A. L. Tarasov

[Microbiological processes in a high-temperature oil field].

Thermophilic sulfate-reducing bacteria (SRB) oxidizing lactate, butyrate, and C12–C16n-alkanes of oil at a temperature of 90°C were isolated from samples of water and oil originating from oil reservoirs of the White Tiger high-temperature oil field (Vietnam). At the same time, no thermophiles were detected in the injected seawater, which contained mesophilic microorganisms and was the site of low-temperature processes of sulfate reduction and methanogenesis. Thermophilic SRB were also found in samples of liquid taken from various engineering reservoirs used for oil storage, treatment, and transportation. These samples also contained mesophilic SRB, methanogens, aerobic oil-oxidizing bacteria, and heterotrophs. Rates of bacterial production of hydrogen sulfide varied from 0.11 to 2069.63 at 30°C and from 1.18 to 173.86 at 70°C μg S/(l day); and those of methane production, varied from 58.4 to 100 629.8 nl CH4/(l day) (at 30°C). The sulfur isotopic compositions of sulfates contained in reservoir waters and of hydrogen sulfide of the accompanying gas indicate that bacterial sulfate reduction might be effective in the depth of the oil field.

A microbiological study of an underground gas storage in the process of gas extraction

The liquid phase of different units of an underground gas storage (UGS) in the period of gas injection was studied with respect to its hydrochemical composition and characterized microbiologically. The presence of viable aerobic and anaerobic bacteria was revealed in the UGS stratal and associated waters. An important source of microorganisms and biogenic elements in the ecosystem studied is water and various technogenic admixtures contained in trace amounts in the gas entering from the gas main in the period of gas injection into the storage. Owing to this fact, the bacterial functional diversity, number, and activity are maximal in the system of gas treatment and purification and considerably lower in the observation well zone. At the terminal stages, the anaerobic transformation of organic matter in the UGS aqueous media occurs via sulfate reduction and methanogenesis; exceptionally high rates of these processes (up to 4.9 × 105 ng S2− l−1 day−1 and 2.8 × 106 nl CH4 l−1 day−1, respectively) were recorded for above-ground technological equipment.

Dynamics of Microbial Processes in the Stratal Waters of the Romashkinskoe Oil Field

The dynamics of the microbial processes developing in parallel with the exploitation of the Romashkinskoe oil field (Tatarstan) were studied in two areas differing in the degree of stratal water freshening. Flooding of the strata, in conjunction with purposeful measures on stratal microflora activation, was shown to increase the microbial population density and activate both methanogenesis and sulfate reduction; the latter process was limited by the low sulfate concentration. Development of anaerobic processes correlated with changes in acetate concentration in the stratal water. High mineralization (over 200 g/l) inhibited the stratal water microflora even if other conditions were favorable. Isotopic analysis of the carbonate carbon showed that the bicarbonate concentration increased in the stratal water due to microbial degradation of oil hydrocarbons and further participation of the biogenic carbon dioxide in dissolution of the carbonate cement of the oil-bearing strata. In strongly desalinated stratal water, the proportion of the newly formed bicarbonate was as high as 80%.

Isolation and investigation of anaerobic microorganisms involved in methanol transformation in an underground gas storage facility

High methanol and acetate concentrations (up to 12 and 14 g l−1, respectively) were found in water samples collected at different objects of the North Stavropol underground gas storage facility (UGSF), and significant seasonal variations in the content of these compounds were revealed. The dominant anaerobic microorganisms isolated from these samples during the study belonged to acetogens, methanogens, and sulfate reducers. The results of 16S rRNA gene sequencing and analysis of the physiological properties showed that the isolates were close to the species of Eubacterium limosum, Sporomusa sphaeroides, Methanosarcina barkeri, Methanobacterium formicicum, and Desulfovibrio desulfuricans. The isolated organisms, except for Methanobacterium formicicum, were capable of methylotrophic growth. All strains were characterized by resistance to high methanol concentrations (up to 40–50 g l−1). Their other energy substrate was hydrogen. The combination of the growth characteristics of these strains (pH, temperature, and salinity ranges) was shown to correspond to the ecological situation observed in the UGSF. The results of investigation of the isolated strains suggest that organic acids (acetate, butyrate) found in high concentrations in the initial samples are metabolic products of the revealed acetogens. Based on the established biological peculiarities of the isolated strains of methanogens, acetogens, and sulfate-reducing bacteria, these microorganisms may be considered as the main agents of anaerobic transformation of methanol and some other organic and inorganic compounds in UGSFs.

Investigation of the trophic relations between anaerobic microorganisms from an underground gas repository during methanol utilization

Joint cultivation of the dominant strains of acetogenic, sulfate-reducing and methanogenic microorganisms isolated from water samples of the North Stavropol underground gas storage facility (UGSF) was carried out for revealing their probable trophic relationships. It was shown that acetogenic strains Eubacterium limosum AG12 and Sporomusa sphaeroides AG8-2 growing on methanol could form a considerable pool of hydrogen, which may support development of hydrogenotrophic cultures, the methanogen Methanobacterium formicicum MG134, or the sulfate reducer Desulfovibrio desulfuricans SR12. Growth of this sulfate-reducing strain was not stimulated under joint cultivation with Methanosarcina barkeri MGZ3 on methanol, probably due to its inability to take up low hydrogen concentrations observed during methanosarcina development. The results show that acetogens in the UGSF system are the most important consumers of methanol and hydrogen and after exhaustion of the latter and switching over to methanol utilization they can supply hydrogen to other microorganisms, including methanogens and sulfate reducers. The role of methanosarcina in the UGSF increases as the hydrogen and CO2 reserves are exhausted, and methanogenesis on methanol becomes the main way of its destruction.

Role of alkylhydroxybenzenes in bacterial adaptation to unfavorable growth conditions

The adaptogenic effect of the chemical analogues of alkylhydroxybenzenes (AHBs), bacterial extracellular autoregulators (the individual compound C7-AHB and its technical preparation Sidovit), was demonstrated for two pseudomonad species, Pseudomonas aeruginosa and P. fluorescens. The protective effect of AHBs resulted in increased growth rate and biomass accumulation in bacteria grown under suboptimal conditions within the species tolerance range. The adaptogenic effect of AHBs (10–50 μmg/l) was more pronounced under more unfavorable growth conditions. In the case of P. fluorescens, the individual compound C7-AHB increased the biomass yield by 30% under alkaline conditions (pH 9.5), when the growth rate decreased by 80–90% compared to the optimum (pH 5.5–7.5). The Sidovit preparation, containing a mixture of natural AHBs with C7-AHB as the main component, increased the growth rate of P. aeruginosa by 40–60% at nonoptimal temperatures (45 and 10°C) or under enhanced salinity (1% NaCl). The action of AHBs as regulators of the rpoS and SOS response stress regulons was demonstrated to be among the mechanisms of their adaptogenic effect, as was demonstrated with the relevant reporter genes in the model strains E. coli C600 thi, thr, leuΔ(pro-lac) with the osmE-lacZ and umuD-lacZ hybrid operons, respectively. AHBs are technologically and economically acceptable as adaptogenic supplements for bacterial preparations used in soil bioremediation and oil spillage removal under conditions unfavorable for microbial growth, including increased salinity, extreme pH, and fluctuating sub- or supraoptimal temperatures.

Antimicrobic features of phenolic lipids

The connection between the efficiency of phenolic lipids and their hydrophobic property (solubility) and hydrophobic property of microorganisms’ cell structure is shown. The mixture of amphiphilic di(oxiphenil)-phenil-methanes, which act bacteriostatically under 15 mg/l, possesses maximal efficiency against Staphylococcus aureus. Against Mycobacterium smegmatis with hydrophobic cell wall, hydrophobic 2,4-dialkylocibenzol 70 mg/l was the most effective. Hexylresorcin (HR) stops the development of gram-positive bacteria in concentrations 20–50 mg/l, that of gram-negative bacteria in concentration 65 mg/l, that of M. smegmatis at 70 mg/l, and that of yeast and fungi at 300 mg/l. HR prevails bacteria spores germination in the concentration 25–100 mg/l. The dependence of antibacterial action of isomers and homologues of alkylresorcins on their structure-number, position, and length of alkyl substituents-is studied.

Sulfate-reducing bacteria of the genus Desulfovibrio from south vietnam seacoast

Nine strains of sulfate-reducing bacteria were isolated from a biofouling of corroded steel samples incubated in a marine environment near Nha Trang, South Vietnam. Sulfate-reducing bacteria were obtained from all samples with black corrosion products (in rust-filled metal cavities, beneath the Balanus and oyster booths, and beneath Bryozoa or algal colonies). Analysis of the 16S rRNA gene sequences of these strains showed that they belonged to the genus Desulfovibrio, with D. salexigens, D. marinisediminis, D. alaskensis, D. bizertensis D. indonesiensis, and D. dechloracetivorans as the closest phylogenetic relatives (98–99% similarity). According to the 16S rRNA gene sequencing, one Desulfovibrio isolate was related to “D. caledoniensis”, although the similarity did not exceed 97.0%. All strains utilized hydrogen (in the presence of acetate and CO2), lactate, pyruvate, formate, and fumarate, but not acetate. Utilization of other substrates varied from strain to strain. Some isolates were capable of slow autotrophic growth with H2 as the sole electron donor. D. indonesiensis and D. alaskensis strains were tolerant to long-term exposure to atmospheric oxygen exposure and could grow in the presence of 0.1% O2 in the gas phase.

Microbial Potential for Cleaning the Oiled Iron Scale

The possibility of using microorganisms to clean oiled iron scale of metallurgical production was investigated with the goal of recuperation. A stable microbial association growing on mineral oil as the sole carbon source was isolated from a sample from oiled iron scale taken directly from a metallurgical plant. For microbial cultures isolated from this association, the taxonomic position, as well as their morphological and cultural characteristics, were determined. The microorganisms belonged to the genera Luteimonas, Alcanivorax, Flavobacterium, and Pseudomonas. Microbial associations oxidizing mineral oil were found to contain some microorganisms incapable of its utilization, which stimulated the hydrocarbon-oxidizing microflora. Application of the isolates, as well as of the strains from microbial collections, resulted in a 58% decrease in residual oil content in treated samples of the oiled iron scale.

Utilization of H2O2 as the Oxygen Source by Bacteria of the Genera Pseudomonas and Rhodococcus

The growth of bacteria of the genera Pseudomonas and Rhodococcus in the presence of hydrogen peroxide as the sole source of oxygen was studied. The toxic effect of H2O2 in the concentration range of 100–200 μg/ml was shown to extend the lag phase by two to three days. Apart from the peroxide toxicity, the bacterial growth was inhibited by the toxic effect of dissolved oxygen in concentrations over 100 μg O2/ml; in the presence of a liquid hydrocarbon phase, this effect was alleviated. Under decreased partial pressure of oxygen in the presence of hydrocarbons (12–15 vol %), culture growth was initiated at high initial concentrations of H2O2 (300 μg/ml). When hydrogen peroxide concentrations exceeded 320 μg/ml, no growth occurred, regardless of how much hydrocarbon was added.