Microbiological Methods for the Enhancement of Oil Recovery

Abstract

Unpredictable adverse results can ensue from blood transfusion therapies because of the donation of cells from a very large number of anonymous volunteers. There is little doubt that transfusable blood cells produced on a large scale in vitro would be candidate materials to replace cells donated from such a large group of anonymous individuals. The recent progress of technology for in vitro production of transfusable red blood cells is reviewed.     

油藏微生物的代谢特征与提高原油采收率技术

油藏是一个高温、高压、少氧、寡营养和封闭的极端环境,油田经过多年注水开发后,在油藏内部形成了相对稳定的微生物群落体系,这些微生物以石油烃分解为起始,形成了一个复杂的食物链,对油藏碳、硫和金属离子的元素地球化学循环起着非常重要的作用。微生物提高原油采收率技术(MEOR)是利用微生物及其代谢产物与油藏和原油发生作用来提高原油采收率的一种新技术,具有成本低、适应性强和环境友好等特点,因此有望成为未来化学驱后油藏和高含水油藏进一步提高采收率的重要手段。对油藏内源微生物及其介导的生化反应,微生物采油原理、发展历程和现场试验进行综述,并提出了未来的发展方向。     

Selection and Application of Microorganisms to Improve Oil Recovery

Microbial enhanced oil recovery (Meor) is an incontestably efficient alternative to improve oil recovery, especially in mature fields and in oil reservoirs with high paraffinic content. This is the case for most oil fields in the Recôncavo basin of Bahia, Brazil. Given the diverse conditions of most oil fields, an approach to apply Meor technology should consider primarily: (i) microbiological studies to select the appropriate microorganisms and (ii) mobilization of oil in laboratory experiments before oil field application. A total of 163 bacterial strains, selectively isolated from various sources, were studied to determine their potential to be used in Meor. A laboratory microbial screening based on physiological and metabolic profiles and growth rates under conditions representative for oil fields and reservoirs revealed that 10 bacterial strains identified as Pseudomonas aeruginosa (2), Bacillus licheniformis (2), Bacillus brevis (1), Bacillus polymyxa (1), Micrococcus varians (1), Micrococcus sp. (1), and two Vibrio species demonstrated potential to be used in oil recovery. Strains of B. licheniformis and B. polymyxa produced the most active surfactants and proved to be the most anaerobic and thermotolerant among the selected bacteria. Micrococcus and B. brevis were the most salt‐tolerant and polymer producing bacteria, respectively, whereas Vibrio sp. and B. polymyxa strains were the most gas‐producing bacteria. Three bacterial consortia were prepared with a mixture of bacteria that showed metabolic and technological complementarity and the ability to grow at a wide range of temperatures and salinity characteristics for the oil fields in Bahia, Brazil. Oil mobilization rates in laboratory column experiments using the three consortia of bacteria varied from 11.2 to 18.3 % [v/v] of the total oil under static conditions. Consortia of B. brevis, B. icheniformis and B. polymyxa exhibited the best oil mobilization rates. Using these consortia under anaerobic conditions could be an interesting alternative for Meor technology because their growth could be easily controlled through the administration of phosphate and inorganic electron acceptors. Bacterial strains selected in this work could be valuable for further application in oil recovery at productive and mature oil well sites as well as for the prevention and control of paraffin deposits.     

Microorganisms of Lake Baikal and Lake Nyasa as Indicators of Anthropogenic Influence: Prospects for Use in Biotechnology

Restriction endonucleases (RENs) were detected in 650 microbial strains isolated from water columns and bottom sediments of deep rift lakes, Baikal (Russia) and Nyasa (Southeastern Africa). They enzymes included unique (Fan I, Aca I, and Sse 91) and very rare (Bsi I, and Cci N I) species not typical of aquatic ecosystems. Water columns, deep cores, and bottom sediments of pure areas of the lakes contained no microorganisms with new RENs. Thus, the inshore areas of Lake Baikal, having been exposed to anthropogenic influences, may contain mutant bacterial strains expressing RENs that have not been described previously.     

Purification of Soil from Oil Pollutants with the Use of Denitrifying Hydrocarbon-Oxidizing Microorganisms

The efficiency of an oil-oxidizing microbial community in the bioremediation of oil-polluted soil was studied under laboratory conditions. A specific feature of the community was its ability to oxidize oil hydrocarbons under both aerobic and anoxic conditions. The degree of oil-hydrocarbon degradation in various bioremediation modes increased as follows: self-remediation (40%) < nitrate application (42%) < introduction of the denitrifying oil-oxidizing community (50%) < introduction of the denitrifying oil-oxidizing community plus nitrate application (60%). The intensification of bioremediation is related to the increase in the population of the hydrocarbon-oxidizing microorganisms, first of all, denitrifying ones, resulting from the introduction of the community.     

利用机油提高琼脂糖凝胶中DNA片段的回收率

目的:发展一种简单快速经济的回收琼脂糖凝胶中DNA的方法.方法:将切的琼脂糖凝胶胶块放入嵌套Eppendorf管中捣碎,加入50μL机油,室温下12 000r/min离心5min,取大Eppendorf管中收集的液体跑胶检验,凝皎上包括代表100bp～2000bp不同大小DNA分子回收率的分子量标准DL2000.结果:DNA回收率可由加机油前的约35%提高为加机油后的45%-90%,回收效率的波动主要取决于DNA片段的大小、切下的含有DNA的胶块的大小及操作者的熟练程度.结论:该方法快速、简便、经济,具有良好的重复性与特异性,比许多国产的试剂盒更可靠.     

Simplified Procedure for Recovery of Lignin Acidolysis Products for Determining the Lignin-Degrading Abilities of Microorganisms

A simplified procedure for the identification and measurement of single-ring aromatic products of lignin acidolysis is described. The procedure employed a 6-h hydrolysis of spruce milled wood lignin in acidic dioxane at 87°C, followed by a series of organic extractions to recover acidolysis products which were quantified by gas chromatography of trimethylsilyl derivatives. Complex gel permeation chromatography procedures utilized by other workers were avoided in the modified procedure, but equivalent results were obtained. The simplified procedure was utilized to hydrolyze sound and actinomycete-decayed spruce milled wood lignins and was shown to be useful as a technique for the rapid screening of microorganisms for their ability to alter lignin.     

Use of Tissue Culture and Biotechnology for the Genetic Improvement of Watermelon

Watermelon is an important vegetable crop world-wide with over 81 million metric tons produced annually. Despite these high production figures, million of metric tons of fruit are lost in fields to disease. Genetic improvement through tissue culture and biotechnology offer potential routes of improving fruit harvest by offering higher quality products, like seedless fruit, or by introducing recombinant genes or generating somaclonal variants with improved resistance to biotic or abiotic stresses. The purpose of this review is to highlight how tissue culture and biotechnology have been used for the genetic improvement of watermelon and provide suggestions for future application of these methods to facilitate further genetic improvement.     

Use of Colicin E3 for Biological Containment of Microorganisms

The genetic determinant of the lethal antibiotic colicin E3 was cloned under the control of a tightly regulated promoter in the absence of the gene for its cognate inhibitor. Combination of this killing cassette with a stringent regulatory element provided a substrate-dependent conditional suicide system that was exploited for the biological containment of a Pseudomonas putida strain. The lethality of a single gene copy and the distinct and universal cellular target of the antibiotic suggest colicin E3 as an ideal candidate for combination with other lethal functions to design highly efficient containment systems for microorganisms.     

Use of Bioluminescence for Detection of Genetically Engineered Microorganisms Released into the Environment

The persistence and movement of strain JS414 of Xanthomonas campestris pv. campestris, which was genetically engineered to bioluminesce, were monitored during a limited field introduction. Bioluminescence and traditional dilution plate counts were determined. Strain JS414 was applied to cabbage plants and surrounding soil by mist inoculation, by wound inoculation, by scattering infested debris among plants, and by incorporating bacteria into the soil. Bioluminescent X. campestris pv. campestris was detected in plant samples and in the rhizosphere up to 6 weeks after inoculation. Movement to uninoculated plants was detected on one occasion, but movement from the immediate release area was not detected. Strain JS414 was detected in soil samples beneath mist- and wound-inoculated plants only at intentionally infested locations and in aerial samples only on the day of inoculation. Our bioluminescence methods proved to be as sensitive as plating methods for detecting the genetically engineered microorganisms in environmental samples. Our results demonstrate that transgenic incorporation of the luxCDABE operon provides a non-labor-intensive, sensitive detection method for monitoring genetically engineered microorganisms in nature.     

噬菌体展示技术的发展及应用

噬菌体展示技术是一种用于筛选和改造功能性多肽的生物技术 ,编码多肽的ＤＮＡ片段与噬菌体表面蛋白的编码基因融合后 ,以融合蛋白的形式在噬菌体的表面表达出多肽序列。这是一种表型与基因型的统一。噬菌体展示技术最初是以Ｍ 13噬菌体为载体的 ,其宿主菌为大肠杆菌。以大肠杆菌为宿主的展示系统还有其他 ,如λ噬菌体和Ｔ4噬菌体等展示系统。还有利用真核细胞的病毒以及酵母菌作为展示系统的。这些展示系统各有各的优势 ,但最常用的仍是Ｍ 13噬菌体表达系统。最初的噬菌体展示系统是将外源肽或蛋白质与噬菌体外壳蛋白ＰⅢ或ＰⅧ的Ｎ末端融…     

油藏极端环境中的微生物

油藏属于高温、高压、高矿化度以及油、气、水共存的极端环境,与之相适应,油藏中孕育着丰富多样的微生物.介绍了油藏极端环境、油藏微生物的一般特征、典型的微生物生理群以及随油田开发微生物分布的变化.     

Use of Genetically Engineered Microorganisms (GEMs) for the Bioremediation of Contaminants

ABSTRACT

This paper presents a critical review of the literature on the application of genetically engineered microorganisms (GEMs) in bioremediation. The important aspects of using GEMs in bioremediation, such as development of novel strains with desirable properties through pathway construction and the modification of enzyme specificity and affinity, are discussed in detail. Particular attention is given to the genetic engineering of bacteria using bacterial hemoglobin (VHb) for the treatment of aromatic organic compounds under hypoxic conditions. The application of VHb technology may advance treatment of contaminated sites, where oxygen availability limits the growth of aerobic bioremediating bacteria, as well as the functioning of oxygenases required for mineralization of many organic pollutants. Despite the many advantages of GEMs, there are still concerns that their introduction into polluted sites to enhance bioremediation may have adverse environmental effects, such as gene transfer. The extent of horizontal gene transfer from GEMs in the environment, compared to that of native organisms including benefits regarding bacterial bioremediation that may occur as a result of such transfer, is discussed. Recent advances in tracking methods and containment strategies for GEMs, including several biological systems that have been developed to detect the fate of GEMs in the environment, are also summarized in this review. Critical research questions pertaining to the development and implementation of GEMs for enhanced bioremediation have been identified and posed for possible future research.     

青海油田微生物采油技术研究

从青海油田地层水中分离培养出4株微生物菌种,对其进行了耐温性、耐盐性等进行了探讨,分析了微生物处理前后原油组份及物性变化情况,并采用岩芯流动试验探讨了微生物的驱油效果。结果表明,这几种微生物菌种均能适应青海油田温度及高矿化度地层环境,作用于原油后对原油中的长链饱和烃类物质有较好的降解作用,能使其分子链变短,并产生有机酸或石油羧酸盐类表面活性剂等低分子物质,使原油的粘度、凝点及蜡含量均出现明显的下降,改善原油的流动性能。提高石油采收率。2口井的现场试验证明,微生物采油具有良好的提高石油采收率的效果和清蜡减阻效果。     

Recovery of Anaerobic Microorganisms from Clinical Specimens in Prereduced Media Versus Recovery by Routine Clinical Laboratory Methods

Prereduced anaerobically sterilized culture media, used with rigid adherence to the cultivation techniques described by Moore and his associates, were capable of recovering more than twice the number of anaerobic bacteria from clinical specimens than could be recovered by the conventional use of fluid thioglycolate medium and of blood-agar plates incubated anaerobically with hydrogen generation packets. No loss of clinical isolates was encountered with the more sensitive methods; however many of the isolates recovered only in prereduced media would not grow when placed into thioglycolate medium. A representative anaerobic isolate placed into aerobic transport broth was unable to survive beyond 30 min. Methods employing prereduced media were not difficult to master and were feasible for clinical laboratory use. Evidence implicating the gingival crevice flora as an important possible source of anaerobic bacteria that become involved in systemic infections was considered.     

On the Mechanism of Radiation Enhancement of Lethal Effect of Sodium Chloride on Microorganisms

In connection with the preceding paper concerning the enhancement of radiation lethal effect by the NaCl treatment, the experimental evidences and the further discussion for the possible mechanism of the enhancement by the post-NaCl treatment are presented.

In order to elucidate the radiation damages of cell constituents and its enhancement by NaCl, the infrared analysis and isotopic experiments were carried out with cells of S. cerevisiae, Z. soya and E. coli.

Consequently, it has been concluded that the possible mechanism may involve the processes of increase in cell permeability by irradiation and of the denaturation of the critical cell constituents by increasing penetration of NaCl and thus the sensitivity of irradiated cells to NaCl may increase. The NaCl sensitive sites are considered not to be restricted to the biological targets for radiation inactivation by the supports of the survival curve analysis based on the target theory and of the isotopic experiment.     

Enhancement of the Over-all Lethal Effect of Ionizing Radiations on Microorganisms by Sodium Chloride

Aiming at the enhancement of total lethal effect of radiations, chemical agent actions on irradiated cells were investigated. It was found that the sensitivity of irradiated cells to NaCl was markedly increased by radiation. In other words, the over-all lethal effect of radiations on microorganisms was enhanced by the pre- or post-treatment with NaCl. Some microbiological aspects of radiation enhancement of NaCl sensitivity were described in this paper, connecting with the possible mechanism. The fact that this phenomenon of radiation enhancement was observed in the case of the cells which gave the one-hit type survival curve suggests that radiation damage responsible for this phenomenon is not restricted to biological targets. Further discussion and experimental evidences will be presented in the subsequent paper.