Biodegradation of binary mixtures of octane with benzene, toluene, ethylbenzene or xylene (BTEX): insights on the potential of Burkholderia, Pseudomonas and Cupriavidus isolates.

The contamination of the environment by crude oil and its by-products, mainly composed of aliphatic and aromatic hydrocarbons, is a widespread problem. Biodegradation by bacteria is one of the processes responsible for the removal of these pollutants. This study was conducted to determine the abilities of Burkholderia sp. B5, Cupriavidus sp. B1, Pseudomonas sp. T1, and another Cupriavidus sp. X5 to degrade binary mixtures of octane (representing aliphatic hydrocarbons) with benzene, toluene, ethylbenzene, or xylene (BTEX as aromatic hydrocarbons) at a final concentration of 100 ppm under aerobic conditions. These strains were isolated from an enriched bacterial consortium (Yabase or Y consortium) that prefer to degrade aromatic hydrocarbon over aliphatic hydrocarbons. We found that B5 degraded all BTEX compounds more rapidly than octane. In contrast, B1, T1 and X5 utilized more of octane over BTX compounds. B5 also preferred to use benzene over octane with varying concentrations of up to 200 mg/l. B5 possesses alkane hydroxylase (alkB) and catechol 2,3-dioxygenase (C23D) genes, which are responsible for the degradation of alkanes and aromatic hydrocarbons, respectively. This study strongly supports our notion that Burkholderia played a key role in the preferential degradation of aromatic hydrocarbons over aliphatic hydrocarbons in the previously characterized Y consortium. The preferential degradation of more toxic aromatic hydrocarbons over aliphatics is crucial in risk-based bioremediation.

Long-Term Biocide Efficacy and Its Effect on a Souring Microbial Community.

Reservoir souring, which is the production of H2S mainly by sulfate-reducing microorganisms (SRM) in oil reservoirs, has been a long-standing issue for the oil industry. While biocides have been frequently applied to control biogenic souring, the effects of biocide treatment are usually temporary, and biocides eventually fail. The reasons for biocide failure and the long-term response of the microbial community remain poorly understood. In this study, one-time biocide treatments with glutaraldehyde (GA) and an aldehyde-releasing biocide (ARB) at low (100 ppm) and high (750 ppm) doses were individually applied to a complex SRM community, followed by 1 year of monitoring of the chemical responses and the microbial community succession. The chemical results showed that souring control failed after 7 days at a dose of 100 ppm regardless of the biocide type and lasting souring control for the entire 1-year period was achieved only with ARB at 750 ppm. Microbial community analyses suggested that the high-dose biocide treatments resulted in 1 order of magnitude lower average total microbial abundance and average SRM abundance, compared to the low-dose treatments. The recurrence of souring was associated with reduction of alpha diversity and with long-term microbial community structure changes; therefore, monitoring changes in microbial community metrics may provide early warnings of the failure of a biocide-based souring control program in the field. Furthermore, spore-forming sulfate reducers (Desulfotomaculum and Desulfurispora) were enriched and became dominant in both GA-treated groups, which could cause challenges for the design of long-lasting remedial souring control strategies. IMPORTANCE Reservoir souring is a problem for the oil and gas industry, because H2S corrodes the steel infrastructure, downgrades oil quality, and poses substantial risks to field personnel and the environment. Biocides have been widely applied to remedy souring, but the long-term performance of biocide treatments is hard to predict or to optimize due to limited understanding of the microbial ecology affected by biocide treatment. This study investigates the long-term biocide performance and associated changes in the abundance, diversity, and structure of the souring microbial community, thus advancing the knowledge toward a deeper understanding of the microbial ecology of biocide-treated systems and contributing to the improvement of current biocide-based souring control practices. The study showcases the potential application of incorporating microbial community analyses to forecast souring, and it highlights the long-term consequences of biocide treatment in the microbial communities, with relevance to both operators and regulators.

Prokaryotic community diversity during bioremediation of crude oil contaminated oilfield soil: effects of hydrocarbon concentration and salinity.

Crude oil extracted from oilfield reservoirs brings together hypersaline produced water. Failure in pipelines transporting this mixture causes contamination of the soil with oil and hypersaline water. Soil salinization is harmful to biological populations, impairing the biodegradation of contaminants. We simulated the contamination of a soil from an oilfield with produced water containing different concentrations of NaCl and crude oil, in order to evaluate the effect of salinity and hydrocarbon concentration on prokaryote community structure and biodegradation activity. Microcosms were incubated in CO2-measuring respirometer. After the incubation, residual aliphatic hydrocarbons were quantified and were performed 16S rRNA gene sequencing. An increase in CO2 emission and hydrocarbon biodegradation was observed with increasing oil concentration up to 100 g kg-1. Alpha diversity decreased in oil-contaminated soils with an increase in the relative abundance of Actinobacteria and reduction of Bacteroidetes with increasing oil concentration. In the NaCl-contaminated soils, alpha diversity, CO2 emission, and hydrocarbon biodegradation decreased with increasing NaCl concentration. There was an increase in the relative abundance of Firmicutes and Proteobacteria and a reduction of Actinobacteria with increasing salt concentration. Our results highlight the need to adopt specific bioremediation strategies in soils impacted by mixtures of crude oil and hypersaline produced water.

Cupidesulfovibrio liaohensis gen. nov., sp. nov., a novel sulphate-reducing bacterium isolated from an oil reservoir and reclassification of Desulfovibrio oxamicus and Desulfovibrio termitidis as Cupidesulfovibrio oxamicus comb. nov. and Cupidesulfovibrio termitidis comb. nov.

A novel sulphate-reducing, Gram-stain-negative, anaerobic strain, isolate XJ01T, recovered from production fluid at the LiaoHe oilfield, PR China, was the subject of a polyphasic study. The isolate together with Desulfovibrio oxamicus NCIMB 9442T and Desulfovibrio termitidis DSM 5308T formed a distinct, well-supported clade in the Desulfovibrionaceae 16S rRNA gene tree. The taxonomic status of the clade was underscored by complementary phenotypic data. The three isolates comprising the clade formed distinct phyletic branches and were distinguished using a combination of physiological features and by low average nucleotide identity and digital DNA-DNA hybridization values. Consequently, it is proposed that isolate XJ01T represents a novel genus and species for which the name Cupidesulfovibrio liaohensis gen. nov., sp. nov. is proposed with the type strain XJ01T (=CGMCC 1.5227T=DSM 107637T). It is also proposed that D. oxamicus and D. termitidis be reclassified as Cupidesulfovibrio oxamicus comb. nov. and Cupidesulfovibrio termitidis comb. nov., respectively.

Isolating, identifying and evaluating of oil degradation strains for the air-assisted microbial enhanced oil recovery process.

Due to the inefficient reproduction of microorganisms in oxygen-deprived environments of the reservoir, the applications of microbial enhanced oil recovery (MEOR) are restricted. To overcome this problem, a new type of air-assisted MEOR process was investigated. Three compounding oil degradation strains were screened using biochemical experiments. Their performances in bacterial suspensions with different amounts of dissolved oxygen were evaluated. Water flooding, microbial flooding and air-assisted microbial flooding core flow experiments were carried out. Carbon distribution curve of biodegraded oil with different oxygen concentration was determined by chromatographic analysis. The long-chain alkanes are degraded by microorganisms. A simulation model was established to take into account the change in oxygen concentration in the reservoir. The results showed that the optimal dissolved oxygen concentration for microbial growth was 4.5~5.5mg/L. The main oxygen consumption in the reservoir happened in the stationary and declining phases of the microbial growth systems. In order to reduce the oxygen concentration to a safe level, the minimum radius of oxygen consumption was found to be about 145m. These results demonstrate that the air-assisted MEOR process can overcome the shortcomings of traditional microbial flooding techniques. The findings of this study can help for better understanding of microbial enhanced oil recovery and improving the efficiency of microbial oil displacement.

Azospirillum oleiclasticum sp. nov, a nitrogen-fixing and heavy oil degrading bacterium isolated from an oil production mixture of Yumen Oilfield.

Two nitrogen-fixing and heavy oil degrading strains, designated RWY-5-1-1T and ROY-1-1-2, were isolated from an oil production mixture from Yumen Oilfield in China. The 16S rRNA gene sequence showed they belong to Azospirillum and have less than 96.1 % pairwise similarity with each species in this genus. The average nucleotide identity and digital DNA-DNA hybridization values between them and other type strains of Azospirillum species were less than 75.69 % and 22.0 %, respectively, both below the species delineation threshold. Pan-genomic analysis showed that the novel isolate RWY-5-1-1T shared 2145 core gene families with other type strains in Azospirillum, and the number of strain-specific gene families was 1623, almost two times more than the number known from other species. Furthermore, genes related to nitrogenase, hydrocarbon degradation and biosurfactant production were found in the isolates' genomes. Also, this strain was capable of reducing acetylene to ethylene at a rate of 22nmol ethylene h-1 (108 cells) and degrading heavy oil at a rate of 36.2 %. The major fatty acids and polar lipids were summed feature 8 (C18:1ω7c/C18:1ω6c), and phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol, and phosphatidylcholine. Furthermore, a combination of phenotypic, chemotaxonomic, phylogenetic and genotypic data clearly indicated that strains RWY-5-1-1T and ROY-1-1-2 represent a novel species, for which the name Azospirillum oleiclasticum sp. nov. is proposed. The type strain is RWY-5-1-1T (=CGMCC 1.13426T =KCTC 72259 T). Azospirillum novel strains with the ability of heavy oil degradation associated with the promotion of plant growth has never been reported to date.

Instigation of indigenous thermophilic bacterial consortia for enhanced oil recovery from high temperature oil reservoirs.

The purpose of the study involves the development of an anaerobic, thermophilic microbial consortium TERIK from the high temperature reservoir of Gujarat for enhance oil recovery. To isolate indigenous microbial consortia, anaerobic baltch media were prepared and inoculated with the formation water; incubated at 65°C for 10 days. Further, the microbial metabolites were analyzed by gas chromatography, FTIR and surface tension. The efficiency of isolated consortia towards enhancing oil recovery was analyzed through core flood assay. The novelty of studied consortia was that, it produces biomass (600 mg/l), bio-surfactant (325 mg/l), and volatile fatty acids (250 mg/l) at 65°C in the span of 10 days, that are adequate to alter the surface tension (70 to 34 mNm -1) and sweep efficiency of zones facilitating the displacement of oil. TERIK was identified as Clostridium sp. The FTIR spectra of biosurfactant indicate the presence of N-H stretch, amides and polysaccharide. A core flooding assay was designed to explore the potential of TERIK towards enhancing oil recovery. The results showed an effective reduction in permeability at residual oil saturation from 2.14 ± 0.1 to 1.39 ± 0.05 mD and 19% incremental oil recovery.

A novel exopolysaccharide-producing and long-chain n-alkane degrading bacterium Bacillus licheniformis strain DM-1 with potential application for in-situ enhanced oil recovery.

A novel Bacillus licheniformis strain (DM-1) was isolated from a mature reservoir in Dagang oilfield of China. DM-1 showed unique properties to utilize petroleum hydrocarbons and agroindustrial by-product (molasses) for exopolysaccharide (EPS) production under oil recovery conditions. The DM-1 EPS was proven to be a proteoglycan with a molecular weight of 568 kDa. The EPS showed shear thinning properties and had high viscosities at dilute concentrations (<1%, w/v), high salinities, and elevated temperatures. Strain DM-1 could degrade long-chain n-alkanes up to C36. Viscosity reduction test have shown that the viscosity of the crude oil was reduced by 40% compared with that before DM-1 treatment. Sand pack flooding test results under simulated reservoir conditions have shown that the enhanced oil recovery efficiency was 19.2% after 7 days of in-situ bioaugmentation with B. licheniformis DM-1. The obtained results indicate that strain DM-1 is a promising candidate for in situ microbial enhanced oil recovery (MEOR).

Genome repository of oil systems: An interactive and searchable database that expands the catalogued diversity of crude oil-associated microbes.

Microbial communities ultimately control the fate of petroleum hydrocarbons (PHCs) that enter the natural environment, but the interactions of microbes with PHCs and the environment are highly complex and poorly understood. Genome-resolved metagenomics can help unravel these complex interactions. However, the lack of a comprehensive database that integrates existing genomic/metagenomic data from oil environments with physicochemical parameters known to regulate the fate of PHCs currently limits data analysis and interpretations. Here, we curated a comprehensive, searchable database that documents microbial populations in natural oil ecosystems and oil spills, along with available underlying physicochemical data, geocoded via geographic information system to reveal their geographic distribution patterns. Analysis of the ~2000 metagenome-assembled genomes (MAGs) available in the database revealed strong ecological niche specialization within habitats. Over 95% of the recovered MAGs represented novel taxa underscoring the limited representation of cultured organisms from oil-contaminated and oil reservoir ecosystems. The majority of MAGs linked to oil-contaminated ecosystems were detectable in non-oiled samples from the Gulf of Mexico but not in comparable samples from elsewhere, indicating that the Gulf is primed for oil biodegradation. The repository should facilitate future work toward a predictive understanding of the microbial taxa and their activities that control the fate of oil spills.

Stimulation of indigenous microbes by optimizing the water cut in low permeability reservoirs for green and enhanced oil recovery.

Low permeability oil reservoirs are a widespread petroleum reservoir type all over the world. Therefore, methods to recover these reservoirs efficiently are of importance to guarantee energy supply. Here we report our novel stimulation of indigenous microbes by optimizing the water cut in low permeability reservoirs for green and enhanced oil recovery. We aimed to investigate the characteristics of indigenous bacterial communities with changes in water cut in reservoirs by high-throughput sequencing technology, and reveal the mechanism and characteristics of the crude oil biotreatment under different crude oil-water ratio conditions and the optimum activation time of indigenous functional microbial groups in reservoirs. The indigenous microbial metabolism products were characterized by gas chromatography mass spectrometry. Results showed that Acinetobacter (47.1%) and Pseudomones (19.8%) were the main functional genus of crude oil degradation at the optimal activation time, and can reduce the viscosity of crude oil from 8.33 to 5.75 mPa·s. The dominant bacteria genus for oil recovery after activation of the production fluids was similar to those in the reservoirs with water cut of 60-80%. Furthermore seven mechanism pathways of enhancing oil recovery by the synergistic of functional microbial groups and their metabolites under different water cut conditions in low permeability reservoirs have been established.

Low-Abundance Dietzia Inhabiting a Water-Flooding Oil Reservoir and the Application Potential for Oil Recovery.

With the development of molecular ecology, increasing low-abundance microbial populations were detected in oil reservoirs. However, our knowledge about the oil recovery potential of these populations is lacking. In this study, the oil recovery potential of low-abundance Dietzia that accounts for less than 0.5% in microbial communities of a water-flooding oil reservoir was investigated. On the one hand, Dietzia sp. strain ZQ-4 was isolated from the water-flooding reservoir, and the oil recovery potential was evaluated from the perspective of metabolisms and oil-displacing test. On the other hand, the strain has alkane hydroxylase genes alkB and P450 CYP153 and can degrade hydrocarbons and produce surfactants. The core-flooding test indicated that displacing fluid with 2% ZQ-4 fermentation broth increased 18.82% oil displacement efficiency, and in situ fermentation of ZQ-4 increased 1.97% oil displacement efficiency. Furthermore, the responses of Dietzia in the reservoir accompanied by the nutrient stimulation process was investigated and showed that Dietzia in some oil production wells significantly increased in the initial phase of nutrient injection and sharply decreased along with the continuous nutrient injection. Overall, this study indicates that Dietzia sp. strain has application potential for enhancing oil recovery through an ex situ way, yet the ability of oil recovery in situ based on nutrient injection is limited.

Changes in the Microbial Community Diversity of Oil Exploitation.

To systematically evaluate the ecological changes of an active offshore petroleum production system, the variation of microbial communities at several sites (virgin field, wellhead, storage tank) of an oil production facility in east China was investigated by sequencing the V3 to V4 regions of 16S ribosomal ribonucleic acid (rRNA) of microorganisms. In general, a decrease of microbial community richness and diversity in petroleum mining was observed, as measured by operational taxonomic unit (OTU) numbers, α (Chao1 and Shannon indices), and ß (principal coordinate analysis) diversity. Microbial community structure was strongly affected by environmental factors at the phylum and genus levels. At the phylum level, virgin field and wellhead were dominated by Proteobacteria, while the storage tank had higher presence of Firmicutes (29.3-66.9%). Specifically, the wellhead displayed a lower presentence of Proteobacteria (48.6-53.4.0%) and a higher presence of Firmicutes (24.4-29.6%) than the virgin field. At the genus level, the predominant genera were Ochrobactrum and Acinetobacter in the virgin field, Lactococcus and Pseudomonas in the wellhead, and Prauseria and Bacillus in the storage tank. Our study revealed that the microbial community structure was strongly affected by the surrounding environmental factors, such as temperature, oxygen content, salinity, and pH, which could be altered because of the oil production. It was observed that the various microbiomes produced surfactants, transforming the biohazard and degrading hydro-carbon. Altering the microbiome growth condition by appropriate human intervention and taking advantage of natural microbial resources can further enhance oil recovery technology.

In situ transformation of ethoxylate and glycol surfactants by shale-colonizing microorganisms during hydraulic fracturing.

In the last decade, extensive application of hydraulic fracturing technologies to unconventional low-permeability hydrocarbon-rich formations has significantly increased natural-gas production in the United States and abroad. The injection of surface-sourced fluids to generate fractures in the deep subsurface introduces microbial cells and substrates to low-permeability rock. A subset of injected organic additives has been investigated for their ability to support biological growth in shale microbial community members; however, to date, little is known on how complex xenobiotic organic compounds undergo biotransformations in this deep rock ecosystem. Here, high-resolution chemical, metagenomic, and proteomic analyses reveal that widely-used surfactants are degraded by the shale-associated taxa Halanaerobium, both in situ and under laboratory conditions. These halotolerant bacteria exhibit surfactant substrate specificities, preferring polymeric propoxylated glycols (PPGs) and longer alkyl polyethoxylates (AEOs) over polyethylene glycols (PEGs) and shorter AEOs. Enzymatic transformation occurs through repeated terminal-end polyglycol chain shortening during co-metabolic growth through the methylglyoxal bypass. This work provides the first evidence that shale microorganisms can transform xenobiotic surfactants in fracture fluid formulations, potentially affecting the efficiency of hydrocarbon recovery, and demonstrating an important association between injected substrates and microbial growth in an engineered subsurface ecosystem.

Isolation, identification, and characterization of diesel-oil-degrading bacterial strains indigenous to Changqing oil field, China.

In the present study, 12 indigenous diesel-oil-degrading bacteria were isolated from the petroleum-contaminated soils of the Changqing oil field (Xi'an, China). Measurement of the diesel-oil degradation rates of these strains by the gravimetric method revealed that they ranged from 42% to 66% within 2 weeks. The highest degradation rates were observed from strains CQ8-1 (66%), CQ8-2 (62.6%), and CQ11 (59%), which were identified as Bacillus thuringiensis, Ochrobactrum anthropi, and Bordetella bronchialis, respectively, based on their 16S rDNA sequences. Moreover, the physiological and biochemical properties of these three strains were analyzed by Gram staining, catalase, oxidase, and Voges-Proskauer tests. Transmission electron microscopy showed that all three strains were rod shaped with flagella. Gas chromatography and mass spectrometric analyses indicated that medium- and long-chain n-alkanes in diesel oil (C11-C29) were degraded to different degrees by B. thuringiensis, O. anthropi, and B. bronchialis, and the degradation rates gradually decreased as the carbon numbers increased. Overall, the results of this study indicate strains CQ8-1, CQ8-2, and CQ11 might be useful for environmentally friendly and cost-effective bioremediation of oil-contaminated soils.

Biosurfactant-Producing Capability and Prediction of Functional Genes Potentially Beneficial to Microbial Enhanced Oil Recovery in Indigenous Bacterial Communities of an Onshore Oil Reservoir.

Microbial enhanced oil recovery (MEOR) is a bio-based technology with economic and environmental benefits. The success of MEOR depends greatly on the types and characteristics of indigenous microbes. The aim of this study was to evaluate the feasibility of applying MEOR at Mae Soon Reservoir, an onshore oil reservoir experiencing a decline in its production rate. We investigated the capability of the reservoir's bacteria to produce biosurfactants, and evaluated the potentials of uncultured indigenous bacteria to support MEOR by means of prediction of MEOR-related functional genes, based on a set of metagenomic 16s rRNA gene data. The biosurfactant-producing bacteria isolated from the oil-bearing sandstones from the reservoir belonged to one species: Bacillus licheniformis, with one having the ability to decrease surface tension from 72 to 32 mN/m. Gene sequences responsible for biosurfactant (licA3), lipase (lipP1) and catechol 2,3-dioxygenase (C23O) were detected in these isolates. The latter two, and other genes encoding MEOR-related functional proteins such as enoyl-CoA hydratase and alkane 1-monooxygenase, were predicted in the bacterial communities residing the reservoir's sandstones. Exposure of these sandstones to nutrients, consisting of KNO3 and NaH2PO4, resulted in an increase in the proportions of some predicted functional genes. These results indicated the potentials of MEOR application at Mae Soon site. Using the approaches demonstrated in this study would also assist evaluation of the feasibility of applying MEOR in oil reservoirs, which may be enhanced by an appropriate nutrient treatment.

Characterizing the microbiome in petroleum reservoir flooded by different water sources.

Petroleum reservoir is an unusual subsurface biosphere, where indigenous microbes lived and evolved for million years. However, continual water injection changed the situation by introduction of new electron acceptors, donors and exogenous microbes. In this study, 16S-rRNA gene sequencing, comparative metagenomics and genomic bins reconstruction were employed to investigate the microbial community and metabolic potential in three typical water-flooded blocks of the Shen84 oil reservoir in Liaohe oil field, China. The results showed significant difference of microbial community compositions and metabolic characteristics existed between the injected water and the produced water/oil mixtures; however, there was considerable uniformity between the produced samples in different blocks. Microbial communities in the produced fluids were dominated by exogenous facultative microbes such as Pseudomonas and Thauera members from Proteobacteria phylum. Metabolic potentials for O2-dependent hydrocarbon degradation, dissimilarly nitrate reduction, and thiosulfate­sulfur oxidation were much more abundant, whereas genes involved in dissimilatory sulfate reduction, anaerobic hydrocarbon degradation and methanogenesis were less abundant in the oil reservoir. Statistical analysis indicated the water composition had an obvious influence on microbial community composition and metabolic potential. The water-flooding process accompanied with introduction of nitrate or nitrite, and dissolved oxygen promoted the alteration of microbiome in oil reservoir from slow-growing anaerobic indigenous microbes (such as Thermotoga, Clostridia, and Syntrophobacter) to fast-growing opportunists as Beta- and Gama- Proteobacteria. The findings of this study shed light on the microbial ecology change in water flooded petroleum reservoir.

Isolation and characterization of biosurfactant producing and oil degrading Bacillus subtilis MG495086 from formation water of Assam oil reservoir and its suitability for enhanced oil recovery.

The strains isolated from the formation water were characterized and screened considering their crude oil degradation capability and biosurfactant production ability. The growth kinetics study of isolated Bacillus subtilis MG495086 was carried out by varying growth parameters i.e. carbon source, temperature, pH and salinity. The biosurfactant production was optimized adopting RSM-CCD considering carbon source (1-5%), pH (3-11) and temperature (25-65 °C) as matrix parameters. The optimum biosurfactant production (6.3 ±â€¯0.1 g/L) and the minimum surface tension 29.85 mN/m were obtained after 96 h of incubation under optimal conditions i.e. 3.8% (v/v) of light-paraffin oil as sole carbon source at 62.4 °C and pH 7.7 with the maximum oil degradation capability of 91.3 ±â€¯5%. Critical micelle concentration value of crude biosurfactant was found to be 40 mg/L with high emulsification activity of 72.45 ±â€¯0.85%. The produced biosurfactant was identified as lipopeptide (Surfactin) and characterized using various analytical techniques to establish its suitability for microbial enhanced oil recovery.

Microbial community composition of a hydrocarbon reservoir 40 years after a CO2 enhanced oil recovery flood.

Injecting CO2 into depleted oil reservoirs to extract additional crude oil is a common enhanced oil recovery (CO2-EOR) technique. However, little is known about how in situ microbial communities may be impacted by CO2 flooding, or if any permanent microbiological changes occur after flooding has ceased. Formation water was collected from an oil field that was flooded for CO2-EOR in the 1980s, including samples from areas affected by or outside of the flood region, to determine the impacts of CO2-EOR on reservoir microbial communities. Archaea, specifically methanogens, were more abundant than bacteria in all samples, while identified bacteria exhibited much greater diversity than the archaea. Microbial communities in CO2-impacted and non-impacted samples did not significantly differ (ANOSIM: Statistic R = -0.2597, significance = 0.769). However, several low abundance bacteria were found to be significantly associated with the CO2-affected group; very few of these species are known to metabolize CO2 or are associated with CO2-rich habitats. Although this study had limitations, on a broad scale, either the CO2 flood did not impact the microbial community composition of the target formation, or microbial communities in affected wells may have reverted back to pre-injection conditions over the ca. 40 years since the CO2-EOR.

Indigenous microorganisms residing in oil sands tailings biodegrade residual bitumen.

The purpose of this study was to determine the capacity of indigenous microbes in tailings to degrade bitumen aerobically, and if acetate biostimulation further improved degradation. Fluid fine tailings, from Base Mine Lake (BML), were used as microbial inocula, and bitumen in the tailings served as a potential carbon source during the experiment. The tailings were capped with 0.22 µm-filtered BML surface water with or without BML bitumen and acetate addition and incubated for 100 days at 20 °C. CO2 production and petroleum hydrocarbon reductions (50-70% for the biostimulation treatment) in the tailings were observed. DNA was extracted directly from the tailings, and increased bacterial density was observed by qPCR targeting the rpoB gene in the biostimulated group. 16 S rRNA sequencing was used to determine microbial composition profiles in each treatment group. The microbial communities indigenous to the tailings shifted after the bitumen was added. Acidovorax, Rhodoferax, Pseudomonas and Pseudoxanthomonas spp. significantly increased compared to the original microbial community and demonstrated tolerance to bitumen-based toxicity. The first three genera showed more potential for biostimulation treatment with acetate and may be important bitumen/hydrocarbon-degraders in an oil sands end pit lake environment.

Prauserella oleivorans sp. nov., a halophilic and thermotolerant crude-oil-degrading actinobacterium isolated from an oil-contaminated mud pit.

A crude-oil-degrading, Gram-stain-positive actinobacterial strain, RIPIT, was isolated from a soil sample collected from an oil-contaminated mud pit in Khangiran oil and gas field, in the north-east of Iran. RIPIT was strictly aerobic, catalase- and oxidase-positive. The strain grew with 0-12.5 % (w/v) NaCl (optimum 3-5 %), at 25-55 °C (optimum 45 °C) and at pH 6.0-9.5 (optimum pH 7.0). The results of 16S rRNA gene sequence comparative analysis indicated that RIPIT represents a member of the genus Prauserella, with high phylogenetic similarity to Prauserella coralliicola SCSIO 11529T (97.5 %), Prauserella endophytica SP28S-3T (97.5 %) and Prauserella marina MS498T (97.2 %). DNA-DNA relatedness values between the novel strain and P. coralliicola DSM 45821T, P. endophytica DSM 46655T and P. marina DSM 45268T were 28 , 19 and 23 %, respectively. The cell wall peptidoglycan of RIPIT contained meso-diaminopimelic acid as the diamino acid and the whole-cell sugars are galactose and arabinose. The polar lipids pattern contained phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylinositol and two unknown phospholipids. Its cellular fatty acids pattern consisted of C17 : 1ω6c, iso-C16 : 0 and summed feature 3 (C16 : 1ω7c and/or iso-C15 : 0 2-OH), and the major respiratory quinone was MK-9(H4). The G+C content of the genomic DNA was 69 mol%. On the basis of polyphasic taxonomic data we propose that RIPIT represents a novel species of the genus Prauserella, for which the name Prauserella oleivorans sp. nov. is proposed. The type strain of Prauserellaoleivorans is RIPIT (=IBRC-M 10906T=LMG 28389T).