Degradation Of Dibenzofuran Research Articles

Characterizing the Contaminant-Adhesion of a Dibenzofuran Degrader Rhodococcus sp.

The adhesion between dibenzofuran (DF) and degrading bacteria is the first step of DF biodegradation and affects the efficient degradation of DF. However, their efficient adhesion mechanism at the molecular level remains unclear. Therefore, this study first examined the adhesive behaviors and molecular mechanisms of Rhodococcus sp. strain p52 upon exposure to DF. The results showed that the adhesion between strain p52 and DF is mediated by extracellular polymeric substances (EPSs). Compared with sodium acetate as a carbon source, the percentages of glucose and proteins related to electron transfer, toxin-antitoxin, and stress responses were elevated, which were analyzed by polysaccharide composition and proteomics, and the contents of extracellular polysaccharides and proteins were increased. Moreover, biofilm analysis suggested an increase in EPS content, and the change in components increased biofilm yield and promoted loose and porous aggregation between the bacteria; this aggregation caused an increase in the specific surface area in contact with DF. The surface characteristics analysis indicated that the production of EPS reduced the absolute value of the zeta potential and increased the hydrophobicity of strain p52, which was beneficial for the adhesion of strain p52 and DF. These findings help us to enhance the understanding of the adhesion mechanisms and bioremediation of polycyclic aromatic hydrocarbons by degrading bacteria.

Mechanisms of nano zero-valent iron in enhancing dibenzofuran degradation by a Rhodococcus sp.: Trade-offs between ATP production and protection against reactive oxygen species

Nano zero-valent iron (nZVI) can enhance pollutants biodegradation, but it displays toxicity towards microorganisms. Gram-positive (G+) bacteria exhibit greater resistance to nZVI than Gram-negative bacteria. However, mechanisms of nZVI accelerating pollutants degradation by G+ bacteria remain unclear. Herein, we explored effects of nZVI on a G+ bacterium, Rhodococcus sp. strain p52, and mechanisms by which nZVI accelerates biodegradation of dibenzofuran, a typical polycyclic aromatic compound. Electron microscopy and energy dispersive spectroscopy analysis revealed that nZVI could penetrate cell membranes, which caused damage and growth inhibition. nZVI promoted dibenzofuran biodegradation at certain concentrations, while higher concentration functioned later due to the delayed reactive oxygen species (ROS) mitigation. Transcriptomic analysis revealed that cells adopted response mechanisms to handle the elevated ROS induced by nZVI. ATP production was enhanced by accelerated dibenzofuran degradation, providing energy for protein synthesis related to antioxidant stress and damage repair. Meanwhile, electron transport chain (ETC) was adjusted to mitigate ROS accumulation, which involved downregulating expression of ETC complex I-related genes, as well as upregulating expression of the genes for the ROS-scavenging cytochrome bd complex and ETC complex II. These findings revealed the mechanisms underlying nZVI-enhanced biodegradation by G+ bacteria, offering insights into optimizing bioremediation strategies involving nZVI.

Different Roles of Dioxin-Catabolic Plasmids in Growth, Biofilm Formation, and Metabolism of Rhodococcus sp. Strain p52.

Microorganisms harbor catabolic plasmids to tackle refractory organic pollutants, which is crucial for bioremediation and ecosystem health. Understanding the impacts of plasmids on hosts provides insights into the behavior and adaptation of degrading bacteria in the environment. Here, we examined alterations in the physiological properties and gene expression profiles of Rhodococcus sp. strain p52 after losing two conjugative dioxin-catabolic megaplasmids (pDF01 and pDF02). The growth of strain p52 accelerated after pDF01 loss, while it decelerated after pDF02 loss. During dibenzofuran degradation, the expression levels of dibenzofuran catabolic genes on pDF01 were higher compared to those on pDF02; accordingly, pDF01 loss markedly slowed dibenzofuran degradation. It was suggested that pDF01 is more beneficial to strain p52 under dibenzofuran exposure. Moreover, plasmid loss decreased biofilm formation, especially after pDF02 loss. Transcriptome profiling revealed different pathways enriched in upregulated and downregulated genes after pDF01 and pDF02 loss, indicating different adaptation mechanisms. Based on the transcriptional activity variation, pDF01 played roles in transcription and anabolic processes, while pDF02 profoundly influenced energy production and cellular defense. This study enhances our knowledge of the impacts of degradative plasmids on native hosts and the adaptation mechanisms of hosts, contributing to the application of plasmid-mediated bioremediation in contaminated environments.

Changes of Nitrate Activity and Byproduct Distribution Characteristics for Synergistic NOx and Dioxin Abatement over V2O5/AC Catalyst.

The simultaneous removal of NOx and dioxins has been considered an economical and effective technology of controlling multipollutant flue gas in the context of "carbon peaking and carbon neutrality". However, this technology has not yet been implemented in practical situations, because the interactive relationship between the selective catalytic reduction (SCR) reaction and dioxin catalytic oxidation lacks a deep understanding, especially on a carbon-based catalyst. In this research, the influence of NO and NH3 on the oxidation characteristics and byproducts distribution of dibenzofuran (DBF) was studied on V2O5/AC catalyst. Results indicated that NH3 has a stronger inhibition effect for DBF catalytic oxidation than NO due to obvious competitive adsorption between NH3 and DBF on the V2O5/AC catalyst. In addition, although both NO and NH3 inhibit the complete degradation of DBF, their effects on the byproduct distribution are not consistent. NO primarily affects the level of oxygen-containing byproducts, while NH3 primarily affects the level of alkane byproducts. Furthermore, the SCR reaction activity demonstrated a reduction when DBF was present. The occupation of V2O5 sites by DBF and its oxidizing intermediates has hindered the production of monodentate nitrate and the reactivity of bridged nitrate, resulting in a decrease in SCR activity via the L-H mechanism. This work aims to provide theoretical guidance for simultaneous removal of NOx and dioxins in industrial fumes.

Metabolic cross-feeding between the competent degrader Rhodococcus sp. strain p52 and an incompetent partner during catabolism of dibenzofuran: Understanding the leading and supporting roles

Microbial interactions, particularly metabolic cross-feeding, play important roles in removing recalcitrant environmental pollutants; however, the underlying mechanisms involved in this process remain unclear. Thus, this study aimed to elucidate the mechanism by which metabolic cross-feeding occurs during synergistic dibenzofuran degradation between a highly efficient degrader, Rhodococcus sp. strain p52, and a partner incapable of utilizing dibenzofuran. A bottom-up approach combined with pairwise coculturing was used to examine metabolic cross-feeding between strain p52 and Arthrobacter sp. W06 or Achromobacter sp. D10. Pairwise coculture not only promoted bacterial pair growth but also facilitated dibenzofuran degradation. Specifically, strain p52, acting as a donor, released dibenzofuran metabolic intermediates, including salicylic acid and gentisic acid, for utilization and growth, respectively, by the partner strains W06 and D10. Both salicylic acid and gentisic acid exhibited biotoxicity, and their accumulation inhibited dibenzofuran degradation. The transcriptional activity of the genes responsible for the catabolism of dibenzofuran and its metabolic intermediates was coordinately regulated in strain p52 and its cocultivated partners, thus achieving synergistic dibenzofuran degradation. This study provides insights into microbial metabolic cross-feeding during recalcitrant environmental pollutant removal.

Roles of n-hexadecane in the degradation of dibenzofuran by a biosurfactant-producing bacterium Rhodococcus sp.

The hydrophobic nature of dioxins is a major obstacle to their bioremediation. The present study attempted to improve the bioaccessibility of dioxins in two-phase partitioning bioreactors and activated sludge reactors. Dibenzofuran and n-hexadecane were used as model compounds for dioxins and non-aqueous phase liquids (NAPLs), respectively. First, biosurfactants produced by dioxin-degrading Rhodococcus sp. strain p52 were analyzed. Then, the role of n-hexadecane during the biodegradation of dibenzofuran was evaluated. The results showed that high NAPL ratios (n-hexadecane ≥40 g/L) in the two liquid-phase systems favored dibenzofuran removal, while the cell surface of strain p52 became extremely hydrophobic (>97%), which promoted its adhesion to NAPLs and subsequent transport of dibenzofuran from NAPL to the cells. In addition, a bioaugmented activated sludge reactor with n-hexadecane as the supplemented carbon source could not only facilitate the aerobic granular sludge formation but also improve dibenzofuran removal and the activated sludge settling properties. Simultaneously, conjugative transfer of catabolic plasmids was promoted. This study demonstrated the potential application of strain p52 with NAPLs for the bioremediation of dioxins.

Dibenzofuran Degradation by Bacterial Community in Landfill Leachate

The contamination of the environment has been a global issue, and bioremediation is proposed as an option to clean up the contamination sites with the promising utilization of bacterial community capabilities. The indigenous bacterial community in the landfill leachate is recognized to carry enzymes for the degradation of contaminants such as dioxin congeners, the dibenzofuran. Environmental factors have been known to influence the process to achieve successful biodegradation, and the optimized conditions may speed up the biodegradation process. Thus, this study was conducted to optimize the substrate availability, temperature, and pH factor for the degradation of dibenzofuran from landfill leachate by the native bacterial community in landfill leachate. This study uses the one-factor at-time (OFAT) approach to measure dibenzofuran degradation. The landfill leachate with enrichment of dibenzofuran (15 to 45 mg L-1) was incubated at temperatures (30°C to 42°C) and pH (5 to 9) for 24 hours before being extracted and analyzed. From the first part of the study, 15 mg L-1 of dibenzofuran, 30°C temperature, and pH 7 have shown the highest dibenzofuran degradation. Later, the optimum condition of dibenzofuran removal (74.40%) was achieved when the landfill leachate was spiked with 15 ppm dibenzofuran at 30°C and pH 7 for 24 hours. This study proposes optimized conditions that give a better result for dibenzofuran degradation, which may enhance bioremediation.

Unravelling the impacts of sulfur dioxide on dioxin catalytic decomposition on V2O5/AC catalysts

Dioxins are high chlorine, toxic, and persistent organic pollutants that exert significant pressure on both human and the environment. From the analysis of current pollutant removal of the whole life cycle, such as integrated removal of NOx, SO2 and dioxins in a system, the dioxins oxidation activity as well as the distribution of oxidation products in the presence of SO2 are still a challenge. In this study, dibenzofuran (DBF) was regarded as a model dioxin compound, and V2O5/AC was used as a catalyst to investigate the impact of SO2 on degradation activity and the degradation path of DBF. Various characterization results showed that SO2 could promote the transformation of DBF to intermediates through a reaction with lattice oxygen and lower the apparent activated energy of DBF catalytic oxidation on V2O5/AC catalysts. The density functional theory (DFT) calculations confirmed that SO2 improved the oxidation ability of lattice oxygen on V2O5/AC. The ethyl hydrogen fumarate intermediate decreased and the small-molecule byproducts increased, providing further evidence that SO2 accelerates the degradation of DBF and its intermediates. However, the formation of VOSO4 would inevitably deteriorate the adsorption and oxidation abilities of V2O5/AC. A model is pioneered to describe the relationship between SO2 promotion and VOSO4 inhibition on DBF catalytic oxidation on a V2O5/AC catalyst. This study is expected to provide theoretical guidance for the collaborative abatement of multi-pollutants in flue gas.

Improved expression of Catechol 1,2-dioxygenase gene from Burkholderia cepacia in Escherichia coli

Catechol 1,2-dioxygenase (CAT1) is a key enzyme for the ortho-cleavage pathway involved in the degradation of dibenzofuran, a dioxin derivative, which is a highly toxic environmental pollutant. The present study aims to investigate appropriate culture conditions for enhancing the expression of the cat1 gene encoding CAT1 enzyme from Burkholderia cepacia DF4 in Escherichia coli M15. The optimized culture conditions for gene expression are cell density at the time of induction, shaking speed, induction temperature, induction time, and inducer concentration. The highest level for CAT1 was obtained at the IPTG concentration of 1.2 mM, 10 hours after induction at 35 °C, shaking speed 200 rpm with cell density at OD600 0.7.

A Pseudomonas sp. strain uniquely degrades PAHs and heterocyclic derivatives via lateral dioxygenation pathways

Polycyclic aromatic hydrocarbons (PAHs) and heterocyclic derivatives are organic pollutants that pose a serious health risk to human beings. In this study, a newly isolated Pseudomonas brassicacearum strain MPDS could effectively degrade PAHs and heterocyclic derivatives, including naphthalene, fluorene, dibenzofuran (DBF) and dibenzothiophene (DBT). Notably, strain MPDS is able to degrade fluorene, DBF and DBT uniquely via a lateral dioxygenation pathway, while most reported strains degrade fluorene, DBF and DBT via an angular dioxygenation pathway or co-metabolize them via a lateral dioxygenation pathway. Strain MPDS completely degraded 50 mg naphthalene (in 50 mL medium) in 84 h, and OD600 reached 1.0–1.1; while, it stabilized at OD600 0.5–0.6 with 5 mg fluorene or DBF or DBT. Meanwhile, 65.7% DBF and 32.1% DBT were degraded in 96 h, and 40.3% fluorene was degraded in 72 h, respectively. Through genomic and transcriptomic analyses, and comparative genomic analysis with another DBF degradation strain, relevant gene clusters were predicted, and a naphthalene-degrading gene cluster was identified. This study provides understanding of degradation of PAHs and their heterocyclic derivatives, as well as new insights into the lateral dioxygenation pathway of relevant contaminants.

Isolation and characterization of a moderate thermophilic Paenibacillus naphthalenovorans strain 4B1 capable of degrading dibenzofuran from dioxin-contaminated soil in Vietnam

A moderate thermophilic dibenzofuran (DF) degrader, strain 4B1, was isolated from dioxin-contaminated soil in Vietnam under thermophilic condition. A 16S rRNA gene sequence analysis assigned the strain to genus Paenibacillus. The optimum growth temperature of strain 4B1 was 45°C with a doubling time of 2.7h in the presence of DF as a sole carbon and energy source. The rate of its growth and DF-degradation were approximately 3-fold higher than those of a reference Paenibacillus sp. strain. The 4B1 strain degraded 89% of 1000mgL-1 DF within 48h cultivation at the optimum temperature. TBLASTN analysis based on its draft genome sequence revealed that this strain possessed a dbf gene cluster. The open reading frames (dbfA1A2RBC) in the cluster shared 99-100% identity with those of Paenibacillus sp. YK5, indicating that DF was likely degraded by an angular dioxygenation pathway in strain 4B1. Four genes in the dbf gene cluster (dbfA1A2BC) were partially induced by DF, which was observed by semi-quantitative RT-PCR. Quantitative PCR analysis of dbfA1 transcripts, encoding the alpha subunit of DF dioxygenase, indicated that dbfA1 was expressed 4-times higher than that of strain YK5 at 45°C. These results suggest that the faster growth and degradation of DF in strain 4B1 could be due to differences in transcriptional regulation of dbf cluster genes.

Characterization of a Dibenzofuran-degrading strain of Pseudomonas aeruginosa, FA-HZ1

Dibenzofuran (DBF) derivatives have caused serious environmental problems, especially those produced by paper pulp bleaching and incineration processes. Prominent for its resilient mutagenicity and toxicity, DBF poses a major challenge to human health. In the present study, a new strain of Pseudomonas aeruginosa, FA-HZ1, with high DBF-degrading activity was isolated and identified. The determined optimum conditions for cell growth of strain FA-HZ1 were a temperature of 30 °C, pH 5.0, rotation rate of 200 rpm and 0.1 mM DBF as a carbon source. The biochemical and physiological features as well as usage of different carbon sources by FA-HZ1 were studied. The new strain was positive for arginine double hydrolase, gelatinase and citric acid, while it was negative for urease and lysine decarboxylase. It could utilize citric acid as its sole carbon source, but was negative for indole and H2S production. Intermediates of DBF 1,2-dihydroxy-1,2-dihydrodibenzofuran, 1,2-dihydroxydibenzofuran, 2-hydroxy-4-(3′-oxo-3′H-benzofuran-2′-yliden)but-2-enoic acid, 2,3-dihydroxybenzofuran, 2-oxo-2-(2′-hydrophenyl)lactic acid, and 2-hydroxy-2-(2′-hydroxyphenyl)acetic acid were detected and identified through liquid chromatography-mass analyses. FA-HZ1 metabolizes DBF by both the angular and lateral dioxygenation pathways. The genomic study identified 158 genes that were involved in the catabolism of aromatic compounds. To identify the key genes responsible for DBF degradation, a proteomic study was performed. A total of 1459 proteins were identified in strain FA-HZ1, of which 100 were up-regulated and 104 were down-regulated. A novel enzyme “HZ6359 dioxygenase”, was amplified and expressed in pET-28a in E. coli BL21(DE3). The recombinant plasmid was successfully constructed, and was used for further experiments to verify its function. In addition, the strain FA-HZ1 can also degrade halogenated analogues such as 2, 8-dibromo dibenzofuran and 4-(4-bromophenyl) dibenzofuran. Undoubtedly, the isolation and characterization of new strain and the designed pathways is significant, as it could lead to the development of cost-effective and alternative remediation strategies. The degradation pathway of DBF by P. aeruginosa FA-HZ1 is a promising tool of biotechnological and environmental significance.

Cometabolic Degradation of Dibenzofuran and Dibenzothiophene by a Naphthalene-Degrading Comamonas sp. JB.

Comamonas sp. JB was used to investigate the cometabolic degradation of dibenzofuran (DBF) and dibenzothiophene (DBT) with naphthalene as the primary substrate. Dehydrogenase and ATPase activity of the growing system with the presence of DBF and DBT were decreased when compared to only naphthalene in the growing system, indicating that the presence of DBF and DBT inhibited the metabolic activity of strain JB. The pathways and enzymes involved in the cometabolic degradation were tested. Examination of metabolites elucidated that strain JB cometabolically degraded DBF to 1,2-dihydroxydibenzofuran, subsequently to 2-hydroxy-4-(3'-oxo-3'H-benzofuran-2'-yliden)but-2-enoic acid, and finally to catechol. Meanwhile, strain JB cometabolically degraded DBT to 1,2-dihydroxydibenzothiophene and subsequently to the ring cleavage product. A series of naphthalene-degrading enzymes including naphthalene dioxygenase, 1,2-dihydroxynaphthalene dioxygenase, salicylaldehyde dehydrogenase, salicylate hydroxylase, and catechol 2,3-oxygenase have been detected, confirming that naphthalene was the real inducer of expression the degradation enzymes and metabolic pathways were controlled by naphthalene-degrading enzymes.

Complete Genome Sequence of Pseudomonas aeruginosa FA-HZ1, an Efficient Dibenzofuran-Degrading Bacterium.

ABSTRACTPseudomonas sp. FA-HZ1, an efficient dibenzofuran-degrading bacterium, was isolated from landfill leachate. Here, we present the complete genome sequence of strain FA-HZ1, which contains only one circular chromosome. The complete genome sequence will be essential for revealing the molecular mechanisms of dibenzofuran degradation.

Effects of inorganic nanoparticles on viability and catabolic activities of Agrobacterium sp. PH-08 during biodegradation of dibenzofuran.

This study investigated the cytotoxicity, genotoxicity, and growth inhibition effects of four different inorganic nanoparticles (NPs) such as aluminum (nAl), iron (nFe), nickel (nNi), and zinc (nZn) on a dibenzofuran (DF) degrading bacterium Agrobacterium sp. PH-08. NP (0-1,000 mg L(-1)) -treated bacterial cells were assessed for cytotoxicity, genotoxicity, growth and biodegradation activities at biochemical and molecular levels. In an aqueous system, the bacterial cells treated with nAl, nZn and nNi at 500 mg L(-1) showed significant reduction in cell viability (30-93.6 %, p < 0.05), while nFe had no significant inhibition on bacterial cell viability. In the presence of nAl, nZn and nNi, the cells exhibited elevated levels of reactive oxygen species (ROS), DNA damage and cell death. Furthermore, NP exposure showed significant (p < 0.05) impairment in DF and catechol biodegradation activities. The reduction in DF biodegradation was ranged about 71.7-91.6 % with single NPs treatments while reached up to 96.3 % with a mixture of NPs. Molecular and biochemical investigations also clearly revealed that NP exposure drastically affected the catechol-2,3-dioxygenase activities and its gene (c23o) expression. However, no significant inhibition was observed in nFe treatment. The bacterial extracellular polymeric materials and by-products from DF degradation can be assumed as key factors in diminishing the toxic effects of NPs, especially for nFe. This study clearly demonstrates the impact of single and mixed NPs on the microbial catabolism of xenobiotic-degrading bacteria at biochemical and molecular levels. This is the first study on estimating the impact of mixed NPs on microbial biodegradation.

Degradation of dibenzofuran via multiple dioxygenation by a newly isolated Agrobacterium sp. PH-08

To demonstrate the biodegradation of dibenzofuran (DF) and its structural analogs by a newly isolated Agrobacterium sp. PH-08. To assess the biodegradation potential of newly isolated Agrobacterium sp. PH-08, various substrates were evaluated as sole carbon sources in growth and biotransformation experiments. ESI LC-MS/MS analysis revealed the presence of angular degrading by-products as well as lateral dioxygenation metabolites in the upper pathway. The metabolites in the lower pathway also were detected. In addition, the cometabolically degraded daughter compounds of DF-related compounds such as BP and dibenzothiophene (DBT) in dual substrate degradation were observed. Strain PH-08 exhibited the evidence of meta-cleavage pathway as confirmed by the activity and gene expression of catechol-2,3-dioxygenase. Newly isolated bacterial strain, Agrobacterium sp. PH-08, grew well with and degraded DF via both angular and lateral dioxygenation as demonstrated by metabolites identified through ESI LC-MS/MS and GC-MS analyses. The other heterocyclic pollutants were also cometabolically degraded. Few reports have described the complete degradation of DF by a cometabolic lateral pathway. Our study demonstrates the novel results that the newly isolated strain utilized the DF as a sole carbon source and mineralized it via multiple dioxygenation.

Cometabolic degradation of dibenzofuran by Comamonas sp. MQ

In this study, strain MQ belonging to the genera Comamonas was used to cometabolically degrade dibenzofuran (DBF) with naphthalene, phenanthrene, benzene, toluene, biphenyl and nitrobenzene, respectively, for the first time. Strain MQ could cometabolically degrade DBF in the growing system using naphthalene as a substrate and the Ki value of strain MQ on naphthalene and DBF was 90.26mgL−1 and 68.34mgL−1, respectively. The degradation rate of DBF by naphthalene-cultivated strain MQ cells (0.080mmolL−1h−1) was 1.05, 1.11, 1.13, 1.18 and 1.27-fold higher than that cultivated by phenanthrene, benzene, toluene, biphenyl and nitrobenzene, respectively. Examination of metabolites indicated that naphthalene-cultivated strain MQ cells degraded DBF to 2-hydroxy-4-(3′-oxo-3′H-benzofuran-2′-yliden)but-2-enoic acid (HOBB) and subsequently to salicylic acid via the lateral dioxygenation and meta cleavage pathway. In contrast, biphenyl-cultivated strain MQ cells degraded DBF to monohydroxydibenzofuran through the lateral dioxygenation without meta cleavage pathway. These results suggested that strain MQ could be useful in the bioremediation of environments contaminated by heterocyclic compounds mixtures with polycyclic aromatic hydrocarbons.

Cloning of dfdA genes from Terrabacter sp. strain DBF63 encoding dibenzofuran 4,4a-dioxygenase and heterologous expression in Streptomyces lividans

A dibenzofuran (DF)-degrader Terrabacter sp. strain DBF63 harbors the dbfA and dbfBC genes for DF degradation and the fln-dbfA, pht, and pca gene clusters for the utilization of fluorene (FN) as a sole carbon source. From this strain, dfdA1, the gene encoding the second DF dioxygenase was detected using degenerate polymerase chain reaction (PCR) and the dfdA1A2A3A4 genes were cloned from a cosmid library of the DBF63 genome. Nucleotide sequencing revealed that the dfdA genes showed considerably high identities with those of other actinobacteria, such as Terrabacter sp. strain YK3 and Rhodococcus sp. strain HA01. In the neighboring region of the dfdA genes, as many as 11 homologs for transposase and integrase genes and the putative extradiol dioxygenase gene disrupted by an insertion sequence (dfdB::ISTesp2) were found, suggesting that repeated gene rearrangement had occurred. Quantitative reverse transcription-PCR analysis revealed that dfdA1 was expressed primarily in the DF-fed strain, whereas dbfA1 was expressed in the FN-cultured strain, apparently indicating that the dfdA genes are induced by DF for the initial hydroxylation of DF in strain DBF63. Furthermore, two polycistronic gene cassettes consisting of either dfdA or dbfA together with the dbfBC gene were constructed and expressed heterologously in Streptomyces lividans, degrading DF to salicylate. Furthermore, the expressed DfdA dioxygenase degraded dibenzo-p-dioxin, carbazole, dibenzothiophene, anthracene, phenanthrene, and biphenyl, thereby exhibiting a broader substrate range than that of the DbfA dioxygenase.

Genome-Wide Analysis of Salicylate and Dibenzofuran Metabolism in Sphingomonas Wittichii RW1

Sphingomonas wittichii RW1 is a bacterium isolated for its ability to degrade the xenobiotic compounds dibenzodioxin and dibenzofuran (DBF). A number of genes involved in DBF degradation have been previously characterized, such as the dxn cluster, dbfB, and the electron transfer components fdx1, fdx3, and redA2. Here we use a combination of whole genome transcriptome analysis and transposon library screening to characterize RW1 catabolic and other genes implicated in the reaction to or degradation of DBF. To detect differentially expressed genes upon exposure to DBF, we applied three different growth exposure experiments, using either short DBF exposures to actively growing cells or growing them with DBF as sole carbon and energy source. Genome-wide gene expression was examined using a custom-made microarray. In addition, proportional abundance determination of transposon insertions in RW1 libraries grown on salicylate or DBF by ultra-high throughput sequencing was used to infer genes whose interruption caused a fitness loss for growth on DBF. Expression patterns showed that batch and chemostat growth conditions, and short or long exposure of cells to DBF produced very different responses. Numerous other uncharacterized catabolic gene clusters putatively involved in aromatic compound metabolism increased expression in response to DBF. In addition, only very few transposon insertions completely abolished growth on DBF. Some of those (e.g., in dxnA1) were expected, whereas others (in a gene cluster for phenylacetate degradation) were not. Both transcriptomic data and transposon screening suggest operation of multiple redundant and parallel aromatic pathways, depending on DBF exposure. In addition, increased expression of other non-catabolic genes suggests that during initial exposure, S. wittichii RW1 perceives DBF as a stressor, whereas after longer exposure, the compound is recognized as a carbon source and metabolized using several pathways in parallel.

Proteomic profiling of the dioxin-degrading bacterium Sphingomonas wittichii RW1.

Sphingomonas wittichii RW1 is a bacterium of interest due to its ability to degrade polychlorinated dioxins, which represent priority pollutants in the USA and worldwide. Although its genome has been fully sequenced, many questions exist regarding changes in protein expression of S. wittichii RW1 in response to dioxin metabolism. We used difference gel electrophoresis (DIGE) and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) to identify proteomic changes induced by growth on dibenzofuran, a surrogate for dioxin, as compared to acetate. Approximately 10% of the entire putative proteome of RW1 could be observed. Several components of the dioxin and dibenzofuran degradation pathway were shown to be upregulated, thereby highlighting the utility of using proteomic analyses for studying bioremediation agents. This is the first global protein analysis of a microorganism capable of utilizing the carbon backbone of both polychlorinated dioxins and dibenzofurans as the sole source for carbon and energy.