developed an improved method for the prediction...

  5. A substrate dependent biological containment systems for Pseudomonas putida based on the Escherichia coli gef gene

    DEFF Research Database (Denmark)

    Jensen, Lars Bogø; Ramos, J. L.; Kaneva, Z.;

    1993-01-01

    A model substrate-dependent suicide system to biologically contain Pseudomonas putida KT2440 is reported. The system consists of two elements. One element carries a fusion between a synthetic lac promoter (PA1-04/03) and the gef gene, which encodes a killing function. This element is contained...... operon (Pm) and the lacI gene, encoding the Lac repressor, plus xylS2, coding for a positive regulator of Pm. In liquid culture under optimal growth conditions and in sterile and nonsterile soil microcosms, P. putida KT2440 (pWWO) bearing the containment system behaves as designed. In the presence...

  6. The Identification and Validation of Novel Small Proteins in Pseudomonas Putida KT-2440

    DEFF Research Database (Denmark)

    Yang, Xiaochen; Long, Katherine

    2014-01-01

    is to establish methods for the identification of short open reading frames (sORFs). A custom bioinformatics tool has been developed to predict sORFs from the genome and sRNA sequencing databases, where sequences containing a Shine-­Dalgarno (SD) sequence and start/stop codon are identified. In a second approach...

  7. Global Transcriptional Responses to Osmotic, Oxidative, and Imipenem Stress Conditions in Pseudomonas putida

    DEFF Research Database (Denmark)

    Bojanovic, Klara; D'Arrigo, Isotta; Long, Katherine

    2017-01-01

    Bacteria cope with and adapt to stress by modulating gene expression in response to specific environmental cues. In this study, the transcriptional response of Pseudomonas putida KT2440 to osmotic, oxidative, and imipenem stress conditions at two time points was investigated via identification of...

  8. Novel small protein identification and quantitative proteomic analysis in Pseudomonas putida KT-­2440

    DEFF Research Database (Denmark)

    Yang, Xiaochen

    .This thesis investigated an industrial bacterium, Pseudomonas putida KT-2440, in two aspects. First, the research focused on discovering novel small proteins (s-proteins) in the bacterium. With large-scale approaches for gene identification, groups of novel s-proteins were identified and validated from...... the genome, transcriptome and proteome of the bacterium. The application of new research approach, ribosome profiling, enabled us to analysis novel open reading frames (ORFs) from different standpoint. Second, by quantitative proteomic approach, the differential expressions of genes were analyzed at proteome...... level under different environmental conditions. The results yield insights intothe adaptation of P. putida KT-2440 in different environments.Based on bioinformatic, proteomic and transcriptomic approaches, global gene expression was analyzed on both transcriptional and translational levels. Our research...

  9. Global transcriptional response of solvent-sensitive and solvent-tolerant Pseudomonas putida strains exposed to toluene.

    Science.gov (United States)

    Molina-Santiago, Carlos; Udaondo, Zulema; Gómez-Lozano, María; Molin, Soren; Ramos, Juan-Luis

    2017-02-01

    Pseudomonas putida strains are generally recognized as solvent tolerant, exhibiting varied sensitivity to organic solvents. Pan-genome analysis has revealed that 30% of genes belong to the core-genome of Pseudomonas. Accessory and unique genes confer high degree of adaptability and capabilities for the degradation and synthesis of a wide range of chemicals. For the use of these microbes in bioremediation and biocatalysis, it is critical to understand the mechanisms underlying these phenotypic differences. In this study, RNA-seq analysis compared the short- and long-term responses of the toluene-sensitive KT2440 strain and the highly tolerant DOT-T1E strain. The sensitive strain activates a larger number of genes in a higher magnitude than DOT-T1E. This is expected because KT2440 bears one toluene tolerant pump, while DOT-T1E encodes three of these pumps. Both strains activate membrane modifications to reduce toluene membrane permeability. The KT2440 strain activates the TCA cycle to generate energy, while avoiding energy-intensive processes such as flagellar biosynthesis. This suggests that KT2440 responds to toluene by focusing on survival mechanisms. The DOT-T1E strain activates toluene degradation pathways, using toluene as source of energy. Among the unique genes encoded by DOT-T1E is a 70 kb island composed of genes of unknown function induced in response to toluene. © 2016 Society for Applied Microbiology and John Wiley & Sons Ltd.

  10. Metabolism of amino acid amides in Pseudomonas putida ATCC 12633

    NARCIS (Netherlands)

    Hermes, H.F.M.; Croes, L.M.; Peeters, W.P.H.; Peters, P.J.H.; Dijkhuizen, L.

    1993-01-01

    The metabolism of the natural amino acid L-valine, the unnatural amino acids D-valine, and D-, L-phenylglycine (D-, L-PG), and the unnatural amino acid amides D-, L-phenylglycine amide (D, L-PG-NH2) and L-valine amide (L-Val-NH2) was studied in Pseudomonas putida ATCC 12633. The organism possessed c

  11. Metabolism of amino acid amides in Pseudomonas putida ATCC 12633

    NARCIS (Netherlands)

    Hermes, H.F.M.; Croes, L.M.; Peeters, W.P.H.; Peters, P.J.H.; Dijkhuizen, L.

    1993-01-01

    The metabolism of the natural amino acid L-valine, the unnatural amino acids D-valine, and D-, L-phenylglycine (D-, L-PG), and the unnatural amino acid amides D-, L-phenylglycine amide (D, L-PG-NH2) and L-valine amide (L-Val-NH2) was studied in Pseudomonas putida ATCC 12633. The organism possessed

  12. A New Extant Respirometric Assay to Estimate Intrinsic Growth Parameters Applied to Study Plasmid Metabolic Burden

    DEFF Research Database (Denmark)

    Seoane, Jose Miguel; Sin, Gürkan; Lardon, Laurent

    2010-01-01

    Start-Lip phenomena in microbial biokinetic assays are not captured by the most commonly used growth-related equations. In this Study we propose a new respirometric experimental design to estimate intrinsic growth parameters that allow us to avoid these limitations without data omission, separate...... mathematical treatment, or wake-up pulses prior to the analysis.. Identifiability and sensitivity analysis were performed to confirm the robustness of the new approach for obtaining unique and accurate estimates of growth kinetic parameters. The new experimental design was applied to establish the. metabolic...... burden caused by the carriage of a pWW0 TOL plasmid in the model organism Pseudomonas putida KT2440; The metabolic,burden associated was manifested as a reduction in the yield and the specific growth rate of the host, with both plasmid maintenance and the over-expression of recombinant proteins from...

  13. A new P. putida instrumental toxicity bioassay.

    Science.gov (United States)

    Figueredo, Federico; Abrevaya, Ximena C; Cortón, Eduardo

    2015-05-01

    Here, we present a new toxicity bioassay (CO2-TOX), able to detect toxic or inhibitory compounds in water samples, based on the quantification of Pseudomonas putida KT2440 CO2 production. The metabolically produced CO2 was measured continuously and directly in the liquid assay media, with a potentiometric gas electrode. The optimization studies were performed using as a model toxicant 3,5-DCP (3,5-dichlorophenol); later, heavy metals (Pb(2+), Cu(2+), or Zn(2+)) and a metalloid (As(5+)) were assayed. The response to toxics was evident after 15 min of incubation and at relatively low concentrations (e.g., 1.1 mg/L of 3,5-DCP), showing that the CO2-TOX bioassay is fast and sensitive. The EC50 values obtained were 4.93, 0.12, 6.05, 32.17, and 37.81 mg/L for 3,5-DCP, Cu(2+), Zn(2+), As(5+), and Pb(2+), respectively, at neutral pH. Additionally, the effect of the pH of the sample and the use of lyophilized bacteria were also analyzed showing that the bioassay can be implemented in different conditions. Moreover, highly turbid samples and samples with very low oxygen levels were measured successfully with the new instrumental bioassay described here. Finally, simulated samples containing 3,5-DCP or a heavy metal mixture were tested using the proposed bioassay and a standard ISO bioassay, showing that our test is more sensible to the phenol but less sensible to the metal mixtures. Therefore, we propose CO2-TOX as a rapid, sensitive, low-cost, and robust instrumental bioassay that could perform as an industrial wastewater-process monitor among other applications.

  14. A new method for determining the metabolic activity of specific bacterial populations in soil using tritiated leucine and immunomagnetic separation

    DEFF Research Database (Denmark)

    Sengeløv, Gitte; Sørensen, Søren Johannes; Frette, Lone

    2000-01-01

    . The method was optimized using Pseudomonas putida KT2440¿:¿:Tc+/TOL::gfp inoculated into soil microcosms. Inoculated soil samples were incubated with [3H]-Leu followed by an immunomagnetic separation to recover the target bacteria. Radiolabel incorporated by the target bacteria was then measured. Incubation......A new assay, using immunomagnetic separation and uptake of tritiated leucine ([3H]-Leu), was developed for measuring the in situ metabolic activity of specific bacterial populations in soil. Such assays are needed to assess the role individual species play in diverse microbial soil communities...... time with immunomagnetic beads was not critical for optimal target cell recovery, but samples needed to be washed at least 5¿times during the immunomagnetic separation to reduce unspecific binding of the indigenous soil bacteria to the magnetic beads. Soil absorption of the polyclonal antibody further...

  15. The Ssr protein (T1E_1405) from Pseudomonas putida DOT-T1E enables oligonucleotide-based recombineering in platform strain P. putida EM42

    DEFF Research Database (Denmark)

    Aparicio, Tomás; Ingemann Jensen, Sheila; Nielsen, Alex Toftgaard

    2016-01-01

    of reference strain KT2440) is still a time-consuming endeavor. In this work we have investigated the in vivo activity of the Ssr protein encoded by the open reading frame T1E_1405 from Pseudomonas putida DOT-T1E, a plausible functional homologue of the β protein of the Red recombination system of λ phage...... of Escherichia coli. A test based on the phenotypes of pyrF mutants of P. putida (the yeast’s URA3 ortholog) was developed for quantifying the ability of Ssr to promote invasion of the genomic DNA replication fork by synthetic oligonucleotides. The efficiency of the process was measured by monitoring...

  16. Characterization of cell lysis in Pseudomonas putida induced upon expression of heterologous killing genes

    DEFF Research Database (Denmark)

    Ronchel, M.C.; Molina, L.; Witte, A.;

    1998-01-01

    Active biological containment systems are based on the controlled expression of killing genes. These systems are of interest for the Pseudomonadaceae because of the potential applications of these microbes as bioremediation agents and biopesticides, The physiological effects that lead to cell death...... upon the induction of expression of two different heterologous killing genes in nonpathogenic Pseudomonas putida KT2440 derivatives have been analyzed, P. putida CMC4 and CMC12 carry in their chromosomes a fusion of the PAl-04/03 promoter to the Escherichia coli gef gene and the phi X174 lysis gene E......, respectively. Expression of the killing genes is controlled by the LacI protein, whose expression is initiated from the XylS-dependent Pm promoter. Under induced conditions, killing of P. putida CMC12 cells mediated by phi X174 lysis protein E was faster than that observed for P. putida CMC4, for which the Gef...

  17. Benzoxazinoids in Root Exudates of Maize Attract Pseudomonas putida to the Rhizosphere

    OpenAIRE

    Neal, Andrew L.; Shakoor Ahmad; Ruth Gordon-Weeks; Jurriaan Ton

    2012-01-01

    Benzoxazinoids, such as 2,4-dihydroxy-7-methoxy-2H-1,4-benzoxazin-3(4H)-one (DIMBOA), are secondary metabolites in grasses. In addition to their function in plant defence against pests and diseases above-ground, benzoxazinoids (BXs) have also been implicated in defence below-ground, where they can exert allelochemical or antimicrobial activities. We have studied the impact of BXs on the interaction between maize and Pseudomonas putida KT2440, a competitive coloniser of the maize rhizosphere w...

  18. Genetic analysis of functions involved in adhesion of Pseudomonas putida to seeds.

    Science.gov (United States)

    Espinosa-Urgel, M; Salido, A; Ramos, J L

    2000-05-01

    Many agricultural uses of bacteria require the establishment of efficient bacterial populations in the rhizosphere, for which colonization of plant seeds often constitutes a critical first step. Pseudomonas putida KT2440 is a strain that colonizes the rhizosphere of a number of agronomically important plants at high population densities. To identify the functions involved in initial seed colonization by P. putida KT2440, we subjected this strain to transposon mutagenesis and screened for mutants defective in attachment to corn seeds. Eight different mutants were isolated and characterized. While all of them showed reduced attachment to seeds, only two had strong defects in their adhesion to abiotic surfaces (glass and different plastics). Sequences of the loci affected in all eight mutants were obtained. None of the isolated genes had previously been described in P. putida, although four of them showed clear similarities with genes of known functions in other organisms. They corresponded to putative surface and membrane proteins, including a calcium-binding protein, a hemolysin, a peptide transporter, and a potential multidrug efflux pump. One other showed limited similarities with surface proteins, while the remaining three presented no obvious similarities with known genes, indicating that this study has disclosed novel functions.

  19. In-silico-driven metabolic engineering of Pseudomonas putida for enhanced production of poly-hydroxyalkanoates

    NARCIS (Netherlands)

    Poblete-Castro, I.; Binger, D.; Rodrigues, A.; Becker, J.; Martins Dos Santos, V.A.P.; Wittmann, C.

    2013-01-01

    Here, we present systems metabolic engineering driven by in-silico modeling to tailor Pseudomonas putida for synthesis of medium chain length PHAs on glucose. Using physiological properties of the parent wild type as constraints, elementary flux mode analysis of a large-scale model of the metabolism

  20. In-silico-driven metabolic engineering of Pseudomonas putida for enhanced production of poly-hydroxyalkanoates

    NARCIS (Netherlands)

    Poblete-Castro, I.; Binger, D.; Rodrigues, A.; Becker, J.; Martins Dos Santos, V.A.P.; Wittmann, C.

    2013-01-01

    Here, we present systems metabolic engineering driven by in-silico modeling to tailor Pseudomonas putida for synthesis of medium chain length PHAs on glucose. Using physiological properties of the parent wild type as constraints, elementary flux mode analysis of a large-scale model of the metabolism

  1. Volatilization of arsenic from polluted soil by Pseudomonas putida engineered for expression of the arsM Arsenic(III) S-adenosine methyltransferase gene.

    Science.gov (United States)

    Chen, Jian; Sun, Guo-Xin; Wang, Xiao-Xue; Lorenzo, Víctor de; Rosen, Barry P; Zhu, Yong-Guan

    2014-09-02

    Even though arsenic is one of the most widespread environmental carcinogens, methods of remediation are still limited. In this report we demonstrate that a strain of Pseudomonas putida KT2440 endowed with chromosomal expression of the arsM gene encoding the As(III) S-adenosylmethionine (SAM) methyltransfase from Rhodopseudomonas palustris to remove arsenic from contaminated soil. We genetically engineered the P. putida KT2440 with stable expression of an arsM-gfp fusion gene (GE P. putida), which was inserted into the bacterial chromosome. GE P. putida showed high arsenic methylation and volatilization activity. When exposed to 25 μM arsenite or arsenate overnight, most inorganic arsenic was methylated to the less toxic methylated arsenicals methylarsenate (MAs(V)), dimethylarsenate (DMAs(V)) and trimethylarsine oxide (TMAs(V)O). Of total added arsenic, the species were about 62 ± 2.2% DMAs(V), 25 ± 1.4% MAs(V) and 10 ± 1.2% TMAs(V)O. Volatilized arsenicals were trapped, and the predominant species were dimethylarsine (Me2AsH) (21 ± 1.0%) and trimethylarsine (TMAs(III)) (10 ± 1.2%). At later times, more DMAs(V) and volatile species were produced. Volatilization of Me2AsH and TMAs(III) from contaminated soil is thus possible with this genetically engineered bacterium and could be instrumental as an agent for reducing the inorganic arsenic content of soil and agricultural products.

  2. Toxicity of graphene oxide on growth and metabolism of Pseudomonas putida.

    Science.gov (United States)

    Combarros, R G; Collado, S; Díaz, M

    2016-06-05

    The increasing consumption of graphene derivatives leads to greater presence of these materials in wastewater treatment plants and ecological systems. The toxicity effect of graphene oxide (GO) on the microbial functions involved in the biological wastewater treatment process is studied, using Pseudomonas putida and salicylic acid (SA) as bacterial and pollutant models. A multiparametric flow cytometry (FC) method has been developed to measure the metabolic activity and viability of P. putida in contact with GO. A continuous reduction in the percentages of viable cells and a slight increase, lower than 5%, in the percentages of damaged and dead cells, suggest that P. putida in contact with GO loses the membrane integrity but preserves metabolic activity. The growth of P. putida was strongly inhibited by GO, since 0.05mgmL(-1) of GO reduced the maximum growth by a third, and the inhibition was considerably greater for GO concentrations higher than 0.1mgmL(-1). The specific SA removal rate decreased with GO concentration up to 0.1mgmL(-1) indicating that while GO always reduces the growth of P. putida, for concentrations higher than 0.1mgmL(-1), it also reduces its activity. Similar behaviour is observed using simulated urban and industrial wastewaters, the observed effects being more acute in the industrial wastewaters.

  3. Small Rna Regulatory Networks In Pseudomonas Putida

    DEFF Research Database (Denmark)

    Bojanovic, Klara; Long, Katherine

    2015-01-01

    chemicals and has a potential to be used as an efficient cell factory for various products. P. putida KT2240 is a genome-sequenced strain and a well characterized pseudomonad. Our major aim is to identify small RNA molecules (sRNAs) and their regulatory networks. A previous study has identified 37 sRNAs...... in this strain, while in other pseudomonads many more sRNAs have been found so far.P. putida KT2440 has been grown in different conditions which are likely to be encountered in industrial fermentations with the aim of using sRNAs for generation of improved cell factories. For that, cells have been grown in LB...... and harvested in different growth phases, as well as osmotic, membrane and oxidative stress conditions. RNA sequencing data has been analysed with the open source software system Rockhopper, and it has revealed over 180 putative sRNAs. Most of them (86%) seem to be novel and uncharacterized. The majority...

  4. Efficient recombinant production of prodigiosin in Pseudomonas putida

    Directory of Open Access Journals (Sweden)

    Andreas eDomröse

    2015-09-01

    Full Text Available Serratia marcescens and several other bacteria produce the red-colored pigment prodigiosin which possesses bioactivities as an antimicrobial, anticancer and immunosuppressive agent. Therefore, there is a great interest to produce this natural compound. Efforts aiming at its biotechnological production have so far largely focused on the original producer and opportunistic human pathogen S. marcescens. Here, we demonstrate efficient prodigiosin production in the heterologous host Pseudomonas putida. Random chromosomal integration of the 21 kb prodigiosin biosynthesis gene cluster of S. marcescens in P. putida KT2440 was employed to construct constitutive prodigiosin production strains. Standard cultivation parameters were optimized such that titers of 94 mg/L culture were obtained upon growth of P. putida at 20 °C using rich medium under high aeration conditions. Subsequently, a novel, fast and effective protocol for prodigiosin extraction and purification was established enabling the straightforward isolation of prodigiosin from P. putida growth medium. In summary, we describe here a highly efficient method for the heterologous biosynthetic production of prodigiosin which may serve as a basis to produce large amounts of this bioactive natural compound and may provide a platform for further in-depth studies of prodiginine biosynthesis.

  5. Does doping with aluminum alter the effects of ZnO nanoparticles on the metabolism of soil pseudomonads?

    Science.gov (United States)

    Fang, Tommy; Watson, Jean-Luc; Goodman, Jordan; Dimkpa, Christian O; Martineau, Nicole; Das, Siddhartha; McLean, Joan E; Britt, David W; Anderson, Anne J

    2013-02-22

    Doping of ZnO nanoparticles (NPs) is being used to increase their commercialization in the optical and semiconductor fields. This paper addresses whether doping with Al alters how ZnO NPs at nonlethal levels modifies the metabolism of soil-borne pseudomonads which are beneficial in performing bioremediation or promoting plant growth. The differences in X-ray diffraction (XRD) patterns, observed between commercial ZnO and Al-doped ZnO NPs indicated the aluminum was present as Al NPs. Both particles aggregated in the bacterial growth medium and formed colloids of different surface charges. They had similar effects on bacterial metabolism: rapid, dose-dependent loss in light output indicative of temporary toxicity in a biosensor constructed in Pseudomonas putida KT2440; increased production of a fluorescent pyoverdine-type siderophore, and decreased levels of indole acetic acid and phenazines in Pseudomonas chlororaphis O6. Solubilization of Zn and Al from the NPs contributed to these responses to different extents. These findings indicate that Al-doping of the ZnO NPs did not reduce the ability of the NPs to alter bacterial metabolism in ways that could influence performance of the pseudomonads in their soil environment.

  6. Modeling the Differences in Biochemical Capabilities of Pseudomonas Species by Flux Balance Analysis: How Good Are Genome-Scale Metabolic Networks at Predicting the Differences?

    Directory of Open Access Journals (Sweden)

    Parizad Babaei

    2014-01-01

    Full Text Available To date, several genome-scale metabolic networks have been reconstructed. These models cover a wide range of organisms, from bacteria to human. Such models have provided us with a framework for systematic analysis of metabolism. However, little effort has been put towards comparing biochemical capabilities of closely related species using their metabolic models. The accuracy of a model is highly dependent on the reconstruction process, as some errors may be included in the model during reconstruction. In this study, we investigated the ability of three Pseudomonas metabolic models to predict the biochemical differences, namely, iMO1086, iJP962, and iSB1139, which are related to P. aeruginosa PAO1, P. putida KT2440, and P. fluorescens SBW25, respectively. We did a comprehensive literature search for previous works containing biochemically distinguishable traits over these species. Amongst more than 1700 articles, we chose a subset of them which included experimental results suitable for in silico simulation. By simulating the conditions provided in the actual biological experiment, we performed case-dependent tests to compare the in silico results to the biological ones. We found out that iMO1086 and iJP962 were able to predict the experimental data and were much more accurate than iSB1139.

  7. Metabolic engineering of Pseudomonas putida for production of docosahexaenoic acid based on a myxobacterial PUFA synthase.

    Science.gov (United States)

    Gemperlein, Katja; Zipf, Gregor; Bernauer, Hubert S; Müller, Rolf; Wenzel, Silke C

    2016-01-01

    Long-chain polyunsaturated fatty acids (LC-PUFAs) can be produced de novo via polyketide synthase-like enzymes known as PUFA synthases, which are encoded by pfa biosynthetic gene clusters originally discovered from marine microorganisms. Recently similar gene clusters were detected and characterized in terrestrial myxobacteria revealing several striking differences. As the identified myxobacterial producers are difficult to handle genetically and grow very slowly we aimed to establish heterologous expression platforms for myxobacterial PUFA synthases. Here we report the heterologous expression of the pfa gene cluster from Aetherobacter fasciculatus (SBSr002) in the phylogenetically distant model host bacteria Escherichia coli and Pseudomonas putida. The latter host turned out to be the more promising PUFA producer revealing higher production rates of n-6 docosapentaenoic acid (DPA) and docosahexaenoic acid (DHA). After several rounds of genetic engineering of expression plasmids combined with metabolic engineering of P. putida, DHA production yields were eventually increased more than threefold. Additionally, we applied synthetic biology approaches to redesign and construct artificial versions of the A. fasciculatus pfa gene cluster, which to the best of our knowledge represents the first example of a polyketide-like biosynthetic gene cluster modulated and synthesized for P. putida. Combination with the engineering efforts described above led to a further increase in LC-PUFA production yields. The established production platform based on synthetic DNA now sets the stage for flexible engineering of the complex PUFA synthase.

  8. Ultrasonic disruption of Pseudomonas putida for the release of arginine deiminase: Kinetics and predictive models.

    Science.gov (United States)

    Patil, Mahesh D; Dev, Manoj J; Tangadpalliwar, Sujit; Patel, Gopal; Garg, Prabha; Chisti, Yusuf; Banerjee, Uttam Chand

    2017-06-01

    The responses of the ultrasound-mediated disruption of Pseudomonas putida KT2440 were modelled as the function of biomass concentration in the cell suspension; the treatment time of sonication; the duty cycle and the acoustic power of the sonicator. For the experimental data, the response surface (RSM), the artificial neural network (ANN) and the support vector machine (SVM) models were compared for their ability to predict the performance parameters. The satisfactory prediction of the unseen data of the responses implied the proficient generalization capabilities of ANN. The extent of the cell disruption was mainly dependent on the acoustic power and the biomass concentration. The cellmass concentration in the slurry most strongly influenced the ADI and total protein release. Nearly 28U/mL ADI was released when a biomass concentration of 300g/L was sonicated for 6min with an acoustic power of 187.5W at 40% duty cycle. Cell disruption obeyed first-order kinetics.

  9. A sirA-like gene, sirA2, is essential for 3-succinoyl-pyridine metabolism in the newly isolated nicotine-degrading Pseudomonas sp. HZN6 strain.

    Science.gov (United States)

    Qiu, Jiguo; Ma, Yun; Chen, Liansheng; Wu, Lifei; Wen, Yuezhong; Liu, Weiping

    2011-12-01

    A novel nicotine-degrading Pseudomonas sp. strain, HZN6, was isolated from a pesticide-wastewater treatment facility in Hangzhou. The strain could grow on nicotine as its sole source of carbon, nitrogen, and energy. The strain's main intermediate metabolites were determined to be pseudooxynicotine, 3-succinoyl-pyridine (SP), and 6-hydroxy-3-succinoyl-pyridine (HSP). A Tn5 transposon mutant was generated in which the degradation pathway was blocked at the SP. A 4,583-bp DNA fragment flanking the transposon insertion site was obtained through self-formed adaptor PCR and analyzed. The mutant gene orfC displays 89% deduced amino acid sequence identity with the sirA-like gene (sirA2, a sulfurtransferase homologue gene) of Pseudomonas stutzeri A1501. The orfC-disrupted strain lost the ability to degrade SP, and the complementation strains with the orfC from the Pseudomonas sp. HZN6 and the sirA2 (PP_1233) from Pseudomonas putida KT2440 recovered the degradation ability. Though the orfC-disrupted strain also lost the xanthine dehydrogenase activity, the effects of tungsten on the degradation of SP and hypoxanthine revealed that the hydroxylation of SP to HSP was not a xanthine dehydrogenase type. These results demonstrated that the orfC gene was essential for the SP metabolism involved in the nicotine metabolic pathway in the Pseudomonas sp. HZN6 strain. This study might advance the understanding of the nicotine metabolic mechanism in Pseudomonas.

  10. The Role of CzcRS Two-Component Systems in the Heavy Metal Resistance of Pseudomonas putida X4

    Directory of Open Access Journals (Sweden)

    Pulin Liu

    2015-07-01

    Full Text Available The role of different czcRS genes in metal resistance and the cross-link between czcRS and czcCBA in Pseudomonas putida X4 were studied to advance understanding of the mechanisms by which P. putida copes with metal stress. Similar to P. putida KT2440, two complete czcRS1 and czcRS2 two-component systems, as well as a czcR3 without the corresponding sensing component were amplified in P. putida X4. The histidine kinase genes czcS1 and czcS2 were inactivated and fused to lacZ by homologous recombination. The lacZ fusion assay revealed that Cd2+ and Zn2+ caused a decrease in the transcription of czcRS1, whereas Cd2+ treatment enhanced the transcription of czcRS2. The mutation of different czcRSs showed that all czcRSs are necessary to facilitate full metal resistance in P. putida X4. A putative gene just downstream of czcR3 is related to metal ion resistance, and its transcription was activated by Zn2+. Data from quantitative real-time polymerase chain reaction (qRT-PCR strongly suggested that czcRSs regulate the expression of czcCBA, and a cross-link exists between different czcRSs.

  11. Fructose 1-phosphate is the preferred effector of the metabolic regulator Cra of Pseudomonas putida.

    Science.gov (United States)

    Chavarría, Max; Santiago, César; Platero, Raúl; Krell, Tino; Casasnovas, José M; de Lorenzo, Víctor

    2011-03-18

    The catabolite repressor/activator (Cra) protein is a global sensor and regulator of carbon fluxes through the central metabolic pathways of gram-negative bacteria. To examine the nature of the effector (or effectors) that signal such fluxes to the protein of Pseudomonas putida, the Cra factor of this soil microorganism has been purified and characterized and its three-dimensional structure determined. Analytical ultracentrifugation, gel filtration, and mobility shift assays showed that the effector-free Cra is a dimer that binds an operator DNA sequence in the promoter region of the fruBKA cluster. Furthermore, fructose 1-phosphate (F1P) was found to most efficiently dissociate the Cra-DNA complex. Thermodynamic parameters of the F1P-Cra-DNA interaction calculated by isothermal titration calorimetry revealed that the factor associates tightly to the DNA sequence 5'-TTAAACGTTTCA-3' (K(D) = 26.3 ± 3.1 nM) and that F1P binds the protein with an apparent stoichiometry of 1.06 ± 0.06 molecules per Cra monomer and a K(D) of 209 ± 20 nM. Other possible effectors, like fructose 1,6-bisphosphate, did not display a significant affinity for the regulator under the assay conditions. Moreover, the structure of Cra and its co-crystal with F1P at a 2-Å resolution revealed that F1P fits optimally the geometry of the effector pocket. Our results thus single out F1P as the preferred metabolic effector of the Cra protein of P. putida.

  12. Genome-wide mapping of transcription start sites yields novel insights into the primary transcriptome of Pseudomonas putida

    DEFF Research Database (Denmark)

    D'Arrigo, Isotta; Bojanovic, Klara; Yang, Xiaochen

    2016-01-01

    was examined using an in vivo assay with GFP-fusion vectors and shown to function via a translational repression mechanism. Furthermore, 56 novel intergenic small RNAs and 8 putative actuaton transcripts were detected, as well as 8 novel open reading frames (ORFs). This study illustrates how global mapping...... elements of P. putida strain KT2440. A total of 7937 putative transcription start sites (TSSs) were identified, where over two-thirds were located either on the opposite strand or internal to annotated genes. For TSSs associated with mRNAs, sequence analysis revealed a clear Shine–Dalgarno sequence...... but a lack of conserved overrepresented promoter motifs. These TSSs defined approximately 50 leaderless transcripts and an abundance of mRNAs with long leader regions of which 18 contain RNA regulatory elements from the Rfam database. The thiamine pyrophosphate riboswitch upstream of the thiC gene...

  13. Isolation of a strain of Pseudomonas putida capable of metabolizing anionic detergent sodium dodecyl sulfate (SDS

    Directory of Open Access Journals (Sweden)

    A Kumar

    2011-05-01

    Full Text Available Background and objectives: Sodium Dodecyl Sulfate (SDS is one of the most widely used anionic detergents. The present study deals with isolation and identification of SDS-degrading bacteria from a detergent contaminated pond situated in Varanasi city, India."nMaterials and Methods: Employing enrichment technique in minimal medium (PBM, SDS-degrading bacteria were isolated from pond water sample. Rate of degradation of SDS was studied in liquid PBM and also degradation of different concentrations of SDS was also studied to find out maximum concentration of SDS degraded by the potent isolates. Alkyl sulfatase activity (key enzyme in SDS degradation was estimated in crude cell extracts and multiplicity of alkyl sulfatase was studied by Native PAGE Zymography. The potent isolate was identified by 16S rRNA sequence analysis."nResults: Using enrichment technique in minimal medium containing SDS as a sole carbon source, initially three SDS degrading isolates were recovered. However, only one isolate, SP3, was found to be an efficient degrader of SDS. It was observed that this strain could completely metabolize 0.1% SDS in 16 h, 0.2% SDS in 20 h and 0.3% SDS in 24 h of incubation. Specific activity of alkyl sulfatase was 0.087±0.004 μmol SDS/mg protein/min and Native PAGE Zymography showed presence of alkyl sulfatase of Rf value of 0.21. This isolate was identified as Pseudomonas putida strain SP3."nConclusion: This is the report of isolation of SDS-degrading strain of P. putida, which shows high rate of SDS degradation and can degrade up to 0.3% SDS. It appears that this isolate can be exploited for bioremediation of this detergent from water systems.

  14. The sigma54 regulon (sigmulon) of Pseudomonas putida.

    Science.gov (United States)

    Cases, Ildefonso; Ussery, David W; de Lorenzo, Víctor

    2003-12-01

    sigma54 is unique among the bacterial sigma factors. Besides not being related in sequence with the rest of such factors, its mechanism of transcription initiation is completely different and requires the participation of a transcription activator. In addition, whereas the rest of the alternative sigma factors use to be involved in transcription of somehow related biological functions, this is not the case for sigma54 and many different and unrelated genes have been shown to be transcribed from sigma54-dependent promoters, ranging from flagellation, to utilization of several different carbon and nitrogen sources, or alginate biosynthesis. These genes have been characterized in many different bacterial species and, only until recently with the arrival of complete genome sequences, we have been able to look at the sigma54 functional role from a genomic perspective. Aided by computational methods, the sigma54 regulon has been studied both in Escherichia coli, Salmonella typhimurium and several species of the Rhizobiaceae. Here we present the analysis of the sigma54 regulon (sigmulon) in the complete genome of Pseudomonas putida KT2440. We have developed an improved method for the prediction of sigma54-dependent promoters which combines the scores of sigma54-RNAP target sequences and those of activator binding sites. In combination with other evidence obtained from the chromosomal context and the similarity with closely related bacteria, we have been able to predict more than 80% of the sigma54-dependent promoters of P. putida with high confidence. Our analysis has revealed new functions for sigma54 and, by means of comparative analysis with the previous studies, we have drawn a potential mechanism for the evolution of this regulatory system.

  15. Establishment of oxidative D-xylose metabolism in Pseudomonas putida S12

    NARCIS (Netherlands)

    Meijnen, J.P.; Winde, J.H.de; Ruijssenaars, H.J.

    2009-01-01

    The oxidative D-xylose catabolic pathway of Caulobacter crescentus, encoded by the xylXABCD operon, was expressed in the gram-negative bacterium Pseudomonas putida S12. This engineered transformant strain was able to grow on D-xylose as a sole carbon source with a biomass yield of 53% (based on g [d

  16. ToMI-FBA: A genome-scale metabolic flux based algorithm to select optimum hosts and media formulations for expressing pathways of interest

    Directory of Open Access Journals (Sweden)

    Hadi Nazem-Bokaee

    2015-09-01

    Full Text Available The Total Membrane Influx constrained Flux Balance Analysis (ToMI-FBA algorithm was developed in this research as a new tool to help researchers decide which microbial host and medium formulation are optimal for expressing a new metabolic pathway. ToMI-FBA relies on genome-scale metabolic flux modeling and a novel in silico cell membrane influx constraint that specifies the flux of atoms (not molecules into the cell through all possible membrane transporters. The ToMI constraint is constructed through the addition of an extra row and column to the stoichiometric matrix of a genome-scale metabolic flux model. In this research, the mathematical formulation of the ToMI constraint is given along with four case studies that demonstrate its usefulness. In Case Study 1, ToMI-FBA returned an optimal culture medium formulation for the production of isobutanol from Bacillus subtilis. Significant levels of L-valine were recommended to optimize production, and this result has been observed experimentally. In Case Study 2, it is demonstrated how the carbon to nitrogen uptake ratio can be specified as an additional ToMI-FBA constraint. This was investigated for maximizing medium chain length polyhydroxyalkanoates (mcl-PHA production from Pseudomonas putida KT2440. In Case Study 3, ToMI-FBA revealed a strategy of adding cellobiose as a means to increase ethanol selectivity during the stationary growth phase of Clostridium acetobutylicum ATCC 824. This strategy was also validated experimentally. Finally, in Case Study 4, B. subtilis was identified as a superior host to Escherichia coli, Saccharomyces cerevisiae, and Synechocystis PCC6803 for the production of artemisinate.

  17. New transposon tools tailored for metabolic engineering of Gram-negative microbial cell factories

    Directory of Open Access Journals (Sweden)

    Esteban eMartínez-García

    2014-10-01

    Full Text Available Re-programming microorganisms to modify their existing functions and/or to bestow bacteria with entirely new-to-Nature tasks have largely relied so far on specialized molecular biology tools. Such endeavors are not only relevant in the burgeoning metabolic engineering arena, but also instrumental to explore the functioning of complex regulatory networks from a fundamental point of view. À la carte modification of bacterial genomes thus calls for novel tools to make genetic manipulations easier. We propose the use of a series of new broad-host-range mini-Tn5 vectors, termed pBAMDs, for the delivery of gene(s into the chromosome of Gram-negative bacteria and for generating saturated mutagenesis libraries in gene function studies. These delivery vectors endow the user with the possibility of easy cloning and subsequent insertion of functional cargoes with three different antibiotic resistance markers (kanamycin, streptomycin, and gentamicin. After validating the pBAMD vectors in the environmental bacterium Pseudomonas putida KT2440, their use was also illustrated by inserting the entire poly(3-hydroxybutyrate (PHB synthesis pathway from Cupriavidus necator in the chromosome of a phosphotransacetylase mutant of Escherichia coli. PHB is a completely biodegradable polyester with a number of industrial applications that make it attractive as a potential replacement of oil-based plastics. The non-selective nature of chromosomal insertions of the biosynthetic genes was evidenced by a large landscape of PHB synthesis levels in independent clones. One clone was selected and further characterized as a microbial cell factory for PHB accumulation, and it achieved polymer accumulation levels comparable to those of a plasmid-bearing recombinant. Taken together, our results demonstrate that the new mini-Tn5 vectors can be used to confer interesting phenotypes in Gram-negative bacteria that would be very difficult to engineer through direct manipulation of the

  18. Metabolic flux analysis of a phenol producing mutant of Pseudomonas putida S12: Verification and complementation of hypotheses derived from transcriptomics

    NARCIS (Netherlands)

    Wierckx, N.; Ruijssenaars, H.J.; Winde, J.H.de; Schmid, A.; Blank, L.M.

    2009-01-01

    The physiological effects of genetic and transcriptional changes observed in a phenol producing mutant of the solvent-tolerant Pseudomonas putida S12 were assessed with metabolic flux analysis. The upregulation of a malate/lactate dehydrogenase encoding gene could be connected to a flux increase fro

  19. Environmental stress speeds up DNA replication in Pseudomonas putida in chemostat cultivations.

    Science.gov (United States)

    Lieder, Sarah; Jahn, Michael; Koepff, Joachim; Müller, Susann; Takors, Ralf

    2016-01-01

    Cellular response to different types of stress is the hallmark of the cell's strategy for survival. How organisms adjust their cell cycle dynamics to compensate for changes in environmental conditions is an important unanswered question in bacterial physiology. A cell using binary fission for reproduction passes through three stages during its cell cycle: a stage from cell birth to initiation of replication, a DNA replication phase and a period of cell division. We present a detailed analysis of durations of cell cycle phases, investigating their dynamics under environmental stress conditions. Applying continuous steady state cultivations (chemostats), the DNA content of a Pseudomonas putida KT2440 population was quantified with flow cytometry at distinct growth rates. Data-driven modeling revealed that under stress conditions, such as oxygen deprivation, solvent exposure and decreased iron availability, DNA replication was accelerated correlated to the severity of the imposed stress (up to 1.9-fold). Cells maintained constant growth rates by balancing the shortened replication phase with extended cell cycle phases before and after replication. Transcriptome data underpin the transcriptional upregulation of crucial genes of the replication machinery. Hence adaption of DNA replication speed appears to be an important strategy to withstand environmental stress.

  20. Use of response surface method for maximizing the production of arginine deiminase by Pseudomonas putida.

    Science.gov (United States)

    Patil, Mahesh D; Shinde, Kiran D; Patel, Gopal; Chisti, Yusuf; Banerjee, Uttam Chand

    2016-06-01

    Statistically designed experiments were used to optimize the production of arginine deiminase (ADI) by Pseudomonas putida KT2440 in batch culture. A Plackett-Burman design involving eleven factors showed that ADI production was most influenced by the initial pH and the initial concentrations of glucose and yeast extract. A central composite experimental design showed that the optimal values of these factors were 8.0, 10 g/L and 12.5 g/L, respectively. The other components of the optimal culture medium were bacto peptone 7.5 g/L, Triton X-100 0.30% (v/v), and arginine 3 g/L, for a culture temperature of 25 °C. Compared with the basal medium, the ADI activity in the optimized medium had nearly 4.5-fold increase (4.31 U/mL). The optimized medium was then used for a further study of ADI production in a 14 L stirred tank bioreactor. The agitation speed and the aeration rates were varied to determine suitable values of these variables.

  1. Use of response surface method for maximizing the production of arginine deiminase by Pseudomonas putida

    Directory of Open Access Journals (Sweden)

    Mahesh D. Patil

    2016-06-01

    Full Text Available Statistically designed experiments were used to optimize the production of arginine deiminase (ADI by Pseudomonas putida KT2440 in batch culture. A Plackett-Burman design involving eleven factors showed that ADI production was most influenced by the initial pH and the initial concentrations of glucose and yeast extract. A central composite experimental design showed that the optimal values of these factors were 8.0, 10 g/L and 12.5 g/L, respectively. The other components of the optimal culture medium were bacto peptone 7.5 g/L, Triton X–100 0.30% (v/v, and arginine 3 g/L, for a culture temperature of 25 °C. Compared with the basal medium, the ADI activity in the optimized medium had nearly 4.5-fold increase (4.31 U/mL. The optimized medium was then used for a further study of ADI production in a 14 L stirred tank bioreactor. The agitation speed and the aeration rates were varied to determine suitable values of these variables.

  2. Effect of silver nanoparticles on Pseudomonas putida biofilms at different stages of maturity.

    Science.gov (United States)

    Thuptimdang, Pumis; Limpiyakorn, Tawan; McEvoy, John; Prüß, Birgit M; Khan, Eakalak

    2015-06-15

    This study determined the effect of silver nanoparticles (AgNPs) on Pseudomonas putida KT2440 biofilms at different stages of maturity. Three biofilm stages (1-3, representing early to late stages of development) were identified from bacterial adenosine triphosphate (ATP) activity under static (96-well plate) and dynamic conditions (Center for Disease Control and Prevention biofilm reactor). Extracellular polymeric substance (EPS) levels, measured using crystal violet and total carbohydrate assays, and expression of the EPS-associated genes, csgA and alg8, supported the conclusion that biofilms at later stages were older than those at earlier stages. More mature biofilms (stages 2 and 3) showed little to no reduction in ATP activity following exposure to AgNPs. In contrast, the same treatment reduced ATP activity by more than 90% in the less mature stage 1 biofilms. Regardless of maturity, biofilms with EPS stripped off were more susceptible to AgNPs than controls with intact EPS, demonstrating that EPS is critical for biofilm tolerance of AgNPs. The findings from this study show that stage of maturity is an important factor to consider when studying effect of AgNPs on biofilms.

  3. Effect of silver nanoparticles on Pseudomonas putida biofilms at different stages of maturity

    Energy Technology Data Exchange (ETDEWEB)

    Thuptimdang, Pumis, E-mail: pumis.th@gmail.com [International Program in Hazardous Substance and Environmental Management, Graduate School, Chulalongkorn University, Bangkok 10330 (Thailand); Center of Excellence on Hazardous Substance Management, Bangkok 10330 (Thailand); Limpiyakorn, Tawan, E-mail: tawan.l@chula.ac.th [Center of Excellence on Hazardous Substance Management, Bangkok 10330 (Thailand); Department of Environmental Engineering, Chulalongkorn University, Bangkok 10330 (Thailand); Research Unit Control of Emerging Micropollutants in Environment, Chulalongkorn University, Bangkok 10330 (Thailand); McEvoy, John, E-mail: john.mcevoy@ndsu.edu [Department of Veterinary and Microbiological Sciences, North Dakota State University, Fargo, ND 58108 (United States); Prüß, Birgit M., E-mail: birgit.pruess@ndsu.edu [Department of Veterinary and Microbiological Sciences, North Dakota State University, Fargo, ND 58108 (United States); Khan, Eakalak, E-mail: eakalak.khan@ndsu.edu [Department of Civil and Environmental Engineering, North Dakota State University, Fargo, ND 58108 (United States)

    2015-06-15

    Highlights: • Biofilm stages in static batch conditions were similar to dynamic conditions. • Expression of csgA gene increased earlier than alg8 gene in biofilm maturation. • AgNPs had higher effect on less mature biofilms. • Removal of extracellular polymeric substance made biofilms susceptible to AgNPs. - Abstract: This study determined the effect of silver nanoparticles (AgNPs) on Pseudomonas putida KT2440 biofilms at different stages of maturity. Three biofilm stages (1–3, representing early to late stages of development) were identified from bacterial adenosine triphosphate (ATP) activity under static (96-well plate) and dynamic conditions (Center for Disease Control and Prevention biofilm reactor). Extracellular polymeric substance (EPS) levels, measured using crystal violet and total carbohydrate assays, and expression of the EPS-associated genes, csgA and alg8, supported the conclusion that biofilms at later stages were older than those at earlier stages. More mature biofilms (stages 2 and 3) showed little to no reduction in ATP activity following exposure to AgNPs. In contrast, the same treatment reduced ATP activity by more than 90% in the less mature stage 1 biofilms. Regardless of maturity, biofilms with EPS stripped off were more susceptible to AgNPs than controls with intact EPS, demonstrating that EPS is critical for biofilm tolerance of AgNPs. The findings from this study show that stage of maturity is an important factor to consider when studying effect of AgNPs on biofilms.

  4. Metabolic pathway involved in 2-methyl-6-ethylaniline degradation by Sphingobium sp. strain MEA3-1 and cloning of the novel flavin-dependent monooxygenase system meaBA.

    Science.gov (United States)

    Dong, Weiliang; Chen, Qiongzhen; Hou, Ying; Li, Shuhuan; Zhuang, Kai; Huang, Fei; Zhou, Jie; Li, Zhoukun; Wang, Jue; Fu, Lei; Zhang, Zhengguang; Huang, Yan; Wang, Fei; Cui, Zhongli

    2015-12-01

    2-Methyl-6-ethylaniline (MEA) is the main microbial degradation intermediate of the chloroacetanilide herbicides acetochlor and metolachlor. Sphingobium sp. strain MEA3-1 can utilize MEA and various alkyl-substituted aniline and phenol compounds as sole carbon and energy sources for growth. We isolated the mutant strain MEA3-1Mut, which converts MEA only to 2-methyl-6-ethyl-hydroquinone (MEHQ) and 2-methyl-6-ethyl-benzoquinone (MEBQ). MEA may be oxidized by the P450 monooxygenase system to 4-hydroxy-2-methyl-6-ethylaniline (4-OH-MEA), which can be hydrolytically spontaneously deaminated to MEBQ or MEHQ. The MEA microbial metabolic pathway was reconstituted based on the substrate spectra and identification of the intermediate metabolites in both the wild-type and mutant strains. Plasmidome sequencing indicated that both strains harbored 7 plasmids with sizes ranging from 6,108 bp to 287,745 bp. Among the 7 plasmids, 6 were identical, and pMEA02' in strain MEA3-1Mut lost a 37,000-bp fragment compared to pMEA02 in strain MEA3-1. Two-dimensional electrophoresis (2-DE) and protein mass fingerprinting (PMF) showed that MEA3-1Mut lost the two-component flavin-dependent monooxygenase (TC-FDM) MeaBA, which was encoded by a gene in the lost fragment of pMEA02. MeaA shared 22% to 25% amino acid sequence identity with oxygenase components of some TC-FDMs, whereas MeaB showed no sequence identity with the reductase components of those TC-FDMs. Complementation with meaBA in MEA3-1Mut and heterologous expression in Pseudomonas putida strain KT2440 resulted in the production of an active MEHQ monooxygenase.

  5. Quantitative 'Omics Analyses of Medium Chain Length Polyhydroxyalkanaote Metabolism in Pseudomonas putida LS46 Cultured with Waste Glycerol and Waste Fatty Acids.

    Directory of Open Access Journals (Sweden)

    Jilagamazhi Fu

    Full Text Available Transcriptomes and proteomes of Pseudomonas putida LS46 cultured with biodiesel-derived waste glycerol or waste free fatty acids, as sole carbon sources, were compared under conditions that were either permissive or non-permissive for synthesis of medium chain length polyhydroxyalkanoates (mcl-PHA. The objectives of this study were to elucidate mechanisms that influence activation of biopolymer synthesis, intra-cellular accumulation, and monomer composition, and determine if these were physiologically specific to the carbon sources used for growth of P. putida LS46. Active mcl-PHA synthesis by P. putida LS46 was associated with high expression levels of key mcl-PHA biosynthesis genes and/or gene products including monomer-supplying proteins, PHA synthases, and granule-associated proteins. 'Omics data suggested that expression of these genes were regulated by different genetic mechanisms in P. putida LS46 cells in different physiological states, when cultured on the two waste carbon sources. Optimal polymer production by P. putida LS46 was primarily limited by less efficient glycerol metabolism during mcl-PHA synthesis on waste glycerol. Mapping the 'Omics data to the mcl-PHA biosynthetic pathway revealed significant variations in gene expression, primarily involved in: 1 glycerol transportation; 2 enzymatic reactions that recycle reducing equivalents and produce key mcl-PHA biosynthesis pathway intermediates (e.g. NADH/NADPH, acetyl-CoA. Active synthesis of mcl-PHAs was observed during exponential phase in cultures with waste free fatty acids, and was associated with the fatty acid beta-oxidation pathway. A putative Thioesterase in the beta-oxidation pathway that may regulate the level of fatty acid beta-oxidation intermediates, and thus carbon flux to mcl-PHA biosynthesis, was highly up-regulated. Finally, the data suggested that differences in expression of selected fatty acid metabolism and mcl-PHA monomer-supplying enzymes may play a role in

  6. Competition triggers plasmid-mediated enhancement of substrate utilisation in Pseudomonas putida.

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    Hiren Joshi

    Full Text Available Competition between species plays a central role in the activity and structure of communities. Stable co-existence of diverse organisms in communities is thought to be fostered by individual tradeoffs and optimization of competitive strategies along resource gradients. Outside the laboratory, microbes exist as multispecies consortia, continuously interacting with one another and the environment. Survival and proliferation of a particular species is governed by its competitive fitness. Therefore, bacteria must be able to continuously sense their immediate environs for presence of competitors and prevailing conditions. Here we present results of our investigations on a novel competition sensing mechanism in the rhizosphere-inhabiting Pseudomonas putida KT2440, harbouring gfpmut3b-modified Kan(R TOL plasmid. We monitored benzyl alcohol (BA degradation rate, along with GFP expression profiling in mono species and dual species cultures. Interestingly, enhanced plasmid expression (monitored using GFP expression and consequent BA degradation were observed in dual species consortia, irrespective of whether the competitor was a BA degrader (Pseudomonas aeruginosa or a non-degrader (E. coli. Attempts at elucidation of the mechanistic aspects of induction indicated the role of physical interaction, but not of any diffusible compounds emanating from the competitors. This contention is supported by the observation that greater induction took place in presence of increasing number of competitors. Inert microspheres mimicking competitor cell size and concentration did not elicit any significant induction, further suggesting the role of physical cell-cell interaction. Furthermore, it was also established that cell wall compromised competitor had minimal induction capability. We conclude that P. putida harbouring pWW0 experience a competitive stress when grown as dual-species consortium, irrespective of the counterpart being BA degrader or not. The immediate

  7. Elucidation of the flavonoid catabolism pathway in Pseudomonas putida PML2 by comparative metabolic profiling.

    Science.gov (United States)

    Pillai, Bhinu V S; Swarup, Sanjay

    2002-01-01

    Flavonoids are 15-carbon plant secondary metabolites exuded in the rhizosphere that hosts several flavonoid-degrading bacteria. We studied flavonoid catabolism in a plant growth-promoting rhizobacterial strain of Pseudomonas by using a combination of biochemical and genetic approaches. Transposants carrying mini-Tn5gfp insertions were screened for flavonoid auxotrophy, and these mutant strains were found to be unable to grow in the flavonols naringenin and quercetin, while their growth in glycerol was comparable to that of the parental strain. In order to understand flavonoid catabolism, culture supernatants, whole-cell fractions, cell lysate, and cell debris of the wild-type and mutant strains were analyzed. Intermediates that accumulated intracellularly and those secreted in the medium were identified by a combination of reversed-phase high-pressure liquid chromatography and electrospray ionization-mass spectrometry. Structures of four key intermediates were confirmed by one-dimensional nuclear magnetic resonance spectroscopy. Comparative metabolic profiling of the compounds in the wild-type and mutant strains allowed us to understand the degradation events and to identify six metabolic intermediates. The first step in the pathway involves 3,3'-didehydroxylation, followed by hydrolysis and cleavage of the C-ring, leading via subsequent oxidations to the formation of protocatechuate. This is the first report on quercetin dehydroxylation in aerobic conditions leading to naringenin accumulation.

  8. Metabolic Fingerprinting of Pseudomonas putida DOT-T1E Strains: Understanding the Influence of Divalent Cations in Adaptation Mechanisms Following Exposure to Toluene

    Directory of Open Access Journals (Sweden)

    Ali Sayqal

    2016-04-01

    Full Text Available Pseudomonas putida strains can adapt and overcome the activity of toxic organic solvents by the employment of several resistant mechanisms including efflux pumps and modification to lipopolysaccharides (LPS in their membranes. Divalent cations such as magnesium and calcium play a crucial role in the development of solvent tolerance in bacterial cells. Here, we have used Fourier transform infrared (FT-IR spectroscopy directly on cells (metabolic fingerprinting to monitor bacterial response to the absence and presence of toluene, along with the influence of divalent cations present in the growth media. Multivariate analysis of the data using principal component-discriminant function analysis (PC-DFA showed trends in scores plots, illustrating phenotypic alterations related to the effect of Mg2+, Ca2+ and toluene on cultures. Inspection of PC-DFA loadings plots revealed that several IR spectral regions including lipids, proteins and polysaccharides contribute to the separation in PC-DFA space, thereby indicating large phenotypic response to toluene and these cations. Finally, the saturated fatty acid ratio from the FT-IR spectra showed that upon toluene exposure, the saturated fatty acid ratio was reduced, while it increased in the presence of divalent cations. This study clearly demonstrates that the combination of metabolic fingerprinting with appropriate chemometric analysis can result in practicable knowledge on the responses of important environmental bacteria to external stress from pollutants such as highly toxic organic solvents, and indicates that these changes are manifest in the bacterial cell membrane. Finally, we demonstrate that divalent cations improve solvent tolerance in P. putida DOT‑T1E strains.

  9. The metabolic response of P. putida KT2442 producing high levels of polyhydroxyalkanoate under single- and multiple-nutrient-limited growth: Highlights from a multi-level omics approach

    Directory of Open Access Journals (Sweden)

    Poblete-Castro Ignacio

    2012-03-01

    Full Text Available Abstract Background Pseudomonas putida KT2442 is a natural producer of polyhydroxyalkanoates (PHAs, which can substitute petroleum-based non-renewable plastics and form the basis for the production of tailor-made biopolymers. However, despite the substantial body of work on PHA production by P. putida strains, it is not yet clear how the bacterium re-arranges its whole metabolism when it senses the limitation of nitrogen and the excess of fatty acids as carbon source, to result in a large accumulation of PHAs within the cell. In the present study we investigated the metabolic response of KT2442 using a systems biology approach to highlight the differences between single- and multiple-nutrient-limited growth in chemostat cultures. Results We found that 26, 62, and 81% of the cell dry weight consist of PHA under conditions of carbon, dual, and nitrogen limitation, respectively. Under nitrogen limitation a specific PHA production rate of 0.43 (g·(g·h-1 was obtained. The residual biomass was not constant for dual- and strict nitrogen-limiting growth, showing a different feature in comparison to other P. putida strains. Dual limitation resulted in patterns of gene expression, protein level, and metabolite concentrations that substantially differ from those observed under exclusive carbon or nitrogen limitation. The most pronounced differences were found in the energy metabolism, fatty acid metabolism, as well as stress proteins and enzymes belonging to the transport system. Conclusion This is the first study where the interrelationship between nutrient limitations and PHA synthesis has been investigated under well-controlled conditions using a system level approach. The knowledge generated will be of great assistance for the development of bioprocesses and further metabolic engineering work in this versatile organism to both enhance and diversify the industrial production of PHAs.

  10. Disruption of Pseudomonas putida by high pressure homogenization: a comparison of the predictive capacity of three process models for the efficient release of arginine deiminase.

    Science.gov (United States)

    Patil, Mahesh D; Patel, Gopal; Surywanshi, Balaji; Shaikh, Naeem; Garg, Prabha; Chisti, Yusuf; Banerjee, Uttam Chand

    2016-12-01

    Disruption of Pseudomonas putida KT2440 by high-pressure homogenization in a French press is discussed for the release of arginine deiminase (ADI). The enzyme release response of the disruption process was modelled for the experimental factors of biomass concentration in the broth being disrupted, the homogenization pressure and the number of passes of the cell slurry through the homogenizer. For the same data, the response surface method (RSM), the artificial neural network (ANN) and the support vector machine (SVM) models were compared for their ability to predict the performance parameters of the cell disruption. The ANN model proved to be best for predicting the ADI release. The fractional disruption of the cells was best modelled by the RSM. The fraction of the cells disrupted depended mainly on the operating pressure of the homogenizer. The concentration of the biomass in the slurry was the most influential factor in determining the total protein release. Nearly 27 U/mL of ADI was released within a single pass from slurry with a biomass concentration of 260 g/L at an operating pressure of 510 bar. Using a biomass concentration of 100 g/L, the ADI release by French press was 2.7-fold greater than in a conventional high-speed bead mill. In the French press, the total protein release was 5.8-fold more than in the bead mill. The statistical analysis of the completely unseen data exhibited ANN and SVM modelling as proficient alternatives to RSM for the prediction and generalization of the cell disruption process in French press.

  11. Analysis of the core genome and pangenome of Pseudomonas putida.

    Science.gov (United States)

    Udaondo, Zulema; Molina, Lázaro; Segura, Ana; Duque, Estrella; Ramos, Juan L

    2016-10-01

    Pseudomonas putida are strict aerobes that proliferate in a range of temperate niches and are of interest for environmental applications due to their capacity to degrade pollutants and ability to promote plant growth. Furthermore solvent-tolerant strains are useful for biosynthesis of added-value chemicals. We present a comprehensive comparative analysis of nine strains and the first characterization of the Pseudomonas putida pangenome. The core genome of P. putida comprises approximately 3386 genes. The most abundant genes within the core genome are those that encode nutrient transporters. Other conserved genes include those for central carbon metabolism through the Entner-Doudoroff pathway, the pentose phosphate cycle, arginine and proline metabolism, and pathways for degradation of aromatic chemicals. Genes that encode transporters, enzymes and regulators for amino acid metabolism (synthesis and degradation) are all part of the core genome, as well as various electron transporters, which enable aerobic metabolism under different oxygen regimes. Within the core genome are 30 genes for flagella biosynthesis and 12 key genes for biofilm formation. Pseudomonas putida strains share 85% of the coding regions with Pseudomonas aeruginosa; however, in P. putida, virulence factors such as exotoxins and type III secretion systems are absent.

  12. The metabolic response of P. putida KT2442 producing high levels of polyhydroxyalkanoate under single- and multiple-nutrient-limited growth: Highlights from a multi-level omics approach

    NARCIS (Netherlands)

    Poblete-Castro, I.; Escapa, I.; Jager, C.; Puchalka, J.; Lam, M.C.; Schomburg, D.; Prieto, M.; Martins Dos Santos, V.A.P.

    2012-01-01

    Background - Pseudomonas putida KT2442 is a natural producer of polyhydroxyalkanoates (PHAs), which can substitute petroleum-based non-renewable plastics and form the basis for the production of tailor-made biopolymers. However, despite the substantial body of work on PHA production by P. putida str

  13. The metabolic response of P. putida KT2442 producing high levels of polyhydroxyalkanoate under single- and multiple-nutrient-limited growth: Highlights from a multi-level omics approach

    NARCIS (Netherlands)

    Poblete-Castro, I.; Escapa, I.; Jager, C.; Puchalka, J.; Lam, M.C.; Schomburg, D.; Prieto, M.; Martins Dos Santos, V.A.P.

    2012-01-01

    Background - Pseudomonas putida KT2442 is a natural producer of polyhydroxyalkanoates (PHAs), which can substitute petroleum-based non-renewable plastics and form the basis for the production of tailor-made biopolymers. However, despite the substantial body of work on PHA production by P. putida str

  14. Comparative genomics of an endophytic Pseudomonas putida isolated from mango orchard

    Directory of Open Access Journals (Sweden)

    Huma Asif

    Full Text Available Abstract We analyzed the genome sequence of an endophytic bacterial strain Pseudomonas putida TJI51 isolated from mango bark tissues. Next generation DNA sequencing and short read de novo assembly generated the 5,805,096 bp draft genome of P. putida TJI51. Out of 6,036 protein coding genes in P. putida TJI51 sequences, 4,367 (72% were annotated with functional specifications, while the remaining encoded hypothetical proteins. Comparative genome sequence analysis revealed that the P. putida TJI51genome contains several regions, not identified in so far sequenced P. putida genomes. Some of these regions were predicted to encode enzymes, including acetylornithine deacetylase, betaine aldehyde dehydrogenase, aldehyde dehydrogenase, benzoylformate decarboxylase, hydroxyacylglutathione hydrolase, and uroporphyrinogen decarboxylase. The genome of P. putida TJI51 contained three nonribosomal peptide synthetase gene clusters. Genome sequence analysis of P. putidaTJI51 identified this bacterium as an endophytic resident. The endophytic fitness might be linked with alginate, which facilitates bacterial colonization in plant tissues. Genome sequence analysis shed light on the presence of a diverse spectrum of metabolic activities and adaptation of this isolate to various niches.

  15. The logic layout of the TOL network of Pseudomonas putida pWW0 plasmid stems from a metabolic amplifier motif (MAM that optimizes biodegradation of m-xylene

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    Silva-Rocha Rafael

    2011-11-01

    Full Text Available Abstract Background The genetic network of the TOL plasmid pWW0 of the soil bacterium Pseudomonas putida mt-2 for catabolism of m-xylene is an archetypal model for environmental biodegradation of aromatic pollutants. Although nearly every metabolic and transcriptional component of this regulatory system is known to an extraordinary molecular detail, the complexity of its architecture is still perplexing. To gain an insight into the inner layout of this network a logic model of the TOL system was implemented, simulated and experimentally validated. This analysis made sense of the specific regulatory topology out on the basis of an unprecedented network motif around which the entire genetic circuit for m-xylene catabolism gravitates. Results The most salient feature of the whole TOL regulatory network is the control exerted by two distinct but still intertwined regulators (XylR and XylS on expression of two separated catabolic operons (upper and lower for catabolism of m-xylene. Following model reduction, a minimal modular circuit composed by five basic variables appeared to suffice for fully describing the operation of the entire system. In silico simulation of the effect of various perturbations were compared with experimental data in which specific portions of the network were activated with selected inducers: m-xylene, o-xylene, 3-methylbenzylalcohol and 3-methylbenzoate. The results accredited the ability of the model to faithfully describe network dynamics. This analysis revealed that the entire regulatory structure of the TOL system enables the action an unprecedented metabolic amplifier motif (MAM. This motif synchronizes expression of the upper and lower portions of a very long metabolic system when cells face the head pathway substrate, m-xylene. Conclusion Logic modeling of the TOL circuit accounted for the intricate regulatory topology of this otherwise simple metabolic device. The found MAM appears to ensure a simultaneous expression

  16. Improved production of medium-chain-length Polyhydroxyalkanotes in glucose-based fed-batch cultivations of metabolically engineered Pseudomonas putida strains

    NARCIS (Netherlands)

    Poblete-Castro, I.; Rodriguez, A.L.; Lam, M.C.; Kessler, W.

    2014-01-01

    One of the major challenges in metabolic engineering for enhanced synthesis of value-added chemicals is to design and develop new strains which can be translated into well-controlled fermentation processes using bioreactors. The aim of this study was to assess the influence of various fed-batch stra

  17. The role of the Crc/Hfq/CrcZ-CrcY global regulatory system on the regulation of metabolic and cellular processes in Pseudomonas putida

    OpenAIRE

    La Rosa, Ruggero

    2015-01-01

    Tesis doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 28-09-2015 Metabolically versatile bacteria have evolved diverse strategies to adapt and to colonize different environments and niches. This ensures high fitness and the ability to withstand changing surrounding conditions and diverse kinds of stresses. These abilities rely on a varied genetic repertoire and on complex regulatory networks t...

  18. Redundancy in putrescine catabolism in solvent tolerant Pseudomonas putida S12.

    Science.gov (United States)

    Bandounas, Luaine; Ballerstedt, Hendrik; de Winde, Johannes H; Ruijssenaars, Harald J

    2011-06-10

    Pseudomonas putida S12 is a promising platform organism for the biological production of substituted aromatic compounds due to its extreme tolerance towards toxic chemicals. Solvent or aromatic stress tolerance may be due to membrane modifications and efflux pumps; however in general, polyamines have also been implicated in stressed cells. Previous transcriptomics results of P. putida strains producing an aromatic compound, or being exposed to the solvent toluene, indicated differentially expressed genes involved in polyamine transport and metabolism. Therefore, the metabolism of the polyamine, putrescine was investigated in P. putida S12, as no putrescine degradation pathways have been described for this strain. Via transcriptome analysis various, often redundant, putrescine-induced genes were identified as being potentially involved in putrescine catabolism via oxidative deamination and transamination. A series of knockout mutants were constructed in which up to six of these genes were sequentially deleted, and although putrescine degradation was affected in some of these mutants, complete elimination of putrescine degradation in P. putida S12 was not achieved. Evidence was found for the presence of an alternative pathway for putrescine degradation involving γ-glutamylation. The occurrence of multiple putrescine degradation routes in the solvent-tolerant P. putida S12 is indicative of the importance of controlling polyamine homeostasis, as well as of the high metabolic flexibility exhibited by this microorganism.

  19. Degradation of phenanthrene and pyrene using genetically engineered dioxygenase producing Pseudomonas putida in soil

    Directory of Open Access Journals (Sweden)

    Mardani Gashtasb

    2016-01-01

    Full Text Available Bioremediation use to promote degradation and/or removal of contaminants into nonhazardous or less-hazardous substances from the environment using microbial metabolic ability. Pseudomonas spp. is one of saprotrophic soil bacterium and can be used for biodegradation of polycyclic aromatic hydrocarbons (PAHs but this activity in most species is weak. Phenanthrene and pyrene could associate with a risk of human cancer development in exposed individuals. The aim of the present study was application of genetically engineered P. putida that produce dioxygenase for degradation of phenanthrene and pyrene in spiked soil using high-performance liquid chromatography (HPLC method. The nahH gene that encoded catechol 2,3-dioxygenase (C23O was cloned into pUC18 and pUC18-nahH recombinant vector was generated and transformed into wild P. putida, successfully. The genetically modified and wild types of P. putida were inoculated in soil and pilot plan was prepared. Finally, degradation of phenanthrene and pyrene by this bacterium in spiked soil were evaluated using HPLC measurement technique. The results were showed elimination of these PAH compounds in spiked soil by engineered P. putida comparing to dishes containing natural soil with normal microbial flora and inoculated autoclaved soil by wild type of P. putida were statistically significant (p0.05 but it was few impact on this process (more than 2%. Additional and verification tests including catalase, oxidase and PCR on isolated bacteria from spiked soil were indicated that engineered P. putida was alive and functional as well as it can affect on phenanthrene and pyrene degradation via nahH gene producing. These findings indicated that genetically engineered P. putida generated in this work via producing C23O enzyme can useful and practical for biodegradation of phenanthrene and pyrene as well as petroleum compounds in polluted environments.

  20. AcEST: BP915128 [AcEST

    Lifescience Database Archive (English)

    Full Text Available lear receptor subfamily 4 group A membe... 29 8.4 sp|Q88I82|DNAE2_PSEPK Error-prone DNA polymerase OS=Pseudo...9 >sp|Q88I82|DNAE2_PSEPK Error-prone DNA polymerase OS=Pseudomonas putida (strain KT2440) GN=dnaE2 PE=3 SV=1

  1. [Mechanism of cyanide and thiocyanate decomposition by an association of Pseudomonas putida and Pseudomonas stutzeri strains].

    Science.gov (United States)

    Grigor'eva, N V; Kondrat'eva, T F; Krasil'nikova, E N; Karavaĭko, G I

    2006-01-01

    The intermediate and terminal products of cyanide and thiocyanate decomposition by individual strains of the genus Pseudomonas, P. putida strain 21 and P. stutzeri strain 18, and by their association were analyzed. The activity of the enzymes of nitrogen and sulfur metabolism in these strains was compared with that of the collection strains P. putida VKM B-2187T and P. stutzeri VKM B-975T. Upon the introduction of CN- and SCN- into cell suspensions of strains 18 and 21 in phosphate buffer (pH 8.8), the production of NH4+ was observed. Due to the high rate of their utilization, NH3, NH4+, and CNO- were absent from the culture liquids of P. putida strain 21 and P. stutzeri strain 18 grown with CN- or SCN-. Both Pseudomonas strains decomposed SCN- via cyanate production. The cyanase activity was 0.75 micromol/(min mg protein) for P. putida strain 21 and 1.26 micromol/(min mg protein) for P. stutzeri strain 18. The cyanase activity was present in the cells grown with SCN- but absent in cells grown with NH4+. Strain 21 of P. putida was a more active CN- decomposer than strain 18 of P. stutzeri. Ammonium and CO2 were the terminal nitrogen and carbon products of CN- and SCN- decomposition. The terminal sulfur products of SCN- decomposition by P. stutzeri strain 18 and P. putida strain 21 were thiosulfate and tetrathionate, respectively. The strains utilized the toxic compounds in the anabolism only, as sources of nitrogen (CN- and SCN-) and sulfur (SCN-). The pathway of thiocyanate decomposition by the association of bacteria of the genus Pseudomonas is proposed based on the results obtained.

  2. A Two-Component Regulatory System Integrates Redox State and Population Density Sensing in Pseudomonas putida▿ †

    OpenAIRE

    Fernández-Piñar, Regina; Ramos, Juan Luis; Rodríguez-Herva, José Juan; Espinosa-Urgel, Manuel

    2008-01-01

    A two-component system formed by a sensor histidine kinase and a response regulator has been identified as an element participating in cell density signal transduction in Pseudomonas putida KT2440. It is a homolog of the Pseudomonas aeruginosa RoxS/RoxR system, which in turn belongs to the RegA/RegB family, described in photosynthetic bacteria as a key regulatory element. In KT2440, the two components are encoded by PP_0887 (roxS) and PP_0888 (roxR), which are transcribed in a single unit. Ch...

  3. Metal Inhibition of Growth and Manganese Oxidation in Pseudomonas putida GB-1

    Science.gov (United States)

    Pena, J.; Sposito, G.

    2009-12-01

    Biogenic manganese oxides (MnO2) are ubiquitous nanoparticulate minerals that contribute to the adsorption of nutrient and toxicant metals, the oxidative degradation of various organic compounds, and the respiration of metal-reducing bacteria in aquatic and terrestrial environments. The formation of these minerals is catalyzed by a diverse and widely-distributed group of bacteria and fungi, often through the enzymatic oxidation of aqueous Mn(II) to Mn(IV). In metal-impacted ecosystems, toxicant metals may alter the viability and metabolic activity of Mn-oxidizing organisms, thereby limiting the conditions under which biogenic MnO2 can form and diminishing their potential as adsorbent materials. Pseudomonas putida GB-1 (P. putida GB-1) is a model Mn-oxidizing laboratory culture representative of freshwater and soil biofilm-forming bacteria. Manganese oxidation in P. putida GB-1 occurs via two single-electron-transfer reactions, involving a multicopper oxidase enzyme found on the bacterial outer membrane surface. Near the onset of the stationary phase of growth, dark brown MnO2 particles are deposited in a matrix of bacterial cells and extracellular polymeric substances, thus forming heterogeneous biomineral assemblages. In this study, we assessed the influence of various transition metals on microbial growth and manganese oxidation capacity in a P. putida GB-1 culture propagated in a nutrient-rich growth medium. The concentration-response behavior of actively growing P. putida GB-1 cells was investigated for Fe, Co, Ni, Cu and Zn at pH ≈ 6 in the presence and absence of 1 mM Mn. Toxicity parameters such as EC0, EC50 and Hillslope, and EC100 were obtained from the sigmoidal concentration-response curves. The extent of MnO2 formation in the presence of the various metal cations was documented 24, 50, 74 and 104 h after the metal-amended medium was inoculated. Toxicity values were compared to twelve physicochemical properties of the metals tested. Significant

  4. Benzene, toluene and xylene biodegradation by Pseudomonas putida CCMI 852 Biodegradação de benzeno, tolueno e xileno pela Pseudomonas putida CCMI 852

    Directory of Open Access Journals (Sweden)

    Marcelo Henrique Otenio

    2005-09-01

    Full Text Available A minimal liquid medium containing benzene (B, toluene (T and xylene (X and mixtures thereof, was used to evaluate degradation activity of Pseudomonas putida CCMI 852 containing a TOL plasmid. Experiments were developed with B, T and X (100 mg L-1, with mixtures of BT, BX, and TX (50 + 50 mg L-1 each and BTX (33.3 + 33.3 + 33.3 mg L-1 each, added to 500 mL of medium. After 18 to 24 hours, the inoculum was added and solvent disappearance was determined after 24 to 25 hours by GC. Results showed that P. putida CCMI 852 was able to metabolize T and X, but B was not metabolized. In a BTX mixture, B was not metabolized and T and X degradation rate decreased 50%.Meio mineral líquido contendo benzeno (B ou tolueno (T ou xileno (X a 100 mg L-1 e suas misturas de BT, BX e TX (50 + 50 mg L-1 cada mistura e BTX (33,3 + 33,3 + 33,3 mg L-1 cada mistura foram utilizados para avaliar a atividade de degradação de B, T e X por Pseudomonas putida CCMI 852 contendo um plasmídeo TOL. Após 18 a 24 horas de homogenização da mistura, o inoculo foi adicionado e o decréscimo da concentração dos solventes foi determinado entre 24 e 25 horas por GC. Pseudomonas putida CCMI 852 foi capaz de metabolizar T e X, mas não B. Na mistura BTX, B não foi metabolizado também e a velocidade de degradação de T e X decresceu cerca de 50% comparado com soluções contendo apenas T ou X.

  5. [Influence of microcystin-LR on cell viability and surface characteristics of Pseudomonas putida].

    Science.gov (United States)

    Deng, Ting-jin; Ye, Jin-shao; Peng, Hui; Liu, Zhi-chen; Liu, Ze-hua; Yin, Hua; Chen, Shuo-na

    2015-01-01

    In microcystin-LR (MC-LR) degradation system, the change in surface characteristics and cell viability of Pseudomonas putida was studied. The purpose of this study was to reveal the influence of MC-LR on P. putida and elucidate the toxicity of MC-LR on microorganisms. The result demonstrated that MC-LR enhanced the cytoplasmic membrane permeability, as well as affected the ion metabolism and protein release of P. putida. The soluble sugar and Na+, Cl-release increased with the rising concentration of MC-LR ranging from 0 mg x L(-1) to 2.0 mg x L(-1). Flow Cytometry Method(FCM) analysis revealed that MC-LR accelerated the death of P. putida, and the death rate increased with the ascending concentration of MC-LR. Compared with the control, the death rate on day 5 increased by nearly 30% when 2.5 mg x L(-1) MC-LR was added. Scanning electron microscopy (SEM) analysis showed that the cells were deformed under the toxicity of MC-LR. After 5-day exposure to 2.5 mg x L(-1) MC-LR, the majority of the cells were ruptured and the intracellular materials flew out. The cellular structure was severely damaged under this condition.

  6. Crude glycerol as feedstock for the sustainable production of p-hydroxybenzoate by Pseudomonas putida S12.

    Science.gov (United States)

    Verhoef, Suzanne; Gao, Nisi; Ruijssenaars, Harald J; de Winde, Johannes H

    2014-01-25

    Crude glycerol is a promising renewable feedstock in bioconversion processes for the production of fuels and chemicals. Impurities present in crude glycerol can however, negatively impact the fermentation process. Successful crude glycerol utilization requires robust microbial production hosts that tolerate and preferably, can utilize such impurities. We investigated utilization of crude, unpurified glycerol as a substrate for the production of aromatic compounds by solvent tolerant Pseudomonas putida S12. In high-cell density fed-batch fermentations, P. putida S12 surprisingly performed better on crude glycerol than on purified glycerol. By contrast, growth of Escherichia coli was severely compromised under these high cell density cultivation conditions on crude glycerol. For P. putida S12 the biomass-to-substrate yield, maximum biomass production rate and substrate uptake rate were consistently higher on crude glycerol. Moreover, production of p-hydroxybenzoate by engineered P. putida S12palB5 on crude glycerol showed a 10% yield improvement over production on purified glycerol. P. putida S12 is a favorable host for bioconversion processes utilizing crude glycerol as a substrate. Its intrinsic stress-tolerance properties provide the robustness required for efficient growth and metabolism on this renewable substrate.

  7. Pseudomonas putida CSV86: a candidate genome for genetic bioaugmentation.

    Directory of Open Access Journals (Sweden)

    Vasundhara Paliwal

    Full Text Available Pseudomonas putida CSV86, a plasmid-free strain possessing capability to transfer the naphthalene degradation property, has been explored for its metabolic diversity through genome sequencing. The analysis of draft genome sequence of CSV86 (6.4 Mb revealed the presence of genes involved in the degradation of naphthalene, salicylate, benzoate, benzylalcohol, p-hydroxybenzoate, phenylacetate and p-hydroxyphenylacetate on the chromosome thus ensuring the stability of the catabolic potential. Moreover, genes involved in the metabolism of phenylpropanoid and homogentisate, as well as heavy metal resistance, were additionally identified. Ability to grow on vanillin, veratraldehyde and ferulic acid, detection of inducible homogentisate dioxygenase and growth on aromatic compounds in the presence of heavy metals like copper, cadmium, cobalt and arsenic confirm in silico observations reflecting the metabolic versatility. In silico analysis revealed the arrangement of genes in the order: tRNA(Gly, integrase followed by nah operon, supporting earlier hypothesis of existence of a genomic island (GI for naphthalene degradation. Deciphering the genomic architecture of CSV86 for aromatic degradation pathways and identification of elements responsible for horizontal gene transfer (HGT suggests that genetic bioaugmentation strategies could be planned using CSV86 for effective bioremediation.

  8. From dirt to industrial applications: Pseudomonas putida as a Synthetic Biology chassis for hosting harsh biochemical reactions.

    Science.gov (United States)

    Nikel, Pablo I; Chavarría, Max; Danchin, Antoine; de Lorenzo, Víctor

    2016-10-01

    The soil bacterium Pseudomonas putida is endowed with a central carbon metabolic network capable of fulfilling high demands of reducing power. This situation arises from a unique metabolic architecture that encompasses the partial recycling of triose phosphates to hexose phosphates-the so-called EDEMP cycle. In this article, the value of P. putida as a bacterial chassis of choice for contemporary, industrially-oriented metabolic engineering is addressed. The biochemical properties that make this bacterium adequate for hosting biotransformations involving redox reactions as well as toxic compounds and intermediates are discussed. Finally, novel developments and open questions in the continuous quest for an optimal microbial cell factory are presented at the light of current and future needs in the area of biocatalysis.

  9. Mutants of Pseudomonas putida affected in poly-3-hydroxyalkanoate synthesis

    NARCIS (Netherlands)

    Ren, Q; Kessler, B; van der Leij, F; Witholt, B.

    1998-01-01

    The generation and characterization of Pseudomonas putida KT2442 mutants affected in poly-3-hydroxyalkanoate (PHA) synthesis are reported. The mutants from P. putida KT2442 carrying several copies of the PHA-polymerase-encoding gene (phaC) were isolated via N-methyl-N'-nitro-N-nitrosoguanidine chemi

  10. Characterization of starvation-induced dispersion in Pseudomonas putida biofilms

    DEFF Research Database (Denmark)

    Gjermansen, Morten; Ragas, Paula Cornelia; Sternberg, Claus;

    2005-01-01

    that they must be able to regulate their ability to form biofilm and to dissolve biofilm. We present an investigation of a biofilm dissolution process occurring in flow-chamber-grown Pseudomonas putida biofilms. Local starvation-induced biofilm dissolution appears to be an integrated part of P. putida biofilm...... development that causes characteristic structural rearrangements. Rapid global dissolution of entire P. putida biofilms was shown to occur in response to carbon starvation. Genetic analysis suggested that the adjacent P. putida genes PP0164 and PP0165 play a role in P. putida biofilm formation and dissolution....... PP0164 encodes a putative periplasmic protein of previously unknown function, and PP0164 mutant bacteria are sticky, and unable to reduce their adhesiveness and dissolve their biofilm in response to carbon starvation. PP0165 encodes a putative transmembrane protein containing GGDEF and EAL domains...

  11. Influence of periplasmic oxidation of glucose on pyoverdine synthesis in Pseudomonas putida S11.

    Science.gov (United States)

    Ponraj, Paramasivan; Shankar, Manoharan; Ilakkiam, Devaraj; Rajendhran, Jeyaprakash; Gunasekaran, Paramasamy

    2013-06-01

    Fluorescent pseudomonads catabolize glucose simultaneously by two different pathways, namely, the oxidative pathway in periplasm and the phosphorylative pathway in cytoplasm. This study provides evidence for the role of glucose metabolism in the regulation of pyoverdine synthesis in Pseudomonas putida S11. We have characterized the influence of direct oxidation of glucose in periplasm on pyoverdine synthesis in P. putida S11. We identified a Tn5 transposon mutant of P. putida S11 showing increased pyoverdine production in minimal glucose medium (MGM). This mutant designated as IST1 had Tn5 insertion in glucose dehydrogenase (gcd) gene. To verify the role of periplasmic oxidation of glucose on pyoverdine synthesis, we constructed mutants S11 Gcd(-) and S11 PqqF(-) by antibiotic cassette mutagenesis. These mutants of P. putida S11 with loss of glucose dehydrogenase gene (gcd) or cofactor pyrroloquinoline quinone biosynthesis gene (pqqF) showed increased pyoverdine synthesis and impaired acid production in MGM. In minimal gluconate medium, the pyoverdine production of wild-type strain S11 and mutants S11 Gcd(-) and S11 PqqF(-) was higher than in MGM indicating that gluconate did not affect pyoverdine synthesis. In MGM containing PIPES-NaOH (pH 7.5) buffer which prevent pH changes due to gluconic acid production, strain S11 produced higher amount of pyoverdine similar to mutants S11 Gcd(-) and S11 PqqF(-). Therefore, it is proposed that periplasmic oxidation of glucose to gluconic acid decreases the pH of MGM and thereby influences pyoverdine synthesis of strain S11. The increased pyoverdine synthesis enhanced biotic and abiotic surface colonization of the strain S11.

  12. Metabolism

    Science.gov (United States)

    ... Surgery? Choosing the Right Sport for You Shyness Metabolism KidsHealth > For Teens > Metabolism Print A A A ... food through a process called metabolism. What Is Metabolism? Metabolism (pronounced: meh-TAB-uh-lih-zem) is ...

  13. Physical forces shape group identity of swimming Pseudomonas putida cells

    Directory of Open Access Journals (Sweden)

    David Rodriguez-Espeso

    2016-09-01

    Full Text Available The often striking macroscopic patterns developed by motile bacterial populations on agar plates are a consequence of the environmental conditions where the cells grow and spread. Parameters such as medium stiffness and nutrient concentration have been reported to alter cell swimming behavior, while mutual interactions among populations shape collective patterns. One commonly observed occurrence is the mutual inhibition of clonal bacteria when moving towards each other, which results in a distinct halt at a finite distance on the agar matrix before having direct contact. The dynamics behind this phenomenon (i.e. intolerance to mix in time and space with otherwise identical others has been traditionally explained in terms of cell-to-cell competition/cooperation regarding nutrient availability. In this work, the same scenario has been revisited from an alternative perspective: the effect of the physical mechanics that frame the process, in particular the consequences of collisions between moving bacteria and the semi-solid matrix of the swimming medium. To this end we set up a simple experimental system in which the swimming patterns of Pseudomonas putida were tested with different geometries and agar concentrations. A computational analysis framework that highlights cell-to-medium interactions was developed to fit experimental observations. Simulated outputs suggested that the medium is compressed in the direction of the bacterial front motion. This phenomenon generates what was termed a compression wave that goes through the medium preceding the swimming population and that determines the visible high-level pattern. Taken together, the data suggested that the mechanical effects of the bacteria moving through the medium created a factual barrier that impedes to merge with neighboring cells swimming from a different site. The resulting divide between otherwise clonal bacteria is thus brought about by physical forces –not genetic or metabolic

  14. Proteomics reveals a core molecular response of Pseudomonas putida F1 to acute chromate challenge

    Directory of Open Access Journals (Sweden)

    McCarthy Andrea T

    2010-05-01

    Full Text Available Abstract Background Pseudomonas putida is a model organism for bioremediation because of its remarkable metabolic versatility, extensive biodegradative functions, and ubiquity in contaminated soil environments. To further the understanding of molecular pathways responding to the heavy metal chromium(VI [Cr(VI], the proteome of aerobically grown, Cr(VI-stressed P. putida strain F1 was characterized within the context of two disparate nutritional environments: rich (LB media and minimal (M9L media containing lactate as the sole carbon source. Results Growth studies demonstrated that F1 sensitivity to Cr(VI was impacted substantially by nutrient conditions, with a carbon-source-dependent hierarchy (lactate > glucose >> acetate observed in minimal media. Two-dimensional HPLC-MS/MS was employed to identify differential proteome profiles generated in response to 1 mM chromate under LB and M9L growth conditions. The immediate response to Cr(VI in LB-grown cells was up-regulation of proteins involved in inorganic ion transport, secondary metabolite biosynthesis and catabolism, and amino acid metabolism. By contrast, the chromate-responsive proteome derived under defined minimal growth conditions was characterized predominantly by up-regulated proteins related to cell envelope biogenesis, inorganic ion transport, and motility. TonB-dependent siderophore receptors involved in ferric iron acquisition and amino acid adenylation domains characterized up-regulated systems under LB-Cr(VI conditions, while DNA repair proteins and systems scavenging sulfur from alternative sources (e.g., aliphatic sulfonates tended to predominate the up-regulated proteome profile obtained under M9L-Cr(VI conditions. Conclusions Comparative analysis indicated that the core molecular response to chromate, irrespective of the nutritional conditions tested, comprised seven up-regulated proteins belonging to six different functional categories including transcription, inorganic ion

  15. Pyoverdine synthesis by the Mn(II-oxidizing bacterium Pseudomonas putida GB-1

    Directory of Open Access Journals (Sweden)

    Dorothy Lundquist Parker

    2014-05-01

    Full Text Available When iron-starved, the Mn(II-oxidizing bacteria Pseudomonas putida strains GB-1 and MnB1 produce pyoverdines (PVDGB-1 and PVDMnB1, siderophores that both influence iron uptake and inhibit manganese(II oxidation by these strains. To explore the properties and genetics of a PVD that can affect manganese oxidation, LC-MS/MS and various siderotyping techniques were used to identify the peptides of PVDGB-1 and PVDMnB1 as being (for both PVDs: chromophore-Asp-Lys-OHAsp-Ser-Gly-aThr-Lys-cOHOrn, resembling a structure previously reported for P. putida CFML 90-51, which does not oxidize Mn. All three strains also produced an azotobactin and a sulfonated PVD, each with the peptide sequence above, but with unknown regulatory or metabolic effects. Bioinformatic analysis of the sequenced genome of P. putida GB-1 suggested that a particular non-ribosomal peptide synthetase, coded by the operon PputGB1_4083-4086, could produce the peptide backbone of PVDGB-1. To verify this prediction, plasmid integration disruption of PputGB1_4083 was performed and the resulting mutant failed to produce detectable PVD. In silico analysis of the modules in PputGB1_4083-4086 predicted a peptide sequence of Asp-Lys-Asp-Ser-Ala-Thr-Lsy-Orn, which closely matches the peptide determined by MS/MS. To extend these studies to other organisms, various Mn(II-oxidizing and non-oxidizing isolates of P. putida, P. fluorescens, P. marincola, P. fluorescens-syringae group, P. mendocina-resinovorans group and P. stutzerii group were screened for PVD synthesis. The PVD producers (12 out of 16 tested strains were siderotyped and placed into four sets of differing PVD structures, some corresponding to previously characterized PVDs and some to novel PVDs. These results combined with previous studies suggested that the presence of OHAsp or the flexibility of the pyoverdine polypeptide may enable efficient binding of Mn(III.

  16. A Pseudomonas putida strain genetically engineered for 1,2,3-trichloropropane bioremediation.

    Science.gov (United States)

    Samin, Ghufrana; Pavlova, Martina; Arif, M Irfan; Postema, Christiaan P; Damborsky, Jiri; Janssen, Dick B

    2014-09-01

    1,2,3-Trichloropropane (TCP) is a toxic compound that is recalcitrant to biodegradation in the environment. Attempts to isolate TCP-degrading organisms using enrichment cultivation have failed. A potential biodegradation pathway starts with hydrolytic dehalogenation to 2,3-dichloro-1-propanol (DCP), followed by oxidative metabolism. To obtain a practically applicable TCP-degrading organism, we introduced an engineered haloalkane dehalogenase with improved TCP degradation activity into the DCP-degrading bacterium Pseudomonas putida MC4. For this purpose, the dehalogenase gene (dhaA31) was cloned behind the constitutive dhlA promoter and was introduced into the genome of strain MC4 using a transposon delivery system. The transposon-located antibiotic resistance marker was subsequently removed using a resolvase step. Growth of the resulting engineered bacterium, P. putida MC4-5222, on TCP was indeed observed, and all organic chlorine was released as chloride. A packed-bed reactor with immobilized cells of strain MC4-5222 degraded >95% of influent TCP (0.33 mM) under continuous-flow conditions, with stoichiometric release of inorganic chloride. The results demonstrate the successful use of a laboratory-evolved dehalogenase and genetic engineering to produce an effective, plasmid-free, and stable whole-cell biocatalyst for the aerobic bioremediation of a recalcitrant chlorinated hydrocarbon. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  17. Characterization of starvation-induced dispersion in Pseudomonas putida biofilms: genetic elements and molecular mechanisms

    DEFF Research Database (Denmark)

    Gjermansen, M.; Nilsson, M.; Yang, Liang;

    2010-01-01

    P>Pseudomonas putida OUS82 biofilm dispersal was previously shown to be dependent on the gene PP0164 (here designated lapG). Sequence and structural analysis has suggested that the LapG geneproduct belongs to a family of cysteine proteinases that function in the modification of bacterial surface...... proteins. We provide evidence that LapG is involved in P. putida OUS82 biofilm dispersal through modification of the outer membrane-associated protein LapA. While the P. putida lapG mutant formed more biofilm than the wild-type, P. putida lapA and P. putida lapAG mutants displayed decreased surface...

  18. Industrial biotechnology of Pseudomonas putida and related species

    NARCIS (Netherlands)

    Poblete-Castro, I.; Becker, J.; Dohnt, K.; Martins Dos Santos, V.A.P.; Wittmann, C.

    2012-01-01

    Since their discovery many decades ago, Pseudomonas putida and related subspecies have been intensively studied with regard to their potential application in industrial biotechnology. Today, these Gram-negative soil bacteria, traditionally known as well-performing xenobiotic degraders, are becoming

  19. Biological production of monoethanolamine by engineered Pseudomonas putida S12

    NARCIS (Netherlands)

    Foti, M.J.; Médici, R.; Ruijssenaars, H.J.

    2013-01-01

    Pseudomonas putida S12 was engineered for the production of monoethanolamine (MEA) from glucose via the decarboxylation of the central metabolite l-serine, which is catalyzed by the enzyme l-serine decarboxylase (SDC).The host was first evaluated for its tolerance towards MEA as well as its

  20. The Transcriptional Landscape of the Production Organism Pseudomonas putida

    DEFF Research Database (Denmark)

    D'Arrigo, Isotta

    Bacterial cell factories represent a valid alternative to fossil fuel-based production. A promising bacterium that can be optimized as cell factory is Pseudomonas putida. However, its development in bioproduction applications poses some challenges including a clear understanding of the bacterial ...

  1. Probing the proteome response to toluene exposure in Pseudomonas putida

    NARCIS (Netherlands)

    Wijte, D.

    2011-01-01

    The toxic effects of organic solvents are a major drawback for their application in biotechnology and for the production of fine chemicals by whole-cell bioconversion and biotransformation. However, through the isolation of some microbial strains, like P. putida S12, which tolerate high concentratio

  2. Photoautotrophic production of polyhydroxyalkanoates in a synthetic mixed culture of Synechococcus elongatus cscB and Pseudomonas putida cscAB.

    Science.gov (United States)

    Löwe, Hannes; Hobmeier, Karina; Moos, Manuel; Kremling, Andreas; Pflüger-Grau, Katharina

    2017-01-01

    One of the major challenges for the present and future generations is to find suitable substitutes for the fossil resources we rely on today. Cyanobacterial carbohydrates have been discussed as an emerging renewable feedstock in industrial biotechnology for the production of fuels and chemicals, showing promising production rates when compared to crop-based feedstock. However, intrinsic capacities of cyanobacteria to produce biotechnological compounds are limited and yields are low. Here, we present an approach to circumvent these problems by employing a synthetic bacterial co-culture for the carbon-neutral production of polyhydroxyalkanoates (PHAs) from CO2. The co-culture consists of two bio-modules: Bio-module I, in which the cyanobacterial strain Synechococcus elongatus cscB fixes CO2, converts it to sucrose, and exports it into the culture supernatant; and bio-module II, where this sugar serves as C-source for Pseudomonas putida cscAB and is converted to PHAs that are accumulated in the cytoplasm. By applying a nitrogen-limited process, we achieved a maximal PHA production rate of 23.8 mg/(L day) and a maximal titer of 156 mg/L. We will discuss the present shortcomings of the process and show the potential for future improvement. These results demonstrate the feasibility of mixed cultures of S. elongatus cscB and P. putida cscAB for PHA production, making room for the cornucopia of possible products that are described for P. putida. The construction of more efficient sucrose-utilizing P. putida phenotypes and the optimization of process conditions will increase yields and productivities and eventually close the gap in the contemporary process. In the long term, the co-culture may serve as a platform process, in which P. putida is used as a chassis for the implementation of synthetic metabolic pathways for biotechnological production of value-added products.

  3. Bioremediation in marine ecosystems: a computational study combining ecological modelling and flux balance analysis

    Directory of Open Access Journals (Sweden)

    Marianna eTaffi

    2014-09-01

    Full Text Available The pressure to search effective bioremediation methodologies for contaminated ecosystems has led to the large-scale identification of microbial species and metabolic degradation pathways. However, minor attention has been paid to the study of bioremediation in marine food webs and to the definition of integrated strategies for reducing bioaccumulation in species. We propose a novel computational framework for analysing the multiscale effects of bioremediation at the ecosystem level, based on coupling food web bioaccumulation models and metabolic models of degrading bacteria. The combination of techniques from synthetic biology and ecological network analysis allows the specification of arbitrary scenarios of contaminant removal and the evaluation of strategies based on natural or synthetic microbial strains.In this study, we derive a bioaccumulation model of polychlorinated biphenyls (PCBs in the Adriatic food web, and we extend a metabolic reconstruction of Pseudomonas putida KT2440 (iJN746 with the aerobic pathway of PCBs degradation. We assess the effectiveness of different bioremediation scenarios in reducing PCBs concentration in species and we study indices of species centrality to measure their importance in the contaminant diffusion via feeding links.The analysis of the Adriatic sea case study suggests that our framework could represent a practical tool in the design of effective remediation strategies, providing at the same time insights into the ecological role of microbial communities within food webs.

  4. Bioremediation in marine ecosystems: a computational study combining ecological modeling and flux balance analysis.

    Science.gov (United States)

    Taffi, Marianna; Paoletti, Nicola; Angione, Claudio; Pucciarelli, Sandra; Marini, Mauro; Liò, Pietro

    2014-01-01

    The pressure to search effective bioremediation methodologies for contaminated ecosystems has led to the large-scale identification of microbial species and metabolic degradation pathways. However, minor attention has been paid to the study of bioremediation in marine food webs and to the definition of integrated strategies for reducing bioaccumulation in species. We propose a novel computational framework for analysing the multiscale effects of bioremediation at the ecosystem level, based on coupling food web bioaccumulation models and metabolic models of degrading bacteria. The combination of techniques from synthetic biology and ecological network analysis allows the specification of arbitrary scenarios of contaminant removal and the evaluation of strategies based on natural or synthetic microbial strains. In this study, we derive a bioaccumulation model of polychlorinated biphenyls (PCBs) in the Adriatic food web, and we extend a metabolic reconstruction of Pseudomonas putida KT2440 (iJN746) with the aerobic pathway of PCBs degradation. We assess the effectiveness of different bioremediation scenarios in reducing PCBs concentration in species and we study indices of species centrality to measure their importance in the contaminant diffusion via feeding links. The analysis of the Adriatic sea case study suggests that our framework could represent a practical tool in the design of effective remediation strategies, providing at the same time insights into the ecological role of microbial communities within food webs.

  5. Metabolism

    Science.gov (United States)

    ... Are More Common in People With Type 1 Diabetes Metabolic Syndrome Your Child's Weight Healthy Eating Endocrine System Blood Test: Basic Metabolic Panel (BMP) Activity: Endocrine System Growth Disorders Diabetes Center Thyroid Disorders Your Endocrine System Movie: Endocrine ...

  6. The Regulation of para-Nitrophenol Degradation in Pseudomonas putida DLL-E4

    Science.gov (United States)

    Chen, Qiongzhen; Tu, Hui; Luo, Xue; Zhang, Biying; Huang, Fei; Li, Zhoukun; Wang, Jue; Shen, Wenjing; Wu, Jiale; Cui, Zhongli

    2016-01-01

    Pseudomonas putida DLL-E4 can efficiently degrade para-nitrophenol and its intermediate metabolite hydroquinone. The regulation of para-nitrophenol degradation was studied, and PNP induced a global change in the transcriptome of P. putida DLL-E4. When grown on PNP, the wild-type strain exhibited significant downregulation of 2912 genes and upregulation of 845 genes, whereas 2927 genes were downregulated and 891 genes upregulated in a pnpR-deleted strain. Genes related to two non-coding RNAs (ins1 and ins2), para-nitrophenol metabolism, the tricarboxylic acid cycle, the outer membrane porin OprB, glucose dehydrogenase Gcd, and carbon catabolite repression were significantly upregulated when cells were grown on para-nitrophenol plus glucose. pnpA, pnpR, pnpC1C2DECX1X2, and pnpR1 are key genes in para-nitrophenol degradation, whereas pnpAb and pnpC1bC2bDbEbCbX1bX2b have lost the ability to degrade para-nitrophenol. Multiple components including transcriptional regulators and other unknown factors regulate para-nitrophenol degradation, and the transcriptional regulation of para-nitrophenol degradation is complex. Glucose utilization was enhanced at early stages of para-nitrophenol supplementation. However, it was inhibited after the total consumption of para-nitrophenol. The addition of glucose led to a significant enhancement in para-nitrophenol degradation and up-regulation in the expression of genes involved in para-nitrophenol degradation and carbon catabolite repression (CCR). It seemed that para-nitrophenol degradation can be regulated by CCR, and relief of CCR might contribute to enhanced para-nitrophenol degradation. In brief, the regulation of para-nitrophenol degradation seems to be controlled by multiple factors and requires further study. PMID:27191401

  7. The Regulation of para-Nitrophenol Degradation in Pseudomonas putida DLL-E4.

    Directory of Open Access Journals (Sweden)

    Qiongzhen Chen

    Full Text Available Pseudomonas putida DLL-E4 can efficiently degrade para-nitrophenol and its intermediate metabolite hydroquinone. The regulation of para-nitrophenol degradation was studied, and PNP induced a global change in the transcriptome of P. putida DLL-E4. When grown on PNP, the wild-type strain exhibited significant downregulation of 2912 genes and upregulation of 845 genes, whereas 2927 genes were downregulated and 891 genes upregulated in a pnpR-deleted strain. Genes related to two non-coding RNAs (ins1 and ins2, para-nitrophenol metabolism, the tricarboxylic acid cycle, the outer membrane porin OprB, glucose dehydrogenase Gcd, and carbon catabolite repression were significantly upregulated when cells were grown on para-nitrophenol plus glucose. pnpA, pnpR, pnpC1C2DECX1X2, and pnpR1 are key genes in para-nitrophenol degradation, whereas pnpAb and pnpC1bC2bDbEbCbX1bX2b have lost the ability to degrade para-nitrophenol. Multiple components including transcriptional regulators and other unknown factors regulate para-nitrophenol degradation, and the transcriptional regulation of para-nitrophenol degradation is complex. Glucose utilization was enhanced at early stages of para-nitrophenol supplementation. However, it was inhibited after the total consumption of para-nitrophenol. The addition of glucose led to a significant enhancement in para-nitrophenol degradation and up-regulation in the expression of genes involved in para-nitrophenol degradation and carbon catabolite repression (CCR. It seemed that para-nitrophenol degradation can be regulated by CCR, and relief of CCR might contribute to enhanced para-nitrophenol degradation. In brief, the regulation of para-nitrophenol degradation seems to be controlled by multiple factors and requires further study.

  8. Metabolism

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    2008255 Serum adiponectin level declines in the elderly with metabolic syndrome.WU Xiaoyan(吴晓琰),et al.Dept Geriatr,Huashan Hosp,Fudan UnivShanghai200040.Chin J Geriatr2008;27(3):164-167.Objective To investigate the correlation between ser-um adiponectin level and metabolic syndrome in the elderly·Methods Sixty-one subjects with metabolic syndrome and140age matched subjects without metabolic

  9. Characterization of the RND family of multidrug efflux pumps: in silico to in vivo confirmation of four functionally distinct subgroups.

    Science.gov (United States)

    Godoy, Patricia; Molina-Henares, Antonio J; de la Torre, Jesús; Duque, Estrella; Ramos, Juan L

    2010-11-01

    We have developed a generalized profile that identifies members of the root-nodulation-cell-division (RND) family of efflux pumps and classifies them into four functional subfamilies. According to Z-score values, efflux pumps can be grouped by their metabolic function, thus making it possible to distinguish pumps involved in antibiotic resistance (group 1) from those involved in metal resistance (group 3). In silico data regarding efflux pumps in group 1 were validated after identification of RND efflux pumps in a number of environmental microbes that were isolated as resistant to ethidium bromide. Analysis of the Pseudomonas putida KT2440 genome identified efflux pumps in all groups. A collection of mutants in efflux pumps and a screening platform consisting of 50 drugs were created to assign a function to the efflux pumps. We validated in silico data regarding efflux pumps in groups 1 and 3 using 9 different mutants. Four mutants belonging to group 2 were found to be more sensitive than the wild-type to oxidative stress-inducing agents such as bipyridyl and methyl viologen. The two remaining mutants belonging to group 4 were found to be more sensitive than the parental to tetracycline and one of them was particularly sensitive to rubidium and chromate. By effectively combining in vivo data with generalized profiles and gene annotation data, this approach allowed the assignment, according to metabolic function, of both known and uncharacterized RND efflux pumps into subgroups, thereby providing important new insight into the functions of proteins within this family.

  10. iTRAQ-based quantitative proteomic analysis of the global response to 17β-estradiol in estrogen-degradation strain Pseudomonas putida SJTE-1

    Science.gov (United States)

    Xu, Jing; Zhang, Lei; Hou, Jingli; Wang, Xiuli; Liu, Huan; Zheng, Daning; Liang, Rubing

    2017-01-01

    Microorganism degradation is efficient to remove the steroid hormones like 17β-estradiol (E2); but their degradation mechanism and metabolic network to these chemicals are still not very clear. Here the global responses of the estrogen-degradation strain Pseudomonas putida SJTE-1 to 17β-estradiol and glucose were analyzed and compared using the iTRAQ (isobaric tags for relative and absolute quantization) strategy combined with LC-MS/MS (liquid chromatography-tandem mass spectrometry). 78 proteins were identified with significant changes in expression; 45 proteins and 33 proteins were up-regulated and down-regulated, respectively. These proteins were mainly involved in the processes of stress response, energy metabolism, transportation, chemotaxis and cell motility, and carbon metabolism, considered probably responding to 17β-estradiol and playing a role in its metabolism. The up-regulated proteins in electron transfer, energy generation and transport systems were thought crucial for efficient uptake, translocation and transformation of 17β-estradiol. The over-expression of carbon metabolism proteins indicated cells may activate related pathway members to utilize 17β-estradiol. Meanwhile, proteins functioning in glucose capture and metabolism were mostly down-regulated. These findings provide important clues to reveal the 17β-estradiol degradation mechanism in P. putida and promote its bioremediation applications. PMID:28155874

  11. Draft Genome Sequence of the Model Naphthalene-Utilizing Organism Pseudomonas putida OUS82

    DEFF Research Database (Denmark)

    Tay, Martin; Roizman, Dan; Cohen, Yehuda

    2014-01-01

    Pseudomonas putida OUS82 was isolated from petrol- and oil-contaminated soil in 1992, and ever since, it has been used as a model organism to study the microbial assimilation of naphthalene and phenanthrene. Here, we report the 6.7-Mb draft genome sequence of P. putida OUS82 and analyze its featu...

  12. Chemostat-based proteomic analysis of toluene-affected Pseudomonas putida S12

    NARCIS (Netherlands)

    Volkers, R.J.M.; Jong, A.L. de; Hulst, A.G.; Baar, B.L.M. van; Bont, J.A.M. de; Wery, J.

    2006-01-01

    The aim of this study was to assess the cellular response of the solvent-tolerant Pseudomonas putida S12 to toluene as the single effector. Proteomic analysis (two-dimensional difference-in-gel-electrophoresis) was used to assess the response of P. putida S12 cultured in chemostats. This approach en

  13. Chemostat-based proteomic analysis of toluene-affected Pseudomonas putida S12

    NARCIS (Netherlands)

    Volkers, R.J.M.; Jong, A.L. de; Hulst, A.G.; Baar, B.L.M. van; Bont, J.A.M. de; Wery, J.

    2006-01-01

    The aim of this study was to assess the cellular response of the solvent-tolerant Pseudomonas putida S12 to toluene as the single effector. Proteomic analysis (two-dimensional difference-in-gel-electrophoresis) was used to assess the response of P. putida S12 cultured in chemostats. This approach en

  14. Synergism of Lumbricus rubellus and Pseudomonas putida Pf-20 in Inducing Resistance to Cucumber Mosaic Virus

    Directory of Open Access Journals (Sweden)

    WIWIEK SRI WAHYUNI

    2006-09-01

    Full Text Available Both Lumbricus rubellus and Pseudomonas putida decompose soil organic matters. The population of P. putida Pf-20 increased if L. rubellus was introduced to the cucumber growth medium. The process of organic decomposition was much better if the medium was introduced with both L. rubellus and P. putida Pf-20, compared to the medium contained only either one of those organisms. The activity of L. rubellus may serve to provide nutrients for both the cucumber and P. putida. The role of P. putida to reduce disease severity was increased if L. rubellus was introduced to the growth medium. The synergism of these two organisms, reduced either the level of disease severity to CMV-48 and C/N ratio of medium, but increased the content of available phosphor and potassium.

  15. A tricistronic heat shock operon is important for stress tolerance of Pseudomonas putida and conserved in many environmental bacteria.

    Science.gov (United States)

    Krajewski, Stefanie S; Joswig, Matthias; Nagel, Miriam; Narberhaus, Franz

    2014-06-01

    Small heat shock proteins (sHsps) including the well-studied IbpA protein from Escherichia coli are molecular chaperones that bind to non-native proteins and prevent them from aggregation. We discovered an entirely unexplored tricistronic small heat shock gene cluster in Pseudomonas putida. The genes pp3314, pp3313 and pp3312 (renamed to hspX, hspY and hspZ respectively) are transcribed in a single transcript. In addition to σ(32) -dependent transcriptional control, translation of the first and second gene of the operon is controlled by RNA thermometers with novel architectures. Biochemical analysis of HspY, HspZ and P. putida IbpA demonstrated that they assemble into homo-oligomers of different sizes whose quaternary structures alter in a temperature-dependent manner. IbpA and HspY are able to prevent the model substrate citrate synthase from thermal aggregation in vitro. Increased stress sensitivity of a P. putida strain lacking HspX, HspY and HspZ revealed an important role of these sHsps in stress adaptation. The hspXYZ operon is conserved among metabolically related bacteria that live in hostile environments including polluted soils. This heat shock operon might act as a protective system to promote survival in such ecological niches.

  16. Molecular Probes: A Tool for Studying Toxicity of VOCs to P.Putida F1

    Science.gov (United States)

    Singh, R.; Olson, M. S.

    2007-12-01

    Volatile Organic Compounds (VOCs) are of great concern in ground water remediation, and are generally present in the form of NAPLs in subsurface environments. Among the various treatment technologies, in situ bioremediation is one of the most effective and low-cost treatment options. Many soil bacteria are reported to degrade these organic contaminants via metabolism (using them as a source of carbon to derive energy) or co- metabolism up to certain concentrations. However, larger concentrations of these contaminants are toxic to bacteria. Thus, in order to achieve successful bioremediation, it is important to determine the optimal concentrations of various contaminants that is beneficial for the activity and survival of degrading bacteria. The purpose of this study is to develop a novel method for toxicity analyses of VOC contaminants to the soil bacteria that degrade them. The present study is based on a two-color fluorescence assay of bacterial viability which facilitates actual counting of live and dead bacteria. Pseudomonas putida F1 cells were labeled with a LIVE/DEAD® BacLightTM bacterial viability kit (Invitrogen), which consists of a mixture of two dyes, SYTO 9 and propidium iodide, each with a different ability to penetrate healthy bacterial cells. Live cells stain green whereas propidium iodide (red dye) only penetrates cells with compromised membranes that are considered dead or dying. Stained cells were exposed to different concentrations of trichloroethylene (TCE) and toluene in sealed vials. Change in the concentrations of green and red cells were monitored over the time using fluorescence microscopy. UTHSCSA ImageTool software was used to count the live and dead cells in the images. It was observed that live (green) cell concentrations decreased and dead/damaged (red) cell concentrations increased over time when cells were exposed to TCE. No significant changes were observed in control experiments. Death rate constants calculated based on live cell

  17. Metabolism

    Science.gov (United States)

    ... a particular food provides to the body. A chocolate bar has more calories than an apple, so ... acid phenylalanine, needed for normal growth and protein production). Inborn errors of metabolism can sometimes lead to ...

  18. Structure characterization of phospholipids and lipid A of Pseudomonas putida KT2442.

    Science.gov (United States)

    Wang, Yuqian; Wang, Jianli; Li, Ye; Wang, Biwen; Tao, Guanjun; Wang, Xiaoyuan

    2015-01-01

    Pseudomonas putida KT2442 is an important bacterium for producing various types of polyhydroxyalkanoate polymers. Phospholipids and lipid A in membranes of P. putida play important roles in stress responses, but detailed structural information of these lipids is not known. In this study, phospholipids and lipid A were isolated from P. putida KT2442, and their structures were analyzed using thin layer chromatography, high performance liquid chromatography, and electrospray ionization/mass spectrometry. Major phospholipids in P. putida KT2442 were phosphatidylethanolamine (79.9%), phosphatidylglycero1 (12.7%), and cardiolipin (7.4%), with C16:1 and/or C18:1 acyl chains. Four lipid A species were found in P. putida KT2442: two are hexa-acylated, and the other two are penta-acylated. Compared with lipid A of P. aeruginosa, P. putida lipid A has less hydroxylation on the secondary acyl chains and less modification. Therefore, P. putida lipid A could be used as a base structure to investigate lipid A modification of P. aeruginosa for understanding its pathogenesis.

  19. Toxicity of titanium dioxide nanoparticles on Pseudomonas putida.

    Science.gov (United States)

    Combarros, R G; Collado, S; Díaz, M

    2016-03-01

    The increasing use of engineered nanoparticles (NPs) in industrial and household applications will very likely lead to the release of such materials into the environment. As wastewater treatment plants (WWTPs) are usually the last barrier before the water is discharged into the environment, it is important to understand the effects of these materials in the biotreatment processes, since the results in the literature are usually contradictory. We proposed the use of flow cytometry (FC) technology to obtain conclusive results. Aqueous solutions of TiO2 nanoparticles (0-2 mg mL(-1)) were used to check its toxicity effect using Pseudomonas putida as simplified model of real sludge over room light. Physiological changes in P. putida from viable to viable but non-culturable cells were observed by flow cytometry in presence of TiO2. The damaged and dead cell concentrations were below 5% in all cases under study. Both FSC and SSC parameter increased with TiO2 dose dependent manner, indicating nanoparticles uptake by the bacteria. The biological removal of salicylic acid (SA) was also significantly impacted by the presence of TiO2 in the medium reducing the efficiency. The use of FC allows also to develop and fit segregated kinetic models, giving the impact of TiO2 nanoparticles in the physiological subpopulations growth and implications for SA removal.

  20. The davDT operon of Pseudomonas putida, involved in lysine catabolism, is induced in response to the pathway intermediate delta-aminovaleric acid

    DEFF Research Database (Denmark)

    Revelles, O.; Espinosa-Urgel, M.; Molin, Søren;

    2004-01-01

    -aminovaleric acid and then further degraded to glutaric acid via the action of the davDT gene products. We show that the davDT genes form an operon transcribed from a single sigma(70)-dependent promoter. The relatively high level of basal expression from the davD promoter increased about fourfold in response...... to the addition of exogenous lysine to the culture medium. However, the true inducer of this operon seems to be delta-aminovaleric acid because in a mutant unable to metabolize lysine to delta-aminovaleric acid, this compound, but not lysine, acted as an effector. Effective induction of the P. putida P...

  1. Comparative genomic and functional analyses: unearthing the diversity and specificity of nematicidal factors in Pseudomonas putida strain 1A00316

    Science.gov (United States)

    Guo, Jing; Jing, Xueping; Peng, Wen-Lei; Nie, Qiyu; Zhai, Yile; Shao, Zongze; Zheng, Longyu; Cai, Minmin; Li, Guangyu; Zuo, Huaiyu; Zhang, Zhitao; Wang, Rui-Ru; Huang, Dian; Cheng, Wanli; Yu, Ziniu; Chen, Ling-Ling; Zhang, Jibin

    2016-01-01

    We isolated Pseudomonas putida (P. putida) strain 1A00316 from Antarctica. This bacterium has a high efficiency against Meloidogyne incognita (M. incognita) in vitro and under greenhouse conditions. The complete genome of P. putida 1A00316 was sequenced using PacBio single molecule real-time (SMRT) technology. A comparative genomic analysis of 16 Pseudomonas strains revealed that although P. putida 1A00316 belonged to P. putida, it was phenotypically more similar to nematicidal Pseudomonas fluorescens (P. fluorescens) strains. We characterized the diversity and specificity of nematicidal factors in P. putida 1A00316 with comparative genomics and functional analysis, and found that P. putida 1A00316 has diverse nematicidal factors including protein alkaline metalloproteinase AprA and two secondary metabolites, hydrogen cyanide and cyclo-(l-isoleucyl-l-proline). We show for the first time that cyclo-(l-isoleucyl-l-proline) exhibit nematicidal activity in P. putida. Interestingly, our study had not detected common nematicidal factors such as 2,4-diacetylphloroglucinol (2,4-DAPG) and pyrrolnitrin in P. putida 1A00316. The results of the present study reveal the diversity and specificity of nematicidal factors in P. putida strain 1A00316. PMID:27384076

  2. METABOLISM

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    Objective: To determine the allele frequencies of genetic variants 373 Ala→Pro and 451 Arg→Gln of cholesteryl ester transfer protein (CETP) and to explore their potential impacts on serum lipid metabolism. Methods: The genotypes in CETP codon 373 and 451 in 91 German healthy students and 409 an-

  3. Less is more: reduced catechol production permits Pseudomonas putida F1 to grow on styrene.

    Science.gov (United States)

    George, Kevin W; Hay, Anthony

    2012-11-01

    Pseudomonas putida F1 is unable to grow on styrene due to the accumulation of 3-vinylcatechol, a toxic metabolite that is produced through the toluene degradation (tod) pathway and causes catechol-2,3-dioxygenase (C23O) inactivation. In this study, we characterized a spontaneous F1 mutant, designated SF1, which acquired the ability to grow on styrene and did not accumulate 3-vinylcatechol. Whereas adaptation to new aromatic substrates has typically been shown to involve increased C23O activity or the acquisition of resistance to C23O inactivation, SF1 retained wild-type C23O activity. Surprisingly, SF1 grew more slowly on toluene, its native substrate, and exhibited reduced toluene dioxygenase (TDO) activity (approximately 50 % of that of F1), the enzyme responsible for ring hydroxylation and subsequent production of 3-vinylcatechol. DNA sequence analysis of the tod operon of SF1 revealed a single base pair mutation in todA (C479T), a gene encoding the reductase component of TDO. Replacement of the wild-type todA allele in F1 with todA(C479T) reduced TDO activity to SF1 levels, obviated vinylcatechol accumulation, and conferred the ability to grow on styrene. This novel 'less is more' strategy - reduced catechol production as a means to expand growth substrate range - sheds light on an alternative approach for managing catechol toxicity during the metabolism of aromatic compounds.

  4. Purification and characterization of benzyl alcohol- and benzaldehyde- dehydrogenase from Pseudomonas putida CSV86.

    Science.gov (United States)

    Shrivastava, Rahul; Basu, Aditya; Phale, Prashant S

    2011-08-01

    Pseudomonas putida CSV86 utilizes benzyl alcohol via catechol and methylnaphthalenes through detoxification pathway via hydroxymethylnaphthalenes and naphthaldehydes. Based on metabolic studies, benzyl alcohol dehydrogenase (BADH) and benzaldehyde dehydrogenase (BZDH) were hypothesized to be involved in the detoxification pathway. BADH and BZDH were purified to apparent homogeneity and were (1) homodimers with subunit molecular mass of 38 and 57 kDa, respectively, (2) NAD(+) dependent, (3) broad substrate specific accepting mono- and di-aromatic alcohols and aldehydes but not aliphatic compounds, and (4) BADH contained iron and magnesium, while BZDH contained magnesium. BADH in the forward reaction converted alcohol to aldehyde and required NAD(+), while in the reverse reaction it reduced aldehyde to alcohol in NADH-dependent manner. BZDH showed low K (m) value for benzaldehyde as compared to BADH reverse reaction. Chemical cross-linking studies revealed that BADH and BZDH do not form multi-enzyme complex. Thus, the conversion of aromatic alcohol to acid is due to low K (m) and high catalytic efficiency of BZDH. Phylogenetic analysis revealed that BADH is a novel enzyme and diverged during the evolution to gain the ability to utilize mono- and di-aromatic compounds. The wide substrate specificity of these enzymes enables strain to detoxify methylnaphthalenes to naphthoic acids efficiently.

  5. Growth of genetically engineered Pseudomonas aeruginosa and Pseudomonas putida in soil and rhizosphere.

    OpenAIRE

    Yeung, K H; Schell, M A; Hartel, P G

    1989-01-01

    The effect of the addition of a recombinant plasmid containing the pglA gene encoding an alpha-1,4-endopolygalacturonase from Pseudomonas solanacearum on the growth of Pseudomonas aeruginosa and Pseudomonas putida in soil and rhizosphere was determined. Despite a high level of polygalacturonase production by genetically engineered P. putida and P. aeruginosa, the results suggest that polygalacturonase production had little effect on the growth of these strains in soil or rhizosphere.

  6. Manganese (Mn) Oxidation Increases Intracellular Mn in Pseudomonas putida GB-1

    Science.gov (United States)

    Banh, Andy; Chavez, Valarie; Doi, Julia; Nguyen, Allison; Hernandez, Sophia; Ha, Vu; Jimenez, Peter; Espinoza, Fernanda; Johnson, Hope A.

    2013-01-01

    Bacterial manganese (Mn) oxidation plays an important role in the global biogeochemical cycling of Mn and other compounds, and the diversity and prevalence of Mn oxidizers have been well established. Despite many hypotheses of why these bacteria may oxidize Mn, the physiological reasons remain elusive. Intracellular Mn levels were determined for Pseudomonas putida GB-1 grown in the presence or absence of Mn by inductively coupled plasma mass spectrometry (ICP-MS). Mn oxidizing wild type P. putida GB-1 had higher intracellular Mn than non Mn oxidizing mutants grown under the same conditions. P. putida GB-1 had a 5 fold increase in intracellular Mn compared to the non Mn oxidizing mutant P. putida GB-1-007 and a 59 fold increase in intracellular Mn compared to P. putida GB-1 ∆2665 ∆2447. The intracellular Mn is primarily associated with the less than 3 kDa fraction, suggesting it is not bound to protein. Protein oxidation levels in Mn oxidizing and non oxidizing cultures were relatively similar, yet Mn oxidation did increase survival of P. putida GB-1 when oxidatively stressed. This study is the first to link Mn oxidation to Mn homeostasis and oxidative stress protection. PMID:24147089

  7. Molecular determinants of azo reduction activity in the strain Pseudomonas putida MET94.

    Science.gov (United States)

    Mendes, Sónia; Pereira, Luciana; Batista, Carlos; Martins, Lígia O

    2011-10-01

    Azo dyes are the major group of synthetic colourants used in industry and are serious environmental pollutants. In this study, Pseudomonas putida MET94 was selected from 48 bacterial strains on the basis of its superior ability to degrade a wide range of structurally diverse azo dyes. P. putida is a versatile microorganism with a well-recognised potential for biodegradation or bioremediation applications. P. putida MET94 removes, in 24 h and under anaerobic growing conditions, more than 80% of the majority of the structurally diverse azo dyes tested. Whole cell assays performed under anaerobic conditions revealed up to 90% decolourisation in dye wastewater bath models. The involvement of a FMN dependent NADPH: dye oxidoreductase in the decolourisation process was suggested by enzymatic measurements in cell crude extracts. The gene encoding a putative azoreductase was cloned from P. putida MET94 and expressed in Escherichia coli. The purified P. putida azoreductase is a 40 kDa homodimer with broad substrate specificity for azo dye reduction. The presence of dioxygen leads to the inhibition of the decolourisation activity in agreement with the results of cell cultures. The kinetic mechanism follows a ping-pong bi-bi reaction scheme and aromatic amine products were detected in stoichiometric amounts by high-performance liquid chromatography. Overall, the results indicate that P. putida MET94 is a promising candidate for bioengineering studies aimed at generating more effective dye-reducing strains.

  8. Biodegradation in a Partially Saturated Sand Matrix: Compounding Effects of Water Content, Bacterial Spatial Distribution, and Motility

    DEFF Research Database (Denmark)

    Dechesne, Arnaud; Owsianiak, Mikolaj; Bazire, Alexis

    2010-01-01

    colonizing these zones or on pollutant mass transfer to neighboring zones containing degraders. In a model system, we quantified the role exerted by water on mineralization rate in the context of a heterogeneously distributed degradation potential. Alginate beads colonized by Pseudomonas putida KT2440 were......Bacterial pesticide degraders are generally heterogeneously distributed in soils, leaving soil volumes devoid of degradation potential. This is expected to have an impact on degradation rates because the degradation of pollutant molecules in such zones will be contingent either on degraders......, partially relieving the diffusion limitation. Dry conditions, however, sustained low mineralization rates through the combined effects of low pollutant diffusivity and limited degrader dispersal....

  9. Integrated organic-aqueous biocatalysis and product recovery for quinaldine hydroxylation catalyzed by living recombinant Pseudomonas putida.

    Science.gov (United States)

    Ütkür, F Ozde; Thanh Tran, Tan; Collins, Jonathan; Brandenbusch, Christoph; Sadowski, Gabriele; Schmid, Andreas; Bühler, Bruno

    2012-07-01

    In an earlier study, biocatalytic carbon oxyfunctionalization with water serving as oxygen donor, e.g., the bioconversion of quinaldine to 4-hydroxyquinaldine, was successfully achieved using resting cells of recombinant Pseudomonas putida, containing the molybdenum-enzyme quinaldine 4-oxidase, in a two-liquid phase (2LP) system (Ütkür et al. J Ind Microbiol Biotechnol 38:1067-1077, 2011). In the study reported here, key parameters determining process performance were investigated and an efficient and easy method for product recovery was established. The performance of the whole-cell biocatalyst was shown not to be limited by the availability of the inducer benzoate (also serving as growth substrate) during the growth of recombinant P. putida cells. Furthermore, catalyst performance during 2LP biotransformations was not limited by the availability of glucose, the energy source to maintain metabolic activity in resting cells, and molecular oxygen, a possible final electron acceptor during quinaldine oxidation. The product and the organic solvent (1-dodecanol) were identified as the most critical factors affecting biocatalyst performance, to a large extent on the enzyme level (inhibition), whereas substrate effects were negligible. However, none of the 13 alternative solvents tested surpassed 1-dodecanol in terms of toxicity, substrate/product solubility, and partitioning. The use of supercritical carbon dioxide for phase separation and an easy and efficient liquid-liquid extraction step enabled 4-hydroxyquinaldine to be isolated at a purity of >99.9% with recoveries of 57 and 84%, respectively. This study constitutes the first proof of concept on an integrated process for the oxyfunctionalization of toxic substrates with a water-incorporating hydroxylase.

  10. Pmr, a histone-like protein H1 (H-NS) family protein encoded by the IncP-7 plasmid pCAR1, is a key global regulator that alters host function.

    Science.gov (United States)

    Yun, Choong-Soo; Suzuki, Chiho; Naito, Kunihiko; Takeda, Toshiharu; Takahashi, Yurika; Sai, Fumiya; Terabayashi, Tsuguno; Miyakoshi, Masatoshi; Shintani, Masaki; Nishida, Hiromi; Yamane, Hisakazu; Nojiri, Hideaki

    2010-09-01

    Histone-like protein H1 (H-NS) family proteins are nucleoid-associated proteins (NAPs) conserved among many bacterial species. The IncP-7 plasmid pCAR1 is transmissible among various Pseudomonas strains and carries a gene encoding the H-NS family protein, Pmr. Pseudomonas putida KT2440 is a host of pCAR1, which harbors five genes encoding the H-NS family proteins PP_1366 (TurA), PP_3765 (TurB), PP_0017 (TurC), PP_3693 (TurD), and PP_2947 (TurE). Quantitative reverse transcription-PCR (qRT-PCR) demonstrated that the presence of pCAR1 does not affect the transcription of these five genes and that only pmr, turA, and turB were primarily transcribed in KT2440(pCAR1). In vitro pull-down assays revealed that Pmr strongly interacted with itself and with TurA, TurB, and TurE. Transcriptome comparisons of the pmr disruptant, KT2440, and KT2440(pCAR1) strains indicated that pmr disruption had greater effects on the host transcriptome than did pCAR1 carriage. The transcriptional levels of some genes that increased with pCAR1 carriage, such as the mexEF-oprN efflux pump genes and parI, reverted with pmr disruption to levels in pCAR1-free KT2440. Transcriptional levels of putative horizontally acquired host genes were not altered by pCAR1 carriage but were altered by pmr disruption. Identification of genome-wide Pmr binding sites by ChAP-chip (chromatin affinity purification coupled with high-density tiling chip) analysis demonstrated that Pmr preferentially binds to horizontally acquired DNA regions. The Pmr binding sites overlapped well with the location of the genes differentially transcribed following pmr disruption on both the plasmid and the chromosome. Our findings indicate that Pmr is a key factor in optimizing gene transcription on pCAR1 and the host chromosome.

  11. Cometabolic degradation kinetics of TCE and phenol by Pseudomonas putida.

    Science.gov (United States)

    Chen, Yan-Min; Lin, Tsair-Fuh; Huang, Chih; Lin, Jui-Che

    2008-08-01

    Modeling of cometabolic kinetics is important for better understanding of degradation reaction and in situ application of bio-remediation. In this study, a model incorporated cell growth and decay, loss of transformation activity, competitive inhibition between growth substrate and non-growth substrate and self-inhibition of non-growth substrate was proposed to simulate the degradation kinetics of phenol and trichloroethylene (TCE) by Pseudomonas putida. All the intrinsic parameters employed in this study were measured independently, and were then used for predicting the batch experimental data. The model predictions conformed well to the observed data at different phenol and TCE concentrations. At low TCE concentrations (TCE concentrations (>6 mg l(-1)), only the model considering self-inhibition can describe the experimental data, suggesting that a self-inhibition of TCE was present in the system. The proposed model was also employed in predicting the experimental data conducted in a repeated batch reactor, and good agreements were observed between model predictions and experimental data. The results also indicated that the biomass loss in the degradation of TCE below 2 mg l(-1) can be totally recovered in the absence of TCE for the next cycle, and it could be used for the next batch experiment for the degradation of phenol and TCE. However, for higher concentration of TCE (>6 mg l(-1)), the recovery of biomass may not be as good as that at lower TCE concentrations.

  12. Enzymatic Production of l-Citrulline by Pseudomonas putida

    Science.gov (United States)

    Kakimoto, Toshio; Shibatani, Takeji; Nishimura, Noriyuki; Chibata, Ichiro

    1971-01-01

    To develop an efficient method for the production of l-citrulline, optimum conditions for the conversion of l-arginine to l-citrulline by microbial l-arginine deiminase and for production of the enzyme were studied. A number of micro-organisms were screened to test their ability to form and accumulate l-citrulline from l-arginine. Pseudomonas putida was selected as the best organism. With this organism, enzyme activity as high as 9.20 units per ml could be produced by a shaking culture at 30 C in a medium containing glucose, ammonium phosphate, l-arginine hydrochloride, yeast extract, peptone, and inorganic salts. Appropriate addition of a surface active agent to the reaction mixture was found to shorten the time required for the conversion. A large amount of l-arginine hydrochloride was converted stoichiometrically to l-citrulline in 62 hr at 37 C. Accumulated l-citrulline was readily isolated in pure form by ordinary procedures with ion-exchange resins. Yields of isolated l-citrulline of over 90.5% from l-arginine hydrochloride were easily attainable. PMID:5137589

  13. Characterization of Pseudomonas putida Genes Responsive to Nutrient Limitation

    Energy Technology Data Exchange (ETDEWEB)

    Syn, Chris K.; Magnuson, Jon K.; Kingsley, Mark T.; Swarup, Sanjay

    2004-06-01

    The low bioavailability of nutrients and oxygen in the soil environment has hampered successful expression of biodegradation/biocontrol genes that are driven by promoters highly active during routine laboratory conditions of high nutrient- and oxygen-availability. Hence, in the present study, expression of the gus-tagged genes in 12 Tn5-gus mutants of the soil microbe Pseudomonas putida PNL-MK25 was examined under various conditions chosen to mimic the soil environment: low carbon, phosphate, nitrate, or oxygen, and in the rhizosphere. Based on their expression profiles, three nutrient-responsive mutant (NRM) strains, NRM5, NRM7, and NRM17, were selected for identification of the tagged genes. In the mutant strain NRM5, expression of the glutamate dehydrogenase (gdhA) gene was increased between 4.9- to 26.4-fold under various low nutrient conditions. In NRM7, expression of the novel NADPH:quinone oxidoreductase-like (nql) gene was consistently amongst the highest and was synergistically upregulated by low nutrient and anoxic conditions. The cyoD gene in NRM17, which encodes the fourth subunit of the cytochrome o ubiquinol oxidase complex, had decreased expression in low nutrient conditions but its absolute expression levels was still amongst the highest. Additionally, it was independent of oxygen availability, in contrast to that in E. coli.

  14. Marine Pseudomonas putida: a potential source of antimicrobial substances against antibiotic-resistant bacteria

    Directory of Open Access Journals (Sweden)

    Palloma Rodrigues Marinho

    2009-08-01

    Full Text Available Bacteria isolated from marine sponges found off the coast of Rio de Janeiro, Brazil, were screened for the production of antimicrobial substances. We report a new Pseudomonas putida strain (designated P. putida Mm3 isolated from the sponge Mycale microsigmatosa that produces a powerful antimicrobial substance active against multidrug-resistant bacteria. P. putida Mm3 was identified on the basis of 16S rRNA gene sequencing and phenotypic tests. Molecular typing for Mm3 was performed by RAPD-PCR and comparison of the results to other Pseudomonas strains. Our results contribute to the search for new antimicrobial agents, an important strategy for developing alternative therapies to treat infections caused by multidrug-resistant bacteria.

  15. Optimization Production of Biosurfactant by Pseudomonas putida Using Crude Palm Oil (CPO) as Substrate

    Science.gov (United States)

    Suryanti, V.; Handayani, D. S.; Masykur, A.; Lindasari

    2017-07-01

    The production of biosurfactant by Pseudomonas putida has been studied. P. putida FNCC 0071 was grown in the nutrient broth medium supplemented with NaCl and crude palm oil (CPO). The effect of CPO concentration and fermentation time on the biosurfactant production were evaluated. The biosurfactant production was evaluated every 24 h for 10 days by optical density, surface tension and emulsification index. The best culture medium was found to be medium containing 5% v/v of CPO with 5 days of incubation time. The biosurfactant was identified as rhamnolipids.

  16. Influence of growth stage on activities of polyhydroxyalkanoate (PHA polymerase and PHA depolymerase in Pseudomonas putida U

    Directory of Open Access Journals (Sweden)

    Zinn Manfred

    2010-10-01

    Full Text Available Abstract Background Medium chain length (mcl- polyhydroxyalkanoates (PHA are synthesized by many bacteria in the cytoplasm as storage compounds for energy and carbon. The key enzymes for PHA metabolism are PHA polymerase (PhaC and depolymerase (PhaZ. Little is known of how mcl-PHA accumulation and degradation are controlled. It has been suggested that overall PHA metabolism is regulated by the β-oxidation pathway of which the flux is governed by intracellular ratios of [NADH]/[NAD] and [acetyl-CoA]/[CoA]. Another level of control could relate to modulation of the activities of PhaC and PhaZ. In order to investigate the latter, assays for in vitro activity measurements of PhaC and PhaZ in crude cell extracts are necessary. Results Two in vitro assays were developed which allow the measurement of PhaC and PhaZ activities in crude cell extracts of Pseudomonas putida U. Using the assays, it was demonstrated that the activity of PhaC decreased 5-fold upon exponential growth on nitrogen limited medium and octanoate. In contrast, the activity of PhaZ increased only 1.5-fold during growth. One reason for the changes in the enzymatic activity of PhaC and PhaZ could relate to a change in interaction with the phasin surface proteins on the PHA granule. SDS-PAGE analysis of isolated PHA granules demonstrated that during growth, the ratio of [phasins]/[PHA] decreased. In addition, it was found that after eliminating phasins (PhaF and PhaI from the granules PhaC activity decreased further. Conclusion Using the assays developed in this study, we followed the enzymatic activities of PhaC and PhaZ during growth and correlated them to the amount of phasins on the PHA granules. It was found that in P. putida PhaC and PhaZ are concomitantly active, resulting in parallel synthesis and degradation of PHA. Moreover PhaC activity was found to be decreased, whereas PhaZ activity increased during growth. Availability of phasins on PHA granules affected the activity of

  17. Metabolite profiling reveals abiotic stress tolerance in Tn5 mutant of Pseudomonas putida.

    Directory of Open Access Journals (Sweden)

    Vasvi Chaudhry

    Full Text Available Pseudomonas is an efficient plant growth-promoting rhizobacteria (PGPR; however, intolerance to drought and high temperature limit its application in agriculture as a bioinoculant. Transposon 5 (Tn5 mutagenesis was used to generate a stress tolerant mutant from a PGPR Pseudomonas putida NBRI1108 isolated from chickpea rhizosphere. A mutant NBRI1108T, selected after screening of nearly 10,000 transconjugants, exhibited significant tolerance towards high temperature and drought. Southern hybridization analysis of EcoRI and XhoI restricted genomic DNA of NBRI1108T confirmed that it had a single Tn5 insertion. The metabolic changes in the polar and non-polar extracts of NBRI1108 and NBRI1108T were examined using 1H, 31P nuclear magnetic resonance (NMR spectroscopy and gas chromatography-mass spectrometry (GC-MS. Thirty six chemically diverse metabolites consisting of amino acids, fatty acids and phospholipids were identified and quantified. Insertion of Tn5 influenced amino acid and phospholipid metabolism and resulted in significantly higher concentration of aspartic acid, glutamic acid, glycinebetaine, glycerophosphatidylcholine (GPC and putrescine in NBRI1108T as compared to that in NBRI1108. The concentration of glutamic acid, glycinebetaine and GPC increased by 34%, 95% and 100%, respectively in the NBRI1108T as compared to that in NBRI1108. High concentration of glycerophosphatidylethanolamine (GPE and undetected GPC in NBRI1108 indicates that biosynthesis of GPE may have taken place via the methylation pathway of phospholipid biosynthesis. However, high GPC and low GPE concentration in NBRI1108T suggest that methylation pathway and phosphatidylcholine synthase (PCS pathway of phospholipid biosynthesis are being followed in the NBRI1108T. Application of multivariate principal component analysis (PCA on the quantified metabolites revealed clear variations in NBRI1108 and NBRI1108T in polar and non-polar metabolites. Identification of abiotic

  18. Metabolite profiling reveals abiotic stress tolerance in Tn5 mutant of Pseudomonas putida.

    Science.gov (United States)

    Chaudhry, Vasvi; Bhatia, Anil; Bharti, Santosh Kumar; Mishra, Shashank Kumar; Chauhan, Puneet Singh; Mishra, Aradhana; Sidhu, Om Prakash; Nautiyal, Chandra Shekhar

    2015-01-01

    Pseudomonas is an efficient plant growth-promoting rhizobacteria (PGPR); however, intolerance to drought and high temperature limit its application in agriculture as a bioinoculant. Transposon 5 (Tn5) mutagenesis was used to generate a stress tolerant mutant from a PGPR Pseudomonas putida NBRI1108 isolated from chickpea rhizosphere. A mutant NBRI1108T, selected after screening of nearly 10,000 transconjugants, exhibited significant tolerance towards high temperature and drought. Southern hybridization analysis of EcoRI and XhoI restricted genomic DNA of NBRI1108T confirmed that it had a single Tn5 insertion. The metabolic changes in the polar and non-polar extracts of NBRI1108 and NBRI1108T were examined using 1H, 31P nuclear magnetic resonance (NMR) spectroscopy and gas chromatography-mass spectrometry (GC-MS). Thirty six chemically diverse metabolites consisting of amino acids, fatty acids and phospholipids were identified and quantified. Insertion of Tn5 influenced amino acid and phospholipid metabolism and resulted in significantly higher concentration of aspartic acid, glutamic acid, glycinebetaine, glycerophosphatidylcholine (GPC) and putrescine in NBRI1108T as compared to that in NBRI1108. The concentration of glutamic acid, glycinebetaine and GPC increased by 34%, 95% and 100%, respectively in the NBRI1108T as compared to that in NBRI1108. High concentration of glycerophosphatidylethanolamine (GPE) and undetected GPC in NBRI1108 indicates that biosynthesis of GPE may have taken place via the methylation pathway of phospholipid biosynthesis. However, high GPC and low GPE concentration in NBRI1108T suggest that methylation pathway and phosphatidylcholine synthase (PCS) pathway of phospholipid biosynthesis are being followed in the NBRI1108T. Application of multivariate principal component analysis (PCA) on the quantified metabolites revealed clear variations in NBRI1108 and NBRI1108T in polar and non-polar metabolites. Identification of abiotic stress

  19. Proteins with GGDEF and EAL domains regulate Pseudomonas putida biofilm formation and dispersal

    DEFF Research Database (Denmark)

    Gjermansen, Morten; Ragas, Paula Cornelia; Tolker-Nielsen, Tim

    2006-01-01

    H protein, which contains an EAL domain, strongly inhibited biofilm formation. Induction of YhjH expression in P. putida cells situated in established biofilms led to rapid dispersion of the biofilms. These results support the emerging theme that GGDEF-domain and EAL-domain proteins are involved...

  20. Active efflux systems in the solvent-tolerant bacterium Pseudomonas putida S12

    NARCIS (Netherlands)

    Kieboom, J.

    2002-01-01

    The aim of the research presented in this thesis was to study the molecular mechanisms of organic solvent tolerance in Pseudomonas putida S12. This bacterium is capable of growth at saturated solvent concentrations, which are lethal to normal bacteria. Organic solve

  1. Bioremediation of Chromium (VI from Textile Industry’s Effluent and Contaminated Soil Using Pseudomonas putida

    Directory of Open Access Journals (Sweden)

    Deepali

    2011-01-01

    Full Text Available Nine bacterial colonies were screened for the Cr(VI removal efficiency and out of these three bacterial strains Pseudomonas putida, Pseudomonas aeruginosa and Bacillus sp. were isolated from soil and used to remove Cr(VI from aqueous solution. The effect of time and concentrations on the removal rate of hexavalent chromium were studied using batch experiment. Maximum Cr (VI removal was noted 75.0% by Bacillus sp. at 10mg/l, 69.70% by Pseudomonas aeruginosa at 40mg/l and 90.88% by Pseudomonas putida at 10mg/l of synthetic solution, during 96 hours. Among these three bacteria, the maximum Cr(VI removal was reported by Pseudomonas putida on lower concentration. On the basis of highest removal rate, Pseudomonas putida was selected and used for further chromium removal from samples. It was found to be removed the highest Cr(VI by 82.92%, from effluent and 74.41% from soil during 96 hours. The present study depicts that bacteria removes chromium efficiently and this could be used for industrial waste management and other environmental contaminants.

  2. Genome sequence of the plant growth-promoting rhizobacterium Pseudomonas putida S11.

    Science.gov (United States)

    Ponraj, Paramasivan; Shankar, Manoharan; Ilakkiam, Devaraj; Rajendhran, Jeyaprakash; Gunasekaran, Paramasamy

    2012-11-01

    Here we report the genome sequence of a plant growth-promoting rhizobacterium, Pseudomonas putida S11. The length of the draft genome sequence is approximately 5,970,799 bp, with a G+C content of 62.4%. The genome contains 6,076 protein-coding sequences.

  3. Effect of culture medium on biocalcification by Pseudomonas Putida, Lysinibacillus Sphaericus and Bacillus Subtilis

    Directory of Open Access Journals (Sweden)

    Márcia Aiko Shirakawa

    2011-06-01

    Full Text Available The objective of this study is to investigate the efficiency of calcium carbonate bioprecipitation by Lysinibacillus sphaericus, Bacillus subtilis and Pseudomonas putida, obtained from the Coleção de Culturas do Instituto Nacional de Controle de Qualidade em Saúde (INCQS, as a first step in determining their potential to protect building materials against water uptake. Two culture media were studied: modified B4 containing calcium acetate and 295 with calcium chloride. Calcium consumption in the two media after incubation with and without the bacterial inoculum was determined by atomic absorption analysis. Modified B4 gave the best results and in this medium Pseudomonas putida INQCS 113 produced the highest calcium carbonate precipitation, followed by Lysinibacillus sphaericus INQCS 414; the lowest precipitation was produced by Bacillus subtilis INQCS 328. In this culture medium XRD analysis showed that Pseudomonas putida and Bacillus subtilis precipitated calcite and vaterite polymorphs while Lysinibacillus sphaericus produced only vaterite. The shape and size of the crystals were affected by culture medium, bacterial strain and culture conditions, static or shaken. In conclusion, of the three strains Pseudomonas putida INQCS 113 in modified B4 medium gave the best results precipitating 96% of the calcium, this strain thus has good potential for use on building materials.

  4. Effect of culture medium on biocalcification by Pseudomonas Putida, Lysinibacillus Sphaericus and Bacillus Subtilis.

    Science.gov (United States)

    Shirakawa, Márcia Aiko; Cincotto, Maria Alba; Atencio, Daniel; Gaylarde, Christine C; John, Vanderley M

    2011-04-01

    The objective of this study is to investigate the efficiency of calcium carbonate bioprecipitation by Lysinibacillus sphaericus, Bacillus subtilis and Pseudomonas putida, obtained from the Coleção de Culturas do Instituto Nacional de Controle de Qualidade em Saúde (INCQS), as a first step in determining their potential to protect building materials against water uptake. Two culture media were studied: modified B4 containing calcium acetate and 295 with calcium chloride. Calcium consumption in the two media after incubation with and without the bacterial inoculum was determined by atomic absorption analysis. Modified B4 gave the best results and in this medium Pseudomonas putida INQCS 113 produced the highest calcium carbonate precipitation, followed by Lysinibacillus sphaericus INQCS 414; the lowest precipitation was produced by Bacillus subtilis INQCS 328. In this culture medium XRD analysis showed that Pseudomonas putida and Bacillus subtilis precipitated calcite and vaterite polymorphs while Lysinibacillus sphaericus produced only vaterite. The shape and size of the crystals were affected by culture medium, bacterial strain and culture conditions, static or shaken. In conclusion, of the three strains Pseudomonas putida INQCS 113 in modified B4 medium gave the best results precipitating 96% of the calcium, this strain thus has good potential for use on building materials.

  5. Regulation of the biosynthesis of cyclic lipopeptides from Pseudomonas putida PCL1445

    NARCIS (Netherlands)

    Dubern, Jean-Frédéric

    2006-01-01

    Pseudomonas putida strain PCL1445 produces two cyclic lipopeptides, named putisolvins I and II, which represent a Novel class of biosurfactants. Putisolvins reduce the surface tension between liquid and air, and disrupt already existing biofilms of several Pseudomonas sp., including those of the

  6. Effect of Pseudomonas putida on Growth and Anthocyanin Pigment in Two Poinsettia (Euphorbia pulcherrima Cultivars

    Directory of Open Access Journals (Sweden)

    Ramon Zulueta-Rodriguez

    2014-01-01

    Full Text Available Pseudomonas putida is plant growth promoting rhizobacteria (PGPR that have the capacity to improve growth in plants. The purpose of this study was to determine growth and anthocyanin pigmentation of the bracts in two poinsettia Euphorbia pulcherrima cultivars (Prestige and Sonora Marble using three strains of P. putida, as well as a mixture of the three (MIX. Comparison with the control group indicated for the most part that Prestige grew better than the Sonora Marble cultivars with the PGPR strains. Prestige with the MIX strain grew better compared to control for the number of cyathia (83 versus 70.4, volume of roots (45 versus 35 cm3, number of leaves (78 versus 58, and area of leaf (1,788 versus 1,331 cm2, except for the number of flowers (8.8 versus 11.6. To the naked eye, coloration of plants appeared identical in color compared to the control group. For all plants with P. putida strains, there was less anthocyanin pigment, but biomass was always greater with PGPR strains. Nevertheless, to the naked eye, the coloration of the plants appeared identical in color compared to the control group. This is the first study reporting the positive effects of P. putida rhizobacteria treatments on growth of poinsettia cultivars.

  7. Biological production of hydroxylated aromatics: Optimization strategies for Pseudomonas putida S12

    NARCIS (Netherlands)

    Verhoef, A.

    2010-01-01

    To replace environmentally unfriendly petrochemical production processes, the demand for bio-based production of organic chemicals is increasing. This thesis focuses on the biological production of hydroxylated aromatics from renewable substrates by engineered P. putida S12 including several cases o

  8. Redundancy in putrescine catabolism in solvent tolerant Pseudomonas putida S12

    NARCIS (Netherlands)

    Bandounas, L.; Ballerstedt, H.; Winde, J.H. de; Ruijssenaars, H.J.

    2011-01-01

    Pseudomonas putida S12 is a promising platform organism for the biological production of substituted aromatic compounds due to its extreme tolerance towards toxic chemicals. Solvent or aromatic stress tolerance may be due to membrane modifications and efflux pumps; however in general, polyamines hav

  9. Engineering Pseudomonas putida S12 for efficient utilization of D-Xylose and L-Arabinose

    NARCIS (Netherlands)

    Meijnen, J.P.; Winde, J.H. de; Ruijssenaars, H.J.

    2008-01-01

    The solvent-tolerant bacterium Pseudomonas putida S12 was engineered to utilize xylose as a substrate by expressing xylose isomerase (XylA) and xylulokinase (XylB) from Escherichia coli. The initial yield on xylose was low (9% [g CDW g substrate−1], where CDW is cell dry weight), and the growth rate

  10. Regulation of the biosynthesis of cyclic lipopeptides from Pseudomonas putida PCL1445

    NARCIS (Netherlands)

    Dubern, Jean-Frédéric

    2006-01-01

    Pseudomonas putida strain PCL1445 produces two cyclic lipopeptides, named putisolvins I and II, which represent a Novel class of biosurfactants. Putisolvins reduce the surface tension between liquid and air, and disrupt already existing biofilms of several Pseudomonas sp., including those of the opp

  11. C1 compounds as auxiliary substrate for engineered Pseudomonas putida S12

    NARCIS (Netherlands)

    Koopman, F.W.; Winde, J.H. de; Ruijssenaars, H.J.

    2009-01-01

    The solvent-tolerant bacterium Pseudomonas putida S12 was engineered to efficiently utilize the C1 compounds methanol and formaldehyde as auxiliary substrate. The hps and phi genes of Bacillus brevis, encoding two key steps of the ribulose monophosphate (RuMP) pathway, were introduced to construct a

  12. Active efflux systems in the solvent-tolerant bacterium Pseudomonas putida S12

    NARCIS (Netherlands)

    Kieboom, J.

    2002-01-01

    The aim of the research presented in this thesis was to study the molecular mechanisms of organic solvent tolerance in Pseudomonas putida S12. This bacterium is capable of growth at saturated solvent concentrations, which are lethal to normal bacteria. Organic

  13. Redundancy in putrescine catabolism in solvent tolerant Pseudomonas putida S12

    NARCIS (Netherlands)

    Bandounas, L.; Ballerstedt, H.; Winde, J.H. de; Ruijssenaars, H.J.

    2011-01-01

    Pseudomonas putida S12 is a promising platform organism for the biological production of substituted aromatic compounds due to its extreme tolerance towards toxic chemicals. Solvent or aromatic stress tolerance may be due to membrane modifications and efflux pumps; however in general, polyamines hav

  14. Functional, genetic and chemical characterization of biosurfactants produced by plant growth-promoting Pseudomonas putida 267

    NARCIS (Netherlands)

    Kruijt, M.; Tran, H.; Raaijmakers, J.M.

    2009-01-01

    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 identif

  15. Biosorption of aluminum through the use of non-viable biomass of Pseudomonas putida.

    Science.gov (United States)

    Boeris, Paola Sabrina; Agustín, María Del Rosario; Acevedo, Diego Fernando; Lucchesi, Gloria Inés

    2016-10-20

    Living and non-living biomass of Pseudomonas putida A (ATCC 12633) was used as biosorbent for the removing of Al(3+) from aqueous solutions. The process was stable with time, efficient at pH 4.3 and between 15°C and 42°C. Two isotherms models were applied to describe the interaction between the biosorbent and Al(3+). Non-living biomass of P. putida A (ATCC 12633) was found to be the most efficient at adsorbing Al(3+) with a maximum sorption capacity of 0.55mg Al(3+)/gr adsorbent and with 36×10(5) binding sites of Al(3+)/microorganisms. Infrared spectroscopy analysis shows that the biosorbent present some vibrational band of functional groups that change in presence of Al(3+): hydroxyl, carboxyl and phosphate. Considering that Al(3+) binds to the phosphate group of phosphatidylcholine, non-viable biomass of P. putida PB01 (mutant lacking phosphatidylcholine) was used. Aluminum adsorption of the parental strain was 30 times higher than values registered in P. putida PB01 (36×10(5) sites/microorganism vs 1.2×10(5) sites/microorganism, respectively). This result evidenced that the absence of phosphatidylcholine significantly affected the availability of the binding sites and consequently the efficiency of the biomass to adsorb Al(3+).

  16. Rhizoplane colonisation of peas by Rhizobium leguminosarum bv. viceae and a deleterious Pseudomonas putida

    NARCIS (Netherlands)

    Berggren, I.; Alstrom, S.; Vuurde, van J.W.L.; Martensson, A.M.

    2005-01-01

    Pseudomonas putida strain angstrom 313, a deleterious rhizosphere bacterium, reduced pea nitrogen content when inoculated alone or in combination with Rhizobium leguminosarum bv. viceae on plants in the presence of soil under greenhouse conditions. When plants were grown gnotobiotically in liquid me

  17. Interaction of Pseudomonas putida ATCC 12633 and Bacteriophage gh-1 in Berea Sandstone Rock.

    Science.gov (United States)

    Chang, P L; Yen, T F

    1985-12-01

    Measurements of the passage of Pseudomonas putida ATCC 12633 and a phage-resistant mutant through Berea sandstone rock were made. When bacteriophage gh-1 was adsorbed within the rock matrix, a reduction in the passage of the susceptible but not the resistant cells through the rock was observed.

  18. Effect of culture medium on biocalcification by Pseudomonas Putida, Lysinibacillus Sphaericus and Bacillus Subtilis

    Science.gov (United States)

    Shirakawa, Márcia Aiko; Cincotto, Maria Alba; Atencio, Daniel; Gaylarde, Christine C.; John, Vanderley M.

    2011-01-01

    The objective of this study is to investigate the efficiency of calcium carbonate bioprecipitation by Lysinibacillus sphaericus, Bacillus subtilis and Pseudomonas putida, obtained from the Coleção de Culturas do Instituto Nacional de Controle de Qualidade em Saúde (INCQS), as a first step in determining their potential to protect building materials against water uptake. Two culture media were studied: modified B4 containing calcium acetate and 295 with calcium chloride. Calcium consumption in the two media after incubation with and without the bacterial inoculum was determined by atomic absorption analysis. Modified B4 gave the best results and in this medium Pseudomonas putida INQCS 113 produced the highest calcium carbonate precipitation, followed by Lysinibacillus sphaericus INQCS 414; the lowest precipitation was produced by Bacillus subtilis INQCS 328. In this culture medium XRD analysis showed that Pseudomonas putida and Bacillus subtilis precipitated calcite and vaterite polymorphs while Lysinibacillus sphaericus produced only vaterite. The shape and size of the crystals were affected by culture medium, bacterial strain and culture conditions, static or shaken. In conclusion, of the three strains Pseudomonas putida INQCS 113 in modified B4 medium gave the best results precipitating 96% of the calcium, this strain thus has good potential for use on building materials. PMID:24031661

  19. Using "Pseudomonas Putida xylE" Gene to Teach Molecular Cloning Techniques for Undergraduates

    Science.gov (United States)

    Dong, Xu; Xin, Yi; Ye, Li; Ma, Yufang

    2009-01-01

    We have developed and implemented a serial experiment in molecular cloning laboratory course for undergraduate students majored in biotechnology. "Pseudomonas putida xylE" gene, encoding catechol 2, 3-dioxygenase, was manipulated to learn molecular biology techniques. The integration of cloning, expression, and enzyme assay gave students…

  20. DETECTION OF PHENOL DEGRADING BACTERIA AND PSEUDOMONAS PUTIDA IN ACTIVATED SLUDGE BY POLYMERASE CHAIN REACTION

    Directory of Open Access Journals (Sweden)

    H. Movahedyan ، H. Khorsandi ، R. Salehi ، M. Nikaeen

    2009-04-01

    Full Text Available Phenol is one of the organic pollutants in various industrial wastewaters especially petrochemical and oil refining. Biological treatment is one of the considerable choices for removing of phenol present in these wastewaters. Identification of effective microbial species is considered as one of the important priorities for production of the biomass in order to achieve desirable kinetic of biological reactions. Basic purpose of this research is identification of phenol-degrading Pseudomonas Putida in activated sludge by polymerase chain reaction (PCR that has high speed and specificity. In this research, 10 various colonies of phenol-degrading bacteria were isolated from municipal activated sludge and the rate of phenol removal and growth rate of these bacteria were assessed in different concentrations of phenol (200 – 900 mg/L. Confirmation of the largest subunit of multicomponent phenol hydroxylase (LmPH gene and gene coding the N fragment in Pseudomonas Putida-derived methyl phenol operon (DmpN gene through PCR were used for general identification of phenol-degrading bacteria and Pseudomonas Putida, respectively. Presence of a 600 bp (base pairs bond in all of isolated strains indicated that they contain phenol hydroxylase gene. 6 of 10 isolated bacteria were Pseudomonas Putida because they produced a 199 bp PCR product by DmpN primers. According to PCR results in this study, the best phenol-degrading bacteria that can utilize 500 – 600 mg/L phenol completely after 48 hours incubation, belong to Pseudomonas Putida strains. It is clear that use of isolated bacteria can lead to considerable decrease of treatment time as well as promotion of phenol removal rate.

  1. Accurate determination of plasmid copy number of flow-sorted cells using droplet digital PCR.

    Science.gov (United States)

    Jahn, Michael; Vorpahl, Carsten; Türkowsky, Dominique; Lindmeyer, Martin; Bühler, Bruno; Harms, Hauke; Müller, Susann

    2014-06-17

    Many biotechnological processes rely on the expression of a plasmid-based target gene. A constant and sufficient number of plasmids per cell is desired for efficient protein production. To date, only a few methods for the determination of plasmid copy number (PCN) are available, and most of them average the PCN of total populations disregarding heterogeneous distributions. Here, we utilize the highly precise quantification of DNA molecules by droplet digital PCR (ddPCR) and combine it with cell sorting using flow cytometry. A duplex PCR assay was set up requiring only 1000 sorted cells for precise determination of PCN. The robustness of this method was proven by thorough optimization of cell sorting, cell disruption, and PCR conditions. When non plasmid-harboring cells of Pseudomonas putida KT2440 were spiked with different dilutions of the expression plasmid pA-EGFP_B, a PCN from 1 to 64 could be accurately detected. As a proof of principle, induced cultures of P. putida KT2440 producing an EGFP-fused model protein by means of the plasmid pA-EGFP_B were investigated by flow cytometry and showed two distinct subpopulations, fluorescent and nonfluorescent cells. These two subpopulations were sorted for PCN determination with ddPCR. A remarkably diverging plasmid distribution was found within the population, with nonfluorescent cells showing a much lower PCN (≤1) than fluorescent cells (PCN of up to 5) under standard conditions.

  2. Simultaneous removal of phosphorus and nitrogen in a sequencing batch biofilm reactor with transgenic bacteria expressing polyphosphate kinase.

    Science.gov (United States)

    Du, Hongwei; Yang, Liuyan; Wu, Jun; Xiao, Lin; Wang, Xiaolin; Jiang, Lijuan

    2012-10-01

    To improve phosphorus removal from wastewater, we constructed a high-phosphate-accumulating microorganism, KTPPK, using Pseudomonas putida KT2440 as a host. The expression plasmid was constructed by inserting and expressing polyphosphate kinase gene (ppk) from Microcystis aeruginosa NIES-843 into broad-host-range plasmid, pBBR1MCS-2. KTPPK was then added to a sequencing batch biofilm reactor (SBBFR) using lava as a biological carrier. The results showed that SBBFR with KTPPK not only efficiently removed COD, NH(3)-N, and NO(3)(-)-N but also had a high removal capacity for PO(4)(3-)-P, resulting in a low phosphorus concentration remaining in the outflow of the SBBFR. The biofilm increased by 30-53% on the lava in the SBBFR that contained KTPPK after 11 days when compared with the reactor that contained P. putida KT2440. Real-time quantitative polymerase chain reaction confirmed that the copy of ppk was maintained at about 3.5 × 10(10) copies per μL general DNA in the biofilm after 20 days. Thus, the transgenic bacteria KTPPK could maintain a high density and promote phosphorus removal in the SBBFR. In summary, this study indicates that the use of SBBFR with transgenic bacteria has the potential to remove phosphorus and nitrogen from wastewater.

  3. Base-Catalyzed Depolymerization of Solid Lignin-Rich Streams Enables Microbial Conversion

    Energy Technology Data Exchange (ETDEWEB)

    Beckham, Gregg T [National Renewable Energy Laboratory (NREL), Golden, CO (United States); Salvachua Rodriguez, Davinia [National Renewable Energy Laboratory (NREL), Golden, CO (United States); Katahira, Rui [National Renewable Energy Laboratory (NREL), Golden, CO (United States); Pleitner, Brenna P [National Renewable Energy Laboratory (NREL), Golden, CO (United States); Cleveland, Nicholas S [National Renewable Energy Laboratory (NREL), Golden, CO (United States); Nolker, Michelle L [National Renewable Energy Laboratory (NREL), Golden, CO (United States); Smith, Holly K [National Renewable Energy Laboratory (NREL), Golden, CO (United States); Rodriguez, Alberto [Sandia National Laboratories; Baidoo, Edward E. K. [Lawrence Berkeley National Laboratory; DOE Joint BioEnergy Institute; Keasling, Jay D. [Lawrence Berkeley National Laboratory; DOE Joint BioEnergy InstituteUniversity of California, Berkeley; Technical University of Denmark; Simmons, Blake A. [Sandia National Laboratories; Lawrence Berkeley National Laboratory; DOE Joint BioEnergy Institute; Technical University of Denmark; Gladden, John M. [Sandia National Laboratories; DOE Joint BioEnergy Institute

    2017-08-01

    Lignin valorization offers significant potential to enhance the economic viability of lignocellulosic biorefineries. However, because of its heterogeneous and recalcitrant nature, conversion of lignin to value-added coproducts remains a considerable technical challenge. In this study, we employ base-catalyzed depolymerization (BCD) using a process-relevant solid lignin stream produced via deacetylation, mechanical refining, and enzymatic hydrolysis to enable biological lignin conversion. BCD was conducted with the solid lignin substrate over a range of temperatures at two NaOH concentrations, and the results demonstrate that the lignin can be partially extracted and saponified at temperatures as low as 60 degrees C. At 120 degrees C and 2% NaOH, the high extent of lignin solubility was accompanied by a considerable decrease in the lignin average molecular weight and the release of lignin-derived monomers including hydroxycinnamic acids. BCD liquors were tested for microbial growth using seven aromatic-catabolizing bacteria and two yeasts. Three organisms (Pseudomonas putida KT2440, Rhodotorula mucilaginosa, and Corynebacterium glutamicum) tolerate high BCD liquor concentrations (up to 90% v/v) and rapidly consume the main lignin-derived monomers, resulting in lignin conversion of up to 15%. Furthermore, as a proof of concept, muconic acid production from a representative lignin BCD liquor was demonstrated with an engineered P. putida KT2440 strain. These results highlight the potential for a mild lignin depolymerization process to enhance the microbial conversion of solid lignin-rich biorefinery streams.

  4. Evaluation of genotoxicity and pro-oxidant effect of the azo dyes: acids yellow 17, violet 7 and orange 52, and of their degradation products by Pseudomonas putida mt-2.

    Science.gov (United States)

    Ben Mansour, Hedi; Corroler, David; Barillier, Daniel; Ghedira, Kamel; Chekir, Leila; Mosrati, Ridha

    2007-09-01

    Acids yellow 17, violet 7 and orange 52, very important commercial azo dyes used in the textile, food, paper and cosmetic industries, were degraded by Pseudomonas putida mt-2 at concentrations up to 100mg/l. The culture media was completely decolorized under static incubation for 60 h, this faster than under continuous shaking incubation. SOS chromotest using Escherichia coli PQ37, with and without metabolic activation (S-9 preparations), was used to assess genotoxicity potential of these dyes before and after biodegradation. None of these dyes or their metabolites was found to be genotoxic in the absence of "Araclor-Induced rat liver microsome" preparations (S-9). However, in presence of the preparation S-9, the genotoxicity of the biodegradation products was highlighted. Metabolites resulting from static cultures were more genotoxic than those obtained in shaken conditions. In addition to genotoxic effects, metabolites have shown a significant ability to induce the formation of superoxide free radical anion (O(2)(*-)). The toxicities generated by the pure azo dyes and the pure azo-reduction products (sulfanilic acid, N,N'-dimethyl-p-phenylenediamine and 4'-aminoacetanilid) were compared. These results suggest that P. putida mt-2 degrades the studied azo dyes in two steps: an azo-reduction followed by an oxygen-dependent metabolization. Some of the derived metabolites would be responsible of genotoxicity and metabolic toxicity.

  5. Attachment of Pseudomonas putida onto differently structured kaolinite minerals: a combined ATR-FTIR and 1H NMR study.

    Science.gov (United States)

    Vasiliadou, Ioanna A; Papoulis, Dimitris; Chrysikopoulos, Constantinos V; Panagiotaras, Dionisios; Karakosta, Eleni; Fardis, Michael; Papavassiliou, Georgios

    2011-06-01

    The attachment of Pseudomonas (P.) putida onto well (KGa-1) and poorly (KGa-2) crystallized kaolinite was investigated in this study. Batch experiments were carried out to determine the attachment isotherms of P. putida onto both types of kaolinite particles. The attachment process of P. putida onto KGa-1 and KGa-2 was adequately described by a Langmuir isotherm. Attenuated Total Reflection Fourier Transform Infrared Spectroscopy and Nuclear Magnetic Resonance were employed to study the attachment mechanisms of P. putida. Experimental results indicated that KGa-2 presented higher affinity and attachment capacity than KGa-1. It was shown that electrostatic interactions and clay mineral structural disorders can influence the attachment capacity of clay mineral particles.

  6. Activity of toluene-degrading Pseudomonas putida in the early growth phase of a biofilm for waste gas treatment

    DEFF Research Database (Denmark)

    Pedersen, A.R.; Møller, S.; Molin, S.

    1997-01-01

    A biological trickling filter for treatment of toluene-containing waste gas was studied. The overall kinetics of the biofilm growth was followed in the early growth phase. A rapid initial colonization took place during the first three days. The biofilm thickness increased exponentially, whereas...... the increase of active biomass and polymers was linear. In order to investigate the toluene degradation, various toluene degraders from the multispecies biofilm were isolated, and a Pseudomonas putida was chosen as a representative of the toluene-degrading population. A specific rRNA oligonucleotide probe...... was used to follow the toluene-degrading P. putida in the multispecies biofilm in the filter by means of number and cellular rRNA content. P. putida appeared to detach from the biofilm during the first three days of growth, after which P. putida was found at a constant level of 10% of the active biomass...

  7. The solvent-tolerant Pseudomonas putida S12 as host for the production of cinnamic acid from glucose

    NARCIS (Netherlands)

    Nijkamp, K.; Luijk, N. van; Bont, J.A.M. de; Wery, J.

    2005-01-01

    A Pseudomonas putida S12 strain was constructed that efficiently produced thefine chemical cinnamic acid from glucose or glycerol via the central metabolite phenylalanine. The gene encoding phenylalanine ammonia lyase from the yeast Rhodosporidium toruloides was introduced. Phenylalanine availabilit

  8. Pseudomonas and Beyond: Polyamine metabolism, lignin degradation and potential applications in industrial biotechnology

    NARCIS (Netherlands)

    Bandounas, L.

    2011-01-01

    Renewable resources such as lignocellulosic biomass are promising feedstocks for the production of bio-fuels and value-added products. Biocatalysts are considered important tools in such processes. Pseudomonas putida S12 has a broad metabolic potential and is exceptionally tolerant towards a range o

  9. Metabolic commensalism and competition in a two-species microbial consortium

    DEFF Research Database (Denmark)

    Christensen, Bjarke Bak; Haagensen, Janus Anders Juul; Heydorn, Arne

    2002-01-01

    We analyzed metabolic interactions and the importance of specific structural relationships in a benzyl alcohol-degrading microbial consortium comprising two species, Pseudomonas putida strain R1 and Acinetobacter strain C6, both of which are able to utilize benzyl alcohol as their sole carbon...... and energy source. The organisms were grown either as surface-attached organisms (biofilms) in How chambers or as suspended cultures in chemostats. The numbers of CFU of P. putida RI and Acinetobacter strain C6 were determined in chemostats and from the effluents of the flow chambers. When the two species...... positioning in the biofilm. In the initial phase of biofilm development, the growth activity of P. putida R1 was shown to be higher near microcolonies of Acinetobacter strain C6. High-pressure liquid chromatography analysis showed that in the effluent of the Acinetobacter strain C6 monoculture biofilm...

  10. Better to light a candle than curse the darkness: illuminating spatial localization and temporal dynamics of rapid microbial growth in the rhizosphere

    Directory of Open Access Journals (Sweden)

    Patrick M Herron

    2013-09-01

    Full Text Available The rhizosphere is a hotbed of microbial activity in ecosystems, fueled by carbon compounds from plant roots. Basic questions about the location and dynamics of plant-spurred microbial growth in the rhizosphere are difficult to answer with standard, destructive soil assays mixing a multitude of microbe-scale microenvironments in a single, often sieved, sample. Soil microbial biosensors designed with the luxCDABE reporter genes fused to a promoter of interest enable continuous imaging of the microbial perception of (and response to environmental conditions in soil. We used the common soil bacterium Pseudomonas putida KT2440 as host to plasmid pZKH2 containing a fusion between the strong constituitive promoter nptII and luxCDABE (coding for light-emitting proteins from Vibrio fischeri. Experiments in liquid media demonstrated that high light production by KT2440/pZKH2 was associated with rapid microbial growth supported by high carbon availability. We applied the biosensors in microcosms filled with non-sterile soil in which corn (Zea mays L., black poplar (Populus nigra L. or tomato (Solanum lycopersicum L. was growing. We detected minimal light production from microbiosensors in the bulk soil, but biosensors reported continuously from around roots for as long as six days. For corn, peaks of luminescence were detected 1-4 and 20-35 mm along the root axis behind growing root tips, with the location of maximum light production moving farther back from the tip as root growth rate increased. For poplar, luminescence around mature roots increased and decreased on a coordinated diel rhythm, but was not bright near root tips. For tomato, luminescence was dynamic, but did not exhibit a diel rhythm, appearing in acropetal waves along roots. KT2440/pZKH2 revealed that root tips are not always the only, or even the dominant, hotspots for rhizosphere microbial growth, and carbon availability is highly variable in space and time around roots.

  11. Synthesis and Physical Properties of Polyhydroxyalkanoate Polymers with Different Monomer Compositions by Recombinant Pseudomonas putida LS46 Expressing a Novel PHA SYNTHASE (PhaC116 Enzyme

    Directory of Open Access Journals (Sweden)

    Parveen K. Sharma

    2017-03-01

    Full Text Available A recombinant of Pseudomonas putida LS461 (deletion of the phaC1phaZphaC2 genes was constructed by introducing cosmid JC123 carrying a novel phaC116 gene from a metagenomic clone. The resulting strain, P. putida LS46123, was able to synthesize polyhydroxyalkanoate (PHA polymers with novel monomer compositions when cultured on glucose or free fatty acids, and accumulated PHAs from 9.24% to 27.09% of cell dry weight. The PHAs synthesized by P. putida LS46123 contained up to 50 mol % short chain length subunits (3-hydroxybutyrate and 3-hydroxyvalerate, with the remaining monomers consisting of various medium chain length subunits. The PhaC116 protein expressed by P. putida LS46123 had an amino acid sequence similarity of 45% with the PhaC1 protein of the parent strain, P. putida LS46. Predicted 3D structures of the PhaC116 proteins from P. putida LS46123 and P. putida LS46 revealed several differences in the numbers and locations of protein secondary structures. The physical and thermal properties of the novel polymers synthesized by P. putida LS46123 cultured with glucose or free fatty acids differed significantly from those produced by P. putida LS46 grown on the same substrates. PHA polymers with different subunit compositions, and hence different physical and thermal properties, can be tailor-made using novel PHA synthase for specific applications.

  12. Induction of the tod Operon by Trichloroethylene in Pseudomonas putida TVA8

    Science.gov (United States)

    Shingleton, Justin T.; Applegate, Bruce M.; Nagel, Aaron C.; Bienkowski, Paul R.; Sayler, Gary S.

    1998-01-01

    Bioluminescence, mRNA levels, and toluene degradation rates in Pseudomonas putida TVA8 were measured as a function of various concentrations of toluene and trichloroethylene (TCE). TVA8 showed an increasing bioluminescence response to increasing TCE and toluene concentrations. Compared to uninduced TVA8 cultures, todC1 mRNA levels increased 11-fold for TCE-treated cultures and 13-fold for toluene-treated cultures. Compared to uninduced P. putida F1 cultures, todC1 mRNA levels increased 4.4-fold for TCE-induced cultures and 4.9-fold for toluene-induced cultures. Initial toluene degradation rates were linearly correlated with specific bioluminescence in TVA8 cultures. PMID:9835608

  13. Aerobic TCE degradation by encapsulated toluene-oxidizing bacteria, Pseudomonas putida and Bacillus spp.

    Science.gov (United States)

    Kim, Seungjin; Bae, Wookeun; Hwang, Jungmin; Park, Jaewoo

    2010-01-01

    The degradation rates of toluene and trichloroethylene (TCE) by Pseudomonas putida and Bacillus spp. that were encapsulated in polyethylene glycol (PEG) polymers were evaluated in comparison with the results of exposure to suspended cultures. PEG monomers were polymerized together with TCE-degrading microorganisms, such that the cells were encapsulated in and protected by the matrices of the PEG polymers. TCE concentrations were varied from 0.1 to 1.5 mg/L. In the suspended cultures of P. putida, the TCE removal rate decreased as the initial TCE concentration increased, revealing TCE toxicity or a limitation of reducing power, or both. When the cells were encapsulated, an initial lag period of about 10-20 h was observed for toluene degradation. Once acclimated, the encapsulated P. putida cultures were more tolerant to TCE at an experimental range of 0.6-1.0 mg/L and gave higher transfer efficiencies (mass TCE transformed/mass toluene utilized). When the TCE concentration was low (e.g., 0.1 mg/L) the removal of TCE per unit mass of cells (specific removal) was significantly lower, probably due to a diffusion limitation into the PEG pellet. Encapsulated Bacillus spp. were able to degrade TCE cometabolically. The encapsulated Bacillus spp. gave significantly higher values than did P. putida in the specific removal and the transfer efficiency, particularly at relatively high TCE concentration of approximately 1.0±0.5 mg/L. The transfer efficiency by encapsulated Bacillus spp. in this study was 0.27 mgTCE/mgToluene, which was one to two orders of magnitude greater than the reported values.

  14. Effect of culture medium on biocalcification by Pseudomonas Putida, Lysinibacillus Sphaericus and Bacillus Subtilis

    OpenAIRE

    Márcia Aiko Shirakawa; Maria Alba Cincotto; Daniel Atencio; Gaylarde,Christine C.; John,Vanderley M.

    2011-01-01

    The objective of this study is to investigate the efficiency of calcium carbonate bioprecipitation by Lysinibacillus sphaericus, Bacillus subtilis and Pseudomonas putida, obtained from the Coleção de Culturas do Instituto Nacional de Controle de Qualidade em Saúde (INCQS), as a first step in determining their potential to protect building materials against water uptake. Two culture media were studied: modified B4 containing calcium acetate and 295 with calcium chloride. Calcium consumption in...

  15. Cloning and expression in Escherichia coli of histidine utilization genes from Pseudomonas putida.

    OpenAIRE

    Consevage, M W; Porter, R D; Phillips, A. T.

    1985-01-01

    A library of the Pseudomonas putida chromosome, prepared through the use of the cosmid pJB8 ligated to a partial Sau3A digest of bacterial DNA, followed by in vitro packaging into bacteriophage lambda particles, was used to construct a strain of Escherichia coli which contained the genes for histidine utilization. This isolate produced a repressor product and all five enzymes required in Pseudomonas spp. for histidine dissimilation, whereas none of these could be detected in the nontransduced...

  16. Effect of gravity on Pseudomonas putida and kaolinite cotransport in water saturated porous media

    Science.gov (United States)

    Vasiliadou, Ioanna A.; Chrysikopoulos, Constantinos V.

    2013-04-01

    Bacterial transport in porous media can be affected by several factors, such as cell concentration, water velocity, and attachment onto the solid matrix or suspended in the aqueous phase soil particles (e.g. clays). Gravity, also may significantly influence bacterial transport behavior in the subsurface. The present study aims to determine the gravity effect on transport and cotransport of bacteria species Pseudomonas (P.) putida and kaolinite colloid particles in porous media. Transport experiments were conducted under horizontal-, up- and down-flow conditions in water saturated columns packed with glass beads. These different flow modes represent different gravity effects, namely: no-, negative- and positive-gravity effect. Initial experiments were performed with bacteria and kaolinite alone in order to evaluate the effect of gravity on their individual transport characteristics. No significant gravity effect was observed on the transport of individual bacterial cells. In contrary, each different flow mode was found to differently affect kaolinite transport. Compared to the horizontal-flow mode, the kaolinite mass recovery was decreased during the up-flow mode, and increased during the down-flow mode. Finally, P. putida and kaolinite particles were injected simultaneously into the packed column in order to investigate their cotransport behavior under different flow modes. The experimental data indicated that the kaolinite-P. putida cotransport behavior was similar to that observed for the transport of individual kaolinite particles. It was observed that the P. putida mass recovery decreased during down-flow conditions. This phenomenon may be caused by the attachment of bacteria onto kaolinite particles, which were adsorbed onto the solid matrix.

  17. Pseudomonas putida and Pseudomonas fluorescens Species Group Recovery from Human Homes Varies Seasonally and by Environment.

    Directory of Open Access Journals (Sweden)

    Susanna K Remold

    Full Text Available By shedding light on variation in time as well as in space, long-term biogeographic studies can help us define organisms' distribution patterns and understand their underlying drivers. Here we examine distributions of Pseudomonas in and around 15 human homes, focusing on the P. putida and P. fluorescens species groups. We describe recovery from 10,941 samples collected during up to 8 visits per home, occurring on average 2.6 times per year. We collected a mean of 141 samples per visit, from sites in most rooms of the house, from the surrounding yards, and from human and pet occupants. We recovered Pseudomonas in 9.7% of samples, with the majority of isolates being from the P. putida and P. fluorescens species groups (approximately 62% and 23% of Pseudomonas samples recovered respectively. Although representatives of both groups were recovered from every season, every house, and every type of environment sampled, recovery was highly variable across houses and samplings. Whereas recovery of P. putida group was higher in summer and fall than in winter and spring, P. fluorescens group isolates were most often recovered in spring. P. putida group recovery from soils was substantially higher than its recovery from all other environment types, while higher P. fluorescens group recovery from soils than from other sites was much less pronounced. Both species groups were recovered from skin and upper respiratory tract samples from healthy humans and pets, although this occurred infrequently. This study indicates that even species that are able to survive under a broad range of conditions can be rare and variable in their distributions in space and in time. For such groups, determining patterns and causes of stochastic and seasonal variability may be more important for understanding the processes driving their biogeography than the identity of the types of environments in which they can be found.

  18. Trichloroethylene degradation by Escherichia coli containing the cloned Pseudomonas putida F1 toluene dioxygenase genes.

    OpenAIRE

    Zylstra, G J; Wackett, L P; Gibson, D T

    1989-01-01

    Toluene dioxygenase from Pseudomonas putida F1 has been implicated as an enzyme capable of degrading trichloroethylene. This has now been confirmed with Escherichia coli JM109(pDTG601) that contains the structural genes (todC1C2BA) of toluene dioxygenase under the control of the tac promoter. The extent of trichloroethylene degradation by the recombinant organism depended on the cell concentration and the concentration of trichloroethylene. A linear rate of trichloroethylene degradation was o...

  19. The c-di-GMP phosphodiesterase BifA regulates biofilm development in Pseudomonas putida.

    Science.gov (United States)

    Jiménez-Fernández, Alicia; López-Sánchez, Aroa; Calero, Patricia; Govantes, Fernando

    2015-02-01

    We previously showed the isolation of biofilmpersistent Pseudomonas putida mutants that fail to undergo biofilm dispersal upon entry in stationary phase. Two such mutants were found to bear insertions in PP0914, encoding a GGDEF/EAL domain protein with high similarity to Pseudomon asaeruginosa BifA. Here we show the phenotypic characterization of a ΔbifA mutant in P. putida KT2442.This mutant displayed increased biofilm and pellicle formation, cell aggregation in liquid medium and decreased starvation-induced biofilm dispersal relative to the wild type. Unlike its P. aeruginosa counterpart, P. putida BifA did not affect swarming motility. The hyperadherent phenotype of the ΔbifA mutant correlates with a general increase in cyclic diguanylate (c-di-GMP) levels, Congo Red-binding exopolyaccharide production and transcription of the adhesin-encoding lapA gene. Integrity of the EAL motif and a modified GGDEF motif (altered to GGDQF)were crucial for BifA activity, and c-di-GMP depletion by overexpression of a heterologous c-di-GMP phosphodiesterase in the ΔbifA mutant restored wild-type biofilm dispersal and lapA expression.Our results indicate that BifA is a phosphodiesterase involved in the regulation of the c-di-GMP pool and required for the generation of the low c-di-GMP signal that triggers starvation-induced biofilm dispersal.

  20. Bioremediation Of Heavy Metals By Pseudomonas Putida Isolated From Groundwater In Egypt.

    Directory of Open Access Journals (Sweden)

    Fawazy

    2015-08-01

    Full Text Available In this present study total four bacterial isolates were obtained from 34 collected groundwater samples in 10th of Ramadan Sharkia governorate Egypt. These isolate were grown on nutrient agar supplemented with 1mgl of iron manganese and combination between them VV. Further testing of the bacterial isolates were grown on nutrient agar supplemented with different concentrations 2 4 5 6 7 8 and 9 mgl of iron and manganese. Out of four isolates one bacterial isolate no.83 has shown the resistance to heavy metals at maximum concentration of 8mgl. Selected isolate no.S83 was identified as Pseudomonas putida S83 according to Bergeys manual depending on morphological and biochemical characteristics. Transmission electron microscopy study of P. putida isolate no. S83 showed accumulation of heavy metal salts within and external to bacterial cells. P. putida S83 have higher removal efficiency of Fe2 94.5 and Mn2 94 at concentration 2 mgl and 96 hours.

  1. Antibiotic resistance determinants in a Pseudomonas putida strain isolated from a hospital.

    Directory of Open Access Journals (Sweden)

    Lázaro Molina

    Full Text Available Environmental microbes harbor an enormous pool of antibiotic and biocide resistance genes that can impact the resistance profiles of animal and human pathogens via horizontal gene transfer. Pseudomonas putida strains are ubiquitous in soil and water but have been seldom isolated from humans. We have established a collection of P. putida strains isolated from in-patients in different hospitals in France. One of the isolated strains (HB3267 kills insects and is resistant to the majority of the antibiotics used in laboratories and hospitals, including aminoglycosides, ß-lactams, cationic peptides, chromoprotein enediyne antibiotics, dihydrofolate reductase inhibitors, fluoroquinolones and quinolones, glycopeptide antibiotics, macrolides, polyketides and sulfonamides. Similar to other P. putida clinical isolates the strain was sensitive to amikacin. To shed light on the broad pattern of antibiotic resistance, which is rarely found in clinical isolates of this species, the genome of this strain was sequenced and analysed. The study revealed that the determinants of multiple resistance are both chromosomally-borne as well as located on the pPC9 plasmid. Further analysis indicated that pPC9 has recruited antibiotic and biocide resistance genes from environmental microorganisms as well as from opportunistic and true human pathogens. The pPC9 plasmid is not self-transmissible, but can be mobilized by other bacterial plasmids making it capable of spreading antibiotic resistant determinants to new hosts.

  2. In situ biosurfactant production and hydrocarbon removal by Pseudomonas putida CB-100 in bioaugmented and biostimulated oil-contaminated soil

    Directory of Open Access Journals (Sweden)

    Martínez-Toledo Ángeles

    2013-01-01

    Full Text Available In situ biosurfactant (rhamnolipid production by Pseudomonas putida CB-100 was achieved during a bioaugmented and biostimulated treatment to remove hydrocarbons from aged contaminated soil from oil well drilling operations. Rhamnolipid production and contaminant removal were determined for several treatments of irradiated and non-irradiated soils: nutrient addition (nitrogen and phosphorus, P. putida addition, and addition of both (P. putida and nutrients. The results were compared against a control treatment that consisted of adding only sterilized water to the soils. In treatment with native microorganisms (non-irradiated soils supplemented with P. putida, the removal of total petroleum hydrocarbons (TPH was 40.6%, the rhamnolipid production was 1.54 mg/kg, and a surface tension of 64 mN/m was observed as well as a negative correlation (R = -0.54; p < 0.019 between TPH concentration (mg/kg and surface tension (mN/m, When both bacteria and nutrients were involved, TPH levels were lowered to 33.7%, and biosurfactant production and surface tension were 2.03 mg/kg and 67.3 mN/m, respectively. In irradiated soil treated with P. putida, TPH removal was 24.5% with rhamnolipid generation of 1.79 mg/kg and 65.6 mN/m of surface tension, and a correlation between bacterial growth and biosurfactant production (R = -0.64; p < 0.009 was observed. When the nutrients and P. putida were added, TPH removal was 61.1%, 1.85 mg/kg of biosurfactants were produced, and the surface tension was 55.6 mN/m. In summary, in irradiated and non-irradiated soils, in situ rhamnolipid production by P. putida enhanced TPH decontamination of the soil.

  3. Identification of camphor oxidation and reduction products in Pseudomonas putida: new activity of the cytochrome P450cam system.

    Science.gov (United States)

    Prasad, Brinda; Rojubally, Adina; Plettner, Erika

    2011-06-01

    P450 enzymes are known for catalyzing hydroxylation reactions of non-activated C-H bonds. For example, P450(cam) from Pseudomonas putida oxidizes (1R)-(+)-camphor to 5-exo-hydroxy camphor and further to 5-ketocamphor. This hydroxylation reaction proceeds via a catalytic cycle in which the reduction of dioxygen (O(2)) is coupled to the oxidation of the substrate. We have observed that under conditions of low oxygen, P. putida and isolated P450(cam) reduce camphor to borneol. We characterized the formation of borneol under conditions of low oxygen or when the catalytic cycle is shunted by artificial oxidants like m-chloro perbenzoic acid, cumene hydroperoxide, etc. We also tested the toxicity of camphor and borneol with P. putida and Escherichia coli. We have found that in P. putida borneol is less toxic than camphor, whereas in E. coli borneol is more toxic than camphor. We discuss a potental ecological advantage of the camphor reduction reaction for P. putida.

  4. Draft Genome Sequence Analysis of a Pseudomonas putida W15Oct28 Strain with Antagonistic Activity to Gram-Positive and Pseudomonas sp. Pathogens

    NARCIS (Netherlands)

    Ye, L.; Hildebrandt, F.; Ballet, S.; Laus, M.S.; Berendsen, Roeland; Cornelis, P.

    2014-01-01

    Pseudomonas putida is a member of the fluorescent pseudomonads known to produce the yellow-green fluorescent pyoverdine siderophore. P. putida W15Oct28, isolated from a stream in Brussels, was found to produce compound(s) with antimicrobial activity against the opportunistic pathogens Staphylococcus

  5. Activity and three-dimensional distribution of toluene-degrading Pseudomonas putida in a multispecies biofilm assessed by quantitative in situ hybridization and scanning confocal laser microscopy

    DEFF Research Database (Denmark)

    Møller, Søren; Pedersen, Anne Rathmann; Poulsen, L.K.

    1996-01-01

    As a representative member of the toluene-degrading population in a biofilter for waste gas treatment, Pseudomonas putida was investigated with a 16S rRNA targeting probe, The three-dimensional distribution of P. putida was visualized in the biofilm matrix by scanning confocal laser microscopy...

  6. Isolation of NDM-1-producing multidrug-resistant Pseudomonas putida from a paediatric case of acute gastroenteritis, India

    Directory of Open Access Journals (Sweden)

    D. Bhattacharya

    2015-05-01

    Full Text Available Pseudomonas putida is an uncommon opportunistic pathogen, usually susceptible to antimicrobial agents. Data concerning resistance to antimicrobial agents in clinical P. putida isolates are limited. To the best of our knowledge we report for the first time the isolation of NDM-1-producing multidrug-resistant P. putida from a case of acute gastroenteritis. The isolate showed resistance to a wide range of antimicrobials, including fluoroquinolones, third-generation cephalosporins and carbapenems. The isolate also exhibited multiple mutations in the quinolone resistance determining region and showed the presence of qepA, blaTEM, blaOXA1 and blaOXA7 genes. The present study highlights the importance of looking for the relatively rare aetiological agents in clinical samples that do not yield common pathogens.

  7. Inhibition of Xanthomonas fragariae, Causative Agent of Angular Leaf Spot of Strawberry, through Iron Deprivation

    Directory of Open Access Journals (Sweden)

    Peter Henry

    2016-10-01

    Full Text Available In commercial production settings, few options exist to prevent or treat angular leaf spot of strawberry, a disease of economic importance and caused by the bacterial pathogen Xanthomonas fragariae (Xfr. In the process of isolating and identifying Xfr bacteria from symptomatic plants, we observed growth inhibition of Xfr by bacterial isolates from the same leaf macerates. Identified as species of Pseudomonas and Rhizobium, these isolates were confirmed to suppress growth of Xfr in agar overlay plates and in microtiter plate cultures, as did our reference strain Pseudomonas putida KT2440. Screening of a transposon mutant library of KT2440 revealed that disruption of the biosynthetic pathway for the siderophore pyoverdine resulted in complete loss of Xfr antagonism, suggesting iron competition as a mode of action. Antagonism could be replicated on plate and in culture by addition of purified pyoverdine or by addition of the chelating agents tannic acid and dipyridyl, while supplementing the medium with iron negated the inhibitory effects of pyoverdine, tannic acid and dipyridyl. When co-inoculated with tannic acid onto strawberry plants, Xfr’s ability to cause foliar symptoms was greatly reduced, suggesting a possible opportunity for iron-based management of angular leaf spot. We discuss our findings in the context of ‘nutritional immunity’, the idea that plant hosts restrict pathogens access to iron, either directly, or indirectly through their associated microbiota.

  8. Pseudomonas putida response in membrane bioreactors under salicylic acid-induced stress conditions

    Energy Technology Data Exchange (ETDEWEB)

    Collado, Sergio; Rosas, Irene; González, Elena; Gutierrez-Lavin, Antonio; Diaz, Mario, E-mail: mariodiaz@uniovi.es

    2014-02-01

    Highlights: • MBR under feed-induced stress conditions: starvation and changing feeding conditions. • High capacity of MBR to withstand high variations in feed loads. • Slow biofilm formation under starvation conditions during the first days. • Observed growth of P. putida for substrate to microorganism ratio higher than 0.6 g/g. • Maximum specific growth rate and growth yield values of around 37.5 h{sup −1} and 0.5 g/g. - Abstract: Starvation and changing feeding conditions are frequently characteristics of wastewater treatment plants. They are typical causes of unsteady-state operation of biological systems and provoke cellular stress. The response of a membrane bioreactor functioning under feed-induced stress conditions is studied here. In order to simplify and considerably amplify the response to stress and to obtain a reference model, a pure culture of Pseudomonas putida was selected instead of an activated sludge and a sole substrate (salicylic acid) was employed. The system degraded salicylic acid at 100–1100 mg/L with a high level of efficiency, showed rapid acclimation without substrate or product inhibition phenomena and good stability in response to unsteady states caused by feed variations. Under starvation conditions, specific degradation rates of around 15 mg/g h were achieved during the adaptation of the biomass to the new conditions and no biofilm formation was observed during the first days of experimentation using an initial substrate to microorganisms ratio lower than 0.1. When substrate was added to the reactor as pulses resulting in rapidly changing concentrations, P. putida growth was observed only for substrate to microorganism ratios higher than 0.6, with a maximum Y{sub X/S} of 0.5 g/g. Biofilm development under changing feeding conditions was fast, biomass detachment only being significant for biomass concentrations on the membrane surface that were higher than 16 g/m{sup 2}.

  9. Screening and optimization of low-cost medium for Pseudomonas putida Rs-198 culture using RSM.

    Science.gov (United States)

    Peng, Yanjie; He, Yanhui; Wu, Zhansheng; Lu, Jianjiang; Li, Chun

    2014-01-01

    The plant growth-promoting rhizobacterial strain Pseudomonas putida Rs-198 was isolated from salinized soils from Xinjiang Province. We optimized the composition of the low-cost medium of P. putida Rs-198 based on its bacterial concentration, as well as its phosphate-dissolving and indole acetic acid (IAA)-producing capabilities using the response surface methodology (RSM), and a mathematical model was developed to show the effect of each medium component and its interactions on phosphate dissolution and IAA production. The model predicted a maximum phosphate concentration in medium containing 63.23 mg/L inorganic phosphate with 49.22 g/L corn flour, 14.63 g/L soybean meal, 2.03 g/L K₂HPO₄, 0.19 g/L MnSO₄ and 5.00 g/L NaCl. The maximum IAA concentration (18.73 mg/L) was predicted in medium containing 52.41 g/L corn flour, 15.82 g/L soybean meal, 2.40 g/L K₂HPO₄, 0.17 g/L MnSO₄ and 5.00 g/L NaCl. These predicted values were also verified through experiments, with a cell density of 10(13) cfu/mL, phosphate dissolution of 64.33 mg/L, and IAA concentration of 18.08 mg/L. The excellent correlation between predicted and measured values of each model justifies the validity of both the response models. The study aims to provide a basis for industrialized fermentation using P. putida Rs-198.

  10. Screening and optimization of low-cost medium for Pseudomonas putida Rs-198 culture using RSM

    Directory of Open Access Journals (Sweden)

    Yanjie Peng

    2014-12-01

    Full Text Available The plant growth-promoting rhizobacterial strain Pseudomonas putida Rs-198 was isolated from salinized soils from Xinjiang Province. We optimized the composition of the low-cost medium of P. putida Rs-198 based on its bacterial concentration, as well as its phosphate-dissolving and indole acetic acid (IAA-producing capabilities using the response surface methodology (RSM, and a mathematical model was developed to show the effect of each medium component and its interactions on phosphate dissolution and IAA production. The model predicted a maximum phosphate concentration in medium containing 63.23 mg/L inorganic phosphate with 49.22 g/L corn flour, 14.63 g/L soybean meal, 2.03 g/L K2HPO4, 0.19 g/L MnSO4 and 5.00 g/L NaCl. The maximum IAA concentration (18.73 mg/L was predicted in medium containing 52.41 g/L corn flour, 15.82 g/L soybean meal, 2.40 g/L K2HPO4, 0.17 g/L MnSO4 and 5.00 g/L NaCl. These predicted values were also verified through experiments, with a cell density of 10(13 cfu/mL, phosphate dissolution of 64.33 mg/L, and IAA concentration of 18.08 mg/L. The excellent correlation between predicted and measured values of each model justifies the validity of both the response models. The study aims to provide a basis for industrialized fermentation using P. putida Rs-198.

  11. Variability in subpopulation formation propagates into biocatalytic variability of engineered Pseudomonas putida strains

    Directory of Open Access Journals (Sweden)

    Martin eLindmeyer

    2015-10-01

    Full Text Available Pivotal challenges in industrial biotechnology are the identification and overcoming of cell-to-cell heterogeneity in microbial processes. While the development of subpopulations of isogenic cells in bioprocesses is well described (intra-population variability, a possible variability between genetically identical cultures growing under macroscopically identical conditions (clonal variability is not. A high such clonal variability has been found for the recombinant expression of the styrene monooxygenase genes styAB from Pseudomonas taiwanensis VLB120 in solvent-tolerant Pseudomonas putida DOT-T1E using the alk-regulatory system from P. putida GPo1. In this study, the oxygenase subunit StyA fused to eGFP was used as readout tool to characterize the population structure in P. putida DOT-T1E regarding recombinant protein content. Flow cytometric analyses revealed that in individual cultures, at least two subpopulations with highly differing recombinant StyA-eGFP protein contents appeared (intra-population variability. Interestingly, subpopulation sizes varied from culture-to-culture correlating with the specific styrene epoxidation activity of cells derived from respective cultures (clonal variability. In addition, flow cytometric cell sorting coupled to plasmid copy number (PCN determination revealed that detected clonal variations cannot be correlated to the PCN, but depend on the combination of the regulatory system and the host strain employed. This is, to the best of our knowledge, the first work reporting that intra-population variability (with differing protein contents in the presented case study causes clonal variability of genetically identical cultures. Respective impacts on bioprocess reliability and performance and strategies to overcome respective reliability issues are discussed.

  12. Kinetics of mercury reduction by Serratia marcescens mercuric reductase expressed by pseudomonas putida strains

    Energy Technology Data Exchange (ETDEWEB)

    Schneider, M.; Deckwer, W.D. [GBF-Gesellschaft fuer Biotechnologische Forschung mbH, Abteilung TU-BCE, Mascheroder Weg 1, D-38124 Braunschweig (Germany)

    2005-10-01

    Mercury (Hg) resistance is widespread among microorganisms and is based on the intracellular transformation of Hg(II) to less toxic elemental Hg(0). The use of microbial consortia to demercurize polluted wastewater streams and environments has been demonstrated. To develop efficient and versatile microbial cleanup strategies requires detailed knowledge of transport and reaction rates. This study focuses on the kinetics of the key enzyme of the microbial transformation, e.g., the mercuric reductase (MerA) under conditions closely resembling the cell interior. To this end, previously constructed and characterized Pseudomonas putida strains expressing MerA from Serratia marcescens were applied. Of the P. putida strains considered in this study P. putida KT2442::mer73 constitutively expressing broad spectrum mercury resistance (merTPAB) yielded the highest mercuric reductase (MerA) activity directly after cell disruption. MerA in the raw extract was further purified (about 100 fold). Reduction rates were measured for various substrates (HgCl{sub 2}, Hg{sub 2}SO{sub 4}, Hg(NO{sub 3}){sub 2} and phenyl mercury acetate) up to high concentrations dependent on the purification grade. In all cases, a pronounced substrate inhibition was found. The kinetic constants determined for the cell raw extract are in agreement with those measured for intact cells. However, the rate data exhibit reduced affinity and inhibition with rising purification grade (specific activity). Therefore, the findings seemingly point to reactions preceding the catalytic reduction. Based on simplified assumptions, a kinetic model is suggested which reasonably describes the experimental findings and can advantageously be applied to the bioreactor design. (Abstract Copyright [2005], Wiley Periodicals, Inc.)

  13. Enhanced Biological Phosphorus Removal with Pseudomonas putida GM6 from Activated Sludge

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    The enhanced biological phosphorus removal (EBPR) method is widely adopted for phosphorus removal from wastewater, yet little is known about its microbiological and molecular mechanisms. Therefore, it is difficult to predict and control the deterioration of the EBPR process in a large-scale municipal sewage treatment plant. This study used a novel strain isolated in the laboratory, Pseudomonas putida GM6, which had a high phosphate accumulating ability and could recover rapidly from the deteriorated system and enhance the capability of phosphorus removal in activated sludge. Strain GM6 marked with gfp gene, which was called GMTR, was delivered into a bench-scale sequencing batch reactor (SBR)of low efficiency, to investigate the colonization of GMTR and removal of phosphorus. After 21 days, the proportion of GMTR in the total bacteria of the sludge reached 9.2%, whereas the phosphorus removal rate was 96%, with an effluent concentration of about 0.2 mg L-1. In the reactor with the addition of GMTR, phosphorus was removed quickly, in 1 h under anaerobic conditions, and in 2 h under aerobic conditions. These evidences were characteristic of EBPR processes.Field testing was conducted at a hospital sewage treatment facility with low phosphorus removal capability. Twentyone days after Pseudononas putida GM6 was added, effluent phosphorus concentration remained around 0.3 mg L-1,corresponding to a removal rate of 96.8%. It was therefore demonstrated that Pseudomonas putida GM6 could be used for a quick startup and enhancement of wastewater biological phosphorus removal, which provided a scientific basis for potential large-scale engineering application.

  14. Pseudomonas putida KT2442 as a platform for the biosynthesis of polyhydroxyalkanoates with adjustable monomer contents and compositions

    DEFF Research Database (Denmark)

    Tripathi, Lakshmi; Wu, Lin-Ping; Dechuan, Meng

    2013-01-01

    The β-oxidation weakened Pseudomonas putida were established as a platform for the production of polyhydroxyalkanoates (PHA) with adjustable monomer compositions and micro-structures. When mutant P. putida KTOYO6ΔC (phaPCJA.c) was cultivated on mixtures of sodium butyrate and sodium hexanoate (C4:C......HD-co-3HDD) consisting of 49 mol% P3HHx and 51 mol% P(3HD-co-3HDD) [35.25 mol% 3HDD (3-hydroxydodecanoate)], which was characterized by (13)C NMR, HMBC NMR, DSC, GPC and universal testing machine....

  15. NahY, a Catabolic Plasmid-Encoded Receptor Required for Chemotaxis of Pseudomonas putida to the Aromatic Hydrocarbon Naphthalene

    OpenAIRE

    1999-01-01

    Pseudomonas putida G7 exhibits chemotaxis to naphthalene, but the molecular basis for this was not known. A new gene, nahY, was found to be cotranscribed with meta cleavage pathway genes on the NAH7 catabolic plasmid for naphthalene degradation. The nahY gene encodes a 538-amino-acid protein with a membrane topology and a C-terminal region that resemble those of chemotaxis transducer proteins. A P. putida G7 nahY mutant grew on naphthalene but was not chemotactic to this aromatic hydrocarbon....

  16. Identification and validation of novel small proteins in Pseudomonas putida

    DEFF Research Database (Denmark)

    Yang, Xiaochen; Ingemann Jensen, Sheila; Wulff, Tune;

    2016-01-01

    Small proteins of fifty amino acids or less have been understudied due to difficulties that impede their annotation and detection. In order to obtain information on small open reading frames (sORFs) in P. putida, bioinformatic and proteomic approaches were used to identify putative small open...... of fourteen sORFs was confirmed, where all except one are novel. All of the validated sORFs except one are located adjacent to annotated genes on the same strand and three are in close proximity to genes with known functions. These include an ABC transporter operon and the two transcriptional regulators Fis...

  17. Regulation of phenylacetic acid uptake is sigma54 dependent in Pseudomonas putida CA-3.

    LENUS (Irish Health Repository)

    O' Leary, Niall D

    2011-10-13

    Abstract Background Styrene is a toxic and potentially carcinogenic alkenylbenzene used extensively in the polymer processing industry. Significant quantities of contaminated liquid waste are generated annually as a consequence. However, styrene is not a true xenobiotic and microbial pathways for its aerobic assimilation, via an intermediate, phenylacetic acid, have been identified in a diverse range of environmental isolates. The potential for microbial bioremediation of styrene waste has received considerable research attention over the last number of years. As a result the structure, organisation and encoded function of the genes responsible for styrene and phenylacetic acid sensing, uptake and catabolism have been elucidated. However, a limited understanding persists in relation to host specific regulatory molecules which may impart additional control over these pathways. In this study the styrene degrader Pseudomonas putida CA-3 was subjected to random mini-Tn5 mutagenesis and mutants screened for altered styrene\\/phenylacetic acid utilisation profiles potentially linked to non-catabolon encoded regulatory influences. Results One mutant, D7, capable of growth on styrene, but not on phenylacetic acid, harboured a Tn5 insertion in the rpoN gene encoding σ54. Complementation of the D7 mutant with the wild type rpoN gene restored the ability of this strain to utilise phenylacetic acid as a sole carbon source. Subsequent RT-PCR analyses revealed that a phenylacetate permease, PaaL, was expressed in wild type P. putida CA-3 cells utilising styrene or phenylacetic acid, but could not be detected in the disrupted D7 mutant. Expression of plasmid borne paaL in mutant D7 was found to fully restore the phenylacetic acid utilisation capacity of the strain to wild type levels. Bioinformatic analysis of the paaL promoter from P. putida CA-3 revealed two σ54 consensus binding sites in a non-archetypal configuration, with the transcriptional start site being resolved by

  18. Investigation Of The Primary Transcriptome Of The Production Organism Pseudomonas Putida

    DEFF Research Database (Denmark)

    D'Arrigo, Isotta; Bojanovic, Klara; Long, Katherine

    2015-01-01

    compounds, a capability that renders the bacterium useful in bioremediation. Finally, P. putida shows a high potential as a cell factory for the production of several compounds. Methods: Here, a differential RNA-sequencing approach (dRNA-seq) is used to gain new insights into the organization of the P...... motifs in promoter sequence. Finally, RNA regulatory elements have been identified including putative sRNAs and riboswitches. Selected candidate riboswitch sequences have been tested for the study of the ligand-dependent regulatory mechanisms. Conclusions: By determination of the 5’-ends of transcripts...

  19. Biosorption of heavy metals and uranium by starfish and Pseudomonas putida

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Jaeyoung [Korea Institute of Science and Technology (KIST), Gangneung Institute, Gangneung 210-340 (Korea, Republic of)], E-mail: jchoi@kist.re.kr; Lee, Ju Young; Yang, Jung-Seok [Korea Institute of Science and Technology (KIST), Gangneung Institute, Gangneung 210-340 (Korea, Republic of)

    2009-01-15

    Biosorption of heavy metals and uranium from contaminated wastewaters may represent an innovative purification process. This study investigates the removal ability of unit mass of Pseudomonas putida and starfish for lead, cadmium, and uranium by quantifying the adsorption capacity. The adsorption of heavy metals and uranium by the samples was influenced by pH, and increased with increasing Pb, Cd, and U concentrations. Dead cells adsorbed the largest quantity of all heavy metals than live cells and starfish. The adsorption capacity followed the order: U(VI) > Pb > Cd. The results also suggest that bacterial membrane cells can be used successfully in the treatment of high strength metal-contaminated wastewaters.

  20. A case report of acute pediatric bacterial meningitis due to the rare isolate, Pseudomonas putida

    Institute of Scientific and Technical Information of China (English)

    Grishma V. Kulkarni

    2016-01-01

    Acute bacterial meningitis (ABM) is the medical emergency which warrants an early diagnosis and an aggressive therapy. Despite the availability of the potent newer antibiotics, the mortality caused by ABM and its complications remain high in India, ranging from 16% to 32%. The aim of this case report is to present the rare isolation ofPseudomonas putida from cerebrospinal lfuid sample. Besides this, the author also emphasizes the importance of correctly identifying the organism and thus the selection of the most accurate antibiotic from the susceptibility proifle to allow for early recovery and to improve the patient outcome and survival.

  1. Modeling of TCE and Toluene Toxicity to Pseudomonas putida F1

    Science.gov (United States)

    Singh, R.; Olson, M. S.

    2009-12-01

    Prediction of viable bacterial distribution with respect to contaminants is important for efficient bioremediation of contaminated ground-water aquifers, particularly those contaminated with residual NAPLs. While bacterial motility and chemotaxis may help situate bacteria close to high concentrations of contaminant thereby enhancing bioremediation, prolonged exposure to high concentrations of contaminates is toxic to contaminant-degrading bacteria. The purpose of this work is to model the toxicity of trichloroethylene and toluene to Pseudomonas putida F1. The Live/Dead® bacterial viability assay was used to determine the toxic effect of chemical contaminants on the viability of P. putida F1 in a sealed zero head-space experimental environment. Samples of bacterial suspensions were exposed to common ground-water pollutants, TCE and toluene, for different durations. Changes in live and dead cell populations were monitored over the course of experiments using fluorescence microscopy. Data obtained from these toxicity experiments were fit to simple linear and exponential bacterial decay models using non-linear regression to describe loss of bacterial viability. TCE toxicity to P. putida F1 was best described with an exponential decay model (Figure 1a), with a decay constant kTCE = 0.025 h-4.95 (r2 = 0.956). Toluene toxicity showed a marginally better fit to the linear decay model (Figure 1b) (r2 = 0.971), with a decay constant ktoluene = 0.204 h-1. Best-fit model parameters obtained for both TCE and toluene were used to predict bacterial viability in toxicity experiments with higher contaminant concentrations and matched well with experimental data. Results from this study can be used to predict bacterial accumulation and viability near NAPL sources, and thus may be helpful in improving bioremediation performance assessment of contaminated sites. Figure 1: Survival ratios (S = N/No) of P. putida F1 in TCE- (a) and toluene- (b) stressed samples (observed (

  2. The Interface Reaction between Pseudomonas putida and Soil Clay Minerals and Its Effect on the Biodegradation of Methyl Parathion%恶臭假单胞菌与土壤矿物的界面反应及其对甲基对硫磷降解影响

    Institute of Scientific and Technical Information of China (English)

    冯伟亮; 贺小敏; 沈敏; 张晓斌; 陈浩; 黄巧云

    2009-01-01

    Adsorption characteristics and metabolism curves of Pseudomonas putida on the surfaces of montmorillonite, kaolinite and goethite were investigated. Biodegradation kinetics of methyl parathion by Pseudomonas putida was also measured in the presence of various clay minerals. The binding strength of Pseudomonas putida on minerals followed the sequence of goethite > kaolinite > montmorillonite. The metabolism abilities of free cells were stronger than that of immobilized ones at a methyl parathion of 10 mg/L over the whole incubation period. However, the percent degradation of methyl parathion by immobilized cells was higher (within 9 h) and then gradually lower than by free ones at high pesticide concentrations (20~40 mg/L). Degradation capabilities of Pseudomonas putida sorbed on various minerals followed the sequence of montmorillonite > kaolinite > goethite. Montmorillonite presented a lower affinity for Pseudomonas putida, and it could enhance the bacterial activity and further stimulate the bioavailability of pesticide. However, goethite exhibited higher sorption capability for the bacterium, and it could display an inhibitory effect on the bacterial activitiy and hinder the substrate biodegradation.%研究了恶臭假单胞菌在蒙脱石、高岭石和针铁矿表面的吸附特征,探讨了细菌在不同粘粒矿物存在下的生长代谢活性,及对甲基对硫磷的降解动力学.结果表明, 三种矿物对细菌的吸附强度为针铁矿>高岭石>蒙脱石.当甲基对硫磷浓度较低时(10 mg/L), 游离菌的降解能力始终比固定菌强;在高浓度(20~40 mg/L)下, 固定菌对农药的降解能力起初(前9 h)高于游离菌, 随后渐渐低于游离菌.不同矿物固定的细菌, 其降解能力为蒙脱石>高岭石>针铁矿.蒙脱石对细菌的亲和力最弱, 但它对细菌的代谢活性有促进作用, 有利于农药的生物降解; 而针铁矿与细菌的结合强度最大, 细菌活性受到抑制, 不利于农药的降解.

  3. Genetic Analysis of the Nature of Genes Coding Early Enzymes in the Metabolism of Cresol in JPT3-4, a Derivative Strain of Pseudomonas Aeroginosis J1.

    Science.gov (United States)

    1982-04-29

    sections, The cresols, phenol, and catechols were purchased from Aldrich. Crystalline catechols were vacuum- sublimated and stored in brown phials under...transmissible plasmid controlling camphor oxidation in Pseudomonas putida. Proc, Nat. Acad. Sci. U.S.A. 70:855-889. Ribbons, D, W. 1966, Metabolism of o

  4. Novel AroA from Pseudomonas putida Confers Tobacco Plant with High Tolerance to Glyphosate

    Science.gov (United States)

    Yan, Hai-Qin; Chang, Su-Hua; Tian, Zhe-Xian; Zhang, Le; Sun, Yi-Cheng; Li, Yan; Wang, Jing; Wang, Yi-Ping

    2011-01-01

    Glyphosate is a non-selective broad-spectrum herbicide that inhibits 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS, also designated as AroA), a key enzyme in the aromatic amino acid biosynthesis pathway in microorganisms and plants. Previously, we reported that a novel AroA (PpAroA1) from Pseudomonas putida had high tolerance to glyphosate, with little homology to class I or class II glyphosate-tolerant AroA. In this study, the coding sequence of PpAroA1 was optimized for tobacco. For maturation of the enzyme in chloroplast, a chloroplast transit peptide coding sequence was fused in frame with the optimized aroA gene (PparoA1optimized) at the 5′ end. The PparoA1optimized gene was introduced into the tobacco (Nicotiana tabacum L. cv. W38) genome via Agrobacterium-mediated transformation. The transformed explants were first screened in shoot induction medium containing kanamycin. Then glyphosate tolerance was assayed in putative transgenic plants and its T1 progeny. Our results show that the PpAroA1 from Pseudomonas putida can efficiently confer tobacco plants with high glyphosate tolerance. Transgenic tobacco overexpressing the PparoA1optimized gene exhibit high tolerance to glyphosate, which suggest that the novel PpAroA1 is a new and good candidate applied in transgenic crops with glyphosate tolerance in future. PMID:21611121

  5. Phenotypic variation of Pseudomonas putida and P. tolaasii affects attachment to Agaricus bisporus mycelium.

    Science.gov (United States)

    Rainey, P B

    1991-12-01

    The effect of phenotypic variation on attachment of Pseudomonas tolaasii and P. putida to Agaricus bisporus mycelium was investigated. Quantitative studies demonstrated the ability of each isolate to attach rapidly and firmly to A. bisporus mycelium and significant differences in attachment of wild-type and phenotypic variant strains were observed. This was most pronounced in P. tolaasii, where the percentage attachment of the wild-type form was always greater than that of the phenotypic variant. The medium upon which the bacteria were cultured, prior to conducting an attachment assay, had a significant effect on their ability to attach. Attachment of the wild-type form of P. putida was enhanced when the assay was performed in the presence of CaCl2, suggesting the involvement of electrostatic forces. No correlation was observed between bacterial hydrophobicity and ability to attach to A. bisporus mycelium. Scanning electron microscopy confirmed the results obtained from the quantitative studies and provided further evidence for marked differences in the ability of the pseudomonads to attach to mycelium. Fibrillar structures and amorphous material were frequently associated with attached cells and appeared to anchor bacteria to each other and to the hyphal surface. A time-course study of attachment using transmission electron microscopy revealed the presence of uneven fibrillar material on the surface of cells. This material stained positive for polysaccharide and may be involved in ensuring rapid, firm attachment of the cells.

  6. Pseudomonas putida response in membrane bioreactors under salicylic acid-induced stress conditions.

    Science.gov (United States)

    Collado, Sergio; Rosas, Irene; González, Elena; Gutierrez-Lavin, Antonio; Diaz, Mario

    2014-02-28

    Starvation and changing feeding conditions are frequently characteristics of wastewater treatment plants. They are typical causes of unsteady-state operation of biological systems and provoke cellular stress. The response of a membrane bioreactor functioning under feed-induced stress conditions is studied here. In order to simplify and considerably amplify the response to stress and to obtain a reference model, a pure culture of Pseudomonas putida was selected instead of an activated sludge and a sole substrate (salicylic acid) was employed. The system degraded salicylic acid at 100-1100mg/L with a high level of efficiency, showed rapid acclimation without substrate or product inhibition phenomena and good stability in response to unsteady states caused by feed variations. Under starvation conditions, specific degradation rates of around 15mg/gh were achieved during the adaptation of the biomass to the new conditions and no biofilm formation was observed during the first days of experimentation using an initial substrate to microorganisms ratio lower than 0.1. When substrate was added to the reactor as pulses resulting in rapidly changing concentrations, P. putida growth was observed only for substrate to microorganism ratios higher than 0.6, with a maximum YX/S of 0.5g/g. Biofilm development under changing feeding conditions was fast, biomass detachment only being significant for biomass concentrations on the membrane surface that were higher than 16g/m(2).

  7. Tracing explosives in soil with transcriptional regulators of Pseudomonas putida evolved for responding to nitrotoluenes

    Science.gov (United States)

    Garmendia, Junkal; De Las Heras, Aitor; Galvão, Teca Calcagno; De Lorenzo, Víctor

    2008-01-01

    Summary Although different biological approaches for detection of anti‐personnel mines and other unexploded ordnance (UXO) have been entertained, none of them has been rigorously documented thus far in the scientific literature. The industrial 2,4,6 trinitrotoluene (TNT) habitually employed in the manufacturing of mines is at all times tainted with a small but significant proportion of the more volatile 2,4 dinitrotoluene (2,4 DNT) and other nitroaromatic compounds. By using mutation‐prone PCR and DNA sequence shuffling we have evolved in vitro and selected in vivo variants of the effector recognition domain of the toluene‐responsive XylR regulator of the soil bacterium Pseudomonas putida that responds to mono‐, bi‐ and trinitro substituted toluenes. Re‐introduction of such variants in P. putida settled the transcriptional activity of the cognate promoters (Po and Pu) as a function of the presence of nitrotoluenes in the medium. When strains bearing transcriptional fusions to reporters with an optical output (luxAB, GFP) were spread on soil spotted with nitrotoluenes, the signal triggered by promoter activation allowed localization of the target compounds on the soil surface. Our data provide a proof of concept that non‐natural transcription factors evolved to respond to nitroaromatics can be engineered in soil bacteria and inoculated on a target site to pinpoint the presence of explosives. This approach thus opens new ways to tackle this gigantic humanitarian problem. PMID:21261843

  8. Global transcriptional response of solvent-sensitive and solvent-tolerant Pseudomonas putida strains exposed to toluene

    DEFF Research Database (Denmark)

    Molina-Santiago, Carlos; Udaondo, Zulema; Gómez Lozano, María

    2017-01-01

    Pseudomonas putida strains are generally recognized as solvent tolerant, exhibiting varied sensitivity to organic solvents. Pan-genome analysis has revealed that 30% of genes belong to the core-genome of Pseudomonas. Accessory and unique genes confer high degree of adaptability and capabilities...

  9. In situ phenol removal from fed-batch fermentations of solvent tolerant Pseudomonas putida S12 by pertraction

    NARCIS (Netherlands)

    Heerema, L.; Wierckx, N.; Roelands, C.P.M.; Hanemaaijer, J.H.; Goetheer, E.L.V.; Verdoes, D.; Keurentjes, J.

    2011-01-01

    In situ phenol pertraction with 1-octanol has been experimentally studied to improve the production of the model component phenol by a recombinant strain of Pseudomonas putida S12. When the phenol concentration in the reactor reaches 2mM, the cells in fermentations without phenol removal are inhibit

  10. TrgI, toluene repressed gene I, a novel gene involved in toluene-tolerance in Pseudomonas putida S12

    NARCIS (Netherlands)

    Volkers, R.J.M.; Ballerstedt, H.; Ruijssenaars, H.; Bont, J.A.M. de; Winde, J.H. de; Wery, J.

    2009-01-01

    Pseudomonas putida S12 is well known for its remarkable solvent tolerance. Transcriptomics analysis of this bacterium grown in toluene-containing chemostats revealed the differential expression of 253 genes. As expected, the genes encoding one of the major solvent tolerance mechanisms, the solvent e

  11. Novel Dehalogenase Mechanism for 2,3-Dichloro-1-Propanol Utilization in Pseudomonas putida Strain MC4

    NARCIS (Netherlands)

    Arif, Muhammad Ilan; Samin, Ghufrana; van Leeuwen, Jan G. E.; Oppentocht, Jantien; Janssen, Dick B.

    2012-01-01

    A Pseudomonas putida strain (MC4) that can utilize 2,3-dichloro-1-propanol (DCP) and several aliphatic haloacids and haloalcohols as sole carbon and energy source for growth was isolated from contaminated soil. Degradation of DCP was found to start with oxidation and concomitant dehalogenation catal

  12. TrgI, toluene repressed gene I, a novel gene involved in toluene-tolerance in Pseudomonas putida S12

    NARCIS (Netherlands)

    Volkers, R.J.M.; Ballerstedt, H.; Ruijssenaars, H.; Bont, J.A.M. de; Winde, J.H. de; Wery, J.

    2009-01-01

    Pseudomonas putida S12 is well known for its remarkable solvent tolerance. Transcriptomics analysis of this bacterium grown in toluene-containing chemostats revealed the differential expression of 253 genes. As expected, the genes encoding one of the major solvent tolerance mechanisms, the solvent e

  13. TrgI, toluene repressed gene I, a novel gene involved in toluene-tolerance in Pseudomonas putida S12

    NARCIS (Netherlands)

    Volkers, R.J.M.; Ballerstedt, H.; Ruijssenaars, H.; De Bont, J.A.M.; De Winde, J.H.; Wery, J.

    2008-01-01

    Pseudomonas putida S12 is well known for its remarkable solvent tolerance. Transcriptomics analysis of this bacterium grown in toluene-containing chemostats revealed the differential expression of 253 genes. As expected, the genes encoding one of the major solvent tolerance mechanisms, the solvent e

  14. Crystal Structure of the Leucine Aminopeptidase from Pseudomonas putida Reveals the Molecular Basis for its Enantioselectivity and Broad Substrate Specificity

    NARCIS (Netherlands)

    Kale, Avinash; Pijning, Tjaard; Sonke, Theo; Dijkstra, Bauke W.; Thunnissen, Andy-Mark W. H.; Guss, M.

    2010-01-01

    The zinc-dependent leucine aminopeptidase from Pseudomonas putida (ppLAP) is an important enzyme for the industrial production of enantiomerically pure amino acids. To provide a better understanding of its structure function relationships, the enzyme was studied by X-ray crystallography. Crystal str

  15. Development of a growth-dependent selection system for identification of L-threonine aldolases.

    Science.gov (United States)

    Bulut, Dalia; Gröger, Harald; Hummel, Werner

    2015-07-01

    Threonine aldolases (TAs) are useful enzymes for the synthesis of β-hydroxy-α-amino acids due to their capability to catalyze asymmetric aldol reactions. Starting from two prochiral compounds, an aldehyde and glycine, two chiral stereocenters were formed in a single step via C-C bond formation. Owing to poor diastereoselectivity and low activity, the enzymatic synthesis of β-hydroxy-α-amino acids by TAs is still a challenge. For identification of new TAs, a growth-dependent selection system in Pseudomonas putida KT2440 has been developed. This bacterium is able to use aromatic compounds such as benzaldehyde, which is the cleavage product of the TA-mediated retro-aldol reaction of phenylserine, as sole carbon source via the β-ketoadipate pathway. With DL-threo-β-phenylserine as sole carbon source, this strain showed only slight growth in minimal medium. This growth deficiency can be restored by introducing and expressing genes encoding TAs. In order to develop a highly efficient selection system, the gene taPp of P. putida KT2440 encoding a TA was successfully deleted by replacement with an antibiotic resistance cassette. Different growth studies were carried out to prove the operability of the selection system. Genes encoding for L- and D-specific TAs (L-TA genes of Escherichia coli (ltaE) and Saccharomyces cerevisiae (gly1) and D-TA gene of Achromobacter xylosoxidans (dtaAX)) were introduced into the selection strain P. putida KT2440ΔtaPp, followed by cultivation on minimal medium supplemented with DL-threo-β-phenylserine. The results demonstrate that only the selection strains with plasmid-encoded L-TAs were able to grow on this racemic amino acid, whereas the corresponding strain harboring the gene coding for a D-specific TA showed no growth. In summary, it can be stated that a powerful screening tool was developed to identify easily by growth new L-specific threonine aldolases or other enzymes from genomic or metagenomic libraries liberating

  16. Activity and three-dimensional distribution of toluene-degrading Pseudomonas putida in a multispecies biofilm assessed by quantitative in situ hybridization and scanning confocal laser microscopy

    DEFF Research Database (Denmark)

    Møller, Søren; Pedersen, Anne Rathmann; Poulsen, L.K.

    1996-01-01

    As a representative member of the toluene-degrading population in a biofilter for waste gas treatment, Pseudomonas putida was investigated with a 16S rRNA targeting probe, The three-dimensional distribution of P. putida was visualized in the biofilm matrix by scanning confocal laser microscopy....... demonstrating that P. putida was present throughout the biofilm. Acridine orange staining revealed a very heterogeneous structure of the fully hydrated biofilm, with cell-free channels extending ft om the surface Into the biofilm. This indicated that toluene may penetrate to deeper layers of the biofilm...

  17. Decolorization of industrial synthetic dyes using engineered Pseudomonas putida cells with surface-immobilized bacterial laccase

    Directory of Open Access Journals (Sweden)

    Wang Wei

    2012-06-01

    Full Text Available Abstract Background Microbial laccases are highly useful in textile effluent dye biodegradation. However, the bioavailability of cellularly expressed or purified laccases in continuous operations is usually limited by mass transfer impediment or enzyme regeneration difficulty. Therefore, this study develops a regenerable bacterial surface-displaying system for industrial synthetic dye decolorization, and evaluates its effects on independent and continuous operations. Results A bacterial laccase (WlacD was engineered onto the cell surface of the solvent-tolerant bacterium Pseudomonas putida to construct a whole-cell biocatalyst. Ice nucleation protein (InaQ anchor was employed, and the ability of 1 to 3 tandemly aligned N-terminal repeats to direct WlacD display were compared. Immobilized WlacD was determined to be surface-displayed in functional form using Western blot analysis, immunofluorescence microscopy, flow cytometry, and whole-cell enzymatic activity assay. Engineered P. putida cells were then applied to decolorize the anthraquinone dye Acid Green (AG 25 and diazo-dye Acid Red (AR 18. The results showed that decolorization of both dyes is Cu2+- and mediator-independent, with an optimum temperature of 35°C and pH of 3.0, and can be stably performed across a temperature range of 15°C to 45°C. A high activity toward AG25 (1 g/l with relative decolorization values of 91.2% (3 h and 97.1% (18 h, as well as high activity to AR18 (1 g/l by 80.5% (3 h and 89.0% (18 h, was recorded. The engineered system exhibited a comparably high activity compared with those of separate dyes in a continuous three-round shake-flask decolorization of AG25/AR18 mixed dye (each 1 g/l. No significant decline in decolorization efficacy was noted during first two-rounds but reaction equilibriums were elongated, and the residual laccase activity eventually decreased to low levels. However, the decolorizing capacity of the system was easily retrieved

  18. Evaluating the efficiency of two phase partitioning stirred tank bio-reactor for treating xylene vapors from the airstreamthrough a bed of Pseudomonas Putida

    Directory of Open Access Journals (Sweden)

    F. Golbabaei

    2015-04-01

    Conclusion: Overall, the results of the present research revealed that the application of two phase stirred tank bioreactors (TPPBs containing pure strains of Pseudomonas putida was successful for treatment of air streams with xylene.

  19. The crystal structure of Pseudomonas putida azoreductase - the active site revisited.

    Science.gov (United States)

    Gonçalves, Ana Maria D; Mendes, Sónia; de Sanctis, Daniele; Martins, Lígia O; Bento, Isabel

    2013-12-01

    The enzymatic degradation of azo dyes begins with the reduction of the azo bond. In this article, we report the crystal structures of the native azoreductase from Pseudomonas putida MET94 (PpAzoR) (1.60 Å), of PpAzoR in complex with anthraquinone-2-sulfonate (1.50 Å), and of PpAzoR in complex with Reactive Black 5 dye (1.90 Å). These structures reveal the residues and subtle changes that accompany substrate binding and release. Such changes highlight the fine control of access to the catalytic site that is required by the ping-pong mechanism, and in turn the specificity offered by the enzyme towards different substrates. The topology surrounding the active site shows novel features of substrate recognition and binding that help to explain and differentiate the substrate specificity observed among different bacterial azoreductases.

  20. Degradation of phenol and TCE using suspended and chitosan-bead immobilized Pseudomonas putida.

    Science.gov (United States)

    Chen, Yan-Min; Lin, Tsair-Fuh; Huang, Chih; Lin, Jui-Che; Hsieh, Feng-Ming

    2007-09-30

    The degradability of phenol and trichloroethene (TCE) by Pseudomonas putida BCRC 14349 in both suspended culture and immobilized culture systems are investigated. Chitosan beads at a size of about 1-2mm were employed to encapsulate the P. putida cells, becoming an immobilized culture system. The phenol concentration was controlled at 100 mg/L, and that of TCE was studied from 0.2 to 20 mg/L. The pH, between 6.7 and 10, did not affect the degradation of either phenol or TCE in the suspended culture system. However, it was found to be an important factor in the immobilized culture system in which the only significant degradation was observed at pH >8. This may be linked to the surface properties of the chitosan beads and its influence on the activity of the bacteria. The transfer yield of TCE on a phenol basis was almost the same for the suspended and immobilized cultures (0.032 mg TCE/mg phenol), except that these yields occurred at different TCE concentrations. The transfer yield at a higher TCE concentration for the immobilized system suggested that the cells immobilized in carriers can be protected from harsh environmental conditions. For kinetic rate interpretation, the Monod equation was employed to describe the degradation rates of phenol, while the Haldane's equation was used for TCE degradation. Based on the kinetic parameters obtained from the two equations, the rate for the immobilized culture systems was only about 1/6 to that of the suspended culture system for phenol degradation, and was about 1/2 for TCE degradation. The slower kinetics observed for the immobilized culture systems was probably due to the slow diffusion of substrate molecules into the beads. However, compared with the suspended cultures, the immobilized cultures may tolerate a higher TCE concentration as much less inhibition was observed and the transfer yield occurred at a higher TCE concentration.

  1. Proteomic analysis of Pseudomonas putida reveals an organic solvent tolerance-related gene mmsB.

    Directory of Open Access Journals (Sweden)

    Ye Ni

    Full Text Available Organic solvents are toxic to most microorganisms. However, some organic-solvent-tolerant (OST bacteria tolerate the destructive effects of organic solvent through various accommodative mechanisms. In this work, we developed an OST adapted strain Pseudomonas putida JUCT1 that could grow in the presence of 60% (v/v cyclohexane. Two-dimensional gel electrophoresis was used to compare and analyze the total cellular protein of P. putida JUCT1 growing with or without 60% (v/v cyclohexane. Under different solvent conditions, five high-abundance protein spots whose intensity values show over 60% discrepancies were identified by MALDI-TOF/TOF spectra. Specifically, they are arginine deiminase, carbon-nitrogen hydrolase family putative hydrolase, 3-hydroxyisobutyrate dehydrogenase, protein chain elongation factor EF-Ts, and isochorismatase superfamily hydrolase. The corresponding genes of the latter three proteins, mmsB, tsf, and PSEEN0851, were separately expressed in Escherichia coli to evaluate their effect on OST properties of the host strain. In the presence of 4% (v/v cyclohexane, E. coli harboring mmsB could grow to 1.70 OD(660, whereas cell growth of E. coli JM109 (the control was completely inhibited by 2% (v/v cyclohexane. Transformants carrying tsf or PSEEN0851 also showed an increased resistance to cyclohexane and other organic solvents compared with the control. Of these three genes, mmsB exhibited the most prominent effect on increasing OST of E. coli. Less oxidation product of cyclohexane was detected because mmsB transformants might help keep a lower intracellular cyclohexane level. This study demonstrates a feasible approach for elucidating OST mechanisms of microorganisms, and provides molecular basis to construct organic-solvent-tolerant strains for industrial applications.

  2. The T7-related Pseudomonas putida phage φ15 displays virion-associated biofilm degradation properties.

    Directory of Open Access Journals (Sweden)

    Anneleen Cornelissen

    Full Text Available Formation of a protected biofilm environment is recognized as one of the major causes of the increasing antibiotic resistance development and emphasizes the need to develop alternative antibacterial strategies, like phage therapy. This study investigates the in vitro degradation of single-species Pseudomonas putida biofilms, PpG1 and RD5PR2, by the novel phage ϕ15, a 'T7-like virus' with a virion-associated exopolysaccharide (EPS depolymerase. Phage ϕ15 forms plaques surrounded by growing opaque halo zones, indicative for EPS degradation, on seven out of 53 P. putida strains. The absence of haloes on infection resistant strains suggests that the EPS probably act as a primary bacterial receptor for phage infection. Independent of bacterial strain or biofilm age, a time and dose dependent response of ϕ15-mediated biofilm degradation was observed with generally a maximum biofilm degradation 8 h after addition of the higher phage doses (10(4 and 10(6 pfu and resistance development after 24 h. Biofilm age, an in vivo very variable parameter, reduced markedly phage-mediated degradation of PpG1 biofilms, while degradation of RD5PR2 biofilms and ϕ15 amplification were unaffected. Killing of the planktonic culture occurred in parallel with but was always more pronounced than biofilm degradation, accentuating the need for evaluating phages for therapeutic purposes in biofilm conditions. EPS degrading activity of recombinantly expressed viral tail spike was confirmed by capsule staining. These data suggests that the addition of high initial titers of specifically selected phages with a proper EPS depolymerase are crucial criteria in the development of phage therapy.

  3. Determinants of Pseudomonas putida WCS358 involved in inducing systemic resistance in plants.

    Science.gov (United States)

    Meziane, Hamid; VAN DER Sluis, Ientse; VAN Loon, Leendert C; Höfte, Monica; Bakker, Peter A H M

    2005-03-01

    SUMMARY Pseudomonas putida WCS358 is a plant growth-promoting rhizobacterium originally isolated from the rhizosphere of potato. It can suppress soil-borne plant diseases by siderophore-mediated competition for iron, but it has also been reported to result in induced systemic resistance (ISR) in Arabidopsis thaliana. Bacterial determinants of this strain involved in inducing systemic resistance in Arabidopsis were investigated using a Tn5 transposon mutant defective in biosynthesis of the fluorescent siderophore pseudobactin, a non-motile Tn5 mutant lacking flagella, and a spontaneous phage-resistant mutant lacking the O-antigenic side chain of the lipopolysaccharides (LPS). When using Pseudomonas syringae pv. tomato as the challenging pathogen, purified pseudobactin, flagella and LPS all triggered ISR. However, the mutants were all as effective as the parental strain, suggesting redundancy in ISR-triggering traits in WCS358. The Botrytis cinerea-tomato, B. cinerea-bean and Colletotrichum lindemuthianum-bean model systems were used to test further the potential of P. putida WCS358 to induce ISR. Strain WCS358 significantly reduced disease development in all three systems, indicating that also on tomato and bean WCS358 can trigger ISR. In both tomato and bean, the LPS mutant had lost the ability to induce resistance, whereas the flagella mutant was still effective. In bean, the pseudobactin mutant was still effective, whereas this mutant has lost its effectivity in tomato. In both bean and tomato, flagella isolated from the parental strain were not effective, whereas LPS or pseudobactin did induce systemic resistance.

  4. Contrasting colonization and plant growth promoting capacity between wild type and gfp-derative of the endophyte Pseudomonas putida W619 in hybrid poplar

    Energy Technology Data Exchange (ETDEWEB)

    Weyens N.; van der Lelie D.; Boulet, J.; Adriaensen, D.; Timmermans, J.-P.; Prinsen, E.; Van Oevelen, S.; D" Haen, J.; Smeets, K.; Taghavi, S.; Vangronsveld, J.

    2011-06-09

    This study aims to investigate the colonization of poplar by the endophyte Pseudomonas putida W619 and its capacity to promote plant growth. Poplar cuttings were inoculated with P. putida W619 (wild-type or gfp-labelled). The colonization of both strains was investigated and morphological, physiological and biochemical parameters were analyzed to evaluate plant growth promotion. Inoculation with P. putida W619 (wild-type) resulted in remarkable growth promotion, decreased activities of antioxidative defence related enzymes, and reduced stomatal resistance, all indicative of improved plant health and growth in comparison with the non-inoculated cuttings. In contrast, inoculation with gfp-labelled P. putida W619 did not promote growth; it even had a negative effect on plant health and growth. Furthermore, compared to the wildtype strain, colonization by the gfp-labelled P. putida W619::gfp1 was much lower; it only colonized the rhizosphere and root cortex while the wild-type strain also colonized the root xylem vessels. Despite the strong plant growth promoting capacity of P. putida W619 (wild-type), after gfp labelling its growth promoting characteristics disappeared and its colonization capacity was strongly influenced; for these reasons gfp labelling should be applied with sufficient caution.

  5. Nitrogen Removal Characteristics of Pseudomonas putida Y-9 Capable of Heterotrophic Nitrification and Aerobic Denitrification at Low Temperature

    Directory of Open Access Journals (Sweden)

    Yi Xu

    2017-01-01

    Full Text Available The cold-adapted bacterium Pseudomonas putida Y-9 was investigated and exhibited excellent capability for nitrogen removal at 15°C. The strain capable of heterotrophic nitrification and aerobic denitrification could efficiently remove ammonium, nitrate, and nitrite at an average removal rate of 2.85 mg, 1.60 mg, and 1.83 mg NL−1 h−1, respectively. Strain Y-9 performed nitrification in preference to denitrification when ammonium and nitrate or ammonium and nitrite coexisted in the solution. Meantime, the presence of nitrate had no effect on the ammonium removal rate of strain Y-9, and yet the presence of high concentration of nitrite would inhibit the cell growth and decrease the nitrification rate. The experimental results indicate that P. putida Y-9 has potential application for the treatment of wastewater containing high concentrations of ammonium along with its oxidation products at low temperature.

  6. Bioremoval of Basic Violet 3 and Acid Blue 93 by Pseudomonas putida and its adsorption isotherms and kinetics.

    Science.gov (United States)

    Arunarani, A; Chandran, Preethy; Ranganathan, B V; Vasanthi, N S; Sudheer Khan, S

    2013-02-01

    Basic Violet 3 and Acid Blue 93 are the most important group of synthetic colourants extensively used in textile industries for dyeing cotton, wool, silk and nylon. Release of these dye pollutants in to the environment adversely affects the human health and aquatic organisms. The present study we used Pseudomonas putida MTCC 4910 for the adsorptive removal of Basic Violet 3 and Acid Blue 93 from the aqueous solutions. The pH (4-9) and NaCl concentrations (1mM-1M) did not influence the adsorption process. The equilibrium adsorption process fitted well to Freundlich model than Langmuir model. The kinetics of adsorption fitted well by pseudo-second-order. Thus in the present study an attempt has been made to exploit the dye removal capability of P. putida MTCC 4910, and it was found to be an efficient microbe that could be used for bio removal of dyes from textile effluents.

  7. Degradation of o-chloronitrobenzene as the sole carbon and nitrogen sources by Pseudomonas putida OCNB-1

    Institute of Scientific and Technical Information of China (English)

    WU Haizhen; WEI Chaohai; WAMG Yaqin; HE Qincong; LIANG Shizhong

    2009-01-01

    A bacterial strain that utilized o-chloronitrobenzene (o-CNB) as the sole carbon, nitrogen and energy sources was isolated from an activated sludge collected from an industrial waste treatment plant. It was identified as Pseudomonas putida based on its morphology, physiological, and biochemical characteristics with an automatic biometrical system and the 16S rRNA sequence analysis. Microcosm study showed that the biodegradation of o-CNB was optimized at culture medium pH 8.0 and temperature of 32℃. At these conditions, the strain degraded 85% of o-CNB at a starting concentration of 1.1 mmol/L in 42 h. o-Chloroaniline was identified as the major metabolite with both high performance liquid chromatography (HPLC) and gas chromatography-mass spectrometry (GC-MS). The study showed that o-chloronitrobenzene degradation by Pseudomonas putida OCNB-1 was initiated by aniline dioxyenase, nitrobenzene reductase and catechol-1,2-dioxygenase.

  8. Characterization of a Pseudomonas putida rough variant evolved in a mixed species biofilm with Acinetobacter sp. strain C6

    DEFF Research Database (Denmark)

    Hansen, Susse Kirkelund; Haagensen, Janus Anders Juul; Gjermansen, Morten

    2007-01-01

    biosynthesis. Here we investigate further the biofilm physiology and the phenotypic characteristics of the selected P. putida rough colony variants. The coexistence of the P. putida population in a mixed-species biofilm with Acinetobacter sp. strain C6 is dependent on the benzoate excreted from Acinetobacter...... was shown to evolve rapidly by natural selection of better-adapted variants in a mixed-species biofilm consortium (S. K. Hansen, P. B. Rainey, J. A. Haagensen, and S. Molin, Nature 445:533-536, 2007). Adaptation was caused by mutations in a wapH homolog (PP4943) involved in core lipopolysaccharide...... during the catabolism of benzyl alcohol, the sole carbon source. Examination of biofilm development and the dynamics of the wild-type consortium revealed that the biofilm environment became oxygen limited, possibly with low oxygen concentrations around Acinetobacter microcolonies. In contrast to P...

  9. Plant Drought Tolerance Enhancement by Trehalose Production of Desiccation-Tolerant Microorganisms

    Science.gov (United States)

    Vílchez, Juan I.; García-Fontana, Cristina; Román-Naranjo, Desireé; González-López, Jesús; Manzanera, Maximino

    2016-01-01

    A collection of desiccation-tolerant xeroprotectant-producing microorganisms was screened for their ability to protect plants against drought, and their role as plant growth-promoting rhizobacteria was investigated in two different crops (tomato and pepper). The most commonly described biochemical mechanisms for plant protection against drought by microorganisms including the production of phytohormones, antioxidants and xeroprotectants were analyzed. In particular, the degree of plant protection against drought provided by these microorganisms was characterized. After studying the findings and comparing them with results of the closest taxonomic relatives at the species and strain levels, we propose that trehalose produced by these microorganisms is correlated with their ability to protect plants against drought. This proposal is based on the increased protection of plants against drought by the desiccation-sensitive microorganism Pseudomonas putida KT2440, which expresses the otsAB genes for trehalose biosynthesis in trans.

  10. The porous surface model, a novel experimental system for online quantitative observation of microbial processes under unsaturated conditions

    DEFF Research Database (Denmark)

    Dechesne, Arnaud; Or, D.; Gulez, Gamze

    2008-01-01

    Water is arguably the most important constituent of microbial microhabitats due to its control of physical and physiological processes critical to microbial activity. In natural environments, bacteria often live on unsaturated surfaces, in thin (micrometric) liquid films. Nevertheless...... of bacterial growth and activity under controlled unsaturated conditions. Bacteria are inoculated on a porous ceramic plate, wetted by a liquid medium. The thickness of the liquid film at the surface of the plate is set by imposing suction, corresponding to soil matric potential, to the liquid medium....... The utility of the PSM was demonstrated using Pseudomonas putida KT2440 tagged with gfp as a model bacterium. Single cells were inoculated at the surface of the PSM, and the rate at which colonies expanded laterally was measured for three matric potentials (–0.5, –1.2, and –3.6 kPa). The matric potential...

  11. Antimicrobial Activity of Commercial Nanoparticles

    Science.gov (United States)

    Gajjar, Priyanka; Pettee, Brian; Britt, David W.; Huang, Wenjie; Johnson, William P.; Anderson, Anne J.

    2009-07-01

    Engineered nanoparticles are finding increased use in applications ranging from biosensors to prophylactic antimicrobials embedded in socks. The release of heavy metal-containing nanoparticles (NP) into the environment may be harmful to the efficacy of beneficial microbes that function in element cycling, pollutant degradation, and plant growth. Antimicrobial activity of commercial NP of Ag, CuO, and ZnO is demonstrated here against the beneficial soil microbe, Pseudomonas putida KT2440, which was modified to serve as a bioluminescent sentinel organism. "As manufactured" preparations of nano- Ag, -CuO, and -ZnO caused rapid, dose dependent loss of light output in the biosensor. Bulk equivalents of these products showed no inhibitory activity, indicating that particle size was determinant in activity.

  12. Plant Drought Tolerance Enhancement by Trehalose Production of Desiccation-Tolerant Microorganisms

    Directory of Open Access Journals (Sweden)

    Juan Ignacio Vilchez Morillas

    2016-09-01

    Full Text Available A collection of desiccation-tolerant xeroprotectant-producing microorganisms was screened for their ability to protect plants against drought, and their role as plant growth-promoting rhizobacteria was investigated in two different crops (tomato and pepper. The most commonly described biochemical mechanisms for plant protection against drought by microorganisms including the production of phytohormones, antioxidants and xeroprotectants were analyzed. In particular, the degree of plant protection against drought provided by these microorganisms was characterized. After studying the findings and comparing them with results of the closest taxonomic relatives at the species and strain levels, we propose that trehalose produced by these microorganisms is correlated with their ability to protect plants against drought. This proposal is based on the increased protection of plants against drought by the desiccation-sensitive microorganism Pseudomonas putida KT2440, which expresses the otsAB genes for trehalose biosynthesis in trans.

  13. Hydration-controlled bacterial motility and dispersal on surfaces

    DEFF Research Database (Denmark)

    Dechesne, Arnaud; Wang, G.; Gulez, Gamze

    2010-01-01

    hydrated habitats, where water dynamics result in fragmented aquatic habitats connected by micrometric films, is debated. Here, we quantify the spatial dynamics of Pseudomonas putida KT2440 and its nonflagellated isogenic mutant as affected by the hydration status of a rough porous surface using......Flagellar motility, a mode of active motion shared by many prokaryotic species, is recognized as a key mechanism enabling population dispersal and resource acquisition in microbial communities living in marine, freshwater, and other liquid-replete habitats. By contrast, its role in variably...... an experimental system that mimics aquatic habitats found in unsaturated soils. The flagellar motility of the model soil bacterium decreased sharply within a small range of water potential (0 to −2 kPa) and nearly ceased in liquid films of effective thickness smaller than 1.5 μm. However, bacteria could rapidly...

  14. Kinetics Modelling of the Biodegradation of Benzene, Toluene and Phenol as Single Substrate and Mixed Substrate by Using Pseudomonas putida

    OpenAIRE

    Mathur, A K; Majumder, C. B.

    2010-01-01

    In the present work, kinetics of the biodegradation of benzene, toluene and phenol by using a pure culture of Pseudomonas putida (MTCC 1194) was determined by measuring the specific growth rate and degradation rate with substrate concentration as a function of time in a batch reactor. In general, the degradation rate of benzene, toluene and phenol increased with the increase in the initial substrate concentration and then decreased after reaching a maximum, showing substrate inhibition kineti...

  15. Dynamics of development and dispersal in sessile microbial communities: examples from Pseudomonas aeruginosa and Pseudomonas putida model biofilms

    DEFF Research Database (Denmark)

    Klausen, M.; Gjermansen, Morten; Kreft, J.-U.;

    2006-01-01

    Surface-associated microbial communities in many cases display dynamic developmental patterns. Model biofilms formed by Pseudomonas aeruginosa and Pseudomonas putida in laboratory flow-chamber setups represent examples of such behaviour. Dependent on the experimental conditions the bacteria...... organisms do not possess comprehensive genetic programs for biofilm development. Instead the bacteria appear to have evolved a number of different mechanisms to optimize surface colonization, of which they express a subset in response to the prevailing environmental conditions. These mechanisms include...

  16. The response of aggregated Pseudomonas putida CP1 cells to UV-C and UV-A/B disinfection.

    Science.gov (United States)

    Maganha de Almeida, Ana C; Quilty, Bríd

    2016-11-01

    UV radiation is a spread method used worldwide for the disinfection of water. However, much of the research on the disinfection of bacterial cells by UV has focused on planktonic cells. Many bacterial cells in nature are present in clumps or aggregates, and these aggregates, which are more resistant to disinfection than their planktonic counterparts, can be problematic in engineered water systems. The current research used Pseudomonas putida (P. putida) CP1, an environmental and non-pathogenic microorganism which autoaggregates when grown under certain conditions, as a model organism to simulate aggregated cells. The study investigated the response of both the planktonic and the aggregated forms of the bacterium to UV-C (λ = 253.7 nm) and UV-A/B (λ > 300 nm) disinfection at laboratory scale in a minimal medium. The planktonic cells of P. putida CP1 were inactivated within 60 s by UV-C and in 60 min by UV-A/B; however, the aggregated cells required 120 min of UV-C treatment and 240 min of UV-A/B radiation to become inactive. The size of the aggregate was reduced following UV treatment. Although all the cells had lost culturability, viability as measured by the LIVE/DEAD(®) stain and epifluorescence microscopy was not completely lost and the cells all demonstrated regrowth after overnight incubation in the dark.

  17. Evaluation of Zosteric Acid for Mitigating Biofilm Formation of Pseudomonas putida Isolated from a Membrane Bioreactor System

    Directory of Open Access Journals (Sweden)

    Andrea Polo

    2014-05-01

    Full Text Available This study provides data to define an efficient biocide-free strategy based on zosteric acid to counteract biofilm formation on the membranes of submerged bioreactor system plants. 16S rRNA gene phylogenetic analysis showed that gammaproteobacteria was the prevalent taxa on fouled membranes of an Italian wastewater plant. Pseudomonas was the prevalent genus among the cultivable membrane-fouler bacteria and Pseudomonas putida was selected as the target microorganism to test the efficacy of the antifoulant. Zosteric acid was not a source of carbon and energy for P. putida cells and, at 200 mg/L, it caused a reduction of bacterial coverage by 80%. Biofilm experiments confirmed the compound caused a significant decrease in biomass (−97% and thickness (−50%, and it induced a migration activity of the peritrichous flagellated P. putida over the polycarbonate surface not amenable to a biofilm phenotype. The low octanol-water partitioning coefficient and the high water solubility suggested a low bioaccumulation potential and the water compartment as its main environmental recipient and capacitor. Preliminary ecotoxicological tests did not highlight direct toxicity effects toward Daphnia magna. For green algae Pseudokirchneriella subcapitata an effect was observed at concentrations above 100 mg/L with a significant growth of protozoa that may be connected to a concurrent algal growth inhibition.

  18. Evaluation of Zosteric Acid for Mitigating Biofilm Formation of Pseudomonas putida Isolated from a Membrane Bioreactor System

    Science.gov (United States)

    Polo, Andrea; Foladori, Paola; Ponti, Benedetta; Bettinetti, Roberta; Gambino, Michela; Villa, Federica; Cappitelli, Francesca

    2014-01-01

    This study provides data to define an efficient biocide-free strategy based on zosteric acid to counteract biofilm formation on the membranes of submerged bioreactor system plants. 16S rRNA gene phylogenetic analysis showed that gammaproteobacteria was the prevalent taxa on fouled membranes of an Italian wastewater plant. Pseudomonas was the prevalent genus among the cultivable membrane-fouler bacteria and Pseudomonas putida was selected as the target microorganism to test the efficacy of the antifoulant. Zosteric acid was not a source of carbon and energy for P. putida cells and, at 200 mg/L, it caused a reduction of bacterial coverage by 80%. Biofilm experiments confirmed the compound caused a significant decrease in biomass (−97%) and thickness (−50%), and it induced a migration activity of the peritrichous flagellated P. putida over the polycarbonate surface not amenable to a biofilm phenotype. The low octanol-water partitioning coefficient and the high water solubility suggested a low bioaccumulation potential and the water compartment as its main environmental recipient and capacitor. Preliminary ecotoxicological tests did not highlight direct toxicity effects toward Daphnia magna. For green algae Pseudokirchneriella subcapitata an effect was observed at concentrations above 100 mg/L with a significant growth of protozoa that may be connected to a concurrent algal growth inhibition. PMID:24879523

  19. Experimental and kinetic study on the cometabolic biodegradation of phenol and 4-chlorophenol by psychrotrophic Pseudomonas putida LY1.

    Science.gov (United States)

    Wang, Qing; Li, Yi; Li, Jing; Wang, Yuming; Wang, Chao; Wang, Peifang

    2015-01-01

    This study investigated the kinetics of phenol and 4-chlorophenol (4-CP) biodegradation by a cold-adapted bacteria, Pseudomonas putida LY1, isolated from Songhua River sediment. The results showed that P. putida LY1 cannot grow on 4-CP as a sole carbon source. P. putida LY1 had the potential to cometabolic biodegrade phenol and 4-CP in a wide range of temperature (varying from 5 to 35 °C) with the optimal temperature around 25 °C. Mixture of phenol and 4-CP were completely removed at two 4-CP concentrations (15 and 40 mg/L) over a wide range of phenol (20-400 mg/L) concentrations, whereby the ratio of 4-CP/biomass (S 2/X) was lower than 0.03. The kinetic models of cometabolic biodegradation of phenol and 4-CP were proposed, considering the growth and nongrowth substrate inhibition. These models successfully simulate the processes of cometabolic degradation of phenol and 4-CP.

  20. Degradation of polyurethane by bacterium isolated from soil and assessment of polyurethanolytic activity of a Pseudomonas putida strain.

    Science.gov (United States)

    Peng, Yu-Huei; Shih, Yang-hsin; Lai, Yen-Chun; Liu, Yuan-Zan; Liu, Ying-Tong; Lin, Nai-Chun

    2014-01-01

    The increasing usage and the persistence of polyester polyurethane (PU) generate significant sources of environmental pollution. The effective and environmental friendly bioremediation techniques for this refractory waste are in high demand. In this study, three novel PU degrading bacteria were isolated from farm soils and activated sludge. Based upon 16S ribosomal RNA gene sequence blast, their identities were determined. Particularly robust activity was observed in Pseudomonas putida; it spent 4 days to degrade 92% of Impranil DLN(TM) for supporting its growth. The optimum temperature and pH for DLN removal by P. putida were 25 °C and 8.4, respectively. The degradation and transformation of DLN investigated by Fourier transformed infrared spectroscopy show the decrease in ester functional group and the emergence of amide group. The polyurethanolytic activities were both presented in the extracellular fraction and in the cytosol. Esterase activity was detected in the cell lysate. A 45-kDa protein bearing polyurethanolytic activity was also detected in the extracellular medium. This study presented high PU degrading activity of P. putida and demonstrated its responsible enzymes during the PU degradation process, which could be applied in the bioremediation and management of plastic wastes.

  1. Evaluation of medium-chain-length polyhydroxyalkanoate production by Pseudomonas putida LS46 using biodiesel by-product streams.

    Science.gov (United States)

    Fu, Jilagamazhi; Sharma, Umesh; Sparling, Richard; Cicek, Nazim; Levin, David B

    2014-07-01

    Medium-chain-length polyhydroxyalkanoate (mcl-PHA) production by Pseudomonas putida LS46 was analyzed in shake-flask-based batch reactions, using pure chemical-grade glycerol (PG), biodiesel-derived "waste" glycerol (WG), and biodiesel-derived "waste" free fatty acids (WFA). Cell growth, substrate consumption, mcl-PHA accumulation within the cells, and the monomer composition of the synthesized biopolymers were monitored. The patterns of mcl-PHA synthesis in P. putida LS46 cells grown on PG and WG were similar but differed from that of cells grown with WFA. Polymer accumulation in glycerol-based cultures was stimulated by nitrogen limitation and plateaued after 48 h in both PG and WG cultures, with a total accumulation of 17.9% cell dry mass and 16.3% cell dry mass, respectively. In contrast, mcl-PHA synthesis was independent of nitrogen concentration in P. putida LS46 cells cultured with WFA, which accumulated to 29% cell dry mass. In all cases, the mcl-PHAs synthesized consisted primarily of 3-hydroxyoctanoate (C(8)) and 3-hydroxydecanoate (C(10)). WG and WFA supported similar or greater cell growth and mcl-PHA accumulation than PG under the experimental conditions used. These results suggest that biodiesel by-product streams could be used as low-cost carbon sources for sustainable mcl-PHA production.

  2. Degradation of Tectilon Yellow 2G by hybrid technique: combination of sonolysis and biodegradation using mutant Pseudomonas putida.

    Science.gov (United States)

    Srinivasan, Raman; Kathiravan, Mathur Nadarajan; Gopinath, Kannappan Panchamoorthy

    2011-02-01

    Degradation of Tectilon Yellow 2G (TY2G), an azo dye has been studied by hybrid technique involving pretreatment by sonochemical method and further biological treatment by Pseudomonas putida mutant. Pretreatment experiments were carried out by sonolysis of the dye solution at different concentrations (100-1000 mg/L). Wild type Gram-negative P. putida species isolated from the textile effluent contaminated soil, which was found to be effective towards dye degradation, has been acclimatized so as to consume TY2G as the sole source of nutrition. Mutant strain was obtained from the acclimatized species by random mutagenesis using the chemical mutagen ethidium bromide for various time intervals (6-30 min). The optimum mutagenesis exposure time for obtaining the most efficient species for dye degradation was found to be 18 min. An efficient mutant strain P. putida ACT 1 has been isolated and was used for growth experiments. The mutant strain showed a better growth compared to the wild strain. The substrate utilization kinetics has been modeled using Monod and Haldane model equations of which the Haldane model provided a better fit. The enzyme kinetics of the mutant and wild species was obtained using Michaelis-Menten equation. The mutated species showed better enzyme kinetics towards the degradation of TY2G.

  3. A Novel Method for Production of 3-Hydroxydecanoic Acid by Recombinant Escherichia coli and Pseudomonas putida

    Institute of Scientific and Technical Information of China (English)

    郑重; 宫强; 陈国强

    2004-01-01

    3-hydroxydecanoic acid (3HD) is an interesting intermediate for chemical synthesis of many valuable compounds. A novel method to produce 3HD by recombinant bacteria was constructed in Escherichia coli HB101 and Pseudomonas putida GPp104, respectively. Simultaneous expression of both phaG encoding (R)-3-hydroxydecanoyl-ACP:CoA transacylase and tesB encoding thioesterase Ⅱ in E. coli HB101 increased 3HD production approximate 1.7-folds compared with the expression of phaG gene alone under identical conditions. In addition, when the tesB gene was introduced into the strain, the polyhydroxyalkanoate synthase negative strain P. putida GPpl04 produced extracellular 3HD. Thus, a novel pathway to produce 3HD by recombinant Pseudomonas was constructed. It was also found that the ratio of carbon source to nitrogen source affected the production of 3HD by recombinant P.putida harboring tesB gene. Nitrogen limitation seemed to promote the extracellular 3HD production.

  4. Characterization of a marine-isolated mercury-resistant Pseudomonas putida strain SP1 and its potential application in marine mercury reduction

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Weiwei; Chen, Lingxin; Liu, Dongyan [Chinese Academy of Sciences, Yantai, SD (China). Yantai Inst. of Coastal Zone Research (YICCAS); Chinese Academy of Sciences, Yantai, SD (China). Shandong Provincial Key Lab. of Coastal Zone Environmental Processes

    2012-02-15

    The Pseudomonas putida strain SP1 was isolated from marine environment and was found to be resistant to 280 {mu}M HgCl{sub 2}. SP1 was also highly resistant to other metals, including CdCl{sub 2}, CoCl{sub 2}, CrCl{sub 3}, CuCl{sub 2}, PbCl{sub 2}, and ZnSO{sub 4}, and the antibiotics ampicillin (Ap), kanamycin (Kn), chloramphenicol (Cm), and tetracycline (Tc). mer operon, possessed by most mercury-resistant bacteria, and other diverse types of resistant determinants were all located on the bacterial chromosome. Cold vapor atomic absorption spectrometry and a volatilization test indicated that the isolated P. putida SP1 was able to volatilize almost 100% of the total mercury it was exposed to and could potentially be used for bioremediation in marine environments. The optimal pH for the growth of P. putida SP1 in the presence of HgCl{sub 2} and the removal of HgCl{sub 2} by P. putida SP1 was between 8.0 and 9.0, whereas the optimal pH for the expression of merA, the mercuric reductase enzyme in mer operon that reduces reactive Hg{sup 2+} to volatile and relatively inert monoatomic Hg{sup 0} vapor, was around 5.0. LD50 of P. putida SP1 to flounder and turbot was 1.5 x 10{sup 9} CFU. Biofilm developed by P. putida SP1 was 1- to 3-fold lower than biofilm developed by an aquatic pathogen Pseudomonas fluorescens TSS. The results of this study indicate that P. putida SP1 is a low virulence strain that can potentially be applied in the bioremediation of HgCl{sub 2} contamination over a broad range of pH. (orig.)

  5. In situ gene expression in mixed-culture biofilms: Evidence of metabolic interactions between community members

    DEFF Research Database (Denmark)

    Møller, Søren; Sternberg, Claus; Andersen, Jens Bo

    1998-01-01

    in both community and pure-culture biofilms, while the Pm promoter was induced in the mixed community but not in a pure-culture biofilm. By sequentially adding community members, induction of Pm was shown to be a consequence of direct metabolic interactions between an Acinetobacter species and P. putida...... are involved in the biodegradation of toluene and related aromatic compounds. The upper-pathway promoter (Pu) and the meta-pathway promoter (Pm) from the TOL plasmid were fused independently to the gene coding for the green fluorescent protein (GFP), and expression from these promoters was studied in P. putida......, which was a dominant community member. Biofilms were cultured in flow chambers, which in combination with scanning confocal laser microscopy allowed direct monitoring of promoter activity with single-cell spatial resolution. Expression from the Pu promoter was homogeneously induced by benzyl alcohol...

  6. Toxicity Testing of Pristine and Aged Silver Nanoparticles in Real Wastewaters Using Bioluminescent Pseudomonas putida

    Directory of Open Access Journals (Sweden)

    Florian Mallevre

    2016-03-01

    Full Text Available Impact of aging on nanoparticle toxicity in real matrices is scarcely investigated due to a lack of suitable methodologies. Herein, the toxicity of pristine and aged silver nanoparticles (Ag NPs to a bioluminescent Pseudomonas putida bioreporter was measured in spiked crude and final wastewater samples (CWs and FWs, respectively collected from four wastewater treatment plants (WWTPs. Results showed lower toxicity of pristine Ag NPs in CWs than in FWs. The effect of the matrix on the eventual Ag NP toxicity was related to multiple physico-chemical parameters (biological oxygen demand (BOD, chemical oxygen demand (COD, total suspended solids (TSS pH, ammonia, sulfide and chloride based on a multivariate analysis. However, no collection site effect was concluded. Aged Ag NPs (up to eight weeks were found less toxic than pristine Ag NPs in CWs; evident increased aggregation and decreased dissolution were associated with aging. However, Ag NPs exhibited consistent toxicity in FWs despite aging; comparable results were obtained in artificial wastewater (AW simulating effluent. The study demonstrates the potency of performing nanoparticle acute toxicity testing in real and complex matrices such as wastewaters using relevant bacterial bioreporters.

  7. Inferring the Chemotactic Strategy of P. putida and E. coli Using Modified Kramers-Moyal Coefficients

    Science.gov (United States)

    Hintsche, Marius; Beta, Carsten; Stark, Holger

    2017-01-01

    Many bacteria perform a run-and-tumble random walk to explore their surrounding and to perform chemotaxis. In this article we present a novel method to infer the relevant parameters of bacterial motion from experimental trajectories including the tumbling events. We introduce a stochastic model for the orientation angle, where a shot-noise process initiates tumbles, and analytically calculate conditional moments, reminiscent of Kramers-Moyal coefficients. Matching them with the moments calculated from experimental trajectories of the bacteria E. coli and Pseudomonas putida, we are able to infer their respective tumble rates, the rotational diffusion constants, and the distributions of tumble angles in good agreement with results from conventional tumble recognizers. We also define a novel tumble recognizer, which explicitly quantifies the error in recognizing tumbles. In the presence of a chemical gradient we condition the moments on the bacterial direction of motion and thereby explore the chemotaxis strategy. For both bacteria we recover and quantify the classical chemotactic strategy, where the tumble rate is smallest along the chemical gradient. In addition, for E. coli we detect some cells, which bias their mean tumble angle towards smaller values. Our findings are supported by a scaling analysis of appropriate ratios of conditional moments, which are directly calculated from experimental data. PMID:28114420

  8. Immobilization and characterization of benzoylformate decarboxylase from Pseudomonas putida on spherical silica carrier.

    Science.gov (United States)

    Peper, Stephanie; Kara, Selin; Long, Wei Sing; Liese, Andreas; Niemeyer, Bernd

    2011-08-01

    If an adequate biocatalyst is identified for a specific reaction, immobilization is one possibility to further improve its properties. The immobilization allows easy recycling, improves the enzyme performance, and it often enhances the stability of the enzyme. In this work, the immobilization of the benzoylformate decarboxylase (BFD) variant, BFD A460I-F464I, from Pseudomonas putida was accomplished on spherical silica. Silicagel is characterized by its high mechanical stability, which allows its application in different reactor types without restrictions. The covalently bound enzyme was characterized in terms of its activity, stability, and kinetics for the formation of chiral 2-hydroxypropiophenone (2-HPP) from benzaldehyde and acetaldehyde. Moreover, temperature as well as pressure dependency of immobilized BFD A460I-F464I activity and enantioselectivity were analyzed. The used wide-pore silicagel shows a good accessibility of the immobilized enzyme. The activity of the immobilized BFD A460I-F464I variant was determined to be 70% related to the activity of the free enzyme. Thereby, the enantioselectivity of the enzyme was not influenced by the immobilization. In addition, a pressure-induced change in stereoselectivity was found both for the free and for the immobilized enzyme. With increasing pressure, the enantiomeric excess (ee) of (R)-2-HPP can be increased from 44% (0.1 MPa) to 76% (200 MPa) for the free enzyme and from 43% (0.1 MPa) to 66% (200 MPa) for the immobilized enzyme.

  9. Purification to Homogeneity and Characterization of a Novel Pseudomonas putida Chromate Reductase

    Science.gov (United States)

    Park, C. H.; Keyhan, M.; Wielinga, B.; Fendorf, S.; Matin, A.

    2000-01-01

    Cr(VI) (chromate) is a widespread environmental contaminant. Bacterial chromate reductases can convert soluble and toxic chromate to the insoluble and less toxic Cr(III). Bioremediation can therefore be effective in removing chromate from the environment, especially if the bacterial propensity for such removal is enhanced by genetic and biochemical engineering. To clone the chromate reductase-encoding gene, we purified to homogeneity (>600-fold purification) and characterized a novel soluble chromate reductase from Pseudomonas putida, using ammonium sulfate precipitation (55 to 70%), anion-exchange chromatography (DEAE Sepharose CL-6B), chromatofocusing (Polybuffer exchanger 94), and gel filtration (Superose 12 HR 10/30). The enzyme activity was dependent on NADH or NADPH; the temperature and pH optima for chromate reduction were 80°C and 5, respectively; and the Km was 374 μM, with a Vmax of 1.72 μmol/min/mg of protein. Sulfate inhibited the enzyme activity noncompetitively. The reductase activity remained virtually unaltered after 30 min of exposure to 50°C; even exposure to higher temperatures did not immediately inactivate the enzyme. X-ray absorption near-edge-structure spectra showed quantitative conversion of chromate to Cr(III) during the enzyme reaction. PMID:10788340

  10. Purification to homogeneity and characterization of a novel Pseudomonas putida chromate reductase

    Energy Technology Data Exchange (ETDEWEB)

    Park, C.H.; Keyhan, M.; Wielinga, B.; Fendorf, S.; Matin, A.

    2000-05-01

    Cr(VI) (chromate) is a widespread environmental contaminant. Bacterial chromate reductases can convert soluble and toxic chromate to the insoluble and less toxic Cr(III). Bioremediation can therefore be effective in removing chromate from the environment, especially if the bacterial propensity for such removal is enhanced by genetic and biochemical engineering. To clone the chromate reductase-encoding gene, the authors purified to homogeneity and characterized a novel soluble chromate reductase from Pseudomonas putida, using ammonium sulfate precipitation, anion-exchange chromatography, chromatofocusing, and gel filtration. The enzyme activity was dependent on NADH or NADPH; the temperature and pH optima for chromate reduction were 80 C and 5, respectively; and the K{sub m} was 374 {micro}M, with a V{sub max} of 1.72 {micro}mol/min/mg of protein. Sulfate inhibited the enzyme activity noncompetitively. The reductase activity remained virtually unaltered after 30 min of exposure to 50 C; even exposure to higher temperatures did not immediately inactivate the enzyme. X-ray absorption near-edge-structure spectra showed quantitative conversion of chromate to Cr(III) during the enzyme reaction.

  11. Pseudomonas fluorescens and Pseudomonas putida for Promoting Growth of Jatropha curcas Seedling Root

    Directory of Open Access Journals (Sweden)

    Sri Sumarsih

    2012-05-01

    Full Text Available Pseudomonas fluorescens and P. putida are Plant Growth Promoting Rhizobacteria (PGPR that can produce growth hormone. The objective of this study is to know the effects of those two combined species of PGPR on seedling root growth of Jatropha curcas. The condition of the seedling root determines the success of dry land cultivation. The root which has wider coverage, is larger in number, and is bigger in diameter makes seedling more resistant to stress in dry land environment. In the experiment, two kinds of plant materials are used for seedling, the Jatropha seed and stem material, which are treated in a mixed culture of PGPR. For the Jatropha seed, this mixed culture of PGPR is given at the same time of cultivating the sprout on the seedling medium. For the stem cutting, the PGPR is poured in together during the first watering of the seedling cultivation medium. In the fourthweek, the observed growth parameters are root length, root diameter, primary and secondary lateral root numbers, Root Length Density (RLD, Frequency of Lateral Root (FLR, and Specific Root Length (SRL. These data are analyzed using analysis of variant with DMRT test at 0.05 level of significance. The result of this study shows that PGPR tend to reduce FLR values on the seedling root made from seeds. On the seedling root made from stem cutting, PGPR increase the root length, primary and secondary lateral root numbers, root diameter, FLR and SRL values as well.

  12. ISOLATION AND IDENTIFICATION OF A THERMOTOLERANT PLANT GROWTH PROMOTING PSEUDOMONAS PUTIDA PRODUCING TREHALOSE SYNTHASE

    Directory of Open Access Journals (Sweden)

    Ali Sk.Z.

    2013-08-01

    Full Text Available A thermotolerant plant growth promoting Pseudomonas isolate growing at 40oC producing trehalose synthase (TreS was isolated from rhizosphere soil under semi arid conditions of India. Trehalose synthase was extracted; purified and enzymatic activity was examined at various temperatures and pH. The optimum temperature and pH was 38oC and pH 7.5 and the activity declined at above or below the optimum pH and temperature. The enzyme was active on maltose and trehalose among saccharides tested. The enzyme had a higher catalytic activity for maltose with a trehalose yield of 72% than for trehalose where 30% yield of maltose was achieved, indicating maltose as preferred substrate. The isolate showed multiple plant growth promoting traits (indole acetic acid (IAA, phosphate solubilization, siderophore and ammonia both at ambient (28oC and high temperature (40oC. Based on phenotypic and 16SrRNA analysis the isolate was identified as Pseudomonas putida (Accession No. GU396283.

  13. Synergistic effect of Pseudomonas putida and Bacillus amyloliquefaciens ameliorates drought stress in chickpea (Cicer arietinum L.).

    Science.gov (United States)

    Kumar, Manoj; Mishra, Sankalp; Dixit, Vijaykant; Kumar, Manoj; Agarwal, Lalit; Chauhan, Puneet Singh; Nautiyal, Chandra Shekhar

    2016-01-01

    Two plant growth promoting rhizobacteria (PGPR) Pseudomonas putida NBRIRA and Bacillus amyloliquefaciens NBRISN13 with ability to tolerate abiotic stress along with multiple PGP traits like ACC deaminase activity, minerals solubilisation, hormones production, biofilm formation, siderophore activity were evaluated for their synergistic effect to ameliorate drought stress in chickpea. Earlier we have reported both the strains individually for their PGP attributes and stress amelioration in host plants. The present study explains in detail the possibilities and benefits of utilizing these 2 PGPR in consortium for improving the chickpea growth under control and drought stressed condition. In vitro results clearly demonstrate that both the PGPR strains are compatible to each other and their synergistic growth enhances the PGP attributes. Greenhouse experiments were conducted to evaluate the effect of inoculation of both strains individually and consortia in drought tolerant and sensitive cultivars (BG362 and P1003). The growth parameters were observed significantly higher in consortium as compared to individual PGPR. Colonization of both PGPR in chickpea rhizosphere has been visualized by using gfp labeling. Apart from growth parameters, defense enzymes, soil enzymes and microbial diversity were significantly modulated in individually PGPR and in consortia inoculated plants. Negative effects of drought stress has been ameliorated and apparently seen by higher biomass and reversal of stress indicators in chickpea cultivars treated with PGPR individually or in consortia. Findings from the present study demonstrate that synergistic application has better potential to improve plant growth promotion under drought stress conditions.

  14. Electricity Generation and Wastewater Treatment of Oil Refinery in Microbial Fuel Cells Using Pseudomonas putida

    Directory of Open Access Journals (Sweden)

    Dip Majumder

    2014-09-01

    Full Text Available Microbial fuel cells (MFCs represent a novel platform for treating wastewater and at the same time generating electricity. Using Pseudomonas putida (BCRC 1059, a wild-type bacterium, we demonstrated that the refinery wastewater could be treated and also generate electric current in an air-cathode chamber over four-batch cycles for 63 cumulative days. Our study indicated that the oil refinery wastewater containing 2213 mg/L (ppm chemical oxygen demand (COD could be used as a substrate for electricity generation in the reactor of the MFC. A maximum voltage of 355 mV was obtained with the highest power density of 0.005 mW/cm2 in the third cycle with a maximum current density of 0.015 mA/cm2 in regard to the external resistor of 1000 Ω. A maximum coulombic efficiency of 6 × 10−2% was obtained in the fourth cycle. The removal efficiency of the COD reached 30% as a function of time. Electron transfer mechanism was studied using cyclic voltammetry, which indicated the presence of a soluble electron shuttle in the reactor. Our study demonstrated that oil refinery wastewater could be used as a substrate for electricity generation.

  15. Electricity generation and wastewater treatment of oil refinery in microbial fuel cells using Pseudomonas putida.

    Science.gov (United States)

    Majumder, Dip; Maity, Jyoti Prakash; Tseng, Min-Jen; Nimje, Vanita Roshan; Chen, Hau-Ren; Chen, Chien-Cheng; Chang, Young-Fo; Yang, Tsui-Chu; Chen, Chen-Yen

    2014-09-22

    Microbial fuel cells (MFCs) represent a novel platform for treating wastewater and at the same time generating electricity. Using Pseudomonas putida (BCRC 1059), a wild-type bacterium, we demonstrated that the refinery wastewater could be treated and also generate electric current in an air-cathode chamber over four-batch cycles for 63 cumulative days. Our study indicated that the oil refinery wastewater containing 2213 mg/L (ppm) chemical oxygen demand (COD) could be used as a substrate for electricity generation in the reactor of the MFC. A maximum voltage of 355 mV was obtained with the highest power density of 0.005 mW/cm² in the third cycle with a maximum current density of 0.015 mA/cm² in regard to the external resistor of 1000 Ω. A maximum coulombic efficiency of 6 × 10⁻²% was obtained in the fourth cycle. The removal efficiency of the COD reached 30% as a function of time. Electron transfer mechanism was studied using cyclic voltammetry, which indicated the presence of a soluble electron shuttle in the reactor. Our study demonstrated that oil refinery wastewater could be used as a substrate for electricity generation.

  16. Suicide inactivation of catechol 2,3-dioxygenase from Pseudomonas putida mt-2 by 3-halocatechols

    Energy Technology Data Exchange (ETDEWEB)

    Bartels, I.; Knackmuss, H.J.; Reineke, W.

    1984-03-01

    The inactivation of catechol 2,3-dioxygenase from Pseudomonas putida mt-2 by 3-chloro- and 3-fluorocatechol and the iron-chelating agent Tiron (catechol-3,5-disulfonate) was studied. Whereas inactivation by Tiron is an oxygen-independent and mostly reversible process, inactivation by the 3-halocatechols was only observed in the presence of oxygen and was largely irreversible. The rate constants for inactivation (K/sub 2/) were 1.62 x 10/sup -3/ sec/sup -1/ for 3-chlorocatechol and 2.38 x 10/sup -3/ sec/sup -1/ for 3-fluorocatechol. The inhibitor constants (K/sub i/) were 23 ..mu..M for 3-chlorocatechol and 17 ..mu..M for 3-fluorocatechol. The kinetic data for 3-fluorocatechol could only be obtained in the presence of 2-mercaptoethanol. Besides inactivated enzyme, some 2-hydroxyhexa-2,4-dienoic acid as the actual suicide product of meta-cleavage. A side product of 3-fluorocatechol cleavage is a yellow compound with the spectral characteristics of a 2-hydroxy-6-oxohexa-2,4-dienoci acid indicating 1,6-cleavage. Rates of inactivation by 3-fluorocatechol were reduced in the presence of superoxide dismutase, catalase, formate, and mannitol, which implies that superoxide anion, hydrogen peroxide, and hydroxyl radical exhibit additional inactivation. 64 references.

  17. Biosorption of cadmium, copper, lead and zinc by inactive biomass of Pseudomonas Putida

    Energy Technology Data Exchange (ETDEWEB)

    Pardo, Rafael; Herguedas, Mar; Barrado, Enrique; Vega, Marisol [Departamento de Quimica Analitica, Universidad de Valladolid, Facultad de Ciencias, Prado de la Magdalena, 47005, Valladolid (Spain)

    2003-05-01

    The accumulation of Cd(II), Cu(II), Pb(II) and Zn(II) at mg L{sup -1} concentration levels by inactive freeze-dried biomass of Pseudomonas Putida has been investigated. These metals could be efficiently removed from diluted aqueous solutions. A contact time of 10 min was sufficient to reach equilibrium. The pH has a strong effect on metal biosorption and the optimal pH values were 6.0, 5.0-6.0, 6.0-6.5 and 7.0-7.5 for Cd(II), Cu(II), Pb(II) and Zn(II) respectively. Under these conditions there was 80% removal for all metals studied. The process of biosorption can be described by a Langmuir-type adsorption model. This model accounts for 98% of the data variance. The K{sub A} and q{sub max} parameters for each metal are strongly correlated (at confidence levels greater than 98%) with the metal acidity, quantified by the constant of the corresponding M(OH){sup +} complex, thus confirming previous assertions by other authors. (orig.)

  18. Lead Sorption from Aqueous Solutions by Pseudomonas putida (p168 and its Composites with Palygorskite and Sepiolite Clays

    Directory of Open Access Journals (Sweden)

    Marzieh tavanaei

    2017-02-01

    Full Text Available Introduction: Heavy metals contamination due to natural and anthropogenic sources is a global environmental concern. Lead (Pb is one of the very toxic heavy metals. Industrial production processes and their emissions, mining operation, smelting, combustion sources and solid waste incinerators are the primary sources of lead. This heavy metal has aberrant effects on the environment and living organisms. Hence, proper treatment of lead from soil and industrial wastewaters is very important. In order to remove toxic heavy metals from contaminated water systems, conventional methods such as chemical precipitation, coagulation, ion exchange, solvent extraction and filtration, evaporation and membrane methods are being used. These conventional methods generally have high costs and technical problems. Therefore, biosorption processes, in which microorganisms are used as sorbents, have been considered as economical and environmentally friendly options for removal of heavy metals from aqueous solution. Clay minerals are another group of sorbents used in removal of heavy metals from polluted environments. Furthermore, bacterial cells can be attached on clay mineral surfaces and form bacteria-mineral composites. These composites adsorb heavy metals and convert them into forms with low mobility and bioavailability. Pseudomonas putida is a unique microorganism with a high tendency to sorb and/or degrade certain environmental pollutants. Palygorskite and sepiolite are the fibrous clay minerals of arid and semiarid regions; their structures consist of ribbons and channels. These fibrous minerals have various applications in industry and the environment because of its large surface area and high adsorption capacity. The present study was conducted in order to determine the ability of Pseudomonas putida (P168, and its composites with palygorskite and sepiolite in lead sorption. Materials and Methods: The bacterial strain used in the present study was Pseudomonas

  19. Assessment of Qualitative and Quantitative Characters of Two Persian Clover Ecotypes Inoculated by Rhizobium leguminosarum biovartrifoli and Pesudomonas putida Bacteria

    Directory of Open Access Journals (Sweden)

    R Azamei

    2016-02-01

    Full Text Available Introduction Over the past decades, world attitude has changed towards the reduction of environmental pollutants. Harmful effects of synthetic fertilizers on environment have been identified. Bio-fertilizers are not harmful to the environment, but also they have favorable effects on plant growth processes. Soil biotechnology can be defined as the study of soil organisms and their metabolic processes which may have positive effects on plant yields. The main goal of this study is to asess the biotechnology fertilizers beneficial effects on soil organisms and their subsequently to maximize the yield. It is also our desire conside the soil quality, hygiene and environmental protection along this process. Among the strain of nitrogen-fixing bacteria, symbiotic bacteria such as rhizobium bacteria are important and essential in planning the sustainable farming systems. Several studies have shown that crop varieties which inoculated with rhizobium and pseudomonas were superior in yield production and performance. Material and Methods An experiment was designed as factorial performed in randomized complete block design (RCBD with three replications in Agricultural Research Center of Golpayegan (Isfahan during 2010 – 2011. the purpose of this study was to evaluate the effects of inoculation of two ecotypes of Persian clover by various strains of Rhizobium leguminosarum. Biovar trifoli bacteria accompanied with plant growth promoting rhizobacteria (PGPR Pseudomonas putida was employed to find certain qualitative and quantitative characteristics of clover yield, The main plots included two local ecotypes of Persian clover; Arak Haft Chin (V1 and Isfahan Haft Chin (V2, the subplots included inoculation by two strain of Rhizobium; Rb-3, Rb-13 and one strain of Pseudomonas; PS -168.4 cuts were performed during the experiment and 60 kg/ha seed was used for cultivation based on local knowledge. According to recommendations of the Institute of Soil and Water

  20. PpoR is a conserved unpaired LuxR solo of Pseudomonas putida which binds N-acyl homoserine lactones

    Directory of Open Access Journals (Sweden)

    Venturi Vittorio

    2009-06-01

    Full Text Available Abstract Background Only a small number of Pseudomonas putida strains possess the typical N-acyl homoserine lactone quorum sensing system (AHL QS that consists of a modular LuxR family protein and its cognate LuxI homolog that produces the AHL signal. Moreover, AHL QS systems in P. putida strains are diverse in the type of AHLs they produce and the phenotypes that they regulate. Results We identified an unpaired LuxR solo (QS luxR homolog that occurs without the corresponding luxI homolog, which is highly conserved in both the AHL producing and non-AHL producing P. putida strains that we analyzed. In this study we report the cloning and functional characterization of this unpaired LuxR homolog designated PpoR. An AHL binding assay showed that PpoR protein binds to 3-oxo-C6-HSL. Studies using a ppoR promoter-lacZ reporter fusion revealed that it exhibits stringent growth phase dependent expression. Functional interaction of PpoR with the endogenous complete AHL QS systems of P. putida WCS358 (PpuI/R system and PpoR was also investigated. Microarray analysis of P. putida WCS358 wild type and a PpoR over-expressing strain revealed several putative target genes that may be directly or indirectly regulated by PpoR. Conclusion Our results indicate that PpoR in P. putida strains may have a conserved role in detecting an AHL signal, either self or foreign, and regulating specific target genes.

  1. Liquid chromatography time of flight mass spectrometry based environmental metabolomics for the analysis of Pseudomonas putida Bacteria in potable water.

    Science.gov (United States)

    Kouremenos, Konstantinos A; Beale, David J; Antti, Henrik; Palombo, Enzo A

    2014-09-01

    Water supply biofilms have the potential to harbour waterborne diseases, accelerate corrosion, and contribute to the formation of tuberculation in metallic pipes. One particular species of bacteria known to be found in the water supply networks is Pseudomonas sp., with the presence of Pseudomonas putida being isolated to iron pipe tubercles. Current methods for detecting and analysis pipe biofilms are time consuming and expensive. The application of metabolomics techniques could provide an alternative method for assessing biofilm risk more efficiently based on bacterial activity. As such, this paper investigates the application of metabolomic techniques and provides a proof-of-concept application using liquid chromatography coupled with time-of-flight mass spectrometry (LC-ToF-MS) to three biologically independent P. putida samples, across five different growth conditions exposed to solid and soluble iron (Fe). Analysis of the samples in +ESI and -ESI mode yielded 887 and 1789 metabolite features, respectively. Chemometric analysis of the +ESI and -ESI data identified 34 and 39 significant metabolite features, respectively, where features were considered significant if the fold change was greater than 2 and obtained a p-value less than 0.05. Metabolite features were subsequently identified according to the Metabolomics Standard Initiative (MSI) Chemical Analysis Workgroup using analytical standards and standard online LC-MS databases. Possible markers for P. putida growth, with and without being exposed to solid and soluble Fe, were identified from a diverse range of different chemical classes of metabolites including nucleobases, nucleosides, dipeptides, tripeptides, amino acids, fatty acids, sugars, and phospholipids. Copyright © 2014 Elsevier B.V. All rights reserved.

  2. C5-technology in Pseudomonas putida S12: Construction, analysis and implementation of D-xylose metabolic pathways

    NARCIS (Netherlands)

    Meijnen, J.P.

    2010-01-01

    The rising price of oil and impending deficit of fossil resources stimulate the development of alternative processes for the production of chemicals. The production of biochemicals from lignocellulosic biomass is a promising alternative. Lignocellulosic biomass consists of a mixture of sugars that c

  3. Cow Dung Substrate for the Potential Production of Alkaline Proteases by Pseudomonas putida Strain AT in Solid-State Fermentation

    OpenAIRE

    Ponnuswamy Vijayaraghavan; Sreekumar Saranya; Samuel Gnana Prakash Vincent

    2014-01-01

    Cow dung and agroresidues were used as the substrates for the production of alkaline proteases by Pseudomonas putida strain AT in solid-state fermentation. Among the various substrates evaluated, cow dung supported maximum (1351±217 U/g) protease production. The optimum conditions for the production of alkaline proteases were a fermentation period of 48 h, 120% (v/w) moisture, pH 9, and the addition of 6% (v/w) inoculum, 1.5% (w/w) trehalose, and 2.0% (w/w) yeast extract to the cow dung subst...

  4. Genomic analysis of Pseudomonas putida phage tf with localized single-strand DNA interruptions.

    Directory of Open Access Journals (Sweden)

    Anatoly S Glukhov

    Full Text Available The complete sequence of the 46,267 bp genome of the lytic bacteriophage tf specific to Pseudomonas putida PpG1 has been determined. The phage genome has two sets of convergently transcribed genes and 186 bp long direct terminal repeats. The overall genomic architecture of the tf phage is similar to that of the previously described Pseudomonas aeruginosa phages PaP3, LUZ24 and phiMR299-2, and 39 out of the 72 products of predicted tf open reading frames have orthologs in these phages. Accordingly, tf was classified as belonging to the LUZ24-like bacteriophage group. However, taking into account very low homology levels between tf DNA and that of the other phages, tf should be considered as an evolutionary divergent member of the group. Two distinguishing features not reported for other members of the group were found in the tf genome. Firstly, a unique end structure--a blunt right end and a 4-nucleotide 3'-protruding left end--was observed. Secondly, 14 single-chain interruptions (nicks were found in the top strand of the tf DNA. All nicks were mapped within a consensus sequence 5'-TACT/RTGMC-3'. Two nicks were analyzed in detail and were shown to be present in more than 90% of the phage population. Although localized nicks were previously found only in the DNA of T5-like and phiKMV-like phages, it seems increasingly likely that this enigmatic structural feature is common to various other bacteriophages.

  5. Phenol biodegradation by immobilized Pseudomonas putida FNCC-0071 cells in alginate beads

    Science.gov (United States)

    Hakim, Lukman Nul; Rochmadi, Sutijan

    2017-06-01

    Phenol is one of industrial liquid waste which is harmful to the environment, so it must be degraded. It can be degraded by immobilized Pseudomonas putida FNCC-0071 cells. It needs the kinetics and mass transfer data to design this process which can be estimated by the proposed dynamic model in this study. This model involves simultaneous diffusion and reaction in the alginate bead and liquid bulk. The preliminary stage of phenol biodegradation process was acclimatization cells. This is the stage where cells were acclimated to phenol as carbon source (substrate). Then the acclimated cells were immobilized in alginate beads by extrusion method. The variation of the initial phenol concentration in the solution is 350 to 850 ppm where 60 g alginate bead contained by cells loaded into its solution in reactor batch, so then biodegradation occurs. In this study, the average radius of alginate bead was 0.152 cm. The occurred kinetic reaction process can be explained by Blanch kinetic model with the decreasing of parameter μmax' while the increasing values of initial phenol concentration in the same time, but the parameters KM, KM', and kt were increasing by the rising values of initial phenol concentration. The value of the parameter β is almost zero. Effective diffusivity of phenol and cells are 1.11 × 10-5±4.5% cm2 s-1 and 1.39 × 10-7± 0.04% cm2 s-1. The partition coefficient of phenol and cells are 0.39 ± 15% and 2.22 ± 18%.

  6. Broad-Host-Range Expression Reveals Native and Host Regulatory Elements That Influence Heterologous Antibiotic Production in Gram-Negative Bacteria.

    Science.gov (United States)

    Zhang, Jia Jia; Tang, Xiaoyu; Zhang, Michelle; Nguyen, Darlene; Moore, Bradley S

    2017-09-05

    Heterologous expression has become a powerful tool for studying microbial biosynthetic gene clusters (BGCs). Here, we extend the transformation-associated recombination cloning and heterologous expression platform for microbial BGCs to include Gram-negative proteobacterial expression hosts. Using a broad-host-range expression platform, we test the implicit assumption that biosynthetic pathways are more successfully expressed in more closely related heterologous hosts. Cloning and expression of the violacein BGC from Pseudoalteromonas luteoviolacea 2ta16 revealed robust production in two proteobacterial hosts, Pseudomonas putida KT2440 and Agrobacterium tumefaciens LBA4404, but very little production of the antibiotic in various laboratory strains of Escherichia coli, despite their closer phylogenetic relationship. We identified a nonclustered LuxR-type quorum-sensing receptor from P. luteoviolacea 2ta16, PviR, that increases pathway transcription and violacein production in E. coli by ∼60-fold independently of acyl-homoserine lactone autoinducers. Although E. coli harbors the most similar homolog of PviR identified from all of the hosts tested, overexpression of various E. coli transcription factors did not result in a statistically significant increase in violacein production, while overexpression of two A. tumefaciens PviR homologs significantly increased production. Thus, this work not only introduces a new genetic platform for the heterologous expression of microbial BGCs, it also challenges the assumption that host phylogeny is an accurate predictor of host compatibility.IMPORTANCE Although Gram-positive heterologous hosts such as Streptomyces have been developed and optimized to support diverse secondary metabolic reactions, there has been comparatively less work on Gram-negative hosts, some of which grow faster and are easier to work with. This work presents a new genetic platform for direct cloning and broad-host-range heterologous expression of BGCs

  7. Isolation of a diphenylamine-degrading bacterium and characterization of its metabolic capacities, bioremediation and bioaugmentation potential.

    Science.gov (United States)

    Perruchon, Chiara; Batianis, Christos; Zouborlis, Stelios; Papadopoulou, Evangelia S; Ntougias, Spyridon; Vasileiadis, Sotirios; Karpouzas, Dimitrios G

    2015-12-01

    The antioxidant diphenylamine (DPA) is used in fruit-packaging plants for the control of the physiological disorder apple scald. Its use results in the production of DPA-contaminated wastewater which should be treated before finally discharged. Biological treatment systems using tailored-made microbial inocula with specific catabolic activities comprise an appealing and sustainable solution. This study aimed to isolate DPA-degrading bacteria, identify the metabolic pathway of DPA and evaluate their potential for future implementation in bioremediation and biodepuration applications. A Pseudomonas putida strain named DPA1 able to rapidly degrade and utilize DPA as the sole C and N source was enriched from a DPA-contaminated soil. The isolated strain degraded spillage-level concentrations of DPA in liquid culture (2000 mg L(-1)) and in contaminated soil (1000 mg kg(-1)) and metabolized DPA via the transient formation of aniline and catechol. Further evidence for the bioremediation and biodepuration potential of the P. putida strain DPA1 was provided by its capacity to degrade the post-harvest fungicide ortho-phenylphenol (OPP), concurrently used by the fruit-packaging plants, although at slower rates and DPA in a wide range of pH (4.5-9) and temperatures (15-37 °C). These findings revealed the high potential of the P. putida strain DPA1 for use in future soil bioremediation strategies and/or as start-up inocula in wastewater biodepuration systems.

  8. Antimicrobial effect of TiO2 doped with Ag and Cu on Escherichia coli and Pseudomonas putida

    Science.gov (United States)

    Angelov, O.; Stoyanova, D.; Ivanova, I.

    2016-10-01

    Antimicrobial effect of TiO2 doped with Ag and Cu on Gram-negative bacteria Escherichia coli and Pseudomonas putida is studied. The thin films are deposited on glass substrates without heating during the deposition by r.f. magnetron co-sputtering of TiO2 target and pieces of Ag and Cu. The studied films, thickness about 65 nm, were as deposited and annealed (5200C, 4h, N2+5%H2, 4Pa). The as deposited thin films TiO2:Ag:Cu have band gap energy of 3.56 eV little higher than the band gap of crystalline anatase TiO2 which can be explained with the quantum effect of the granular structure of r.f. magnetron sputtered films. The annealed samples have band gap of 2.52 eV due to formation of donor levels from Ag and Cu atoms near the bottom of the conduction band. The toxic effect was determined through the classical Koch's method and the optical density measurements at λ=610 nm. The as deposited TiO2:Ag:Cu thin films demonstrate stronger inhibition effect - bactericidal for P. putida and bacteriostatic for E. coli (up to the 6th hour) in comparison with the annealed samples. The both methods of study show the same trends of the bacterial growth independently of their different sensitivity which confirms the observed effect.

  9. Toxicities effects of pharmaceutical, olive mill and textile wastewaters before and after degradation by Pseudomonas putida mt-2

    Directory of Open Access Journals (Sweden)

    Mansour Hedi

    2012-02-01

    Full Text Available Abstract Background Removal of numerous classes of chemical pollutants from the industrial wastewater such as textile, pharmaceutical and olive mill using conventional wastewater treatment, is incomplete and several studies suggested that improvement of this situation would require the application of biological treatment techniques. Dyes, polyphenols and drugs are an environmental pollutants extremely toxics to plants and other living organisms including humans. These effluents were previously treated by Pseudomonas putida. The main of this work was to evaluate the in vivo toxicity of the three wastewaters. Methods Writhes and convulsant effect of effluents were carried out and were compared to the treated effluents. Only pharmaceutical wastewater was exhibited a convulsant effect which observed in mice treated by effluent. On the other hand, all industrial wastewater induced significantly an algogenic effects particularly when mice were treated by the pharmaceutical wastewater (Number of writhes = 44. Conclusion Toxicity was totally removed when mice were treated by the bio remediated effluent. This indicates that P. putida was able to completely detoxify the toxic industrial effluent.

  10. N-acyl Homoserine Lactone-Producing Pseudomonas putida Strain T2-2 from Human Tongue Surface

    Directory of Open Access Journals (Sweden)

    Yeun Mun Choo

    2013-09-01

    Full Text Available Bacterial cell-to-cell communication (quorum sensing refers to the regulation of bacterial gene expression in response to changes in microbial population density. Quorum sensing bacteria produce, release and respond to chemical signal molecules called autoinducers. Bacteria use two types of autoinducers, namely autoinducer-1 (AI-1 and autoinducer-2 (AI-2 where the former are N-acylhomoserine lactones and the latter is a product of the luxS gene. Most of the reported literatures show that the majority of oral bacteria use AI-2 for quorum sensing but rarely the AI-1 system. Here we report the isolation of Pseudomonas putida strain T2-2 from the oral cavity. Using high resolution mass spectrometry, it is shown that this isolate produced N-octanoylhomoserine lactone (C8-HSL and N-dodecanoylhomoserine lactone (C12-HSL molecules. This is the first report of the finding of quorum sensing of P. putida strain T2-2 isolated from the human tongue surface and their quorum sensing molecules were identified.

  11. Antimicrobial effects of selected plant essential oils on the growth of a Pseudomonas putida strain isolated from meat.

    Science.gov (United States)

    Oussalah, Mounia; Caillet, Stéphane; Saucier, Linda; Lacroix, Monique

    2006-06-01

    The inhibitory effect of 60 different essential oils was evaluated on a Pseudomonas putida strain of meat origin, associated with meat spoilage. Essential oils were tested at concentrations from 0.003 to 0.8% (wt/vol) to determine minimum inhibitory and maximal tolerated concentrations (MIC and MTC, respectively) using an agar medium culture. Of the 60 samples tested, Corydothymus capitatus essential oil was the most active showing a MIC of 0.025% and a MTC of 0.06%. Seven essential oils (Cinnamomum cassia, Origanum compactum, Origanum heracleoticum, Satureja hortensis, Satureja montana, Thymus vulgaris carvacroliferum, Thymus vulgaris thymoliferum) have shown a strong antimicrobial activity against P. putida with a MIC of 0.05% and a MTC ranging from 0.013% to 0.025%. Ten other oils (Cinnamomum verum (leaf and bark), Eugenia caryophyllus, Cymbopogon martinii var. motia, Cymbopogon nardus, Melaleuca linariifolia, Origanum majorana, Pimenta dioica, Thymus satureoides, Thymus serpyllum) showed a high antimicrobial activity showing a MIC ranging from 0.1% to 0.4%, while the remaining were less active showing a MIC⩾0.8%.

  12. Role of extracellular polymeric substances (EPS from Pseudomonas putida strain MnB1 in dissolution of natural rhodochrosite

    Directory of Open Access Journals (Sweden)

    H. Wang

    2014-05-01

    Full Text Available Microbially mediated oxidation of Mn(II to Mn oxides have been demonstrated in previous studies, however, the mechanisms of bacteria how to dissolve and oxidize using a solid Mn(II origin are poorly understood. In this study, we examined the role of extracellular polymeric substances (EPS from P. putida strain MnB1 in enhancing dissolution of natural rhodochrosite. The results showed that P. putida strain MnB1 cell can effectively dissolve and oxidize natural rhodochrosite to generate Mn oxides, and EPS were found to play an important role in increasing dissolution of natural rhodochrosite. Compared with EPS-free treatment, dissolution rate of natural rhodochrosite in the presence of bacterial EPS was significantly increased with decreasing initial pH and increasing EPS concentration, ionic strength and rhodochrosite dosage (p < 0.05. The fourier-transform infrared spectroscopy (FTIR analysis implies that the functional groups like N-H, C=O and C-H in EPS contributed to the dissolution of natural rhodochrosite. This study is helpful for understanding the mechanisms of the formation of biogenic Mn oxides using a solid Mn(II origin.

  13. Expression of recombinant Pseudomonas stutzeri di-heme cytochrome c(4) by high-cell-density fed-batch cultivation of Pseudomonas putida

    DEFF Research Database (Denmark)

    Thuesen, Marianne Hallberg; Nørgaard, Allan; Hansen, Anne Merete

    2003-01-01

    The gene of the di-heme protein cytochrome c(4) from Pseudomonas stutzeri was expressed in Pseudomonas putida. High-yield expression of the protein was achieved by high-cell-density fed-batch cultivation using an exponential glucose feeding strategy. The recombinant cytochrome c(4) protein...

  14. Potential of the TCE-degrading endophyte Pseudomonas putida W619-TCE to improve plant growth and reduce TCE phytotoxicity and evapotranspiration in poplar cuttings

    Energy Technology Data Exchange (ETDEWEB)

    Weyens, N.; van der Lelie, D.; Truyens, S.; Dupae, J.; Newman, L.; Taghavi, S.; Carleer, R.; Vangronsveld, J.

    2010-09-01

    The TCE-degrading poplar endophyte Pseudomonas putida W619-TCE was inoculated in poplar cuttings, exposed to 0, 200 and 400 mg l{sup -1} TCE, that were grown in two different experimental setups. During a short-term experiment, plants were grown hydroponically in half strength Hoagland nutrient solution and exposed to TCE for 3 days. Inoculation with P. putida W619-TCE promoted plant growth, reduced TCE phytotoxicity and reduced the amount of TCE present in the leaves. During a mid-term experiment, plants were grown in potting soil and exposed to TCE for 3 weeks. Here, inoculation with P. putida W619-TCE had a less pronounced positive effect on plant growth and TCE phytotoxicity, but resulted in strongly reduced amounts of TCE in leaves and roots of plants exposed to 400 mg l{sup -1} TCE, accompanied by a lowered evapotranspiration of TCE. Dichloroacetic acid (DCAA) and trichloroacetic acid (TCAA), which are known intermediates of TCE degradation, were not detected. The endophyte P. putida W619-TCE degrades TCE during its transport through the xylem, leading to reduced TCE concentrations in poplar, and decreased TCE evapotranspiration.

  15. Characterization of a novel blaIMP gene, blaIMP-58, using whole genome sequencing in a Pseudomonas putida isolate detected in Denmark

    DEFF Research Database (Denmark)

    Holmgaard, Dennis Back; Hansen, Frank; Hasman, Henrik

    2017-01-01

    A multidrug-resistant strain of Pseudomonas putida was isolated from the urine of a 65-year-old women hospitalized for serious clinical conditions. Using whole genome sequencing a novel blaIMP gene, blaIMP-58 was discovered and characterized....

  16. Involvement of specialized DNA polymerases Pol II, Pol IV and DnaE2 in DNA replication in the absence of Pol I in Pseudomonas putida

    Energy Technology Data Exchange (ETDEWEB)

    Sidorenko, Julia; Jatsenko, Tatjana; Saumaa, Signe; Teras, Riho; Tark-Dame, Mariliis; Horak, Rita [Department of Genetics, Institute of Molecular and Cell Biology, Tartu University and Estonian Biocentre, 23 Riia Street, 51010 Tartu (Estonia); Kivisaar, Maia, E-mail: maiak@ebc.ee [Department of Genetics, Institute of Molecular and Cell Biology, Tartu University and Estonian Biocentre, 23 Riia Street, 51010 Tartu (Estonia)

    2011-09-01

    The majority of bacteria possess a different set of specialized DNA polymerases than those identified in the most common model organism Escherichia coli. Here, we have studied the ability of specialized DNA polymerases to substitute Pol I in DNA replication in Pseudomonas putida. Our results revealed that P. putida Pol I-deficient cells have severe growth defects in LB medium, which is accompanied by filamentous cell morphology. However, growth of Pol I-deficient bacteria on solid rich medium can be restored by reduction of reactive oxygen species in cells. Also, mutants with improved growth emerge rapidly. Similarly to the initial Pol I-deficient P. putida, its adapted derivatives express a moderate mutator phenotype, which indicates that DNA replication carried out in the absence of Pol I is erroneous both in the original Pol I-deficient bacteria and the adapted derivatives. Analysis of the spectra of spontaneous Rif{sup r} mutations in P. putida strains lacking different DNA polymerases revealed that the presence of specialized DNA polymerases Pol II and Pol IV influences the frequency of certain base substitutions in Pol I-proficient and Pol I-deficient backgrounds in opposite ways. Involvement of another specialized DNA polymerase DnaE2 in DNA replication in Pol I-deficient bacteria is stimulated by UV irradiation of bacteria, implying that DnaE2-provided translesion synthesis partially substitutes the absence of Pol I in cells containing heavily damaged DNA.

  17. Multivariate analysis of microarray data by principal component discriminant analysis: Prioritizing relevant transcripts linked to the degradation of different carbohydrates in Pseudomonas putida S12

    NARCIS (Netherlands)

    Werf, M.J. van der; Pieterse, B.; Luijk, N. van; Schuren, F.; Werff van der - Vat, B. van der; Overkamp, K.; Jellema, R.H.

    2006-01-01

    The value of the multivariate data analysis tools principal component analysis (PCA) and principal component discriminant analysis (PCDA) for prioritizing leads generated by microarrays was evaluated. To this end, Pseudomonas putida S12 was grown in independent triplicate fermentations on four

  18. The regulatory logic of m-xylene biodegradation by Pseudomonas putida mt-2 exposed by dynamic modelling of the principal node Ps/Pr of the TOL plasmid

    NARCIS (Netherlands)

    Koutinas, M.; Lam, M.C.; Kiparissides, A.; Silva-Rocha, R.; Godinho, M.; Livingston, A.G.; Pistikopoulos, E.N.; Lorenzo, de V.; Martins Dos Santos, V.A.P.; Mantalaris, A.

    2010-01-01

    P>The structure of the extant transcriptional control network of the TOL plasmid pWW0 born by Pseudomonas putida mt-2 for biodegradation of m-xylene is far more complex than one would consider necessary from a mere engineering point of view. In order to penetrate the underlying logic of such a ne

  19. Conversion of 3-chlorocatechol by various catechol 2,3-dioxygenases and sequence analysis of the chlorocatechol dioxygenase region of Pseudomonas putida GJ31

    NARCIS (Netherlands)

    Mars, Astrid E.; Kingma, Jaap; Kaschabek, Stefan R.; Reineke, Walter; Janssen, Dick B.

    1999-01-01

    Pseudomonas putida GJ31 contains an unusual catechol 2,3-dioxygenase that converts 3-chlorocatechol and 3-methylcatechol, which enables the organism to use both chloroaromatics and methylaromatics for growth, A 3.1-kb region of genomic DNA of strain GJ31 containing the gene for this chlorocatechol 2

  20. Multivariate analysis of microarray data by principal component discriminant analysis: Prioritizing relevant transcripts linked to the degradation of different carbohydrates in Pseudomonas putida S12

    NARCIS (Netherlands)

    Werf, M.J. van der; Pieterse, B.; Luijk, N. van; Schuren, F.; Werff van der - Vat, B. van der; Overkamp, K.; Jellema, R.H.

    2006-01-01

    The value of the multivariate data analysis tools principal component analysis (PCA) and principal component discriminant analysis (PCDA) for prioritizing leads generated by microarrays was evaluated. To this end, Pseudomonas putida S12 was grown in independent triplicate fermentations on four diffe

  1. Field release of genetically modified Pseudomonas putida WCS358r : molecular analysis of effects on microbial communities in the rhizosphere of wheat

    NARCIS (Netherlands)

    Viebahn, Mareike

    2005-01-01

    Genetically modified microorganisms (GMMs) with enhanced biocontrol activity are attractive to apply in agriculture. To investigate potential ecological risks of field introduction of GMMs, effects of P. putida strain WCS358r and two genetically modified derivatives of this strain on the indigenous

  2. Optimization of the solvent-tolerant Pseudomonas putida S12 as host for the production of p-coumarate from glucose

    NARCIS (Netherlands)

    Nijkamp, K.; Westerhof, R.G.M.; Ballerstedt, H.; Bont, J.A.M.de; Wery, J.

    2007-01-01

    A Pseudomonas putida S12 strain was constructed that is able to convert glucose to p-coumarate via the central metabolite l-tyrosine. Efficient production was hampered by product degradation, limited cellular L-tyrosine availability, and formation of the by-product cinnamate via L-phenylalanine. The

  3. Draft genome sequence analysis of a Pseudomonas putida W15Oct28 strain with antagonistic activity to Gram-positive and Pseudomonas sp. pathogens.

    Science.gov (United States)

    Ye, Lumeng; Hildebrand, Falk; Dingemans, Jozef; Ballet, Steven; Laus, George; Matthijs, Sandra; Berendsen, Roeland; Cornelis, Pierre

    2014-01-01

    Pseudomonas putida is a member of the fluorescent pseudomonads known to produce the yellow-green fluorescent pyoverdine siderophore. P. putida W15Oct28, isolated from a stream in Brussels, was found to produce compound(s) with antimicrobial activity against the opportunistic pathogens Staphylococcus aureus, Pseudomonas aeruginosa, and the plant pathogen Pseudomonas syringae, an unusual characteristic for P. putida. The active compound production only occurred in media with low iron content and without organic nitrogen sources. Transposon mutants which lost their antimicrobial activity had the majority of insertions in genes involved in the biosynthesis of pyoverdine, although purified pyoverdine was not responsible for the antagonism. Separation of compounds present in culture supernatants revealed the presence of two fractions containing highly hydrophobic molecules active against P. aeruginosa. Analysis of the draft genome confirmed the presence of putisolvin biosynthesis genes and the corresponding lipopeptides were found to contribute to the antimicrobial activity. One cluster of ten genes was detected, comprising a NAD-dependent epimerase, an acetylornithine aminotransferase, an acyl CoA dehydrogenase, a short chain dehydrogenase, a fatty acid desaturase and three genes for a RND efflux pump. P. putida W15Oct28 genome also contains 56 genes encoding TonB-dependent receptors, conferring a high capacity to utilize pyoverdines from other pseudomonads. One unique feature of W15Oct28 is also the presence of different secretion systems including a full set of genes for type IV secretion, and several genes for type VI secretion and their VgrG effectors.

  4. Antimicrobial effect of Al2O3, Ag and Al2O3/Ag thin films on Escherichia coli and Pseudomonas putida

    Science.gov (United States)

    Angelov, O.; Stoyanova, D.; Ivanova, I.; Todorova, S.

    2016-10-01

    The influence of Al2O3, Ag and Al2O3/Ag thin films on bacterial growth of Gramnegative bacteria Pseudomonas putida and Escherichia coli is studied. The nanostructured thin films are deposited on glass substrates without intentional heating through r.f. magnetron sputtering in Ar atmosphere of Al2O3 and Ag targets or through sequential sputtering of Al2O3 and Ag targets, respectively. The individual Ag thin films (thickness 8 nm) have a weak bacteriostatic effect on Escherichia coli expressed as an extended adaptive phase of the bacteria up to 5 hours from the beginning of the experiment, but the final effect is only 10 times lower bacterial density than in the control. The individual Al2O3 film (20 nm) has no antibacterial effect against two strains E. coli - industrial and pathogenic. The Al2O3/Ag bilayer films (Al2O3 20 nm/Ag 8 nm) have strong bactericidal effect on Pseudomonas putida and demonstrate an effective time of disinfection for 2 hours. The individual films Al2O3 and Ag have not pronounced antibacterial effect on Pseudomonas putida. A synergistic effect of Al2O3/Ag bilayer films in formation of oxidative species on the surface in contact with the bacterial suspension could be a reason for their antimicrobial effect on E. coli and P. putida.

  5. Organo-mineral interactions in Pseudomonas putida-birnessite assemblages: Impact on mineral reactivity

    Science.gov (United States)

    Simanova, Anna; Kroll, Alexandra; Pena, Jasquelin

    2016-04-01

    The ability of microorganisms to precipitate biogenic birnessite nanoparticles is widely spread in the bacterial and fungal trees of life, with this process accounting largely for the formation of birnessite in nature. Birnessite minerals occur typically as nanoparticles that exhibit significant chemical and structural disorder. Furthermore, the mineral is embedded within a biomass matrix composed of microbial cells and extracellular polymeric substances, where the biomass not only provides reactive surfaces but can mediate electron transfer reactions. The overarching question guiding our research is: How do nanoscale properties and admixing with microbial biomass modify the reactivity of Mn oxide minerals? In this study, we investigate the biomass-birnessite composites of Pseudomonas putida GB-1 biomass and δ-MnO2 nanoparticles. We characterized the structure and composition of the mineral fraction using X-ray diffraction, Mn K-edge X-ray absorption spectroscopy and wet-chemical methods. To characterize the biomass fraction, we employed FTIR spectroscopy and size-exclusion chromatography analysis of the extracellular polymeric substances. Finally, we measured Ni(II) sorption isotherms at pH 6 and Ni K-edge EXAFS spectra to determine the extent and mechanism of Ni sorption in the biomass-mineral composites and in biomass-only and mineral-only systems. This approach provided direct and indirect evidence for the extent of organo-mineral interactions in the composites, as well as a direct measure of sorption reactivity in the composites relative to biomass-only and mineral-only systems. We found that admixing of mineral nanoparticles with biomass reduced the reactivity of the edge sites of birnessite particles towards Ni(II) through the attachment of organic moieties to the mineral particles and/or modification of the assemblage surface charge properties. In addition, the interaction of biomass components with MnO2 particles leads to partial Mn(IV) reduction and

  6. Study on the efficiency of the two phase partitioning stirred tank bioreactor on the toluene filtration from the airstream by Pseudomonas putida via

    Directory of Open Access Journals (Sweden)

    2013-02-01

    Full Text Available Introduction: There are different methods for controlling gaseous pollutants formed from air pollution sources that one of the most economical and efficient of them, is bio-filtration. The purpose of this study is Toluene removal from airstream by using the pure Pseudomonas putida bacteria as a fluidized bed in a two phase partitioning stirred tank bioreactor.Toluene ( Metyle benzene is one of the aromatic compounds which uses as a chemical solvent.low to moderate concentration of Toluene causes fatigue, dizziness, weakness,unbalance behaviour, memory loss, insomnia, loss of appetite, loss of vision and hearing. .Material and Method: In this experimental study at first, pure Pseudomonas putida in an aqueous phase containing nutrients and trace elements solution was duplicated and accustomed with Toluene. then solution contained microorganisms with 10% silicon oil was entered to bioreactor. The amount of CO2 and pollutant concentrations in the entrance and exhaust of bioreactor containing Pseudomonas putida was studied during 17 days for each variable. .Result: Experimental findings showed that in the 0.06 m3/h and 0.12 m3/h flow rate, the efficiency of bioreactor containing Pseudomonas putida in the concentration ranges of 283 Mg/m3 to 4710 Mg/m3 was at least 97% and 25% respectively. Statistical analysis (ANOVA showed that in two flow rates of 0.06 m3/h and 0.12 m3/h removal efficiency and mineralization percentage had significant differences .(Pvalue =0.01. .Conclusion: Achieving high efficiencies in pollutants removal was because of the prepared optimum conditions for Pseudomonas putida in the two phase partitioning stirred tank bioreactor with 10% organic phase.

  7. A facile approach for surface alteration of Pseudomonas putida I3 by supplying K2SO4 into growth medium: Enhanced removal of Pb(II) from aqueous solution.

    Science.gov (United States)

    Xu, Xingjian; Li, Haiyan; Wang, Quanying; Li, Dandan; Han, Xuerong; Yu, Hongwen

    2017-02-12

    A new sight of obtaining a high efficient biosorbent by supplying specific salts into bacterial growth medium was investigated in this study for Pb(II). Among a series of salts including Na2SO4, Na2S2O3, KCl, and K2SO4, the highest Pb(II) removal efficiency was observed by psychrotrophilic Pseudomonas putida I3 grown in the presence of 30g/L K2SO4 (KSI3-30) with biosorption capacity of 62.89mg/g under cold condition (15°C), which was increased by 42.35% as compared to control (without any additive, RI3). This stimulation effect was ascribed to the increase of potassium and sulfur containing groups on KSI3-30 surface via metabolic dependent ways. The probable mechanism for Pb(II) adsorption was ion-exchange and chemical complexation. The thermal and kinetic data well fitted to Langmuir adsorption model and pseudo-second order and intraparticle diffusion kinetic model. Good recyclability and effectively dealing with real wastewater suggested KSI3-30 was a promising biosorbent for Pb-contaminated wastewater treatment.

  8. Influence of siderophore pyoverdine synthesis and iron-uptake on abiotic and biotic surface colonization of Pseudomonas putida S11.

    Science.gov (United States)

    Ponraj, Paramasivan; Shankar, Manoharan; Ilakkiam, Devaraj; Gunasekaran, Paramasamy

    2012-12-01

    Fluorescent pseudomonads produce a characteristic fluorescent pigment, pyoverdines as their primary siderophore for iron acquisition under iron-limiting conditions. Here, we report the identification of a random transposon mutant IST3 of Pseudomonas putida S11 showing tolerance to iron starvation stress condition and increased pyoverdine production. The insertion of the Tn5 transposon was found to be in pstS gene of pstSR operon encoding sensor histidine kinase protein of the two-component signal transduction system. A pyoverdine negative derivative of IST3 mutant constructed was sensitive to iron stress condition. It indicated that increased survival of IST3 under iron-limiting condition was due to higher pyoverdine production. The iron starvation tolerant mutant (IST3) exhibited enhanced pyoverdine-mediated iron uptake in minimal medium which significantly improved its biofilm formation, seed adhesion and competitive root colonization.

  9. Estudo da capacidade de sorção de cobre por Pseudomonas putida sp. em reator.

    OpenAIRE

    2014-01-01

    Bactérias aclimatadas a cobre foram isoladas a partir de amostras de solo e água coletadas na região da Mina de Sossego (Vale, Carajás, PA). Pseudomonas putida sp. foi escolhida, pois, apresentou a maior capacidade sortiva de Cu2+, Q = 40 mg/g. O cultivo em regime de batelada, meio mineral, com glicerol como fonte de carbono, resultou fator de conversão de glicerol a células, YX/S, de 0,49 g/g e velocidade específica máxima de crescimento, mmax, de 0,11 h-1. Alta concentração celular, 90 g/L,...

  10. Production of Medium Chain Length Polyhydroxyalkanoates From Oleic Acid Using Pseudomonas putida PGA1 by Fed Batch Culture

    Directory of Open Access Journals (Sweden)

    Sidik Marsudi

    2010-10-01

    Full Text Available Bacterial polyhydroxyalkanoates (PHAs are a class of p0lymers currently receiving much attention because of their potential as renewable and biodegradable plastics. A wide variety of bacteria has been reported to produce PHAs including Pseudomonas strains. These strains are known as versatile medium chain length PHAs (PHAs-mcl producers using fatty acids as carbon source. Oleic acid was used to produce PHAs-mcl using Pseudomonas putida PGA 1 by continuous feeding of both nitrogen and carbon source, in a fed batch culture. During cell growth, PHAs also accumulated, indicating that PHA production in this organism is growth associated. Residual cell increased until the nitrogen source was depleted. At the end of fermentation, final cell concentration, PHA content, and roductivity were 30.2 g/L, 44.8 % of cell dry weight, and 0.188 g/l/h, respectively.

  11. Mechanisms of trace metal sorption in Pseudomonas putida-birnessite assemblages

    Science.gov (United States)

    Peña, J.; Kwon, K. D.; Bargar, J. R.; Sposito, G.

    2012-04-01

    Biogenic manganese oxides (MnO2) are ubiquitous nanoparticulate minerals that contribute strongly to the adsorption of nutrient and toxicant metals in aquatic and terrestrial environments. The formation of these minerals is catalyzed by a diverse and widely-distributed group of bacteria and fungi, often through the enzymatic oxidation of aqueous Mn(II) to Mn(IV). The biogenic Mn(IV) oxide found in field settings, as well as that produced by model bacteria in laboratory culture, is typically layer-type hexagonal birnessite containing abundant cation vacancy sites and enmeshed in an organic matrix of bacterial cells and extracellular polymeric substances. In this talk I summarize the results from laboratory-scale research designed to investigate the mechanisms of metal sorption by the bacterial biomass-birnessite assemblages formed by Pseudomonas putida GB-1 when grown in the presence of 1 mM Mn(II) at circumneutral pH values. The goals of this research were first, to identify the structure of the surface complexes formed by trace metals (e.g., Ni, Cu and Zn) on biogenic birnessite and second, to determine the conditions under which the bacterial cell surfaces and extracellular polymeric substances contribute to metal sorption. Macroscopic and spectroscopic experiments were performed at varying pH values (6 - 8) and over a wide-range of metal concentrations. Extended X-ray absorption fine structure (EXAFS) spectroscopy and first-principles calculations based on density functional theory showed that cation vacancy sites in birnessite drive mineral reactivity, but that surface speciation varies from metal to metal. For, Ni we identified two species, Ni bonded to three surface oxygen atoms vacancy sites as a triple-corner-sharing (TCS) complex and Ni incorporated at vacancy sites, with surface speciation varying with pH and surface loading. Zinc formed TCS complexes at vacancy sites, with the proportion of Zn in tetrahedral or octahedral coordination geometry influenced

  12. Metabolic acidosis

    Science.gov (United States)

    Acidosis - metabolic ... Metabolic acidosis occurs when the body produces too much acid. It can also occur when the kidneys are not ... the body. There are several types of metabolic acidosis. Diabetic acidosis develops when acidic substances, known as ...

  13. Metabolic neuropathies

    Science.gov (United States)

    Neuropathy - metabolic ... can be caused by many different things. Metabolic neuropathy may be caused by: A problem with the ... one of the most common causes of metabolic neuropathies. People who are at the highest risk for ...

  14. Biosensor for Direct Determination of Fenitrothion and EPN Using Recombinant Pseudomonas putida JS444 with Surface Expressed Organophosphorus Hydrolase. 1. Modified Clark Oxygen Electrode

    Directory of Open Access Journals (Sweden)

    Ashok Mulchandani

    2006-04-01

    Full Text Available This paper reports a first microbial biosensor for rapid and cost-effectivedetermination of organophosphorus pesticides fenitrothion and EPN. The biosensorconsisted of recombinant PNP-degrading/oxidizing bacteria Pseudomonas putida JS444anchoring and displaying organophosphorus hydrolase (OPH on its cell surface asbiological sensing element and a dissolved oxygen electrode as the transducer. Surface-expressed OPH catalyzed the hydrolysis of fenitrothion and EPN to release 3-methyl-4-nitrophenol and p-nitrophenol, respectively, which were oxidized by the enzymaticmachinery of Pseudomonas putida JS444 to carbon dioxide while consuming oxygen,which was measured and correlated to the concentration of organophosphates. Under theoptimum operating conditions, the biosensor was able to measure as low as 277 ppb offenitrothion and 1.6 ppm of EPN without interference from phenolic compounds and othercommonly used pesticides such as carbamate pesticides, triazine herbicides andorganophosphate pesticides without nitrophenyl substituent. The applicability of thebiosensor to lake water was also demonstrated.

  15. Biosensor for Direct Determination of Fenitrothion and EPN Using Recombinant Pseudomonas putida JS444 with Surface Expressed Organophosphorus Hydrolase. 1. Modified Clark Oxygen Electrode

    OpenAIRE

    2006-01-01

    This paper reports a first microbial biosensor for rapid and cost-effective determination of organophosphorus pesticides fenitrothion and EPN. The biosensor consisted of recombinant PNP-degrading/oxidizing bacteria Pseudomonas putida JS444 anchoring and displaying organophosphorus hydrolase (OPH) on its cell surface as biological sensing element and a dissolved oxygen electrode as the transducer. Surface-expressed OPH catalyzed the hydrolysis of fenitrothion and EPN to release 3-methyl-4-nitr...

  16. A newly isolated Pseudomonas putida S-1 strain for batch-mode-propanethiol degradation and continuous treatment of propanethiol-containing waste gas

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Dong-Zhi, E-mail: cdz@zjut.edu.cn [College of Biological and Environmental Engineering, Zhejiang University of Technology, Hangzhou 310032 (China); Sun, Yi-Ming; Han, Li-Mei [College of Biological and Environmental Engineering, Zhejiang University of Technology, Hangzhou 310032 (China); Chen, Jing [College of Food and Pharmacy, Zhejiang Ocean University, Zhoushan 316004 (China); Ye, Jie-Xu; Chen, Jian-Meng [College of Biological and Environmental Engineering, Zhejiang University of Technology, Hangzhou 310032 (China)

    2016-01-25

    Highlights: • A novel strain capable of effectively degrading 1-propanethiol (PT) was isolated. • Cells could be feasibly cultured in nutrition-rich media for PT degradation. • A possible pathway for PT degradation was proposed. • Pseudomonas putida S-1 could degrade mixed pollutants with diauxic growth. • Continuous removal of gaseous PT with or without isopropanol was demonstrated. - Abstract: Pseudomonas putida S-1 was isolated from activated sludge. This novel strain was capable of degrading malodorous 1-propanethiol (PT). PT degradation commenced with no lag phase by cells pre-grown in nutrition-rich media, such as Luria–Bertani (LB), and PT-contained mineral medium at specific growth rates of 0.10–0.19 h{sup −1}; this phenomenon indicated the operability of a large-scale cell culture. A possible PT degradation pathway was proposed on the basis of the detected metabolites, including dipropyl disulfide, 3-hexanone, 2-hexanone, 3-hexanol, 2-hexanol, S{sup 0}, SO{sub 4}{sup 2−}, and CO{sub 2}. P. putida S-1 could degrade mixed pollutants containing PT, diethyl disulfide, isopropyl alcohol, and acetaldehyde, and LB-pre-cultured cells underwent diauxic growth. Waste gas contaminated with 200–400 mg/m{sup 3} PT was continuously treated by P. putida S-1 pre-cultured in LB medium in a completely stirred tank reactor. The removal efficiencies exceeded 88% when PT stream was mixed with 200 mg/m{sup 3} isopropanol; by contrast, the removal efficiencies decreased to 60% as the empty bed residence time was shortened from 40 s to 20 s.

  17. Co-culture with Listeria monocytogenes within a dual-species biofilm community strongly increases resistance of Pseudomonas putida to benzalkonium chloride.

    Directory of Open Access Journals (Sweden)

    Efstathios Giaouris

    Full Text Available Biofilm formation is a phenomenon occurring almost wherever microorganisms and surfaces exist in close proximity. This study aimed to evaluate the possible influence of bacterial interactions on the ability of Listeria monocytogenes and Pseudomonas putida to develop a dual-species biofilm community on stainless steel (SS, as well as on the subsequent resistance of their sessile cells to benzalkonium chloride (BC used in inadequate (sub-lethal concentration (50 ppm. The possible progressive adaptability of mixed-culture biofilms to BC was also investigated. To accomplish these, 3 strains per species were left to develop mixed-culture biofilms on SS coupons, incubated in daily renewable growth medium for a total period of 10 days, under either mono- or dual-species conditions. Each day, biofilm cells were exposed to disinfection treatment. Results revealed that the simultaneous presence of L. monocytogenes strongly increased the resistance of P. putida biofilm cells to BC, while culture conditions (mono-/dual-species did not seem to significantly influence the resistance of L. monocytogenes biofilm cells. BC mainly killed L. monocytogenes cells when this was applied against the dual-species sessile community during the whole incubation period, despite the fact that from the 2nd day this community was mainly composed (>90% of P. putida cells. No obvious adaptation to BC was observed in either L. monocytogenes or P. putida biofilm cells. Pulsed field gel electrophoresis (PFGE analysis showed that the different strains behaved differently with regard to biofilm formation and antimicrobial resistance. Such knowledge on the physiological behavior of mixed-culture biofilms could provide the information necessary to control their formation.

  18. Co-culture with Listeria monocytogenes within a dual-species biofilm community strongly increases resistance of Pseudomonas putida to benzalkonium chloride.

    Science.gov (United States)

    Giaouris, Efstathios; Chorianopoulos, Nikos; Doulgeraki, Agapi; Nychas, George-John

    2013-01-01

    Biofilm formation is a phenomenon occurring almost wherever microorganisms and surfaces exist in close proximity. This study aimed to evaluate the possible influence of bacterial interactions on the ability of Listeria monocytogenes and Pseudomonas putida to develop a dual-species biofilm community on stainless steel (SS), as well as on the subsequent resistance of their sessile cells to benzalkonium chloride (BC) used in inadequate (sub-lethal) concentration (50 ppm). The possible progressive adaptability of mixed-culture biofilms to BC was also investigated. To accomplish these, 3 strains per species were left to develop mixed-culture biofilms on SS coupons, incubated in daily renewable growth medium for a total period of 10 days, under either mono- or dual-species conditions. Each day, biofilm cells were exposed to disinfection treatment. Results revealed that the simultaneous presence of L. monocytogenes strongly increased the resistance of P. putida biofilm cells to BC, while culture conditions (mono-/dual-species) did not seem to significantly influence the resistance of L. monocytogenes biofilm cells. BC mainly killed L. monocytogenes cells when this was applied against the dual-species sessile community during the whole incubation period, despite the fact that from the 2nd day this community was mainly composed (>90%) of P. putida cells. No obvious adaptation to BC was observed in either L. monocytogenes or P. putida biofilm cells. Pulsed field gel electrophoresis (PFGE) analysis showed that the different strains behaved differently with regard to biofilm formation and antimicrobial resistance. Such knowledge on the physiological behavior of mixed-culture biofilms could provide the information necessary to control their formation.

  19. Draft genome sequence analysis of a Pseudomonas putida W15Oct28 strain with antagonistic activity to Gram-positive and Pseudomonas sp. pathogens.

    Directory of Open Access Journals (Sweden)

    Lumeng Ye

    Full Text Available Pseudomonas putida is a member of the fluorescent pseudomonads known to produce the yellow-green fluorescent pyoverdine siderophore. P. putida W15Oct28, isolated from a stream in Brussels, was found to produce compound(s with antimicrobial activity against the opportunistic pathogens Staphylococcus aureus, Pseudomonas aeruginosa, and the plant pathogen Pseudomonas syringae, an unusual characteristic for P. putida. The active compound production only occurred in media with low iron content and without organic nitrogen sources. Transposon mutants which lost their antimicrobial activity had the majority of insertions in genes involved in the biosynthesis of pyoverdine, although purified pyoverdine was not responsible for the antagonism. Separation of compounds present in culture supernatants revealed the presence of two fractions containing highly hydrophobic molecules active against P. aeruginosa. Analysis of the draft genome confirmed the presence of putisolvin biosynthesis genes and the corresponding lipopeptides were found to contribute to the antimicrobial activity. One cluster of ten genes was detected, comprising a NAD-dependent epimerase, an acetylornithine aminotransferase, an acyl CoA dehydrogenase, a short chain dehydrogenase, a fatty acid desaturase and three genes for a RND efflux pump. P. putida W15Oct28 genome also contains 56 genes encoding TonB-dependent receptors, conferring a high capacity to utilize pyoverdines from other pseudomonads. One unique feature of W15Oct28 is also the presence of different secretion systems including a full set of genes for type IV secretion, and several genes for type VI secretion and their VgrG effectors.

  20. Development of a visual loop-mediated isothermal amplification method for rapid detection of the bacterial pathogen Pseudomonas putida of the large yellow croaker (Pseudosciaena crocea).

    Science.gov (United States)

    Mao, Zhijuan; Qiu, Yangyu; Zheng, Lei; Chen, Jigang; Yang, Jifang

    2012-06-01

    In recent years, the large yellow croaker (Pseudosciaena crocea), an important marine fish farmed in the coastal areas of Zhejiang province, east China, has become severely endangered as a result of the bacterial pathogen Pseudomonas putida. This paper reports the development of a visual loop-mediated isothermal amplification (LAMP) assay for rapid detection of the pathogen. Four primers, F3, B3, FIP and BIP, were designed on the basis of DNA sequence of the rpoN gene of P. putida. After optimization of the reaction conditions, the detection limit of LAMP assay was 4.8cfu per reaction, 10-fold higher than that of conventional PCR. The assay showed high specificity to discriminate all P. putida isolates from nine other Gram-negative bacteria. The assay also successfully detected the pathogen DNA in the tissues of infected fish. For visual LAMP without cross-contamination, SYBR Green I was embedded in a microcrystalline wax capsule and preset in the reaction tubes; after the reaction the wax was melted at 85°C to release the dye and allow intercalation with the amplicons. The simple, highly sensitive, highly specific and cost-effective characteristics of visual LAMP may encourage its application in the rapid diagnosis of this pathogen.

  1. Potential of the TCE-degrading endophyte Pseudomonas putida W619-TCE to improve plant growth and reduce TCE phytotoxicity and evapotranspiration in poplar cuttings.

    Science.gov (United States)

    Weyens, Nele; Truyens, Sascha; Dupae, Joke; Newman, Lee; Taghavi, Safiyh; van der Lelie, Daniel; Carleer, Robert; Vangronsveld, Jaco

    2010-09-01

    The TCE-degrading poplar endophyte Pseudomonas putida W619-TCE was inoculated in poplar cuttings, exposed to 0, 200 and 400 mg l(-1) TCE, that were grown in two different experimental setups. During a short-term experiment, plants were grown hydroponically in half strength Hoagland nutrient solution and exposed to TCE for 3 days. Inoculation with P. putida W619-TCE promoted plant growth, reduced TCE phytotoxicity and reduced the amount of TCE present in the leaves. During a mid-term experiment, plants were grown in potting soil and exposed to TCE for 3 weeks. Here, inoculation with P. putida W619-TCE had a less pronounced positive effect on plant growth and TCE phytotoxicity, but resulted in strongly reduced amounts of TCE in leaves and roots of plants exposed to 400 mg l(-1) TCE, accompanied by a lowered evapotranspiration of TCE. Dichloroacetic acid (DCAA) and trichloroacetic acid (TCAA), which are known intermediates of TCE degradation, were not detected.

  2. Cloning, nucleotide sequence, and overexpression of the gene coding for delta 5-3-ketosteroid isomerase from Pseudomonas putida biotype B.

    Science.gov (United States)

    Kim, S W; Kim, C Y; Benisek, W F; Choi, K Y

    1994-11-01

    The structural gene coding for the delta 5-3-ketosteroid isomerase (KSI) of Pseudomonas putida biotype B has been cloned, and its entire nucleotide sequence has been determined by a dideoxynucleotide chain termination method. A 2.1-kb DNA fragment containing the ksi gene was cloned from a P. putida biotype B genomic library in lambda gt11. The open reading frame of ksi encodes 393 nucleotides, and the amino acid sequence deduced from the nucleotide sequence agrees with the directly determined amino acid sequence (K. Linden and W. F. Benisek, J. Biol. Chem. 261:6454-6460, 1986). A putative purine-rich ribosome binding site was found 8 bp upstream of the ATG start codon. Escherichia coli BL21(DE3) transformed with the pKK-KSI plasmid containing the ksi gene expressed a high level of isomerase activity when induced by isopropyl-beta-D-thiogalactopyranoside. KSI was purified to homogeneity by a simple and rapid procedure utilizing fractional precipitation and an affinity column of deoxycholate-ethylenediamine-agarose as a major chromatographic step. The molecular weight of KSI was 14,535 (calculated, 14,536) as determined by electrospray mass spectrometry. The purified KSI showed a specific activity (39,807 mumol min-1 mg-1) and a Km (60 microM) which are close to those of KSI originally obtained from P. putida biotype B.

  3. Pseudomonas putida MnB1氧化分解菱锰矿的实验研究%Experimental Study on Microbial Oxiadation of Rhodochrosite by Pseudomonas putida MnB1

    Institute of Scientific and Technical Information of China (English)

    陈笑夜; 陆现彩; 李娟; 刘欢; 向婉丽; 张蕊

    2015-01-01

    Rhodochrosite (MnCO3) is a solid Mn(Ⅱ) origin mineral and its oxidation into Mn oxides is a very common phenomenon. Although microbially mediated oxidation of Mn(II) to Mn oxides have been demonstrated in previous studies, the mechanisms of bacteria how to dissolve and oxidize using a solid Mn(II) origin are poorly understood. In this study, we examined the role of Pseudomonas putida MnB1 cell in enhancing dissolution and oxidation of rhodochrosite from Wuzhou, Guangxi province. By using Field Emission Scanning Electron Microscopy (FESEM), Scanning Transmission X-ray Microscopy imaging analysis (STXM) as well as other techniques, the changes of the mineral surfaces and the distribution of manganese in cells were identified. The result show that bacteria significantly promote the dissolution of the rhodochrosite. Moreover, the contribution of bacteria to the oxidation of rhodochrosite was discussed. The results will enrich the research of secondary manganese deposits.%自然界中,菱锰矿氧化形成锰的氧化物矿物是非常普遍的现象,在菱锰矿被氧化分解发生物相转变的过程中,碳酸盐溶解和锰的氧化往往同时发生,微生物可能起着催化作用。选取锰氧化模式菌株Pseudomonas putida MnB1和广西梧州菱锰矿,通过菱锰矿在该细菌作用下发生转变的实验,利用场发射扫描电镜、扫描透射X射线显微成像等分析方法,研究了矿石表面形貌变化以及锰元素在细胞上的分布特征。结果表明细菌显著促进的菱锰矿的溶解,在此基础上,进一步探讨了细菌在菱锰矿氧化过程中的贡献,本实验结果丰富了次生锰矿床的微生物成因研究。

  4. Overexpression and characterization of medium-chain-length polyhydroxyalkanoate granule bound polymerases from Pseudomonas putida GPo1

    Directory of Open Access Journals (Sweden)

    Ren Qun

    2009-11-01

    Full Text Available Abstract Background Polyhydroxyalkanoates (PHA are synthesized by many bacteria in the cytoplasm as storage compounds for energy and carbon. The key enzymes for PHA biosynthesis are PHA polymerases, which catalyze the covalent linkage of 3-hydroxyacyl coenzymeA thioesters by transesterification with concomitant release of CoA. Pseudomonas putida GPo1 and many other Pseudomonas species contain two different class II polymerases, encoded by phaC1 and phaC2. Although numerous studies have been carried out on PHA polymerases and they are well characterized at the molecular level, the biochemical properties of the class II polymerases have not been studied in detail. Previously we and other groups purified the polymerases, however, the activities of the purified enzymes were several magnitude lower than the granule-bound enzymes. It is problematic to study the intrinsic properties of these enzymes with such low activities, although they are pure. Results PHA polymerase 1 (PhaC1 and PHA polymerase 2 (PhaC2 from P. putida GPo1 were overexpressed in the PHA-negative host P. putida GPp104 and purified from isolated PHA granules. Only minor activity (two to three orders of magnitude lower than that of the granule bound proteins could be recovered when the enzymes were purified to homogeneity. Therefore, kinetic properties and substrate ranges were determined for the granule bound polymerases. The polymerases differed significantly with respect to their association with PHA granules, enzyme kinetics and substrate specificity. PhaC2 appeared to bind PHA granules more tightly than PhaC1. When R-3-hydroxyoctanoic acid was used as substrate, the granule-bound PhaC1 exhibited a Km of 125 (± 35 μM and a Vmax of 40.8 (± 6.2 U/mg PhaC1, while a Km of 37 (± 10 μM and a Vmax of 2.7 (± 0.7 U/mg PhaC2 could be derived for the granule-bound PhaC2. Granule-bound PhaC1 showed a strong preference for medium chain length (mcl- 3-hydroxyacly-CoAs, with highest

  5. Effect of silver nanoparticles and silver ions on growth and adaptive response mechanisms of Pseudomonas putida mt-2.

    Science.gov (United States)

    Hachicho, Nancy; Hoffmann, Philipp; Ahlert, Kristin; Heipieper, Hermann J

    2014-06-01

    The distribution and use of nanoparticles increased rapidly during the last years, while the knowledge about mode of action, ecological tolerance and biodegradability of these chemicals is still insufficient. The effect of silver nanoparticles (AgNP) and free silver ions (Ag(+) , AgNO3 ) on Pseudomonas putida mt-2 as one of the best described bacterial strains for stress response were investigated. The effective concentration (EC50) causing 50% growth inhibition for AgNP was about 250 mg L(-1) , whereas this was only 0.175 mg L(-1) for AgNO3 . However, when calculating the amount of free silver ions released from AgNP both tested compounds showed very similar results. Therefore, the antibacterial activity of AgNP can be explained and reduced, respectively, to the amount of silver ions released from the nanoparticles. Both tested compounds showed a strong activation of the unique membrane adaptive response of Pseudomonas strains, the cis-trans isomerization of unsaturated fatty acids, whereas another important adaptive response of these bacteria, changes in cell surface hydrophobicity, measured as water contact angle, was not activated. These results are important informations for the estimation of environmental tolerance of newly developed, active ingredients like silver nanoparticles. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  6. U(VI) biosorption by bi-functionalized Pseudomonas putida @ chitosan bead: Modeling and optimization using RSM.

    Science.gov (United States)

    Sohbatzadeh, Hozhabr; Keshtkar, Ali Reza; Safdari, Jaber; Fatemi, Faezeh

    2016-08-01

    In this work, Pseudomonas putida cells immobilized into chitosan beads (PICB) were synthesized to investigate the impact of microorganism entrapment on biosorption capacity of prepared biosorbent for U(VI) biosorption from aqueous solutions. Response Surface Methodology (RSM) based on Central Composite Design (CCD) was utilized to evaluate the performance of the PICB in comparison with chitosan beads (CB) under batch mode. Performing experiments under optimal condition sets viz. pH 5, initial U(VI) concentration 500mg/L, biosorbent dosage 0.4g/L and 20wt.% bacterial cells showed that the observed biosorption capacity enhanced by 1.27 times from 398mg/g (CB) to 504mg/g (PICB) that confirmed the effectiveness of cells immobilization process. FTIR and potentiometric titration were then utilized to characterize the prepared biosorbents. While the dominant functional group in the binding process was NH3(+) (4.78meq/g) in the CB, the functional groups of NH3(+), NH2, OH, COOH (6.00meq/g) were responsible for the PICB. The equilibrium and kinetic studies revealed that the Langmuir isotherm model and the pseudo-second-order kinetic model were in better fitness with the CB and PICB experimental data. In conclusion, the present study indicated that the PICB could be a suitable biosorbent for uranium (VI) biosorption from aqueous solutions.

  7. The role of amino acid residues in the active site of L-methionine γ-lyase from Pseudomonas putida.

    Science.gov (United States)

    Fukumoto, Mitsuki; Kudou, Daizou; Murano, Shouko; Shiba, Tomoo; Sato, Dan; Tamura, Takashi; Harada, Shigeharu; Inagaki, Kenji

    2012-01-01

    Cys116, Lys240*, and Asp241* (asterisks indicate residues from the second subunit of the active dimer) at the active site of L-methionine γ-lyase of Pseudomonas putida (MGL_Pp) are highly conserved among heterologous MGLs. In a previous study, we found that substitution of Cys116 for His led to a drastic increase in activity toward L-cysteine and a decrease in that toward L-methionine. In this study, we examined some properties of the C116H mutant by kinetic analysis and 3D structural analysis. We assumed that substitution of Cys116 for His broke the original hydrogen-bond network and that this induced a significant effect of Tyr114 as a general acid catalyst, possibly due to the narrow space in the active site. The C116H mutant acquired a novel β-elimination activity and lead a drastic conformation change in the histidine residue at position 116 by binding the substrate, suggesting that this His residue affects the reaction specificity of C116H. Furthermore, we suggest that Lys240* is important for substrate recognition and structural stability and that Asp241* is also involved in substrate specificity in the elimination reaction. Based on this, we suggest that the hydrogen-bond network among Cys116, Lys240*, and Asp241* contributes to substrate specificity that is, to L-methionine recognition at the active site in MGL_Pp.

  8. Growth stimulation of Bacillus cereus and Pseudomonas putida using nanostructured ZnO thin film as transducer element

    Energy Technology Data Exchange (ETDEWEB)

    Loukanov, Alexandre, E-mail: loukanov@mail.saitama-u.ac.jp [Saitama University, Department of Chemistry, Faculty of Science (Japan); Filipov, Chavdar [University of Forestry, Department of Infectious pathology, hygiene, technology and control of food stuffs of animal origin, Faculty of Veterinary Medicine (Bulgaria); Valcheva, Violeta [Bulgarian Academy of Science, Department of Infectious Diseases, Institute of microbiology (Bulgaria); Lecheva, Marta [University of Mining and Geology “St. Ivan Rilski”, Laboratory of Engineering NanoBiotechnology, Department of Engineering Geoecology (Bulgaria); Emin, Saim [University of Nova Gorica, Materials Research Laboratory (Slovenia)

    2015-04-15

    The semiconductor zinc oxide nanomaterial (ZnO or ZnO:H) is widely used in advanced biosensor technology for the design of highly-sensitive detector elements for various applications. In the attempt to evaluate its effect on common microorganisms, two types of nanostructured transducer films have been used (average diameter 600–1000 nm). They have been prepared by using both wet sol–gel method and magnetron sputtering. Their polycrystalline structure and specific surface features have been analyzed by X-ray diffraction (XRD), scanning electron microscope, and atomic force microscope. The assessment of growth stimulation of bacteria was determined using epifluorescent microscope by cell staining with Live/Dead BacLight kit. In our experiments, the growth stimulation of Gram-positive and Gram-negative bacteria on nanostructured ZnO film is demonstrated by Bacillus cereus and Pseudomonas putida. These two bacterial species have been selected, because they are well known and studied in biosensor technologies, with structural difference of their cell walls. These pathogens are easy for with common source in the liquid food or some commercial products. Our data has revealed that the method of transducer film preparation influences strongly bacterial inhibition and division. These results present the transforming signal precisely, when ZnO is used in biosensor applications.

  9. Growth kinetics, effect of carbon substrate in biosynthesis of mcl-PHA by Pseudomonas putida Bet001

    Directory of Open Access Journals (Sweden)

    A.M. Gumel

    2014-06-01

    Full Text Available Growth associated biosynthesis of medium chain length poly-3-hydroxyalkanoates (mcl-PHA in Pseudomonas putida Bet001 isolated from palm oil mill effluent was studied. Models with substrate inhibition terms described well the kinetics of its growth. Selected fatty acids (C8:0 to C18:1 and ammonium were used as carbon and nitrogen sources during growth and PHA biosynthesis, resulting in PHA accumulation of about 50 to 69% (w/w and PHA yields ranging from 10.12 g L-1 to 15.45 g L-1, respectively. The monomer composition of the PHA ranges from C4 to C14, and was strongly influenced by the type of carbon substrate fed. Interestingly, an odd carbon chain length (C7 monomer was also detected when C18:1 was fed. Polymer showed melting temperature (Tm of 42.0 (± 0.2 °C, glass transition temperature (Tg of -1.0 (± 0.2 °C and endothermic melting enthalpy of fusion (ΔHf of 110.3 (± 0.1 J g-1. The molecular weight (Mw range of the polymer was relatively narrow between 55 to 77 kDa.

  10. Inhibited transport of graphene oxide nanoparticles in granular quartz sand coated with Bacillus subtilis and Pseudomonas putida biofilms.

    Science.gov (United States)

    He, Jian-Zhou; Wang, Deng-Jun; Fang, Huan; Fu, Qing-Long; Zhou, Dong-Mei

    2017-02-01

    Increasing production and use of graphene oxide nanoparticles (GONPs) boost their wide dissemination in the subsurface environments where biofilms occur ubiquitously, representative of the physical and chemical heterogeneities. This study aimed at investigating the influence of Gram-positive Bacillus subtilis (BS) and Gram-negative Pseudomonas putida (PP) biofilms on the transport of GONPs under different ionic strengths (0.1, 0.5, and 1.0 mM CaCl2) at neutral pH 7.2 in water-saturated porous media. Particularly, the X-ray micro-computed tomography was used to quantitatively characterize the pore structures of sand columns in the presence and absence of biofilms. Our results indicated that the presence of biofilms reduced the porosity and narrowed down the pore sizes of packed columns. Transport experiments in biofilm-coated sand showed that biofilms, irrespective of bacterial species, significantly inhibited the mobility of GONPs compared to that in cleaned sand. This could be due to the Ca(2+) complexation, increased surface roughness and charge heterogeneities of collectors, and particularly enhanced physical straining caused by biofilms. The two-site kinetic retention model-fitted value of maximum solid-phase concentration (Smax2) for GONPs was higher for biofilm-coated sand than for cleaned sand, demonstrating that biofilms act as favorable sites for GONPs retention. Our findings presented herein are important to deepen our current understanding on the nature of particle-collector interactions.

  11. Extracellular expression of natural cytosolic arginine deiminase from Pseudomonas putida and its application in the production of L-citrulline.

    Science.gov (United States)

    Su, Lingqia; Ma, Yue; Wu, Jing

    2015-11-01

    The Pseudomonas putida arginine deiminase (ADI), a natural cytosolic enzyme, and Thermobifida fusca cutinase were co-expressed in Escherichia coli, and the optimized cutinase gene was used for increasing its expression level. 90.9% of the total ADI protein was released into culture medium probably through a nonspecific leaking mechanism caused by the co-expressed cutinase. The enzymatic properties of the extracellular ADI were found to be similar to those of ADI prepared by conventional cytosolic expression. Extracellular production of ADI was further scaled up in a 3-L fermentor. When the protein expression was induced by IPTG (25.0μM) and lactose (0.1gL(-1)h(-1)) at 30°C, the extracellular ADI activity reached 101.2UmL(-1), which represented the highest ADI production ever reported. In addition, the enzymatic synthesis of l-citrulline was performed using the extracellularly expressed ADI, and the conversion rate reached 100% with high substrate concentration at 650gL(-1).

  12. The Repetitive Detection of Toluene with Bioluminescence Bioreporter Pseudomonas putida TVA8 Encapsulated in Silica Hydrogel on an Optical Fiber

    Directory of Open Access Journals (Sweden)

    Gabriela Kuncová

    2016-06-01

    Full Text Available Living cells of the lux-based bioluminescent bioreporter Pseudomonas putida TVA8 were encapsulated in a silica hydrogel attached to the distal wider end of a tapered quartz fiber. Bioluminescence of immobilized cells was induced with toluene at high (26.5 mg/L and low (5.3 mg/L concentrations. Initial bioluminescence maxima were achieved after >12 h. One week after immobilization, a biofilm-like layer of cells had formed on the surface of the silica gel. This resulted in shorter response times and more intensive bioluminescence maxima that appeared as rapidly as 2 h after toluene induction. Considerable second bioluminescence maxima were observed after inductions with 26.5 mg toluene/L. The second and third week after immobilization the biosensor repetitively and semiquantitatively detected toluene in buffered medium. Due to silica gel dissolution and biofilm detachment, the bioluminescent signal was decreasing 20–32 days after immobilization and completely extinguished after 32 days. The reproducible formation of a surface cell layer on the wider end of the tapered optical fiber can be translated to various whole cell bioluminescent biosensor devices and may serve as a platform for in-situ sensors.

  13. Gene Sequence, Soluble Expression and Homologous Comparison of a D-Hydantoinase from Pseudomonas putida YZ-26

    Institute of Scientific and Technical Information of China (English)

    SHI Ya-wei; ZHAO Li-xia; NIU Li-xi; FENG Xia; YUAN Jing-ming

    2005-01-01

    A 1440bp open-reading frame encoding D-hydantoinase from Pseudomonas putida YZ-26 was cloned and sequenced(GenBank AY387829). The DNA fragment was inserted into Nde and BamHI sites of vector pET-3a, yielding a recombinant plasmid, pET-HDT. After being transferred into the host strain, the artificial strain, pET-HDT/E.coli BL21 can express the D-hydantoinase as the soluble form in the Lura-Bertani medium without addition of any inducers. The activity of the enzyme toward substrate DL-hydantoin can reach 3000-4000 IU per cells from one-liter bacterial culture incubated at 30 ℃ for 10-12 h. By the comparison of amino acid sequence homology, hydrophobic residues analysis and secondary structure prediction, it was found that D-hydantoinase reported herein is quite similar to that from Pseudomonas putdia CCRC12857, and alike to that from Pseudomonas putdia DSM84 or other bacteria. A rapid and efficient purification procedure of the enzyme was performed by a three-step procedure: ammonium sulfate fractionation, phenyl Sepharose hydrophobic interaction chromatography and Sephacryl S-200 gel filtration. The molecular mass of the monomeric enzyme is 52042 Da as determined by MALDI-TOF mass spectrometry.

  14. Ni2+ -uptake in Pseudomonas putida strain S4: a possible role of Mg2+ -uptake pump

    Indian Academy of Sciences (India)

    V N Tripathi; S Srivastava

    2006-03-01

    Essential metal ion homeostasis is based on regulated uptake of metal ions, both during its scarcity and abundance. Pseudomonas putida strain S4, a multimetal resistant bacterium, was employed to investigate Ni2+ entry into cells. It was observed that Mg2+ regulates the entry of Ni2+ and by this plays a protective role to minimize Ni2+ toxicity in this strain. This protection was evident in both growth as well as viability. Intracellular accumulation of Ni2+ varied in accordance with Mg2+ concentrations in the medium. It was hypothesized that Ni2+ enters the cell using a broad Mg2+ pump, i.e. the CorA system, as the CorA inhibitor, i.e. Co(III) Hex, also inhibits Ni2+ uptake. This led to the inference that Mg2+-based protection was basically due to competitive inhibition of Ni2+ uptake. We also show that Zn2+ can further regulate the entry of Ni2+.

  15. Metabolic Panel

    Science.gov (United States)

    ... basic metabolic panel (BMP) and comprehensive metabolic panel (CMP). The BMP checks your blood sugar, calcium, and ... as creatinine to check your kidney function. The CMP includes all of those tests, as well as ...

  16. Metabolic Disorders

    Science.gov (United States)

    ... as your liver, muscles, and body fat. A metabolic disorder occurs when abnormal chemical reactions in your body ... that produce the energy. You can develop a metabolic disorder when some organs, such as your liver or ...

  17. Metabolic Syndrome

    Science.gov (United States)

    Metabolic syndrome is a group of conditions that put you at risk for heart disease and diabetes. These ... doctors agree on the definition or cause of metabolic syndrome. The cause might be insulin resistance. Insulin is ...

  18. [Metabolic syndrome].

    Science.gov (United States)

    Mitsuishi, Masanori; Miyashita, Kazutoshi; Itoh, Hiroshi

    2009-02-01

    Metabolic syndrome, which is consisted of hypertension, dyslipidemia and impaired glucose tolerance, is one of the most significant lifestyle-related disorders that lead to cardiovascular diseases. Among many upstream factors that are related to metabolic syndrome, obesity, especially visceral obesity, plays an essential role in its pathogenesis. In recent studies, possible mechanisms which connect obesity to metabolic syndrome have been elucidated, such as inflammation, abnormal secretion of adipokines and mitochondrial dysfunction. In this review, we focus on the relationship between obesity and metabolic syndrome; and illustrate how visceral obesity contributes to, and how the treatments for obesity act on metabolic syndrome.

  19. Effect of mutation of chemotaxis signal transduction gene cheA in Pseudomonas putida DLL-1 on its chemotaxis and methyl parathion biodegradation%趋化信号转导基因cheA突变对Pseudomonas putida DLL-1甲基对硫磷的趋化性及原位降解的影响

    Institute of Scientific and Technical Information of China (English)

    文阳; 蒋建东; 邓海华; 蓝鸿; 李顺鹏

    2007-01-01

    Pseudomonas putida DLL-1是一株甲基对硫磷(MP)高效降解菌株,同时对MP具有趋化性.cheA基因是菌株趋化信号转导过程中负责编码组氨酸激酶的基因,为了研究菌株趋化性在农药原位降解中的作用,通过基因打靶的方式使P.putida DLL-1染色体上单拷贝的cheA基因失活,成功地获得了MP的趋化突变株P.putida DAK,突变株与野生菌株生长能力没有显著差异.通过土壤盆钵试验(MP浓度为50mg/kg),发现在灭菌与未灭菌土壤中趋化突变株对MP的降解能力低于原始出发菌株DLL-1约20%~30%,说明菌株DLL-1趋化性的丧失会减慢其对农药的降解,趋化性在农药的原位降解过程中发挥重要作用.

  20. In situ detection of aromatic compounds with biosensor Pseudomonas putida cells preserved and delivered to soil in water-soluble gelatin capsules.

    Science.gov (United States)

    de las Heras, Aitor; de Lorenzo, Víctor

    2011-05-01

    While many types of bacteria have been engineered to produce an optical output in response to given analytes in a culture, their use for extensive, in situ monitoring of distinct chemical species in soil is hampered by a dearth of practicable spreading schemes. In this work, we report and validate a comprehensive system for the long-term preservation of Pseudomonas putida cells genetically designed for biosensing benzene, toluene, ethylbenzene, and xylenes (BTEX) in soil, along with a procedure to formulate, spread, and vigorously activate such bacteria at the desired site and occasion. To this end, various known lyoprotectants were tested for promoting the long-term maintenance of biosensor cells with quite variable outcomes. While a formulation of inositol and maltodextrines was optimal for preservation of freeze-dried BTEX-sensing bacteria, adsorption of P. putida cells to corncob powder (an abundant residue of the corn industry) endowed the resulting material with a lasting viability at ambient conditions. In any case, the thereby preserved bacterial biomass acquired physical and mechanical properties adequate for formulating the biosensor agent in water-soluble but otherwise hard dry gelatine capsules with a long shelf life. When such capsules were spread in a soil microcosm and subsequently liquefied with water or high humidity, the released microorganisms formed spots that gave an intense luminiscent signal upon exposure to effectors of the sensor circuit implanted in the chromosome of the P. putida strain. We argue that the procedures described here can facilitate implementation of wide-area biological detection strategies for revealing the location of toxic or perilous chemicals.

  1. Surface display of monkey metallothionein {alpha} tandem repeats and EGFP fusion protein on Pseudomonas putida X4 for biosorption and detection of cadmium

    Energy Technology Data Exchange (ETDEWEB)

    He, Xiaochuan; Chen, Wenli; Huang, Qiaoyun [Huazhong Agricultural Univ., Wuhan (China). State Key Lab. of Agricultural Microbiology

    2012-09-15

    Monkey metallothionein {alpha} domain tandem repeats (4mMT{alpha}), which exhibit high cadmium affinity, have been displayed for the first time on the surface of a bacterium using ice nucleation protein N-domain (inaXN) protein from the Xanthomonas campestris pv (ACCC - 10049) as an anchoring motif. The shuttle vector pIME, which codes for INAXN-4mMT{alpha}-EGFP fusion, was constructed and used to target 4mMT{alpha} and EGFP on the surface of Pseudomonas putida X4 (CCTCC - 209319). The surface location of the INAXN-4mMT{alpha}-EGFP fusion was further verified by western blot analysis and immunofluorescence microscopy. The growth of X4 showed resistance to cadmium presence. The presence of surface-exposed 4mMT{alpha} on the engineered strains was four times higher than that of the wild-type X4. The Cd{sup 2+} accumulation by X4/pIME was not only four times greater than that of the original host bacterial cells but was also remarkably unaffected by the presence of Cu{sup 2+} and Zn{sup 2+}. Moreover, the surface-engineered strains could effectively bind Cd{sup 2+} under a wide range of pH levels, from 4 to 7. P. putida X4/pIME with surface-expressed 4mMT{alpha}-EGFP had twice the cadmium binding capacity as well as 1.4 times the fluorescence as the cytoplasmic 4mMTa-EGFP. These results suggest that P. putida X4 expressing 4mMT{alpha}-EGFP with the INAXN anchor motif on the surface would be a useful tool for the remediation and biodetection of environmental cadmium contaminants. (orig.)

  2. Identification and characterization of genes encoding polycyclic aromatic hydrocarbon dioxygenase and polycyclic aromatic hydrocarbon dihydrodiol dehydrogenase in Pseudomonas putida OUS82.

    OpenAIRE

    Takizawa, N; Kaida, N; Torigoe, S; Moritani,T.; Sawada, T.; Satoh, S.; Kiyohara, H

    1994-01-01

    Naphthalene and phenanthrene are transformed by enzymes encoded by the pah gene cluster of Pseudomonas putida OUS82. The pahA and pahB genes, which encode the first and second enzymes, dioxygenase and cis-dihydrodiol dehydrogenase, respectively, were identified and sequenced. The DNA sequences showed that pahA and pahB were clustered and that pahA consisted of four cistrons, pahAa, pahAb, pahAc, and pahAd, which encode ferredoxin reductase, ferredoxin, and two subunits of the iron-sulfur prot...

  3. Elevated mutation frequency in surviving populations of carbon-starved rpoS-deficient Pseudomonas putida is caused by reduced expression of superoxide dismutase and catalase.

    Science.gov (United States)

    Tarassova, Kairi; Tegova, Radi; Tover, Andres; Teras, Riho; Tark, Mariliis; Saumaa, Signe; Kivisaar, Maia

    2009-06-01

    RpoS is a bacterial sigma factor of RNA polymerase which is involved in the expression of a large number of genes to facilitate survival under starvation conditions and other stresses. The results of our study demonstrate that the frequency of emergence of base substitution mutants is significantly increased in long-term-starved populations of rpoS-deficient Pseudomonas putida cells. The increasing effect of the lack of RpoS on the mutation frequency became apparent in both a plasmid-based test system measuring Phe(+) reversion and a chromosomal rpoB system detecting rifampin-resistant mutants. The elevated mutation frequency coincided with the death of about 95% of the cells in a population of rpoS-deficient P. putida. Artificial overexpression of superoxide dismutase or catalase in the rpoS-deficient strain restored the survival of cells and resulted in a decline in the mutation frequency. This indicated that, compared to wild-type bacteria, rpoS-deficient cells are less protected against damage caused by reactive oxygen species. 7,8-Dihydro-8-oxoguanine (GO) is known to be one of the most stable and frequent base modifications caused by oxygen radical attack on DNA. However, the spectrum of base substitution mutations characterized in rpoS-deficient P. putida was different from that in bacteria lacking the GO repair system: it was broader and more similar to that identified in the wild-type strain. Interestingly, the formation of large deletions was also accompanied by a lack of RpoS. Thus, the accumulation of DNA damage other than GO elevates the frequency of mutation in these bacteria. It is known that oxidative damage of proteins and membrane components, but not that of DNA, is a major reason for the death of cells. Since the increased mutation frequency was associated with a decline in the viability of bacteria, we suppose that the elevation of the mutation frequency in the surviving population of carbon-starved rpoS-deficient P. putida may be caused both by

  4. Pseudomonas putida attunes morphophysiological, biochemical and molecular responses in Cicer arietinum L. during drought stress and recovery.

    Science.gov (United States)

    Tiwari, Shalini; Lata, Charu; Chauhan, Puneet Singh; Nautiyal, Chandra Shekhar

    2016-02-01

    Drought is one of the most important abiotic stresses that adversely affect plant growth and yield potential. However, some drought resistant rhizosphere competent bacteria are known to improve plant health and promote growth during abiotic stresses. Present study showed the role of Pseudomonas putida MTCC5279 (RA) in ameliorating drought stress on cv. BG-362 (desi) and cv. BG-1003 (kabuli) chickpea cultivars under in vitro and green house conditions. Polyethylene glycol-induced drought stress severely affected seed germination in both cultivars which was considerably improved on RA-inoculation. Drought stress significantly affected various growth parameters, water status, membrane integrity, osmolyte accumulation, ROS scavenging ability and stress-responsive gene expressions, which were positively modulated upon application of RA in both chickpea cultivars. Quantitative real-time (qRT)-PCR analysis showed differential expression of genes involved in transcription activation (DREB1A and NAC1), stress response (LEA and DHN), ROS scavenging (CAT, APX, GST), ethylene biosynthesis (ACO and ACS), salicylic acid (PR1) and jasmonate (MYC2) signalling in both chickpea cultivars exposed to drought stress and recovery in the presence or absence of RA. The observations imply that RA confers drought tolerance in chickpea by altering various physical, physiological and biochemical parameters, as well as by modulating differential expression of at least 11 stress-responsive genes. To the best of our knowledge, this is the first report on detailed analysis of plant growth promotion and stress alleviation in one month old desi and kabuli chickpea subjected to drought stress for 0, 1, 3 and 7 days and recovery in the presence of a PGPR.

  5. Biosynthesis and characterization of polyhydroxyalkanoates copolymers produced by Pseudomonas putida Bet001 isolated from palm oil mill effluent.

    Directory of Open Access Journals (Sweden)

    Ahmad Mohammed Gumel

    Full Text Available The biosynthesis and characterization of medium chain length poly-3-hydroxyalkanoates (mcl-PHA produced by Pseudomonas putida Bet001 isolated from palm oil mill effluent was studied. The biosynthesis of mcl-PHA in this newly isolated microorganism follows a growth-associated trend. Mcl-PHA accumulation ranging from 49.7 to 68.9% on cell dry weight (CDW basis were observed when fatty acids ranging from octanoic acid (C(8:0 to oleic acid (C(18:1 were used as sole carbon and energy source. Molecular weight of the polymer was found to be ranging from 55.7 to 77.7 kDa. Depending on the type of fatty acid used, the (1H NMR and GCMSMS analyses of the chiral polymer showed a composition of even and odd carbon atom chain with monomer length of C4 to C14 with C8 and C10 as the principal monomers. No unsaturated monomer was detected. Thermo-chemical analyses showed the accumulated PHA to be semi-crystalline polymer with good thermal stability, having a thermal degradation temperature (T(d of 264.6 to 318.8 (± 0.2 (oC, melting temperature (T(m of 43. (± 0.2 (oC, glass transition temperature (T(g of -1.0 (± 0.2 (oC and apparent melting enthalpy of fusion (ΔH(f of 100.9 (± 0.1 J g(-1.

  6. Nucleotide Metabolism

    DEFF Research Database (Denmark)

    Martinussen, Jan; Willemoës, M.; Kilstrup, Mogens

    2011-01-01

    Metabolic pathways are connected through their utilization of nucleotides as supplier of energy, allosteric effectors, and their role in activation of intermediates. Therefore, any attempt to exploit a given living organism in a biotechnological process will have an impact on nucleotide metabolism....... The aim of this article is to provide knowledge of nucleotide metabolism and its regulation to facilitate interpretation of data arising from genetics, proteomics, and transcriptomics in connection with biotechnological processes and beyond....

  7. Nucleotide Metabolism

    DEFF Research Database (Denmark)

    Martinussen, Jan; Willemoës, M.; Kilstrup, Mogens

    2011-01-01

    Metabolic pathways are connected through their utilization of nucleotides as supplier of energy, allosteric effectors, and their role in activation of intermediates. Therefore, any attempt to exploit a given living organism in a biotechnological process will have an impact on nucleotide metabolism....... The aim of this article is to provide knowledge of nucleotide metabolism and its regulation to facilitate interpretation of data arising from genetics, proteomics, and transcriptomics in connection with biotechnological processes and beyond....

  8. In-vitro antibacterial activities of the essential oils of aromatic plants against Erwinia herbicola (Lohnis and pseudomonas putida (Kris Hamilton

    Directory of Open Access Journals (Sweden)

    Pandey Abhay K.

    2012-01-01

    Full Text Available This study was designed to examine in vitro antibacterial activities of essential oils extracted from 53 aromatic plants of Gorakhpur Division (UP, INDIA for the control of two phytopathogenic bacteria namely Erwinia herbicola and Pseudomonas putida causing several post-harvest diseases in fruits and vegetables. Out of 53 oils screened, 8 oils such as Chenopodium ambrosioides, Citrus aurantium, Clausena pentaphylla, Hyptis suaveolens, Lippia alba, Mentha arvensis, Ocimum sanctum and Vitex negundo completely inhibited the growth of test bacteria. Furthermore MIC & MBC values of C. ambrosioides oil were least for Erw. herbicola (0.25 & 2.0 μl/ml and Ps. putida (0.12 & 1.0 μl/ml respectively than other 7 oils as well as Agromycin and Streptomycin drugs used in current study. GC and GC-MS analysis of Chenopodium oil revealed presence of 125 major and minor compounds, out of them, 14 compounds were recognized. The findings concluded that Chenopodium oil may be regarded as safe antibacterial agent for the management of post-harvest diseases of fruits and vegetables.

  9. Functional characterization of a StyS sensor kinase reveals distinct domains associated with intracellular and extracellular sensing of styrene in P. putida CA-3

    Science.gov (United States)

    O'Leary, Niall D; Mooney, Aisling; O'Mahony, Mark; Dobson, Alan DW

    2014-01-01

    Bacterial two-component systems (TCSs) are of vital importance in the translation of rapidly changing environmental conditions into appropriate cellular regulatory responses enabling adaptation, growth, and survival. The diverse range of environmental signals that TCSs can process, coupled with discrete modular domains within TCS proteins, offers considerable potential for the rational design of bio-sensor and/or bio-reporter strains. In this study we functionally characterize the multi-domain StyS sensor kinase associated with sensing of the aromatic pollutant styrene by Pseudomonas putida CA-3. Deletion analysis of discrete domains was performed and the ability of the truncated StyS sensor proteins to activate a cognate reporter system in an E. coli host assessed. The essential histidine kinase and PAS input domains were identified for StyS dependent activation of the reporter system. However, co-expression of an ABC-transporter protein StyE, previously linked to styrene transport in P. putida CA-3, enabled activation of the reporter system with a StyS construct containing a non-essential PAS input domain, suggesting a novel role for intracellular detection and/or activation. Site directed mutagenesis and amino acid deletions were employed to further characterize the PAS sensing domains of both input regions. The potential implications of these findings in the use of multi-domain sensor kinases in rational design strategies and the potential link between transport and intracellular sensing are discussed. PMID:24637704

  10. Biosensor for direct determination of fenitrothion and EPN using recombinant Pseudomonas putida JS444 with surface-expressed organophosphorous hydrolase. 2. Modified carbon paste electrode.

    Science.gov (United States)

    Lei, Yu; Mulchandani, Priti; Chen, Wilfred; Mulchandani, Ashok

    2007-03-01

    A whole cell-based amperometric biosensor for highly selective, sensitive, rapid, and cost-effective determination of the organophosphate pesticides fenitrothion and ethyl p-nitrophenol thio-benzene phosphonate (EPN) is discussed. The biosensor comprised genetically engineered p-nitrophenol (PNP)-degrading bacteria Pseudomonas putida JS444 anchoring and displaying organophosphorous hydrolase (OPH) on its cell surface as biological sensing element and carbon paste electrode as the amperometric transducer. Surface-expressed OPH catalyzed the hydrolysis of organophosphorous pesticides such as fenitrothion and EPN to release PNP and 3-methyl-4- nitrophenol, respectively, which were subsequently degraded by the enzymatic machinery of P. putida JS444 through electrochemically active intermediates to the TCA cycle. The electro-oxidization current of the intermediates was measured and correlated to the concentration of organophosphates. Operating at optimum conditions, 0.086 mg dry wt of cell operating at 600 mV of applied potential (vs Ag/AgCl reference) in 50 mM citrate phosphate buffer, pH 7.5, with 50 muM CoCl2 at room temperature, the biosensor measured as low as 1.4 ppb of fenitrothion and 1.6 ppb of EPN. There was no interference from phenolic compounds, carbamate pesticides, triazine herbicides, or organophosphate pesticides without nitrophenyl substituent. The service life of the biosensor and the applicability to lake water were also demonstrated.

  11. Kemampuan Pseudomonas putida Pf-20 dan 24.7B untuk Memperbaiki Sifat Kimia Media Tumbuh dan Ketahanan Terinduksi Tembakau H877 terhadap Cucumber mosaic virus

    Directory of Open Access Journals (Sweden)

    W. S. Wahyuni

    2005-12-01

    Full Text Available P. putida Pf-20 from medium M1 (paddy soil, leaves compost and manure compost, 1:3:2 has a higher affinity and more rapid to solubilize phosphate on Pikovskaya medium, rather than P. putida 27.4B. Those affinities of both strains from the media M1 with CMV-48 were much greater than that of medium without CMV. After 64 days planted, the content of N total, C-organic, CEC, K20, and P205 were better on medium M1, but the pH was little bit decreased. The reduction of P04 and Fe in medium M1 could due to the bacterial growth, and for producing siderophore, particularly on the media with viral treatment. These bacteria strains did not affect on the plant height and number of leaves on media M1, M2 or M3, however, these bacteria affected on the total root length and root densities in the medium M1 with virus. The more available nutrient in the medium, the more rapid bacteria colonized rhizosphere and roots. Both of these bacteria were effective to reduce the disease severity of CMV on tobacco H877. The medium M1 was the best medium for bacterial and tobacco growth.

  12. Pb remobilization by bacterially mediated dissolution of pyromorphite Pb5(PO4)3Cl in presence of phosphate-solubilizing Pseudomonas putida.

    Science.gov (United States)

    Topolska, Justyna; Latowski, Dariusz; Kaschabek, Stefan; Manecki, Maciej; Merkel, Broder J; Rakovan, John

    2014-01-01

    Remediation of lead (Pb)-contaminated sites with phosphate amendments is one of the best studied and cost-effective methods for in situ immobilization. In this treatment, a very stable mineral, pyromorphite Pb5(PO4)3Cl, is formed. Several studies propose to improve this treatment method with the addition of phosphate-solubilizing bacteria (PSB). The effect of bacteria on solubilization of pyromorphite is unknown. In this study, the effect of the soil microorganisms on the stability of pyromorphite Pb5(PO4)3Cl has been investigated in a set of batch solution experiments. The mineral was reacted with Pseudomonas putida, a common soil microorganism. Dissolution of pyromorphite was enhanced by the presence of P. putida, resulting in an elevated Pb concentration in the solution. This occurred even when the bacteria were provided with an additional source of phosphate in the solution. Pyromorphite has been shown to be a potential source of nutrient phosphorus for common soil bacteria. Thus, the use of PSB in remediation treatments of Pb contaminated sites may have adverse long-term impacts on Pb immobilization. Conscious phosphate management is suggested for long-term sustainability of the in situ Pb immobilization by pyromorphite formation.

  13. Two-dimensional electrophoresis analysis of protein production during growth of Pseudomonas putida F1 on toluene, phenol, and their mixture.

    Science.gov (United States)

    Reardon, Kenneth F; Kim, Kee-Hong

    2002-07-01

    The protein profiles of Pseudomonas putida F1 during growth on toluene, phenol, and their mixture were examined by two-dimensional polyacrylamide gel electrophoresis. Although this bacterium uses the same catabolic pathway for both substrates, P. putida F1 produced specific sets of proteins in response to toluene and phenol as single or mixed substrates. Proteins associated with growth on these substrates could be classified into three categories: ten Group T proteins were associated with the degradation of toluene, seventeen Group P proteins were associated with the degradation of phenol, and one Group M protein was observed to be associated only with toluene-phenol mixture degradation. During growth on the mixture, the protein profile of the cells shifted from Group T proteins to Group P proteins. This correlated well with the substrate consumption pattern, in which toluene was consumed first and growth on phenol did not begin until the medium was nearly depleted of toluene. Individual Group T and Group P protein intracellular concentrations had different transients as the cells grew on the mixture; seven protein levels increased, four decreased, and sixteen reached a maximum and then declined. The Group M protein reached a concentration maximum near the time when growth on phenol began. Variations in the maintenance of these proteins were also noted. These results demonstrate that cells growing on a mixture of substrates undergo significant physiological changes. Further investigation of these changes is expected to shed light on the unusual biodegradation kinetics previously observed with this mixed-substrate system.

  14. Microcalorimetric and potentiometric titration studies on the adsorption of copper by P. putida and B. thuringiensis and their composites with minerals

    Energy Technology Data Exchange (ETDEWEB)

    Fang Linchuan [State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070 (China); Key Laboratory of Subtropical Agriculture and Environment, Ministry of Agriculture, College of Resources and Environment, Huazhong Agricultural University, Wuhan 430070 (China); Cai Peng; Li Pengxiang; Wu Huayong; Liang Wei; Rong Xingmin [Key Laboratory of Subtropical Agriculture and Environment, Ministry of Agriculture, College of Resources and Environment, Huazhong Agricultural University, Wuhan 430070 (China); Chen Wenli [State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070 (China); Huang Qiaoyun, E-mail: qyhuang@mail.hzau.edu.cn [State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070 (China); Key Laboratory of Subtropical Agriculture and Environment, Ministry of Agriculture, College of Resources and Environment, Huazhong Agricultural University, Wuhan 430070 (China)

    2010-09-15

    In order to have a better understanding of the interactions of heavy metals with bacteria and minerals in soil and associated environments, isothermal titration calorimetry (ITC), potentiometric titration and equilibrium sorption experiments were conducted to investigate the adsorption behavior of Cu(II) by Bacillus thuringiensis, Pseudomonas putida and their composites with minerals. The interaction of montmorillonite with bacteria increased the reactive sites and resulted in greater adsorption for Cu(II) on their composites, while decreased adsorption sites and capacities for Cu(II) were observed on goethite-bacteria composites. A gram-positive bacterium B. thuringiensis played a more important role than a gram-negative bacterium P. putida in determining the properties of the bacteria-minerals interfaces. The enthalpy changes ({Delta}H{sub ads}) from endothermic (6.14 kJ mol{sup -1}) to slightly exothermic (-0.78 kJ mol{sup -1}) suggested that Cu(II) is complexed with the anionic oxygen ligands on the surface of bacteria-mineral composites. Large entropies (32.96-58.89 J mol{sup -1} K{sup -1}) of Cu(II) adsorption onto bacteria-mineral composites demonstrated the formation of inner-sphere complexes in the presence of bacteria. The thermodynamic data implied that Cu(II) mainly bound to the carboxyl and phosphoryl groups as inner-sphere complexes on bacteria and mineral-bacteria composites.

  15. Metabolic acidosis.

    Science.gov (United States)

    Lim, Salim

    2007-01-01

    Acute metabolic acidosis is frequently encountered in critically ill patients. Metabolic acidosis can occur as a result of either the accumulation of endogenous acids that consumes bicarbonate (high anion gap metabolic acidosis) or loss of bicarbonate from the gastrointestinal tract or the kidney (hyperchloremic or normal anion gap metabolic acidosis). The cause of high anion gap metabolic acidosis includes lactic acidosis, ketoacidosis, renal failure and intoxication with ethylene glycol, methanol, salicylate and less commonly with pyroglutamic acid (5-oxoproline), propylene glycole or djenkol bean (gjenkolism). The most common causes of hyperchloremic metabolic acidosis are gastrointestinal bicarbonate loss, renal tubular acidosis, drugs-induced hyperkalemia, early renal failure and administration of acids. The appropriate treatment of acute metabolic acidosis, in particular organic form of acidosis such as lactic acidosis, has been very controversial. The only effective treatment for organic acidosis is cessation of acid production via improvement of tissue oxygenation. Treatment of acute organic acidosis with sodium bicarbonate failed to reduce the morbidity and mortality despite improvement in acid-base parameters. Further studies are required to determine the optimal treatment strategies for acute metabolic acidosis.

  16. Draft Genome Sequences of Pseudomonas fluorescens Strains PA4C2 and PA3G8 and Pseudomonas putida PA14H7, Three Biocontrol Bacteria against Dickeya Phytopathogens.

    Science.gov (United States)

    Cigna, Jérémy; Raoul des Essarts, Yannick; Mondy, Samuel; Hélias, Valérie; Beury-Cirou, Amélie; Faure, Denis

    2015-01-29

    Pseudomonas fluorescens strains PA4C2 and PA3G8 and Pseudomonas putida strain PA14H7 were isolated from potato rhizosphere and show an ability to inhibit the growth of Dickeya phytopathogens. Here, we report their draft genome sequences, which provide a basis for understanding the molecular mechanisms involved in antibiosis against Dickeya.

  17. Metabolic encephalopathies.

    Science.gov (United States)

    Angel, Michael J; Young, G Bryan

    2011-11-01

    Kinnier Wilson coined the term metabolic encephalopathy to describe a clinical state of global cerebral dysfunction induced by systemic stress that can vary in clinical presentation from mild executive dysfunction to deep coma with decerebrate posturing; the causes are numerous. Some mechanisms by which cerebral dysfunction occurs in metabolic encephalopathies include focal or global cerebral edema, alterations in transmitter function, the accumulation of uncleared toxic metabolites, postcapillary venule vasogenic edema, and energy failure. This article focuses on common causes of metabolic encephalopathy, and reviews common causes, clinical presentations and, where relevant, management.

  18. Metabolic Syndrome

    Science.gov (United States)

    ... hypertension, hypertriglyceridemia, insulin resistance syndrome, low HDL cholesterol, Metabolic Syndrome, overweight, syndrome x, type 2 diabetes Family Health, Kids and Teens, Men, Women January 2005 Copyright © American Academy of Family PhysiciansThis ...

  19. Systems analysis of solvent tolerance mechanisms in Pseudomonas putida S12 - Importance of energy metabolism and functional identification of the TrgI regulator

    NARCIS (Netherlands)

    Volkers, R.J.M.

    2012-01-01

    Hydrophobic organic solvents, like benzene and toluene, are mainly toxic for bacteria because of their accumulation in the membranes of the cells. The accumulation of solvent molecules in a membrane lowers its rigidity and increases its fluidity and permeability, resulting in an increased rate of ce

  20. Metabolic syndrome

    Directory of Open Access Journals (Sweden)

    Gogia Atul

    2006-02-01

    Full Text Available The Metabolic syndrome is a widely prevalent and multi-factorial disorder. The syndrome has been given several names, including- the metabolic syndrome, the insulin resistance syndrome, the plurimetabolic syndrome, and the deadly quartet. With the formulation of NCEP/ATP III guidelines, some uniformity and standardization has occurred in the definition of metabolic syndrome and has been very useful for epidemiological purposes. The mechanisms underlying the metabolic syndrome are not fully known; however resistance to insulin stimulated glucose uptake seems to modify biochemical responses in a way that predisposes to metabolic risk factors. The clinical relevance of the metabolic syndrome is related to its role in the development of cardiovascular disease. Management of the metabolic syndrome involves patient-education and intervention at various levels. Weight reduction is one of the main stays of treatment. In this article we comprehensively discuss this syndrome- the epidemiology, pathogenesis, clinical relevance and management. The need to do a comprehensive review of this particular syndrome has arisen in view of the ever increasing incidence of this entitiy. Soon, metabolic syndrome will overtake cigarette smoking as the number one risk factor for heart disease among the US population. Hardly any issue of any primary care medical journal can be opened without encountering an article on type 2 diabetes, dyslipidemia or hypertension. It is rare to see type 2 diabetes, dyslipidemia, obesity or hypertension in isolation. Insulin resistance and resulting hyperinsulinemia have been implicated in the development of glucose intolerance (and progression to type 2 diabetes, hypertriglyceridemia, hypertension, polycystic ovary yndrome, hypercoagulability and vascular inflammation, as well as the eventual development of atherosclerotic cardiovascular disease manifested as myocardial infarction, stroke and myriad end organ diseases. Conversely

  1. Lipid Metabolism

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    2008393 Effects of angiotensin Ⅱ type 1 receptor blocker on triglyceride metabolism in the liver: experiment with Zucker fatty rats. RAN Jianmin(冉建民), et al. Dept Endocrinol, Guangzhou Red Cross Hosp, 4th Hosp Med Coll, Jinan Univ, Guangzhou 510220. Natl Med J China 2008;88(22):1557-1561. Objective To investigate the effects of angiotensin receptor blocker (ARB) on triglyceride (TG) metabolism and mechanism thereof.

  2. Metabolic Syndrome (For Parents)

    Science.gov (United States)

    ... Old Feeding Your 1- to 2-Year-Old Metabolic Syndrome KidsHealth > For Parents > Metabolic Syndrome A A A ... this is a condition called metabolic syndrome . About Metabolic Syndrome Not to be confused with metabolic disease (which ...

  3. Simultaneous removal of Cr(Ⅵ) and phenol in consortium culture of Bacillus sp. and Pseudomonas putida Migula (CCTCC AB92019)

    Institute of Scientific and Technical Information of China (English)

    LIU Yun-guo; PAN Cui; XIA Wen-bin; ZENG Guang-ming; ZHOU Ming; LIU Yuan-yuan; KE Jie; HUANG Chao

    2008-01-01

    The simultaneous removal of Cr(Ⅵ) and phenol in a consortium culture containing Cr(Ⅵ) reducer, Bacillus sp. and phenol degrader, Pseudomonas putida Migula (CCTCC AB92019) was studied. Phenol was used as the sole carbon source. Bacillus sp. utilized metabolites formed from phenol degradation as electron donors and energy source for Cr(Ⅵ) reduction. Optimum Cr(Ⅵ) reduction was observed at a phenol concentration of 150 mg/L and an initial Cr(Ⅵ) concentration of 15 mg/L. Both the Cr(Ⅵ) reduction and phenol degradation were influenced by the cell composition of the culture, but the phenol degradation was not significantly affected by the content of Bacillus sp. The experiments also showed that the amount of phenol degraded was more than that stoichiometrically required for Cr(Ⅵ) reduction.

  4. Whole-cell bio-oxidation of n-dodecane using the alkane hydroxylase system of P. putida GPo1 expressed in E. coli

    DEFF Research Database (Denmark)

    Grant, Chris; Woodley, John; Baganz, Frank

    2011-01-01

    The alkane-1-monoxygenase (alkB) complex of Pseudomonas putida GPo1 has been extensively studied in the past and shown to be capable of oxidising aliphatic C5–C12 alkanes to primary alcohols both in the wild-type organism by growth on C5–C12 alkanes as sole carbon source and in vitro. Despite this......-octane (21μmol/min/gdcw); similar to the 5-fold difference observed between wild-type growth rates using the two respective alkanes as sole carbon source. Despite this, both total volumetric rate and final yield exceeded n-octane oxidation by 3.5-fold under the same conditions, due to the lower toxicity of n...

  5. Nitrogen regulation of the xyl genes of Pseudomonas putida mt-2 propagates into a significant effect of nitrate on m-xylene mineralization in soil

    DEFF Research Database (Denmark)

    Svenningsen, Nanna Bygvraa; Nicolaisen, Mette Haubjerg; Hansen, Hans Chr. Bruun;

    2016-01-01

    in the test tube is propagated into actual catabolism of, e.g. m-xylene in soil, the natural habitat of this bacterium. To address this issue, we have developed a test-tube-to-soil model system that exposes the end-effects of remediation practices influencing gene expression of P. putida mt-2. We found...... that NO3(-) compared with NH4(+) had a stimulating effect on xyl gene expression in pure culture as well as in soil, and that this stimulation was translated into increased m-xylene mineralization in soil. Furthermore, expression analysis of the nitrogen-regulated genes amtB and gdhA allowed us to monitor...... nitrogen sensing status in both experimental systems. Hence, for nitrogen sources, regulatory patterns that emerge in soil reflect those observed in liquid cultures. The current study shows how distinct regulatory traits can lead to discrete environmental consequences; and it underpins that attempts...

  6. Hidrólise ácida e enzimática de pectina para crescimento celular de Cupriavidus necator e Pseudomonas putida

    OpenAIRE

    Locatelli, Gabriel Olivo; Dias da Silva, Gabriella; Finkler, Leandro; Lamenha Luna Finkler, Christine

    2012-01-01

    Substâncias pécticas são encontradas em grandes quantidades nos resíduos da industrialização de sucos de frutas. Esses compostos são susceptíveis à hidrólise por métodos ácidos e enzimáticos, podendo ser empregados como substratos de baixo custo em processos de conversão biológica para obtenção de bioprodutos, como os biopolímeros. Cupriavidus necator e Pseudomonas putida são micro-organismos conhecidos por sua capacidade de acumular poli-hidroxialcanoatos (PHAs) como fonte de carbono e energ...

  7. The toxicity of the quaternary ammonium compound benzalkonium chloride alone and in mixtures with other anionic compounds to bacteria in test systems with Vibrio fischeri and Pseudomonas putida.

    Science.gov (United States)

    Sütterlin, H; Alexy, R; Kümmerer, K

    2008-10-01

    Mixtures of chemicals are present in the aquatic environment but standard testing methods assess only single compounds. One aspect of this question is the importance of the formation of ionic pairs, for example from quaternary ammonium compounds with organic anions, and the significance of the ionic pairs for bacterial toxicity in the aquatic environment. The aim of the present study was to investigate the toxicity of the cationic quaternary ammonium compound benzalkonium chloride (BAC) against aquatic bacteria in the presence of substances commonly found in wastewater, such as the anionic surfactant linear alkylbenzene sulfonate (LAS), naphthalene sulfonic acid (NSA), sodium dodecylsulfonate (SDS), and benzene sulfonic acid (BSA). The growth inhibition test with Pseudomonas putida and the Vibrio fischeri luminescent inhibition test were used to determine the toxicity of single compounds and compound mixtures. The results found in this study indicate that ion pair formation is of minor significance under the test conditions applied here.

  8. Expression of Fap amyloids in Pseudomonas aeruginosa, P. fluorescens, and P. putida results in aggregation and increased biofilm formation

    DEFF Research Database (Denmark)

    Dueholm, Morten S; Søndergaard, Mads; Nilsson, Martin

    2013-01-01

    resulted in highly aggregative phenotypes and in increased biofilm formation. Detailed biophysical investigations of purified fibrils confirmed FapC as the main fibril monomer and supported the role of FapB as a minor, nucleating constituent as also indicated by bioinformatic analysis. Bioinformatics......The fap operon, encoding functional amyloids in Pseudomonas (Fap), is present in most pseudomonads, but so far the expression and importance for biofilm formation has only been investigated for P. fluorescens strain UK4. In this study, we demonstrate the capacity of P. aeruginosa PAO1, P....... fluorescens Pf-5, and P. putida F1 to express Fap fibrils, and investigated the effect of Fap expression on aggregation and biofilm formation. The fap operon in all three Pseudomonas species conferred the ability to express Fap fibrils as shown using a recombinant approach. This Fap overexpression consistently...

  9. Molecular level biodegradation of phenol and its derivatives through dmp operon of Pseudomonas putida: A bio-molecular modeling and docking analysis.

    Science.gov (United States)

    Ray, Sujay; Banerjee, Arundhati

    2015-10-01

    Participation of Pseudomonas putida-derived methyl phenol (dmp) operon and DmpR protein in the biodegradation of phenol or other harmful, organic, toxic pollutants was investigated at a molecular level. Documentation documents that P. putida has DmpR protein which positively regulates dmp operon in the presence of inducers; like phenols. From the operon, phenol hydroxylase encoded by dmpN gene, participates in degrading phenols after dmp operon is expressed. For the purpose, the 3-D models of the four domains from DmpR protein and of the DNA sequences from the two Upstream Activation Sequences (UAS) present at the promoter region of the operon were demonstrated using discrete molecular modeling techniques. The best modeled structures satisfying their stereo-chemical properties were selected in each of the cases. To stabilize the individual structures, energy optimization was performed. In the presence of inducers, probable interactions among domains and then the two independent DNA structures with the fourth domain were perused by manifold molecular docking simulations. The complex structures were made to be stable by minimizing their overall energy. Responsible amino acid residues, nucleotide bases and binding patterns for the biodegradation, were examined. In the presence of the inducers, the biodegradation process is initiated by the interaction of phe50 from the first protein domain with the inducers. Only after the interaction of the last domain with the DNA sequences individually, the operon is expressed. This novel residue level study is paramount for initiating transcription in the operon; thereby leading to expression of phenol hydroxylase followed by phenol biodegradation.

  10. Elimination of manganese(II,III) oxidation in Pseudomonas putida GB-1 by a double knockout of two putative multicopper oxidase genes.

    Science.gov (United States)

    Geszvain, Kati; McCarthy, James K; Tebo, Bradley M

    2013-01-01

    Bacterial manganese(II) oxidation impacts the redox cycling of Mn, other elements, and compounds in the environment; therefore, it is important to understand the mechanisms of and enzymes responsible for Mn(II) oxidation. In several Mn(II)-oxidizing organisms, the identified Mn(II) oxidase belongs to either the multicopper oxidase (MCO) or the heme peroxidase family of proteins. However, the identity of the oxidase in Pseudomonas putida GB-1 has long remained unknown. To identify the P. putida GB-1 oxidase, we searched its genome and found several homologues of known or suspected Mn(II) oxidase-encoding genes (mnxG, mofA, moxA, and mopA). To narrow this list, we assumed that the Mn(II) oxidase gene would be conserved among Mn(II)-oxidizing pseudomonads but not in nonoxidizers and performed a genome comparison to 11 Pseudomonas species. We further assumed that the oxidase gene would be regulated by MnxR, a transcription factor required for Mn(II) oxidation. Two loci met all these criteria: PputGB1_2447, which encodes an MCO homologous to MnxG, and PputGB1_2665, which encodes an MCO with very low homology to MofA. In-frame deletions of each locus resulted in strains that retained some ability to oxidize Mn(II) or Mn(III); loss of oxidation was attained only upon deletion of both genes. These results suggest that PputGB1_2447 and PputGB1_2665 encode two MCOs that are independently capable of oxidizing both Mn(II) and Mn(III). The purpose of this redundancy is unclear; however, differences in oxidation phenotype for the single mutants suggest specialization in function for the two enzymes.

  11. Synthesis of Diblock copolymer poly-3-hydroxybutyrate -block-poly-3-hydroxyhexanoate [PHB-b-PHHx] by a β-oxidation weakened Pseudomonas putida KT2442

    Directory of Open Access Journals (Sweden)

    Tripathi Lakshmi

    2012-04-01

    Full Text Available Abstract Background Block polyhydroxyalkanoates (PHA were reported to be resistant against polymer aging that negatively affects polymer properties. Recently, more and more attempts have been directed to make PHA block copolymers. Diblock copolymers PHB-b-PHHx consisting of poly-3-hydroxybutyrate (PHB block covalently bonded with poly-3-hydroxyhexanoate (PHHx block were for the first time produced successfully by a recombinant Pseudomonas putida KT2442 with its β-oxidation cycle deleted to its maximum. Results The chloroform extracted polymers were characterized by nuclear magnetic resonance (NMR, thermo- and mechanical analysis. NMR confirmed the existence of diblock copolymers consisting of 58 mol% PHB as the short chain length block with 42 mol% PHHx as the medium chain length block. The block copolymers had two glass transition temperatures (Tg at 2.7°C and −16.4°C, one melting temperature (Tm at 172.1°C and one cool crystallization temperature (Tc at 69.1°C as revealed by differential scanning calorimetry (DSC, respectively. This is the first microbial short-chain-length (scl and medium-chain-length (mcl PHA block copolymer reported. Conclusions It is possible to produce PHA block copolymers of various kinds using the recombinant Pseudomonas putida KT2442 with its β-oxidation cycle deleted to its maximum. In comparison to a random copolymer poly-3-hydroxybutyrate-co-3-hydroxyhexanoate (P(HB-co-HHx and a blend sample of PHB and PHHx, the PHB-b-PHHx showed improved structural related mechanical properties.

  12. Display of a thermostable lipase on the surface of a solvent-resistant bacterium, Pseudomonas putida GM730, and its applications in whole-cell biocatalysis

    Directory of Open Access Journals (Sweden)

    Kwon Seok-Joon

    2006-04-01

    Full Text Available Abstract Background Whole-cell biocatalysis in organic solvents has been widely applied to industrial bioprocesses. In two-phase water-solvent processes, substrate conversion yields and volumetric productivities can be limited by the toxicity of solvents to host cells and by the low mass transfer rates of the substrates from the solvent phase to the whole-cell biocatalysts in water. Results To solve the problem of solvent toxicity, we immobilized a thermostable lipase (TliA from Pseudomonas fluorescens on the cell surface of a solvent-resistant bacterium, Pseudomonas putida GM730. Surface immobilization of enzymes eliminates the mass-transfer limitation imposed by the cell wall and membranes. TliA was successfully immobilized on the surface of P. putida cells using the ice-nucleation protein (INP anchoring motif from Pseudomonas syrinage. The surface location was confirmed by flow cytometry, protease accessibility and whole-cell enzyme activity using a membrane-impermeable substrate. Three hundred and fifty units of whole-cell hydrolytic activity per gram dry cell mass were obtained when the enzyme was immobilized with a shorter INP anchoring motif (INPNC. The surface-immobilized TliA retained full enzyme activity in a two-phase water-isooctane reaction system after incubation at 37°C for 12 h, while the activity of the free form enzyme decreased to 65% of its initial value. Whole cells presenting immobilized TliA were shown to catalyze three representative lipase reactions: hydrolysis of olive oil, synthesis of triacylglycerol and chiral resolution. Conclusion In vivo surface immobilization of enzymes on solvent-resistant bacteria was demonstrated, and appears to be useful for a variety of whole-cell bioconversions in the presence of organic solvents.

  13. Prediction of the adaptability of Pseudomonas putida DOT-T1E to a second phase of a solvent for economically sound two-phase biotransformations.

    Science.gov (United States)

    Neumann, Grit; Kabelitz, Nadja; Zehnsdorf, Andreas; Miltner, Anja; Lippold, Holger; Meyer, Daniel; Schmid, Andreas; Heipieper, Hermann J

    2005-11-01

    The strain Pseudomonas putida DOT-T1E was tested for its ability to tolerate second phases of different alkanols for their use as solvents in two-liquid-phase biotransformations. Although 1-decanol showed an about 10-fold higher toxicity to the cells than 1-octanol, the cells were able to adapt completely to 1-decanol only and could not be adapted in order to grow stably in the presence of a second phase of 1-octanol. The main explanation for this observation can be seen in the higher water and membrane solubility of 1-octanol. The hydrophobicity (log P) of a substance correlates with a certain partitioning of that compound into the membrane. Combining the log P value with the water solubility, the maximum membrane concentration of a compound can be calculated. With this simple calculation, it is possible to predict the property of an organic chemical for its potential applicability as a solvent for two-liquid-phase biotransformations with solvent-tolerant P. putida strains. Only compounds that show a maximum membrane concentration of less than 400 mM, such as 1-decanol, seem to be tolerated by these bacterial strains when applied in supersaturating concentrations to the medium. Taking into consideration that a solvent for a two-liquid-phase system should possess partitioning properties for potential substrates and products of a fine chemical synthesis, it can be seen that 1-decanol is a suitable solvent for such biotransformation processes. This was also demonstrated in shake cultures, where increasing amounts of a second phase of 1-decanol led to bacteria tolerating higher concentrations of the model substrate 3-nitrotoluene. Transferring this example to a 5-liter-scale bioreactor with 10% (vol/vol) 1-decanol, the amount of 3-nitrotoluene tolerated by the cells is up to 200-fold higher than in pure aqueous medium. The system demonstrates the usefulness of two-phase biotransformations utilizing solvent-tolerant bacteria.

  14. Rationally rewiring the connectivity of the XylR/Pu regulatory node of the m-xylene degradation pathway in Pseudomonas putida.

    Science.gov (United States)

    de Las Heras, Aitor; Martínez-García, Esteban; Domingo-Sananes, Maria Rosa; Fraile, Sofia; de Lorenzo, Víctor

    2016-04-18

    The XylR/Pu regulatory node of the m-xylene biodegradation pathway of Pseudomonas putida mt-2 is one of the most intricate cases of processing internal and external cues into a single controlling element. Despite this complexity, the performance of the regulatory system is determined in vivo only by the occupation of Pu by m-xylene-activated XylR and σ(54)-RNAP. The stoichiometry between these three elements defines natural system boundaries that outline a specific functional space. This space can be expanded artificially following different strategies that involve either the increase of XylR or σ(54) or both elements at the same time (each using a different inducer). In this work we have designed a new regulatory architecture that drives the system to reach a maximum performance in response to one single input. To this end, we first explored using a simple mathematical model whether the output of the XylR/Pu node could be amended by simultaneously increasing σ(54) and XylR in response to only natural inducers. The exacerbation of Pu activity in vivo was tested in strains bearing synthetic transposons encoding xylR and rpoN (the σ(54) coding gene) controlled also by Pu, thereby generating a P. putida strain with the XylR/Pu output controlled by two intertwined feed forward loops (FFLs). The lack of a negative feedback loop in the expression node enables Pu activity to reach its physiological maximum in response to a single input. Only competition for cell resources might ultimately check the upper activity limit of such a rewired m-xylene sensing device.

  15. Oxidation of methyl tert-butyl ether by alkane hydroxylase in dicyclopropylketone-induced and n-octane-grown Pseudomonas putida GPo1.

    Science.gov (United States)

    Smith, Christy A; Hyman, Michael R

    2004-08-01

    The alkane hydroxylase enzyme system in Pseudomonas putida GPo1 has previously been reported to be unreactive toward the gasoline oxygenate methyl tert-butyl ether (MTBE). We have reexamined this finding by using cells of strain GPo1 grown in rich medium containing dicyclopropylketone (DCPK), a potent gratuitous inducer of alkane hydroxylase activity. Cells grown with DCPK oxidized MTBE and generated stoichiometric quantities of tert-butyl alcohol (TBA). Cells grown in the presence of DCPK also oxidized tert-amyl methyl ether but did not appear to oxidize either TBA, ethyl tert-butyl ether, or tert-amyl alcohol. Evidence linking MTBE oxidation to alkane hydroxylase activity was obtained through several approaches. First, no TBA production from MTBE was observed with cells of strain GPo1 grown on rich medium without DCPK. Second, no TBA production from MTBE was observed in DCPK-treated cells of P. putida GPo12, a strain that lacks the alkane-hydroxylase-encoding OCT plasmid. Third, all n-alkanes that support the growth of strain GPo1 inhibited MTBE oxidation by DCPK-treated cells. Fourth, two non-growth-supporting n-alkanes (propane and n-butane) inhibited MTBE oxidation in a saturable, concentration-dependent process. Fifth, 1,7-octadiyne, a putative mechanism-based inactivator of alkane hydroxylase, fully inhibited TBA production from MTBE. Sixth, MTBE-oxidizing activity was also observed in n-octane-grown cells. Kinetic studies with strain GPo1 grown on n-octane or rich medium with DCPK suggest that MTBE-oxidizing activity may have previously gone undetected in n-octane-grown cells because of the unusually high K(s) value (20 to 40 mM) for MTBE.

  16. Kinetics studies of p-cresol biodegradation by using Pseudomonas putida in batch reactor and in continuous bioreactor packed with calcium alginate beads.

    Science.gov (United States)

    Mathur, A K; Bala, Shashi; Majumder, C B; Sarkar, S

    2010-01-01

    Present study deals with the biodegradation of p-cresol by using Pseudomonas putida in a batch reactor and a continuous bioreactor packed with calcium alginate beads. The maximum specific growth rate of 0.8121 h(-1) was obtained at 200 mg L(-1) concentration of p-cresol in batch reactor. The maximum p-cresol degradation rate was obtained 6.598 mg L(-1) h(-1) at S(o)=200 mg L(-1) and 62.8 mg L(-1) h(-1) at S(o)=500 mg L(-1) for batch reactor and a continuous bioreactor, respectively. The p-cresol degradation rate of continuous bioreactor was 9 to 10-fold higher than those of the batch reactor. It shows that the continuous bioreactor could tolerate a higher concentration of p-cresol. A Haldane model was also used for p-cresol inhibition in batch reactor and a modified equation similar to Haldane model for continuous bioreactor. The Haldane parameters were obtained as µ(max) 0.3398 h(-1), K(s) 110.9574 mg L(-1), and K(I) 497.6169 mg L(-1) in batch reactor. The parameters used in continuous bioreactor were obtained as D(max) 91.801 mg L(-1) h(-1), K(s) 131.292 mg L(-1), and K(I) 1217.7 mg L(-1). The value K(I) of continuous bioreactor is approximately 2.5 times higher than the batch reactor. Higher K(I) value of continuous bioreactor indicates P. putida can grow at high range of p-cresol concentration. The ability of tolerance of higher p-cresol concentrations may be one reason for biofilm attachment on the packed bed in the continuous operation.

  17. Metabolic syndrome

    Institute of Scientific and Technical Information of China (English)

    Charles Shaeffer

    2004-01-01

    @@ The emergence of cardiac disease as the number one world-wide cause of death justifies efforts to identify individuals at higher risk for preventive therapy. The metabolic syndrome, originally described by Reaven, 1 has been associated with higher cardiovascular disease risk. 2 Type Ⅱ diabetes is also a frequent sequela. 3

  18. Oil sands to the rescue: oil sand microbial communities can degrade recalcitrant alkyl phenyl alkanoic acids

    Energy Technology Data Exchange (ETDEWEB)

    Whitby, Corinne [University of Essex (Canada)], email: cwhitby@essex.ac.uk

    2011-07-01

    Almost half of all global oil reserves are found as biodegraded heavy oils found in vast tar sand deposits located in North and South America and these account for 47% of Canadian oil production. Oil sand extraction generates large amounts of toxic waste water, known as oil sand process waters (OSPW), that are stored in large tailing ponds that contain toxic compounds like naphthenic acids (NAs). The presence of NAs creates problems like toxicity, corrosion, and the formation of calcium napthenate deposits which block pipelines and other infrastructure and need to be removed. This paper presents oil sand microbial communities that can degrade these NAs. The approach is to apply new aliphatic and aromatic NAs as substrates to supplement and identify NA degrading microbes and also to identify the metabolites produced and explain NA degradation pathways and the functional genes involved. The chemistry and the processes involved are explained. From the results, it is suggested that pure cultures of P. putida KT2440 be used against NAs.

  19. Enhanced yield of medium-chain-length polyhydroxyalkanoates from nonanoic acid by co-feeding glucose in carbon-limited, fed-batch culture.

    Science.gov (United States)

    Sun, Zhiyong; Ramsay, Juliana; Guay, Martin; Ramsay, Bruce

    2009-09-25

    Medium-chain-length polyhydroxyalkanoates (MCL-PHAs) were produced in carbon-limited, single-stage, fed-batch fermentations of Pseudomonas putida KT2440 by co-feeding nonanoic acid (NA) and glucose (G) to enhance the yield of PHA from NA. An exponential (mu=0.25 h(-1)) followed by a linear feeding strategy at a NA:G ratio of 1:1 (w/w) achieved 71 g l(-1) biomass containing 56% PHA. Although the same overall PHA productivity (1.44 g l(-1) h(-1)) was obtained when NA alone was fed at the same specific growth rate, the overall yield of PHA from NA increased by 25% (0.66 g PHA g NA(-1) versus 0.53 g g(-1)) with glucose co-feeding. Further increasing glucose in the feed (NA:G=1:1.5) resulted in a slightly higher yield (0.69 g PHA g NA(-1)) but lower PHA content (48%) and productivity (1.16 g l(-1) h(-1)). There was very little change in the PHA composition.

  20. Identification of flgZ as a flagellar gene encoding a PilZ domain protein that regulates swimming motility and biofilm formation in Pseudomonas.

    Science.gov (United States)

    Martínez-Granero, Francisco; Navazo, Ana; Barahona, Emma; Redondo-Nieto, Miguel; González de Heredia, Elena; Baena, Irene; Martín-Martín, Irene; Rivilla, Rafael; Martín, Marta

    2014-01-01

    Diguanylate cyclase and phosphodiesterase enzymatic activities control c-di-GMP levels modulating planktonic versus sessile lifestyle behavior in bacteria. The PilZ domain is described as a sensor of c-di-GMP intracellular levels and the proteins containing a PilZ domain represent the best studied class of c-di-GMP receptors forming part of the c-di-GMP signaling cascade. In P. fluorescens F113 we have found two diguanylate cyclases (WspR, SadC) and one phosphodiesterase (BifA) implicated in regulation of swimming motility and biofilm formation. Here we identify a flgZ gene located in a flagellar operon encoding a protein that contains a PilZ domain. Moreover, we show that FlgZ subcellular localization depends on the c-di-GMP intracellular levels. The overexpression analysis of flgZ in P. fluorescens F113 and P. putida KT2440 backgrounds reveal a participation of FlgZ in Pseudomonas swimming motility regulation. Besides, the epistasis of flgZ over wspR and bifA clearly shows that c-di-GMP intracellular levels produced by the enzymatic activity of the diguanylate cyclase WspR and the phosphodiesterase BifA regulates biofilm formation through FlgZ.

  1. Identification of flgZ as a flagellar gene encoding a PilZ domain protein that regulates swimming motility and biofilm formation in Pseudomonas.

    Directory of Open Access Journals (Sweden)

    Francisco Martínez-Granero

    Full Text Available Diguanylate cyclase and phosphodiesterase enzymatic activities control c-di-GMP levels modulating planktonic versus sessile lifestyle behavior in bacteria. The PilZ domain is described as a sensor of c-di-GMP intracellular levels and the proteins containing a PilZ domain represent the best studied class of c-di-GMP receptors forming part of the c-di-GMP signaling cascade. In P. fluorescens F113 we have found two diguanylate cyclases (WspR, SadC and one phosphodiesterase (BifA implicated in regulation of swimming motility and biofilm formation. Here we identify a flgZ gene located in a flagellar operon encoding a protein that contains a PilZ domain. Moreover, we show that FlgZ subcellular localization depends on the c-di-GMP intracellular levels. The overexpression analysis of flgZ in P. fluorescens F113 and P. putida KT2440 backgrounds reveal a participation of FlgZ in Pseudomonas swimming motility regulation. Besides, the epistasis of flgZ over wspR and bifA clearly shows that c-di-GMP intracellular levels produced by the enzymatic activity of the diguanylate cyclase WspR and the phosphodiesterase BifA regulates biofilm formation through FlgZ.

  2. Effect of red clay on diesel bioremediation and soil bacterial community.

    Science.gov (United States)

    Jung, Jaejoon; Choi, Sungjong; Hong, Hyerim; Sung, Jung-Suk; Park, Woojun

    2014-08-01

    Red clay is a type of soil, the red color of which results from the presence of iron oxide. It is considered an eco-friendly material, with many industrial, cosmetic, and architectural uses. A patented method was applied to red clay in order to change its chemical composition and mineral bioavailability. The resulting product was designated processed red clay. This study evaluates the novel use of red clay and processed red clay as biostimulation agents in diesel-contaminated soils. Diesel biodegradation was enhanced in the presence of red clay and processed red clay by 4.9- and 6.7-fold, respectively, and the number of culturable bacterial cells was correlated with the amount of diesel biodegradation. The growth of Acinetobacter oleivorans DR1, Pseudomonas putida KT2440, and Cupriavidus necator was promoted by both types of red clays. Culture-independent community analysis determined via barcoded pyrosequencing indicated that Nocardioidaceae, Xanthomonadaceae, Pseudomonadaceae, and Caulobacteraceae were enriched by diesel contamination. Bacterial strain isolation from naphthalene- and liquid paraffin-amended media was affiliated with enriched taxa based on 16S rRNA gene sequence identity. We suggest that the biostimulating mechanism of red clay and processed red clay is able to support bacterial growth without apparent selection for specific bacterial species.

  3. Characterization of rhamnolipids by liquid chromatography/mass spectrometry after solid-phase extraction.

    Science.gov (United States)

    Behrens, Beate; Engelen, Jeannine; Tiso, Till; Blank, Lars Mathias; Hayen, Heiko

    2016-04-01

    Rhamnolipids are surface-active agents with a broad application potential that are produced in complex mixtures by bacteria of the genus Pseudomonas. Analysis from fermentation broth is often characterized by laborious sample preparation and requires hyphenated analytical techniques like liquid chromatography coupled to mass spectrometry (LC-MS) to obtain detailed information about sample composition. In this study, an analytical procedure based on chromatographic method development and characterization of rhamnolipid sample material by LC-MS as well as a comparison of two sample preparation methods, i.e., liquid-liquid extraction and solid-phase extraction, is presented. Efficient separation was achieved under reversed-phase conditions using a mixed propylphenyl and octadecylsilyl-modified silica gel stationary phase. LC-MS/MS analysis of a supernatant from Pseudomonas putida strain KT2440 pVLT33_rhlABC grown on glucose as sole carbon source and purified by solid-phase extraction revealed a total of 20 congeners of di-rhamnolipids, mono-rhamnolipids, and their biosynthetic precursors 3-(3-hydroxyalkanoyloxy)alkanoic acids (HAAs) with different carbon chain lengths from C8 to C14, including three rhamnolipids with uncommon C9 and C11 fatty acid residues. LC-MS and the orcinol assay were used to evaluate the developed solid-phase extraction method in comparison with the established liquid-liquid extraction. Solid-phase extraction exhibited higher yields and reproducibility as well as lower experimental effort.

  4. APPLICATION OF PSEUDOMONAS PUTIDA AND RHODOCOCCUS SP. BY BIODEGRADATION OF PAH(S, PCB(S AND NEL SOIL SAMPLES FROM THE HAZARDOUS WASTE DUMP IN POZĎÁTKY (CZECH REPUBLIC

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    Radmila Kucerova

    2006-12-01

    Full Text Available The objective of the project was a laboratory check of biodegradation of soil samples contaminated by PAH(s, PCB(s and NEL from the hazardous waste dump in the Pozďátky locality. For the laboratory check, pure bacterial cultures of Rhodococcus sp. and Pseudomonas putida have been used. It is apparent from the laboratory experiments results that after one-month bacterial leaching, applying the bacterium of Rhodococcus sp. there is a 83 % removal of NEL, a 79 % removal of PAH(s and a 14 % removal of PCB(s. Applying a pure culture of Pseudomonas putida there is a 87 % removal of NEL, a 81 % removal of PAH(s and a 14 % removal of PCB(s.

  5. Oligomerization mechanisms of an H-NS family protein, Pmr, encoded on the plasmid pCAR1 provide a molecular basis for functions of H-NS family members.

    Science.gov (United States)

    Suzuki, Chiho; Kawazuma, Kohei; Horita, Shoichiro; Terada, Tohru; Tanokura, Masaru; Okada, Kazunori; Yamane, Hisakazu; Nojiri, Hideaki

    2014-01-01

    Enterobacterial H-NS-like proteins and Pseudomonas MvaT-like proteins share low homology at the amino acid sequence level, but both can function as xenogeneic silencers and are included in the H-NS family of proteins. H-NS family members have dimerization/oligomerization and DNA-binding domains connected by a flexible linker and form large nucleoprotein complexes using both domains. Pmr, an MvaT-like protein encoded on the IncP-7 carbazole-degradative plasmid pCAR1, is a key regulator of an interaction between pCAR1 and its host Pseudomonas putida KT2440. KT2440 has two transcribed genes that encode the MvaT-like proteins TurA and TurB. Our previous transcriptome analyses suggested that the functions of Pmr, TurA and TurB are non-equivalent, although the detailed underlying mechanisms remain unclear. In this study, we focused on the protein-protein interactions of Pmr, and assessed the homo-oligomerization capacity of various substituted and truncated Pmr derivatives by protein-protein cross-linking analysis. Six of the seven residues identified as important for homo-oligomerization in Pmr were located near the N-terminus, and the putative flexible linker or the region near that was not involved in homo-oligomerization, suggesting that Pmr homo-oligomerization is different from that of enterobacterial H-NS and that the functional mechanism differs between H-NS-like and MvaT-like proteins. In addition, we assessed homo- and hetero-oligomerization of Pmr by surface plasmon resonance analysis and found that the coupling ratio of TurB-Pmr oligomers is smaller than that of Pmr-Pmr or TurA-Pmr oligomers. These results raised the possibility that composition of the hetero-oligomers of Pmr, TurA, and TurB could explain why the different gene sets were affected by either pmr, turA, or turB disruption in our previous studies.

  6. The Potential of the Ni-Resistant TCE-Degrading Pseudomonas putida W619-TCE to Reduce Phytotoxicity and Improve Phytoremediation Efficiency of Poplar Cuttings on A Ni-TCE Co-Contamination.

    Science.gov (United States)

    Weyens, Nele; Beckers, Bram; Schellingen, Kerim; Ceulemans, Reinhart; van der Lelie, Daniel; Newman, Lee; Taghavi, Safiyh; Carleer, Robert; Vangronsveld, Jaco

    2015-01-01

    To examine the potential of Pseudomonas putida W619-TCE to improve phytoremediation of Ni-TCE co-contamination, the effects of inoculation of a Ni-resistant, TCE-degrading root endophyte on Ni-TCE phytotoxicity, Ni uptake and trichloroethylene (TCE) degradation of Ni-TCE-exposed poplar cuttings are evaluated. After inoculation with P. putida W619-TCE, root weight of non-exposed poplar cuttings significantly increased. Further, inoculation induced a mitigation of the Ni-TCE phytotoxicity, which was illustrated by a diminished exposure-induced increase in activity of antioxidative enzymes. Considering phytoremediation efficiency, inoculation with P. putida W619-TCE resulted in a 45% increased Ni uptake in roots as well as a slightly significant reduction in TCE concentration in leaves and TCE evapotranspiration to the atmosphere. These results indicate that endophytes equipped with the appropriate characteristics can assist their host plant to deal with co-contamination of toxic metals and organic contaminants during phytoremediation. Furthermore, as poplar is an excellent plant for biomass production as well as for phytoremediation, the obtained results can be exploited to produce biomass for energy and industrial feedstock applications in a highly productive manner on contaminated land that is not suited for normal agriculture. Exploiting this land for biomass production could contribute to diminish the conflict between food and bioenergy production.

  7. Metabolic Syndrome

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    Sevil Ikinci

    2010-10-01

    Full Text Available Metabolic Syndrome is a combination of risk factors including common etiopathogenesis. These risk factors play different roles in occurence of atherosclerotic diseases, type 2 diabetes, and cancers. Although a compromise can not be achieved on differential diagnosis for MS, the existence of any three criterias enable to diagnose MS. These are abdominal obesity, dislipidemia (hypertrigliceridemia, hypercholesterolemia, and reduced high density lipoprotein hypertension, and elevated fasting blood glucose. According to the results of Metabolic Syndrome Research (METSAR, the overall prevalence of MS in Turkey is 34%; in females 40%, and in males it is 28%. As a result of “Western” diet, and increased frequency of obesity, MS is observed in children and in adolescents both in the world and in Turkey. Resulting in chronic diseases, it is thought that the syndrome can be prevented by healthy lifestyle behaviours. [TAF Prev Med Bull 2010; 9(5.000: 535-540

  8. What is Metabolic Syndrome?

    Science.gov (United States)

    ... from the NHLBI on Twitter. What Is Metabolic Syndrome? Metabolic syndrome is the name for a group of ... that may play a role in causing metabolic syndrome. Outlook Metabolic syndrome is becoming more common due to a ...

  9. Pentachlorophenol dechlorination and simultaneous Cr6+ reduction by Pseudomonas putida SKG-1 MTCC (10510): characterization of PCP dechlorination products, bacterial structure, and functional groups.

    Science.gov (United States)

    Garg, Satyendra Kumar; Tripathi, Manikant; Singh, Santosh Kumar; Singh, Anamika

    2013-04-01

    It is the first report in which a novel psychrotrophic Pseudomonas putida SKG-1 strain was evaluated for simultaneous bioremediation of pentachlorophenol and Cr(6+) under various cultural and nutritional conditions. Pentachlorophenol (PCP) dechlorination products, bacterial structure, and functional groups were characterized by gas chromatography and mass spectrometry (GC-MS), scanning electron microscope and energy dispersive X-ray spectroscopy (SEM-EDS), and Fourier-transform infrared (FTIR) techniques. The strain was extremely tolerant to excessively higher individual concentration of PCP (1,400 mg l(-1)) and Cr(6+) (4,300 mg l(-1)). Increasing concentration of PCP and Cr(6+) exerted inhibitory effect on bacterial growth and toxicants' removal. The strain exhibited growth, and concomitantly remediated both the pollutants simultaneously over a broad pH (7.0-9.0) and temperature (28-32 °C) range; maximum growth, PCP dechlorination (87.5%), and Cr(6+) removal (80.0%) occurred at optimum pH 8.0 and 30 °C (from initial PCP 100 mg l(-1) and Cr(6+) 500 mg l(-1)) under shaking (150 rpm) within 72 h incubation. Optimization of agitation (125 rpm) and aeration (0.4 vvm) in bioreactor further enhanced PCP dechlorination by ~10% and Cr(6+) removal by 2%. A direct correlation existed between growth and bioremediation of both the toxicants. Among other heavy metals, mercury exerted maximum and cobalt minimum inhibitory effect on PCP dechlorination and Cr(6+) removal. Chromate reductase activity was mainly associated with the supernatant and cytosolic fraction of bacterial cells. GC-MS analysis revealed the formation of tetrachloro-p-hydroquinone, 2,4,6-trichlorophenol, and 2,6-dichlorophenol as PCP dechlorination products. FTIR spectrometry indicated likely involvement of carbonyl and amide groups in Cr(3+) adsorption, and SEM-EDS showed the presence of chromium on P. putida surface. Thus, our promising isolate can be ecofriendly employed for biotreatment of various

  10. The catabolism of 2,4-xylenol and p-cresol share the enzymes for the oxidation of para-methyl group in Pseudomonas putida NCIMB 9866.

    Science.gov (United States)

    Chen, Yan-Fei; Chao, Hongjun; Zhou, Ning-Yi

    2014-02-01

    Pseudomonas putida NCIMB 9866 utilizes p-cresol or 2,4-xylenol as a sole carbon and energy source. Enzymes catalyzing the oxidation of the para-methyl group of p-cresol have been studied in detail. However, those responsible for the oxidation of the para-methyl group in 2,4-xylenol catabolism are still not reported. In this study, real-time quantitative PCR analysis indicated pchC- and pchF-encoded p-cresol methylhydroxylase (PCMH) and pchA-encoded p-hydroxybenzaldehyde dehydrogenase (PHBDD) in p-cresol catabolism were also likely involved in the catabolism of 2,4-xylenol. Enzyme activity assays and intermediate identification indicated that the PCMH and PHBDD catalyzed the oxidations of 2,4-xylenol to 4-hydroxy-3-methylbenzaldehyde and 4-hydroxy-3-methylbenzaldehyde to 4-hydroxy-3-methylbenzoic acid, respectively. Furthermore, the PCMH-encoding gene pchF was found to be necessary for the catabolism of 2,4-xylenol, whereas the PHBDD-encoding gene pchA was not essential for the catabolism by gene knockout and complementation. Analyses of the maximum specific growth rate (μ m) and specific activity of the gene-knockout strain to different intermediates revealed the presence of other enzyme(s) with PHBDD activity in strain 9866. However, PHBDD played a major role in the catabolism of 2,4-xylenol in contrast to the other enzyme(s).

  11. Isolation of oxygenase genes for indigo-forming activity from an artificially polluted soil metagenome by functional screening using Pseudomonas putida strains as hosts.

    Science.gov (United States)

    Nagayama, Hirofumi; Sugawara, Tomonori; Endo, Ryo; Ono, Akira; Kato, Hiromi; Ohtsubo, Yoshiyuki; Nagata, Yuji; Tsuda, Masataka

    2015-05-01

    Metagenomes contain the DNA from many microorganisms, both culturable and non-culturable, and are a potential resource of novel genes. In this study, a 5.2-Gb metagenomic DNA library was constructed from a soil sample (artificially polluted with four aromatic compounds, i.e., biphenyl, phenanthrene, carbazole, and 3-chlorobenzoate) in Escherichia coli by using a broad-host-range cosmid vector. The resultant library was introduced into naphthalene-degrading Pseudomonas putida-derived strains having deficiencies in their naphthalene dioxygenase components, and indigo-forming clones on the indole-containing agar plates were screened. Cosmids isolated from 29 positive clones were classified by their various properties (original screening hosts, hosts showing indigo-forming activity, and digestion patterns with restriction enzymes), and six representative cosmids were chosen. Sequencing and in vitro transposon mutagenesis of the six cosmids resulted in the identification of genes encoding putative class B and D flavoprotein monooxygenases, a multicomponent hydroxylase, and a reductase that were responsible for the indigo-forming activity in the host cells. Among them, the genes encoding the multicomponent hydroxylase were demonstrated to be involved in phenol degradation. Furthermore, two genes encoding ring-cleavage dioxygenases were also found adjacent to the genes responsible for the indigo formation, and their functions were experimentally confirmed.

  12. Decaffeination and measurement of caffeine content by addicted Escherichia coli with a refactored N-demethylation operon from Pseudomonas putida CBB5.

    Science.gov (United States)

    Quandt, Erik M; Hammerling, Michael J; Summers, Ryan M; Otoupal, Peter B; Slater, Ben; Alnahhas, Razan N; Dasgupta, Aurko; Bachman, James L; Subramanian, Mani V; Barrick, Jeffrey E

    2013-06-21

    The widespread use of caffeine (1,3,7-trimethylxanthine) and other methylxanthines in beverages and pharmaceuticals has led to significant environmental pollution. We have developed a portable caffeine degradation operon by refactoring the alkylxanthine degradation (Alx) gene cluster from Pseudomonas putida CBB5 to function in Escherichia coli. In the process, we discovered that adding a glutathione S-transferase from Janthinobacterium sp. Marseille was necessary to achieve N 7 -demethylation activity. E. coli cells with the synthetic operon degrade caffeine to the guanine precursor, xanthine. Cells deficient in de novo guanine biosynthesis that contain the refactored operon are ″addicted″ to caffeine: their growth density is limited by the availability of caffeine or other xanthines. We show that the addicted strain can be used as a biosensor to measure the caffeine content of common beverages. The synthetic N-demethylation operon could be useful for reclaiming nutrient-rich byproducts of coffee bean processing and for the cost-effective bioproduction of methylxanthine drugs.

  13. Hydrolysis of triglyceride by the whole cell of Pseudomonas putida 3SK in two-phase batch and continuous reactors systems.

    Science.gov (United States)

    Lee, S Y; Rhee, J S

    1994-08-05

    Batch and continuous hydrolysis of olive oil in an organic-aqueous two-phase system using the live whole cell of Pseudomonas putida 3SK as a source of a lipase is investigated. The strain was not only fully viable and grown well, but also produced extracellular lipase simultaneously. The degree of hydrolysis, depending on olive oil concentration in the solvents, was maximal at 13.5% (w/v) and decreased with the increase of the substrate concentration. At the optimal condition, a degree of hydrolysis higher than 95% was achieved with 24 h at 30 degrees C when the reaction was carried out in a two-phase batch stirred reactor. For long-term operation a continuous stirred reactor was designed. When the reaction was carried out in a continuous stirred reactor, the degree was hydrolysis reached 86% at a dilution rate of 0.2 h(-1). Satisfactory performance of a two-phase bioreactor was obtained in a long-term continous operation, which lasted for at least 30 days by feeding organic solvent containing olive oil and aqueous media separately. (c) 1994 John Wiley & Sons, Inc.

  14. Heterofunctional Magnetic Metal-Chelate-Epoxy Supports for the Purification and Covalent Immobilization of Benzoylformate Decarboxylase From Pseudomonas Putida and Its Carboligation Reactivity.

    Science.gov (United States)

    Tural, Servet; Tural, Bilsen; Demir, Ayhan S

    2015-09-01

    In this study, the combined use of the selectivity of metal chelate affinity chromatography with the capacity of epoxy supports to immobilize poly-His-tagged recombinant benzoylformate decarboxylase from Pseudomonas putida (BFD, E.C. 4.1.1.7) via covalent attachment is shown. This was achieved by designing tailor-made magnetic chelate-epoxy supports. In order to selectively adsorb and then covalently immobilize the poly-His-tagged BFD, the epoxy groups (300 µmol epoxy groups/g support) and a very small density of Co(2+)-chelate groups (38 µmol Co(2+)/g support) was introduced onto magnetic supports. That is, it was possible to accomplish, in a simple manner, the purification and covalent immobilization of a histidine-tagged recombinant BFD. The magnetically responsive biocatalyst was tested to catalyze the carboligation reactions. The benzoin condensation reactions were performed with this simple and convenient heterogeneous biocatalyst and were comparable to that of a free-enzyme-catalyzed reaction. The enantiomeric excess (ee) of (R)-benzoin was obtained at 99 ± 2% for the free enzyme and 96 ± 3% for the immobilized enzyme. To test the stability of the covalently immobilized enzyme, the immobilized enzyme was reused in five reaction cycles for the formation of chiral 2-hydroxypropiophenone (2-HPP) from benzaldehyde and acetaldehyde, and it retained 96% of its original activity after five reaction cycles.

  15. A novel Pseudomonas aeruginosa bacteriophage, Ab31, a chimera formed from temperate phage PAJU2 and P. putida lytic phage AF: characteristics and mechanism of bacterial resistance.

    Directory of Open Access Journals (Sweden)

    Libera Latino

    Full Text Available A novel temperate bacteriophage of Pseudomonas aeruginosa, phage vB_PaeP_Tr60_Ab31 (alias Ab31 is described. Its genome is composed of structural genes related to those of lytic P. putida phage AF, and regulatory genes similar to those of temperate phage PAJU2. The virion structure resembles that of phage AF and other lytic Podoviridae (S. enterica Epsilon 15 and E. coli phiv10 with similar tail spikes. Ab31 was able to infect P. aeruginosa strain PA14 and two genetically related strains called Tr60 and Tr162, out of 35 diverse strains from cystic fibrosis patients. Analysis of resistant host variants revealed different phenotypes, including induction of pigment and alginate overproduction. Whole genome sequencing of resistant variants highlighted the existence of a large deletion of 234 kbp in two strains, encompassing a cluster of genes required for the production of CupA fimbriae. Stable lysogens formed by Ab31 in strain Tr60, permitted the identification of the insertion site. During colonization of the lung in cystic fibrosis patients, P. aeruginosa adapts by modifying its genome. We suggest that bacteriophages such as Ab31 may play an important role in this adaptation by selecting for bacterial characteristics that favor persistence of bacteria in the lung.

  16. ISOLATION AND CHARACTERIZATION OF A MOLYBDENUM-REDUCING, PHENOL- AND CATECHOL-DEGRADING PSEUDOMONAS PUTIDA STRAIN AMR-12 IN SOILS FROM EGYPT

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    M. Abd. AbdEl-Mongy

    2016-02-01

    Full Text Available Sites contaminated with both heavy metals and organic xenobiotic pollutants warrants the effective use of either a multitude of bacterial degraders or bacteria having the capacity to detoxify numerous toxicants simultaneously. A molybdenum-reducing bacterium with the capacity to degrade phenolics is reported. Molybdenum (sodium molybdate reduction was optimum between pH 6.0 and 7.0 and between 20 and 30 °C. The most suitable electron donor was glucose. A narrow range of phosphate concentrations between 5.0 and 7.5 mM was required for optimal reduction, while molybdate between 20 and 30 mM were needed for optimal reduction. The scanning absorption spectrum of the molybdenum blue produced indicated that Mo-blue is a reduced phosphomolybdate. Molybdenum reduction was inhibited by the heavy metals mercury, silver and chromium. Biochemical analysis identified the bacterium as Pseudomonas putida strain Amr-12. Phenol and phenolics cannot support molybdenum reduction. However, the bacterium was able to grow on the phenolic compounds (phenol and catechol with observable lag periods. Maximum growth on phenol and catechol occurred around the concentrations of 600 mg∙L-1. The ability of this bacterium to detoxify molybdenum and grown on toxic phenolic makes this bacterium an important tool for bioremediation.

  17. Alkanols and chlorophenols cause different physiological adaptive responses on the level of cell surface properties and membrane vesicle formation in Pseudomonas putida DOT-T1E.

    Science.gov (United States)

    Baumgarten, Thomas; Vazquez, José; Bastisch, Christian; Veron, Wilfried; Feuilloley, Marc G J; Nietzsche, Sandor; Wick, Lukas Y; Heipieper, Hermann J

    2012-01-01

    In order to cope with the toxicity imposed by the exposure to environmental hydrocarbons, many bacteria have developed specific adaptive responses such as modifications in the cell envelope. Here we compared the influence of n-alkanols and chlorophenols on the surface properties of the solvent-tolerant bacterium Pseudomonas putida DOT-T1E. In the presence of toxic concentrations of n-alkanols, this strain significantly increased its cell surface charge and hydrophobicity with changes depending on the chain length of the added n-alkanols. The adaptive response occurred within 10 min after the addition of the solvent and was demonstrated to be of physiological nature. Contrary to that, chlorophenols of similar hydrophobicity and potential toxicity as the corresponding alkanols caused only minor effects in the surface properties. To our knowledge, this is the first observation of differences in the cellular adaptive response of bacteria to compound classes of quasi equal hydrophobicity and toxicity. The observed adaptation of the physico-chemical surface properties of strain DOT-T1E to the presence of alkanols was reversible and correlated with changes in the composition of the lipopolysaccharide content of the cells. The reaction is explained by previously described reactions allowing the release of membrane vesicles that was demonstrated for cells affected by 1-octanol and heat shock, whereas no membrane vesicles were released after the addition of chlorophenols.

  18. Application of the freeze-dried bioluminescent bioreporter Pseudomonas putida mt-2 KG1206 to the biomonitoring of groundwater samples from monitoring wells near gasoline leakage sites.

    Science.gov (United States)

    Ko, Kyung-Seok; Kong, In Chul

    2017-02-01

    This study examined the applicability of a freeze-dried bioluminescent bioreporter, Pseudomonas putida mt-2 KG1206 (called KG1206), to the biomonitoring of groundwater samples. Samples were collected from the monitoring wells of gas station tanks or old pipeline leakage sites in Korea. In general, the freeze-dried strain in the presence of pure inducer chemicals showed low bioluminescence activity and a different activity order compared with that of the subcultured strain. The effects of KNO3 as a bioluminescence stimulant were observed on the pure inducers and groundwater samples. The stimulation rates varied according to the type of inducers and samples, ranging from 2.2 to 20.5 times (for pure inducers) and from 1.1 to 11 times (for groundwater samples) the total bioluminescence of the control. No considerable correlations were observed between the bioluminescence intensity of the freeze-dried strain and the inducer concentrations in the samples (R (2) < 0.1344). However, samples without a high methyl tertiary butyl ether (MTBE) level and those from the gas station leakage site showed reasonable correlations with the bioluminescence activity with R (2) values of 0.3551 and 0.4131, respectively. These results highlight the potential of using freeze-dried bioluminescent bacteria as a rapid, simple, and portable tool for the preliminary biomonitoring of specific pollutants at contaminated sites.

  19. Biodegradation of benzene, toluene, ethylbenzene, and o-xylene by a coculture of Pseudomonas putida and Pseudomonas fluorescens immobilized in a fibrous-bed bioreactor.

    Science.gov (United States)

    Shim, H; Yang, S T

    1999-01-22

    A fibrous-bed bioreactor containing the coculture of Pseudomonas putida and P. fluorescens immobilized in a fibrous matrix was developed to degrade benzene (B), toluene (T), ethylbenzene (E), and o-xylene (X) in synthetic waste streams. The kinetics of BTEX biodegradation by immobilized cells adapted in the fibrous-bed bioreactor and free cells grown in serum bottles were studied. In general, the BTEX biodegradation rate increased with increasing substrate concentration and then decreased after reaching a maximum, showing substrate-inhibition kinetics. However, for immobilized cells, the degradation rate was much higher than that of free cells. Compared to free cells, immobilized cells in the bioreactor tolerated higher concentrations (> 1000 mg l-1) of benzene and toluene, and gave at least 16-fold higher degradation rates for benzene, ethylbenzene, and o-xylene, and a 9-fold higher degradation rate for toluene. Complete and simultaneous degradation of BTEX mixture was achieved in the bioreactor under hypoxic conditions. Cells in the bioreactor were relatively insensitive to benzene toxicity; this insensitivity was attributed to adaptation of the cells in the bioreactor. Compared to the original seeding culture, the adapted cells from the fibrous-bed bioreactor had higher specific growth rate, benzene degradation rate, and cell yield when the benzene concentration was higher than 100 mg l-1. Cells in the fibrous bed had a long, slim morphology, which is different from the normal short-rod shape found for suspended cells in solution.

  20. Interactive effects of phosphorus and Pseudomonas putida on chickpea (Cicer arietinum L.) growth, nutrient uptake, antioxidant enzymes and organic acids exudation.

    Science.gov (United States)

    Israr, Dania; Mustafa, Ghulam; Khan, Khalid Saifullah; Shahzad, Muhammad; Ahmad, Niaz; Masood, Sajid

    2016-11-01

    Phosphorus (P) availability in alkaline soils of arid and semi-arid regions is a major constraint for decreased crop productivity. Use of plant growth promoting rhizobacteria (PGPR) may enhance plant growth through the increased plant antioxidation activity. Additionally, PGPR may increase nutrient uptake by plants as a result of induced root exudation and rhizosphere acidification. The current study was aimed to investigate combined effects of P and Pesudomonas putida (PGPR) on chickpea growth with reference to antioxidative enzymatic activity and root exudation mediated plant nutrient uptake, particularly P. Half of the seeds were soaked in PGPR solution, whereas others in sterile water and latter sown in soils. Plants were harvested 8 weeks after onset of experiment and analyzed for leaf nutrient contents, antioxidant enzymes activities and organic acids concentrations. Without PGPR, P application (+P) increased various plant growth attributes, plant uptake of P and Ca, soil pH, citric acid and oxalic acid concentrations, whereas decreased the leaf POD enzymatic activity as compared to the P-deficiency. PGPR supply both under -P and +P improved the plant growth, plant uptake of N, P, and K, antioxidative activity of SOD and POD enzymes and concentrations of organic acids, whereas reduced the rhizosphere soil pH. Growth enhancement by PGPR supply was related to higher plant antioxidation activity as well as nutrient uptake of chickpea including P as a result of root exudation mediated rhizosphere acidification. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  1. The regulatory logic of m-xylene biodegradation by Pseudomonas putida mt-2 exposed by dynamic modelling of the principal node Ps/Pr of the TOL plasmid.

    Science.gov (United States)

    Koutinas, Michalis; Lam, Ming-Chi; Kiparissides, Alexandros; Silva-Rocha, Rafael; Godinho, Miguel; Livingston, Andrew G; Pistikopoulos, Efstratios N; de Lorenzo, Victor; Dos Santos, Vitor A P Martins; Mantalaris, Athanasios

    2010-06-01

    The structure of the extant transcriptional control network of the TOL plasmid pWW0 born by Pseudomonas putida mt-2 for biodegradation of m-xylene is far more complex than one would consider necessary from a mere engineering point of view. In order to penetrate the underlying logic of such a network, which controls a major environmental cleanup bioprocess, we have developed a dynamic model of the key regulatory node formed by the Ps/Pr promoters of pWW0, where the clustering of control elements is maximal. The model layout was validated with batch cultures estimating parameter values and its predictive capability was confirmed with independent sets of experimental data. The model revealed how regulatory outputs originated in the divergent and overlapping Ps/Pr segment, which expresses the transcription factors XylS and XylR respectively, are computed into distinct instructions to the upper and lower catabolic xyl operons for either simultaneous or stepwise consumption of m-xylene and/or succinate. In this respect, the model reveals that the architecture of the Ps/Pr is poised to discriminate the abundance of alternative and competing C sources, in particular m-xylene versus succinate. The proposed framework provides a first systemic understanding of the causality and connectivity of the regulatory elements that shape this exemplary regulatory network, facilitating the use of model analysis towards genetic circuit optimization.

  2. Influence of pH on dynamics of microbial enhanced oil recovery processes using biosurfactant producing Pseudomonas putida: Mathematical modelling and numerical simulation.

    Science.gov (United States)

    Sivasankar, P; Suresh Kumar, G

    2017-01-01

    In present work, the influence of reservoir pH conditions on dynamics of microbial enhanced oil recovery (MEOR) processes using Pseudomonas putida was analysed numerically from the developed mathematical model for MEOR processes. Further, a new strategy to improve the MEOR performance has also been proposed. It is concluded from present study that by reversing the reservoir pH from highly acidic to low alkaline condition (pH 5-8), flow and mobility of displaced oil, displacement efficiency, and original oil in place (OOIP) recovered gets significantly enhanced, resulting from improved interfacial tension (IFT) reduction by biosurfactants. At pH 8, maximum of 26.1% of OOIP was recovered with higher displacement efficiency. The present study introduces a new strategy to increase the recovery efficiency of MEOR technique by characterizing the biosurfactants for IFTmin/IFTmax values for different pH conditions and subsequently, reversing the reservoir pH conditions at which the IFTmin/IFTmax value is minimum. Copyright © 2016 Elsevier Ltd. All rights reserved.

  3. Numerical modelling of biophysicochemical effects on multispecies reactive transport in porous media involving Pseudomonas putida for potential microbial enhanced oil recovery application.

    Science.gov (United States)

    Sivasankar, P; Rajesh Kanna, A; Suresh Kumar, G; Gummadi, Sathyanarayana N

    2016-07-01

    pH and resident time of injected slug plays a critical role in characterizing the reservoir for potential microbial enhanced oil recovery (MEOR) application. To investigate MEOR processes, a multispecies (microbes-nutrients) reactive transport model in porous media was developed by coupling kinetic and transport model. The present work differs from earlier works by explicitly determining parametric values required for kinetic model by experimental investigations using Pseudomonas putida at different pH conditions and subsequently performing sensitivity analysis of pH, resident time and water saturation on concentrations of microbes, nutrients and biosurfactant within reservoir. The results suggest that nutrient utilization and biosurfactant production are found to be maximum at pH 8 and 7.5 respectively. It is also found that the sucrose and biosurfactant concentrations are highly sensitive to pH rather than reservoir microbial concentration, while at larger resident time and water saturation, the microbial and nutrient concentrations were lesser due to enhanced dispersion. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. Dynamic Response of Pseudomonas putida S12 to Sudden Addition of Toluene and the Potential Role of the Solvent Tolerance Gene trgI.

    Science.gov (United States)

    Volkers, Rita J M; Snoek, L Basten; Ruijssenaars, Harald J; de Winde, Johannes H

    2015-01-01

    Pseudomonas putida S12 is exceptionally tolerant to various organic solvents. To obtain further insight into this bacterium's primary defence mechanisms towards these potentially harmful substances, we studied its genome wide transcriptional response to sudden addition of toluene. Global gene expression profiles were monitored for 30 minutes after toluene addition. During toluene exposure, high oxygen-affinity cytochrome c oxidase is specifically expressed to provide for an adequate proton gradient supporting solvent efflux mechanisms. Concomitantly, the glyoxylate bypass route was up-regulated, to repair an apparent toluene stress-induced redox imbalance. A knock-out mutant of trgI, a recently identified toluene-repressed gene, was investigated in order to identify TrgI function. Remarkably, upon addition of toluene the number of differentially expressed genes initially was much lower in the trgI-mutant than in the wild-type strain. This suggested that after deletion of trgI cells were better prepared for sudden organic solvent stress. Before, as well as after, addition of toluene many genes of highly diverse functions were differentially expressed in trgI-mutant cells as compared to wild-type cells. This led to the hypothesis that TrgI may not only be involved in the modulation of solvent-elicited responses but in addition may affect basal expression levels of large groups of genes.

  5. The impact of succinate trace on pWW0 and ortho-cleavage pathway transcription in Pseudomonas putida mt-2 during toluene biodegradation.

    Science.gov (United States)

    Tsipa, Argyro; Koutinas, Michalis; Vernardis, Spyros I; Mantalaris, Athanasios

    2017-06-01

    Toluene is a pollutant catabolised through the interconnected pWW0 (TOL) and ortho-cleavage pathways of Pseudomonas putida mt-2, while upon succinate and toluene mixtures introduction in batch cultures grown on M9 medium, succinate was previously reported as non-repressing. The effect of a 40 times lower succinate concentration, as compared to literature values, was explored through systematic real-time qPCR monitoring of transcriptional kinetics of the key TOL Pu, Pm and ortho-cleavage PbenR, PbenA promoters in mixed-substrate experiments. Even succinate trace inhibited transcription leading to bi-modal promoters expression. Potential carbon catabolite repression mechanisms and novel expression patterns of promoters were unfolded. Lag phase was shortened and biomass growth levels increased compared to sole toluene biodegradation suggesting enhanced pollutant removal efficiency. The study stressed the noticeable effect of a preferred compound's left-over on the main route of a bioprocess, revealing the beneficiary supply of low preferred substrates concentrations to design optimal bioremediation strategies. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. 3-Chloro-1,2-propanediol biodegradation by Ca-alginate immobilized Pseudomonas putida DSM 437 cells applying different processes: mass transfer effects.

    Science.gov (United States)

    Konti, Aikaterini; Mamma, Diomi; Hatzinikolaou, Dimitios G; Kekos, Dimitris

    2016-10-01

    3-Chloro-1,2-propanediol (3-CPD) biodegradation by Ca-alginate immobilized Pseudomonas putida cells was performed in batch system, continuous stirred tank reactor (CSTR), and packed-bed reactor (PBR). Batch system exhibited higher biodegradation rates and 3-CPD uptakes compared to CSTR and PBR. The two continuous systems (CSTR and PBR) when compared at 200 mg/L 3-CPD in the inlet exhibited the same removal of 3-CPD at steady state. External mass-transfer limitations are found negligible at all systems examined, since the observable modulus for external mass transfer Ω ≪ 1 and the Biot number Bi > 1. Intra-particle diffusion resistance had a significant effect on 3-CPD biodegradation in all systems studied, but to a different extent. Thiele modulus was in the range of 2.5 in batch system, but it was increased at 11 when increasing cell loading in the beads, thus lowering significantly the respective effectiveness factor. Comparing the systems at the same cell loading in the beads PBR was less affected by internal diffusional limitations compared to CSTR and batch system, and, as a result, exhibited the highest overall effectiveness factor.

  7. The oxygenating constituent of 3,6-diketocamphane monooxygenase from the CAM plasmid of Pseudomonas putida: the first crystal structure of a type II Baeyer–Villiger monooxygenase

    Energy Technology Data Exchange (ETDEWEB)

    Isupov, Michail N.; Schröder, Ewald; Gibson, Robert P.; Beecher, Jean; Donadio, Giuliana; Saneei, Vahid; Dcunha, Stephlina A.; McGhie, Emma J.; Sayer, Christopher; Davenport, Colin F. [University of Exeter, Stocker Road, Exeter EX4 4QD (United Kingdom); Lau, Peter C. [National Research Council Canada, 6100 Royalmount Avenue, Montreal, QC H4P 2R2 (Canada); Hasegawa, Yoshie; Iwaki, Hiroaki [Kansai University (Japan); Kadow, Maria; Balke, Kathleen; Bornscheuer, Uwe T. [Greifswald University, Felix-Hausdorff-Strasse 4, 17487 Greifswald (Germany); Bourenkov, Gleb [European Molecular Biology Laboratory (EMBL), Hamburg Outstation, Notkestrasse 85, 22607 Hamburg (Germany); Littlechild, Jennifer A., E-mail: j.a.littlechild@exeter.ac.uk [University of Exeter, Stocker Road, Exeter EX4 4QD (United Kingdom)

    2015-10-31

    The first crystal structure of a type II Baeyer–Villiger monooxygenase reveals a different ring orientation of its FMN cofactor compared with other related bacterial luciferase-family enzymes. The three-dimensional structures of the native enzyme and the FMN complex of the overexpressed form of the oxygenating component of the type II Baeyer–Villiger 3,6-diketocamphane monooxygenase have been determined to 1.9 Å resolution. The structure of this dimeric FMN-dependent enzyme, which is encoded on the large CAM plasmid of Pseudomonas putida, has been solved by a combination of multiple anomalous dispersion from a bromine crystal soak and molecular replacement using a bacterial luciferase model. The orientation of the isoalloxazine ring of the FMN cofactor in the active site of this TIM-barrel fold enzyme differs significantly from that previously observed in enzymes of the bacterial luciferase-like superfamily. The Ala77 residue is in a cis conformation and forms a β-bulge at the C-terminus of β-strand 3, which is a feature observed in many proteins of this superfamily.

  8. Identification of anthranilate and benzoate metabolic operons of Pseudomonas fluorescens and functional characterization of their promoter regions

    Directory of Open Access Journals (Sweden)

    Lee Vincent D

    2006-01-01

    Full Text Available Abstract Background In an effort to identify alternate recombinant gene expression systems in Pseudomonas fluorescens, we identified genes encoding two native metabolic pathways that were inducible with inexpensive compounds: the anthranilate operon (antABC and the benzoate operon (benABCD. Results The antABC and benABCD operons were identified by homology to the Acinetobacter sp. anthranilate operon and Pseudomonas putida benzoate operon, and were confirmed to be regulated by anthranilate or benzoate, respectively. Fusions of the putative promoter regions to the E. coli lacZ gene were constructed to confirm inducible gene expression. Each operon was found to be controlled by an AraC family transcriptional activator, located immediately upstream of the first structural gene in each respective operon (antR or benR. Conclusion We have found the anthranilate and benzoate promoters to be useful for tightly controlling recombinant gene expression at both small (

  9. Structure of a putative BenF-like porin from Pseudomonas fluorescens Pf-5 at 2.6 A resolution

    Energy Technology Data Exchange (ETDEWEB)

    Sampathkumar, P.; Swaminathan, S.; Lu, F.; Zhao, X.; Li, Z.; Gilmore, J.; Bain, K.; Rutter, M. E.; Gheyi, T.; Schwinn, D.; Bonanno, J. B.; Pieper, U.; Fajardo, J. E.; Fiser, A.; Almo, S. C.; Chance, M. R.; Baker, D.; Atwell, S.; Thompson, D. A.; Emtage, J. S.; Wasserman, S. R.; Sali, A.; Sauder, J. M.; Burley, S. K.

    2010-11-01

    of benzoate. BLAST analysis of the amino acid sequence of P. fluorescens Pf-5 gene PFL1329 (Uniprot id: http://www.uniprot.org/uniprot/Q4KH25) against P. putida KT2440 strain identified 20 related porins. The top six hits include P. putida KT2440 genes PP1383 (annotated as BenF-like), PP2517 (annotated as BenF-like), and PP3168 (annotated as BenF), which share sequence identities of 76%, 66%, and 44% with PFL1329, respectively. The precise functions of these genes are not yet known. Therefore, we refer to the protein product of gene PFL1329 as PflBenF-like, which reflects its current annotation in the Uniprot database. Crystal structures of OprD and OpdK (vanillate specific porin), both from P. aeruginosa (designated below as PaOprD and PaOpdK, respectively) have been determined. Herein, we report the crystal structure of a putative BenF-like porin from P. fluorescens Pf-5 (PflBenF-like). For the sake of brevity, all subsequent references to the PflBenF-like porin will be made using PflBenF. X-ray crystallography revealed a canonical 18-stranded {beta}-barrel fold that forms a central pore with a diameter of {approx}4.6 E. We describe detailed comparisons of the PflBenF structure with those of PaOprD and PaOpdK.

  10. Carbohydrate Metabolism Disorders

    Science.gov (United States)

    ... you eat. Food is made up of proteins, carbohydrates, and fats. Chemicals in your digestive system (enzymes) ... metabolic disorder, something goes wrong with this process. Carbohydrate metabolism disorders are a group of metabolic disorders. ...

  11. Cancer stem cell metabolism

    National Research Council Canada - National Science Library

    Peiris-Pagès, Maria; Martinez-Outschoorn, Ubaldo E; Pestell, Richard G; Sotgia, Federica; Lisanti, Michael P

    2016-01-01

    .... Cancer stem cells also seem to adapt their metabolism to microenvironmental changes by conveniently shifting energy production from one pathway to another, or by acquiring intermediate metabolic phenotypes...

  12. The Complete Nucleotide Sequence of the Carbapenem Resistance-Conferring Conjugative Plasmid pLD209 from a Pseudomonas putida Clinical Strain Reveals a Chimeric Design Formed by Modules Derived from Both Environmental and Clinical Bacteria

    Science.gov (United States)

    Marchiaro, Patricia M.; Brambilla, Luciano; Morán-Barrio, Jorgelina; Revale, Santiago; Pasteran, Fernando; Vila, Alejandro J.; Viale, Alejandro M.

    2014-01-01

    The complete sequence of the carbapenem-resistance-conferring conjugative plasmid pLD209 from a Pseudomonas putida clinical strain is presented. pLD209 is formed by 3 well-defined regions: an adaptability module encompassing a Tn402-like class 1 integron of clinical origin containing blaVIM-2 and aacA4 gene cassettes, partitioning and transfer modules, and a replication module derived from plasmids of environmental bacteria. pLD209 is thus a mosaic of modules originating in both the clinical and environmental (nonclinical) microbiota. PMID:24395220

  13. Bacteria mediated dissolution of pyromorphite Pb5(PO4)3Cl in presence of Pseudomonas putida bacteria - an effect on Pb remobilization in the environment

    Science.gov (United States)

    Flis, Justyna; Manecki, Maciej; Merkel, Broder J.; Latowski, Dariusz

    2010-05-01

    The objective of the study was to determine the mechanisms of microbially enhanced dissolution of lead phosphate-pyromorphite Pb5(PO4)3Cl). Contrary to the current literature, the results of our experiments indicate a great potential for Pb remobilization in the environment by an aerobic microorganism acquiring phosphates. Broad knowledge exists about the role of Pb-apatites in regulating the behavior and the bioavailability of Pb in soils and wastewater. In situ Pb immobilization is one of the methods now routinely applied for the reclamation of Pb-contaminated soils, including shallow unconfined aquifers (Magalhaes & Silva, 2003; Magalhaes, 2002; Ma et al. 1993). This method is based on the principle that aqueous phosphates added to soil pore solutions form a very stable (insoluble) mineral pyromorphite (Pb-apatite) Pb5(PO4)3Cl. Bioavailability of aqueous Pb is thus minimized due to the very low solubility and the high thermodynamic stability of pyromorphite (Flis, 2007; Nriagu, 1974). Several reports have examined the ability of different bacterial species including Pseudomonas to solubilise insoluble inorganic phosphate compounds for example apatites (Welch et al., 2002; Maurice et al., 1999; Rodriguez and Fraga, 1999 ). Various species of Pseudomonas genera are encountered as common inhabitants of soils and wastes in the industrial areas under strong pollution influence. To date, there has not been any published evidence of microbial dissolution of pyromorphite. The major objective of the project was to study Pseudomonas putida growth in the presence of Pb-apatite (Pb5(PO4)3Cl) as the sole source of phosphate. It was to test the hypothesis that in the phosphate deficient environment bacteria are able to actively scavenge for P from the Pb-apatite which results in remobilization of Pb in the environment. The bacteria growth was investigated at 22oC. Commercially available Pseudomonas putida strain was used throughout. The experiment and its controls were run in

  14. Inactivation of Pseudomonas putida by pulsed electric field treatment: a study on the correlation of treatment parameters and inactivation efficiency in the short-pulse range.

    Science.gov (United States)

    Frey, Wolfgang; Gusbeth, Christian; Schwartz, Thomas

    2013-10-01

    An important issue for an economic application of the pulsed electric field treatment for bacterial decontamination of wastewater is the specific treatment energy needed for effective reduction of bacterial populations. The present experimental study performed in a field amplitude range of 40 > E > 200 kV/cm and for a suspension conductivity of 0.01 = κ(e) > 0.2 S/m focusses on the application of short pulses, 25 ns > T > 10 μs, of rectangular, bipolar and exponential shape and was made on Pseudomonas putida, which is a typical and widespread wastewater microorganism. The comparison of inactivation results with calculations of the temporal and azimuthal membrane charging dynamics using the model of Pauly and Schwan revealed that for efficient inactivation, membrane segments at the cell equator have to be charged quickly and to a sufficiently high value, on the order of 0.5 V. After fulfilling this basic condition by an appropriate choice of pulse field strength and duration, the log rate of inactivation for a given suspension conductivity of 0.2 S/m was found to be independent of the duration of individual pulses for constant treatment energy expenditure. Moreover, experimental results suggest that even pulse shape plays a minor role in inactivation efficiency. The variation of the suspension conductivity resulted in comparable inactivation performance of identical pulse parameters if the product of pulse duration and number of pulses was the same, i.e., required treatment energy can be linearly downscaled for lower conductivities, provided that pulse amplitude and duration are selected for entire membrane surface permeabilization.

  15. HipH Catalyzes the Hydroxylation of 4-Hydroxyisophthalate to Protocatechuate in 2,4-Xylenol Catabolism by Pseudomonas putida NCIMB 9866.

    Science.gov (United States)

    Chao, Hong-Jun; Chen, Yan-Fei; Fang, Ti; Xu, Ying; Huang, Wei E; Zhou, Ning-Yi

    2015-11-13

    In addition to growing on p-cresol, Pseudomonas putida NCIMB 9866 is the only reported strain capable of aerobically growing on 2,4-xylenol, which is listed as a priority pollutant by the U.S. Environmental Protection Agency. Several enzymes involved in the oxidation of the para-methyl group, as well as the corresponding genes, have previously been reported. The enzyme catalyzing oxidation of the catabolic intermediate 4-hydroxyisophthalate to the ring cleavage substrate protocatechuate was also purified from strain NCIMB 9866, but its genetic determinant is still unavailable. In this study, the gene hipH, encoding 4-hydroxyisophthalate hydroxylase, from strain NCIMB 9866 was cloned by transposon mutagenesis. Purified recombinant HipH-His6 was found to be a dimer protein with a molecular mass of approximately 110 kDa. HipH-His6 catalyzed the hydroxylation of 4-hydroxyisophthalate to protocatechuate with a specific activity of 1.54 U mg(-1) and showed apparent Km values of 11.40 ± 3.05 μM for 4-hydroxyisophthalate with NADPH and 11.23 ± 2.43 μM with NADH and similar Km values for NADPH and NADH (64.31 ± 13.16 and 72.76 ± 12.06 μM, respectively). The identity of protocatechuate generated from 4-hydroxyisophthalate hydroxylation by HipH-His6 has also been confirmed by high-performance liquid chromatography and mass spectrometry. Gene transcriptional analysis, gene knockout, and complementation indicated that hipH is essential for 2,4-xylenol catabolism but not for p-cresol catabolism in this strain. This fills a gap in our understanding of the gene that encodes a critical step in 2,4-xylenol catabolism and also provides another example of biochemical and genetic diversity of microbial catabolism of structurally similar compounds.

  16. The 3,4-dihydroxyphenylacetic acid catabolon, a catabolic unit for degradation of biogenic amines tyramine and dopamine in Pseudomonas putida U.

    Science.gov (United States)

    Arcos, Mario; Olivera, Elías R; Arias, Sagrario; Naharro, Germán; Luengo, José M

    2010-06-01

    Degradation of tyramine and dopamine by Pseudomonas putida U involves the participation of twenty one proteins organized in two coupled catabolic pathways, Tyn (tynABFEC tynG tynR tynD, 12 338 bp) and Hpa (hpaR hpaBC hpaHI hpaX hpaG1G2EDF hpaA hpaY, 12 722 bp). The Tyn pathway catalyses the conversion of tyramine and dopamine into 4-hydroxyphenylacetic acid (4HPA) and 3,4-dihydroxyphenylacetic acid (3,4HPA) respectively. Together, the Tyn and Hpa pathways constitute a complex catabolic unit (the 3,4HPA catabolon) in which 3,4HPA is the central intermediate. The genes encoding Tyn proteins are organized in four consecutive transcriptional units (tynABFEC, tynG, tynR and tynD), whereas those encoding Hpa proteins constitute consecutive operons (hpaBC, hpaG1G2EDF, hpaX, hpaHI) and three independent units (hpaA, hpaR and hpaY). Genetic engineering approaches were used to clone tyn and hpa genes and then express them, either individually or in tandem, in plasmids and/or bacterial chromosomes, resulting in recombinant bacterial strains able to eliminate tyramine and dopamine from different media. These results enlarge our biochemical and genetic knowledge of the microbial catabolic routes involved in the degradation of aromatic bioamines. Furthermore, they provide potent biotechnological tools to be used in food processing and fermentation as well as new strategies that could be used for pharmacological and gene therapeutic applications in the near future.

  17. High Resolution Structures of Periplasmic Glucose-binding Protein of Pseudomonas putida CSV86 Reveal Structural Basis of Its Substrate Specificity.

    Science.gov (United States)

    Pandey, Suman; Modak, Arnab; Phale, Prashant S; Bhaumik, Prasenjit

    2016-04-01

    Periplasmic substrate-binding proteins (SBPs) bind to the specific ligand with high affinity and mediate their transport into the cytoplasm via the cognate inner membrane ATP-binding cassette proteins. Because of low sequence identities, understanding the structural basis of substrate recognition by SBPs has remained very challenging. There are several structures available for the ligand-bound sugar SBPs, but very few unliganded structures are reported. No structural data are available for sugar SBPs fromPseudomonassp. to date. This study reports the first high resolution crystal structures of periplasmic glucose-binding protein fromPseudomonas putidaCSV86 (ppGBP) in unliganded form (2.5 Å) and complexed with glucose (1.25 Å) and galactose (1.8 Å). Asymmetric domain closure of ppGBP was observed upon substrate binding. The ppGBP was found to have an affinity of ∼ 0.3 μmfor glucose. The structural analysis showed that the sugars are bound to the protein mainly by hydrogen bonds, and the loss of two strong hydrogen bonds between ppGBP and galactose compared with glucose may be responsible for lowering its affinity toward galactose. The higher stability of ppGBP-glucose complex was also indicated by an 8 °C increase in the melting temperature compared with unliganded form and ppGBP-galactose complex. ppGBP binds to monosaccharide, but the structural features revealed it to have an oligosaccharide-binding protein fold, indicating that during evolution the sugar binding pocket may have undergone structural modulation to accommodate monosaccharide only.

  18. Characterization of a phosphate solubilizing and antagonistic strain of Pseudomonas putida (B0) isolated from a sub-alpine location in the Indian Central Himalaya.

    Science.gov (United States)

    Pandey, Anita; Trivedi, Pankaj; Kumar, Bhavesh; Palni, Lok Man S

    2006-08-01

    The morphological, biochemical, and physiological characteristics of a phosphate solubilizing and antagonistic bacterial strain, designated as B0, isolated from a sub-alpine Himalayan forest site have been described. The isolate is gram negative, rod shaped, 0.8 x 1.6 microm in size, and psychrotrophic in nature that could grow from 0 to 35 degrees C (optimum temp. 25 degrees C). It exhibited tolerance to a wide pH range (3-12; optimum 8.0) and salt concentration up to 4% (w/v). Although it was sensitive to kanamycin, gentamicin, and streptomycin (1000 microg mL(-1)). The isolate showed maximum similarity with Pseudomonas putida based on 16S rRNA analysis. It solubilized tricalcium phosphate under in vitro conditions. The phosphate solubilization was estimated along a temperature range (4-28 degrees C), and maximum activity (247 microg mL(-1)) was recorded at 21 degrees C after 15 days of incubation. The phosphate solubilizing activity coincided with a concomitant decrease in pH of the medium. The isolate also exhibited antifungal activity against phytopathogenic fungi in Petri dish assays and produced chitinase, ss-l,3-glucanase, salicylic acid, siderophore, and hydrogen cyanide. The plant growth promotion and antifungal properties were demonstrated through a maize-based bioassay under greenhouse conditions. Although the bacterial inoculation was found to result in significant increment in plant biomass, it stimulated bacterial and suppressed fungal counts in the rhizosphere. The present study is important with respect to enumerating microbial diversity of the colder regions as well as understanding the potential biotechnological applications of native microbes.

  19. Profiling metabolic networks to study cancer metabolism.

    Science.gov (United States)

    Hiller, Karsten; Metallo, Christian M

    2013-02-01

    Cancer is a disease of unregulated cell growth and survival, and tumors reprogram biochemical pathways to aid these processes. New capabilities in the computational and bioanalytical characterization of metabolism have now emerged, facilitating the identification of unique metabolic dependencies that arise in specific cancers. By understanding the metabolic phenotype of cancers as a function of their oncogenic profiles, metabolic engineering may be applied to design synthetically lethal therapies for some tumors. This process begins with accurate measurement of metabolic fluxes. Here we review advanced methods of quantifying pathway activity and highlight specific examples where these approaches have uncovered potential opportunities for therapeutic intervention.

  20. Metabolism disrupting chemicals and metabolic disorders.

    Science.gov (United States)

    Heindel, Jerrold J; Blumberg, Bruce; Cave, Mathew; Machtinger, Ronit; Mantovani, Alberto; Mendez, Michelle A; Nadal, Angel; Palanza, Paola; Panzica, Giancarlo; Sargis, Robert; Vandenberg, Laura N; Vom Saal, Frederick

    2017-03-01

    The recent epidemics of metabolic diseases, obesity, type 2 diabetes(T2D), liver lipid disorders and metabolic syndrome have largely been attributed to genetic background and changes in diet, exercise and aging. However, there is now considerable evidence that other environmental factors may contribute to the rapid increase in the incidence of these metabolic diseases. This review will examine changes to the incidence of obesity, T2D and non-alcoholic fatty liver disease (NAFLD), the contribution of genetics to these disorders and describe the role of the endocrine system in these metabolic disorders. It will then specifically focus on the role of endocrine disrupting chemicals (EDCs) in the etiology of obesity, T2D and NAFLD while finally integrating the information on EDCs on multiple metabolic disorders that could lead to metabolic syndrome. We will specifically examine evidence linking EDC exposures during critical periods of development with metabolic diseases that manifest later in life and across generations.

  1. Metabolic engineering of Arabidopsis for remediation of different polycyclic aromatic hydrocarbons using a hybrid bacterial dioxygenase complex.

    Science.gov (United States)

    Peng, Rihe; Fu, Xiaoyan; Tian, Yongsheng; Zhao, Wei; Zhu, Bo; Xu, Jing; Wang, Bo; Wang, Lijuan; Yao, Quanhong

    2014-11-01

    The widespread presence of polycyclic aromatic hydrocarbons (PAHs) and their potential harm to various organisms has generated interest in efficiently eliminating these compounds from the environment. Phytoremediation is an efficient technology for cleaning up pollutants. However, unlike microorganisms, plants lack the catabolic pathway for complete degradation of these dangerous groups of compounds. One way to enhance the potential of plants for remediation of these compounds is by transferring genes involved in xenobiotic degradation from microbes to plants. In this paper, four genes, namely nidA and nidB (encoding the large and small subunits of naphthalene dioxygenase of Mycobacterium vanbaalenii PYR-1) as well as NahAa and NahAb (encoding flavoprotein reductase and ferredoxin of the electron-transport chain of the Pseudomonas putida G7 naphthalene dioxygenase system), were transferred and ectopically expressed in Arabidopsis thaliana. Transgenic Arabidopsis plants overexpressing the heterozygous naphthalene dioxygenase system exhibited enhanced tolerance toward 2-4 rings PAHs. Transgenic plants assimilated PAHs from the culture media faster and accumulated less in vivo than wild-type plants. Furthermore, examination of metabolic intermediates by gas chromatography-mass spectrometry revealed that the naphthalene metabolic pathway in transgenic plants mainly involves the dioxygenase pathway. Taken together, our findings suggest that grafting the naphthalene dioxygenase complex into plants is a possible strategy to breed PAH-tolerant plants to efficiently degrade PAHs in the environment. Copyright © 2014 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  2. Mangiferin modulation of metabolism and metabolic syndrome.

    Science.gov (United States)

    Fomenko, Ekaterina Vladimirovna; Chi, Yuling

    2016-09-10

    The recent emergence of a worldwide epidemic of metabolic disorders, such as obesity and diabetes, demands effective strategy to develop nutraceuticals or pharmaceuticals to halt this trend. Natural products have long been and continue to be an attractive source of nutritional and pharmacological therapeutics. One such natural product is mangiferin (MGF), the predominant constituent of extracts of the mango plant Mangifera indica L. Reports on biological and pharmacological effects of MGF increased exponentially in recent years. MGF has documented antioxidant and anti-inflammatory effects. Recent studies indicate that it modulates multiple biological processes involved in metabolism of carbohydrates and lipids. MGF has been shown to improve metabolic abnormalities and disorders in animal models and humans. This review focuses on the recently reported biological and pharmacological effects of MGF on metabolism and metabolic disorders. © 2016 BioFactors, 42(5):492-503, 2016.

  3. Cold-induced metabolism

    NARCIS (Netherlands)

    Lichtenbelt, W. van Marken; Daanen, H.A.M.

    2003-01-01

    Purpose of review Cold response can be insulative (drop in peripheral temperature) or metabolic (increase in energy expenditure). Nonshivering thermogenesis by sympathetic, norepinephrine-induced mitochondrial heat production in brown adipose tissue is a well known component of this metabolic

  4. Cold-induced metabolism

    NARCIS (Netherlands)

    van Marken Lichtenbelt, W.D.; Daanen, A.M.

    2003-01-01

    Cold-induced metabolism. van Marken Lichtenbelt WD, Daanen HA. Department of Human Biology, Maastricht University, Maastricht, The Netherlands. PURPOSE OF REVIEW: Cold response can be insulative (drop in peripheral temperature) or metabolic (increase in energy expenditure). Nonshivering thermogenesi

  5. Lipid Metabolism Disorders

    Science.gov (United States)

    ... metabolic disorder, something goes wrong with this process. Lipid metabolism disorders, such as Gaucher disease and Tay-Sachs disease, involve lipids. Lipids are fats or fat-like substances. They ...

  6. Mineral metabolism in cats

    OpenAIRE

    Pineda Martos, Carmen María

    2014-01-01

    The present Doctoral Thesis wa metabolism in the feline species. Through a series of studies, the relationship between calcium metabolism and the main hormones involved in it has been determined metabolism during the juvenile stage of growing cats effects linked to feeding calculolytic diets on feline mineral metabolism. The first part of the work was aimed the quantification of intact (I-PTH) and whole PTH) and to characterize the dynamics of PTH secretion, including ...

  7. Engineering Cellular Metabolism

    DEFF Research Database (Denmark)

    Nielsen, Jens; Keasling, Jay

    2016-01-01

    of metabolic engineering and will discuss how new technologies can enable metabolic engineering to be scaled up to the industrial level, either by cutting off the lines of control for endogenous metabolism or by infiltrating the system with disruptive, heterologous pathways that overcome cellular regulation....

  8. Metabolic Engineering X Conference

    Energy Technology Data Exchange (ETDEWEB)

    Flach, Evan [American Institute of Chemical Engineers

    2015-05-07

    The International Metabolic Engineering Society (IMES) and the Society for Biological Engineering (SBE), both technological communities of the American Institute of Chemical Engineers (AIChE), hosted the Metabolic Engineering X Conference (ME-X) on June 15-19, 2014 at the Westin Bayshore in Vancouver, British Columbia. It attracted 395 metabolic engineers from academia, industry and government from around the globe.

  9. Catabolism of caffeine and purification of a xanthine oxidade responsible for methyluric acids production in Pseudomonas putida L Catabolismo de cafeína e purificação de xantina oxidase responsável pela produção de ácidos metilúricos em Pseudomonas putida L

    Directory of Open Access Journals (Sweden)

    Dirce Mithico Yamaoka-Yano

    1999-01-01

    Full Text Available Caffeine catabolism and a xanthine oxidase involved in the alkaloid breakdown were studied in Pseudomonas putida L, a strain displaying high ability to grow on this substrate. Cells cultured with unlabelled caffeine and 14C labeled caffeine and xanthine showed that this alkaloid was broken-down via theobromine/paraxanthine -> 7-methylxanthine -> xanthine -> uric acid -> allantoin -> allantoic acid. Methyluric acids were formed from the oxidation of theobromine, paraxanthine and 7-methylxanthine, although no bacterial growth was observed on these compounds, indicating that this might be due to a wide substrate specificity of xanthine oxidase. This was confirmed by activity staining in PAGE where activity was observed with theophylline and 3-methylxanthine, which are not involved in the alkaloid breakdown. A single band of activity was detected without addition of NAD+, showing an oxidase form of the enzyme. The enzyme optimum temperature and pH were 30oC and 7.0, respectively. The determined Km was 169 µM, and the pI 3.1 - 4.0. The molecular weight determined by side by side comparison of activity staining of the enzyme in PAGE and PAGE of BSA was 192 kDa, which was coincident with the sum (198.4 kDa of three subunits (71, 65.6 and 61.8 kDa of the purified protein.O catabolismo de cafeína e a enzima xantina oxidase, envolvida na sua degradação, foram estudados em Pseudomonas putida L, uma linhagem com alta capacidade para utilizar este substrato como fonte de energia. Células crescidas na presença de cafeína e xantina marcadas com 14C, e cafeína não marcada, mostraram que este alcalóide foi degradado via teobromina/paraxantina -> 7-metilxantina -> xantina -> ácido úrico -> alantoína -> ácido alantóico. Ácidos metilúricos foram formados a partir de teobromina, paraxantina e 7-metilxantina, embora nenhum crescimento bacteriano ter sido observado quando estes compostos foram usados como substratos, indicando que a xantina oxidase

  10. Systems Biology of Metabolism.

    Science.gov (United States)

    Nielsen, Jens

    2017-06-20

    Metabolism is highly complex and involves thousands of different connected reactions; it is therefore necessary to use mathematical models for holistic studies. The use of mathematical models in biology is referred to as systems biology. In this review, the principles of systems biology are described, and two different types of mathematical models used for studying metabolism are discussed: kinetic models and genome-scale metabolic models. The use of different omics technologies, including transcriptomics, proteomics, metabolomics, and fluxomics, for studying metabolism is presented. Finally, the application of systems biology for analyzing global regulatory structures, engineering the metabolism of cell factories, and analyzing human diseases is discussed.

  11. Sustained metabolic scope.

    Science.gov (United States)

    Peterson, C C; Nagy, K A; Diamond, J

    1990-03-01

    Sustained metabolic rates (SusMR) are time-averaged metabolic rates that are measured in free-ranging animals maintaining constant body mass over periods long enough that metabolism is fueled by food intake rather than by transient depletion of energy reserves. Many authors have suggested that SusMR of various wild animal species are only a few times resting (basal or standard) metabolic rates (RMR). We test this conclusion by analyzing all 37 species (humans, 31 other endothermic vertebrates, and 5 ectothermic vertebrates) for which SusMR and RMR had both been measured. For all species, the ratio of SusMR to RMR, which we term sustained metabolic scope, is less than 7; most values fall between 1.5 and 5. Some of these values, such as those for Tour de France cyclists and breeding birds, are surely close to sustainable metabolic ceilings for the species studied. That is, metabolic rates higher than 7 times RMR apparently cannot be sustained indefinitely. These observations pose several questions: whether the proximate physiological causes of metabolic ceilings reside in the digestive tract's ability to process food or in each tissue's metabolic capacity; whether ceiling values are independent of the mode of energy expenditure; whether ceilings are set by single limiting physiological capacities or by coadjusted clusters of capacities (symmorphosis); what the ultimate evolutionary causes of metabolic ceilings are; and how metabolic ceilings may limit animals' reproductive effort, foraging behavior, and geographic distribution.

  12. Metabolic enzymes link morphine withdrawal with metabolic disorder

    Institute of Scientific and Technical Information of China (English)

    Xi Jiang; Jing Li; Lan Ma

    2007-01-01

    @@ Energy metabolism is a fundamental biological process that is vital for the survival of all species. Disorders in the metabolic system result in deficiency or redundancy of certain nutrients, including carbohydrates, lipids, amino acids, etc. Abnormality of the energy metabolism system leads to a number of metabolic diseases, such as the metabolic syndrome. Broadly speaking, the term "metabolic diseases" now tends to be widened to the category that refers to all diseases with metabolism disorder.

  13. Engineering Cellular Metabolism.

    Science.gov (United States)

    Nielsen, Jens; Keasling, Jay D

    2016-03-10

    Metabolic engineering is the science of rewiring the metabolism of cells to enhance production of native metabolites or to endow cells with the ability to produce new products. The potential applications of such efforts are wide ranging, including the generation of fuels, chemicals, foods, feeds, and pharmaceuticals. However, making cells into efficient factories is challenging because cells have evolved robust metabolic networks with hard-wired, tightly regulated lines of communication between molecular pathways that resist efforts to divert resources. Here, we will review the current status and challenges of metabolic engineering and will discuss how new technologies can enable metabolic engineering to be scaled up to the industrial level, either by cutting off the lines of control for endogenous metabolism or by infiltrating the system with disruptive, heterologous pathways that overcome cellular regulation.

  14. Metabolic syndrome and migraine

    Directory of Open Access Journals (Sweden)

    Amit eSachdev

    2012-11-01

    Full Text Available Migraine and metabolic syndrome are highly prevaleirnt and costly conditions.The two conditions coexist, but it is unclear what relationship may exist between the two processes. Metabolic syndrome involves a number of findings, including insulin resistance, systemic hypertension, obesity, a proinflammatory state, and a prothrombotic state. Only one study addresses migraine in metabolic syndrome, finding significant differences in the presentation of metabolic syndrome in migraineurs. However, controversy exists regarding the contribution of each individual risk factor to migraine pathogensis and prevalence. It is unclear what treatment implications, if any, exist as a result of the concomitant diagnosis of migraine and metabolic syndrome. The cornerstone of migraine and metabolic syndrome treatments is prevention, relying heavily on diet modification, sleep hygiene, medication use, and exercise.

  15. Metabolic disorders in menopause

    OpenAIRE

    Grzegorz Stachowiak; Tomasz Pertyński; Magdalena Pertyńska-Marczewska

    2015-01-01

    Metabolic disorders occurring in menopause, including dyslipidemia, disorders of carbohydrate metabolism (impaired glucose tolerance – IGT, type 2 diabetes mellitus – T2DM) or components of metabolic syndrome, constitute risk factors for cardiovascular disease in women. A key role could be played here by hyperinsulinemia, insulin resistance and visceral obesity, all contributing to dyslipidemia, oxidative stress, inflammation, alter coagulation and atherosclerosis observed during the menopaus...

  16. METABOLISM OF IRON STORES

    OpenAIRE

    Saito, Hiroshi

    2014-01-01

    ABSTRACT Remarkable progress was recently achieved in the studies on molecular regulators of iron metabolism. Among the main regulators, storage iron, iron absorption, erythropoiesis and hepcidin interact in keeping iron homeostasis. Diseases with gene-mutations resulting in iron overload, iron deficiency, and local iron deposition have been introduced in relation to the regulators of storage iron metabolism. On the other hand, the research on storage iron metabolism has not advanced since th...

  17. Genetic and metabolic engineering

    OpenAIRE

    Yang,Yea-Tyng; Bennett, George N.; San, Ka-yiu

    1998-01-01

    Recent advances in molecular biology techniques, analytical methods and mathematical tools have led to a growing interest in using metabolic engineering to redirect metabolic fluxes for industrial and medical purposes. Metabolic engineering is referred to as the directed improvement of cellular properties through the modification of specific biochemical reactions or the introduction of new ones, with the use of recombinant DNA technology (Stephanopoulos, 1999). This multidisciplinary field dr...

  18. Inflammation and metabolic cardiomyopathy.

    Science.gov (United States)

    Nishida, Kazuhiko; Otsu, Kinya

    2017-03-15

    Excessive feeding is associated with an increase in the incidence of chronic metabolic diseases, such as obesity, insulin resistance, and type 2 diabetes. Metabolic disturbance induces chronic low-grade inflammation in metabolically-important organs, such as the liver and adipose tissue. Many of the inflammatory signalling pathways are directly triggered by nutrients. The pro-inflammatory mediators in adipocytes and macrophages infiltrating adipose tissue promote both local and systemic pro-inflammatory status. Metabolic cardiomyopathy is a chronic metabolic disease characterized by structural and functional alterations and interstitial fibrosis without coronary artery disease or hypertension. In the early stage of metabolic cardiomyopathy, metabolic disturbance is not accompanied by substantial changes in myocardial structure and cardiac function. However, metabolic disturbance induces subcellular low-grade inflammation in the heart, and in turn, subcellular component abnormalities, such as oxidative stress, mitochondrial dysfunction, endoplasmic reticulum stress, and impaired calcium handling, leading to impaired myocardial relaxation. In the advanced stage, the vicious cycle of subcellular component abnormalities, inflammatory cell infiltration, and neurohumoral activation induces cardiomyocyte injury and death, and cardiac fibrosis, resulting in impairment of both diastolic and systolic functions. This review discusses some recent advances in understanding involvement of inflammation in metabolic cardiomyopathy. Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2017. For permissions, please email: journals.permissions@oup.com.

  19. What is Nutrition & Metabolism?

    Science.gov (United States)

    Feinman, Richard D; Hussain, M Mahmood

    2004-08-17

    A new Open Access journal, Nutrition & Metabolism (N&M) will publish articles that integrate nutrition with biochemistry and molecular biology. The open access process is chosen to provide rapid and accessible dissemination of new results and perspectives in a field that is of great current interest. Manuscripts in all areas of nutritional biochemistry will be considered but three areas of particular interest are lipoprotein metabolism, amino acids as metabolic signals, and the effect of macronutrient composition of diet on health. The need for the journal is identified in the epidemic of obesity, diabetes, dyslipidemias and related diseases, and a sudden increase in popular diets, as well as renewed interest in intermediary metabolism.

  20. Mathematical modelling of metabolism

    DEFF Research Database (Denmark)

    Gombert, Andreas Karoly; Nielsen, Jens

    2000-01-01

    Mathematical models of the cellular metabolism have a special interest within biotechnology. Many different kinds of commercially important products are derived from the cell factory, and metabolic engineering can be applied to improve existing production processes, as well as to make new processes...... available. Both stoichiometric and kinetic models have been used to investigate the metabolism, which has resulted in defining the optimal fermentation conditions, as well as in directing the genetic changes to be introduced in order to obtain a good producer strain or cell line. With the increasing...... availability of genomic information and powerful analytical techniques, mathematical models also serve as a tool for understanding the cellular metabolism and physiology....

  1. Fluoroacetylcarnitine: metabolism and metabolic effects in mitochondria

    Energy Technology Data Exchange (ETDEWEB)

    Bremer, J.; Davis, E.J.

    1973-01-01

    The metabolism and metabolic effects of fluoroacetylcarnitine have been investigated. Carnitineacetyltransferase transfers the fluoro-acetyl group of fluoroacetylcarnitine nearly as rapidly to CoA as the acetyl group of acetylcarnitine. Fluorocitrate is then formed by citrate synthase, but this second reaction is relatively slow. The fluorocitrate formed intramitochondrially inhibits the metabolism of citrate. In heart and skeletal muscle mitochondria the accumulated citrate inhibits citrate synthesis and the ..beta..-oxidation of fatty acids. Free acetate is formed, presumably because accumulated acetyl-CoA is hydrolyzed. In liver mitochondria the accumulation of citrate leads to a relatively increased rate of ketogenesis. Increased ketogenesis is obtained also upon the addition of citrate to the reaction mixture.

  2. The two-step electrochemical oxidation of alcohols using a novel recombinant PQQ alcohol dehydrogenase as a catalyst for a bioanode.

    Science.gov (United States)

    Takeda, Kouta; Matsumura, Hirotoshi; Ishida, Takuya; Samejima, Masahiro; Igarashi, Kiyohiko; Nakamura, Nobuhumi; Ohno, Hiroyuki

    2013-12-01

    A bioanode has been developed based on the oxidation of ethanol by the recombinant pyrroloquinoline quinone (PQQ) dependent alcohol dehydrogenase from Pseudomonas putidaKT2440 heterologously expressed in Pichia pastoris. The apo form of the recombinant protein (PpADH) was purified and displayed catalytic activity for binding PQQ in the presence of Ca(2+). PpADH exhibited broad substrate specificity towards various alcohols and aldehydes. The Km values for the aldehydes of PpADH were increased compared to those for the alcohols, whereas the kcat values were unaltered. For instance, the Km values at T=298.15K (25 °C) for ethanol and acetaldehyde were 0.21 (± 0.02)mM and 5.8 (± 0.60)mM, respectively. The kcat values for ethanol and acetaldehyde were 24.8 (± 1.2) s(-1) and 31.1 (± 1.2) s(-1), respectively. The aminoferrocene was used as an electron transfer mediator between PpADH and the electrode during electrochemical experiments. The catalytic currents for the oxidation of alcohol and acetaldehyde by PpADH were also observed in this system. The electric charge for the oxidation of ethanol (Q = 2.09 × 10(-3) · C) was increased two-fold compared to that for the oxidation of acetaldehyde (Q = 0.95 × 10(-3) · C), as determined by chronoamperometric measurements. Thus, we have electrochemically demonstrated the two-step oxidation of ethanol to acetate using only PpADH. © 2013.

  3. Bacterial attraction and quorum sensing inhibition in Caenorhabditis elegans exudates.

    Science.gov (United States)

    Kaplan, Fatma; Badri, Dayakar V; Zachariah, Cherian; Ajredini, Ramadan; Sandoval, Francisco J; Roje, Sanja; Levine, Lanfang H; Zhang, Fengli; Robinette, Steven L; Alborn, Hans T; Zhao, Wei; Stadler, Michael; Nimalendran, Rathika; Dossey, Aaron T; Brüschweiler, Rafael; Vivanco, Jorge M; Edison, Arthur S

    2009-08-01

    Caenorhabditis elegans, a bacterivorous nematode, lives in complex rotting fruit, soil, and compost environments, and chemical interactions are required for mating, monitoring population density, recognition of food, avoidance of pathogenic microbes, and other essential ecological functions. Despite being one of the best-studied model organisms in biology, relatively little is known about the signals that C. elegans uses to interact chemically with its environment or as defense. C. elegans exudates were analyzed by using several analytical methods and found to contain 36 common metabolites that include organic acids, amino acids, and sugars, all in relatively high abundance. Furthermore, the concentrations of amino acids in the exudates were dependent on developmental stage. The C. elegans exudates were tested for bacterial chemotaxis using Pseudomonas putida (KT2440), a plant growth promoting rhizobacterium, Pseudomonas aeruginosa (PAO1), a soil bacterium pathogenic to C. elegans, and Escherichia coli (OP50), a non-motile bacterium tested as a control. The C. elegans exudates attracted the two Pseudomonas species, but had no detectable antibacterial activity against P. aeruginosa. To our surprise, the exudates of young adult and adult life stages of C. elegans exudates inhibited quorum sensing in the reporter system based on the LuxR bacterial quorum sensing (QS) system, which regulates bacterial virulence and other factors in Vibrio fischeri. We were able to fractionate the QS inhibition and bacterial chemotaxis activities, thus demonstrating that these activities are chemically distinct. Our results demonstrate that C. elegans can attract its bacterial food and has the potential of partially regulating the virulence of bacterial pathogens by inhibiting specific QS systems.

  4. Up-dating the Cholodny method using PET films to sample microbial communities in soil

    Directory of Open Access Journals (Sweden)

    Kordium V. A.

    2011-04-01

    Full Text Available The aim of this work was to investigate the use of PET (polyethylene terephtalate films as a modern development of Cholodny’s glass slides, to enable microscopy and molecular-based analysis of soil communities where spatial detail at the scale of microbial habitats is essential to understand microbial associations and interactions in this complex environment. Methods. Classical microbiological methods; attachment assay; surface tension measurements; molecular techniques: DNA extraction, PCR; confocal laser scanning microscopy (CLSM; micro-focus X-ray computed tomography (µCT. Results. We first show, using the model soil and rhizosphere bacteria Pseudomonas fluorescens SBW25 and P. putida KT2440, that bacteria are able to attach and detach from PET films, and that pre-conditioning with a filtered soil suspension improved the levels of attachment. Bacteria attached to the films were viable and could develop substantial biofilms. PET films buried in soil were rapidly colonised by microorganisms which could be investigated by CLSM and recovered onto agar plates. Secondly, we demonstrate that µCT can be used to non-destructively visualise soil aggregate contact points and pore spaces across the surface of PET films buried in soil. Conclusions. PET films are a successful development of Cholodny’s glass slides and can be used to sample soil communities in which bacterial adherence, growth, biofilm and community development can be investigated. The use of these films with µCT imaging in soil will enable a better understanding of soil micro-habitats and the spatially-explicit nature of microbial interactions in this complex environment.

  5. Bacterial repetitive extragenic palindromic sequences are DNA targets for Insertion Sequence elements

    Directory of Open Access Journals (Sweden)

    Pareja Eduardo

    2006-03-01

    Full Text Available Abstract Background Mobile elements are involved in genomic rearrangements and virulence acquisition, and hence, are important elements in bacterial genome evolution. The insertion of some specific Insertion Sequences had been associated with repetitive extragenic palindromic (REP elements. Considering that there are a sufficient number of available genomes with described REPs, and exploiting the advantage of the traceability of transposition events in genomes, we decided to exhaustively analyze the relationship between REP sequences and mobile elements. Results This global multigenome study highlights the importance of repetitive extragenic palindromic elements as target sequences for transposases. The study is based on the analysis of the DNA regions surrounding the 981 instances of Insertion Sequence elements with respect to the positioning of REP sequences in the 19 available annotated microbial genomes corresponding to species of bacteria with reported REP sequences. This analysis has allowed the detection of the specific insertion into REP sequences for ISPsy8 in Pseudomonas syringae DC3000, ISPa11 in P. aeruginosa PA01, ISPpu9 and ISPpu10 in P. putida KT2440, and ISRm22 and ISRm19 in Sinorhizobium meliloti 1021 genome. Preference for insertion in extragenic spaces with REP sequences has also been detected for ISPsy7 in P. syringae DC3000, ISRm5 in S. meliloti and ISNm1106 in Neisseria meningitidis MC58 and Z2491 genomes. Probably, the association with REP elements that we have detected analyzing genomes is only the tip of the iceberg, and this association could be even more frequent in natural isolates. Conclusion Our findings characterize REP elements as hot spots for transposition and reinforce the relationship between REP sequences and genomic plasticity mediated by mobile elements. In addition, this study defines a subset of REP-recognizer transposases with high target selectivity that can be useful in the development of new tools for

  6. Lignin depolymerization by fungal secretomes and a microbial sink

    Energy Technology Data Exchange (ETDEWEB)

    Salvachúa, Davinia; Katahira, Rui; Cleveland, Nicholas S.; Khanna, Payal; Resch, Michael G.; Black, Brenna A.; Purvine, Samuel O.; Zink, Erika M.; Prieto, Alicia; Martínez, María J.; Martínez, Angel T.; Simmons, Blake A.; Gladden, John M.; Beckham, Gregg T.

    2016-08-25

    In Nature, powerful oxidative enzymes secreted by white rot fungi and some bacteria catalyze lignin depolymerization and some microbes are able to catabolize the resulting aromatic compounds as carbon and energy sources. Taken together, these two processes offer a potential route for microbial valorization of lignin. However, many challenges remain in realizing this concept, including that oxidative enzymes responsible for lignin depolymerization also catalyze polymerization of low molecular weight (LMW) lignin. Here, multiple basidiomycete secretomes were screened for ligninolytic enzyme activities in the presence of a residual lignin solid stream from a corn stover biorefinery, dubbed DMR-EH (Deacetylation, Mechanical Refining, and Enzymatic Hydrolysis) lignin. Two selected fungal secretomes, with high levels of laccases and peroxidases, were utilized for DMR-EH lignin depolymerization assays. The secretome from Pleurotus eryngii, which exhibited the highest laccase activity, reduced the lignin average molecular weight by 63% and 75% at pH 7 compared to the Mw of the control treated at the same conditions and the initial DMR-EH lignin, respectively, and was applied in further depolymerization assays as a function of time. As repolymerization was observed after 3 days of incubation, an aromatic-catabolic microbe (Pseudomonas putida KT2440) was incubated with the fungal secretome and DMR-EH lignin. These experiments demonstrated that the presence of the bacterium enhances lignin depolymerization, likely due to bacterial catabolism of LMW lignin, which may partially prevent repolymerization. In addition, proteomics was also applied to the P. eryngii secretome to identify the enzymes present in the fungal cocktail utilized for the depolymerization assays, which highlighted a significant number of glucose/ methanol/choline (GMC) oxidoreductases and laccases. Overall, this study demonstrates that ligninolytic enzymes can be used to partially depolymerize a solid, high

  7. Lignin Depolymerization by Fungal Secretomes and a Microbial Sink

    Energy Technology Data Exchange (ETDEWEB)

    Salvachua, Davinia; Katahira, Rui; Cleveland, Nicholas S.; Khanna, Payal; Resch, Michael G.; Black, Brenna A.; Purvine, Samuel O.; Zink, Erika M.; Prieto, Alicia; Martinez, Maria J.; Martinez, Angel T.; Simmons, Blake A.; Gladden, John M.; Beckham, Gregg T.

    2016-11-21

    In Nature, powerful oxidative enzymes secreted by white rot fungi and some bacteria catalyze lignin depolymerization and some microbes are able to catabolize the resulting aromatic compounds as carbon and energy sources. Taken together, these two processes offer a potential route for microbial valorization of lignin. However, many challenges remain in realizing this concept, including that oxidative enzymes responsible for lignin depolymerization also catalyze polymerization of low molecular weight (LMW) lignin. Here, multiple basidiomycete secretomes were screened for ligninolytic enzyme activities in the presence of a residual lignin solid stream from a corn stover biorefinery, dubbed DMR-EH (Deacetylation, Mechanical Refining, and Enzymatic Hydrolysis) lignin. Two selected fungal secretomes, with high levels of laccases and peroxidases, were utilized for DMR-EH lignin depolymerization assays. The secretome from Pleurotus eryngii, which exhibited the highest laccase activity, reduced the lignin average molecular weight (Mw) by 63% and 75% at pH 7 compared to the Mw of the control treated at the same conditions and the initial DMR-EH lignin, respectively, and was applied in further depolymerization assays as a function of time. As repolymerization was observed after 3 days of incubation, an aromatic-catabolic microbe (Pseudomonas putida KT2440) was incubated with the fungal secretome and DMR-EH lignin. These experiments demonstrated that the presence of the bacterium enhances lignin depolymerization, likely due to bacterial catabolism of LMW lignin, which may partially prevent repolymerization. In addition, proteomics was also applied to the P. eryngii secretome to identify the enzymes present in the fungal cocktail utilized for the depolymerization assays, which highlighted a significant number of glucose/methanol/choline (GMC) oxidoreductases and laccases. Overall, this study demonstrates that ligninolytic enzymes can be used to partially depolymerize a solid

  8. Attractor metabolic networks.

    Directory of Open Access Journals (Sweden)

    Ildefonso M De la Fuente

    Full Text Available BACKGROUND: The experimental observations and numerical studies with dissipative metabolic networks have shown that cellular enzymatic activity self-organizes spontaneously leading to the emergence of a Systemic Metabolic Structure in the cell, characterized by a set of different enzymatic reactions always locked into active states (metabolic core while the rest of the catalytic processes are only intermittently active. This global metabolic structure was verified for Escherichia coli, Helicobacter pylori and Saccharomyces cerevisiae, and it seems to be a common key feature to all cellular organisms. In concordance with these observations, the cell can be considered a complex metabolic network which mainly integrates a large ensemble of self-organized multienzymatic complexes interconnected by substrate fluxes and regulatory signals, where multiple autonomous oscillatory and quasi-stationary catalytic patterns simultaneously emerge. The network adjusts the internal metabolic activities to the external change by means of flux plasticity and structural plasticity. METHODOLOGY/PRINCIPAL FINDINGS: In order to research the systemic mechanisms involved in the regulation of the cellular enzymatic activity we have studied different catalytic activities of a dissipative metabolic network under different external stimuli. The emergent biochemical data have been analysed using statistical mechanic tools, studying some macroscopic properties such as the global information and the energy of the system. We have also obtained an equivalent Hopfield network using a Boltzmann machine. Our main result shows that the dissipative metabolic network can behave as an attractor metabolic network. CONCLUSIONS/SIGNIFICANCE: We have found that the systemic enzymatic activities are governed by attractors with capacity to store functional metabolic patterns which can be correctly recovered from specific input stimuli. The network attractors regulate the catalytic patterns

  9. Cold-induced metabolism

    NARCIS (Netherlands)

    Lichtenbelt, W. van Marken; Daanen, H.A.M.

    2003-01-01

    Purpose of review Cold response can be insulative (drop in peripheral temperature) or metabolic (increase in energy expenditure). Nonshivering thermogenesis by sympathetic, norepinephrine-induced mitochondrial heat production in brown adipose tissue is a well known component of this metabolic respon

  10. Circadian Systems and Metabolism

    NARCIS (Netherlands)

    Roenneberg, Till; Merrow, Martha

    1999-01-01

    Circadian systems direct many metabolic parameters and, at the same time, they appear to be exquisitely shielded from metabolic variations. Although the recent decade of circadian research has brought insights into how circadian periodicity may be generated at the molecular level, little is known ab

  11. Metabolic Engineering VII Conference

    Energy Technology Data Exchange (ETDEWEB)

    Kevin Korpics

    2012-12-04

    The aims of this Metabolic Engineering conference are to provide a forum for academic and industrial researchers in the field; to bring together the different scientific disciplines that contribute to the design, analysis and optimization of metabolic pathways; and to explore the role of Metabolic Engineering in the areas of health and sustainability. Presentations, both written and oral, panel discussions, and workshops will focus on both applications and techniques used for pathway engineering. Various applications including bioenergy, industrial chemicals and materials, drug targets, health, agriculture, and nutrition will be discussed. Workshops focused on technology development for mathematical and experimental techniques important for metabolic engineering applications will be held for more in depth discussion. This 2008 meeting will celebrate our conference tradition of high quality and relevance to both industrial and academic participants, with topics ranging from the frontiers of fundamental science to the practical aspects of metabolic engineering.

  12. Disorders of fructose metabolism.

    Science.gov (United States)

    Froesch, E R

    1976-11-01

    There are fundamental differences between the metabolic fate of fructose and of glucose. Whereas the metabolism of glucose is controlled by hormones such as insulin, fructose uptake and phosphorylation in the liver occurs independently of hormones and its ultimate metabolic fate is unpredictable. Essential fructosuria, a harmless inherited anomaly of fructose metabolism, is the least harmful of the disorders of fructose metabolism. Hereditary fructose intolerance and fructose-1,6-diphosphatase deficiency are discussed in greater detail with regard to biochemical abnormalities and clinical aspects. HFI is most serious in bottle-fed infants who cannot reject their sucrose-containing diet. Patients with HFI will have no clinical symptoms if kept on a fructose-free diet. In contrast, patients with fructose-1,6-diphosphatase deficiency can tolerate frucose. However, severe infections precipitate attacks of hypoglycaemia and lactic acidosis.

  13. Primary Metabolic Pathways and Metabolic Flux Analysis

    DEFF Research Database (Denmark)

    2015-01-01

    his chapter introduces the metabolic flux analysis (MFA) or stoichiometry-based MFA, and describes the quantitative basis for MFA. It discusses the catabolic pathways in which free energy is produced to drive the cell-building anabolic pathways. An overview of these primary pathways provides...

  14. 一种通过重组大肠杆菌和重组恶臭假单胞菌生产3-羟基癸酸的新方法%A Novel Method for Production of 3-Hydroxydecanoic Acid by Recombinant Escherichia coli and Pseudomonas putida

    Institute of Scientific and Technical Information of China (English)

    郑重; 宫强; 陈国强

    2004-01-01

    3-hydroxydecanoic acid (3HD) is an interesting intermediate for chemical synthesis of many valuable compounds. A novel method to produce 3HD by recombinant bacteria was constructed in Escherichia coli HB101 and Pseudomonas putida GPp104, respectively. Simultaneous expression of both phaG encoding (R)-3-hydroxydecanoylACP:CoA transacylase and tesB encoding thioesterase Ⅱ in E. coli HB101 increased 3HD production approximate1.7-folds compared with the expression of phaG gene alone under identical conditions. In addition, when the tesB gene was introduced into the strain, the polyhydroxyalkanoate synthase negative strain P. putida GPp104 produced extracellular 3HD. Thus, a novel pathway to produce 3HD by recombinant Pseudomonas was constructed. It was also found that the ratio of carbon source to nitrogen source affected the production of 3HD by recombinant P.putida harboring tesB gene. Nitrogen limitation seemed to promote the extracellular 3HD production.

  15. Metabolic disorders in menopause

    Directory of Open Access Journals (Sweden)

    Grzegorz Stachowiak

    2015-04-01

    Full Text Available Metabolic disorders occurring in menopause, including dyslipidemia, disorders of carbohydrate metabolism (impaired glucose tolerance – IGT, type 2 diabetes mellitus – T2DM or components of metabolic syndrome, constitute risk factors for cardiovascular disease in women. A key role could be played here by hyperinsulinemia, insulin resistance and visceral obesity, all contributing to dyslipidemia, oxidative stress, inflammation, alter coagulation and atherosclerosis observed during the menopausal period. Undiagnosed and untreated, metabolic disorders may adversely affect the length and quality of women’s life. Prevention and treatment preceded by early diagnosis should be the main goal for the physicians involved in menopausal care. This article represents a short review of the current knowledge concerning metabolic disorders (e.g. obesity, polycystic ovary syndrome or thyroid diseases in menopause, including the role of a tailored menopausal hormone therapy (HT. According to current data, HT is not recommend as a preventive strategy for metabolic disorders in menopause. Nevertheless, as part of a comprehensive strategy to prevent chronic diseases after menopause, menopausal hormone therapy, particularly estrogen therapy may be considered (after balancing benefits/risks and excluding women with absolute contraindications to this therapy. Life-style modifications, with moderate physical activity and healthy diet at the forefront, should be still the first choice recommendation for all patients with menopausal metabolic abnormalities.

  16. Metabolic disorders in menopause.

    Science.gov (United States)

    Stachowiak, Grzegorz; Pertyński, Tomasz; Pertyńska-Marczewska, Magdalena

    2015-03-01

    Metabolic disorders occurring in menopause, including dyslipidemia, disorders of carbohydrate metabolism (impaired glucose tolerance - IGT, type 2 diabetes mellitus - T2DM) or components of metabolic syndrome, constitute risk factors for cardiovascular disease in women. A key role could be played here by hyperinsulinemia, insulin resistance and visceral obesity, all contributing to dyslipidemia, oxidative stress, inflammation, alter coagulation and atherosclerosis observed during the menopausal period. Undiagnosed and untreated, metabolic disorders may adversely affect the length and quality of women's life. Prevention and treatment preceded by early diagnosis should be the main goal for the physicians involved in menopausal care. This article represents a short review of the current knowledge concerning metabolic disorders (e.g. obesity, polycystic ovary syndrome or thyroid diseases) in menopause, including the role of a tailored menopausal hormone therapy (HT). According to current data, HT is not recommend as a preventive strategy for metabolic disorders in menopause. Nevertheless, as part of a comprehensive strategy to prevent chronic diseases after menopause, menopausal hormone therapy, particularly estrogen therapy may be considered (after balancing benefits/risks and excluding women with absolute contraindications to this therapy). Life-style modifications, with moderate physical activity and healthy diet at the forefront, should be still the first choice recommendation for all patients with menopausal metabolic abnormalities.

  17. Binary combination of epsilon-poly-L-lysine and isoeugenol affect progression of spoilage microbiota in fresh turkey meat, and delay onset of spoilage in Pseudomonas putida challenged meat.

    Science.gov (United States)

    Hyldgaard, Morten; Meyer, Rikke L; Peng, Min; Hibberd, Ashley A; Fischer, Jana; Sigmundsson, Arnar; Mygind, Tina

    2015-12-23

    Proliferation of microbial population on fresh poultry meat over time elicits spoilage when reaching unacceptable levels, during which process slime production, microorganism colony formation, negative organoleptic impact and meat structure change are observed. Spoilage organisms in raw meat, especially Gram-negative bacteria can be difficult to combat due to their cell wall composition. In this study, the natural antimicrobial agents ε-poly-L-lysine (ε-PL) and isoeugenol were tested individually and in combinations for their activities against a selection of Gram-negative strains in vitro. All combinations resulted in additive interactions between ε-PL and isoeugenol towards the bacteria tested. The killing efficiency of different ratios of the two antimicrobial agents was further evaluated in vitro against Pseudomonas putida. Subsequently, the most efficient ratio was applied to a raw turkey meat model system which was incubated for 96 h at spoilage temperature. Half of the samples were challenged with P. putida, and the bacterial load and microbial community composition was followed over time. CFU counts revealed that the antimicrobial blend was able to lower the amount of viable Pseudomonas spp. by one log compared to untreated samples of challenged turkey meat, while the single compounds had no effect on the population. However, the compounds had no effect on Pseudomonas spp. CFU in unchallenged meat. Next-generation sequencing offered culture-independent insight into population diversity and changes in microbial composition of the meat during spoilage and in response to antimicrobial treatment. Spoilage of unchallenged turkey meat resulted in decreasing species diversity over time, regardless of whether the samples received antimicrobial treatment. The microbiota composition of untreated unchallenged meat progressed from a Pseudomonas spp. to a Pseudomonas spp., Photobacterium spp., and Brochothrix thermosphacta dominated food matrix on the expense of low

  18. Regulation of lipid metabolism

    Institute of Scientific and Technical Information of China (English)

    Peng LI

    2011-01-01

    @@ Lipids including cholesterol, phospholipids, fatty acids and triacylglycerols are important cellular constituents involved in membrane structure, energy homeostasis and many biological processes such as signal transduction, organelle development and cell differentiation.Recently, the area of lipid metabolism has drawn a great deal of attention due to its emerging role in the development of metabolic disorders such as obesity, diabetes, atherosclerosis and liver steatosis.We decided to organize a special issue of Frontiers in Biology focusing on our current understanding of lipid metabolism.

  19. A Metabolic Switch

    DEFF Research Database (Denmark)

    Hjorth, Poul G.

    Our muscles are metabolically flexible, i.e., they are capable of `switching' between two types of oxidation: (1) when fasting, a predominantly lipid oxidation with high rates of fatty acid uptake, and (2) when fed, suppression of lipid oxidation in favour of increased glucose uptake, oxidation...... and storage, in response to insulin. One of the many manifestations of obesity and Type 2 diabetes is an insulin resistance of the skeletal muscles, which suppresses this metabolic switch. This talk describes recent development of a low-dimensional system of ODEs that model the metabolic switch, displaying...

  20. Sirtuins, Metabolism and Cancer

    Directory of Open Access Journals (Sweden)

    Barbara eMartinez-Pastor

    2012-02-01

    Full Text Available More than a decade ago, sirtuins were discovered as a highly conserved family of NAD+-dependent enzymes that extend lifespan in lower organisms. In mammals, sirtuins are key regulators of stress responses and metabolism, influencing a range of diseases, including diabetes, neurodegeneration and cancer. In recent years, new functions of sirtuins have been characterized, uncovering the underlying mechanisms of their multifaceted role in metabolism. Here, we specifically review recent progress on the role of sirtuins in DNA repair and energy metabolism, further discussing the implication of sirtuins in the biology of cancer.

  1. Vitamin D metabolism

    Energy Technology Data Exchange (ETDEWEB)

    Deluca, H.F.

    1977-01-01

    During the past decade, an explosion of information has become available on the metabolism and function of vitamin D which is of great importance to clinicians in the treatment of metabolic bone disease. We have learned that vitamin D is the precursor of at least one hormone and that this hormone carries out functions in calcium and phosphorus metabolism bringing about mineralization of bone on one hand, and the prevention of hypocalcaemic tetany on the other. It may also function in the prevention of such degenerative bone diseases as osteoporosis. An important analogue of this hormone, 1..cap alpha..-hydroxyvitamin D/sub 3/ has been prepared and is used successfully in the treatment of a variety of clinical conditions. This presentation will summarize these findings and their possible implications in these metabolic bone diseases.

  2. What is Nutrition & Metabolism?

    Directory of Open Access Journals (Sweden)

    Feinman Richard D

    2004-08-01

    Full Text Available Abstract A new Open Access journal, Nutrition & Metabolism (N&M will publish articles that integrate nutrition with biochemistry and molecular biology. The open access process is chosen to provide rapid and accessible dissemination of new results and perspectives in a field that is of great current interest. Manuscripts in all areas of nutritional biochemistry will be considered but three areas of particular interest are lipoprotein metabolism, amino acids as metabolic signals, and the effect of macronutrient composition of diet on health. The need for the journal is identified in the epidemic of obesity, diabetes, dyslipidemias and related diseases, and a sudden increase in popular diets, as well as renewed interest in intermediary metabolism.

  3. Metabolism. Part III: Lipids.

    Science.gov (United States)

    Bodner, George M.

    1986-01-01

    Describes the metabolic processes of complex lipids, including saponification, activation and transport, and the beta-oxidation spiral. Discusses fatty acid degradation in regard to biochemical energy and ketone bodies. (TW)

  4. Amino Acid Metabolism Disorders

    Science.gov (United States)

    ... this process. One group of these disorders is amino acid metabolism disorders. They include phenylketonuria (PKU) and maple syrup urine disease. Amino acids are "building blocks" that join together to form ...

  5. Sleep & the metabolic syndrome

    National Research Council Canada - National Science Library

    Lam, Jamie C M; Ip, Mary S M

    2010-01-01

    Sleep is an essential part of our daily living, and sleep disturbances may intervene with the biological and physiological processes in human body leading to the development of metabolic dysfunction...

  6. Epigenetics and Cellular Metabolism

    Science.gov (United States)

    Xu, Wenyi; Wang, Fengzhong; Yu, Zhongsheng; Xin, Fengjiao

    2016-01-01

    Living eukaryotic systems evolve delicate cellular mechanisms for responding to various environmental signals. Among them, epigenetic machinery (DNA methylation, histone modifications, microRNAs, etc.) is the hub in transducing external stimuli into transcriptional response. Emerging evidence reveals the concept that epigenetic signatures are essential for the proper maintenance of cellular metabolism. On the other hand, the metabolite, a main environmental input, can also influence the processing of epigenetic memory. Here, we summarize the recent research progress in the epigenetic regulation of cellular metabolism and discuss how the dysfunction of epigenetic machineries influences the development of metabolic disorders such as diabetes and obesity; then, we focus on discussing the notion that manipulating metabolites, the fuel of cell metabolism, can function as a strategy for interfering epigenetic machinery and its related disease progression as well. PMID:27695375

  7. [Metabolic syndrome and melatonin].

    Science.gov (United States)

    Rapoport, S I; Molchanov, A Iu; Golichenkov, V A; Burlakova, O V; Suprunenko, E S; Savchenko, E S

    2013-01-01

    Metabolic syndrome (MS) is characterized by the following symptoms: obesity, AH, dyslipidemia, insulin resistance. Pathophysiologically, MS is underlain by disorders of many biochemical and physiological processes, such as elevated levels of low density lipoproteins, hyperstimulation of pancreatic b-cells, increased insulin secretion, substitution of lipid metabolism for carbohydrate one, overgrowth of adipose tissue, excess production of adiponectin, leptin and other signal molecules and a rise in their local intravascular concentration, weight gain. Endogenous and exogenous melatonin inhibits these pathophysiological mechanisms, normalizes metabolism, equilibrates insulin secretion, prevents pancreatic hyperfunction, phosphorylates insulin receptors, inactivates active oxygen and nitrogen species including those produced in LDLP metabolism. Melatonin has specific MT1 and MT2 receptors localized in all body cells. Due to this, it exerts combined preventive action in patients with MS. Recently, melatonin has been reported to have therapeutic effect in MS; it may be recommended to treat this condition.

  8. Endocrine and Metabolic Disorders

    Science.gov (United States)

    ... for disorders of endocrine glands other than the thyroid, compared to 3.1 percent of visits made by women. The rate of visits due to metabolic and immunity disorders was also higher among men than women (2. ...

  9. Hypothalamic Hormones and Metabolism

    OpenAIRE

    Thio, Liu Lin

    2012-01-01

    The ketogenic diet is an effective treatment for medically intractable epilepsy and may have antiepileptogenic, neuroprotective, and antitumor properties. While on a ketogenic diet, the body obtains most of its calories from fat rather than carbohydrates. This dramatic change in caloric composition results in a unique metabolic state. In turn, these changes in caloric composition and metabolism alter some of the neurohormones that participate in the complex neuronal network regulating energy ...

  10. Tumor cell metabolism

    Science.gov (United States)

    Romero-Garcia, Susana; Lopez-Gonzalez, Jose Sullivan; B´ez-Viveros, José Luis; Aguilar-Cazares, Dolores

    2011-01-01

    Cancer is a genetic disease that is caused by mutations in oncogenes, tumor suppressor genes and stability genes. The fact that the metabolism of tumor cells is altered has been known for many years. However, the mechanisms and consequences of metabolic reprogramming have just begun to be understood. In this review, an integral view of tumor cell metabolism is presented, showing how metabolic pathways are reprogrammed to satisfy tumor cell proliferation and survival requirements. In tumor cells, glycolysis is strongly enhanced to fulfill the high ATP demands of these cells; glucose carbons are the main building blocks in fatty acid and nucleotide biosynthesis. Glutaminolysis is also increased to satisfy NADPH regeneration, whereas glutamine carbons replenish the Krebs cycle, which produces metabolites that are constantly used for macromolecular biosynthesis. A characteristic feature of the tumor microenvironment is acidosis, which results from the local increase in lactic acid production by tumor cells. This phenomenon is attributed to the carbons from glutamine and glucose, which are also used for lactic acid production. Lactic acidosis also directs the metabolic reprogramming of tumor cells and serves as an additional selective pressure. Finally, we also discuss the role of mitochondria in supporting tumor cell metabolism. PMID:22057267

  11. Evolutionary dynamics of metabolic adaptation

    NARCIS (Netherlands)

    van Hoek, M.J.A.

    2008-01-01

    In this thesis we study how organisms adapt their metabolism to a changing environment. Metabolic adaptation occurs at different timescales. Organisms adapt their metabolism via metabolic regulation, which happens in the order of minutes to hours and via evolution, which takes many generations. Here

  12. Evolutionary dynamics of metabolic adaptation

    NARCIS (Netherlands)

    van Hoek, M.J.A.

    2008-01-01

    In this thesis we study how organisms adapt their metabolism to a changing environment. Metabolic adaptation occurs at different timescales. Organisms adapt their metabolism via metabolic regulation, which happens in the order of minutes to hours and via evolution, which takes many generations. Here

  13. Metabolism, temperature, and ventilation.

    Science.gov (United States)

    Mortola, Jacopo P; Maskrey, Michael

    2011-10-01

    In mammals and birds, all oxygen used (VO2) must pass through the lungs; hence, some degree of coupling between VO2 and pulmonary ventilation (VE) is highly predictable. Nevertheless, VE is also involved with CO2 elimination, a task that is often in conflict with the convection of O2. In hot or cold conditions, the relationship between VE and VO2 includes the participation of the respiratory apparatus to the control of body temperature and water balance. Some compromise among these tasks is achieved through changes in breathing pattern, uncoupling changes in alveolar ventilation from VE. This article examines primarily the relationship between VE and VO2 under thermal stimuli. In the process, it considers how the relationship is influenced by hypoxia, hypercapnia or changes in metabolic level. The shuffling of tasks in emergency situations illustrates that the constraints on VE-VO2 for the protection of blood gases have ample room for flexibility. However, when other priorities do not interfere with the primary goal of gas exchange, VE follows metabolic rate quite closely. The fact that arterial CO2 remains stable when metabolism is changed by the most diverse circumstances (moderate exercise, cold, cold and exercise combined, variations in body size, caloric intake, age, time of the day, hormones, drugs, etc.) makes it unlikely that VE and metabolism are controlled in parallel by the condition responsible for the metabolic change. Rather, some observations support the view that the gaseous component of metabolic rate, probably CO2, may provide the link between the metabolic level and VE.

  14. 恶臭假单胞菌产精氨酸脱亚胺酶发酵条件及特性的研究%Fermentation conditions and properties of arginine deiminase from Pseudomonas putida UN0705

    Institute of Scientific and Technical Information of China (English)

    张媛媛; 刘晓蓉; 周爱芳

    2011-01-01

    对恶臭假单胞菌N0705产精氨酸脱亚胺酶的发酵条件及部分酶学性质进行了研究.结果表明,在培养温度28℃,培养基初始pH值为6.6,接种量4%,装液量40mL/250mL,发酵时间50h时,精氨酸脱亚胺酶酶活最大.精氨酸脱亚胺酶发酵粗酶液在温度37℃及pH 6.0~7.0时有较好的稳定性.%The fermentation conditions and the some properties of arginine deiminase (ADI) produced by Pseudomonas putida UN0705 were studied. The optimal fermentation conditions were as follows: liquid volume 40ml/250ml, initial pH value 6.6, inoculum 4%, culture temperature 28℃ and culture time Soh. Under these conditions, the high activity of arginine deiminase was obtained. The crude arginine deiminase was highly stable at 37℃ and in the pH range from 6.0 to 7.0.

  15. Effects of TX-100 on Biphenol A Degradation in Biofilm by Pseudomonas Putida%曲拉通(TX-100)对恶臭假单胞菌降解生物膜中双酚A的影响

    Institute of Scientific and Technical Information of China (English)

    李都峰; 赵文晋; 黎娜; 王宪恩

    2009-01-01

    研究非离子型表面活性剂曲拉通(TX-100)对恶臭假单胞菌降解生物膜中双酚A的影响. 结果表明, 低浓度(小于临界胶束浓度, CMC)TX-100抑制生物膜中双酚A的降解, 而高浓度(大于CMC)TX-100可促进生物膜中双酚A的降解, 并且随着生物膜中TX-100浓度的增加, 双酚A的降解率逐渐增大;同时发现TX-100存在条件下生物膜主要化学组分的选择性萃取分离可导致恶臭假单胞菌降解双酚A的能力下降.%Effect of non-ionic surfactant TX-100 on biphenol A (BPA) degradation in the natural surface coatings by Pseudomonas putida was investigated. TX-100 inhibits the biodegradation of BPA in biofilm at low concentrations ( 1.0 times of CMC), and the degradation rate of BPA increases gradually with the increase of BPA concentration. The removal of the main components of the natural surface coatings by the selective extraction techniques has an effect on the BPA degradation, leading to a significant decrease in the biodegradation of BPA.

  16. Metabolic surgery: quo vadis?

    Science.gov (United States)

    Ramos-Leví, Ana M; Rubio Herrera, Miguel A

    2014-01-01

    The impact of bariatric surgery beyond its effect on weight loss has entailed a change in the way of regarding it. The term metabolic surgery has become more popular to designate those interventions that aim at resolving diseases that have been traditionally considered as of exclusive medical management, such as type 2 diabetes mellitus (T2D). Recommendations for metabolic surgery have been largely addressed and discussed in worldwide meetings, but no definitive consensus has been reached yet. Rates of diabetes remission after metabolic surgery have been one of the most debated hot topics, with heterogeneity being a current concern. This review aims to identify and clarify controversies regarding metabolic surgery, by focusing on a critical analysis of T2D remission rates achieved with different bariatric procedures, and using different criteria for its definition. Indications for metabolic surgery for patients with T2D who are not morbidly obese are also discussed. Copyright © 2013 SEEN. Published by Elsevier Espana. All rights reserved.

  17. Nutrition and metabolic syndrome.

    Directory of Open Access Journals (Sweden)

    Albornoz López, Raúl

    2012-12-01

    Full Text Available The metabolic syndrome comprises a cluster of metabolic abnormalities that increase the risk for cardiovascular disease and type 2 diabetes mellitus. The exact etiology is unclear, although it is known thatthere is a complex interaction between genetic, metabolic and environmental factors. Among the environmental factors, dietary habits play an important role in the treatment and prevention of this condition. General classic recommendations include control of obesity, increased physical activity, decreased intake of saturated fat and cholesterol, reduced intake of simple sugars and increased intake of fruits and vegetables. It has been studied the influence of diets low in carbohydrates, diets rich in polyunsaturated and monounsaturated fatty acids, fiber intake, the Mediterranean diet and the glycemic index in relation to metabolic syndrome.Other nutrients recently studied are the micronutrients (magnesium and calcium, soy and other phytochemicals. Evidence suggests that a healthy diet like the Mediterranean protects against metabolic syndrome,caracterized for a low content in saturated and trans fat, high in monounsaturated and polyunsaturated fatty acids, balanced intake of carbohydrates and high in fiber, fruits and vegetables. There is more controversy about the type of diet of choice for the control ofmetabolic syndrome (low-carbohydrate diets or lowfat, needing more studies on the role of soy and other phytochemicals.

  18. Apelin and energy metabolism

    Directory of Open Access Journals (Sweden)

    Chantal eBertrand

    2015-04-01

    Full Text Available A wide range of adipokines identified over the past years has allowed considering white adipose tissue as a secretory organ closely integrated into overall physiological and metabolic control. Apelin, an ubiquitous peptide was known to exert different physiological effects mainly on the cardiovascular system and the regulation of fluid homeostasis until its identification as an adipokine. This has increased its broad range of action and apelin now appears clearly as a new player in energy metabolism alongside leptin and adiponectin. Apelin has been shown to act on glucose and lipid metabolism but also to modulate insulin secretion. Moreover, different studies in both animals and humans have shown that plasma apelin concentrations are usually increased during obesity and type 2 diabetes. This mini-review will focus on the various systemic apelin effects on energy metabolism by addressing its mechanisms of action. The advances concerning the role of apelin in metabolic diseases in relation with the recent reports on apelin concentrations in obese and/or diabetic subjects will also be discussed.

  19. Metabolism of phencyclidine

    Energy Technology Data Exchange (ETDEWEB)

    Hoag, M.K.P.

    1987-01-01

    Phencyclidine (PCP) is a drug of abuse which may produce, in some users, a persistent schizophreniform psychosis. The possibility that long term effects of PCP are mediated by metabolic activation of the parent compound to reactive species is consistent with the demonstration of metabolism-dependent covalent binding of radiolabeled PCP in vivo and in vitro to macromolecules in rodent lung, liver, and kidney. Formation of the electrophilic iminium ion metabolite of PCP is believed to be critical for covalent binding since binding was inhibited by cyanide ion at concentrations which did not inhibit metabolism of PCP but did trap the iminium ion to form the corresponding alpha-aminonitrile. The present studies were designed to characterize further the biological fate of PCP by identifying possible macromolecular targets of the reactive metabolite(s).

  20. Analytics for Metabolic Engineering.

    Science.gov (United States)

    Petzold, Christopher J; Chan, Leanne Jade G; Nhan, Melissa; Adams, Paul D

    2015-01-01

    Realizing the promise of metabolic engineering has been slowed by challenges related to moving beyond proof-of-concept examples to robust and economically viable systems. Key to advancing metabolic engineering beyond trial-and-error research is access to parts with well-defined performance metrics that can be readily applied in vastly different contexts with predictable effects. As the field now stands, research depends greatly on analytical tools that assay target molecules, transcripts, proteins, and metabolites across different hosts and pathways. Screening technologies yield specific information for many thousands of strain variants, while deep omics analysis provides a systems-level view of the cell factory. Efforts focused on a combination of these analyses yield quantitative information of dynamic processes between parts and the host chassis that drive the next engineering steps. Overall, the data generated from these types of assays aid better decision-making at the design and strain construction stages to speed progress in metabolic engineering research.