scholarly journals Multiple Roles for Two Efflux Pumps in the Polycyclic Aromatic Hydrocarbon-Degrading Pseudomonas putida Strain B6-2 (DSM 28064)

2017 ◽  
Vol 83 (24) ◽  
Author(s):  
Xuemei Yao ◽  
Fei Tao ◽  
Kunzhi Zhang ◽  
Hongzhi Tang ◽  
Ping Xu
Keyword(s):  
Antibiotic Resistance ◽  
Polycyclic Aromatic Hydrocarbon ◽  
Aromatic Hydrocarbon ◽  
Pseudomonas Putida ◽  
Mutant Strain ◽  
Efflux Pump ◽  
Efflux Pumps ◽  
Content Type ◽  
Pah Degradation ◽  
Polycyclic Aromatic

ABSTRACTMicrobial bioremediation is a promising approach for the removal of polycyclic aromatic hydrocarbon (PAH) contaminants. Many degraders of PAHs possess efflux pump genes in their genomes; however, their specific roles in the degradation of PAHs have not been clearly elucidated. In this study, two efflux pumps, TtgABC and SrpABC, were systematically investigated to determine their functions in a PAH-degradingPseudomonas putidastrain B6-2 (DSM 28064). The disruption of genesttgABCorsrpABCresulted in a defect in organic solvent tolerance. TtgABC was found to contribute to antibiotic resistance; SrpABC only contributed to antibiotic resistance under an artificial overproduced condition. Moreover, a mutant strain withoutsrpABCdid not maintain its activity in long-term biphenyl (BP) degradation, which correlated with the loss of cell viability. The expression of SrpABC was significantly upregulated in the course of BP degradation. BP, 2-hydroxybiphenyl, 3-hydroxybiphenyl, and 2,3-dihydroxybiphenyl (2,3-DHBP) were revealed to be the inducers ofsrpABC. 2,3-DHBP was verified to be a substrate of pump SrpABC; SrpABC can enhance the tolerance to 2,3-DHBP by pumping it out. The mutant strain B6-2ΔsrpSprolonged BP degradation with the increase ofsrpABCexpression. These results suggest that the pump SrpABC of strain B6-2 plays a positive role in BP biodegradation by pumping out metabolized toxic substances such as 2,3-DHBP. This study provides insights into the versatile physiological functions of the widely distributed efflux pumps in the biodegradation of PAHs.IMPORTANCEPolycyclic aromatic hydrocarbons (PAHs) are notorious for their recalcitrance to degradation in the environment. A high frequency of the occurrence of the efflux pump genes was observed in the genomes of effective PAH degraders; however, their specific roles in the degradation of PAHs are still obscure. The significance of our study is in the identification of the function and mechanism of the efflux pump SrpABC ofPseudomonas putidastrain B6-2 (DSM 28064) in the biphenyl degradation process. SrpABC is crucial for releasing the toxicity caused by intermediates that are unavoidably produced in PAH degradation, which enables an understanding of how cells maintain the intracellular balance of materials. The findings from this study provide a new perspective on PAH recalcitrance and shed light on enhancing PAH degradation by genetic engineering.

scholarly journals Complete Genome Sequence of Pseudomonas putida BS3701, a Promising Polycyclic Aromatic Hydrocarbon-Degrading Strain for Bioremediation Technologies

2020 ◽  
Vol 9 (40) ◽  
Author(s):  
Andrey Filonov ◽  
Yanina Delegan ◽  
Irina Puntus ◽  
Leonid Valentovich ◽  
Artur Akhremchuk ◽  
...  
Keyword(s):  
Polycyclic Aromatic Hydrocarbons ◽  
Polycyclic Aromatic Hydrocarbon ◽  
Aromatic Hydrocarbon ◽  
Pseudomonas Putida ◽  
Russian Federation ◽  
Aromatic Hydrocarbons ◽  
Complete Genome Sequence ◽  
Circular Chromosome ◽  
Content Type ◽  
Polycyclic Aromatic

ABSTRACT The strain Pseudomonas putida BS3701 was isolated from soil contaminated with coke by-product waste (Moscow Region, Russian Federation). It is capable of degrading crude oil and polycyclic aromatic hydrocarbons (PAHs). The P. putida BS3701 genome consists of a 6,337,358-bp circular chromosome and two circular plasmids (pBS1141 with 107,388 bp and pBS1142 with 54,501 bp).

scholarly journals Knockout of Extracytoplasmic Function Sigma Factor ECF-10 Affects Stress Resistance and Biofilm Formation in Pseudomonas putida KT2440

2014 ◽  
Vol 80 (16) ◽  
pp. 4911-4919 ◽  
Author(s):  
Beatrix Tettmann ◽  
Andreas Dötsch ◽  
Olivier Armant ◽  
Christopher D. Fjell ◽  
Joerg Overhage
Keyword(s):  
Antibiotic Resistance ◽  
Pseudomonas Putida ◽  
Mutant Strain ◽  
Stress Resistance ◽  
Sigma Factor ◽  
Biofilm Formation ◽  
Efflux Pump ◽  
Pseudomonas Putida Kt2440 ◽  
Content Type ◽  
Extracytoplasmic Function Sigma Factor

ABSTRACTPseudomonas putidais a Gram-negative soil bacterium which is well-known for its versatile lifestyle, controlled by a large repertoire of transcriptional regulators. Besides one- and two-component regulatory systems, the genome ofP. putidareveals 19 extracytoplasmic function (ECF) sigma factors involved in the adaptation to changing environmental conditions. In this study, we demonstrate that knockout of extracytoplasmic function sigma factor ECF-10, encoded by open reading frame PP4553, resulted in 2- to 4-fold increased antibiotic resistance to quinolone, β-lactam, sulfonamide, and chloramphenicol antibiotics. In addition, the ECF-10 mutant exhibited enhanced formation of biofilms after 24 h of incubation. Transcriptome analysis using Illumina sequencing technology resulted in the detection of 12 genes differentially expressed (>2-fold) in the ECF-10 knockout mutant strain compared to their levels of expression in wild-type cells. Among the upregulated genes werettgA,ttgB, andttgC, which code for the major multidrug efflux pump TtgABC inP. putidaKT2440. Investigation of an ECF-10 andttgAdouble-knockout strain and attgABC-overexpressing strain demonstrated the involvement of efflux pump TtgABC in the stress resistance and biofilm formation phenotypes of the ECF-10 mutant strain, indicating a new role for this efflux pump beyond simple antibiotic resistance inP. putidaKT2440.

scholarly journals Heterologous Expression of Polycyclic Aromatic Hydrocarbon Ring-Hydroxylating Dioxygenase Genes from a Novel Pyrene-Degrading Betaproteobacterium

2012 ◽  
Vol 78 (10) ◽  
pp. 3552-3559 ◽  
Author(s):  
David R. Singleton ◽  
Jing Hu ◽  
Michael D. Aitken
Keyword(s):  
Polycyclic Aromatic Hydrocarbon ◽  
Aromatic Hydrocarbon ◽  
Enrichment Culture ◽  
Metagenomic Library ◽  
Stable Isotope Probing ◽  
Content Type ◽  
Close Proximity ◽  
Polycyclic Aromatic ◽  
Dominant Member ◽  
Pah Metabolism

ABSTRACTA betaproteobacterium within the familyRhodocyclaceaepreviously identified as a pyrene degrader via stable-isotope probing (SIP) of contaminated soil (designated pyrene group 1 or PG1) was cultivated as the dominant member of a mixed bacterial culture. A metagenomic library was constructed, and the largest contigs were analyzed for genes associated with polycyclic aromatic hydrocarbon (PAH) metabolism. Eight pairs of genes with similarity to the α- and β-subunits of ring-hydroxylating dioxygenases (RHDs) associated with aerobic bacterial PAH degradation were identified and linked to PG1 through PCR analyses of a simplified enrichment culture. In tandem with a ferredoxin and reductase found in close proximity to one pair of RHD genes, six of the RHDs were cloned and expressed inEscherichia coli. Each cloned RHD was tested for activity against nine PAHs ranging in size from two to five rings. Despite differences in their predicted protein sequences, each of the six RHDs was capable of transforming phenanthrene and pyrene. Three RHDs could additionally transform naphthalene and fluorene, and these genotypes were also associated with the ability of theE. coliconstructs to convert indole to indigo. Only one of the six cloned RHDs was capable of transforming anthracene and benz[a]anthracene. None of the tested RHDs were capable of significantly transforming fluoranthene, chrysene, or benzo[a]pyrene.

scholarly journals Complete Genome Sequence of the Pyrene-Degrading Bacterium Cycloclasticus sp. Strain P1

2012 ◽  
Vol 194 (23) ◽  
pp. 6677-6677 ◽  
Author(s):  
Qiliang Lai ◽  
Weiwei Li ◽  
Baojiang Wang ◽  
Zhiwei Yu ◽  
Zongze Shao
Keyword(s):  
Polycyclic Aromatic Hydrocarbon ◽  
Aromatic Hydrocarbon ◽  
Complete Genome Sequence ◽  
Deep Sea ◽  
Complete Genome ◽  
Content Type ◽  
Degrading Bacterium ◽  
The Pacific ◽  
Polycyclic Aromatic ◽  
The Pacific Ocean

ABSTRACTCycloclasticussp. strain P1 was isolated from deep-sea sediments of the Pacific Ocean and characterized as a unique bacterium in the degradation of pyrene, a four-ring polycyclic aromatic hydrocarbon (PAH). Here we report the complete genome of P1 and genes associated with PAH degradation.

Comparative study of five polycyclic aromatic hydrocarbon degrading bacterial strains isolated from contaminated soils

1997 ◽  
Vol 43 (4) ◽  
pp. 368-377 ◽  
Author(s):  
Fadi Dagher ◽  
Eric Déziel ◽  
Patricia Lirette ◽  
Gilles Paquette ◽  
Jean-Guy Bisaillon ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Polycyclic Aromatic Hydrocarbon ◽  
Aromatic Hydrocarbon ◽  
Contaminated Soils ◽  
Bacterial Strains ◽  
Chain Reaction ◽  
Pah Degradation ◽  
Catabolic Genes ◽  
Polycyclic Aromatic ◽  
Polymerase Chain

Five polycyclic aromatic hydrocarbon (PAH) degrading bacterial strains, Pseudomonas putida 34, Pseudomonas fluorescens 62, Pseudomonas aeruginosa 57, Sphingomonas sp. strain 107, and the unidentified strain PL1, were isolated from two contaminated soils and characterized for specific features regarding PAH degradation. Degradation efficiency was determined by the rapidity to form clearing zones around colonies when sprayed with different PAH solutions and the growth in liquid medium with different PAHs as sole source of carbon and energy. The presence of plasmids, the production of biosurfactants, the effect of salicylate on PAH degradation, the transformation of indole to indigo indicating the presence of an aromatic ring dioxygenase activity, and the hybridization with the SphAb probe representing a sequence highly homologous to the naphthalene dioxygenase ferredoxin gene nahAb were examined. The most efficient strain in terms of substrate specificity and rapidity to degrade different PAHs was Sphingomonas sp. strain 107, followed by strain PL1 and P. aeruginosa 57. The less efficient strains were P. putida 34 and P. fluorescens 62. Each strain transformed indole to indigo, except strain PL1. Biosurfactants were produced by P. aeruginosa 57 and P. putida 34, and a bioemulsifier was produced by Sphingomonas sp. strain 107. The presence of salicylate in solid medium has accelerated the formation of clearing zones and the transformation of indole by Sphingomonas sp. strain 107 and P. aeruginosa 57 colonies. Plasmids were found in Sphingomonas sp. strain 107 and strain PL1. The SphAb probe hybridized with DNA extracted from each strain. However, hybridization signals were detected only in the plasmidic fraction of Sphingomonas sp. strain 107 and strain PL1. Using a polymerase chain reaction (PCR) approach, we determined that several genes encoding enzymes involved in the upper catabolic pathway of naphthalene were present in each strain. Sequencing of PCR DNA fragments revealed that, for all the five strains, these genes are highly homologous with respective genes found in the pah, dox, and nah opérons, and are arranged in a polycistronic operon. Results suggest that these genes are ordered in the five selected strains like the pah, nah, and dox opérons.Key words: polycyclic aromatic hydrocarbons, biodegradation, polymerase chain reaction, naphthalene catabolic genes.

scholarly journals Draft Genome Sequence of Marinobacter hydrocarbonoclasticus Strain STW2, a Polycyclic Aromatic Hydrocarbon-Degrading and Denitrifying Bacterium from the Rhizosphere of Seagrass Enhalus acodoides

2017 ◽  
Vol 5 (8) ◽  
Author(s):  
Juan Ling ◽  
Liyun Lin ◽  
Yanying Zhang ◽  
Xiancheng Lin ◽  
Manzoor Ahamad ◽  
...  
Keyword(s):  
Polycyclic Aromatic Hydrocarbon ◽  
Aromatic Hydrocarbon ◽  
Genome Sequence ◽  
Draft Genome ◽  
Draft Genome Sequence ◽  
Marine Microbes ◽  
Content Type ◽  
Future Studies ◽  
Polycyclic Aromatic ◽  
Marinobacter Hydrocarbonoclasticus

ABSTRACT Here, we report the draft genome sequence of Marinobacter hydrocarbonoclasticus strain STW2, which was isolated from the rhizosphere of seagrass Enhalus acodoides. This study will facilitate future studies on the genetic pathways of marine microbes capable of both polycyclic aromatic hydrocarbon degradation and nitrate reduction.

scholarly journals Isolation and Characterization of Polycyclic Aromatic Hydrocarbon-Degrading Bacteria Associated with the Rhizosphere of Salt Marsh Plants

2001 ◽  
Vol 67 (6) ◽  
pp. 2683-2691 ◽  
Author(s):  
L. L. Daane ◽  
I. Harjono ◽  
G. J. Zylstra ◽  
M. M. Häggblom
Keyword(s):  
Salt Marsh ◽  
Polycyclic Aromatic Hydrocarbon ◽  
Aromatic Hydrocarbon ◽  
Rrna Gene ◽  
Gram Positive ◽  
Salt Marsh Plants ◽  
Pah Degradation ◽  
Isolation And Characterization ◽  
Degrading Bacteria ◽  
Polycyclic Aromatic

ABSTRACT Polycyclic aromatic hydrocarbon (PAH)-degrading bacteria were isolated from contaminated estuarine sediment and salt marsh rhizosphere by enrichment using either naphthalene, phenanthrene, or biphenyl as the sole source of carbon and energy. Pasteurization of samples prior to enrichment resulted in isolation of gram-positive, spore-forming bacteria. The isolates were characterized using a variety of phenotypic, morphologic, and molecular properties. Identification of the isolates based on their fatty acid profiles and partial 16S rRNA gene sequences assigned them to three main bacterial groups: gram-negative pseudomonads; gram-positive, non-spore-forming nocardioforms; and the gram-positive, spore-forming group,Paenibacillus. Genomic digest patterns of all isolates were used to determine unique isolates, and representatives from each bacterial group were chosen for further investigation. Southern hybridization was performed using genes for PAH degradation fromPseudomonas putida NCIB 9816-4, Comamonas testosteroni GZ42, Sphingomonas yanoikuyae B1, andMycobacterium sp. strain PY01. None of the isolates from the three groups showed homology to the B1 genes, only two nocardioform isolates showed homology to the PY01 genes, and only members of the pseudomonad group showed homology to the NCIB 9816-4 or GZ42 probes. The Paenibacillus isolates showed no homology to any of the tested gene probes, indicating the possibility of novel genes for PAH degradation. Pure culture substrate utilization experiments using several selected isolates from each of the three groups showed that the phenanthrene-enriched isolates are able to utilize a greater number of PAHs than are the naphthalene-enriched isolates. Inoculating two of the gram-positive isolates to a marine sediment slurry spiked with a mixture of PAHs (naphthalene, fluorene, phenanthrene, and pyrene) and biphenyl resulted in rapid transformation of pyrene, in addition to the two- and three-ringed PAHs and biphenyl. This study indicates that the rhizosphere of salt marsh plants contains a diverse population of PAH-degrading bacteria, and the use of plant-associated microorganisms has the potential for bioremediation of contaminated sediments.

scholarly journals Vitamin K3 Induces the Expression of the Stenotrophomonas maltophilia SmeVWX Multidrug Efflux Pump

2017 ◽  
Vol 61 (5) ◽  
Author(s):  
P. Blanco ◽  
F. Corona ◽  
M. B. Sánchez ◽  
J. L. Martínez
Keyword(s):  
Antibiotic Resistance ◽  
Vitamin K ◽  
Stenotrophomonas Maltophilia ◽  
Fluorescent Protein ◽  
Efflux Pump ◽  
Yellow Fluorescent Protein ◽  
Efflux Pumps ◽  
Fluorescence Signal ◽  
Multidrug Efflux ◽  
Content Type

ABSTRACT Stenotrophomonas maltophilia is an opportunistic pathogen with increasing prevalence, which is able to cause infections in immunocompromised patients or in those with a previous pathology. The treatment of the infections caused by this bacterium is often complicated due to the several intrinsic antibiotic resistance mechanisms that it presents. Multidrug efflux pumps are among the best-studied mechanisms of S. maltophilia antibiotic resistance. Some of these efflux pumps have a basal expression level but, in general, their expression is often low and only reaches high levels when the local regulator is mutated or bacteria are in the presence of an effector. In the current work, we have developed a yellow fluorescent protein (YFP)-based sensor with the aim to identify effectors able to trigger the expression of SmeVWX, an efflux pump that confers resistance to quinolones, chloramphenicol, and tetracycline when it is expressed at high levels. With this purpose in mind, we tested a variety of different compounds and analyzed the fluorescence signal given by the expression of YFP under the control of the smeVWX promoter. Among the tested compounds, vitamin K3, which is a compound belonging to the 2-methyl-1,4-naphthoquinone family, is produced by plants in defense against infection, and has increasing importance in human therapy, was able to induce the expression of the SmeVWX efflux pump. In addition, a decrease in the susceptibility of S. maltophilia to ofloxacin and chloramphenicol was observed in the presence of vitamin K3, in both wild-type and smeW-deficient strains.

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