Effect of photosynthetic bacteria and compost on degradation of petroleum products in soil

Addition of diesel fuel and waste engine oil to soil was found to cause biostimulation of hydrocarbon-oxidizing microorganisms. Corynebacteria constitute a large group of hydrocarbon-oxidizing microorganisms. Addition of a liquid culture of photosynthetic bacteria to soil not only facilitates degradation of petroleum products, but also stimulates growth of hydrocarbon-oxidizing microorganisms. Combined addition of photosynthetic bacteria and compost to soil polluted with petroleum products causes even a more significant increase in the count of hydrocarbon-oxidizing bacteria and substantially increases the rate of pollutant degradation.     

Ratio [13C]/[12C] as an index for express estimation of hydrocarbon-oxidizing potential of microbiota in soil polluted with crude oil

The hydrocarbon-oxidizing potential of soil microbiota and hydrocarbon-oxidizing microorganisms introduced into soil was studied based on the quantitative and isotopic characteristics of carbon in products formed in microbial degradation of oil hydrocarbons. Comparison of CO2 production rates in native soil and that polluted with crude oil showed the intensity of microbial mineralization of soil organic matter (SOM) in the presence of oil hydrocarbons to be higher as compared with non-polluted soil, that is, revealed a priming effect ofoil. The amount of carbon of newly synthesized organic products (cell biomass and exometabolites) due to consumed petroleum was shown to significantly exceed that of SOM consumed for production of CO2. The result of microbial processes in oil-polluted soil was found to be a potent release of carbon dioxide to the atmosphere.     

Ratio [13C]/[12C] as an index for express estimation of hydrocarbon-oxidizing potential of microbiota in soil polluted with crude oil

The hydrocarbon-oxidizing potential of soil microbiota and hydrocarbon-oxidizing microorganisms introduced into soil was studied based on the quantitative and isotopic characteristics of carbon in products formed in microbial degradation of oil hydrocarbons. Comparison of CO₂ production rates in native soil and that polluted with crude oil showed the intensity of microbial mineralization of soil organic matter (SOM) in the presence of oil hydrocarbons to be higher as compared with non-polluted soil, that is, revealed a priming effect of oil. The amount of carbon of newly synthesized organic products (cell biomass and exometabolites) due to consumed petroleum was shown to significantly exceed that of SOM consumed for production of CO₂. The result of microbial processes in oil-polluted soil was found to be a potent release of carbon dioxide to the atmosphere.     

Catalase activity of hydrocarbon-oxidizing bacteria

The dynamics of catalase activity of the hydrocarbon-oxidizing bacteria Gordona terrae, Rhodococcus rubropertinctus, and Rhodococcus erythropolis during petroleum product destruction has been studied. A direct relationship between decreasing catalase activity of hydrocarbon-oxidizing microorganisms and the intensity of petroleum product destruction has been established experimentally. The revealed dependence allows one to consider the catalase activity of bacteria as an indicator of the initial stage of petroleum product oxidation and may be used for choosing destructor strains to construct biopreparations suitable for natural ecosystem remediation.     

油气微生物勘探机理及应用

【目的】微生物油气勘探技术是基于油气藏的轻烃微渗漏原理衍生的地表勘探技术。油气藏中的轻烃部分(C1-C5)以微渗漏的方式通过上覆的沉积层,在近地表土壤中诱导专门利用轻烃的微生物繁殖与生长,油气区的微生物种类与浓度有别于下伏没有油气藏的地区。通过分析微生物的浓度异常特征,对油气富集区及油气藏进行研究和预测。【方法】在人工模拟条件下研究油气微生物数量和群落异常特征,在此基础上,以海相碳酸盐岩气田普光气田为研究对象,进一步开展微生物勘探研究,鉴定油气藏上方气态烃氧化过程的微生物驱动者,提取土壤中的微生物异常信息。【结果】人工模拟条件下发现Lacibacter cauensis、Methylococcaceae、Methylophilaceae与甲烷气体培养正相关(气指示菌),而未培养的硫氧化微生物等则与丁烷培养正相关(油指示菌)。【结论】进一步在普光气田原位研究中进行验证,发现地表油气微生物数量和群落异常与油气藏有较好的关联性;与油气化探指标对比后发现,油气藏上方微生物正异常和轻烃负异常具有较为明显的互补关系。本研究深化了对典型油气藏上方气态烃氧化微生物转化机制的认识,为油气微生物勘探技术提供理论与实践依据。     

The properties of hydrocarbon-oxidizing bacteria isolated from the oilfields of Tatarstan, Western Siberia, and Vietnam

Eleven strains of hydrocarbon-oxidizing bacteria, isolated from oilfields, representing the genera Rhodococcus, Gordonia, Dietzia, and Pseudomonas, were characterized as mesophiles and neutrophiles. Rhodococci were halotolerant microorganisms growing in a media containing up to 15% NaCl. All the strains oxidized n-alkanes of crude oil. An influence of the cultivation temperatures (28 or 45 degrees C) and organic supplements on the degradation of C12-C30 n-alkanes in oxidized oil by two bacterial strains of the genus Pseudomonas was shown. The introduction of acetate, propionate, butyrate, ethanol, and sucrose led mainly to the decreased oxidation of petroleum paraffins. At certain cultivation temperatures, the addition of volatile fatty acid salts increased the content of individual n-alkanes in oxidized vs. crude oil.     

Effect of butyric acid on physiologic activity of carbohydrate-oxidizing rhodococci]

Laboratory experiments showed that butyric acid not only fails to meet the trophic requirements of hydrocarbon-oxidizing microorganisms, but even specifically inhibits their assimilatory and dissimilatory activity. Therefore, butyric acid can be referred to as growth inhibitors. The combined mineralization of carbohydrates and hydrocarbons can be described as follows. Plants polymers are converted to monosugars by heterotrophic soil microorganisms. As the concentration of the monosugars grows and oxygen becomes deficient, the monosugars are no longer oxidized completely but are fermented. As a result, glucose transforms to butyric acid, which inhibits hydrocarbon-oxidizing bacteria. It is concluded that, to be efficient, the cleanup of oil-contaminated soils must include measures to intensify the mineralization of carbohydrates and to inhibit their fermentation.     

Effect of Butyric Acid on the Physiological Activity of Hydrocarbon-Oxidizing Rhodococci

Laboratory experiments showed that butyric acid not only fails to meet the trophic requirements of hydrocarbon-oxidizing microorganisms, but even specifically inhibits their assimilatory and dissimilatory activity. Therefore, butyric acid can be referred to as growth inhibitors. The combined mineralization of carbohydrates and hydrocarbons can be described as follows. Plants polymers are converted to monosugars by heterotrophic soil microorganisms. As the concentration of the monosugars grows and oxygen becomes deficient, the monosugars are no longer oxidized completely but are fermented. As a result, glucose transforms to butyric acid, which inhibits hydrocarbon-oxidizing bacteria. It is concluded that, to be efficient, the cleanup of oil-contaminated soils must include measures to intensify the mineralization of carbohydrates and to inhibit their fermentation.     

小黑麦对石油污染盐碱土壤细菌群落与石油烃降解的影响

为了研究小黑麦对石油污染盐碱土壤中的细菌群落与石油烃降解率的影响,采用高通量测序技术,设置0 g/kg,1 g/kg和5 g/kg三个石油浓度,以未种植小黑麦的土壤作为对照,对6组不同处理的盐碱土壤样品的细菌群落结构及其多样性进行测定,并分析土壤中的石油烃降解率。结果表明:在土壤石油浓度为1 g/kg和5 g/kg时,小黑麦根际土壤中的石油烃降解率相较对照组分别提高了36.67%和33.20%。从6个土壤样品中分别获得21398—27899条测序序列。在石油污染土壤中,小黑麦根际土壤的细菌群落多样性和丰度均大于对照组的土壤。同时,在"门","纲","属"的分类水平上,小黑麦根际土壤细菌群落中的一些根际细菌的相对丰度增加了,主要包括变形菌门(Proteobacteria)、酸杆菌门(Acidobacteria)、γ-变形菌纲(Gammaproteobacteria)、烷烃降解菌科-未命名菌属(Alcanivoracaceae_norank)、黄单胞菌属(Xanthomonas)、亚硝化单胞菌-不可培养菌属(Nitrosomonadaceae_unculture)等。有一些相对丰度增加的根际细菌是以石油及石油分解物为碳源的微生物。本研究证明种植小黑麦改变了石油污染盐碱土壤根际土壤细菌群落结构组成和多样性,促进了降解石油微生物群落的构建,显著提高了盐碱土壤石油污染的降解效果。研究结果为石油污染盐碱土壤的植物修复奠定了理论基础。     

Hydrocarbon-oxidizing microorganisms in the waters of certain regions of the Western and Central Atlantic

The incidence of hydrocarbon-oxidizing microorganisms in water was determined by the method of plating on solid media. Vertical distribution of hydrocarbon-oxidizing microflora was different near the shore and in the open ocean; the incidence of the microorganisms was low in the surface water layer but increased at depths of 25 and 75 m in the open ocean in contrast to regions near the shore. Pure bacterial and fungal cultures were isolated and their properties were described. The cultures were grown in a liquid mineral medium with diesel fuel and 50 cultures out of 66 were found to be true hydrocarbon-oxidizing microorganisms surviving under the laboratory conditions. It was shown that bacteria and fungi have grown on tarballs collected from the surface of the ocean during their incubation at 30 degrees C.     

Taxonomic diversity of aerobic organotrophic bacteria from clean vietnamese soils and their capacity for oxidation of petroleum hydrocarbons

The dominant species and abundance of the cultured aerobic organotrophic bacteria were determined in the clean soils of the Republic of Vietnam. The total number of organotrophs varied from 2.0 × 10⁵ to 5.8 × 10⁸ CFU/g soil. A considerable fraction of the bacterial population (1.1 × 10⁵–9.5 × 10⁶ CFU/g soil) was able to utilize petroleum hydrocarbons as the sole carbon and energy source. Most of the organisms obtained in pure cultures were gram-positive bacteria; over 70% were hydrocarbon-oxidizing organisms. Analysis of 16S rRNA gene sequences resulted in tentative determination of the taxonomic position of 22 strains, with 12 belonging to the Firmicutes, 4, to the Proteobacteria, and 6 to the Actinobacteria. The most common bacteria capable of hydrocarbon oxidation belonged to the genera Acinetobacter, Bacillus, Brevibacillus, Chromobacterium, Cupriavidus, Gordonia, Microbacterium, Mycobacterium, and Rhodococcus. Some of the isolated Bacillus and Staphylococcus strains, as well as one Pseudomonas and one Sinomonas strain, did not utilize hydrocarbons. Gram-positive degraders, especially members of the order Actinomycetales, which exhibited high hydrocarbon-oxidizing activity, gained competitive advantage in the presence of hydrocarbons. This microbial group probably plays an important role in hydrocarbon degradation in tropical soils. Thus, Vietnamese soils, which had no history of petroleum contamination, support numerically significant and taxonomically diverse populations of h ydrocarbon-oxidizing bacteria.     

Development of an oil-degrading biopreparation by activation of aboriginal hydrocarbon-oxidizing microflora

A method of activation of aboriginal hydrocarbon-oxidizing microorganisms for remediation of soil and water basins polluted with oil products was developed. The optimum composition of activating additives was found (g/l): mineral components, 10.0; oil, 5.0; and a synthetic detergent, 0.2. The resulting biopreparations increased the degree of purification by factors of 4-8 in soil and 18-24 in water when applied at a concentration of 10(7) cells/g(ml).     

Role of Humic Compounds in Viability Prolongation of the Cells of Hydrocarbon-Oxidizing Bacteria

Microbiology - Addition of humic compounds (9 preparations) to the cultures of hydrocarbon-oxidizing bacteria was shown to result in up to 15-fold increase in the number of viable cells in the...     

Optimization of the purification of soil and water objects from oil using biosorbents

Oil biosorbents (patents 2299181, 2318736) were obtained using immobilization of oil-oxidizing microorganisms into the hydrophobic sorbent Sorbonaft, which is manufactured using special technology at the Press-Torf Company. Two associations of aboriginal hydrocarbon-oxidizing microorganisms were used for this purpose: a fungal association and a bacterial and yeast association. The application of biosorbents resulted in a substantial acceleration of the process of purification from oil. The decrease in the amount of oil in the water and soil during 1 month was 30–44% in the variants with the products, against 5% in the control.     

Screening method for detection of hydrocarbon-oxidizing bacteria in oil-contaminated water and soil specimens

Toxic action of crude oil on the living world and ecosystems in general is a global problem of both aquatic and terrestrial environments. Bearing in mind the possibility of biodegradation of this toxicant, the procedures of determining counts and activity of cultivable microorganisms, and especially of bacteria responsible for degradation processes, are of great significance. The aim of this work was to study the possibility of modifying some solid media by adding triphenyltetrazolium chloride reagent as an indicator of the dehydrogenase activity, to develop a simple screening method for a simultaneous assessment of the count and activity of cultivable hydrocarbon-oxidizing bacteria in the oil-contaminated environments. The modified method appeared to be rapid and very suitable for the intended purposes.     

Microbial communities and their ability to oxidize n-alkanes in the area of release of gas- and oil-containing fluids in Mid-Baikal (Cape Gorevoi Utes)

The microbial community in the area of oil seep in Mid-Baikal (Cape Gorevoi Utes) was studied. The number of microorganisms that oxidize normal hydrocarbons, petroleum, and easily accessible organic matter in the water mass of the lake, bottom sediments, and bitumen structures was studied in 2005?C2009. The high heterogeneity of the distribution of microorganisms associated with the deparaffination of oil in the areas of oil seeps was noted. The maximum concentrations of hydrocarbon-oxidizing microorganisms in the samples of bottom water above bitumen structures (up to 2200 ± 175 CFU/mL) and in bitumen structures themselves (up to 170 000 ± 13 000 CFU/g) were determined. A model experiment showed that in the conditions of low temperatures (4°C) the degradation of the fraction of oil n-alkanes by the natural microbial community reaches 90% over a period of 60 days.     

Development of an Oil-Degrading Biopreparation by Activation of Aboriginal Hydrocarbon-Oxidizing Microflora

A method of activation of aboriginal hydrocarbon-oxidizing microorganisms for remediation of soil and water basins polluted with oil products was developed. The optimum composition of activating additives was found (g/l): mineral components, 10.0; oil, 5.0; and a synthetic detergent, 0.2. The resulting biopreparations increased the degree of purification by factors of 4–8 in soil and 18–24 in water when applied at a concentration of 10⁷ cells/g(ml).     

Distribution of microorganisms in the oil-polluted ground of vadose and saturation zones

The ground polluted with oil products was analyzed at a depth of 0.5–7.8 m (loamy soil), 11.5–13.0 m (gravelstone), 13.0–15.0 m (siltstone). It was shown that the distribution of oil products and microorganisms in the ground over the profile depends on the hydrogeological properties of the rock (porosity, hydraulic conductivity). The number of aerobic heterotrophic microorganisms varied from 10⁶–10⁷ CFU/g, the fraction of hydrocarbon-oxidizing ones increased with depth from 30 to 85%. The number of anaerobic microorganisms was comparable to the number of aerobic ones. The number of psychrotrophs and psychrophiles increases with depth; in the lower horizon these organisms prevail over the number of mesophiles.     

Verification of Degradation of <Emphasis Type="Italic">n</Emphasis>-Alkanes in Diesel Oil by <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> Strain WatG in Soil Microcosms

Degradation of n-alkanes in diesel oil by Pseudomonas aeruginosa strain WatG (WatG) was verified in soil microcosms. The total petroleum hydrocarbon (TPH) degradation level in two bioaugmentation samples was 51% and 46% for 1 week in unsterilized and sterilized soil microcosms, respectively. The TPH degradation in the biostimulation was of control level (15%). The TPH degradation in aeration-limited samples was clearly reduced when compared with that in aeration-unlimited ones under both sterilized and unsterilized conditions. Addition of WatG into soil microcosms was accompanied by dirhamnolipid production only in the presence of diesel oil. These findings suggest that degradation of n-alkanes in diesel oil in soil microcosms would be facilitated by bioaugmentation of WatG, with production of dirhamnolipid, and also by participation of biostimulated indigenous soil bacteria.