Mechanism of phenanthrene degradation by the halophilic Pelagerythrobacter sp. N7.

PAHs has shown worldwide accumulation and causes a significant environmental problem especially in saline and hypersaline environments. Moderately halophilic bacteria could be useful for the bioremediation of PAH pollution in hypersaline environments. Pelagerythrobacter sp. N7 was isolated from the PAH-degrading consortium 5H, which was enriched from mixed saline soil samples collected in Shanxi Province, China. 16S rRNA in the genomic DNA revealed that strain N7 belonged to Pelagerythrobacter. Strain N7 exhibited a high tolerance to a wide range of salinities (1-10%) and was highly efficient under neutral to weak alkaline conditions (pH 6-9). The whole genome of strain N7 was sequenced and analyzed, revealing an abundance of catabolic genes. Using the whole genome information, we conducted preliminary research on key enzymes and gene clusters involved in the upstream and downstream PAH degradation pathways of strain N7, thereby inferring its degradation pathway for phenanthrene and naphthalene. This study adds to our understanding of PAH degradation in hypersaline environments and, for the first time, identifies a Pelagerythrobacter with PAH-degrading capability. Strain N7, with its high efficiency in phenanthrene degradation, represents a promising resource for the bioremediation of PAHs in hypersaline environments.

The discrepant metabolic pathways of PAHs by facultative anaerobic bacteria under aerobic and nitrate-reducing conditions.

Studies regarding the facultative anaerobic biodegradation of polycyclic aromatic hydrocarbons (PAHs) were still in the initial stage. In this study, a facultative anaerobe which was identified as Bacillus Firmus and named as PheN7 was firstly isolated from the mixed petroleum-polluted soil samples using phenanthrene and nitrate as the solo carbon resource and electron acceptor under anaerobic condition. The degradation rates of PheN7 towards phenanthrene were detected as 33.17 µM/d, 13.81 µM/d and 7.11 µM/d at the initial phenanthrene concentration of 250.17 µM with oxygen, nitrate and sulfate as the electron acceptor, respectively. The metabolic pathways toward phenanthrene by PheN7 were deduced combining the metagenome analysis of PheN7 and intermediate metabolites of phenanthrene under aerobic and nitrate-reducing conditions. Dioxygenation and carboxylation were inferred as the initial activation reactions of phenanthrene degradation in these two pathways. This study highlighted the significance of facultative anaerobic bacteria in natural PAHs biodegradation, revealing the discrepant metabolic fates of PAHs by one solo bacteria under aerobic and anaerobic environments.

The negative effects of the excessive nitrite accumulation raised by anaerobic bioaugmentation on bioremediation of PAH-contaminated soil.

Nitrite accumulation in anaerobic bioaugmentation and its side effects on remediation efficiency of polycyclic aromatic hydrocarbon (PAH)-contaminated soil were investigated in this study. Four gradient doses of PAH-degrading inoculum (10^4, 10^5, 10^6 and 10^7 cells/g soil) were separately supplied to the actual PAH-contaminated soil combining with nitrate as the biostimulant. Although bioaugmented with higher dose of inoculum could effectively improve the biodegradation efficiencies in the initial stage than sole nitrate addition but also accelerated the accumulation of nitrite in soil. The inhibition effects of nitrite swiftly occurred following the rapid accumulation of nitrite in each experiment group, restraining the PAH-degrading functionality by inhibiting the growth of total biomass and denitrifying functions in soil. This study revealed the side effects of nitrite accumulation raised by bioaugmentation on soil microorganisms, contributing to further improving the biodegrading efficiencies in the actual site restoration.

Anaerobic biodegradation of pyrene and benzo[a]pyrene by a new sulfate-reducing Desulforamulus aquiferis strain DSA.

The study of anaerobic high molecular weight polycyclic aromatic hydrocarbons (HMW-PAHs) biodegradation under sulfate-reducing conditions by microorganisms, including microbial species responsible for biodegradation and relative metabolic processes, remains in its infancy. Here, we found that a new sulfate-reducer, designated as Desulforamulus aquiferis strain DSA, could biodegrade pyrene and benzo[a]pyrene (two kinds of HMW-PAHs) coupled with the reduction of sulfate to sulfide. Interestingly, strain DSA could simultaneously biodegrade pyrene and benzo[a]pyrene when they co-existed in culture. Additionally, the metabolic processes for anaerobic pyrene and benzo[a]pyrene biodegradation by strain DSA were newly proposed in this study based on the detection of intermediates, quantum chemical calculations and analyses of the genome and RTqPCR. The initial activation step for anaerobic pyrene and benzo[a]pyrene biodegradation by strain DSA was identified as the formation of pyrene-2-carboxylic acid and benzo[a]pyrene-11-carboxylic acid by carboxylation Thereafter, CoA ligase, ring reduction through hydrogenation, and ring cracking occurred, and short-chain fatty acids and carbon dioxide were identified as the final products. Additionally, DSA could also utilize benzene, naphthalene, anthracene, phenanthrene, and benz[a]anthracene as carbon sources. Our study can provide new guidance for the anaerobic HMW-PAHs biodegradation under sulfate-reducing conditions.

Novel synergistic metabolic processes for phenanthrene biodegradation by a nitrate-reducing phenanthrene-degrading culture and an anammox culture.

The synergistic metabolism by anammox cultures and nitrate-reducers for anaerobic PAH biodegradation is largely unknown, including whether anammox culture and which kind of anammox bacterium can perform nitrogen metabolism in the anaerobic PAH biodegradation processes, the inhibitory effect of PAH on anammox activity and nitrite on PAH-degrading nitrate-reducers activity, and the synergistic metabolic processes. Herein, an anammox culture that can eliminate nitrite accumulation and decrease inorganic carbon emission during anaerobic phenanthrene (a model of PAH in this study) biodegradation, the synergistic mechanism for phenanthrene biodegradation by a nitrate-reducer and such anammox culture, and the inhibition effect of phenanthrene on such anammox culture and nitrite on a phenanthrene-degrading nitrate-reducer were newly discussed. The results showed that nitrite largely accumulated during anaerobic phenanthrene biodegradation (nitrate accumulation is a common phenomenon for the biodegradation of refractory matter, including PAHs, by nitrate-reducers) by a nitrate-reducer, PheN2, which mineralizes phenanthrene to inorganic carbon, and nitrite was verified as an inhibiting factor for further biodegradation. Anaerobic phenanthrene biodegradation rates and nitrite concentrations (0-7 mM) appeared to have a negative linear correlation. The anammox culture that mainly contained Candidatus Kuenenia was newly found to efficiently reduce nitrite accumulation and inorganic carbon emissions and significantly promote biodegradation efficiency by ∼1.94-fold. Our results showed that phenanthrene absorbed in and on anammox cells had a more direct relationship with the inhibitory effect on anammox activity than phenanthrene in the environment, and 15.2 mg/gVSS phenanthrene absorbed in and on the cells (4 mM concentration in the culture) showed nearly complete inhibition of anammox culture in this study. In addition, few (less than 2% abundance) anammox bacteria were found to be enough for the removal of nitrite produced from anaerobic phenanthrene biodegradation. In an ideal world, co-pollutants of ammonia, nitrate, phenanthrene, and nitrite could be converted to nitrogen gas and biomass by the synergistic metabolism of anammox cultures and nitrate reducers. Our study reveals a new synergistic process that may exist in our environments for PAH elimination by an anammox culture and a nitrate-reducer, which provides a new strategy for the bioremediation of PAH-polluted anoxic zones.

Distribution of the new functional marker gene (pahE) of aerobic polycyclic aromatic hydrocarbon (PAHs) degrading bacteria in different ecosystems.

Understanding the degradation potentials in PAHs-contaminated sites is significant for formulating effective bioremediation strategies. pahE encoding PAHs hydratase-aldolase has been proven as a better new functional marker gene of aerobic PAHs-degrading bacteria to assess the biodegradation potential of indigenous PAHs-degrading bacterial population. However, the distribution of pahE and its relationship with environmental factors remain unknown. The present study observed spatial variations in the diversity and abundance of pahE across oilfield soils, mangrove sediments, and urban roadside soils. nahE from Pseudomonas, bphE from Hyphomonas oceanitis, nagE from Comamonas testosterone, and novel pahE genes were widely present in these PAHs-polluted ecosystems. The abundance of pahE in PAHs-contaminated sites was in the range of 105-106 copies·g-1 (dry weight). Redundancy analysis and Pearson's correlation analysis implied that the distribution of pahE in the PAHs-contaminated environment was mainly shaped by environmental factors such as PAHs pollution level, nutrient level, salinity, and water content. This work was the first to explore the distribution of the new functional marker gene (pahE) and its links with environmental parameters, which provided new insights into the ecophysiology and distribution of indigenous aerobic PAHs-degrading bacteria in contaminated sites.

Degradation of phenanthrene by consortium 5H under hypersaline conditions.

PAHs have been widely detected to accumulate in saline and hypersaline environments. Moderately halophilic microbes are considered the most suitable player for the elimination of PAHs in such environments. In this study, consortium 5H was enriched under 5% salinity and completely degraded phenanthrene in 5 days. By high-throughput sequencing, consortium 5H was identified as being mainly composed of Methylophaga, Marinobacter and Thalassospira. Combined with the investigation of intermediates and enzymatic activities, the degradation pathway of consortium 5H on phenanthrene was proposed. Consortium 5H was identified as having the ability to tolerate a wide range of salinities (1%-10%) and initial PAH concentrations (50 mg/L to 400 mg/L). It was also able to function under neutral to weak alkaline conditions (pH from 6 to 9) and the phytotoxicity of the produced intermediates showed no significant difference with distilled water. Furthermore, the metagenome of consortium 5H was measured and analyzed, which showed a great abundance of catabolic genes contained in consortium 5H. This study expanded the knowledge of PAH-degradation under hypersaline environments and consortium 5H was proposed to have good potential for the elimination of PAH pollution in saline/hypersaline environments.

Bioaugmentation of polycyclic aromatic hydrocarbon (PAH)-contaminated soil with the nitrate-reducing bacterium PheN7 under anaerobic condition.

The remediation of polycyclic aromatic hydrocarbon (PAH)-contaminated soil under anaerobic condition is still a huge challenge. In this study, an anaerobic Bacillus firmus strain named PheN7 was firstly isolated from mixture of contaminated soil and sludge samples with phenanthrene as the sole carbon resource under nitrate reducing environment. The anaerobic strain was then inoculated combining with nitrate into the phenanthrene-spiked PAH-contaminated soil to investigate the remediation efficiency by anaerobic bioaugmentation (BA). Results showed that the synergy between PheN7 and indigenous degrading bacteria promoted the remediation efficiency of soil. The average removal efficiencies of phenanthrene in 56 days were 1.73 mg/kg soil·d in BA group, much higher than biostimulation group (sole nitrate addition) and natural degradation which achieved 1.48 mg/kg soil·d and 1.24 mg/kg soil·d of degradation rate, respectively. The outstanding adaptability of PheN7 made it become the dominant species in soil in the terminal period, but the invasion of PheN7 also resulted in the decline of diversity of the indigenous microbial community. Phylogenetic Investigation of Communities by Reconstruction of Unobserved States 2 (PICRUSt 2) results showed that a series of functional genes encoding anaerobic phenanthrene degradation and nitrate reductase enzymes in soil were remarkably strengthened with the addition of PheN7. This study confirmed the contribution of PheN7 as the anaerobic inoculum in PAH-contaminated soil remediation, further evaluating the practical applicability of anaerobic bioaugmentation technology in on-site remediation of real PAH-contaminated sites.

Anaerobic phenanthrene biodegradation by a new salt-tolerant/halophilic and nitrate-reducing Virgibacillus halodenitrificans strain PheN4 and metabolic processes exploration.

The biodegradation of polycyclic aromatic hydrocarbons (PAHs) under hypersaline environments has received increasing attention, whereas the study of anaerobic PAH biodegradation under hypersaline environments is still lacking. Here, we found a pure culture designated PheN4, which was affiliated with Virgibacillus halodenitrificans and could degrade phenanthrene with nitrate as the terminal electron acceptor and a wide range of salinities (from 0.3% to 20%) under anaerobic environments. The optimal salinity for biodegradation of phenanthrene by PheN4 was 5%, which could degrade 93.5% of 0.62 ± 0.04 mM phenanthrene within 10 days with the initial inoculum of 0.01 gVSS/L. Meanwhile, an increased microbial amount could efficiently promote the phenanthrene biodegradation rate. The metabolic processes of anaerobic phenanthrene biodegradation under hypersaline conditions by PheN4 were proposed based on intermediates and genome analyses. Phenanthrene was initially activated via methylation to form 2-methylphenanthrene. Next, fumarate addition and ß-oxidation or direct oxidation of the methyl group, ring reduction and ring cleavage were identified as the midstream and downstream steps. In addition, PheN4 could utilize benzene, naphthalene, and anthracene as carbon sources, but Benz[a]anthracene, pyrene, and Benzo[a]pyrene could not be biodegraded by PheN4. This study could provide some guidance for the bioremediation of PAH pollutants in anaerobic and hypersaline zones.

Shifts of the new functional marker gene (pahE) of polycyclic aromatic hydrocarbons (PAHs) degrading bacterial population and its relationship with PAHs biodegradation.

Identification of polycyclic aromatic hydrocarbons (PAHs) degrading bacterial populations and understanding their responses to PAHs are crucial for the designing of appropriate bioremediation strategies. In this study, the responses of PAHs-degrading bacterial populations to different PAHs were studied in terms of the compositions and abundance variations of their new functional marker gene (pahE) by gene-targeted metagenomic and qPCR analysis. Overall, PAHs species significantly affected the composition and abundance of pahE gene within the PAHs-degrading bacteria in each treatment and different pahE of PAHs-degrading bacteria involved in the different stages of PAHs degradation. Noted that new pahE genotypes were also discovered in all PAHs treatment groups, indicating that some potential new PAHs-degrading bacterial genera were also involved in PAHs degradation. Besides, all three PAH removal rates were significantly positively related with pahE gene abundances (R2 = 0.908 ~ 0.922, p < 0.01), demonstrating that pahE could be a good indicator of PAHs degradation activity or potential. This is the first study focusing on the dynamic changes of the pahE gene within PAHs-degrading bacterial community during the degradation of PAHs in mangrove sediment, providing novel insights into the use of pahE gene as the functional marker to indicate PAH degradation.

Insights into effects of algae on decay and distribution of bacterial pathogens in recreational water: Implications for microbial risk management.

The decay and distribution of bacterial pathogens in water is an important information for the health risk assessment to guide water safety management, and suspended algae might affect bacterial pathogens in water. This study established microcosms to investigate the effects of algae-related factors on the representative indicators and opportunistic pathogen species in water. We found that suspended algae increased the persistence of targeted species by 1-2 orders of magnitude of concentrations compared to microcosms without algae; and the effect of algae on microbial survival was affected by water nutrient levels (i.e., carbon, nitrogen and phosphorus), as the increased microbial persistence were correlated to the increased algae concentrations with more nutrient supplies. Moreover, decay and distribution profiles of representative species were determined. The three opportunistic pathogen species (Pseudomonas aeruginosa, Aeromonas hydrophila and Staphylococcus aureus) showed lower decay rates (0.82-0.98/day, 0.76-0.98/day, 0.63-0.87/day) largely affected by algae-related factors, while the enteric species (Escherichia coli and Enterococcus faecalis) had higher decay rates (0.94-1.31/day, 0.89-1.21/day) with little association with algae, indicating the propensity for attachment to algae is an important parameter in microbial fate. Together results suggest suspended algae played an evident role in the decay and distribution of bacterial pathogens, providing important implications regarding microbial safety in recreational water.

Functional microbial community structures and chemical properties indicated mechanisms and potential risks of urban river eco-remediation.

To investigate the mechanisms and potential risks of river eco-remediation, river water, sediment, and biofilms in remediation facilities were sampled from a 2-year full scale eco-remediation site in an urban river in southeastern China. The samples from both remediated and adjacent control areas were analyzed for chemical properties and functional microbial community structures. The eco-remediation significantly changed the community structures in the river and introduced much more diverse functional microorganisms in facility biofilms. Corresponding to effective reduction of organics and ammonium in river water, some labile-organics-degrading and ammonia-oxidizing gene families showed higher abundances in river water of remediated area than control area, and were obviously more abundant in facility biofilms than in river water and sediment. The eco-remediation facilities showed obvious absorption of N, P, and heavy metals (Mn, CrVI, Fe, Al, As, Co), contributing to nutrients and metals removal from river water. The eco-remediation also increased transparency and sedimentation of some heavy metals (Cu, Pb, Zn), which probably associated with colloids breakdown. Various metal-resistance microorganisms showed different abundances between facility biofilms and sediment, in accordance with relative metals. Most detected pathogens were not significantly affected by eco-remediation. However, our measurements in sediment and facilities showed heavy metals accumulation and development of some pathogens and several antibiotic-resistance pathogens, alerting us to investigate and control these potential risks to ecosystem and human health.

Exploration of the biotransformation of phenanthrene degradation coupled with methanogensis by metabolites and enzyme analyses.

The ubiquitous environmental contaminants, polycyclic aromatic hydrocarbons (PAHs), can be aerobically biodegraded. Strategies for biodegradation of PAHs are needed for the persisted character of it in anoxic environments. In current study, we obtained a highly enriched anaerobic, PAHs-degrading co-culture DYM1, from petroleum-polluted soil. DYM1 significantly degrades a range of PAHs in 4 days without supplementary terminal electron acceptors. Co-culture DYM1 is consists of two microorganisms (a degrading bacterium Paracoccus sp. strain PheM1 and an aceticlastic methanogen Methanosaeta concilii.) that utilize different carbon sources in a syntrophic metabolic process of phenanthrene. About 93% of phenanthrene (104.5 µM) has been removed under methanogenic conditions after incubation with co-culture DYM1 for 4 d, and produced 33.68 µmol CH4. Carboxylation, which is catalyzed by UbiD-like carboxylase, was proposed as the initial steps of methanogenic phenanthrene-degrading pathway based upon the detection of 2-phenanthroic acid and 4-phenanthrene acid. Reduction and hydration of the benzene rings were followed by the initial reaction. Hydrated phenanthroic acid metabolites were newly detected and characterized under anaerobic conditions. Anaerobic degradation of phenanthrene without terminal electron acceptor addition not only sheds light on a poorly understood and environmentally relevant biological process, but also supply a novel approach to recover the energy of toxic pollutant in forms of methane.

Effects of Gleditsia sinensis pod powder, coconut shell biochar and rice husk biochar as additives on bacterial communities and compost quality during vermicomposting of pig manure and wheat straw.

This study investigated the effectiveness of Gleditsia sinensis pod powder (GSPP), coconut shell biochar (CSB), rice husk biochar (RHB) and their mixtures on vermicomposting of pig manure and wheat straw using Eisenia fetida. The results indicated that the addition of GSPP or/and CSB and RHB could greatly enhance the relative abundance of Bacteroidetes, Actinobacteria, and Firmicutes, as well as the activities of celluloses, protease, and alkaline phosphatase. However, the earthworm biomass was increased in the GSPP and/or CSB addition treatments but decreased in RHB addition treatments compared with the control. Compared with the control, addition of 4%GSPP+8%CSB significantly (P < 0.05) accelerated the degradation of organic matter and increased the concentration of nutrients (total N, P, K), NO3--N in final vermicompost. Germination and growth of tomato seedings were also higher (P < 0.05) in vermicompost produced with the addition of 4%GSPP+8%CSB than in control. Consequently, 4%GSPP+8%CSB addition was suggested as an efficient method to improve the vermicomposting of pig manure and wheat straw.

Anaerobic phenanthrene biodegradation by a newly isolated sulfate-reducer, strain PheS1, and exploration of the biotransformation pathway.

Phenanthrene is a widespread and harmful polycyclic aromatic hydrocarbon that is difficult to anaerobically biodegrade. Current challenges in anaerobic phenanthrene bioremediation are a lack of degrading cultures and limited knowledge of biotransformation pathways. Under sulfate-reducing conditions, pure-cultures and biotransformation processes for anaerobic phenanthrene biodegradation are poorly understood. In this study, strain PheS1, which is phylogenetically closely related to Desulfotomaculum, was found to be a sulfate-reducing phenanthrene-degrading bacterium. Anaerobic phenanthrene biodegradation using PheS1 was proposed based on metabolite and genome analyses, and the initial step was identified as carboxylation based on the detection of 2-phenanthroic acid, [13C]-2-phenanthroic acid, and [D9]-2- phenanthroic acid when phenanthrene+HCO3-, phenanthrene+H13CO3-, and [D10]-phenanthrene+HCO3- were used as the substrate, respectively. PheS1 genome ubiD gene encoding of carboxylase putatively involved in the biodegradation was performed. Next, benzene ring reduction and cleavage that produced benzene compounds and cyclohexane derivative were reported to occur in the downstream biotransformation processes. Additionally, benzene, naphthalene, benz[a]anthracene, and anthracene can be utilised by PheS1, whereas pyrene and benz[a]pyrene cannot. We discovered a new phenanthrene-degrading sulfate-reducer and provided the anaerobic phenanthrene biotransformation pathway under sulfate-reducing conditions, which can act as a reference for practical applications in bioremediation and for studying the molecular mechanisms of phenanthrene in anaerobic zones.

Pilot-scale fermentation of urban food waste for volatile fatty acids production: The importance of pH.

Here, a pilot-scale volatile fatty acids (VFAs) production system was established using food waste (FW) as feedstock under acidic conditions. The effects of pH (uncontrolled, 4.5, 5.5, and 6.5) on the FW acidification system were investigated. The results showed that VFAs concentration increased from 8419 to 15048 mg COD/L with pH level increasing from 4.5 to 6.5, and the highest VFA production yield (0.79 mgCOD/mgCOD) was obtained at a pH of 6.5. A larger proportion of butyric acid (52.9%) was observed, accompanied by a 23% decrease of acetic acid when pH was elevated to 6.5. Microbial analysis showed that Clostridium sensu stricto 1, Sporanaerobacter, and Proteiniphilum were dominant, which not only positively influence the hydrolysis and acidogenesis processes but also play an essential role in the conversion of acetic acid to butyric acid. In summary, this study provides a valuable reference for large-scale FW treatment to recover valuable resources.

Investigation of anaerobic biodegradation of phenanthrene by a sulfate-dependent Geobacter sulfurreducens strain PheS2.

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous and harmful contaminants, which can be degraded aerobically. However, the persistence of PAHs in anoxic environments indicates that anaerobic biodegradation of PAHs should also be investigated. Pure-culture and biotransformation processes for anaerobic phenanthrene biodegradation with sulfate as a terminal electron acceptor remains in its infancy. In this study, we investigated anaerobic biodegradation of PAHs by PheS2, an isolated phenanthrene-utilizing sulfate-reducer, using phenanthrene as a model compound. PheS2 was phylogenetically closely related to Geobacter sulfurreducens and reduced sulfate to sulfide during anaerobic phenanthrene biodegradation. Phenanthrene biodegradation processes were detected using gas chromatography-mass spectrometry, genome, and reverse transcription quantitative PCR analyses. Carboxylation was the initial step of anaerobic phenanthrene biodegradation based upon detection of 2- and 4-phenanthroic acid, its isotopically labeled analogs when using 13C-labeled bicarbonate and fully deuterated-phenanthrene (C14D10), and genes encoding enzymes putatively involved in the biodegradation. Further, ring-system reducing and cleavage occurred, and substituted benzene series and cyclohexane derivatives were detected in downstream biotransformation metabolites. Additionally, PheS2 can degrade benzene, naphthalene, anthracene, and benz[a]anthracene, but not pyrene and benz[a]pyrene. This study describes the isolation of an anaerobic phenanthrene-degrading sulfate-reducer, the first pure-culture evidence of phenanthrene biotransformation processes with sulfate as an electron acceptor.

Exploration of the biotransformation processes in the biodegradation of phenanthrene by a facultative anaerobe, strain PheF2, with Fe(III) or O2 as an electron acceptor.

The study of biodegradation of polycyclic aromatic hydrocarbons (PAHs) with metal ions as electron acceptors is still in its infancy. Here, a pure culture of PheF2 sharing 99.79% 16S rRNA-sequence similarity with Trichococcus alkaliphilus, which was recently reported to degrade PAHs, was isolated and found to degrade PAHs with Fe (III) or O2 reduction. Phenanthrene was selected as a model of PAH to study the biodegradation process by PheF2 with Fe (III) or O2 as an electron acceptor. PheF2 exhibited nearly 100%, 37.1%, and 28.5% anaerobic biodegradation of phenanthrene at initial concentrations of 280.7 µM, 280.6 µM, and 281.3 µM, respectively, within 10 days under anaerobic conditions with XAD-7 as a carrier, heptamethylnonane (HMN) as a solution, and nothing, respectively. PheF2 could degrade nearly 100% of the initial phenanthrene concentration of 283.4 µM under aerobic conditions within three days. The initial step of phenanthrene biodegradation by PheF2 involved carboxylation and dioxygenation under anaerobic and aerobic conditions, respectively. The biotransformation processes of phenanthrene degradation by PheF2 with Fe(III) or O2 as an electron acceptor were explored by metabolite and genome analysis. These findings provide an important theoretical support for evaluation of PAHs fate and for PAHs pollution control or remediation in anaerobic and aerobic environments.

Anaerobic biodegradation of phenanthrene by a newly isolated nitrate-dependent Achromobacter denitrificans strain PheN1 and exploration of the biotransformation processes by metabolite and genome analyses.

Polycyclic aromatic hydrocarbons (PAHs) are widespread and harmful contaminants and are more persistent under anaerobic conditions. The bioremediation of PAHs in anaerobic zones has been enhanced by treating the contamination with nitrate, which is thermodynamically favourable, cost-effective, and highly soluble. However, anaerobic PAHs biotransformation processes that employ nitrate as an electron acceptor have not been fully explored. In this study, we investigated the anaerobic biotransformation of PAHs by strain PheN1, a newly isolated phenanthrene-degrading denitrifier, using phenanthrene as a model compound. PheN1 is phylogenetically closely related to Achromobacter denitrificans and reduces nitrate to nitrite (not N2 ) during the anaerobic phenanthrene degradation process. Phenanthrene biotransformation processes were detected using gas chromatography-mass spectrometry and were further examined by reverse transcription-quantitative PCR and genome analyses. Carboxylation and methylation were both found to be the initial steps in the phenanthrene degradation process. Downstream biotransformation processed benzene compounds and cyclohexane derivatives. This study describes the isolation of an anaerobic phenanthrene-degrading bacterium along with the pure-culture evidence of phenanthrene biotransformation processes with nitrate as an electron acceptor. The findings in this study can improve our understanding of anaerobic PAHs biodegradation processes and guide PAHs bioremediation by adding nitrate to anaerobic environments.

Investigation of anaerobic phenanthrene biodegradation by a highly enriched co-culture, PheN9, with nitrate as an electron acceptor.

In this study, we developed a highly enriched phenanthrene-degrading co-culture, PheN9, which uses nitrate as an electron acceptor under anaerobic conditions, and the processes mediating biodegradation were proposed. The dominant bacteria populations included Pseudomonas stutzeri (91.7% relative abundance), which shared 98% 16S rRNA-sequence similarity with the naphthalene-degrading, nitrate-reducing strain NAP-3-1, and Candidatus_Kuenenia (2.3% relative abundance), which is a type of anammox bacteria. Enrichment transformed 54% of the added phenanthrene, reduced nitrate, and generated significant amounts of nitrite. Enrichment also result in partial consumption of the produced nitrite by the anammox bacteria. The key initial steps of anaerobic phenanthrene biodegradation by PheN9 were methylation and carboxylation, which were identified for detection of metabolic products, as well as carboxylase and methyltransferase activities. The methylation product was then oxidized to 2-naphthoic acid and then underwent sequential biodegradation steps. Then, ring-system reducing occurred, and the metabolic products were identified as dihydro-, tetrahydro-, hexahydro-, and octahydro-2-phenanthroic acid. Downstream degradation proceeded via a substituted benzene series and cyclohexane derivatives. This study employed anaerobic phenanthrene-biodegradation processes with nitrate as an electron acceptor. These findings can improve our understanding of anaerobic polycyclic aromatic hydrocarbon (PAH) biodegradation processes and guide PAH bioremediation by adding nitrate to anaerobic environments.