Catechol Dioxygenase Research Articles

Degradation of phenol from water by Rhodococcus ruber promoted by MgO nanoparticles

How to improve the treatment efficiency has always been a key problem that needs to be solved in the bio-degradation of phenolic pollutant. The nanomaterials could improve the activity and tolerance of microorganisms, as well as the metabolic efficiency for pollutants under certain conditions. In this research, the effect of nano magnesium oxide (Nano-MgO) on the degradation of phenol by Rhodococcus ruber ATCC 31338 was studied. Moreover, the degradation mechanism of Rhodococcus ruber and Nano-MgO composite system was studied. Results showed that the addition of Nano-MgO could improve the phenol removal efficiency. With the presence of Nano-MgO at 100 mg/L to 300 mg/L, Rhodococcus ruber showed better growth and efficiency in removing phenol. However, when the concentration of Nano-MgO achieved 400 mg/L, the growth of Rhodococcus ruber was inhibited. Enzyme activity experiments suggested that the presence of Nano-MgO had a significant positive effect on the enzyme activity of catechol 1,2-dioxygenase (C12O) and catechol 2,3-dioxygenase (C23O), especially to C23O. The present of Nano-MgO might change the cleavage pathway of catechol. In addition, phosphorus is a limiting factor for the growth of Rhodococcus ruber. The addition of MgO-P materials prepared by encapsulating phosphorus on Nano-MgO can achieve efficient degradation of phenol by Rhodococcus ruber in the phosphorus loss system. This result also proved that nanomaterials can enhance the removal efficiency of organic matter by microorganisms. This study could provide a new solution for improving the microbial degradation rate of phenolic substances.

Influence of humic acid on the p-tert-Butylphenol removal efficiency by Spirodela polyrhiza-Tas13 association

Plant-microbe remediation technique is considered as a promising technology in removal of organic pollutants and its remediation efficiency is largely affected by a variety of surrounding environmental factors. Humic acid (HA) is the complex organic substance ubiquitous in environment, which characterized by its surfactant-like micelle microstructure and various reaction activity. In our study, a plant-microbe association with high p-tert-Butylphenol (PTBP) degradation potential constructed by Spirodela polyrhiza and Sphingobium phenoxybenzoativorans Tas13 has been used, and the influence of HA on the PTBP degradation efficiency of S. polyrhiza-Tas13 association was investigated. The result showed that the presence of HA greatly improved PTBP removal efficiency of S. polyrhiza-Tas13. The reason accounted for this may be due to the presence of HA promoted bacterial cell propagation, altered bacterial cell wall permeability, increased catechol 2,3-dioxygenase (C23O) enzyme activity of strain Tas13, rather than increasing the colonization ability of strain Tas13 on to the root surface. This study will greatly facilitate the application of aquatic plant-microbe association in environmental remediation.

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.

Optimizing carbon sources regulation in the biochemical treatment systems for coal chemical wastewater: Aromatic compounds biodegradation and microbial response strategies

The complex composition of coal chemical wastewater (CCW), marked by numerous highly toxic aromatic compounds, induces the destabilization of the biochemical treatment system, leading to suboptimal treatment efficacy. In this study, a biochemical treatment system was established to efficiently degrade aromatic compounds by quantitatively regulating the dosage of co-metabolized substrates (specifically, the chemical oxygen demand (COD) Glucose: COD Sodium acetate = 3:1, 1:3, and 1:1). The findings demonstrated that the system achieved optimal performance under the condition that the ratio of COD Glucose to COD Sodium acetate was 3:1. When the co-metabolized substrate was added to the system at an optimal ratio, examination of pollutant removal and cumulative effects revealed that the removal efficiencies for COD and total organic carbon (TOC) reached 94.61 % and 86.40 %, respectively. The removal rates of benzene series, nitrogen heterocyclic compounds, polycyclic aromatic hydrocarbons, and phenols were 100 %, 100 %, 63.58 %, and 94.12 %, respectively. Research on the physiological response of microbial cells showed that, under optimal ratio regulation, co-metabolic substrates led to a substantial rise in microbial extracellular polymeric substances (EPS) secretion, particularly extracellular proteins. When the system reached the end of its operation, the contents of loosely bound EPS (LB-EPS) and tightly bound EPS (TB-EPS) for proteins in the optimal group were 7.12 mg/g-SS and 152.28 mg/g-SS, respectively. Meanwhile, the ratio of α-Helix / (β-Sheet + Random coil) and the proportion of intermolecular interaction forces were also increased in the optimal group. At system completion, the ratio of α-Helix / (β-Sheet + Random coil) reached 0.717 (LB-EPS) and 0.618 (TB-EPS), respectively. Additionally, the proportion of intermolecular interaction forces reached 74.83 % (LB-EPS) and 55.03 % (TB-EPS). An in-depth analysis of the metabolic regulation of microorganisms indicated that the introduction of optimal ratios of co-metabolic substrates contributed to a noteworthy upregulation in the expression of Catechol 2,3-dioxygenase (C23O) and Dehydrogenase (DHA). The expression levels of C23O and DHA were measured at 0.029 U/mg Pro·g MLSS and 75.25 mg TF·(g MLSS·h)−1 (peak value), respectively. Correspondingly, enrichment of aromatic compound-degrading bacteria, including Thauera, Saccharimonadales, and Candidatus_Competibacter, occurred, along with the upregulation of associated functional genes such as Catechol 1,2-dioxygenase, Catechol 2,3-dioxygenase, Protocatechuate 3,4-dioxygenase, and Protocatechuate 4,5-dioxygenase. Considering the intricate system of multiple coexisting aromatic compounds in real CCW, this study not only obtained an optimal ratio for carbon source addition but also enhanced the efficient utilization of carbon sources and improved the capability of the system to effectively degrade aromatic compounds. Additionally, this paper established a theoretical foundation for metabolic regulation and harmless treatment within the biochemical treatment of intricate systems, exemplified by real CCW.

Decolorization and detoxification of Brilliant Crocein GR by a newly enriched thermophilic consortium

The environmental pollution caused by azo dyes at high temperatures has become an urgent problem. However, little attention has been paid to decolorizing azo dyes by thermophilic consortiums. In this study, a thermophilic bacterial consortium (BCGR-T) mainly composed of two genera, namely, Caldibacillus (70.90%) and Aeribacillus (17.63%) was first enriched, which can decolorize Brilliant Crocein GR (BCGR) at high temperatures (50–75 °C), pH values of 6∼8, dye concentrations (100–400 mg/L) and salinities (1–5%, w/v). The enzyme activity results showed that the azoreductase activity was nearly 8.8 times that of the control (p < 0.01), and the intracellular lignin peroxidase was also highly expressed with enzyme activity of 5.64 U (min−1 mg−1 protein) (p < 0.05), indicated that both azoreductase and intracellular lignin peroxidase played an important part in the decolorization process. Furthermore, seven new intermediate metabolic products, including aniline, phthalic acid, 2-carboxy benzaldehyde, phenylacetic acid, benzoic acid, toluene, and 4-methyl-hexanoic acid, were identified. In addition, functional genes related with the azo dye decolorization, such as those encoding the azoreductase, laccase, FMN reductase, NADPH-/NADH-quinone oxidoreductases and NADPH-/NADH dehydrogenases, catechol dioxygenase, homogentisate 1,2-dioxygenase, protocatechuate 3,4-dioxygenase, gentisate 1,2-dioxygenase, azobenzene reductase, naphthalene 1,2-dioxygenase, benzoate/toluate 1,2-dioxygenase, and anthranilate 1,2-dioxygenase and so on were found in the metagenome of the consortium BCGR-T. Finally, a new decolorization pathway of the thermophilic consortium BCGR-T was proposed. In addition, the phototoxicity of BCGR decreased after decolorization. Overall, the thermophilic consortium BCGR-T could be a promising candidate in the treatment of high concentration azo dye wastewater at high temperatures.

Batch-mode degradation of high-strength phenolic pollutants by Pseudomonas aeruginosa strain STV1713 immobilized on single and hybrid matrices.

Controlled environments are pivotal in all bioconversion processes, influencing the efficacy of biocatalysts. In this study, we designed a batch bioreactor system with a packed immobilization column and a decontamination chamber to enhance phenol and 2,4-dichlorophenol degradation using the hyper-tolerant bacterium Pseudomonas aeruginosa STV1713. When free cells were employed to degrade phenol and 2,4-DCP at a concentration of 1000mg/L, the cells completely removed the pollutants within 28h and 66h, respectively. Simultaneous reductions in chemical oxygen demand and biological oxygen demand were observed (phenol: 30.21mg/L/h and 16.92mg/L/h, respectively; 2,4-dichlorophenol: 12.85mg/L/h and 7.21mg/L/h, respectively). After assessing the degradation capabilities, the bacterium was immobilized on various matrices (sodium alginate, alginate-chitosan-alginate and polyvinyl alcohol-alginate) to enhance pollutant removal. Hybrid immobilized cells exhibited greater tolerance and degradation capabilities than those immobilized in a single matrix. Among them, polyvinyl alcohol-alginate immobilized cells displayed the highest degradation capacities (up to 2000mg/L for phenol and 2500mg/L for 2,4-dichlorophenol). Morphological analysis of the immobilized cells revealed enhanced cell preservation in hybrid matrices. Furthermore, the elucidation of the metabolic pathway through the catechol dioxygenase enzyme assay indicated higher activity of the catechol 1,2-dioxygenase enzyme, suggesting that the bacterium employed an ortho-degradation mechanism for pollutant removal. Additionally, enzyme zymography confirmed the presence of catechol 1,2-dioxygenase, with the molecular weight of the enzyme determined as 245kDa.

Acidovorax benzenivorans sp. nov., a novel aromatic hydrocarbon-degrading bacterium isolated from a xylene-degrading enrichment culture.

A Gram-stain-negative strain, designated as D2M1T was isolated from xylene-degrading enrichment culture and characterized using a polyphasic approach to determine its taxonomic position. The 16S rRNA gene sequence analysis revealed that strain D2M1T belongs to the genus Acidovorax, with the highest 16S rRNA gene similarity to Acidovorax delafieldii DSM 64T (99.93 %), followed by Acidovorax radicis DSM 23535T (98.77 %) and Acidovorax kalamii MTCC 12652T (98.76 %). The draft genome sequence of strain D2M1T is 5.49 Mb long, and the G+C content of the genome is 64.2 mol%. Orthologous average nucleotide identity and digital DNA-DNA hybridization relatedness values between strain D2M1T and its closest relatives were below the threshold values for species demarcation confirming that strain D2M1T is distinctly separated from its closest relatives. The whole genome analysis of the strain revealed a phenol degradation gene cluster, encoding a multicomponent phenol hydroxylase (mPH) together with a complete meta-cleavage pathway including an I.2.C-type catechol 2,3-dioxygenase (C23O) gene. The strain was able to degrade benzene and ethylbenzene as sole sources of carbon and energy under aerobic and microaerobic conditions. Cells were facultatively aerobic rods and motile with a single polar flagellum. The predominant fatty acids (>10 % of the total) of strain D2M1T were summed feature 3 (C16 : 1 ω7c/C16 : 1 ω6c), C16 : 0 and summed feature 8 (C18 : 1 ω7c/C18 : 1 ω6c). The major ubiquinone of strain D2M1T was Q8, while the major polar lipids were diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine. Based on polyphasic data, it is concluded that strain D2M1T represents a novel species of the genus Acidovorax, for which the name of Acidovorax benzenivorans sp. nov. is proposed. The type strain of the species is strain D2M1T (=DSM 115238T=NCAIM B.02679T).

Degradation and metagenomic analysis of 4-chlorophenol utilizing multiple metal tolerant bacterial consortium.

Chlorophenols are persistent environmental pollutants used in synthesizing dyes, drugs, pesticides, and other industrial products. The chlorophenols released from these processes seriously threaten the environment and human health. The present study describes 4-chlorophenol (4-CP) degradation activity and metagenome structure of a bacterial consortium enriched in a 4-CP-containing medium. The consortium utilized 4-CP as a single carbon source at a wide pH range, temperature, and in the presence of heavy metals. The immobilized consortium retained its degradation capacity for an extended period. The 4-aminoantipyrine colorimetric analysis revealed complete mineralization of 4-CP up to 200mg/L concentration and followed the zero-order kinetics. The addition of glycerol and yeast extract enhanced the degradation efficiency. The consortium showed both ortho- and meta-cleavage activity of catechol dioxygenase. Whole genome sequence (WGS) analysis revealed the microbial compositions and functional genes related to xenobiotic degradation pathways. The identified genes were mapped on the KEGG database to construct the 4-CP degradation pathway. The results exhibited the high potential of the consortium for bioremediation of 4-CP contaminated sites. To our knowledge, this is the first report on WGS analysis of a 4-CP degrading bacterial consortium.

Uncovering applicability of Navicula permitis algae in removing phenolic compounds: A promising solution for olive mill wastewater treatment

This work identifies Navicula permitis, freshwater diatom algae, by analyzing its 18S DNAr and testing how well it could grow and remove phenolic compounds at varying concentrations from 50 to 250 mg/L. The changes in the culture's chlorophyll fluorescence were measured when being exposed to stress. The biodegradation of phenolic compounds was tested by analyzing the enzyme activities of phenol hydroxlase and catechol dioxygenase in N. permitis. It was found that N. permitis could withstand up to 145.9 mg/L of phenol concentration. The response surface methodology was used to optimize conditions for N. permitis to degrade phenol with 50.08 mg/L of concentration, 106 cells/mL of N. permitis, and 11.38 days of treatment. Phenol removal by the Navicula permitis followed a zero-order kinetic model, and N. permitis used PHase to breakdown the pollutant. The ortho-pathway played a role in phenol metabolism. While degrading phenol, N. permitis produced biomass, which makes it a promising option for environmental remediation.

Nuclear Resonance Vibrational Spectroscopy Definition of Peroxy Intermediates in Catechol Dioxygenases: Factors that Determine Extra- versus Intradiol Cleavage.

The extradiol dioxygenases (EDOs) and intradiol dioxygenases (IDOs) are nonheme iron enzymes that catalyze the oxidative aromatic ring cleavage of catechol substrates, playing an essential role in the carbon cycle. The EDOs and IDOs utilize very different FeII and FeIII active sites to catalyze the regiospecificity in their catechol ring cleavage products. The factors governing this difference in cleavage have remained undefined. The EDO homoprotocatechuate 2,3-dioxygenase (HPCD) and IDO protocatechuate 3,4-dioxygenase (PCD) provide an opportunity to understand this selectivity, as key O2 intermediates have been trapped for both enzymes. Nuclear resonance vibrational spectroscopy (in conjunction with density functional theory calculations) is used to define the geometric and electronic structures of these intermediates as FeII-alkylhydroperoxo (HPCD) and FeIII-alkylperoxo (PCD) species. Critically, in both intermediates, the initial peroxo bond orientation is directed toward extradiol product formation. Reaction coordinate calculations were thus performed to evaluate both the extra- and intradiol O-O cleavage for the simple organic alkylhydroperoxo and for the FeII and FeIII metal catalyzed reactions. These results show the FeII-alkylhydroperoxo (EDO) intermediate undergoes facile extradiol O-O bond homolysis due to its extra e-, while for the FeIII-alkylperoxo (IDO) intermediate the extradiol cleavage involves a large barrier and would yield the incorrect extradiol product. This prompted our evaluation of a viable mechanism to rearrange the FeIII-alkylperoxo IDO intermediate for intradiol cleavage, revealing a key role in the rebinding of the displaced Tyr447 ligand in this rearrangement, driven by the proton delivery necessary for O-O bond cleavage.

Cumulative effects and metabolic characteristics of aromatic compounds in microbial cells during the biochemical treatment process of coal chemical wastewater

This study aimed to investigate the effectiveness of a biochemical treatment system for coal chemical wastewater (CCW) in degrading aromatic compounds. The study examined the degradation sequence of different compounds, including benzene series (BTEX), phenols, polycyclic aromatic hydrocarbons (PAHs), and nitrogen heterocyclic compounds (NHCs), in the system. The study also analyzed the metabolic characteristics of accumulated aromatic organic compounds in the cells of microorganisms. The results showed that the aromatic compounds were first adsorbed from water by extracellular polymers (EPS) before entering the cells. The removal rates of COD and TOC in CCW reached 95% after system operation for 8 h. BTEX and NHCs were completely degraded outside the cells within 2 h, whereas PAHs and some phenols accumulated inside the cells. The metabolic characteristics of the accumulated compounds were analyzed and found to be gradually degraded through the action of catechol dioxygenase. The study findings not only demonstrate the potential for achieving efficient biodegradation of aromatic compounds in CCW but also provide the essential theoretical underpinning for the regulation and environmentally benign treatment of such wastewater within a biochemical treatment system.

Microbial machinery dealing diverse aromatic compounds: Decoded from pelagic sediment ecogenomics in the gulfs of Kathiawar Peninsula and Arabian Sea

Aromatic hydrocarbons are persistent pollutants in aquatic systems as endocrine disruptors, significantly impacting natural ecosystems and human health. Microbes perform as natural bioremediators to remove and regulate aromatic hydrocarbons in the marine ecosystem. The present study focuses upon the comparative diversity and abundance of various hydrocarbon-degrading enzymes and their pathways from deep sediments along the Gulf of Kathiawar Peninsula and Arabian Sea, India. The elucidation of large number of degradation pathways in the study area under the presence of a wide range of pollutants whose fate needs to be addressed. Sediment core samples were collected, and the whole microbiome was sequenced. Analysis of the predicted ORFs (open reading frames) against the AromaDeg database revealed 2946 aromatic hydrocarbon-degrading enzyme sequences. Statistical analysis portrayed that the Gulfs were more diverse in degradation pathways compared to the open sea, with the Gulf of Kutch being more prosperous and more diverse than the Gulf of Cambay. The vast majority of the annotated ORFs belonged to groups of dioxygenases that included catechol, gentisate, and benzene dioxygenases, along with Rieske (2Fe–2S) and vicinal oxygen chelate (VOC) family proteins. From the sampling sites, only 960 of the total predicted genes were given taxonomic annotations, which mention the presence of many under-explored marine microorganism-derived hydrocarbon degrading genes and pathways. Through the present study, we tried to unveil the array of catabolic pathways of aromatic hydrocarbon degradation and genes from a marine ecosystem that upholds economic and ecological significance in India. Thus, this study provides vast opportunities and strategies for microbial resource recovery in marine ecosystems, which can be investigated to explore aromatic hydrocarbon degradation and their potential mechanisms under various oxic or anoxic environments. Future studies should focus on aromatic hydrocarbon degradation by considering degradation pathways, biochemical analysis, enzymatic, metabolic, and genetic systems, and regulations.

Reduction of phenolics in faba bean meal using recombinantly produced and purified Bacillus ligniniphilus catechol 2,3-dioxygenase

Pulse meal should be a valuable product in the animal feed industry based on its strong nutritional and protein profiles. However, it contains anti-nutritional compounds including phenolics (large and small molecular weight), which must be addressed to increase uptake by the industry. Microbial fermentation is currently used as a strategy to decrease larger molecular weight poly-phenolics, but results in the undesirable accumulation of small mono-phenolics. Here, we investigate cell-free biocatalytic reduction of phenolic content in faba bean (Vicia faba L.) meal. A representative phenolic ring-breaking catechol dioxygenase, Bacillus ligniniphilus L1 catechol 2,3-dioxygenase (BLC23O) was used in this proof-of concept based on its known stability and broad substrate specificity. Initially, large-scale fermentative recombinant production and purification of BLC23O was carried out, with functionality validated by in vitro kinetic analysis. When applied to faba bean meal, BLC23O yielded greatest reductions in phenolic content in a coarse air classified fraction (high carbohydrate), compared to either a fine fraction (high protein) or the original unfractionated meal. However, the upstream hydrolytic release of phenolics from higher molecular weight species (e.g. tannins, or complexes with proteins and carbohydrates) likely remains a rate limiting step, in the absence of other enzymes or microbial fermentation. Consistent with this, when applied to a selection of commercially available purified phenolic compounds, known to occur in faba bean, BLC23O was found to have high activity against monophenolic acids and little if any detectable activity against larger molecular weight compounds. Overall, this study highlights the potential viability of the biocatalytic processing of pulse meals, for optimization of their nutritional and economical value in the animal feed industry.Graphical

Micro-aeration assisted with electrogenic respiration enhanced the microbial catabolism and ammonification of aromatic amines in industrial wastewater

Improvement of refractory nitrogen-containing organics biodegradation is crucial to meet discharged nitrogen standards and guarantee aquatic ecology safety. Although electrostimulation accelerates organic nitrogen pollutants amination, it remains uncertain how to strengthen ammonification of the amination products. This study demonstrated that ammonification was remarkably facilitated under micro-aerobic conditions through the degradation of aniline, an amination product of nitrobenzene, using an electrogenic respiration system. The microbial catabolism and ammonification were significantly enhanced by exposing the bioanode to air. Based on 16S rRNA gene sequencing and GeoChip analysis, our results indicated that aerobic aniline degraders and electroactive bacteria were enriched in suspension and inner electrode biofilm, respectively. The suspension community had a significantly higher relative abundance of catechol dioxygenase genes contributing to aerobic aniline biodegradation and reactive oxygen species (ROS) scavenger genes to protect from oxygen toxicity. The inner biofilm community contained obviously higher cytochrome c genes responsible for extracellular electron transfer. Additionally, network analysis indicated the aniline degraders were positively associated with electroactive bacteria and could be the potential hosts for genes encoding for dioxygenase and cytochrome, respectively. This study provides a feasible strategy to enhance nitrogen-containing organics ammonification and offers new insights into the microbial interaction mechanisms of micro-aeration assisted with electrogenic respiration.

Isolation and characterization of Indigenous Bacilli strains from an oil refinery wastewater with potential applications for phenol/cresol bioremediation

Phenolic compounds are frequently occurring in wastewaters from various industrial processes at high concentrations, imposing prominent risk to aquatic biosphere and human health. Bioremediation has been proven to be an effective approach to remove these compounds, and hunting for functional organisms is still of primary importance to develop efficient processes. In this study, we report several newly isolated bacillus strains with superior performances in metabolizing phenols, one of which showed paramount efficiencies to metabolize phenol at concentrations up to 1200 mg L−1 and could simultaneously degrade a wide range of other phenolic compounds. The genes encoding for phenol hydroxylase (PH) and catechol-2,3-dioxygenase (C23O) have been detected and characterized, evidencing that phenol degradation occurs via the meta pathway. The GC level of the PH gene was found to be much higher than that of genes from other Bacilli but was quite close to that of the genes from Rhodococcus, and the induction of both enzymes by phenols was confirmed by RT-PCR experiments. We intend to believe this novel strain might be promising to serve as preferred organisms for developing more robust and efficient bioremediation processes of degrading phenolic compounds due to its validated performance.

Heavy Crude Oil Biodegradation: Catechol Dioxygenase Gene Copy Number Variation Determination by Droplet Digital Polymerase Chain Reaction

The crude oil reserves in Oman mainly consist of heavy oil. Microbial enhanced heavy oil recovery (MEOR) has been proved to be an efficient technique in the tertiary heavy oil recovery. Five Bacillus species potential for enhanced heavy oil recovery (EHOR) were isolated and the biodegradation ability of these isolates was studied. As heavy crude oil comprises of aromatic hydrocarbons rather than aliphatic ones, the aromatic catabolism gene, catechol 2,3-dioxygenase (C23O) and catechol 1,2-dioxygenase (C12O) were the genes of interest in this study along with the reference gene, 16S rDNA. The copy number variation of these genes was determined using droplet digital PCR (ddPCR). The primers and probes for ddPCR assay were designed targeting these genes. It was observed that the heavy crude oil biodegradation potential of the isolates correlated with the copy number of C23O gene in the microbial genomes. The isolate, Paenibacillus ehimensis BS1 had the highest C23O gene copy number (1.057) followed by Bacillus firmus BG4 (0.895) and Bacillus halodurans BG5 (0.031) as demonstrated by their biodegradation potential. This is one of the few studies deploying ddPCR in the field of heavy crude oil biodegradation by spore forming bacteria.

Risk assessment and biodegradation potential of PAHs originating from Map Ta Phut Industrial Estate, Rayong, Thailand

ABSTRACT Petroleum hydrocarbon contamination is a serious concern across the globe. Here, the capability of native bacterial consortium enriched from sediment samples of Map Ta Phut Industrial Estate (MTPIE), Rayong, Thailand was described. The distribution of PAHs was assessed from the sediment samples collected from MTPIE by GC-FID and the toxic unit (TU) was calculated to assess the potential ecological risk to the surrounding biota. This study investigated the degradation potential and determined the PAH-degrading bacterial cultures by enriching collected sediments in PAHs mixtures (naphthalene, phenanthrene, and pyrene). The TPH degradation capacity of each bacterial consortium was validated in a soil microcosm using aged crude oil-contaminated soil. The MTPIE sediments were highly contaminated with PAHs (843.99-3904.39 ng g-1) and posed extremely high ecological risks to benthic biota (TU > 1). The consortium S5-P most significantly removed naphthalene (90.03%) and phenanthrene (88.14%) while the highest removal of pyrene was achieved by the S3-P consortium. Other consortia only partially degraded the PAHs. The dominant microbes in the consortia were determined using PCR-DGGE, it was found that the PAH degrading consortia were known PAH degraders such as Annwoodia, Bacillus, Brevibacillus, Lysinibacillus, Paracoccus, Rhodococcus, Sphingopyxis, Sulfurovum, and Sulfurimonas species and unknown PAH degraders such as Lithuaxuella species. The consortium S5-P showed the highest degradation capacity, removing 74.99% of TPHs in the soil microcosm. Furthermore, the inoculation of PAH-biodegrading bacterial consortia significantly promoted the catechol-2,3-dioxygenase (C23O) and dehydrogenase (DHA) activities which directly correlated with the degradation efficiency of petroleum hydrocarbons (p < 0.05).

Bacterial degradation of mixed-PAHs and expression of PAH-catabolic genes.

Polyaromatic hydrocarbons (PAHs) are hazardous organic compounds with established toxicity, carcinogenicity, and mutagenicity, ubiquitous distribution, and persistence in different environmental matrices. In the present study, degradation of the mixture of PAHs (phenanthrene, anthracene, fluorene, and pyrene) by Kocuria flava and Rhodococcus pyridinivorans was investigated. The individual strains and consortium of both degraded 55.6%, 59.5%, and 59.1% of 10mgL-1 of mixed PAHs, respectively, within 15days. The participation of catabolic enzymes [catechol 2,3-dioxygenase (C23O), dehydrogenase (DH), and peroxidase (POD)] was confirmed during catalytic oxidation through meta-cleavage of mixed PAHs in this study. The catabolic gene expression of naphthalene dioxygenase (NAH) and catechol 2,3-dioxygenase (C23O) during degradation was confirmed using RT-qPCR in the present study. This is the first study that shows significant gene expression of the catabolic genes during degradation of mixed PAHs by selected bacterial strains. The C23O gene showed a 6.02 log fold higher expression in Kocuria flava in comparison to Rhodococcus pyridinivorans whereas NAH gene exhibited a 7.9 log fold higher expression in Rhodococcus pyridinivorans in comparison to Kocuria flava. Hence it is likely to conclude that combination of Kocuria flava and Rhodococcus pyridinivorans can effectively remove hazardous mixture of PAHs from the contaminated environmental matrix.

Synchronously enhanced biofilm formation and m-dichlorobenzene removal in biotrickling filters by rhamnolipid chelating rare earth elements

ABSTRACT The effects of rhamnolipid chelating La (III) or Nd (III) on m-dichlorobenzene removal and biofilm growth were investigated. The rhamnolipid chelating La (III) or Nd (III) significantly increased the average m-dichlorobenzene elimination capacity by 24% and 29%, respectively, on day 70–130, and the average wet weights of biomass ranged from 11 kg/m3 to 15 kg/m3 in BTFs on day 100–180 at an inlet load of at 47–98 gm3/h and an EBRT of 60s-90s. Microbial community functional genes about biofilm growth and m-dichlorobenzene removal were also increased. In addition, the rhamnolipid chelating La (III) or Nd (III) can improve the membrane permeability, extracellular polymeric substance (EPS) production, catechol 2,3 dioxygenase (C23O) enzyme, and chlorobenzene dioxygenase activity. These results demonstrate that rhamnolipid chelating La (III) or Nd (III) have the potential to improve the performance of BTFs treating hydrophobic and highly toxic VOCs.

Adaptation mechanism of a spider mite population to tea plants

In herbivores, adaptation to different plants often occurs during their ecological speciation. Plants have a variety of defensive systems, especially toxic and repellent secondary metabolites, to cope with herbivore attacks, while herbivores often enhance their xenobiotic metabolism for detoxifying the defensive compounds. Once a population acquires the adaptation ability to plant defense, it will occupy the plant as a host. This reduces interaction with other populations even within the same species, causing the population to diverge, and consequently, reproductive isolation occurs. The Kanzawa spider mite (KSM), Tetranychus kanzawai Kishida (Trombidiformes: Tetranychidae), is a polyphagous arthropod that causes serious economic damage to agricultural and horticultural crops. KSM is an important pest of tea plants, Camellia sinensis (L.) Kuntze. There are adapted and non-adapted KSM populations to tea plants (adapted and non-adapted KSM). Both populations may have undergone ecological speciation through the interaction with tea plants. However, the mechanism underlying the differences between two KSM populations remains unclarified. The two-spotted spider mite (TSSM), Tetranychus urticae Koch (Trombidiformes: Tetranychidae), is closely related to KSM. Based on the highly-annotated TSSM genome sequence, we conducted comparative transcriptome and proteome analyses among adapted and non-adapted KSM, and TSSM on tea plants and the preferable host bean plants. Xenobiotic-metabolizing enzymes containing catechol dioxygenase, carboxylesterase, ABC transporter, Cathepsin L, and UDP-glycosyltransferase were upregulated in adapted KSM on tea plants. Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that pathways related to oxidoreductase activity were particularly enriched in adapted KSM on tea plants. These enzymes should be dealing with catechins which are known as secondary metabolites of tea plants and to be toxic to spider mites. The survival rate in adapted KSM on tea plants was significantly reduced by RNAi targeting the gene encoding a catechol dioxygenase (TkCTD). These results suggest that TkCTD-mediated detoxification is involved in the adaptation of KSM to tea plants.