Characterization of bacterial strains able to grow on high molecular mass residues from crude oil processing

Oil residues containing high molecular mass hydrocarbons, rich in polyaromatic compounds, are frequent end-products of crude oil processing and are poorly biodegradable. Their disposal poses an environmental problem. Through batch-enrichments from contaminated soils we have isolated and characterized seven bacterial strains that can use a residue from crude oil processing as a source of carbon and energy. The residue was a complex mixture of high molecular mass compounds, including saturated, aromatic and polycyclic aromatic hydrocarbons (PAHs). Analysis of the metabolic profiles of the strains isolated showed that they could all metabolize long-chain-length alkanes efficiently, but not PAHs. Strains degrading naphthalene, a simple PAH, did exist in the soil inocula used, but could be isolated only when enrichments were performed using pure naphthalene as the sole carbon source. All strains tested emulsified the oil residue and their ability to produce surfactants was studied.     

Bioprocessing-Based Approach for Bitumen/Water/Fines Separation and Hydrocarbon Recovery from Oil Sands Tailings

Oil sands are a major source of oil, but their industrial processing generates tailings ponds that are an environmental hazard. The main concerns are mature fine tailings (MFT) composed of residual hydrocarbons, water, and fine clay. Tailings ponds include toxic contaminants such as heavy metals, and toxic organics including naphthenics. Naphthenic acids and polyaromatic hydrocarbons (PAHs) degrade very slowly and pose a long-term threat to surface and groundwater, as they can be transported in the MFT. Research into improved technologies that would enable densification and settling of the suspended particles is ongoing. In batch tests, BioTiger?, a microbial consortium that can metabolize PAHs, demonstrated improved oil sands tailings settling from a Canadian tailings pond. Results also showed, depending on the timing of the measurements, lower suspended solids and turbidity. Elevated total organic carbon was observed in the first 48 hours in the BioTiger?-treated columns and then decreased in overlying water. Oil sands tailings mixed with BioTiger? showed a two-fold reduction in suspended solids within 24 hours as compared to abiotic controls. The tailings treated with BioTiger? increased in microbial densities three orders of magnitude from 8.5 × 105 CFU/mL to 1.2 × 108 CFU/mL without any other carbon or energy source added, indicating metabolism of hydrocarbons and other available nutrients. Results demonstrated that bioaugmentation of BioTiger? increased separation of organic carbon from particles in oil sands and enhanced settling with tailings with improved water quality. Journal style is for Abstract to be less than 200 words, and contain no citations to other sources; please edit as needed     

石油污染土壤自然衰减过程多环芳烃降解基因动态特征

新疆石油污染土壤中微生物多环芳烃(polycyclic aromatic hydrocarbons,PAHs)降解功能基因研究甚少,且环境因子和功能基因之间相关性仍不清楚。【目的】揭示新疆石油污染砂质土壤自然衰减过程中多环芳烃降解关键基因结构和变化规律。【方法】以新疆准东油田为研究区,分析同一采油区不同石油污染年限土壤理化因子和多环芳烃含量变化,采用扩增子测序研究石油自然衰减过程中多环芳烃降解酶基因结构变化规律,利用Mental检验探讨其环境驱动因子。【结果】石油污染时间1年和3年的土壤中有多项理化指标与背景土存在显著性差异,而污染5年土壤与背景土之间仅2项指标具有显著性差异,随石油自然衰减逐渐恢复至正常。石油污染1年的土壤中16种多环芳烃除苊烯和?以外,其余14种多环芳烃均高于石油污染3年和5年土壤,多环芳烃总量和含油率污染1年土壤均显著高于污染3年和5年的土壤,多环芳烃会在污染后短时间内迅速被降解。扩增子测序结果显示,萘双加氧酶基因分类操作单元(operational taxonomic units,OTUs)序列随污染年限延长逐渐增多;芳环羟化双加氧酶基因OTUs序列BLAST(...     

Degradation of Petroleum by an Alga, Prototheca Zopfii

Prototheca zopfii is an achlorophyllous alga which degrades oil. It has been found to degrade 10 and 40% of a motor oil and crude oil, respectively, when tested under appropriate conditions. Degradation of the crude oil observed in this study compares well with the amount of degradation accomplished by bacteria. P. zopfii was found to degrade a greater percentage of the aromatic hydrocarbons in motor oil than of the saturated hydrocarbons and a greater percentage of saturated hydrocarbons in crude oil than of aromatic hydrocarbons. Resins and asphaltens were produced during degradation of motor oil, whereas these fractions in crude oil were degraded. P. zopfii did not demonstrate preferential utilization of lower homologues of cycloalkanes and aromatics as has been observed with bacteria.     

Enhanced Biodegradation of Casablanca Crude Oil by A Microbial Consortium in Presence of a Rhamnolipid Produced by Pseudomonas Aeruginosa AT10

The biodegradation of oil products in the environment is often limited by their low water solubility and dissolution rate. Rhamnolipids produced by Pseudomonas aeruginosa AT10 were investigated for their potential to enhance bioavailability and hence the biodegradation of crude oil by a microbial consortium in liquid medium. The characterization of the rhamnolipids produced by strain AT10 showed the effectiveness of emulsification of complex mixtures. The addition of rhamnolipids accelerates the biodegradation of total petroleum hydrocarbons from 32% to 61% at 10 days of incubation. Nevertheless, the enhancement of biosurfactant addition was more noticeable in the case of the group of isoprenoids from the aliphatic fraction and the alkylated polycyclic aromatic hydrocarbons (PHAS) from the aromatic fraction. The biodegradation of some targeted isoprenoids increased from 16% to 70% and for some alkylated PAHs from 9% to 44%.     

A Perspective on the Toxicity of Petrogenic PAHs to Developing Fish Embryos Related to Environmental Chemistry

Numerous studies demonstrate polynuclear aromatic hydrocarbons (PAHs) dissolved from weathered crude oil adversely affect fish embryos at 0.5 to 23 μg/l. This conclusion has been challenged by studies that claim (1) much lower toxicity of weathered aqueous PAHs; (2) direct contact with dispersed oil droplets plays a significant role or is required for toxicity; (3) that uncontrolled factors (oxygen, ammonia, and sulfides) contribute substantively to toxicity; (4) polar compounds produced by microbial metabolism are the major cause of observed toxicity; and (5) that based on equilibrium models and toxic potential, water contaminated with weathered oil cannot be more toxic per unit mass than effluent contaminated with fresh oil. In contrast, several studies demonstrate high toxicity of weathered oil; shifts in PAH composition were consistent with dissolution (not particle ablation), embryos accumulated dissolved PAHs at low concentrations and were damaged, and assumed confounding factors were inconsequential. Consistent with previous empirical observations of mortality and weathering, temporal shifts in PAH composition (oil weathering) indicate that PAHs dissolved in water should (and do) become more toxic per unit mass with weathering because high molecular weight PAHs are more persistent and toxic than the more abundant low molecular weight PAHs in whole oil.     

Identifying polycyclic aromatic hydrocarbon-degrading bacteria in oil-contaminated surface waters at Deepwater Horizon by cultivation, stable isotope probing and pyrosequencing

Polycyclic aromatic hydrocarbons (PAHs) are an important class of chemical pollutants that constitute a major component of total hydrocarbons in crude oils. Based on their poor water solubility, toxicity, persistence and potential to bioaccumulate, these compounds are recognized as high-priority pollutants in the environment and are of significant concern for human health. At oil-contaminated sites, PAH-degrading bacteria perform a critical role in the degradation and ultimate removal of these compounds. In April 2010, enormous quantities of PAHs entered the Gulf of Mexico from the thousands of tons of oil that were released from the ill-fated drilling rig Deepwater Horizon. In the ensuing months after the spill, intense research efforts were devoted to characterizing the microorganisms responsible for degrading the oil, particularly in deep waters where a large oil plume, enriched with aliphatic and low molecular-weight aromatic hydrocarbons, was found in the range of 1,000–1,300 m. PAHs, however, were found mainly confined to surface waters. This paper discusses efforts utilizing DNA-based stable isotope probing, cultivation-based techniques and metagenomics to characterize the bacterial guild associated with PAH degradation in oil-contaminated surface waters at Deepwater Horizon.     

Genome Sequence of Pseudomonas aeruginosa DQ8, an Efficient Degrader of n-Alkanes and Polycyclic Aromatic Hydrocarbons

Pseudomonas aeruginosa DQ8, which was isolated from the crude oil polluted soil in the Daqing oilfield of China, can efficiently degrade diesel, crude oil, n-alkanes, and polycyclic aromatic hydrocarbons (PAHs). Here, we present a 6.8-Mb assembly of its genome sequence. We have annotated 23 coding sequences (CDSs) responsible for catabolism of n-alkanes and PAHs.     

Compositional changes during landfarming of weathered michigan crude oil‐contaminated soii

Laboratory landfarming experiments were conducted to study the bioremediation potential of weathered Michigan crude oil‐contaminated soils. It was found that landfarming was successful in removing up to 90% of the total petroleum hydrocarbons (TPH) in the soil within 22 weeks of treatment. Boiling point analyses of untreated and treated soils indicate a significant removal of TPH compounds independent of molecular weight or carbon number. Up to 85% of heavy petroleum hydrocarbons with carbon numbers above 44 were biode‐graded. In addition, approximately 93% of saturated and 79% of aromatic compounds of the TPH were biodegraded during the 22 week treatment period. The use of polyethylene sheeting as a landfarm cover does not appear to adversely affect biodegradation kinetics under laboratory conditions. Finally, equilibrium leachate concentrations for BTEX and regulated (in Michigan) polynuclear aromatics (PNAs) were below the respective detection limits for each compound. It can be concluded that landfarming of these weathered soils will be highly successful in removing petroleum hydrocarbons while not adversely impacting either ground‐water or surface water quality.     

Polycyclic Aromatic Hydrocarbons in Sediments: An Overview of Risk-Related Issues

This series of articles address site-specific issues associated with evaluating exposure and toxicity of polycyclic aromatic hydrocarbons (PAHs) in sediments. These include factors that influence the opportunity to come into contact with PAHs in sediments (bioaccessibility) and those that affect the potential for transfer from sediment to ecological and human receptors (bioavailability). Although organic carbon is viewed as an important matrix for sorbing PAHs, studies have shown that there are various forms of carbon, some of which are highly sorptive. These latter forms vary on a site-specific basis and including this form of carbon in the assessment can reduce the uncertainty associated with estimating exposure. Other site-specific factors associated with water clarity, depth, and light penetration can result in enhanced toxicity of PAHs as a result of photoactivation. Chemical analyses of sediments increasingly include the alkylated PAH compounds. Although analyses of this suite of PAH compounds is being conducted to support ecological assessments and forensic analyses, there is little guidance on how to interpret the alkylated PAHs with respect to human health risks. An approach for accomplishing this is suggested.     

Biodegradation of a crude oil by three microbial consortia of different origins and metabolic capabilities

Microbial consortia were obtained three by sequential enrichment using different oil products. Consortium F1AA was obtained on a heavily saturated fraction of a degraded crude oil; consortium TD, by enrichment on diesel and consortium AM, on a mixture of five polycyclic aromatic hydrocarbons [PAHs]. The three consortia were incubated with a crude oil in order to elucidate their metabolic capabilities and to investigate possible differences in the biodegradation of these complex hydrocarbon mixtures in relation to their origin. The efficiency of the three consortia in removing the saturated fraction was 60% (F1AA), 48% (TD) and 34% (AM), depending on the carbon sources used in the enrichment procedures. Consortia F1AA and TD removed 100% of n-alkanes and branched alkanes, whereas with consortium AM, 91% of branched alkanes remained. Efficiency on the polyaromatic fraction was 19% (AM), 11% (TD) and 7% (F1AA). The increase in aromaticity of the polyaromatic fraction during degradation of the crude oil by consortium F1AA suggested that this consortium metabolized the aromatic compounds primarily by oxidation of the alkylic chains. The 500-fold amplification of the inocula from the consortia by subculturing in rich media, necessary for use of the consortia in bioremediation experiments, showed no significant decrease in their degradation capability. Journal of Industrial Microbiology & Biotechnology (2002) 28, 252–260 DOI: 10.1038/sj/jim/7000236 Received 12 July 2001/ Accepted in revised form 11 November 2001     

Bacteria belonging to the genus cycloclasticus play a primary role in the degradation of aromatic hydrocarbons released in a marine environment

To identify the bacteria that play a major role in the aerobic degradation of petroleum polynuclear aromatic hydrocarbons (PAHs) in a marine environment, bacteria were enriched from seawater by using 2-methylnaphthalene, phenanthrene, or anthracene as a carbon and energy source. We found that members of the genus Cycloclasticus became predominant in the enrichment cultures. The Cycloclasticus strains isolated in this study could grow on crude oil and degraded PAH components of crude oil, including unsubstituted and substituted naphthalenes, dibenzothiophenes, phenanthrenes, and fluorenes. To deduce the role of Cycloclasticus strains in a coastal zone oil spill, propagation of this bacterial group on oil-coated grains of gravel immersed in seawater was investigated in beach-simulating tanks that were 1 m wide by 1.5 m long by 1 m high. The tanks were two-thirds filled with gravel, and seawater was continuously introduced into the tanks; the water level was varied between 30 cm above and 30 cm below the surface of the gravel layer to simulate a 12-h tidal cycle. The number of Cycloclasticus cells associated with the grains was on the order of 10(3) cells/g of grains before crude oil was added to the tanks and increased to 3 x 10(6) cells/g of grains after crude oil was added. The number increased further after 14 days to 10(8) cells/g of grains when nitrogen and phosphorus fertilizers were added, while the number remained 3 x 10(6) cells/g of grains when no fertilizers were added. PAH degradation proceeded parallel with the growth of Cycloclasticus cells on the surfaces of the oil-polluted grains of gravel. These observations suggest that bacteria belonging to the genus Cycloclasticus play an important role in the degradation of petroleum PAHs in a marine environment.     

Varietal Differences in Rubidium Uptake Efficiency of Barley Roots

The rate of light saturated photosynthesis of Nitzschia palea was reduced by crude oil, naphthalene and benzene. A decrease in the rate of photosynthesis at weak irradiance was also found with crude oil and naphthalene and high concentrations of benzene. On a mg/1 basis naphthalene decreased photosynthesis to a greater extent than did crude oil and crude oil to a greater extent than did benzene. A linear proportionality was found between the decrease in light saturated photosynthesis and the concentrations of aromatic hydrocarbons. The effects on photosynthesis were generally reversible, but a concentration of 700 mg benzene/1 stopped photosynthesis completely and irreversibly.     

Response of sulfate-reducing bacteria to an artificial oil-spill in a coastal marine sediment

In situ mesocosm experiments using a calcareous sand flat from a coastal area of the island of Mallorca in the Mediterranean Sea were performed in order to study the response of sulfate-reducing bacteria (SRB) to controlled crude oil contamination, or heavy contamination with naphthalene. Changes in the microbial community caused by the contamination were monitored by a combination of comparative sequence analysis of 16S rRNA genes, fluorescence in situ hybridization, cultivation approaches and metabolic activity rates. Our results showed that crude oil and naphthalene negatively influenced the total microbial community as the natural increase in cell numbers due to the seasonal dynamics was attenuated. However, both contaminants enhanced the sulfate reduction rates, as well as the culturability of SRB. Our results suggested the presence of autochthonous deltaproteobacterial SRBs that were able to degrade crude oil or polycyclic aromatic hydrocarbons such as naphthalene in anaerobic sediment layers.     

Microbial degradation of crude oil in sea water in continuous culture

Summary The degradation of crude oil in continuous culture of a mixed bacteria population has been studied. The degradation percentage reaches 83 % with a 0.05 h^-1 dilution rate and a 6 g 1^-1 crude oil concentration. The different crude oil compounds : saturated, aromatic, polar hydrocarbons and asphaltenes are degraded at 97 %, 81 %, 52 % and 74 % respectively.     

The reduction of wax precipitation in waxy crude oils by <Emphasis Type="Italic">Pseudomonas</Emphasis> species

Crude oil with different concentrations was subjected to Pseudomonas species at 37 degrees C and various incubation periods. The results showed that Pseudomonas species grew faster at 1% (v/v) concentration of crude oil and exhibited high biodegradation ability within 1 week. On measuring the emulsification activity and emulsion stability during different stages of growth, in various immiscible hydrocarbons, it appeared that the species was able to produce a stable emulsion with a maximum at the end of stationary phase of growth. The gas chromatography analysis of the saturated hydrocarbons of crude oil showed that, an increase in concentration of iso-alkanes in the range of C15-C20, and a bioconversion of heavy iso-alkanes in the range of C21-C22+. Chemical analysis of crude oil by liquid chromatographic technique before and after growth showed that, the saturated alkanes were more degradable than aromatic and asphaltenic compounds. Treatment by Pseudomonas species may possibly be an effective method for the biodegradation of heavy paraffinic hydrocarbon leading to an enhancement in crude oil properties.     

Effects of carbon substrate enrichment and DOC concentration on biodegradation of PAHs in soil

AIMS: Two common reasons to explain slow environmental biodegradation of polycyclic aromatic hydrocarbons (PAHs), namely lack of appropriate carbon sources for microbial growth and limited bioavailability of PAHs, were tested in a laboratory bioassay using a creosote-contaminated soil. METHODS AND RESULTS: The soil, containing a total of 8 mg g-1 of 16 PAHs, was sieved and incubated in bottles for 45 days. The first explanation was tested by enrichment with the analogue anthracene and the non-analogue myristic acid, and both failed to stimulate degradation of all PAHs except anthracene. The second explanation was tested by addition of different concentrations of dissolved organic carbon (DOC), with effects depending on the DOC concentration and the molecular size of the PAH. The degradation was enhanced from 10 to 35% for 12 PAHs when the soil was saturated. The degraded amounts of individual PAHs were proportional to their concentration in the soil. CONCLUSIONS: The slow in situ degradation of PAHs was enhanced by more than three times by adding water as a solvent. Addition of DOC facilitated the degradation of four- to six-ring PAHs. SIGNIFICANCE AND IMPACT OF STUDY: Bioremediation of PAH-contaminated sites may be facilitated by creating water-saturated conditions but retarded by addition of other carbon substrates, such as analogue compounds.     

Accumulation of Hydrocarbons by Maize (Zea mays L.) in Remediation of Soils Contaminated with Crude Oil

This study has investigated the use of screened maize for remediation of soil contaminated with crude oil. Pots experiment was carried out for 60 days by transplanting maize seedlings into spiked soils. The results showed that certain amount of crude oil in soil (≤2 147 mg·kg^?1) could enhance the production of shoot biomass of maize. Higher concentration (6 373 mg·kg^?1) did not significantly inhibit the growth of plant maize (including shoot and root). Analysis of plant shoot by GC-MS showed that low molecular weight polycyclic aromatic hydrocarbons (PAHs) were detected in maize tissues, but PAHs concentration in the plant did not increase with higher concentration of crude oil in soil. The reduction of total petroleum hydrocarbon in planted soil was up to 52.21–72.84%, while that of the corresponding controls was only 25.85–34.22% in two months. In addition, data from physiological and biochemical indexes demonstrated a favorable adaptability of maize to crude oil pollution stress. This study suggested that the use of maize (Zea mays L.) was a good choice for remediation of soil contaminated with petroleum within a certain range of concentrations.     

Effect of modified Fenton's reaction on microbial activity and removal of PAHs in creosote oil contaminated soil

This study describes the removal of polycyclic aromatic hydrocarbons (PAHs) from creosote oil contaminated soil by modified Fenton's reaction in laboratory-scale column experiments and subsequent aerobic biodegradation of PAHs by indigenous bacteria during incubation of the soil. The effect of hydrogen peroxide addition for 4 and 10 days and saturation of soil with H(2)O(2) on was studied. In both experiments the H(2)O(2) dosage was 0.4 g H(2)O(2)/g soil. In completely H(2)O(2)-saturated soil the removal of PAHs (44% within 4 days) by modified Fenton reaction was uniform over the entire soil column. In non-uniformly saturated soil, PAH removal was higher in completely saturated soil (52% in 10 days) compared to partially saturated soil, with only 25% in 10 days. The effect of the modified Fenton's reaction on the microbial activity in the soil was assessed based on toxicity tests towards Vibrio fischeri, enumeration of viable and dead cells, microbial extracellular enzyme activity, and oxygen consumption and carbon dioxide production during soil incubation. During the laboratory-scale column experiments, the toxicity of column leachate towards Vibrio fischeri increased as a result of the modified Fenton's reaction. The activities of the microbial extracellular enzymes acetate- and acidic phosphomono-esterase were lower in the incubated modified Fenton's treated soil compared to extracellular enzyme activities in untreated soil. Abundance of viable cells was lower in incubated modified Fenton treated soil than in untreated soil. Incubation of soil in serum bottles at 20 degrees C resulted in consumption of oxygen and formation of carbon dioxide, indicating aerobic biodegradation of organic compounds. In untreated soil 20-30% of the PAHs were biodegraded during 2 months of incubation. Incubation of chemically treated soil slightly increased PAH-removal compared to PAH-removal in untreated soil.     

Emulsifier of Arthrobacter RAG-1: specificity of hydrocarbon substrate.

The purified extracellular emulsifying factor produced by Arthrobacter RAG-1 (EF-RAG) emulsified light petroleum oil, diesel oil, and a variety of crude oils and gas oils. Although kerosine and gasoline were emulsified poorly by EF-RAG, they were converted into good substrates for emulsification by addition of aromatic compounds, such as 2-methylnaphthalene. Neither aromatic nor aliphatic fractions of crude oil were emulsified by EF-RAG; however, mixtures containing both fractions were emulsified. Pure aliphatic or aromatic hydrocarbons were emulsified poorly by EF-RAG. Binary mixtures containing an aliphatic and an aromatic hydrocarbon, however, were excellent substrates for EF-RAG-induced emulsification. Of a variety of alkylcyclohexane and alkylbenzene derivatives tested, only hexyl- or heptylbenzene and octyl- or decylcyclohexane were effectively emulsified by EF-RAG. These data indicate that for EF-RAG to induce emulsification of hydrocarbons in water, the hydrocarbon substrate must contain both aliphatic and cyclic components. With binary mixtures of methylnaphthalene and hexadecane, maximum emulsion was obtained with 25% hexadecane.