Degradation of hexane and other recalcitrant hydrocarbons by a novel isolate, <Emphasis Type="Italic">Rhodococcus</Emphasis> sp. EH831

Background, aim, and scope  

Hexane, a representative VOC, is used as a solvent for extraction and as an ingredient in gasoline. The degradation of hexane by bacteria is relatively slow due to its low solubility. Moreover, the biodegradation pathway of hexane under aerobic conditions remains to be investigated; therefore, a study relating to aerobic biodegradation mechanisms is required. Consequently, in this study, an effective hexane degrader was isolated and the biodegradation pathway examined for the first time. In addition, the degradation characteristics of a variety of recalcitrant hydrocarbons were qualitatively and quantitatively investigated using the isolate.     

Role of cyanobacteria in the biodegradation of crude oil by a tropical cyanobacterial mat

Cyanobacterial mats are ubiquitous in tropical petroleum-polluted environments. They form a high biodiversity microbial consortium that contains efficient hydrocarbons degraders. A cyanobacterial mat collected from a petroleum-contaminated environment located in Indonesia was studied for its biodegradation potential. In the field, the natural mat was shown to degrade efficiently the crude oil present in the environment. This natural mat demonstrated also a strong activity of degradation on model crude oil under laboratory conditions. In axenic cultures, the monospecific cyanobacterium Phormidium animale that constitute the bulk of the biomass did not exhibit any degradative capacity on hydrocarbons in the range of C13-C35 carbon atom number either in autotrophic or heterotrophic conditions. It was concluded that this cyanobacterial strain living on a heavily contaminated site had no direct effect on biodegradation of crude oil, the degradation activity being exclusively achieved by the other microorganisms present in the microbial consortium of the mat.     

Characteristics of styrene degradation by Rhodococcus pyridinovorans isolated from a biofilter

A novel strain (PYJ-1) of Rhodococcus pyridinovorans that was isolated from a biofilter was able to degrade styrene at a maximum rate of 0.16 mg (mg protein)(-1) h(-1) in batch culture at 97 mg l(-1) of initial styrene gas concentration. The optimum pH and temperature for styrene degradation were 7 and 32 degrees C, respectively. The degradation kinetic constants were obtained using substrate inhibition kinetics. In a perlite-packed biofilter the maximum styrene removal rate by the strain was 279 gm(-3)h(-1). Styrene removal in the biofilter was more sensitive to the temperature than in the batch culture.     

Microbial degradation of 17β -estradiol and 17α -ethinylestradiol followed by a validated HPLC-DAD method

This work aimed at studying the biodegradation of two estrogens, 17α -estradiol (E2) and 17β -ethinylestradiol (EE2), and their potential metabolism to estrone (E1) by microbial consortia. The biodegradation studies were followed by High Performance Liquid Chromatography–Diode Array Detector (HPLC–DAD) using a specifically developed and validated method. Biodegradation studies of the estrogens (E2 and EE2) were carried out with activated sludge (consortium A, CA) obtained from a Wastewater Treatment Plant (WWTP) and with a microbial consortium able to degrade recalcitrant compounds, namely fluorobenzene (consortium B, CB). E2 was more extensively degraded than EE2 by CA whereas CB was only able to degrade E2. The addition of acetate as a supplementary carbon source led to a faster biodegradation of E2 and EE2. E1 was detected as a metabolite only during the degradation of E2. The 16S rRNA gene sequence analyses of strains recovered from the degrading cultures revealed the presence of the genera Pseudomonas, Chryseobacterium and Alcaligenes. The genera Pseudomonas and Chryseobacterium were retrieved from cultures supplied with E2 and EE2, while the genus Alcaligenes was found in the presence of E2, suggesting that they might be involved in the degradation of these compounds.     

Isolation of a selected microbial consortium capable of degrading methyl parathion and p-nitrophenol from a contaminated soil site

A bacterial consortium with the ability to degrade methyl parathion and p-nitrophenol, using these compounds as the only carbon source, was obtained by selective enrichment in a medium with methyl parathion. Samples were taken from Moravia, Medellin; an area that is highly contaminated, owing to the fact that it was used as a garbage dump from 1974 to 1982. Acinetobacter sp, Pseudomonas putida, Bacillus sp, Pseudomonas aeruginosa Citrobacter freundii, Stenotrophomonas sp, Flavobacterium sp, Proteus vulgaris, Pseudomonas sp, Acinetobacter sp, Klebsiella sp and Proteus sp were the microorganisms identified within the consortium. In culture, the consortium was able to degrade 150 mg L?1 of methyl-parathion and p-nitrophenol in 120 h, but after adding glucose or peptone to the culture, the time of degradation decreased to 24 h. In soil, the consortium was also able to degrade 150 mg L?1 of methyl parathion in 120 h at different depths and also managed to decrease the toxicity.     

Biological treatment of a contaminated gaseous emission from a leather industry in a suspended-growth bioreactor

A suspended-growth bioreactor (SGB) was operated for the treatment of a gaseous stream mimicking emissions generated at a leather industrial company. The main volatile organic compounds (VOCs) present in the gaseous stream consisted of 1-methoxy-2-propanol, 2,6-dimethyl-4-heptanone, 2-butoxyethanol, toluene and butylacetate. A microbial consortium able to degrade these VOCs was successfully enriched. A laboratory-scale SGB was established and operated for 210-d with an 8h cycle period and with shutdowns at weekends. Along this period, the SGB was exposed to organic loads (OL) between 6.5 and 2.3 x 10(2) g h(-1) m(-3). Most of the compounds were not detected at the outlet of the SGB. The highest total VOC removal efficiency (RE) (ca 99%) was observed when an OL of 1.6 x 10(2) g h(-1) m(-3) was fed to the SGB. The maximum total VOC elimination capacity (1.8 x 10(2) g h(-1) m(-3)) was achieved when the OL applied to the SGB was 2.3 x 10(2) g h(-1) m(-3). For all the operating conditions, the SGB showed high levels of degradation of toluene and butylacetate (RE approximately equal to 100%). This study also revealed that recirculation of the gaseous effluent improved the performance of the SGB. Overall, the SGB was shown to be robust, showing high performance after night and weekend shutdown periods.     

Bioremediation of acidic oily sludge-contaminated soil by the novel yeast strain Candida digboiensis TERI ASN6

Background, aim, and scope

Primitive wax refining techniques had resulted in almost 50,000 tonnes of acidic oily sludge (pH 1–3) being accumulated inside the Digboi refinery premises in Assam state, northeast India. A novel yeast species Candida digboiensis TERI ASN6 was obtained that could degrade the acidic petroleum hydrocarbons at pH 3 under laboratory conditions. The aim of this study was to evaluate the degradation potential of this strain under laboratory and field conditions.

Materials and methods

The ability of TERI ASN6 to degrade the hydrocarbons found in the acidic oily sludge was established by gravimetry and gas chromatography–mass spectroscopy. Following this, a feasibility study was done, on site, to study various treatments for the remediation of the acidic sludge. Among the treatments, the application of C. digboiensis TERI ASN6 with nutrients showed the highest degradation of the acidic oily sludge. This treatment was then selected for the full-scale bioremediation study conducted on site, inside the refinery premises.

Results

The novel yeast strain TERI ASN6 could degrade 40 mg of eicosane in 50 ml of minimal salts medium in 10 days and 72% of heneicosane in 192 h at pH 3. The degradation of alkanes yielded monocarboxylic acid intermediates while the polycyclic aromatic hydrocarbon pyrene found in the acidic oily sludge yielded the oxygenated intermediate pyrenol. In the feasibility study, the application of TERI ASN6 with nutrients showed a reduction of solvent extractable total petroleum hydrocarbon (TPH) from 160 to 28.81 g kg^?1 soil as compared to a TPH reduction from 183.85 to 151.10 g kg^?1 soil in the untreated control in 135 days. The full-scale bioremediation study in a 3,280-m² area in the refinery showed a reduction of TPH from 184.06 to 7.96 g kg^?1 soil in 175 days.

Discussion

Degradation of petroleum hydrocarbons by microbes is a well-known phenomenon, but most microbes are unable to withstand the low pH conditions found in Digboi refinery. The strain C. digboiensis could efficiently degrade the acidic oily sludge on site because of its robust nature, probably acquired by prolonged exposure to the contaminants.

Conclusions

This study establishes the potential of novel yeast strain to bioremediate hydrocarbons at low pH under field conditions.

Recommendations and perspectives

Acidic oily sludge is a potential environmental hazard. The components of the oily sludge are toxic and carcinogenic, and the acidity of the sludge further increases this problem. These results establish that the novel yeast strain C. digboiensis was able to degrade hydrocarbons at low pH and can therefore be used for bioremediating soils that have been contaminated by acidic hydrocarbon wastes generated by other methods as well.     

Analysis of polycyclic aromatic hydrocarbons in sediment reference materials by microwave-assisted extraction

The microwave-assisted extraction (MAE) of polycyclic aromatic hydrocarbons (PAHs) from harbor sediment reference material EC-1, marine sediment reference material HS-2 and PAH-spiked river bed soil was conducted. The extraction conditions for EC-1 were carried out at 70 degrees C and 100 degrees C under pressure in closed vessels with cyclohexane acetone (1:1), cyclohexane-water (3:1), hexane acetone (1:1), and hexane-water (3:1) for 10 min. A comparison between MAE and a 16-h Soxhlet extraction (SX) method showed that both techniques gave comparable results with certified values. MAE has advantages over the currently used Soxhlet technique due to a faster extraction time and lower quantity of solvent used. The consumption of organic solvent of the microwave method was less than one-tenth compared to Soxhlet.     

Biodegradation of polycyclic aromatic hydrocarbons by a mixed culture

We investigated the potential biodegradation of polycyclic aromatic hydrocarbons (PAHs) by an aerobic mixed culture utilizing phenanthrene as its carbon source. Following a 3-5 h post-treatment lag phase, complete degradation of 5 mg/l phenanthrene occurred within 28 h (optimal conditions determined as 30 degrees C and pH 7.0). Phenanthrene degradation was enhanced by the individual addition of yeast extract, acetate, glucose or pyruvate. Results show that the higher the phenanthrene concentration, the slower the degradation rate. While the mixed culture was also capable of efficiently degrading pyrene and acenaphthene, it failed to degrade anthracene and fluorene. In samples containing a mixture of the five PAHs, treatment with the aerobic culture increased degradation rates for fluorene and anthracene and decreased degradation rates for acenaphthene, phenanthrene and pyrene. Finally, it was observed that when nonionic surfactants were present at levels above critical micelle concentrations (CMCs), phenanthrene degradation was completely inhibited by the addition of Brij 30 and Brij 35, and delayed by the addition of Triton X100 and Triton N101.     

Degradation characteristics of toluene,benzene, ethylbenzene,and xylene by Stenotrophomonas maltophilia T3-c

Stenotrophomonas maltophilia T3-c, isolated from a biofilter for the removal of benzene, toluene, ethylbenzene, and xylene (BTEX), could grow in a mineral salt medium containing toluene, benzene, or ethylbenzene as the sole source of carbon. The effect of environmental factors such as initial toluene mass, medium pH, and temperature on the degradation rate of toluene was investigated. The cosubstrate interactions in the BTEX mixture by the isolate were also studied. Within the range of initial toluene mass (from 23 to 70 pmol), an increased substrate concentration increased the specific degradation of toluene by S. maltophilia T3-c. The toluene degradation activity of S. maltophilia T3-c could be maintained at a broad pH range from 5 to 8. The rates at 20 and 40 degrees C were 43 and 83%, respectively, of the rate at 30 degrees C. The specific degradation rates of toluene, benzene, and ethylbenzene by strain T3-c were 2.38, 4.25, and 2.06 micromol/g-DCW/hr. While xylene could not be utilized as a growth substrate by S. maltophilia T3-c, the presence of toluene resulted in the cometabolic degradation of xylene. The specific degradation rate of toluene was increased by the presence of benzene, ethylbenzene, or xylene in binary mixtures. The presence of toluene or xylene in binary mixtures with benzene increased the specific degradation rate of benzene. The presence of ethylbenzene in binary mixtures with benzene inhibited benzene degradation. The presence of more than three kinds of substrates inhibited the specific degradation rate of benzene. All BTEX mixtures, except tri-mixtures of benzene, ethylbenzene, and xylene or mixtures of all four substrates, had little effect on the degradation of ethylbenzene by S. maltophilia T3-c. The utilization preference of the substrates by S. maltophilia T3-c was as follows: ethylbenzene was degraded fastest, followed by toluene and benzene. However, the specific degradation rates of substrates, in order, were benzene, toluene, and ethylbenzene.     

Slurry-phase biodegradation of weathered oily sludge waste

We assessed the biodegradation of a typical oily sludge waste (PB401) in Mexico using several regimes of indigenous microbial consortium and relevant bioremediation strategies in slurry-phase system. Abiotic loss of total petroleum hydrocarbons (TPH) in the PB401 was insignificant, and degradation rates under the various treatment conditions ranged between 666.9 and 2168.7 mg kg(-1) day(-1) over a 15 days reaction period, while viable cell count peaked at between log(10)5.7 and log(10)7.4 cfu g(-1). Biostimulation with a commercial fertilizer resulted in 24% biodegradation of the TPH in the oily waste and a corresponding peak cell density of log(10)7.4 cfu g(-1). Addition of non-indigenous adapted consortium did not appear to enhance the removal of TPH from the oily waste. It would appear that the complexities of the components of the alkylaromatic fraction of the waste limited biodegradation rate even in a slurry system.     

Biodegradation of phenanthrene in soil.

We investigated the potential of an aerobic polycyclic aromatic hydrocarbon (PAH)-adapted consortium to degrade phenanthrene in soil. Optimal degradation conditions were determined as pH7.0 and 30 degrees C with a water content of 100% wt soil/wt water (w/w). At a concentration of 5 microg/g, phenanthrene degradation (k1) was measured at 0.0269 l/hr with a half-life (t(1/2)) of 25.8 hrs. Our results show that the higher the phenanthrene concentration, the slower the degradation rates. Phenanthrene degradation was enhanced by treatment with yeast extract, glucose, or pyruvate, but was not significantly improved by the addition of acetate. Degradation was delayed by the addition of either compost or potassium nitrate and enhanced by the addition of nonionic surfactants (Brij30, Brij35, Triton X100 or Triton N101) at critical micelle concentration (CMC). Phenanthrene degradation was delayed at levels above CMC.     

Biodesulfurization of dibenzothiophene by growing cells of Pseudomonas putida CECT 5279 in biphasic media

Several studies have proven that natural or genetically modified bacteria, such as Pseudomonas putida strain, degrade recalcitrant organic sulfur compounds. However, from a practical point of view, the biodesulfurization (BDS) process has to be performed with really high proportions of organic solvents. In this work, the dibenzothiophene (DBT) was selected as recalcitrant model compound, and hexadecane as model organic solvent. It has been observed that P. putida CECT 5279 was able to desulfurize DBT even in the presence of 50% (v/v) of hexadecane. A concentration of 400 ppm of DBT was converted at a specific rate of generation of desulfurized final product, 2-hydroxybiphenyl (HBP), of 2.3 and 1.5 mg HBP L-1 (g DC L-1 h)-1 for 27% and 50% (v/v) of hexadecane, respectively. Finally, the Haldane kinetic model was used to describe the process evolution. The study is relevant as it has been proven that the strain CECT 5279 is a potential biocatalyst for developing an efficient BDS process.     

MBR处理低浓度稠油废水

为解决稠油废水达标排放问题,构建了一套中试实验装置,以经过除油、浮选和过滤预处理的富含溶解性有机化合物、氮磷缺乏的低浓度难生化稠油废水为原水,进行了187 d的连续运行,结果表明,经过厌氧和好氧生物处理后,出水COD可降至80~100 mg/L以下,再经膜过滤后COD降至60~80 mg/L,加入少量的粉末活性炭进行吸附处理后,出水COD可稳定在50 mg/L以下;水力停留时间从72 h降至30 h时,出水COD基本无变化;气相色谱-质谱分析表明该系统容易去除的有机物为酮类、醇类等物质,而烷烃(C17~C25)和芳烃等为本工艺难降解物质,通过膜生物反应器工艺优化或选择专门降解菌如Acinetobacter spp.进行处理可进一步降低出水COD的浓度。     

不适宜生长条件对混茵降解苯胺的影响

分别从台州和衢州某化工厂的好氧池中分离筛选得到2株苯胺降解菌TZl和JH1,经16SrDNA测序鉴定为Comamonassp．TZ1和Pseudomonassp．JH1,均具有较强的苯胺降解能力,培养24h后,可使初始浓度为800mg／L的苯胺去除率达到96．4％～98．4％。在此基础上,按体积比l：1将2株菌液进行混合构建了混合菌体系,进而对比考察了苯胺初始浓度、pH、盐度和重金属等环境因子对单一菌和混合菌生长量及降解苯胺效果的影响,重点探讨混合菌对不适宜生长环境的适应性及其对苯胺的降解特性。通过单一菌和混合菌对比实验发现,在适宜苯胺初始浓度、pH和盐度条件下,混合菌的生长量略高于单一菌;在不适宜生长的高浓度苯胺、pH和盐度条件下,混合菌也表现出了更强的适应性和苯胺矿化能力。Zn2＋和Cr6＋耐受实验则表明,对于Cr6＋混合菌表现出了更强的耐受能力,而对于zn2＋并没有表现出更强的耐受能力。     

Biodegradation of benzene, toluene, and xylene (BTX) in liquid culture and in soil by Bacillus subtilis and Pseudomonas aeruginosa strains and a formulated bacterial consortium

Purpose

The major aromatic constituents of petroleum products viz. benzene, toluene, and mixture of xylenes (BTX) are responsible for environmental pollution and inflict serious public concern. Therefore, BTX biodegradation potential of individual as well as formulated bacterial consortium was evaluated. This study highlighted the role of hydrogen peroxide (H₂O₂), nitrate, and phosphate in stimulating the biodegradation of BTX compounds under hypoxic condition.

Materials and methods

The individual bacterium viz. Bacillus subtilis DM-04 and Pseudomonas aeruginosa M and NM strains and a consortium comprising of the above bacteria were inoculated to BTX-containing liquid medium and in soil. The bioremediation experiment was carried out for 120?h in BTX-containing liquid culture and for 90?days in BTX-contaminated soil. The kinetics of BTX degradation either in presence or absence of H₂O₂, nitrate, and phosphate was analyzed using biochemical and gas chromatographic (GC) technique.

Results

Bacterial consortium was found to be superior in degrading BTX either in soil or in liquid medium as compared to degradation of same compounds by individual strains of the consortium. The rate of BTX biodegradation was further enhanced when the liquid medium/soil was exogenously supplemented with 0.01?% (v/v) H₂O₂, phosphate, and nitrate_. The GC analysis of BTX biodegradation (90?days post-inoculation) in soil by bacterial consortium confirmed the preferential degradation of benzene compared to m-xylene and toluene.

Conclusions

It may be concluded that the bacterial consortium in the present study can degrade BTX compounds at a significantly higher rate as compared to the degradation of the same compounds by individual members of the consortium. Further, addition of H₂O₂ in the culture medium as an additional source of oxygen, and nitrate and phosphate as an alternative electron acceptor and macronutrient, respectively, significantly enhanced the rate of BTX biodegradation under oxygen-limited condition.     

Two-phase partitioning bioreactor for the biodegradation of high concentrations of pentachlorophenol using Sphingobium chlorophenolicum DSM 8671

An organic-aqueous two-liquid-phase partitioning system was developed to degrade high concentrations of pentachlorophenol (PCP). Dioctyl sebacate was selected among 12 non-aqueous phases as the most suitable solvent to control the delivery of PCP to the aqueous phase for being non-biodegradable and biocompatible. In shake-flask experiments, the two-phase system was able to support the removal of 1g PCP l(-1) of total liquid phase. The performance of the two-liquid phase partitioning system (TLPPS) in shake-flask was evaluated under different conditions. At the initial biomass concentrations of 7, 25, and 58 mg dry weight l(-1), the volumetric removal rates of PCP obtained were 25.7+/-0.5, 32.1+/-0.1, and 39.3+/-2.9 mg PCP l(-1)h(-1), respectively. Higher performance was observed at lower organic-aqueous phase ratios (16% and 28%) than higher ones (37% and 44%). In a 2-l TLPPS, the degradation of 10 g PCP was completed in less than 100 h at a total volumetric rate of 142 mg l(-1) h(-1). Kinetics study using Monod model showed that compared to monophasic systems, the biphasic system significantly enhanced the maximum specific growth rate and PCP removal rate. Results of this biphasic system showed no accumulation of unknown by-product(s) which has been reported for physical-pretreatment or high-performance biphasic systems of PCP degradation.     

Degradation of volatile organic compounds with thermally activated persulfate oxidation

This study investigated the extent and treatability of the degradation of 59 volatile organic compounds (VOCs) listed in the EPA SW-846 Method 8260B with thermally activated persulfate oxidation. Data on the degradation of the 59 VOCs (in mixture) reacted with sodium persulfate in concentrations of 1 g l(-1) and 5 g l(-1) and at temperatures of 20 degrees C, 30 degrees C, and 40 degrees C were obtained. The results indicate that persulfate oxidation mechanisms are effective in degrading many VOCs including chlorinated ethenes (CEs), BTEXs and trichloroethanes that are frequently detected in the subsurface at contaminated sites. Most of the targeted VOCs were rapidly degraded under the experimental conditions while some showed persistence to the persulfate oxidation. Compounds with "CC" bonds or with benzene rings bonded to reactive functional groups were readily degraded. Saturated hydrocarbons and halogenated alkanes were much more stable and difficult to degrade. For those highly persulfate-degradable VOCs, degradation was well fitted with a pseudo first-order decay model. Activation energies of reactions of CEs and BTEXs with persulfate were determined. The degradation rates increased with increasing reaction temperature and oxidant concentration. Nevertheless, to achieve complete degradation of persulfate-degradable compounds, the systems required sufficient amounts of persulfate to sustain the degradation reaction.     

Degradation of 3,4-dichloro- and 3,4-difluoroaniline by Pseudomonas fluorescens 26-K

3,4-Dichloro- and 3,4-difluoroanilines were degraded by Pseudomonas fluorescens 26-K under aerobic conditions. In the presence of glucose strain degraded 170 mg/L of 3,4-dichloroaniline (3,4-DCA) during 2-3 days. Increasing of toxicant concentration up to 250 mg/L led to degradation of 3,4-DCA during 4 days and its intermediates during 5-7 days. Without cosubstrate and nitrogen source degradation of 3,4-DCA took place too, but more slowly--about 40% of toxicant at initial concentration 75 mg/L was degraded during 15 days. 3,4-Difluoroaniline (3,4-DFA) (initial concentration 170 mg/L) was degraded by Pseudomonas fluorescens 26-K during 5-7 days. The strain was able to completely degrade up to 90 mg/L of 3,4-DFA, without addition of cosubstrate and nitrogen during 15 days. Degradation of fluorinated aniline was accompanied by intensive defluorination. Activity of catechol 2,3-dioxygenase (C2,3DO) (0.230 micromol/min/mg of protein) was found in the culture liquid of the strain, grown with 3,4-DCA and glucose. This fact, as well as, the presence of 3-chloro-4-hydroxyaniline as a metabolite suggested that 3,4-DCA degradation pathway includes dehalogenation and hydroxylation of aromatic ring followed by its subsequent cleaving by C2,3DO. On the contrary, activity of catechol 1,2-dioxygenase (C1,2DO) (0.08 micromol/min/mg of protein) was found in the cell-free extract of biomass grown on 3,4-DFA. 3-Fluoro-4-hydroxyaniline as intermediate was found in this cell-free extract.     

Alginate immobilized enrichment culture for atrazine degradation in soil and water system

An atrazine degrading enrichment culture, a consortium of bacteria of genus Bacillus along with Pseudomonas and Burkholderia, was immobilized in sodium alginate and was used to study atrazine degradation in mineral salts medium (MSM), soil and wastewater effluent. Sodium alginate immobilized consortium, when stored at room temperature (24 ± 5°C), was effective in degrading atrazine in MSM up to 90 days of storage. The survival of bacteria in alginate beads, based on colony formation unit (CFU) counts, suggested survival up to 90 days and population counts decreased to 1/5^th on 120 days. Comparison of atrazine degrading ability of the freely suspended enrichment culture and immobilized culture suggested that the immobilized culture took longer time for complete degradation of atrazine as a lag phase of 2 days was observed in the MSM inoculated with alginate immobilized culture. The free cells resulted in complete degradation of atrazine within 6 days, while immobilized cells took 10 days for 100% atrazine degradation. Further, immobilized cultures were able to degrade atrazine in soil and wastewater effluent. Alginate beads were stable and effective in degrading atrazine till 3rd transfer and disintegrated thereafter. The study suggested that immobilized enrichment culture, due to its better storage and application, can be used to degrade atrazine in soil water system.