Biodegradation Of Phenanthrene Research Articles

Pseudomonas and Pseudarthrobacter are the key players in synergistic phenanthrene biodegradation at low temperatures

Hydrocarbon contamination, including contamination with polycyclic aromatic hydrocarbons (PAHs), is a major concern in Antarctica due to the toxicity, recalcitrance and persistence of these compounds. Under the Antarctic Treaty, nonindigenous species are not permitted for use in bioremediation at polluted sites in the Antarctic region. In this study, three bacterial consortia (C13, C15, and C23) were isolated from Antarctic soils for phenanthrene degradation. All isolated bacterial consortia demonstrated phenanthrene degradation percentages ranging from 45 to 85% for 50 mg/L phenanthrene at 15 ℃ within 5 days. Furthermore, consortium C13 exhibited efficient phenanthrene degradation potential across a wide range of environmental conditions, including different temperature (4–30 ℃) and water availability (without polyethylene glycol (PEG) 6000 or 30% PEG 6000 (w/v)) conditions. Sequencing analysis of 16S rRNA genes revealed that Pseudomonas and Pseudarthrobacter were the dominant genera in the phenanthrene-degrading consortia. Moreover, six cultivable strains were isolated from these consortia, comprising four strains of Pseudomonas, one strain of Pseudarthrobacter, and one strain of Paeniglutamicibacter. These isolated strains exhibited the ability to degrade 50 mg/L phenanthrene, with degradation percentages ranging from 4 to 22% at 15 ℃ within 15 days. Additionally, the constructed consortia containing Pseudomonas spp. and Pseudarthrobacter sp. exhibited more effective phenanthrene degradation (43–52%) than did the individual strains. These results provide evidence that Pseudomonas and Pseudarthrobacter can be potential candidates for synergistic phenanthrene degradation at low temperatures. Overall, our study offers valuable information for the bioremediation of PAH contamination in Antarctic environments.

Unraveling the mechanism of interaction: accelerated phenanthrene degradation and rhizosphere biofilm/iron plaque formation influenced by phenolic root exudates.

Phenolic root exudates (PREs) secreted by wetland plants facilitate the accumulation of iron in the rhizosphere, potentially providing the essential active iron required for the generation of enzymes that degrade polycyclic aromatic hydrocarbon, thereby enhancing their biodegradation. However, the underlying mechanisms involved are yet to be elucidated. This study focuses on phenanthrene (PHE), a typical polycyclic aromatic hydrocarbon pollutant, utilizing representative PREs from wetland plants, including p-hydroxybenzoic, p-coumaric, caffeic, and ferulic acids. Using hydroponic experiments, 16S rRNA sequencing, and multiple characterization techniques, we aimed to elucidate the interaction mechanism between the accelerated degradation of PHE and the formation of rhizosphere biofilm/iron plaque influenced by PREs. Although all four types of PREs altered the biofilm composition and promoted the formation of iron plaque on the root surface, only caffeic acid, possessing a similar structure to the intermediate metabolite of PHE (catechol), could accelerate the PHE degradation rate. Caffeic acid, notable for its catechol structure, plays a significant role in enhancing PHE degradation through two main mechanisms: (a) it directly boosts PHE co-metabolism by fostering the growth of PHE-degrading bacteria, specifically Burkholderiaceae, and by facilitating the production of the key metabolic enzyme catechol 1,2-dioxygenase (C12O) and (b) it indirectly supports PHE biodegradation by promoting iron plaque formation on root surfaces, thereby enriching free iron for efficient microbial synthesis of C12O, a crucial factor in PHE decomposition.

Major facilitator superfamily transporter PhaT modulates the efflux and degradation of polycyclic aromatic hydrocarbons in Novosphingobium pentaromativorans US6-1

Polycyclic aromatic hydrocarbons (PAHs) are a class of persistent and toxic organic pollutants that are widely distributed in nature. Although catabolic pathways are essential for bacteria to effectively utilize PAHs, transporting PAHs and their intermediate metabolites across the bacterial cell envelope is a key factor in PAH catabolism and bacterial survival. In this study, we identified a major facilitator superfamily (MFS) transporter member, PhaT, in a PAH-degrading plasmid of Novosphingobium pentaromativorans US6–1. Evolutionary analyses indicate that PhaT analogs are widely distributed in PAH-degrading bacteria, including the genera Novosphingobium and Sphingobium. Interestingly, deletion of the phaT gene upregulated the levels of PAH-degrading enzymes and PAH-degrading gene expression, thereby accelerating phenanthrene biodegradation. Further studies demonstrated that a deficiency of PhaT impaired the efflux of phenanthrene and its benzene ring-containing intermediate metabolites. Furthermore, the deficiency of PhaT also promoted the biodegradation of naphthalene but not of benzo[a]pyrene. Molecular docking and site-directed mutagenesis revealed the essential roles of hydrophobic amino acids in the PhaT central cavity for binding to benzene ring-containing substrates. Collectively, these findings indicate that PhaT plays a pivotal role in facilitating the efflux of PAHs and their benzene ring-containing intermediate metabolites.

Enhanced phenanthrene biodegradation in river sediments by harnessing calcium peroxide nanoparticles and minerals in Sphingomonas sp. DSM 7526 cultivation

ABSTRACT Coupling chemical oxidation and biodegradation to remediate polycyclic aromatic hydrocarbon (PAH)-contaminated sediment has recently gained significant attention. In this study, calcium peroxide nanoparticles (nCaO2) were utilized as an innovative oxygen-releasing compound for in-situ chemical oxidation. The study investigates the bioremediation of phenanthrene (PHE)-contaminated sediment inoculated with Sphingomonas sp. DSM 7526 bacteria and treated with either aeration or nCaO2. Using three different culture media, the biodegradation efficiencies of PHE-contaminated anoxic sediment, aerobic sediment, and sediment treated with 0.2% w/w nCaO2 ranged from 57.45% to 63.52%, 69.87% to 71.00%, and 92.80% to 94.67%, respectively. These values were significantly higher compared to those observed in non-inoculated sediments. Additionally, the type of culture medium had a prominent effect on the amount of PHE removal. The presence of minerals in the culture medium increased the percentage of PHE removal compared to distilled water by about 2–10%. On the other hand, although the application of CaO2 nanoparticles negatively impacted the abundance of sediment bacteria, resulting in a 30–42% decrease in colony-forming units after 30 days of treatment, the highest PHE removal was obtained when coupling biodegradation and chemical oxidation. These findings demonstrate the successful application of bioaugmentation and chemical oxidation processes for treating PAH-contaminated sediment.

Cultivation and characterization of functional-yet-uncultivable phenanthrene degraders by stable-isotope-probing and metagenomic-binning directed cultivation (SIP-MDC)

High-throughput identification and cultivation of functional-yet-uncultivable microorganisms is a fundamental goal in environmental microbiology. It remains as a critical challenge due to the lack of routine and effective approaches. Here, we firstly proposed an approach of stable-isotope-probing and metagenomic-binning directed cultivation (SIP-MDC) to isolate and characterize the active phenanthrene degraders from petroleum-contaminated soils. From SIP and metagenome, we assembled 13 high-quality metagenomic bins from 13C-DNA, and successfully obtained the genome of an active PHE degrader Achromobacter (genome-MB) from 13C-DNA metagenomes, which was confirmed by gyrB gene comparison and average nucleotide/amino identity (ANI/AAI), as well as the quantification of PAH dioxygenase and antibiotic resistance genes. Thereinto, we modified the traditional cultivation medium with antibiotics and specific growth factors (e.g., vitamins and metals), and separated an active phenanthrene degrader Achromobacter sp. LJB-25 via directed isolation. Strain LJB-25 could degrade phenanthrene and its identity was confirmed by ANI/AAI values between its genome and genome-MB (>99 %). Our results hinted at the feasibility of SIP-MDC to identify, isolate and cultivate functional-yet-uncultivable microorganisms (active phenanthrene degraders) from their natural habitats. Our findings developed a state-of-the-art SIP-MDC approach, expanded our knowledge on phenanthrene biodegradation mechanisms, and proposed a strategy to mine functional-yet-uncultivable microorganisms.

Metagenomic insights into phenanthrene biodegradation in electrical field-governed biofilms for groundwater bioremediation

Electrical fields (EFs)-assisted in-situ bioremediation of petroleum-contaminated groundwater, such as polycyclic aromatic hydrocarbons, has drawn increasing attention. However, the long-term stability, the EFs influence, and metabolic pathways are still poorly understood, hindering the further development of robust technology design. Herein, a series of EFs was applied to the phenanthrene-contaminated groundwater, and the corresponding system performance was investigated. The highest removal capacity of phenanthrene (phe) (7.63 g/(m3·d)) was achieved with EF_0.8 V biofilm at a hydrolytic retention time of 0.5 d. All the biofilms with four EFs exhibited a high removal efficiency of phe over 80% during a 100-d continuous-flow operation. Intermediates analysis revealed the main pathways of phe degradation: phthalate and salicylate via hydroxylation, methylation, carboxylation, and ring cleavage steps. Synergistic effects between phe-degraders (Dechloromonas), fermentative bacteria (Delftia), and electroactive microorganisms (Geobacter) were the main contributors to the complete phe mineralization. Genes encoding various proteins of methyl-accepting (mcp), response regulator (cheABDRY), and type IV pilus (pilABCMQV) were dominant, revealing the importance of cell motility and extracellular electron transfer. Metagenomics analysis unveiled phe-degrading genes, including ring reduction enzymes (bamBCDE), carboxylase of aromatics (ubiD), and methyltransferase protein (ubiE, pcm). These findings offered a molecular understanding of refractory organics’ decompositions in EFs-governed biotechnology.

A multi-omics approach to unravelling the coupling mechanism of nitrogen metabolism and phenanthrene biodegradation in soil amended with biochar

The presence of polycyclic aromatic hydrocarbons (PAHs) in soil negatively affects the environment and the degradation of these contaminants is influenced by nitrogen metabolism. However, the mechanisms underlying the interrelationships between the functional genes involved in nitrogen metabolism and phenanthrene (PHE) biodegradation, as well as the effects of biochar on these mechanisms, require further study. Therefore, this study utilised metabolomic and metagenomic analysis to investigate primary nitrogen processes, associated functional soil enzymes and functional genes, and differential soil metabolites in PHE-contaminated soil with and without biochar amendment over a 45-day incubation period. Results showed that dissimilatory nitrate reduction to ammonium (DNRA) and denitrification were the dominant nitrogen metabolism processes in PHE-contaminated soil. The addition of biochar enhanced nitrogen modules, exhibiting discernible temporal fluctuations in denitrification and DNRA proportions. Co-occurrence networks and correlation heatmap analysis revealed potential interactions among functional genes and enzymes responsible for PHE biodegradation and nitrogen metabolism. Notably, enzymes associated with denitrification and DNRA displayed significant positive correlation with enzymes involved in downstream phenanthrene degradation. Of particular interest was stronger correlation observed with the addition of biochar. However, biochar amendment inhibited the 9-phenanthrol degradation pathway, resulting in elevated levels of glutathione (GSH) in response to environmental stress. These findings provide new insights into the interactions between nitrogen metabolism and PHE biodegradation in soil and highlight the dual effects of biochar on these processes.

Enhancing Anaerobic Biodegradation of Phenanthrene in Polluted Soil by Bioaugmentation and Biostimulation: Focus on the Distribution of Phenanthrene and Microbial Community Analysis

The remediation of polycyclic aromatic hydrocarbon (PAH)-contaminated soils has received much attention in recent years, and most of the contaminated sites are in anaerobic environments, such as deep soils and flooded soils. We simulated the natural flooded soil environment, selected phenanthrene (PHE) as a model PAH contaminant, and designed batch experiments run for 63 days to comprehensively investigate the effects of the combined addition of anaerobic sludge and granular biochar on microbial community and function and the anaerobic biodegradation of PHE. Firstly, the residue, distribution, and removal of PHE in the flooded soil environment were quantified for each group. Secondly, the effects of bioaugmentation of soil indigenous microorganisms by the addition of anaerobic activated sludge and biostimulation of biochar on the removal of PHE from the soil were analyzed against each other. Lastly, the changes in the structure of the microbial community under the effect of bioaugmentation and biostimulation were illustrated by sequencing analyses. The results of this study showed that the removal efficiency of PHE reached 72.0% after the addition of anaerobic activated sludge. The incorporation of anaerobic activated sludge and biochar resulted in a 25.3% increase in PHE removal compared to a single soil, suggesting that the combination of bioaugmentation and biostimulation can have a synergistic effect on the anaerobic biodegradation of PHE in contaminated soils. The results of sequencing analysis further indicated that the introduction of an exogenous microbial community changed the dominant genera associated with PHE degradation and introduced methanogenic archaea, which enriched the metabolic pathways of the carbon cycle in the system. On this basis, the addition of biochar resulted in higher anaerobic microbial community diversity, functional dominant species were enriched, and the direct interspecies electron transfer (DIET) process between electroactive bacteria (Bacteroides, f_Geobacteraceae) and Methanosaeta was facilitated, which accelerated the degradation of PHE by anaerobic microbial communities. The results of this study provide regulatory tools and basic data support for enhanced bioremediation of PAHs in flooded soils.

Overcoming methanogenesis barrier to acid inhibition and enhancing PAHs removal by granular biochar during anaerobic digestion

Alleviating acid inhibition is the most important for optimizing anaerobic digestion (AD), but the recalcitrant organic pollutants in digestates have long been ignored. In this study, we aimed to simultaneously enhance both organics bio-methanation and recalcitrant polycyclic aromatic hydrocarbons (PAHs) removal in AD system by using granule biochar and short-term ethanol stimulation. The results showed that adding biochar effectively alleviated volatile fatty acids (VFAs) accumulation and achieved 313.6 mL CH4/d•g COD average yield. Meanwhile, the efficiency of phenanthrene (PHE) biodegradation, as the model PAHs pollutant, was significantly promoted by 37.1%. Additionally, more and compact extracellular polymeric substances improved electron transfer and mass transfer among microbes, and higher enzymes (ATPase, NADH and α-Amylase) activities met the physiological demanding of efficient microbial metabolism in biochar-added reactor. Microbiological analysis revealed that biochar-mediated direct interspecies electron transfer (DIET) selectively enriched exoelectrogens (Bacteroides, Proteiniphilum, f_Christensenellaceae, etc.) and acetoclastic Methanosaeta, which contributed to increasing biomethane production. The promoted PAHs biodegradation was mainly dependent on the cooperation between fermenting f_Paludibacteraceae and hydrogenotrophic Methanolinea, and potentially related to biochar-mediated DIET. Metabolic analysis indicated that the metabolic pathways of acetogenesis and methanogenesis were significantly strengthened by biochar, thus alleviating acid inhibition. Also, biochar stimulated functional genes expression in PAHs anaerobic biodegradation. These findings were further verified by a series of short-term metabolism tests.

Biochar inoculated with Pseudomonas putida alleviates its inhibitory effect on biodegradation pathways in phenanthrene-contaminated soil

Controversial results are reported whereby biodegradation of polycyclic aromatic hydrocarbons (PAHs) can be promoted or inhibited by biochar amendment of soil. Metabolomics was applied to analyze the metabolic profiles of amendment with biochar (BB) and biochar inoculated with functional bacteria (Pseudomonas putida) (BP) involved in phenanthrene (PHE) degradation. Additionally, metagenomic analysis was utilized to assess the impact of different treatments on PHE degradation by soil microorganisms. Results indicated that BB treatment decreased the PHE biodegradation of the soil indigenous bacterial consortium, but BP treatment alleviated this inhibitory effect. Metabolomics revealed the differential metabolite 9-phenanthrol was absent in the BB treatment, but was found in the control group (CK), and in the treatment inoculated with the Pseudomonas putida (Ps) and the BP treatment. Metagenomic analysis showed that biochar decreased the abundance of the cytochrome P450 monooxygenase (CYP116), which was detected in the Pseudomonas putida, thus alleviating the inhibitory effect of biochar on PHE degradation. Moreover, a noticeable delayed increase of functional gene abundance and enzymes abundance in the BB treatment was observed in the PHE degradation pathway. Our findings elucidate the mechanism of inhibition with biochar amendment and the alleviating effect of biochar inoculated with degrading bacteria.

Evaluation of Azolla filiculoides potential in pyrene and phenanthrene accumulation and phytoremediation in contaminated waters

Polycyclic aromatic hydrocarbons (PAHs) are a serious threat to the health of the environment. This study investigated the potential of Azolla filiculoides for the uptake, accumulation, and biodegradation of phenanthrene and pyrene. A- filiculoides plants were treated with 10 and 30 mg L−1 concentrations of phenanthrene and pyrene for the experimental duration of ten days. Phenanthrene and pyrene concentrations were measured using the high-performance liquid chromatography (HPLC) technique. Identification of the intermediate by-products resulting from the biological degradation of PAHs was performed by gas chromatography-mass spectrometry (GC/MS). The quantities of phenanthrene and pyrene in the ten-day treatments with 10 and 30 mg L−1 were 0.007 and 0.011 mg g−1 FW, and 0.048 and 0.079 mg g−1 FW, respectively. The growth parameters in the plants such as fresh weight, dry weight and RFN as well as the content of photosynthetic pigment of the plant decreased significantly compared to the control sample (p < 0.05). Ten compounds were identified from the plant tissue during the decomposition of pyrene and phenanthrene, and none of the PAHs were identified in the aquatic environment. Therefore, the use of A-filiculoides for phytoremediation of water resources contaminated with PAHs is an effective and promising method.

Enhanced biodegradation of phenanthrene and anthracene using a microalgal-bacterial consortium.

Polycyclic aromatic hydrocarbons (PAHs) are chemicals that are released into the environment during activities of the petroleum industry. The bioaccumulation, carcinogenic and mutagenic potential of PAHs necessitates the bioremediation of these contaminants. However, bioremediation of PAHs has a number of limitations including the inability of a single microbe to degrade all of the PAH fraction's environmental constituents. Therefore, a different paradigm, employing microalgal-bacterial consortium (MBC), may be used to effectively remove PAHs contaminants. In this type of interaction, the microalgae and bacteria species in the consortium work together in a way that enhances the overall performance of the MBC. Bacterial species in the consortium provide essential nutrients or growth factors by degrading toxic substances and provide these to microalgae, while the microalgae species provide organic carbon for the bacterial species to grow. For the first time, the ability of Gonium pectorale (G. pectorale) microalgae to break down phenanthrene (PHE) and anthracene (ANT) was investigated. Phenanthrene was shown to be more effectively degraded by G. pectorale (98%) as compared to Bacillus licheniformis (B. licheniformis) 19%. Similarly, G. pectorale has effectively degrade anthracene (98%) as compared with B. licheniformis (45%). The consortia of G. pectorale and B. licheniformis has shown a slight increase in the degradation of PHE (96%) and ANT (99%). Our findings show that B. licheniformis did not inhibit the growth of G. pectorale and in the consortia has effectively eliminated the PAHs from the media. Therefore G. pectorale has a tremendous potential to remove PAHs from the polluted environment. Future research will be conducted to assess Gonium's capacity to eliminate PAHs that exhibit high molar masses than that of PHE and ANT.

Integrating metal-organic framework ZIF-8 with green modifier empowered bacteria with improved bioremediation

Suspended microorganisms often experience diminished efficacy in the bioremediation of polycyclic aromatic hydrocarbons (PAHs). In this study, the potential of zeolite imidazolate framework-8 (ZIF-8) and the eco-friendly modifier citric acid (CA) was harnessed to generate a biomimetic mineralized protective shell on the surface of Bacillus subtilis ZL09–26, resulting in an enhanced capability for PAH degradation. This investigation encompassed the integrated responses of B. subtilis ZL09–26 to ZIF-8 and ZIF-8-CA at both cellular and proteomic levels. The amalgamation of ZIF-8 and CA not only stimulated the growth and bolstered the cell viability of B. subtilis ZL09–26, but also counteracted the toxic effects of phenanthrene (PHE) stress. Remarkably, the bioremediation prowess of B. subtilis ZL09–26@ZIF-8-CA surpassed that of ZL09–26@ZIF-8 and ZL09–26, achieving a PHE removal rate of 94.14 % within 6 days. After undergoing five cycles, ZL09–26@ZIF-8-CA demonstrated an enduring PHE removal rate exceeding 83.31 %. A complex interplay of various metabolic pathways orchestrated cellular responses, enhancing PHE transport and degradation. These pathways encompassed direct PHE biodegradation, central carbon metabolism, oxidative phosphorylation, purine metabolism, and aminoacyl-tRNA biosynthesis. This study not only extends the potential applications of biomineralized organisms but also offers alternative strategies for effective contaminant management.

Coupled Fe(III) reduction and phenanthrene degradation by marine-derived Kocuria oceani FXJ8.057 under aerobic condition

Diverse aerobic actinobacteria possess the capacity to degrade polycyclic aromatic hydrocarbons (PAHs) and have recently been shown to reduce Fe(III). However, the coupling of the two processes under oxic conditions remains unclear. Here, the co-metabolism of phenanthrene (PHE) and Fe(III) by marine-derived Kocuria oceani FXJ8.057 was realized under aerobic condition. In the presence of both PHE and Fe(III), the rates of PHE degradation (83.91 %) and Fe(III) reduction (50.00 %) were synchronously enhanced, compared to those with PHE (67.34 %) or Fe(III) (38.00 %) alone. Transcriptome analysis detected upregulation of PHE biodegradation and riboflavin biosynthesis in the strain cultured with both PHE and Fe(III) compared to that with PHE alone. Metabolite analysis indicated that, with the addition of Fe(III), the strain could efficiently degrade PHE via three pathways. Moreover, the strain secreted riboflavin, which acted as a shuttle to promote electron transfer from PHE to Fe(III). It also secreted organic acids that could delay Fe(II) reoxidation. Finally, H2O2 secreted by the strain caused extracellular Fenton reaction to generate •OH, which also played a minor role in the PHE degradation. These findings provide the first example of an aerobic bacterium that couples PAH degradation to Fe(III) reduction and extend our understanding of Fe(III)-reducing microorganisms.

Biodegradation of phenanthrene by piezotolerant Bacillus subtilis EB1 and genomic insights for bioremediation

A marine strain B. subtilis EB1, isolated from Equator water, showed excellent degradation towards a wide range of hydrocarbons. Degradation studies revealed dense growth with 93 % and 83 % removal of phenanthrene within 72 h at 0.1 and 20 MPa, respectively. The identification of phenanthrene degradation metabolites by GC–MS combined with its whole genome analysis provided the pathway involved in the degradation process. Whole genome sequencing indicated a genome size of 3,983,989 bp with 4331 annotated genes. The genome provided the genetic compartments, which includes monooxygenase, dioxygenase, dehydrogenase, biosurfactant synthesis catabolic genes for the biodegradation of aromatic compounds. Detailed COG and KEGG pathway analysis confirmed the genes involved in the oxygenation reaction of hydrocarbons, piezotolerance, siderophores, chemotaxis and transporter systems which were specific to adaptation for survival in extreme marine habitat. The results of this study will be a key to design an optimal bioremediation strategy for oil contaminated extreme marine environment.

Anaerobic biodegradation of phenanthrene and pyrene by sulfate-reducing cultures enriched from contaminated freshwater lake sediments

Our current understanding of the susceptibility of hazardous polycyclic aromatic hydrocarbons (PAHs) to anaerobic microbial degradation is very limited. In the present study, we obtained phenanthrene- and pyrene-degrading strictly anaerobic sulfate-reducing enrichments using contaminated freshwater lake sediments as the source material. The highly enriched phenanthrene-degrading culture, MMKS23, was dominated (98%) by a sulfate-reducing bacterium belonging to the genus Desulfovibrio. While Desulfovibrio sp. was also predominant (79%) in the pyrene-degrading enrichment culture, MMKS44, an anoxygenic purple non-sulfur bacterium, Rhodopseudomonas sp., constituted a significant fraction (18%) of the total microbial community. Phenanthrene or pyrene biodegradation by the enrichment cultures was coupled with sulfate reduction, as evident from near stoichiometric consumption of sulfate and accumulation of sulfide. Also, there was almost complete inhibition of substrate degradation in the presence of an inhibitor of sulfate reduction, i.e., 20 mM MoO42−, in the culture medium. After 180 days of incubation, about 79.40 μM phenanthrene was degraded in the MMKS23 culture, resulting in the consumption of 806.80 μM sulfate and accumulation of 625.80 μM sulfide. Anaerobic pyrene biodegradation by the MMKS44 culture was relatively slow. About 22.30 μM of the substrate was degraded after 180 days resulting in the depletion of 239 μM sulfate and accumulation of 196.90 μM sulfide. Biodegradation of phenanthrene by the enrichment yielded a metabolite, phenanthrene-2-carboxylic acid, suggesting that carboxylation could be a widespread initial step of phenanthrene activation under sulfate-reducing conditions. Overall, this novel study demonstrates the ability of sulfate-reducing bacteria (SRB), dwelling in contaminated freshwater sediments to anaerobically biodegrade three-ringed phenanthrene and highly recalcitrant four-ringed pyrene. Our findings suggest that SRB could play a crucial role in the natural attenuation of PAHs in anoxic freshwater sediments.

Cometabolic degradation of pyrene with phenanthrene as substrate: assisted by halophilic Pseudomonas stutzeri DJP1.

At present, cometabolic degradation is an extensive method for the biological removal of high molecular weight polycyclic aromatic hydrocarbons (HMW-PAHs) in the marine environment. However, due to the refractory to degradation and high toxicity, there are few studies on pyrene (PYR) cometabolic degradation with phenanthrene (PHE) as substrate. In this study, a Pseudomonas stutzeri DJP1 strain isolated from sediments was used in the cometabolic system of PHE and PYR. The biomass and the activity of key enzymes such as dehydrogenase and catechol 12 dioxygenase of strain were improved, but the enhancement of biotoxicity resulted in the inhibition of cometabolism simultaneously. Seven metabolites were identified respectively in PYR, PHE degradation cultures. It was speculated that the cometabolism of PHE and PYR had a common phthalic acid pathway, and the degradation pathway of PHE was included in the downstream pathway of PYR. The functional genes such as PhdF, NidD and CatA involved in DJP1 degradation were revealed by Genome analysis. This study provides a reference for the biodegradation of PYR and PHE in real marine environment.

Aerobic biodegradation of phenanthrene by a newly isolated Klebsiella sp. DS-1 from wastewater

Representative polycyclic aromatic hydrocarbons (PAHs) compounds such as phenanthrene (PHE) derived from multiple anthropogenic activities are contributing to a growing number of concerns about its threats to the global water environment and human health. Microbial degradation is considered to be an increasingly promising approach for PHE removal, while effective PHE biodegraders from in situ environments are yet to be identified and their underlying degradation mechanisms still remain to be understood. Herein, a Klebsiella sp. strain DS-1 was isolated from petrochemical wastewater using PHE as the sole carbon source. The degradation efficiency of PHE was up to 88.4 % at 30 °C and pH 7 within a maximum of 9 days. DS-1 grew actively in wide ranges of environmental factors, such as pH values (5–9), temperature (20 °C–37 °C), and initial concentrations of PHE (20–200 mg/L), with the maintenance of PHE degradability. Genome analysis revealed 14 genes that were involved in PHE biodegradation by DS-1. Detection of metabolic intermediates revealed that strain DS-1 degraded PHE via the phthalic acid pathway. These findings have expanded the knowledge of Klebsiella sp. as an active PHE degrader in wastewater and indicate its potential in the bioremediation of PHE-contaminated aquatic environments.

Whole-genome sequence analysis reveals phenanthrene and pyrene degradation pathways in newly isolated bacteria Klebsiella michiganensis EF4 and Klebsiella oxytoca ETN19

Polycyclic aromatic hydrocarbons (PAHs) are diverse pollutants of significant environmental concerns, requiring effective biodegradation. This study used different bioinformatics tools to conduct whole-genome sequencing of two novel bacterial strains, Klebsiella michiganensis EF4 and K. oxytoca ETN19, to improve our understanding of their many genomic functions and degradation pathways of phenanthrene and pyrene. After 28 days of cultivation, strain EF4 degraded approximately 80% and 60% of phenanthrene and pyrene, respectively. However, their combinations (EF4 +ETN19) showed tremendous phenanthrene degradation efficiency, supposed to be at the first-level kinetic model with a t1/2 value of approximately 6 days. In addition, the two bacterial genomes contained carbohydrate-active enzymes and secondary metabolites biosynthetic gene clusters associated with PAHs degradation. The two genomes contained the bZIP superfamily of transcription factors, primarily the cAMP-response element-binding protein (CREB), which could regulate the expression of several PAHs degradation genes and enzymes. Interestingly, the two genomes were found to uniquely degrade phenanthrene through a putative pathway that catabolizes 2-carboxybenzalpyruvate into the TCA cycle. An operon containing multicomponent proteins, including a novel gene (JYK05_14550) that could initiate the beginning step of phenanthrene and pyrene degradation, was found in the EF4 genome. However, the degradation pathway of ETN19 showed that the yhfP gene encoding putative quinone oxidoreductase was associated with phenanthrene and pyrene catabolic processes. Furthermore, the significant expression of catechol 1,2-dioxygenase and quinone oxidoreductase genes in EF4 +ETN19 and ETN19 following the quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis confirmed the ability of the bacteria combination to degrade pyrene and phenanthrene effectively. These findings present new insight into the possible co-metabolism of the two bacterial species in the rapid biodegradation of phenanthrene and pyrene in soil environments.

Genome analysis for the identification of genes involved in phenanthrene biodegradation pathway in Stenotrophomonas indicatrix CPHE1. Phenanthrene mineralization in soils assisted by integrated approaches.

Phenanthrene (PHE) is a highly toxic compound, widely present in soils. For this reason, it is essential to remove PHE from the environment. Stenotrophomonas indicatrix CPHE1 was isolated from an industrial soil contaminated by polycyclic aromatic hydrocarbons (PAHs) and was sequenced to identify the PHE degrading genes. Dioxygenase, monooxygenase, and dehydrogenase gene products annotated in S. indicatrix CPHE1 genome were clustered into different trees with reference proteins. Moreover, S. indicatrix CPHE1 whole-genome sequences were compared to genes of PAHs-degrading bacteria retrieved from databases and literature. On these basis, reverse transcriptase-polymerase chain reaction (RT-PCR) analysis pointed out that cysteine dioxygenase (cysDO), biphenyl-2,3-diol 1,2-dioxygenase (bphC), and aldolase hydratase (phdG) were expressed only in the presence of PHE. Therefore, different techniques have been designed to improve the PHE mineralization process in five PHE artificially contaminated soils (50mgkg-1), including biostimulation, adding a nutrient solution (NS), bioaugmentation, inoculating S. indicatrix CPHE1 which was selected for its PHE-degrading genes, and the use of 2-hydroxypropyl-β-cyclodextrin (HPBCD) as a bioavailability enhancer. High percentages of PHE mineralization were achieved for the studied soils. Depending on the soil, different treatments resulted to be successful; in the case of a clay loam soil, the best strategy was the inoculation of S. indicatrix CPHE1 and NS (59.9% mineralized after 120days). In sandy soils (CR and R soils) the highest percentage of mineralization was achieved in presence of HPBCD and NS (87.3% and 61.3%, respectively). However, the combination of CPHE1 strain, HPBCD, and NS showed to be the most efficient strategy for sandy and sandy loam soils (LL and ALC soils showed 35% and 74.6%, respectively). The results indicated a high degree of correlation between gene expression and the rates of mineralization.