scholarly journals Enhanced Tolerance to Naphthalene and Enhanced Rhizoremediation Performance for Pseudomonas putida KT2440 via the NAH7 Catabolic Plasmid

2012 ◽  
Vol 78 (15) ◽  
pp. 5104-5110 ◽  
Author(s):  
Matilde Fernández ◽  
José Luis Niqui-Arroyo ◽  
Susana Conde ◽  
Juan Luis Ramos ◽  
Estrella Duque
Keyword(s):  
Pseudomonas Putida ◽  
Cellular Damage ◽  
Pseudomonas Putida Kt2440 ◽  
Content Type ◽  
Naphthalene Degradation ◽  
Repair Enzymes ◽  
Damage Repair ◽  
Catabolic Plasmid

ABSTRACTIn this work, we explore the potential use of thePseudomonas putidaKT2440 strain for bioremediation of naphthalene-polluted soils.Pseudomonas putidastrain KT2440 thrives in naphthalene-saturated medium, establishing a complex response that activates genes coding for extrusion pumps and cellular damage repair enzymes, as well as genes involved in the oxidative stress response. The transfer of the NAH7 plasmid enables naphthalene degradation byP. putidaKT2440 while alleviating the cellular stress brought about by this toxic compound, without affecting key functions necessary for survival and colonization of the rhizosphere.Pseudomonas putidaKT2440(NAH7) efficiently expresses the Nah catabolic pathwayin vitroandin situ, leading to the complete mineralization of [14C]naphthalene, measured as the evolution of14CO2, while the rate of mineralization was at least 2-fold higher in the rhizosphere than in bulk soil.

scholarly journals Small-Molecule Inhibition of Choline Catabolism in Pseudomonas aeruginosa and Other Aerobic Choline-Catabolizing Bacteria

2011 ◽  
Vol 77 (13) ◽  
pp. 4383-4389 ◽  
Author(s):  
Liam F. Fitzsimmons ◽  
Stevenson Flemer ◽  
A. Sandy Wurthmann ◽  
P. Bruce Deker ◽  
Indra Neil Sarkar ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Burkholderia Cepacia ◽  
Sinorhizobium Meliloti ◽  
Growth Inhibitor ◽  
Bioinformatic Analysis ◽  
Content Type ◽  
Genes Encoding ◽  
Dependent Growth

ABSTRACTCholine is abundant in association with eukaryotes and plays roles in osmoprotection, thermoprotection, and membrane biosynthesis in many bacteria. Aerobic catabolism of choline is widespread among soil proteobacteria, particularly those associated with eukaryotes. Catabolism of choline as a carbon, nitrogen, and/or energy source may play important roles in association with eukaryotes, including pathogenesis, symbioses, and nutrient cycling. We sought to generate choline analogues to study bacterial choline catabolismin vitroandin situ. Here we report the characterization of a choline analogue, propargylcholine, which inhibits choline catabolism at the level of Dgc enzyme-catalyzed dimethylglycine demethylation inPseudomonas aeruginosa. We used genetic analyses and13C nuclear magnetic resonance to demonstrate that propargylcholine is catabolized to its inhibitory form, propargylmethylglycine. Chemically synthesized propargylmethylglycine was also an inhibitor of growth on choline. Bioinformatic analysis suggests that there are genes encoding DgcA homologues in a variety of proteobacteria. We examined the broader utility of propargylcholine and propargylmethylglycine by assessing growth of other members of the proteobacteria that are known to grow on choline and possess putative DgcA homologues. Propargylcholine showed utility as a growth inhibitor inP. aeruginosabut did not inhibit growth in other proteobacteria tested. In contrast, propargylmethylglycine was able to inhibit choline-dependent growth in all tested proteobacteria, includingPseudomonas mendocina,Pseudomonas fluorescens,Pseudomonas putida,Burkholderia cepacia,Burkholderia ambifaria, andSinorhizobium meliloti. We predict that chemical inhibitors of choline catabolism will be useful for studying this pathway in clinical and environmental isolates and could be a useful tool to study proteobacterial choline catabolismin situ.

scholarly journals Stability of a Pseudomonas putida KT2440 Bacteriophage-Carried Genomic Island and Its Impact on Rhizosphere Fitness

2012 ◽  
Vol 78 (19) ◽  
pp. 6963-6974 ◽  
Author(s):  
Jose M. Quesada ◽  
María Isabel Soriano ◽  
Manuel Espinosa-Urgel
Keyword(s):  
Pseudomonas Putida ◽  
Genomic Island ◽  
Mobile Element ◽  
Direct Repeat ◽  
Promoter Sequence ◽  
Repeat Sequence ◽  
Genomic Islands ◽  
Pseudomonas Putida Kt2440 ◽  
Bacterial Populations ◽  
Content Type

ABSTRACTThe stability of seven genomic islands ofPseudomonas putidaKT2440 with predicted potential for mobilization was studied in bacterial populations associated with the rhizosphere of corn plants by multiplex PCR. DNA rearrangements were detected for only one of them (GI28), which was lost at high frequency. This genomic island of 39.4 kb, with 53 open reading frames, shows the characteristic organization of genes belonging to tailed phages. We present evidence indicating that it corresponds to the lysogenic state of a functional bacteriophage that we have designated Pspu28. Integrated and rarely excised forms of Pspu28 coexist in KT2440 populations. Pspu28 is self-transmissible, and an excisionase is essential for its removal from the bacterial chromosome. The excised Pspu28 forms a circular element that can integrate into the chromosome at a specific location,attsites containing a 17-bp direct repeat sequence. Excision/insertion of Pspu28 alters the promoter sequence and changes the expression level of PP_1531, which encodes a predicted arsenate reductase. Finally, we show that the presence of Pspu28 in the lysogenic state has a negative effect on bacterial fitness in the rhizosphere under conditions of intraspecific competition, thus explaining why clones having lost this mobile element are recovered from that environment.

scholarly journals Mechanisms of Resistance to Chloramphenicol in Pseudomonas putida KT2440

2011 ◽  
Vol 56 (2) ◽  
pp. 1001-1009 ◽  
Author(s):  
Matilde Fernández ◽  
Susana Conde ◽  
Jesús de la Torre ◽  
Carlos Molina-Santiago ◽  
Juan-Luis Ramos ◽  
...  
Keyword(s):  
Pseudomonas Putida ◽  
Efflux Pump ◽  
Protein Biosynthesis ◽  
Cellular Stress Response ◽  
Knockout Mutant ◽  
Pseudomonas Putida Kt2440 ◽  
Altered Expression ◽  
Content Type ◽  
Reading Frame ◽  
A Genome

ABSTRACTPseudomonas putidaKT2440 is a chloramphenicol-resistant bacterium that is able to grow in the presence of this antibiotic at a concentration of up to 25 μg/ml. Transcriptomic analyses revealed that the expression profile of 102 genes changed in response to this concentration of chloramphenicol in the culture medium. The genes that showed altered expression include those involved in general metabolism, cellular stress response, gene regulation, efflux pump transporters, and protein biosynthesis. Analysis of a genome-wide collection of mutants showed that survival of a knockout mutant in the TtgABC resistance-nodulation-division (RND) efflux pump and mutants in the biosynthesis of pyrroloquinoline (PQQ) were compromised in the presence of chloramphenicol. The analysis also revealed that an ABC extrusion system (PP2669/PP2668/PP2667) and the AgmR regulator (PP2665) were needed for full resistance toward chloramphenicol. Transcriptional arrays revealed that AgmR controls the expression of thepqqgenes and the operon encoding the ABC extrusion pump from the promoter upstream of open reading frame (ORF) PP2669.

scholarly journals Contribution of Bacillomycin D in Bacillus amyloliquefaciens SQR9 to Antifungal Activity and Biofilm Formation

2012 ◽  
Vol 79 (3) ◽  
pp. 808-815 ◽  
Author(s):  
Zhihui Xu ◽  
Jiahui Shao ◽  
Bing Li ◽  
Xin Yan ◽  
Qirong Shen ◽  
...  
Keyword(s):  
Biofilm Formation ◽  
Bacillus Amyloliquefaciens ◽  
Root Colonization ◽  
Antagonistic Activity ◽  
Vital Role ◽  
Content Type ◽  
Reverse Transcription Pcr ◽  
Bacillomycin D

ABSTRACTBacillus amyloliquefaciensstrains are capable of suppressing soilborne pathogens through the secretion of an array of lipopeptides and root colonization, and biofilm formation ability is considered a prerequisite for efficient root colonization. In this study, we report that one of the lipopeptide compounds (bacillomycin D) produced by the rhizosphere strainBacillus amyloliquefaciensSQR9 not only plays a vital role in the antagonistic activity againstFusarium oxysporumbut also affects the expression of the genes involved in biofilm formation. When the bacillomycin D and fengycin synthesis pathways were individually disrupted, mutant SQR9M1, which was deficient in the production of bacillomycin D, only showed minor antagonistic activity againstF. oxysporum, but another mutant, SQR9M2, which was deficient in production of fengycin, showed antagonistic activity equivalent to that of the wild-type strain ofB. amyloliquefaciensSQR9. The results fromin vitro, rootin situ, and quantitative reverse transcription-PCR studies demonstrated that bacillomycin D contributes to the establishment of biofilms. Interestingly, the addition of bacillomycin D could significantly increase the expression levels ofkinCgene, but KinC activation is not triggered by leaking of potassium. These findings suggest that bacillomycin D contributes not only to biocontrol activity but also to biofilm formation in strainB. amyloliquefaciensSQR9.

scholarly journals Analysis of Pseudomonas putida KT2440 Gene Expression in the Maize Rhizosphere: In Vitro Expression Technology Capture and Identification of Root-Activated Promoters

2005 ◽  
Vol 187 (12) ◽  
pp. 4033-4041 ◽  
Author(s):  
María Isabel Ramos-González ◽  
María Jesús Campos ◽  
Juan L. Ramos
Keyword(s):  
Pseudomonas Putida ◽  
Defense Mechanism ◽  
Essential Gene ◽  
Cell Envelope ◽  
Original Position ◽  
Pseudomonas Putida Kt2440 ◽  
Maize Rhizosphere ◽  
Semialdehyde Dehydrogenase

ABSTRACT 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-β-semialdehyde dehydrogenase (asd), which was unable to survive in the rhizosphere, and plasmid pOR1, which carries a promoterless 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.

scholarly journals In situ enhancement of pulmonary surfactant function using temporary flow reversal

2012 ◽  
Vol 112 (1) ◽  
pp. 149-158 ◽  
Author(s):  
Henry W. Glindmeyer ◽  
Bradford J. Smith ◽  
Donald P. Gaver
Keyword(s):  
Fluid Dynamic ◽  
The United States ◽  
Cellular Damage ◽  
Dynamic Simulations ◽  
Airway Patency ◽  
Surfactant Replacement Therapy ◽  
Starting Point ◽  
Microscale Transport

Acute respiratory distress syndrome is a pulmonary disease with a mortality rate of ∼40% and 75,000 deaths annually in the United States. Mechanical ventilation restores airway patency and gas transport but leads to ventilator-induced lung injury. Furthermore, surfactant replacement therapy is ineffective due to surfactant delivery difficulties and deactivation by vascular proteins leaking into the airspace. Here, we demonstrated that surfactant function can be substantially improved (up to 50%) in situ in an in vitro pulmonary airway model using unconventional flows that incorporate a short-term retraction of the air-liquid interface, leading to a net decrease in cellular damage. Computational fluid dynamic simulations provided insights into this method and demonstrated the physicochemical hydrodynamic foundation for the improved surfactant microscale transport and mobility. This study may provide a starting point for developing novel ventilation waveforms to improve surfactant function in edematous airways.

scholarly journals Outer membrane vesicles catabolize lignin-derived aromatic compounds in Pseudomonas putida KT2440

2020 ◽  
Vol 117 (17) ◽  
pp. 9302-9310 ◽  
Author(s):  
Davinia Salvachúa ◽  
Allison Z. Werner ◽  
Isabel Pardo ◽  
Martyna Michalska ◽  
Brenna A. Black ◽  
...  
Keyword(s):  
Pseudomonas Putida ◽  
Outer Membrane ◽  
Aromatic Compounds ◽  
Value Added ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
Pseudomonas Putida Kt2440 ◽  
Rich Media

Lignin is an abundant and recalcitrant component of plant cell walls. While lignin degradation in nature is typically attributed to fungi, growing evidence suggests that bacteria also catabolize this complex biopolymer. However, the spatiotemporal mechanisms for lignin catabolism remain unclear. Improved understanding of this biological process would aid in our collective knowledge of both carbon cycling and microbial strategies to valorize lignin to value-added compounds. Here, we examine lignin modifications and the exoproteome of three aromatic–catabolic bacteria: Pseudomonas putida KT2440, Rhodoccocus jostii RHA1, and Amycolatopsis sp. ATCC 39116. P. putida cultivation in lignin-rich media is characterized by an abundant exoproteome that is dynamically and selectively packaged into outer membrane vesicles (OMVs). Interestingly, many enzymes known to exhibit activity toward lignin-derived aromatic compounds are enriched in OMVs from early to late stationary phase, corresponding to the shift from bioavailable carbon to oligomeric lignin as a carbon source. In vivo and in vitro experiments demonstrate that enzymes contained in the OMVs are active and catabolize aromatic compounds. Taken together, this work supports OMV-mediated catabolism of lignin-derived aromatic compounds as an extracellular strategy for nutrient acquisition by soil bacteria and suggests that OMVs could potentially be useful tools for synthetic biology and biotechnological applications.

scholarly journals The putative phosphate transporter PitB (PP1373) is involved in tellurite uptake in Pseudomonas putida KT2440

Microbiology ◽  
2020 ◽  
Author(s):  
Rafael Montenegro ◽  
Sofía Vieto ◽  
Daniela Wicki-Emmenegger ◽  
Felipe Vásquez-Castro ◽  
Carolina Coronado-Ruiz ◽  
...  
Keyword(s):  
Pseudomonas Putida ◽  
Type Strain ◽  
Type Species ◽  
Wild Type Strain ◽  
Phosphate Transporter ◽  
Transport Systems ◽  
Wild Type ◽  
Pseudomonas Putida Kt2440 ◽  
Content Type ◽  
Link Type

Tellurium oxyanions are chemical species of great toxicity and their presence in the environment has increased because of mining industries and photovoltaic and electronic waste. Recovery strategies for this metalloid that are based on micro-organisms are of interest, but further studies of the transport systems and enzymes responsible for implementing tellurium transformations are required because many mechanisms remain unknown. Here, we investigated the involvement in tellurite uptake of the putative phosphate transporter PitB (PP1373) in soil bacterium Pseudomonas putida KT2440. For this purpose, through a method based on the CRISPR/Cas9 system, we generated a strain deficient in the pitB gene and characterized its phenotype on exposing it to varied concentrations of tellurite. Growth curves and transmission electronic microscopy experiments for the wild-type and ΔpitB strains showed that both were able to internalize tellurite into the cytoplasm and reduce the oxyanion to black nano-sized and rod-shaped tellurium particles, although the ΔpitB strain showed an increased resistance to the tellurite toxic effects. At a concentration of 100 μM tellurite, where the biomass formation of the wild-type strain decreased by half, we observed a greater ability of ΔpitB to reduce this oxyanion with respect to the wild-type strain (~38 vs ~16 %), which is related to the greater biomass production of ΔpitB and not to a greater consumption of tellurite per cell. The phenotype of the mutant was restored on over-expressing pitB in trans. In summary, our results indicate that PitB is one of several transporters responsible for tellurite uptake in P. putida KT2440.

scholarly journals Effects of Three Different Nucleoid-Associated Proteins Encoded on IncP-7 Plasmid pCAR1 on Host Pseudomonas putida KT2440

2015 ◽  
Vol 81 (8) ◽  
pp. 2869-2880 ◽  
Author(s):  
Chiho Suzuki-Minakuchi ◽  
Ryusuke Hirotani ◽  
Masaki Shintani ◽  
Toshiharu Takeda ◽  
Yurika Takahashi ◽  
...  
Keyword(s):  
Pseudomonas Putida ◽  
Structural Instability ◽  
Metabolic Phenotype ◽  
Pseudomonas Putida Kt2440 ◽  
Host Chromosome ◽  
Content Type ◽  
Inorganic Ion ◽  
Segregational Stability ◽  
Genes Encoding ◽  
Associated Proteins

ABSTRACTNucleoid-associated proteins (NAPs), which fold bacterial DNA and influence gene transcription, are considered to be global transcriptional regulators of genes on both plasmids and the host chromosome. Incompatibility P-7 group plasmid pCAR1 carries genes encoding three NAPs: H-NS family protein Pmr, NdpA-like protein Pnd, and HU-like protein Phu. In this study, the effects of single or double disruption ofpmr,pnd, andphuwere assessed in hostPseudomonas putidaKT2440. Whenpmrandpndorpmrandphuwere simultaneously disrupted, both the segregational stability and the structural stability of pCAR1 were markedly decreased, suggesting that Pmr, Pnd, and Phu act as plasmid-stabilizing factors in addition to their established roles in replication and partition systems. The transfer frequency of pCAR1 was significantly decreased in these double mutants. The segregational and structural instability of pCAR1 in the double mutants was recovered by complementation ofpmr, whereas no recovery of transfer deficiency was observed. Comprehensive phenotype comparisons showed that the host metabolism of carbon compounds, which was reduced by pCAR1 carriage, was restored by disruption of the NAP gene(s). Transcriptome analyses of mutants indicated that transcription of genes for energy production, conversion, inorganic ion transport, and metabolism were commonly affected; however, how their products altered the phenotypes of mutants was not clear. The findings of this study indicated that Pmr, Pnd, and Phu act synergistically to affect pCAR1 replication, maintenance, and transfer, as well as to alter the host metabolic phenotype.

scholarly journals Response of Pseudomonas putida KT2440 to Increased NADH and ATP Demand

2011 ◽  
Vol 77 (18) ◽  
pp. 6597-6605 ◽  
Author(s):  
Birgitta E. Ebert ◽  
Felix Kurth ◽  
Marcel Grund ◽  
Lars M. Blank ◽  
Andreas Schmid
Keyword(s):  
Pseudomonas Putida ◽  
Atp Hydrolysis ◽  
Nadh Oxidase ◽  
Cell Metabolism ◽  
Pseudomonas Putida Kt2440 ◽  
Nadh Regeneration ◽  
Regeneration Rate ◽  
Content Type ◽  
Oxidation Rates ◽  
The Impact

ABSTRACTAdenosine phosphate and NAD cofactors play a vital role in the operation of cell metabolism, and their levels and ratios are carefully regulated in tight ranges. Perturbations of the consumption of these metabolites might have a great impact on cell metabolism and physiology. Here, we investigated the impact of increased ATP hydrolysis and NADH oxidation rates on the metabolism ofPseudomonas putidaKT2440 by titration of 2,4-dinitrophenol (DNP) and overproduction of a water-forming NADH oxidase, respectively. Both perturbations resulted in a reduction of the biomass yield and, as a consequence of the uncoupling of catabolic and anabolic activities, in an amplification of the net NADH regeneration rate. However, a stimulation of the specific carbon uptake rate was observed only whenP. putidawas challenged with very high 2,4-dinitrophenol concentrations and was comparatively unaffected by recombinant NADH oxidase activity. This behavior contrasts with the comparably sensitive performance described, for example, forEscherichia coliorSaccharomyces cerevisiae. The apparent robustness ofP. putidametabolism indicates that it possesses a certain buffering capacity and a high flexibility to adapt to and counteract different stresses without showing a distinct phenotype. These findings are important, e.g., for the development of whole-cell redox biocatalytic processes that impose equivalent burdens on the cell metabolism: stoichiometric consumption of (reduced) redox cofactors and increased energy expenditures, due to the toxicity of the biocatalytic compounds.

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