一株石油烃降解菌的细胞疏水性及其乳化性质

【目的】从新疆油田石油污染土壤中分离到一株在25 °C条件下利用烃类产生生物表面活性剂的菌株红球菌(Rhodococcus sp.) HL-6, 对其菌体细胞疏水性及所产表面活性剂进行研究。【方法】通过细胞粘附性、表面张力及乳化活性测定对菌株所产表面活性剂进行性质研究。【结果】菌株HL-6在亲水性和疏水性基质中均能产生生物表面活性剂, 在疏水性基质中可以将培养液表面张力由初始的62.487 mN/m降到30.667 mN/m, 培养液在pH 6?9及NaCl浓度1%?5%范围内乳化效果良好, 在4 °C到55 °C范围内乳化效果均为100%, 菌株对柴油的耐受能力很高, 在30%柴油浓度下依然生长良好并且有44%的乳化活性。【结论】HL-6菌株的细胞表面具有很强的疏水性, 这有助于菌体细胞对烃类的摄取。该菌株能够利用烃类基质生产生物表面活性剂, 可以明显降低培养液表面张力并且对石油烃具有良好的乳化作用。说明菌株HL-6能够适应海洋滩涂石油污染的环境, 并可用于严重石油污染区域的生物修复。     

一株油藏耐热原油降解菌的分离鉴定与特性分析

从中原油田高温采出液中分离到一株能在55℃高温条件下利用石油烃类生长的耐高温菌株P98-18A1.基于表型、生理生化特性和16S rDNA序列分析,初步鉴定该菌株为芽孢杆菌属(Bacillus sp.).通过菌株对原油及部分烷烃降解试验得出,该菌株在适宜条件下,能有效地利用C6-C16的正构烷烃生长,对链长大于C16的烷烃在某种程度上具有一定的选择性利用能力.     

陕北地区石油污染土壤中不动杆菌属的筛选、鉴定及降解性能

从陕北地区石油污染土壤中分离鉴定得到两株不动杆菌属(Acinetobacter sp.)的高效石油降解菌A.sp 1和A.sp 2,分别从盐浓度、pH值、氮源、磷源和接种量等因素进行研究以确定其最佳石油降解条件,并进一步通过GC-MS(Gas ChromatographyMass Spectrometer)方法分析其在最佳条件下对原油组分的不同降解性能。结果显示:A.sp 1在盐浓度为1%、pH值为6—7、磷源为KH2PO4和K2HPO4、氮源为尿素和接种量为4%的条件下,最高降解率可达到60%。A.sp 2在盐浓度为1%、pH值为7—9、磷源为KH2PO4和K2HPO4、氮源为硝酸铵和接种量为8%的条件下,最高降解率可达到67%。GC-MS分析结果表明,菌株A.sp 1对石油烃类C21—C25有明显的降解效果,菌株A.sp 2对石油烃类C20—C30的降解效果较好。     

鼠李糖脂对微生物降解石油烃废水的影响

目的:研究鼠李糖脂对微生物降解石油烃废水的影响.方法:通过测定生物量和观察菌株表面来研究鼠李糖脂对菌株的影响;通过正交实验设计,确定石油烃降解率影响因素.通过石油烃降解率的测定,探讨鼠李糖脂与H2O2深度氧化协同作用对微生物降解石油烃的影响.结果:菌株对石油烃的降解率达53%,在相同条件下,添加鼠李糖脂的石油烃降解率提高了12%-20%.添加鼠李糖脂后菌株的生物量明显增多,菌株细胞表面疏水.正交设计表明,影响石油烃降解的主导因子是培养温度,其次是培养时间和鼠李糖脂的添加量.正交设计得到最佳组合为A3B2C1,即培养时间为7d;温度为35℃,鼠李糖脂浓度为60mg/L.3个因素的最佳组合下,石油烃降解率为82%.加入200 mg/L的H2O2时,降解率从82%提高到97%.结论:鼠李糖脂能促进菌株的生长.鼠李糖脂与H2O2深度氧化协同作用有助于微生物对石油烃类污染物降解效率的提高.     

一株产甘露糖赤藓糖醇脂菌株的筛选及其产物分析

【目的】解决石油长链烃类物质引起的环境污染问题,筛选可以高效降解石油烃的产糖脂类生物表面活性剂菌株。【方法】采用血平板、油平板法,从葡萄皮表面分离到6株产糖脂类的真菌,比较各菌株的排油性能,通过PCR扩增合成糖脂类表面活性剂的关键基因,筛选到一株具有emtl序列的真菌K6。经形态学、生理生化测定和分子系统发育分析(5.8S,ITS1,ITS2)对菌株进行鉴定,而且通过TLC和HPLC分析该菌株的代谢产物。【结果】经鉴定,该菌为Pseudozyma churashimaensis,可产甘露糖赤藓糖醇脂。石油烃降解实验表明,菌株K6具有很强的乳化性能和降解石油烃的能力,其石油烃降解率可达70.17%。【结论】菌株K6具有产生物表面活性剂和降解长链石油烃类的能力,其对石油污染环境的生物修复具有重要的现实意义。     

一株假单胞菌HZ-1降解石油烃的初步研究

从石油废水活性污泥中分离到一株可以较好降解石油烃的假单胞菌（Pseudomonas sp）HZ-1,采用生物膜法对该菌株在30℃条件下降解某炼油厂废水中的烃类物质进行了研究.研究结果表明不同营养盐NH4NO3、NaH2PO4、NH4Cl对菌株HZ-1处理石油废水的作用不同,终浓度为1000mg/L的NH4NO3对菌株HZ-1处理石油废水效果最佳.并在30℃和pH 8.0的条件下,初步研究了该菌株降解萘的情况,在萘浓度低于78 mg/L的情况下,120 rpm好氧振荡培养144h,这株菌对萘的降解率在86%以上.     

陕北地区提高石油采收率菌株筛选及其降解性能评价

筛选出可提高石油采收率的降解菌株。采用富集培养和稀释平板法,从陕北某采油厂附近长期受原油污染的水域和油污泥中筛选获得以原油为唯一碳源的降油菌株,并对其生长条件、排油圈能力、代谢产物表面活性物质、表面张力与乳化性及原油降解实验进行一系列的研究。含油废水和油污泥中筛选出10株细菌,从中选择了6株优势菌作为供试菌株进行后续研究,6株供试菌株均可使原油的理化性质发生变化。菌株B与原油作用后,使原油黏度降低了49.02%,菌株A-08、菌株5-13、菌株1-2及菌株A和菌株A-08复合菌群可使原油乳化为细小的圆型颗粒,与原油作用后,降解率达到43.25%-60.00%。     

中高温油藏微生物单井激活现场试验

针对胜利油田中高温油藏微生物群落结构特点,开展室内模拟试验,对耐高温激活剂进行筛选和性能评价。室内筛选出的激活剂适应温度范围在60℃以上,该激活体系与稠油作用后乳化分散效果显著,激活后菌密度从1.3×10~5个/mL增至3×10~7~4×10~7个/mL,气压达到0.05 MPa以上,原油乳化指数达到90%以上。根据室内模拟条件,对中高温油藏单井进行现场激活试验,开井后,总菌密度由激活前的10~4~10~5个/mL增至3.2×10~7个/mL,在有效期136 d内,单井累计增油超过180 t,含水率降低约15%。试验证明,室内筛选出的激活剂可以激活中高温油藏內源微生物,提高单井产油量,为实施中高温油藏微生物驱油提供理论依据和技术支持。     

驱油功能微生物的初步筛选

目的对大庆油田本源菌筛选得到高效驱油备用菌株。方法采用血平板实验、排油活性测定、原油凝固点测定、乳化性能测定等方法。结果筛选得到能够利用石油为唯一碳源生长的菌株13株,其中5株菌株分别在不同驱油性能方面有突出作用。结论综合筛选结果得到2株在产表面活性剂、较强的排油性能和乳化性能方面均具有较好效果的菌株。     

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...     

Insight into heterogeneity in cell-surface hydrophobicity and ability to degrade hydrocarbons among cells of two hydrocarbon-degrading bacterial populations

The sequential bacterial adherence to hydrocarbons (BATH) of successive generations of hydrophobic fractions of Paenibacillus sp. R0032A and Burkholderia cepacia gave rise to bacterial populations of increasing cell-surface hydrophobicity. Thus, hydrophobicity of the first generation (H1) was less than that of the second generation (H2), which was less than that of the third generation (H3). Beyond H3, the hydrophobic populations became less stable and tended to lyse in hexadecane after violent (vortex) agitation, resulting in an apparent decline in BATH value. The exhaustively fractionated aqueous-phase population (L) was very hydrophilic. The overall cell-surface distribution of the population was L < parental strain < H1 < H2 < H3. The ability to degrade crude oil, hexadecane, or phenanthrene matched the degree of cell-surface hydrophobicity: L < P < H1 < H2 < H3. Thus, in natural populations of hydrocarbon-degrading Paenibacillus sp. R0032A and B. cepacia, there is a heterogeneity in the hydrophobic surface characteriistics that affects the ability of cells to use various hydrocarbon substrates.     

Modulation of cell surface hydrophobicity in the benthic cyanobacterium Phormidium J-1

A shift from cell-surface hydrophobicity to hydrophilicity was experimentally induced in the benthic hydrophobic cyanobacterium Phormidium sp. strain J-1, by mechanical shearing, chloramphenicol, and proteolytic treatment after preincubation with sodium dodecyl sulfate (SDS). Treatment with SDS alone, while releasing large amounts of protein and carbohydrates from the cell wall, did not affect cell surface hydrophobicity.Ultrastructural analysis showed the cells, to be enveloped by a double-layered minicapsule. Treatments affecting cellsurface hydrophobicity also caused changes in capsular components. A model, describing cell-surface structure, composition and properties in Phormidium J-1, was constructed by correlating ultrastructural data with surface properties.Abbreviations SDS Sodium dodecyl sulfate - DCMU 3(3,4-dichlorophenyl)-1,1-dimethylurea This paper is contributed in honor of Prof. G. Drews on the occasion of his sixtieth birthday     

Cell-surface charge and cell-surface hydrophobicity of collagen-bindingAeromonas andVibrio strains

Partitioning in aqueous polymer two-phase systems of polyethylene glycol and dextran was used to detect and compare cell-surface charge and cell-surface hydrophobicity of Aeromonas hydrophila, A. caviae, A. sobria, Vibrio cholerae, and V. anguillarum strains. These strains have cell-surface components that bound either native or thermally denatured type I collagen (i.e., a mixture of the α1+α2 chains) and gelatin immobilized on latex beads. Our goals were: (1) to compare the possible relationship between the cell-surface charge/hydrophobicity and binding to collagen and (2) to evaluate the influence of the culture media on the expression of surface properties. There was no apparent relationship between cell-surface charge, cell-surface hydrophobicity, and binding to collagen. The expression of surface properties was dependent on the culture media. There was no relationship between binding to immobilized collagen and binding to soluble ¹²⁵I-labeled collagen. Particle-agglutination reactivity differed when using various collagen-coated microbead preparations. There were general differences in the particle-agglutination reactivity when collagen-coated latex beads were prepared using different coating procedures. The negative charge and hydrophobicity of the various collagen-coated microbead preparations were also studied by partitioning in the two-phase system of polyethylene glycol and dextran. Under these conditions, the α1+α2 collagen-chain mixture covalently immobilized on carboxy-modified latex beads was less hydrophobic and negatively charged than gelatin and native collagen immobilized on the same kind of latex beads. For latex beads passively coated with collagen preparations, the α1+α2 collagen-chain mixture was more hydrophobic than gelatin and native collagen. We suggest that for screening collagen-binding among Vibrio and Aeromonas strains, a reliable and sensitive particle-agglutination assay should consider the collagen preparation and the coating procedure for the immobilization of collagen onto the latex beads. In this regard, carboxy-modified latex beads coated with an α1+α2 collagen-chain mixture gave the best results. Received: 9 January 1995 / Accepted: 30 May 1995     

Hydrophobicity development,alkane oxidation,and crude-oil emulsification in a Rhodococcus species

The relationship between the phenomena alkane oxidation, extreme hydrophobicity of the cell surface, and crude-oil emulsification in Rhodococcus sp. strain 094 was investigated. Compounds that induce the emulsifying ability simultaneously induced the cytochrome P450-containing alkane oxidizing system and the transition from low to high cell-surface hydrophobicity. Exposed to inducers of crude-oil emulsification, the cells developed a strong hydrophobic character during exponential growth, which was rapidly lost when entering stationary phase. The loss in hydrophobicity coincided in time with the crude-oil emulsification, indicating that the components responsible for the formation of cell-surface hydrophobicity act as excellent emulsion stabilisers only after release from the cells. Rhodococcus sp. strain 094 possessed three distinct levels of cell-surface hydrophobicity. One level of low hydrophobicity was characteristic of cells in late stationary phase and was independent of growth substrate. A second and more hydrophobic level was observed for cells in exponential phase grown on water-soluble substrates, while a third level, characterised by extreme cell hydrophobicity, was observed for cells in exponential phase cultivated on hydrophobic substrates such as hexadecane. The production of the oil-emulsifying agents seems to require external sources of nitrogen and phosphate.     

Standardization of salt aggregation test for reproducible determination of cell-surface hydrophobicity with special reference to Staphylococcus species

The laboratory conditions for reproducible routine determination of staphylococcal cell-surface hydrophobicity by the salt aggregation test were standardized. Fresh bacterial suspensions standardized to 5 x 10(9) cfu/ml gave the most reproducible results with both Staphylococcus aureus and coagulase-negative staphylococci. For relatively hydrophobic strains a 5-min reading time was necessary to detect bacterial aggregation in ammonium sulphate solutions ranging from 0.1 M to 1.5 M, pH 6.8. A x 10 hand lens facilitated reading aggregations. Overnight storage of bacterial suspensions at 20 degrees C reduced cell-surface hydrophobicity of all species, while storage at 4 degrees C reduced the hydrophobic nature of Staph. aureus strains. The hydrophobicity of coagulase-negative staphylococci rarely changed at 4 degrees C. A 10-fold dilution of fresh, standardized bacterial suspensions made it impossible to detect bacterial aggregation in ammonium sulphate solutions even with a hand lens. Under standardized conditions three types of staphylococcal cell aggregations were observed. The first looked like the slide agglutination for O antigens of Enterobacteriaceae, the second resembled H-agglutination, while the third had a filamentous appearance. These patterns indicated that more than one component might contribute to cell-surface hydrophobicity of both Staph. aureus and coagulase-negative staphylococci, or the same component might have different position on the cell surface.     

Toxin profiles, and cell-surface properties of Salmonella strains isolated from Swedish travelers.

Toxin production, cell-surface hydrophobicity and fibronectin-binding properties of 21 Salmonella strains of different species, isolated from Swedish travelers to different parts of the world, were studied. Cell sonicate supernatants from blood agar grown cultures of 80% of the strains induced rabbit skin permeability reaction in the form of induration and/or blueing while 33% of the strains also produced cell necrotizing factor on rabbit skin. Four strains were negative in the rabbit skin permeability test, while only two were negative when tested on CHO cells. When cultured on blood agar, a majority of the strains (17/21) showed low cell-surface hydrophobicity, showing no aggregation even at 1.5 M ammonium sulfate concentration in salt aggregation test (SAT), while only four strains showed high cell-surface hydrophobicity. Furthermore, these strains could be classified as low fibronectin binders due to their poor interaction with fibronectin or its 29 kDa N-terminal fragment.     

Sequential hydrophobic partitioning of cells of Pseudomonas aeruginosa gives rise to variants of increasing cell-surface hydrophobicity

The partitioning of bacterial cells in a dual aqueous-solvent phase system leads to separation into 'hydrophilic' and hydrophobic functions. Sequential multistep partitioning, accompanied by successive enrichment, gives rise to several cycles of hydrophobic and hydrophilic cell populations which possess different cell-surface hydrophobicity characteristics. Characterization of the cell-surface hydrophobicity by several methods (salting-out aggregation test, bacterial adherence to hydrocarbon, polystyrene binding and hydrophobic interaction chromatography) was carried out. The cell-surface hydrophobicity varied in the order: hydrophilic fraction < parental strain < first cycle hydrophobic variant < second cycle hydrophobic variant < third cycle hydrophobic variant. Electron microscopy showed that the most hydrophobic variant was densely covered by hydrophobic structures - fimbriae - whereas the parental strain was covered by a mixture of surface structures. The hydrophilic variant was covered by an amorphous exopolymeric substance, which is a polysaccharide, shown by its reaction with Alcian blue.     

Characterization of a novel long-chain n-alkane-degrading strain, Dietzia sp. E1

The newly isolated strain E1, identified as a Dietzia sp., proved to have an excellent ability to degrade n-C12 to n-C38 alkane components of crude oil. The preferred substrate was the very long-chain alkane n-eicosane at an optimal temperature of 37 degrees C and an optimal pH of 8 under aerobic conditions. The growth and substrate uptake kinetics were monitored during the n-alkane fermentation process, and Dietzia sp. E1 cells were found to possess three distinct levels of cell-surface hydrophobicity. Gas chromatographic/mass spectrometric analysis revealed that intracellular substrate mineralization occurred through the conversion of n-alkane to the corresponding n-alkanal. The monoterminal oxidation pathway was presumably initiated by AlkB and CYP153 terminal alkane hydroxylases, both of their partial coding sequences were successfully detected in the genome of strain E1, a novel member of the Dietzia genus.     

Hydrocarbon degradation in relation to cell-surface hydrophobicity among bacterial hydrocarbon degraders from petroleum-contaminated Kuwait desert environment

Forty six bacterial isolates able to grow on crude oil were isolated from various hydrocarbon-contaminated sites in Kuwait. The extent of crude oil degradation varied over a wide range (1–87%) among the isolates. Isolates were predominantly Gram-positive bacteria (79% of total isolates) belonging to the genera Bacillus (93%) and Paenibacillus (7%). Among the few Gram-negative isolates were from the genera Acinetobacter, Alcaligenes, Klebsiella, Burkholderia, Pseudomonas, and Williamsia. Analyses of their cell-surface hydrophobicity (CSH) by various methods equally showed a wide variation among the isolates. About 74% of isolates that degraded significant amounts of crude oil (>40% degradation) possessed high level of CSH, while 58% of all the isolates exhibited high levels of CSH. Statistical analyses showed significantly high correlation between the ability to degrade crude oil and CSH. The ability of the isolates to bind to polystyrene and salt-aggregation test as measures of CSH were more strongly correlated with hydrocarbon-degrading ability than adherence to hydrocarbons.