Petroleum Degradation Research Articles

Experimentation on Degradation of Petroleum in Contaminated Soils in the Root Zone of Maize (Zea Mays L.) Inoculated with Piriformospora Indica

ABSTRACTPlant-based methods such as rhizodegradation are very promising for the remediation of petroleum-contaminated soils. Associations of plants with endophytes can further enhance their phytoremediation potential. In this study, a rhizobox experiment was conducted to investigate whether inoculation with the root-colonizing fungus Piriformospora indica could further enhance the degradation of petroleum hydrocarbons in the root zone of maize (Zea mays L.). The rhizoboxes were subdivided into compartments in accordance with distance from the plants. After filling the boxes with soil from a petroleum-contaminated site, seedlings that had either been inoculated with P. indica or not were grown in the middle compartments of the rhizoboxes and grown for 64 days. A plant-free treatment was included for control. The presence of roots strongly increased the counts of total and petroleum-degrading soil bacteria, respiration, dehydrogenase activity, water-soluble phenols and petroleum degradation. All these effects were also found in the soil adjacent to the middle compartments of the rhizoboxes, but strongly decreased further away from it. Inoculation with P. indica further enhanced all the recorded parameters without changing the spatial pattern of the effects. Inoculated plants also produced around 40% more root and shoot biomass than noninoculated plants and had greener leaves. Together, the results indicate that the treatment effects on the recorded soil microbial and biochemical parameters including petroleum hydrocarbon degradation were primarily due to increased root exudation. Irrespectively of this, they show that maize can be used to accelerate the rhizodegradation of petroleum hydrocarbons in soil and that inoculation with P. indica can substantially enhance the phytoremediation performance of maize.

Petroleum residues degradation in laboratory-scale by rhizosphere bacteria isolated from the mangrove ecosystem

This research is about petroleum bioremediation experiment to obtain bacterial isolate from mangrove ecosystem which potentially degrade petroleum. It was conducted in an Erlenmeyer batch system filled with growth medium of Stone Mineral Salt Solution (SMSS) plus petroleum residue, placed in an incubator shaker with a rotation speed of 120 rpm, temperature 3000C, for 14 research days. Indigenous bacteria that have been isolated and identified from the roots of mangrove plants are Ochrobactrum anthropi and Bacillus sp., Ralstonia pickettii and Bacillus circulans. Those bacteriain both monoculture and consortium form (mixed culture) are incorporated into erlenmeyer as remediator agents. All bacteria can utilize hydrocarbon compounds, but Ralstonia pickettii and Bacillus circulans reached exponential phase faster with more cell count than other bacteria. Compared to single cultures, petroleum degradation by a bacterial consortium provides a higher TPH reduction efficiency, i.e. at 5%, 10%, and 15% of initial TPH of 94.4%, 72%, and 80.3%, respectively. This study proved that all bacteria could optimize hydrocarbon compounds up to 15% TPH load.

Microbial Activity in Marine Sediments Exposed to Hexadecane: A Laboratory Study

Bacteria have developed metabolic pathways that allow them to use hydrocarbons as the source of carbon and energy, so that they now play a key role throughout the degradation of petroleum in natural environments. However, knowledge regarding the degradation of petroleum by microorganisms is far larger in soils than in marine environments. Hexadecane (HXD) is one of the most representative alkanes present in petroleum, and one of the most frequently used in laboratory trials to evaluate the degradation of hydrocarbons by microorganisms. In this study, the capacity for HXD degradation by microorganisms present in marine sediments collected at 15 and 200 m depth is determined experimentally. For 42 days, kinetics studies are carried out to measure the consumption of HXD. Every 7 days the number of colony forming units (CFU) and HXD concentration are determined. Additionally, a biomass sample for pyrosequencing is collected at the beginning of the experiment (t0), the time of maximum bacterial growth (t7 or t21) and at the end of the experiment (t42). A higher HXD consumption rate is observed in sediments from 15 m with respect to sediments from 200 m depth. The major HXD consumption occurs within the first 7 days for the sediments from 15 m and within 21 days for the sediments from 200 m depth; the biomass concentration CFU follows the same profile. The non‐metric multidimensional scaling analysis of the non‐cultivable bacteria indicates differences between bacteria species composition at different depths, which is confirmed by bacterial identification by pyrosequencing.

English

Hydrocarbon containing compounds like diesel, crude oil and petrol cause environmental pollution. Hydrocarbon contaminated sites contain large amount of microbial community adapted for utilizing these compound as a source of carbon and other metabolic activities. A study was conducted in order to isolate the micro-organism from hydrocarbon contaminated sites for petroleum degradation. The soil sample was collected from the bus terminal of Punjab University, Lahore. The isolate (Micrococcus sp.) was characterized microbiologically by using microbiological techniques such as staining and biochemical testing based on Bergey’s manual. Isolated organism has an ability to degrade the petrol as well as paraffin wax as a sole carbon source. This ability shows clear evidence that genome of this isolate harbors gene for degradation. By combined both properties, we can reduce petroleum product from hydrocarbon impacted environment. Key words: Petrol degradation, paraffin wax, biochemical tests, wax ball.

Petroleum degradation by endophytic Streptomyces spp. isolated from plants grown in contaminated soil of southern Algeria

Petroleum hydrocarbons are well known by their high toxicity and recalcitrant properties. Their increasing utilization around worldwide led to environmental contamination. Phytoremediation using plant-associated microbe is an interesting approach for petroleum degradation and actinobacteria have a great potential for that. For this purpose, our study aimed to isolate, characterize, and assess the ability of endophytic actinobacteria to degrade crude petroleum, as well as to produce plant growth promoting traits. Seventeen endophytic actinobacteria were isolated from roots of plants grown naturally in sandy contaminated soil. Among them, six isolates were selected on the basis of their tolerance to petroleum on solid minimal medium and characterized by 16S rDNA gene sequencing. All petroleum-tolerant isolates belonged to the Streptomyces genus. Determination by crude oil degradation by gas chromatorgraph-flame ionization detector revealed that five strains could use petroleum as sole carbon and energy source and the petroleum removal achieved up to 98% after 7 days of incubation. These isolates displayed an important role in the degradation of the n-alkanes (C6-C30), aromatic and polycyclic aromatic hydrocarbons.All strains showed a wide range of plant growth promoting features such as siderophores, phosphate solubilization, 1-aminocyclopropane-1-carboxylate deaminase, nitrogen fixation and indole-3-acetic acid production as well as biosurfactant production. This is the first study highlighting the petroleum degradation ability and plant growth promoting attributes of endophytic Streptomyces. The finding suggests that the endophytic actinobacteria isolated are promising candidates for improving phytoremediation efficiency of petroleum contaminated soil.

The responses of two native plant species to soil petroleum contamination in the Yellow River Delta, China.

Petroleum contamination is a significant environmental problem in the Yellow River Delta. The responses of two native salt-tolerant plant species, alfalfa (Medicago sativa) and bristle grass (Setaria uiridis Beauv), to soil petroleum contamination were investigated at five levels between 0 and 2.0% (w/w). Results showed that the total, aboveground and underground plant biomasses of both species were significantly reduced by petroleum contamination (p<0.05), with the inhibition enhanced with increased petroleum levels. However, the emergence rate of bristle grass was promoted by petroleum contamination. Following 100days of exposure, the number of soil petroleum degraders increased greatly, with a trend of initial increase followed by a decrease at 1.5% contamination or higher. Compared to bulk soils, bacteria-degrading alkanes, total hydrocarbons and PAHs in alfalfa rhizosphere soils increased by 1.33-4.18-, 0.85-3.01- and 4.12-12.75-fold, respectively, with an increase of 2.80-10.00-, 4.42-14.44- and 7.30-26.00-fold in bristle grass rhizosphere soils, respectively. The greatest number of petroleum degraders in bristle grass rhizosphere soils resulted in the highest petroleum degradation rate. Bristle grass may be the optimal species for petroleum remediation in the studied area.

Exploration of Up-regulated Key Proteins in Pseudomonas Aeruginosa for High-efficiency Petroleum Degradation by Proteomic Analysis.

In this work, proteomic analysis was used to identify the up-regulated key proteins of Pseudomonas aeruginosa (P6), a bacteria used in petroleum degradation, responsible for its high efficiency in degrading crude oil. Seventeen proteins were identified as up-regulated proteins by proteomic analysis and classified by bioinformatics analysis. The results indicated that most of the up-regulated proteins were responsible for P. aeruginosa (P6) survival under harsh environmental conditions and utilization crude oil as carbon source in a better way. The physiological processes, chemotaxis to carbon sources, terminal oxidation of carbons, carbon source uptake and nutrients transport, were associated with the up-regulated proteins in the study. The findings revealed the most influential proteins and set a clear direction for future research.

Effective bioremediation of a petroleum-polluted saline soil by a surfactant-producing Pseudomonas aeruginosa consortium

Bacteria able to produce biosurfactants can use petroleum-based hydrocarbons as a carbon source. Herein, four biosurfactant-producing Pseudomonas aeruginosa strains, isolated from oil-contaminated saline soil, were combined to form a bacterial consortium. The inoculation of the consortium to contaminated soil alleviated the adverse effects of salinity on biodegradation and increased the rate of degradation of petroleum hydrocarbon approximately 30% compared to the rate achieved in non-treated soil. In saline condition, treatment of polluted soil with the consortium led to a significant boost in the activity of dehydrogenase (approximately 2-fold). A lettuce seedling bioassay showed that, following the treatment, the soil's level of phytotoxicity was reduced up to 30% compared to non-treated soil. Treatment with an appropriate bacterial consortium can represent an effective means of reducing the adverse effects of salinity on the microbial degradation of petroleum and thus provides enhancement in the efficiency of microbial remediation of oil-contaminated saline soils.

Bioremediation of Diesel-contaminated Soil by Composting with Locally Generated Bulking Agents

ABSTRACTHerein, we conducted a study toward understanding the impact of composting of the diesel-contaminated soil with some locally available bulking agents (rice husks (RHs), sawdust (SD), and wood chips (WCs)). In order to ascertain the effectiveness of petroleum degradation by the process assayed, we compared the protocols with monitored natural attenuation (MNA). The overall degradation pattern was modeled with non-linear regression by comparing the experimental data with first and second-order kinetic equations. At the end of the six-week study, the amount of total petroleum hydrocarbon removed from contaminated soil was 98.26 ± 1.33% (amendment with SD + RHs + WCs), 96.89 ± 1.20 (RHs amendment), 96.55 ± 1.29% (amendment with SD), 90.01 ± 0.22% (WCs amendment), and 85.02 ± 0.21% (MNA). The degradation of TPH trends followed a second-order kinetic model for all the four compost treatments while the MNA was found to follow a first-order (slower) degradation pattern. In general, the results of the parameter estimate showed that amendment with mixture of the three bulking agents was 1.08 (8%) slower (k2 = (1.289 ± 0.16) × 10−5 (g mg−1 d−1), r2 = 0.991) than SD amendment alone (k2 = (1.392 ± 0.14) × 10−5 (g mg−1 d−1), r2 = 0.995). However, the mixture of the bulking agents was found to be 1.67, 1.41, and 2.4 times faster than amendments with WCs, rice, and MNA, respectively. The phytotoxicity test revealed that all the compost treatments except WCs resulted in germination index of ≥80% after six weeks of bioremediation tests. The outcome of the current investigation confirms the effectiveness of bulking agents (especially when combined) in the supply of nutrients for the bioremediation of diesel-impacted soil.

Fingerprinting of Weathered Oil Residues from the Deepwater Horizon Oil Spill: The Importance of Multiple Lines of Investigation

ABSTRACT 2017-189 The crude oil released from the Macondo Well, also known for its location in Mississippi Canyon area as the MC252 well during the Deepwater Horizon Oil Spill, entered an environment already containing a complex mixture of hydrocarbons from both natural and anthropogenic sources, many of which have closely related chemical profiles. To understand the impact of the released oil in offshore areas, a method was needed to distinguish MC252-related hydrocarbons from other sources. A multiple lines of evidence approach was developed to identify weathered MC252 oil in offshore sediments in the Gulf of Mexico. Chemical data for alkanes, PAH, petroleum biomarkers and metals were combined with spatial, temporal, and observational information to examine the fingerprints for more than 4,000 sediment samples collected over the span of five years. The unique conditions of the Gulf of Mexico (GOM), with many natural petroleum seeps and tepid seas, provided an ideal environment to support microbial degradation of petroleum. As a result of these conditions, the initial fingerprint of the MC252 was rapidly and extensively altered in the environment including depletion of petroleum biomarkers, usually assumed to be recalcitrant and often used in ratios to identify petroleum residues. Revised biomarker match criteria were defined to account for biodegradation within this fraction. Applying this methodology to the offshore sediment data from the GOM provided a comprehensive understanding of the distribution of the MC252 oil in offshore sediment and an understanding of the various transport pathways which conveyed the oil to the sediments.

BIODEGRADACIÓN AEROBIA DE FRACCIONES DE HIDROCARBUROS PROVENIENTES DE LA ACTIVIDAD PETROLERA EN UN SUELO DE LA REGIÓN PATAGONIA NORTE, ARGENTINA

Recent techniques for assessing hydrocarbon contaminated sites are mainly based on risk analysis methodology. The effects of chemicals present in contaminated sites on the human health and the ecosystem are usually estimated as the addition of individual risks. In this sense, the hydrocarbon concentration is a relevant data. Bioremediation is a sustainable technology that accelerates the full or partial degradation of petroleum in contaminated soils and it shows a suitable cost-benefit ratio over other remediation methods. This work describes a laboratory scale study of the degradation rate for petroleum fractions by the incorporation of “biosolids” (sewage sludge) in an oil contaminated Patagonian soil. The determination of the total hydrocarbon concentration and the count of microorganisms in soil were performed both at the beginning and the end of the bioremediation process. The hydrocarbon degradation was analyzed at regular intervals during the study. The observed results showed that alkanes and lower molecular weight aromatic hydrocarbon concentrations were negligible after 14 days of treatment. The heavy hydrocarbon fraction, considered recalcitrant to biodegradation, was also significantly reduced during the biodegradation process. Since heavy hydrocarbons have low volatility and scarse migration by leaching, the observed decrease indicates that heavy hydrocarbon elimination could be mainly due to the native microorganism activity in the north Patagonian soils. Comparison with conventional treatment indicates that addition of “biosolids” enhances hydrocarbon remediation and that the biodiversity of microorganisms in soils is an important factor in achieving significant petroleum degradation.

Hydrocarbon Degradation in Caspian Sea Sediment Cores Subjected to Simulated Petroleum Seepage in a Newly Designed Sediment-Oil-Flow-Through System.

The microbial community response to petroleum seepage was investigated in a whole round sediment core (16 cm length) collected nearby natural hydrocarbon seepage structures in the Caspian Sea, using a newly developed Sediment-Oil-Flow-Through (SOFT) system. Distinct redox zones established and migrated vertically in the core during the 190 days-long simulated petroleum seepage. Methanogenic petroleum degradation was indicated by an increase in methane concentration from 8 μM in an untreated core compared to 2300 μM in the lower sulfate-free zone of the SOFT core at the end of the experiment, accompanied by a respective decrease in the δ13C signal of methane from -33.7 to -49.5‰. The involvement of methanogens in petroleum degradation was further confirmed by methane production in enrichment cultures from SOFT sediment after the addition of hexadecane, methylnapthalene, toluene, and ethylbenzene. Petroleum degradation coupled to sulfate reduction was indicated by the increase of integrated sulfate reduction rates from 2.8 SO42-m-2 day-1 in untreated cores to 5.7 mmol SO42-m-2 day-1 in the SOFT core at the end of the experiment, accompanied by a respective accumulation of sulfide from 30 to 447 μM. Volatile hydrocarbons (C2–C6 n-alkanes) passed through the methanogenic zone mostly unchanged and were depleted within the sulfate-reducing zone. The amount of heavier n-alkanes (C10–C38) decreased step-wise toward the top of the sediment core and a preferential degradation of shorter (<C14) and longer chain n-alkanes (>C30) was seen during the seepage. This study illustrates, to the best of our knowledge, for the first time the development of methanogenic petroleum degradation and the succession of benthic microbial processes during petroleum passage in a whole round sediment core.

Research of Isolation and Degradation Conditions of Petroleum Degrading Marine

A novel petroleum-degrading microbial strain was isolated from sediment samples in estuary of Bohai Sea estuary beaches. The strain was primarily identified as Alcanivorax sp. and named Alcanivorax sp. H34. Effect of PH values, temperature, nitrogen and phosphorus concentrations on degradation of H34 were investigated. The paraffinic components average degradation rate of H34 ungrowth cells under optimized conditions was studied. The results showed that the optimal growth conditions of H34 are were temperature of 30°C, initial PH of 7.0, nitrogen concentration of 3g/L, phosphorus concentration of 3g/L, and paraffinic components average degradation rates of H34 ungrowth cells was 41.6%, while total degradation rate was 45.5%.

Soil and Water Microbiological Characteristics of Olo Flowstation and Rumuekpe Metering Station, Rivers State, Nigeria

This study evaluates the soil and microbiological properties of Olo Flowstation and Rumuekpe Metering Station. Standard sampling and analytical methods were adopted. Results of the soil microbiological composite samples from the random samples showed that the percent Hydrocarbon Utilizing Bacteria (HUB) was 40.0% and 27.27% for the top and subsoil, respectively. Though these values were high, they were below expected values in view of the high level of petroleum hydrocarbon spilled on the soils collected. The moderately high amount of hydrocarbon degraders in these soils compared to the control indicated that these soils might have been in contact with petroleum products which might have polluted the soils. The poor drainage resulting from stagnant water might have leached the petroleum products down the soil along transect, resulting into higher HUB% in the subsoil than the top soil. Other sampling stations had better top soil drainage hence lower % HUB down the profile. All the water samples, both surface and groundwater showed significant range of bacteria. The colonies were within 30 to 300. The growth for bacteria ranges from 1.5 × 105 cfu/ml at Elele-Alimini (up-stream) to 3.0 × 105 cfu/ml at the waste pit, Sombreiro River and flare pit. The heterotrophic organisms isolated were Bacillins sp., Chromobacterium sp., Micrococcus sp, Enterobacter sp., and Psendomonas sp. The presence of the bacteria in the water bodies (effluent) indicated a regrowth and general bacterial composition of the water. The petroleum degraders isolated in the surface waters where Chromobacterium sp., Micrococcus sp., Psendomonas sp. and Bacillus sp. The microorganisms population range from 1.0 × 102 at Elele-Alimini down-stream to 6.0 × 102 at discharge point-Waste pit and the borrow pit at Rumuekpe. The preponderance of petroleum utilizing bacteria in these water samples indicated the presence of hydrocarbons. The fungal count was generally low and was found in Elele-Alimini up-stream and borrow pit near the metering station at Rumuekpe. The mean count of E. coli per 100 ml of water was 180+ meaning that all the water samples both surface and groundwater were polluted. The presence of E. coli showed faecal pollution of the water and there could be possible presence of pathogenic organisms in the water. This study therefore will be a useful guide to farmers in the areas around the flow stations. There is also need for treatment of surface and groundwater in the area. It is recommended that regular monitoring of the geo-environmental media be carried out.

Reclamation of Smaller Volumes of Petroleum Hydrocarbon Contaminated Soil Using an Innovative Reactor System: A Case Study Evaluation of the Design

Petroleum products contamination is a world-wide problem that threatens polluting groundwater and surface water systems. However, the problem is not only large-scale in scope when viewed from a case-by-case basis. Many fueling, construction, agricultural, and industrial activities result in the problem of managing smaller quantities of these soils from an ecological safety perspective. Landfilling has been the disposal method of choice in the US; however, this option is becoming economically prohibitive and it does not really offer a true degradation fate for the pollutants. This study focused on the proving of an innovative biocell design that afforded a high level of petroleum degradation within a simple and cost effective design. Additionally, the design offered a remediation solution for sites not easily accessed. Soil contaminated with both diesel fuel and gasoline collected from a former filling station was used in this on-site remediation case study. Rapid biodegradation of the petroleum products were observed at the initiation of the study with rates leveling off as the study progressed with the final total petroleum hydrocarbon concentration being 10 mg/kg at Day 90. Oxygen uptake rates were monitored and found to nicely track both microbial activity and pollutant removal dynamics. The biocell design met all expectations by being effective, yet simple to build and operate.

Copper effect in petroleum hydrocarbons biodegradation by microorganisms associated to Juncus maritimus: role of autochthonous bioaugmentation

The bioremediation potential of microorganisms from a saltmarsh plant rhizosphere and application of bioaugmentation in estuarine sediment co-contaminated were investigated. Rhizosediment (sediment in contact with plant roots) of Juncus maritimus was contaminated with copper and/or petroleum, inoculated with different autochthonous microbial consortia (resistant to copper and/or with petroleum degraders) and put in vessels to which plants were transplanted. Vessels were irrigated through a system that simulated estuarine tides. After 5 months, vessels were dismantled and copper and petroleum content in rhizosediments were determined. Copper’s presence reduced the potential of the microorganisms associated to J. maritimus rhizosphere for bioremediation of petroleum hydrocarbons in co-contaminated sediment. Indeed, hydrocarbons removal decreased from 39 to 25% when copper was present. In addition, bioaugmentation was not effective to overcome metal negative effects on petroleum hydrocarbons degradation, and the same removal rate was being observed (ca. 25%). Different methodologies for the formulation of consortia must be tested in this situation of co-contamination. Obtained results should be taken in consideration when planning the recovery of moderately impacted estuaries, aiming an effective protection and management of these areas, in the case of co-contamination.

Transformation of Polycyclic Aromatic Hydrocarbons at Degradation of Petroleum in Soil

Transformation of Polycyclic Aromatic Hydrocarbons at Degradation of Petroleum in Soil

Growth of four tropical tree species in petroleum-contaminated soil and effects of crude oil contamination.

Under greenhouse conditions, we evaluated establishment of four tree species and their capacity to degrade crude oil recently incorporated into the soil; the species were as follows: Cedrela odorata (tropical cedar), Haematoxylum campechianum (tinto bush), Swietenia macrophylla (mahogany), and Tabebuia rosea (macuilis). Three-month-old plants were planted in soil with three treatments of heavy petroleum and a control (C0 0mgkg-1; C1 18,000mgkg-1; C2 31,700mgkg-1; C3 47,100mgkg-1) with four repetitions per treatment and species; the experiment was carried out for 245days. Height and biomass of all species significantly diminished as petroleum concentration increased, although plant survival was not affected. The quantity of colony-forming units (CFU) of rhizospheric bacteria varied among tree species and treatments; petroleum stimulated bacterial CFU for S. macrophylla. The number of fungi CFU for S. macrophylla and T. rosea was significantly greater in C0 than in soil with petroleum, but among species and among different concentrations, no significant differences were found. The greatest percentage of total petroleum hydrocarbon (TPH) degradation was found in C1 for soil without plants (45%). Differences from the remaining treatments (petroleum concentrations in soil and plant species) were not significant (P<0.05). Among all trees, H. campechianum had the greatest TPH degradation (32.5% in C2). T. rosea (C1) and H. campechianum (C2) resulted in petroleum degradation at levels ranging from 20.5 to 32.5%. On the basis of this experiment, the tree species used did not improve TPH degradation. However, all of them showed high rates of survival and vigor. So, as tree species provide goods and services, experiments with inoculation of hydrocarbonclastic microorganisms, addition of fertilizers, and mixture of tree and grasses are recommended.

Responses of hydrochemical parameters, community structures, and microbial activities to the natural biodegradation of petroleum hydrocarbons in a groundwater–soil environment

Responses of hydrochemical parameters, bacterial community structures, and microbial activities during the natural biodegradation of hydrocarbons leaking from a crude oil pipeline at a site in northeastern China were studied using the hydrochemical method, the polymerase chain reaction amplification of 16S rDNA genes, and denaturing gradient gel electrophoresis (DGGE).The concentrations of PHs and the main physicochemical characteristics (Eh and pH) were increased gradually at longitudinal ZK1–ZK3–ZK6 section in June 2010, suggesting that the degree of biodegradation decreased gradually from the pollutant source to downstream. The electron acceptor (SO42−) participated in biodegradation process responds identical trend to the degree of biodegradation at the same section while the biodegradation reduction products (Fe2+, Mn2+, and HCO3−) respond to it in opposite trend. The concentrations of Fe2+, Mn2+, and SO42− at wells ZK1 and ZK3 indicate that sulfate-reducing biodegradation was the dominant process experienced at contaminated sites rather than iron–manganese reduction processes. The dynamic changes of hydrochemical parameters suggest that the degree of biodegradation would be intensified at the center of pollutant plume and iron–manganese reduction processes experienced earlier than sulfate-reducing biodegradation. The community diversities and microbial activities in different soil depths were responded to PHs, but the relationship is not simply linear. The community diversities and microbial activities increased significantly with rise in PH concentration of soil until about 20 mg/kg and then peaked off sharply, suggesting that community succession would be occurred in microhabitat when the PH concentration in soil exceeds 20 mg/kg. The major DGGE bands found in the soil samples were sequenced. One sequence affiliated with γ-Proteobacteria showed high similarities to uncultured bacteria which had been found in offshore oil pollution environment. These bacterial species were found to have similar petroleum degradation functions to the bacteria at the sites studied here.

Functional and genetic characterization of hydrocarbon biodegrader and exopolymer-producing clones from a petroleum reservoir metagenomic library

ABSTRACTMicrobial degradation of petroleum is a worldwide issue, which causes physico-chemical changes in its compounds, diminishing its commercial value. Biosurfactants are chemically diverse molecules that can be produced by several microorganisms and can enable microbial access to hydrocarbons. In order to investigate both microbial activities, function-driven screening assays for biosurfactant production and hydrocarbon biodegradation were carried out from a metagenomic fosmid library. It was constructed from the total DNA extracted from aerobic and anaerobic enrichments from a Brazilian biodegraded petroleum sample. A sum of 10 clones were selected in order to evaluate their ability to produce exopolymers (EPS) with emulsifying activity, as well as to characterize the gene sequences, harbored by the fosmid clones, through 454 pyrosequencing. Functional analyses confirmed the ability of some clones to produce surfactant compounds. Regarding hydrocarbon as microbial carbon sources, n-alkane (in mixture or not) and naphthalene were preferentially consumed as substrates. Analysis of sequence data set revealed the presence of genes related to xenobiotics biodegradation and carbohydrate metabolism. These data were corroborated by the results of hydrocarbon biodegradation and biosurfactant production detected in the evaluated clones.