Chemotaxis Toward Crude Oil by an Oil-Degrading Pseudomonas aeruginosa 6-1B Strain

The chemotactic properties of an oil-degrading Pseudomonas aeruginosa strain 6-1B, isolated from Daqing Oilfield, China, have been investigated. The strain 6-1B could grow well in crude oil with a specific rhamnolipid biosurfactant production. Furthermore, it exhibits chemotaxis toward various substrates, including glycine, glycerol, glucose, and sucrose. Compared with another oil-degrading strain, T7-2, the strain 6-1B presented a better chemotactic response towards crude oil and its vital component, n-alkenes. Based on the observed distribution of the strain 6-1B cells around the oil droplet in the chemotactic assays, the potential chemotaxis process of bacteria toward crude oil could be summarized in the following steps: searching, moving and consuming.     

Yield production,chemical composition,and functional properties of emulsifier H28 synthesized by <Emphasis Type="Italic">Halomonas eurihalina </Emphasis>strain H-28 in media containing various hydrocarbons

Halomonas eurihalina strain H-28 is a moderately halophilic bacterium that produces an extracellular polysaccharide not only in media with glucose but also in media supplemented with hydrocarbons (n-tetradecane, n-hexadecane, n-octane, xylene, mineral light oil, mineral heavy oil, petrol, or crude oil). In this study we investigated yield production, chemical composition, viscosity, and emulsifying activity of exopolysaccharides (EPS) extracted from the different media used. The largest amounts of biopolymer were synthesized in media with glucose and n-hexadecane. Chemical composition varied with culture conditions; thus EPS from cultures grown in the presence of hydrocarbons had lower contents of carbohydrates and proteins than EPS from media with glucose. However, the percentages of uronic acids, acetyls, and sulfates were always higher than glucose EPS. Crude oil was the substrate most effectively emulsified. All EPS were capable of emulsifying crude oil more efficiently than the three control surfactants tested (Tween 20, Tween 80, and Triton X-100). All polymers gave low viscosity solutions. EPS H28 could be attractive for application in the oil industry and/or in bioremediation processes, bearing in mind not only its functional properties, but also the capacity of producer strain H-28 to grow in the presence of high salt concentrations and oil substrates.     

Sphingobacterium bambusaue及其紫外诱变菌株的石油降解功能

【目的】研究Sphingobacterium bambusaue及其紫外诱变菌株的石油降解功能。【方法】紫外诱变后筛选石油降解高效菌株; 以不同石油浓度、pH值及盐浓度优化培养条件, 用重量法检测高效菌株石油降解率。【结果】发现菌株S. bambusaue在石油降解培养基中培养5 d的石油降解率为25.86%, UV诱变高效菌株IBFC2009-S3培养5 d的石油降解为42.85%, 比始发菌株提高65.7%; UV诱变菌株IBFC2009-S3的优化培养条件为石油浓度0.5 g/L、pH值7.0以及NaCl质量浓度为10 g/L, 其石油降解率可达50.51%。【结论】首次报道S. bambusaue具有石油降解功能; 紫外诱变获得的菌株S3的石油降解能力较强。     

Naphthalene degradation and biosurfactant activity by Bacillus cereus 28BN

Biosurfactant activity and naphthalene degradation by a new strain identified as Bacillus cereus 28BN were studied. The strain grew well and produced effective biosurfactants in the presence of n-alkanes, naphthalene, crude oil and vegetable oils. The biosurfactants were detected by the surface tension lowering of the medium, thin layer chromatography and infrared spectra analysis. With (2%) naphthalene as the sole carbon source, high levels of rhamnolipids at a concentration of 2.3 g 1(-1) were determined in the stationary growth. After 20 d of incubation 72 +/- 4% of the initial naphthalene was degraded. This is the first report for a Bacillus cereus rhamnolipid producing strain that utilized naphthalene under aerobic conditions. The strain looks promising for application in environmental technologies.     

石油降解细菌的表型特性和系统发育分析

从3种不同土壤中分离和纯化得到10个石油降解细菌菌株,分离菌株均为好氧菌、革兰氏阴性菌、不形成芽孢的杆菌,10个菌株均能利用中等链长的烷烃、柴油和原油作为碳源,而不能以单环芳烃和多环芳烃为碳源。根据其生理生化性状和16SrDNA序列分析结果表明,分离菌株EVA5,EVA6,EVA7,EVA8和EVA9为假单胞菌（Pseudomonas spp.）,EVA10、EVA11、EVA12、EVA13和EVA14为不动杆菌（Acinetobacter spp.）,均属于Proteobac-teria的γ亚群。     

海绵中石油降解微生物的分离筛选及降解特性研究

从大连湾原油污染海域生长的海绵中分离筛选到一株原油降解菌OA58,根据其生长形态、培养特征、16S rRNA序列相似性比对分析和生化指标检测,初步鉴定为脱叶链霉菌（Streptomyces exfoliatus）。同时,考察了该链霉菌对原油的降解效果,OA58能以原油为唯一碳源生长,在人工海水培养基中,14 d内对原油（初始浓度为1 g/L）的平均降解率为83%,是一株具有开发潜力的原油降解放线菌。     

Genome analysis provides insights into crude oil degradation and biosurfactant production by extremely halotolerant Halomonas desertis G11 isolated from Chott El-Djerid salt-lake in Tunisian desert

Here, we report the genomic features and the bioremediation potential of Halomonas desertis G11, a new halophilic species which uses crude oil as a carbon and energy source and displays intrinsic resistance to salt stress conditions (optimum growth at 10% NaCl). G11 genome (3.96 Mb) had a mean GC content of 57.82%, 3622 coding sequences, 480 subsystems and 64 RNA genes. Annotation predicted 38 genes involved in osmotic stress including the biosynthesis of osmoprotectants glycine-betaine, ectoine and osmoregulated periplasmic glucans. Genome analysis revealed also the versatility of the strain for emulsifying crude oil and metabolizing hydrocarbons. The ability of G11 to degrade crude oil components and to secrete a glycolipid biosurfactant with satisfying properties was experimentally confirmed and validated. Our results help to explain the exceptional capacity of G11 to survive at extreme desertic conditions, and highlight the metabolic features of this organism that has biotechnological and ecological potentialities.     

Screening and Evaluation of Biosurfactant-Producing Strains Isolated from Oilfield Wastewater

The six biosurfactant-producing strains, isolated from oilfield wastewater in Daqing oilfield, were screened. The production of biosurfactant was verified by measuring the diameter of the oil spreading, measuring the surface tension value and emulsifying capacity against xylene, n-pentane, kerosene and crude oil. The experimental result showed three strains (S2, S3, S6) had the better surface activity. Among the three strains, the best results were achieved when using S2 strain. The diameter of the oil spreading of the biosurfactant produced by S2 strain was 14 cm, its critical micelle concentration (CMC) was 21.8 mg/l and the interfacial tension between crude oil and biosurfactant solution produced by S2 strain reduced to 25.7 mN/m. The biosurfactant produced by S2 strain was capable of forming stable emulsions with various hydrocarbons, such as xylene, n-pentane, kerosene and crude oil. After S2 strain treatment, the reduction rate of oil viscosity was 51 % and oil freezing point reduced by 4 °C.     

Biosurfactant production by Mucor circinelloides: Environmental applications and surface‐active properties

Biosurfactants are structurally a diverse group of surface‐active molecules widely used for various purposes in industry. In this study, among 120 fungal isolates, M‐06 was selected as a superior biosurfactant producer, based on different standard methods, and was identified as Mucor circinelloides on the basis of its nucleotide sequence of the internal transcribed spacer (ITS) gene. M. circinelloides reduced the surface tension to 26 mN/m and its EI₂₄ index was determined to be 66.6%. The produced biosurfactant exhibited a high degree of stability at a high temperature (121°C), salinity (40 g/L), and acidic pH (2–8). The fermentation broth's ability to recover oil from contaminated sand was 2 and 1.8 times higher than those of water and Tween 80, respectively. The ability of biosurfactant to emulsify crude oil in the sea and fresh water was 64.9 and 48% respectively. This strain could remove 87.6% of crude oil in the Minimal Salt Medium (MSM) crude oil as the sole carbon source. The results from a primary chemical characterization of crude biosurfactant suggest that it is of a glycolipid nature. The strain and its biosurfactant could be used as a potent candidate in bioremediation of oil‐contaminated water, soil, and for oil recovery processes.     

Characteristics of bioemulsifier V2-7 synthesized in culture media added of hydrocarbons: chemical composition, emulsifying activity and rheological properties

The bioemulsifier V2-7 is an exopolysaccharide (EPS) synthesized by strain F2-7 of Halomonas eurihalina and it has the property of emulsifying a wide range of hydrocarbons i.e. n-tetradecane, n-hexadecane, n-octane, xylene mineral light and heavy oils, petrol and crude oil. Characteristics of exopolysaccharide V2-7 produced in media supplemented with various hydrocarbons (n-tetradecane, n-hexadecane, n-octane, xylene, mineral light oil, mineral heavy oil, petrol or crude oil) were studied. Yield production varied from 0.54 to 1.45 g L(-1) according to the hydrocarbon added, in the same way chemical composition, viscosity and emulsifying activity of EPS varied with the culture conditions. Respect to chemical composition, percentage of uronic acids found in exopolymers produced in hydrocarbon media was always higher than that described for V2-7 EPS (1.32%) obtained with glucose. This large amount of uronic acid present could be useful in biodetoxification and waste water treatment. On the other hand, the highest amount of biopolymer was synthesized with mineral light oil, while the most active emulsifiers were those obtained from media added with petrol and n-octane. Furthermore, all EPS were capable of emulsifying crude oil more efficiently than the three chemical surfactants tested as control (Tween 20, Tween 80 and Triton X-100). The capacity of strain F2-7 to grow and produce bioemulsifier in presence of oil hydrocarbons together with the high emulsifying activity and low viscosity power of the biopolymers synthesized in hydrocarbons media could be considered highly beneficial for application of both bioemulsifier and producing strain in bioremediation of oil pollutants.     

一株可乳化降解原油的Compostibacillus的分离及特性

【背景】通过实施多轮次微生物采油,华北油藏产出液菌浓达到了106个/mL以上,油藏内部已经形成了较稳定的微生物发酵场,从其中筛选出能够乳化降解原油的微生物,并在地面对其进行扩大培养,然后再应用到微驱油藏,以进一步提高微生物采油实施效果。【目的】筛选乳化降解原油性能良好的菌株,对其进行多相分类学鉴定和性能评价。【方法】利用原油为底物筛选乳化降解性能良好的菌株,通过形态特征观察、生理生化测定、16S rRNA基因序列分析等确定菌株的分类地位。通过乳化能力、降解率等方法确定菌株的原油乳化降解特性。【结果】从华北油田采集的地层水样品中分离得到一株乳化原油的菌株BLG74,经多相分类鉴定表明其是土壤堆肥芽孢杆菌(Compostibacillus humi)的新菌株,亲源性99.6%。该菌株的生长温度为30-60℃ (最适温度45℃),pH6.5-9.5(最适pH7.0),NaCl浓度0%-7%(质量体积比)。菌株BLG74在玉米浆培养基中培养,其发酵液的表面张力为56.3 mN/m,乳化力约95%,在初始原油质量浓度0.5%、温度45℃的条件下培养20d,对原油的降解率可达40.8%。【结论】菌株BLG74是可乳化降解原油的新成员,其在热盐条件下乳化降解原油的特性在石油开采中有一定的潜力。     

Petroleum Degradation,Biosurfactant and Laccase Production by Fusarium neocosmosporiellum RH-10: A Microcosm Study

The aim of this study was to evaluate the potential of crude oil removal by fungal strains isolated from Arak refinery. The results showed that the RH10 strain is a potent strain as a surfactant producer and degrader of petrochemical hydrocarbons. The strain was identified as a Fusarium neocosmosporiellum and could degrade 58% of hydrocarbons in the minimal medium and reduce the surface tension from 45 to 26.5 mN m^?1. Moreover, residual crude oil analysis with Fourier transform infrared spectrophotometry showed that this strain was able to degrade 50% of aliphatic compounds. To investigate the mechanism of degradation, oxidase enzymes were assayed and it was found that F. neocosmosporiellum can produce 1.94 U L^?1 of laccase in 10 g L^?1 crude oil. Carbon, nitrogen, phosphorus and soil pattern optimization in a microcosm study showed that this strain removed 44% and 27% of the crude oil from contaminated soil in 1% and 5% crude oil concentrations, respectively. Under optimum condition, 9.66 g kg^?1 crude oil was removed by F. neocosmosporiellum when the initial oil concentration was 50 g kg^?1, at the end of 150 days microcosm experiment. The results demonstrated the promising potential of fungi strain for cleaning of contaminated soil.     

Potential of <Emphasis Type="Italic">Burkholderia cepacia</Emphasis> RQ1 in the biodegradation of heavy crude oil

The potential of Burkholderia cepacia strain RQ1 in the biodegradation of heavy crude oil (Maya) was assessed to develop an active indigenous bacterial consortium for the bioremediation of crude oil-polluted systems in Nigeria. The heavy crude oil (Maya) was utilized as sole source of carbon, attaining maximum cell densities of 10(8) cfu ml(-1) from an initial 10(5) cfu ml(-1) in 15 days. Biomass also increased with oil concentrations up to 0.8% (w/v). Growth rates ranged from 0.028 h(-1) to 0.036 h(-1) and degradation rates decreased with increasing concentrations of oil from 0.009 day(-1) to 0.004 day(-1). The quantity of oil metabolized increased significantly (P < 0.05) with increasing concentrations of oil. However, the growth of the bacterium was inhibited at crude oil concentrations beyond 6% (w/v). The pH of the culture media also dropped significantly (P < 0.05) during the 15-day test period, while the non-asphaltic fractions of the oil were significantly reduced (by about 89%) during the same period. The bacterium harbours a plasmid of about 10 kb that lacks restriction sites for the endonucleases Asp718, BamHI and PstI.     

Isolation and characterization of a sulfur-oxidizing chemolithotroph growing on crude oil under anaerobic conditions

Molecular approaches have shown that a group of bacteria (called cluster 1 bacteria) affiliated with the epsilon subclass of the class Proteobacteria constituted major populations in underground crude-oil storage cavities. In order to unveil their physiology and ecological niche, this study isolated bacterial strains (exemplified by strain YK-1) affiliated with the cluster 1 bacteria from an oil storage cavity at Kuji in Iwate, Japan. 16S rRNA gene sequence analysis indicated that its closest relative was Thiomicrospira denitrificans (90% identity). Growth experiments under anaerobic conditions showed that strain YK-1 was a sulfur-oxidizing obligate chemolithotroph utilizing sulfide, elemental sulfur, thiosulfate, and hydrogen as electron donors and nitrate as an electron acceptor. Oxygen also supported its growth only under microaerobic conditions. Strain YK-1 could not grow on nitrite, and nitrite was the final product of nitrate reduction. Neither sugars, organic acids (including acetate), nor hydrocarbons could serve as carbon and energy sources. A typical stoichiometry of its energy metabolism followed an equation: S(2-) + 4NO(3)(-) --> SO(4)(2-) + 4NO(2)(-) (Delta G(0) = -534 kJ mol(-1)). In a difference from other anaerobic sulfur-oxidizing bacteria, this bacterium was sensitive to NaCl; growth in medium containing more than 1% NaCl was negligible. When YK-1 was grown anaerobically in a sulfur-depleted inorganic medium overlaid with crude oil, sulfate was produced, corresponding to its growth. On the contrary, YK-1 could not utilize crude oil as a carbon source. These results suggest that the cluster 1 bacteria yielded energy for growth in oil storage cavities by oxidizing petroleum sulfur compounds. Based on its physiology, ecological interactions with other members of the groundwater community are discussed.     

Genome Sequence of Pseudomonas aeruginosa Strain SJTD-1, a Bacterium Capable of Degrading Long-Chain Alkanes and Crude Oil

Pseudomonas aeruginosa strain SJTD-1 can utilize long-chain alkanes, diesel oil, and crude oil as sole carbon sources. We report the draft genome sequence of strain SJTD-1 (6,074,058 bp, with a GC content of 66.83%) and major findings from its annotation, which could provide insights into its petroleum biodegradation mechanism.     

Characteristics of bioemulsifiers synthesised in crude oil media by <Emphasis Type="Italic">Halomonas eurihalina</Emphasis> and their effectiveness in the isolation of bacteria able to grow in the presence of hydrocarbons

Halomonas eurihalina strains F2-7, H28, H96, H212 and H214 were capable of producing large amounts of exopolysaccharides (EPS) in MY medium with added crude oil. The biopolymers showed lower carbohydrate and protein content than those synthesised in control medium without oil. Nevertheless, the percentages of uronic acids, acetyls and sulphates were higher. The emulsifying activity of biopolymers was measured; crude oil was the substrate most efficiently emulsified. Furthermore, all the EPS tested emulsified higher volumes of crude oil than the commercial surfactants used as controls. We have also proved the effectiveness of both Halomonas eurihalina strains and their EPS to select indigenous bacteria able to grow in the presence of polycyclic aromatic hydrocarbons (naphthalene, phenanthrene and pyrene) from waste crude oil. The majority of isolated strains belonged to the genus Bacillus.     

Investigation of alkane biodegradation using the microtiter plate method and correlation between biofilm formation, biosurfactant production and crude oil biodegradation

Among 25 crude oil-degrading bacteria isolated from a marine environment, four strains, which grew well on crude oil, were selected for more study. All the four isolated had maximum growth on 2.5% of crude oil and strain BC (Pseudomonas) could remove crude oil by 83%. The drop collapse method and microtiter assay show that this strain produces more biosurfactant, and its biofilm formation is higher compared to other strains. Bacterial adhesions to crude oil for strains CS-2 (Pseudomonas), BC, PG-5 (Rhodococcus) and H (Bacillus) were 30%, 46%, 10% and 1%, respectively. Therefore, strain H with a low production of biosurfactant and biofilm formation had showed the least growth on these compounds. PCR analysis of these four strains showed that all isolates had alk-B genes from group (III) alkane hydroxylase. All isolate strains could utilize cyclohexan, octane, hexadecane, octadecan and diesel fuel oil; however, the microtiter plate assay showed that strain BC had more growth, respiration and biofilm formation on octadecan.     

Biotechnological Potential of Bacillus salmalaya 139SI: A Novel Strain for Remediating Water Polluted with Crude Oil Waste

Environmental contamination by petroleum hydrocarbons, mainly crude oil waste from refineries, is becoming prevalent worldwide. This study investigates the bioremediation of water contaminated with crude oil waste. Bacillus salamalaya 139SI, a bacterium isolated from a private farm soil in the Kuala Selangor in Malaysia, was found to be a potential degrader of crude oil waste. When a microbial population of 10⁸ CFU ml^-1 was used, the 139SI strain degraded 79% and 88% of the total petroleum hydrocarbons after 42 days of incubation in mineral salt media containing 2% and 1% of crude oil waste, respectively, under optimum conditions. In the uninoculated medium containing 1% crude oil waste, 6% was degraded. Relative to the control, the degradation was significantly greater when a bacteria count of 99 × 10⁸ CFU ml^-1 was added to the treatments polluted with 1% oil. Thus, this isolated strain is useful for enhancing the biotreatment of oil in wastewater.     

Evaluation of bioemulsifier mediated Microbial Enhanced Oil Recovery using sand pack column

Bacillus licheniformis K125, isolated from an oil reservoir, produces an effective bioemulsifier. The crude bioemulsifier showed 66% emulsification activity (E(24)) and reduced the surface tension of water from 72 to 34 mN/m. It contains substantial amount of polysaccharide, protein and lipid. This bioemulsifier is pseudoplastic non-Newtonian in nature. It forms oil in water emulsion which remains stable at wide range of pH, temperature and salinity. It gave 43+/-3.3% additional oil recovery upon application to a sand pack column designed to simulate an oil reservoir. This is 13.7% higher than that obtained from crude lipopeptide biosurfactants produced by the standard strain, Bacillus mojavensis JF2 and 8.5% higher than hot water spring isolate, Bacillus licheniformis TT42. The increased oil recovery obtained by using the crude bioemulsifier can be attributed to its combined surface and emulsification activity. Its mechanism of oil recovery must be similar to the mechanism exhibited by surfactant-polymer flooding process of chemical enhanced oil recovery.     

Isolation and application of Gordonia sp. JC11 for removal of boat lubricants

Boat lubricants are continuously released into the marine environment and thereby cause chronic oil pollution. This study aims to isolate lubricant-degrading microorganisms from Thai coastal areas as well as to apply a selected strain for removal of boat lubricants. Ten microorganisms in the genera of Gordonia, Microbacterium, Acinetobacter, Pseudomonas, Brucella, Enterococcus and Candida were initially isolated by crude oil enrichment culture techniques. The lubricant-removal activity of these isolates was investigated with mineral-based lubricants that had been manufactured for the 4-stroke diesel engines of fishing boats. Gordonia sp. JC11, the most effective strain was able to degrade 25-55% of 1,000 mg L(-1) total hydrocarbons in six tested lubricants, while only 0-15% of the lubricants was abiotically removed. The bacterium had many characteristics that promoted lubricant degradation such as hydrocarbon utilization ability, emulsification activity and cell surface hydrophobicity. For bioaugmentation treatment of lubricant contaminated seawater, the inoculum of Gordonia sp. JC11 was prepared by immobilizing the bacterium on polyurethane foam (PUF). PUF-immobilized Gordonia sp. JC11 was able to remove 42-56% of 100-1,000 mg L(-1) waste lubricant No. 2 within 5 days. This lubricant removal efficiency was higher than those of free cells and PUF without bacterial cells. The bioaugmentation treatment significantly increased the number of lubricant-degrading microorganisms in the fishery port seawater microcosm and resulted in rapid removal of waste lubricant No. 2.