Successful phytoremediation of crude-oil contaminated soil at an oil exploration and production company by plants-bacterial synergism

Phytoremediation is a promising approach for the cleanup of soil contaminated with petroleum hydrocarbons. This study aimed to develop plant-bacterial synergism for the successful remediation of crude oil-contaminated soil. A consortia of three endophytic bacteria was augmented to two grasses, Leptochloa fusca and Brachiaria mutica, grown in oil-contaminated soil (46.8 g oil kg^?1 soil) in the vicinity of an oil exploration and production company. Endophytes augmentation improved plant growth, crude oil degradation, and soil health. Maximum oil degradation (80%) was achieved with B. mutica plants augmented with the endophytes and it was significantly (P < 0.05) higher than the use of plants or bacteria individually. Moreover, endophytes showed more persistence, the abundance and expression of alkB gene in the rhizosphere as well as in the endosphere of the tested plants than in unvegetated soil. A positive relationship (r = 0.70) observed between gene expression and crude oil reduction indicates that catabolic gene expression is important for hydrocarbon mineralization. This investigation showed that the use of endophytes with appropriate plant is an effective strategy for the cleanup of oil-contaminated soil under field conditions.     

Relating Hybrid Poplar Fine Root Production,Soil Nutrients,and Hydrocarbon Contamination

Quantifying the effects of hydrocarbon contamination on hybrid poplar fine root dynamics provides information about how well these trees tolerate the adverse conditions imposed by the presence of petroleum in the soil. The objective of this research was to investigate the relationship between the varying concentrations of total petroleum hydrocarbons (TPH) and nutrients across a hydrocarbon-contaminated site, and quantify the effects of these properties on the spatial and temporal patterns of fine root production of hybrid poplar (P. deltoides × P. petrowskyana C. V. Griffin) Twelve minirhizotron tubes were distributed across a TPH-contaminated site at Hendon, SK, Canada, and facilitated quantification of fine root production in areas of varying contamination levels. Residual hydrocarbon contamination was positively correlated with soil total C and N, which may suggest that the hydrocarbons remaining in the soil are associated with organic forms of these nutrients. Fine root production was stimulated by small amounts of hydrocarbon contamination at the field site. Nonlinear regression described fine root production as increasing linearly up to approximately 500 mg kg^{? 1} TPH, then remaining constant as contamination increased. Stimulation of hybrid poplar fine root production in hydrocarbon-contaminated soil could to lead to enhanced contaminant degradation as a result of stimulated microbial activity via a greater rhizosphere effect.     

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.     

Bioremediation of oil-polluted soils: Using the [13C]/[12C] ratio to characterize microbial products of oil hydrocarbon biodegradation

We compared data on the extent of bioremediation in soils polluted with oil. The data were obtained using conventional methods of hydrocarbon determination: extraction gas chromatography-mass spectrometry, extraction IR spectroscopy, and extraction gravimetry. Due to differences in the relative abundances of the stable carbon isotopes (¹³C/¹²C) in oil and in soil organic matter, these ratios could be used as natural isotopic labels of either substance. Extraction gravimetry in combination with characteristics of the carbon isotope composition of organic products in the soil before and after bioremediation was shown to be the most informative approach to an evaluation of soil bioremediation. At present, it is the only method enabling quantification of the total petroleum hydrocarbons in oil-polluted soil, as well as of the amounts of hydrocarbons remaining after bioremediation and those microbially transformed into organic products and biomass.     

Biodegradation of Petroleum Hydrocarbons in a Soil Polluted Sample by Oil-Based Drilling Cuttings

Microbial degradation of hydrocarbons in soils polluted by oil-based drilling mud and cuttings has been investigated by static methods such as composting or biopiling. Bioremediation of polluted soils by oil-based drilling cuttings through a slurry bioreactor has not previously been reported. The main aim of this work is to monitor hydrocarbon biodegradation in slurry of drilling cuttings and unpolluted soils and the effects of nutrients on it. Indigenous, bacterial-mixed culture isolated from a polluted soil by drilling cuttings adapted to drilling mud concentrations up to 15% (v/v) was done during a 15-month program. The total petroleum hydrocarbons’ (TPHs) removal efficiency in C/N/P 100/5/1 ratio was 90.5 and 79.85% under experimental and control conditions, respectively. The microbial count on the first day, 15 × 10⁷ CFUg^?1, reached 20 × 10⁹ CFUg^?1on the twenty-first day at experimental conditions. The TPH removal efficiency in C/N/P 100/10/2 was 92.5 and 82.25% at experiment and control, respectively. Increasing nitrogen and phosphorous amount couldn't increase microbial count in comparison with C/N/P ratio 100/5/1. The measured biomass contents and microbial counts in experiments were significantly higher than the control and confirmed hydrocarbons’ biodegradation during the time. Results showed that slurry bioreactors could accelerate the biodegradation of TPHs and reduce remediation time in soil polluted by oil-based drilling cuttings.     

Enhancement of Hydrocarbon Removal in a Clay and Drilling-Waste Polluted Soil

The application of biological processes in restoring oil polluted sites is growing due to their efficiency in removing different classes of pollutants. The aim of this study was to determine the ability of microorganisms present in a drilling-waste polluted soil (36,200 mg TPH kg^?1 soil) to remove weathered hydrocarbons under stimulated and non-stimulated soil conditions. The hypothesis under study was whether petroleum hydrocarbons removal could be enhanced by manipulating C/N ratio, water content and addition of three agroindustrial wastes. A Box-Behnken design was employed to evaluate the effect of each variable. Results demonstrated that, for orange peels and banana trunk treatments, the variable with the largest effect (p < 0.01) on hydrocarbon removal was the C/N ratio, indicating that higher ratio (100/3) improved removal (79.5–82%). The largest effect (p < 0.001) on hydrocarbon removal for pineapple wastes was observed with higher water content (60%) achieving the highest removal (89%). After 90 days of experimentation, the type of agricultural waste and the agricultural waste/soil ratio were not statistically significant in any treatment. However, their addition was important relative to non-stimulated soil, which showed a hydrocarbon removal of 17%. Data reported in this study showed the application of bioremediation in clay and drilling waste-polluted soils.     

Optimization of <Emphasis Type="Italic">Enterobacter cloacae</Emphasis> (KU923381) for diesel oil degradation using response surface methodology (RSM)

Efficiency of Enterobacter cloacae KU923381 isolated from petroleum hydrocarbon contaminated soil was evaluated in batch culture and bioreactor mode. The isolate were screened for biofilm formation using qualitative and quantitative assays. Response surface methodology (RSM) was used to study the effect of pH, temperature, glucose concentration, and sodium chloride on diesel degradation. The predicted values for diesel oil degradation efficiency by the statistical designs are in a close agreement with experimental data (R ² = 99.66%). Degradation efficiency is increased by 36.78% at pH = 7, temperature = 35°C, glucose = 5%, and sodium chloride concentration = 5%. Under the optimized conditions, the experiments were performed for diesel oil degradation by gas chromatographic mass spectrometric analysis (GC-MS). GC-MS analysis confirmed that E. cloacae had highly degrade hexadecane, heptadecane, tridecane, and docosane by 99.71%, 99.23%, 99.66%, and 98.34% respectively. This study shows that rapid bioremoval of hydrocarbons in diesel oil is acheived by E. cloacae with abet of biofilm formation. The potential use of the biofilms for preparing trickling filters (gravel particles) for the degradation of hydrocarbons from petroleum wastes before their disposal in the open environment is highly suggested. This is the first successful attempt for artificially establishing petroleum hydrocarbon degrading bacterial biofilm on solid substrates in bioreactor.     

Petroleum bioremediation — a multiphase problem

Microbial degradation of hydrocarbons is a multiphase reaction, involving oxygen gas, water-insoluble hydrocarbons, water, dissolved salts and microorganisms. The fact that the first step in hydrocarbon catabolism involves a membrane-bound oxygenase makes it essential for microorganisms to come into direct contact with the hydrocarbon substrate. Growth then proceeds on the hydrocarbon/water interface. Bacteria have developed two general strategies for enhancing contact with water-insoluble hydrocarbons: specific adhesion mechanisms and production of extracellular emulsifying agents. Since petroleum is a complex mixture of many different classes of hydrocarbons, of which any particular microorganism has the potential to degrade only part, it follows that the microorganisms must also have a mechanism for desorbing from used' oil droplets.The major limitations in bioremediation of hydrocarbon-contaminated water and soil is available sources of nitrogen and phosphorus. The usual sources of these materials, e.g. ammonium sulfate and phosphate salts, have a high water solubility which reduces their effectiveness in open systems because of rapid dilution. We have attempted to overcome this problem by the use of a new controlled-release, hydrophobic fertilizer, F-1, which is a modified urea-formaldehyde polymer containing 18% N and 10% P as P₂O₅. Microorganisms were obtained by enrichment culture that could grow on crude oil as the carbon and energy source and F-1 as the nitrogen and phosphorus source. The microorganisms and the F-1 adhered to the oil/water interface, as observed microscopically and by the fact that degradation proceeded even when the water phase was removed and replaced seven times with unsupplemented water — a simulated open system. Strains which can use F-1 contain a cell-bound, inducible enzyme which depolymerizes F-1.After optimizing conditions in the laboratory for the use of F-1 and the selected bacteria for degrading crude oil, a field trial was performed on an oil contaminated sandy beach between Haifa and Acre, Israel, in the summer of 1992. The sand was treated with 5 g F-1 per kg sand and inoculated with the selected bacteria; the plot was watered with sea water and plowed daily. After 28 days the average hydrocarbon content of the sand decreased from 5.1 mg per g sand to 0.6 mg per g sand. Overall, there was an approx. 86% degradation of pentane extractables as demonstrated by dry weight, I.R. and GLC analyses. An untreated control plot showed only a 15% decrease in hydrocarbons. During the winter of 1992, the entire beach (approx. 200 tons of crude oil) was cleaned using the F-1 bacteria technology. The rate of degradation was 0.06 mg g^-1 sand day^-1 (10°C) compared to 0.13 mg g^-1 sand day^-1 during the summer (25°C).     

The impact of petroleum hydrocarbons (diesel) on periphyton in an impacted tropical estuary based on in situ microcosms

The distribution of petroleum hydrocarbons and their effects on the periphytic algal biomass using in situ microcosms were investigated in Ponggol estuary located on the northeastern coast of Singapore. Dissolved or dispersed petroleum hydrocarbon (DDPH) concentrations in the surface and bottom waters and absorbed or adsorbed petroleum hydrocarbon (AAPH) concentrations in sediments were monitored from July 1999 to June 2000. Results showed concentrations ranging from 4.42 to 248.94 μg l⁻¹, from 0.35 to 1099.65 μg l⁻¹, and from 20.55 to 541.01 mg kg⁻¹ for DDPH in surface and bottom waters and AAPH in sediments, respectively. Accidental spillages of fuel from dredgers operating in the estuary, fuel and engine oil from recreational boats, shipping operations in the adjacent strait, and runoff monsoon drains in the vicinity were some of the possible sources of petroleum hydrocarbons in the estuary. An assessment of environmentally realistic concentrations of petroleum hydrocarbons on periphytic algal biomass using in situ microcosms revealed signs of acute toxicity. A reduction in periphytic algal biomass (with respect to controls) of 68-93% was observed for various treatments exposed to diesel.     

Uptake modes of octadecane by Pseudomonas sp. DG17 and synthesis of biosurfactant

Aims: In order to gain more insight into the uptake modes of octadecane by bacteria. Methods and Results: A strain that could utilize octadecane well was isolated from crude oil contaminated soil, and named as Pseudomonas sp. DG17 by 16S rDNA analysis. Culture growth result showed that Pseudomonas sp. DG17 grew well in the addition of 200 and 400 mg l^?1 of octadecane, which showed that physical contact between substrate and bacteria was important in the substrate biodegradation. Meanwhile, Pseudomonas sp. DG17 produced rhamnolipids biosurfactant that contains 10 congeners, thus causing the surface tension of the culture medium decline and facilitating the contact between hydrocarbon and bacteria. Scanning‐electron‐microscopy results showed that a disruption of the surface membranes in certain zones was observed in some of the cells grown in 400 mg l^?1 octadecane at 176 h compared with the cells in exponential phase at 72 h due to the production of biosurfactant‐rhamnolipid. Conclusions: These results indicated the possibility that the direct contact with insoluble octadecane droplets occurred before the contact with pseudosolubilization smaller oil droplets. Significance: This report throws more light on the uptake mechanisms of octadecane by bacteria, and proposes the possibility that role of biosurfactant is to increase the contact between hydrocarbon and bacteria by changing the cell membrane structure which needs studied in depth. Impact of Study: Results of this study are useful in the bioremediation of petroleum polluted soil.     

Contaminated Soil Phytoremediation by Cyperus Laxus Lam. Cytochrome P450 Erod-Activity Induced by Hydrocarbonsin Roots

Laboratory and greenhouse experiments with Cyperus laxus Lam were conducted to determine the rate and extent of phytoremediation and the effect of hydrocarbons on the cytochrome P450 EROD (7-ethoxyresorufin-O-deethylase) enzymatic activity in roots. Plants were cultivated on hydrocarbon-contaminated soil (HCS) and spiked perlite. Phytoremediation was evaluated using 6.5 kg HCS (173 ± 15 mg total petroleum hydrocarbons [TPH] g^?1 of dry soil) pots at different moisture contents; the average removal rate was 3.46 ± 0.25 mg TPH g^?1 dry soil month^?1 and 48% was removed when moisture was kept at 60%. The aromatic hydrocarbon fraction was the mostly removed, 60%; aliphatic, 51%; and polar 24% after 24-month experiments. In unplanted pots, TPH concentration did not exhibit significant differences with respect to the initial concentration. We confirmed that the presence of hydrocarbons induced ERODactivity up to 6.5-fold. Moreover, short-term experiments (up to 13 d) with spiked perlite demonstrated that two EROD activities in roots contributed to the total detected; 60% was found in the cytosolic and 40% in the microsomal fraction. To our knowledge, this is the first work that tries to build links between the hydrocarbon-inducible character of ERODactivity in roots and the phytoremediation ability of C. laxus in highly contaminated soils.     

The role of microbial populations in the containment of aromatic hydrocarbons in the subsurface

A survey of soil gases associated with gasoline stations on theSwan Coastal Plain of Western Australia has shown that 20% leak detectable amountsof petroleum. The fates of volatile hydrocarbons in the vadose zone at one contaminatedsite, and dissolved hydrocarbons in groundwater at another site were followed in anumber of studies which are herein reviewed. Geochemical evidence from a plume ofhydrocarbon-contaminated groundwater has shown that sulfate reduction rapidly developedas the terminal electron accepting process. Toluene degradation but not benzene degradationwas linked to sulfate reduction. The sulfate-reducing bacteria isolated from the plumerepresented a new species, Desulfosporosinus meridiei. Strains of the speciesdo not mineralise ¹⁴C-toluene in pure culture. The addition of large numbersof cells and sulfate to microcosms did stimulate toluene mineralisation but not benzenemineralisation. Attempts to follow populations of sulfate-reducing bacteria byphospholipid signatures, or Desulfosporosinus meridiei by FISH in the plume were unsuccessful, but fluorescently-labeled polyclonal antibodies were successfully used.In the vadose zone at a different site, volatile hydrocarbons were consumed in thetop 0.5 m of the soil profile. The fastest measured rate of mineralisation of ¹⁴C-benzenein soils collected from the most active zone (6.5 mg kg^-1 day^-1) could accountfor the majority of the flux of hydrocarbon vapour towards the surface. The studiesconcluded that intrinsic remediation by subsurface microbial populations in groundwateron the Swan Coastal Plain can control transport of aromatic hydrocarbon contamination,except for the transport of benzene in groundwater. In the vadose zone, intrinsicremediation by the microbial populations in the soil profile can contain the transportof aromatic hydrocarbons, provided the physical transport of gases, inparticular oxygen from the atmosphere, is not impeded by structures.     

TECHNICAL NOTE: NITROGEN FERTILIZATION EFFECTS ON THE DEGRADATION OF AGED DIESEL OIL IN COMPOSTED DRILLING WASTES

Hydrocarbon-contaminated wastes generated from oil and gas drilling activities may be used as a soil amendment once composted and further decomposition of residual hydrocarbons can be accomplished after the composts are applied to soils. To test if N fertilization may enhance hydrocarbon decomposition, we investigated the effects of N application on hydrocarbon degradation in different-aged composts (1-, 2-, 3-, and 4-year-old composts, coded as 1Y, 2Y, 3Y, and 4Y composts, respectively) through a pot experiment planted with white spruce (Picea glauca [Moench] Voss) seedlings. The percentage degradation of total petroleum hydrocarbon (TPH, C11 to C40) in the composts without N fertilization was correlated to initial NH₄ ⁺ concentrations (R = 0.99, P < 0.001). The percentage degradation of TPH was highest in the 3Y compost (41.1%) that had an initial level of 325.3 mg NH₄ ⁺-N kg^?1 and the lowest in the 1Y compost (9.3%) that had an initial level of 8.3 mg NH₄ ⁺-N kg^?1. The degradation of TPH was enhanced by N fertilization in the 1Y (from 9.3 to 15.3%) and 4Y composts (from 14.3 to 22.6%) that had low initial NH₄ ⁺ concentrations. Our results show that application of NH₄ ⁺-based fertilizers may enhance the degradation of TPH when initial NH₄ ⁺ concentrations in the compost are low.     

Phytoremediation of Light Crude Oil by Maize (Zea mays L.) Bio-Augmented with Plant Growth Promoting Bacteria

The aim of this study was to evaluate the converged effect of maize and plant growth promoting bacteria on degradation of petroleum hydrocarbons under axenic conditions. Artificially spiked sand with 10 g kg^?1 light crude oil was planted with maize alone and in combination with eight bacterial isolates having plant growth promotion and bioremediation potential to observe the dissipation of petroleum hydrocarbons. Results showed remarkable suppression of maize growth and biomass production due to phytotoxicity of the crude oil contamination. However, bio-augmentation of plants with bacteria having ACC-deaminase activity significantly compensated the reduction in plant growth compared to uninoculated plants. The results revealed that plants bio-augmented with PM32Y exhibited significant increase in root length (75%), plant height (74%), and biomass (67%) as compared to uninoculated plants after 60 days of planting. The same bacterium in convergence with maize caused 43% degradation of petroleum hydrocarbons as compared to the unplanted and uninoculated control. Amplification, sequencing and phylogenetic analysis of 16S rRNA gene sequence identified PM32Y bacterium as Bacillus subtilis strain. It is concluded that bio-augmentation of plants with plant growth promoting bacteria having bioremediation potential and ACC-deaminase activity can successfully be used in phytoremediation of petroleum hydrocarbons.     

Effects of diurnal temperature variation on microbial community and petroleum hydrocarbon biodegradation in contaminated soils from a sub‐Arctic site

Contaminated soils are subject to diurnal and seasonal temperature variations during on‐site ex‐situ bioremediation processes. We assessed how diurnal temperature variations similar to that in summer at the site from which petroleum hydrocarbon‐contaminated soil was collected affect the soil microbial community and the extent of biodegradation of petroleum hydrocarbons compared with constant temperature regimes. Microbial community analyses for 16S rRNA and alkB genes by pyrosequencing indicated that the microbial community for soils incubated under diurnal temperature variation from 5°C to 15°C (VART5‐15) evolved similarly to that for soils incubated at constant temperature of 15°C (CST15). In contrast, under a constant temperature of 5°C (CST5), the community evolved significantly different. The extent of biodegradation of C10–C16 hydrocarbons in the VART5‐15 systems was 48%, comparable with the 41% biodegradation in CST15 systems, but significantly higher than CST5 systems at 11%. The enrichment of Gammaproteobacteria was observed in the alkB gene‐harbouring communities in VART5‐15 and CST15 but not in CST5 systems. However, the Actinobacteria was abundant at all temperature regimes. The results suggest that changes in microbial community composition as a result of diurnal temperature variations can significantly influence petroleum hydrocarbon bioremediation performance in cold regions.     

Bioremediation of experimental petroleum spills on mineral soils in the Vestfold Hills,Antarctica

The effect of nutrient and water enhancement on the biodegradation of petroleum was tested in Antarctic mineral soils. Nitrogen, phosphorus and potassium were applied in solution, with or without gum xanthan or plastic covers, to sites artificially contaminated with distillate. The effectiveness of these procedures was assessed by measuring changes in total petroleum hydrocarbons; heptadecane/pristane and octadecane/phytane ratios; in concentrations of major hydrocarbon components and in microbial numbers and activity.Significantly lower hydrocarbon concentrations were recorded after one year in soils treated with fertilizer solutions, but only in the surface 3 cm. These soils also showed lowered heptadecane/pristane and octadecane/ phytane ratios and had the highest levels of microbial activity relative to other plots. Soils treated with gum xanthan. or covered with plastic had the highest residual hydrocarbon levels. Both treatments inhibited evaporative loss of hydrocarbon, and there were indications that gum xanthan was utilized by the microbiota as an alternative carbon source to distillate. Higher temperatures were recordecd under the plastic but no stimulation of biodegradation was detected.Estimated numbers of metabolically active bacteria were in the range 10⁷ to 10⁸ g^–1 dry weight of soil, with an estimated biomass of 0.03 to 0.26 mg g^–1 soil. Estimated numbers of amoebae were in the range 10⁶ to 10⁷ g^–1 soil (biomass of 2 to 4 mgg^–1). The highest populations were recorded in fertilized, contaminated soils, the only soils where petroleum degradation was demonstrated.     

Remarkable impact of PAHs and TPHs on the richness and diversity of bacterial species in surface soils exposed to long-term hydrocarbon pollution

Nowadays, because of substantial use of petroleum-derived fuels the number and extension of hydrocarbon polluted terrestrial ecosystems is in growth worldwide. In remediation of aforementioned sites bioremediation still tends to be an innovative, environmentally attractive technology. Although huge amount of information is available concerning the hydrocarbon degradation potential of cultivable hydrocarbonoclastic bacteria little is known about the in situ long-term effects of petroleum derived compounds on the structure of soil microbiota. Therefore, in this study our aim was to determine the long-term impact of total petroleum hydrocarbons (TPHs), volatile petroleum hydrocarbons (VPHs), total alkyl benzenes (TABs) as well as of polycyclic aromatic hydrocarbons (PAHs) on the structure of bacterial communities of four different contaminated soil samples. Our results indicated that a very high amount of TPH affected positively the diversity of hydrocarbonoclastic bacteria. This finding was supported by the occurrence of representatives of the α-, β-, γ-Proteobacteria, Actinobacteria, Flavobacteriia and Bacilli classes. High concentration of VPHs and TABs contributed to the predominance of actinobacterial isolates. In PAH impacted samples the concentration of PAHs negatively correlated with the diversity of bacterial species. Heavily PAH polluted soil samples were mainly inhabited by the representatives of the β-, γ-Proteobacteria (overwhelming dominance of Pseudomonas sp.) and Actinobacteria.     

Mobility and Persistence of Petroleum Hydrocarbons in Peat Soils of Southeastern Mexico

Spilled crude petroleum from oil wells contains numerous hydrocarbons, some of which are toxic and threaten life. We have studied the mobility and persistence of hydrocarbons in waterlogged soils that contain large proportions of fermented organic matter (Histosols) and large concentrations of dissolved organic carbon (DOC) in the State of Tabasco, Mexico. We sampled soil and phreatic water at sites polluted by oil spills for several decades, as well as at sites that had only recently (few weeks) been polluted, and compared their hydrocarbon contents with those of unaffected sites in the same area. Samples were analyzed for 16 non-alkylated polyaromatic hydrocarbons (PAHs) and n-alkanes from nC9 to nC34. The spilled hydrocarbons had remained predominantly in the organic surface horizons of the soil where spillage occurred; there was little evidence of movement within the soil. The fraction of low molecular weight compounds was larger at sites of recent spills than where spills happened several decades ago. Nevertheless, sites of old spills still contained large concentrations of hydrocarbons, among which those of low molecular weight represented from 30 to 49% of total PAHs and from 50 to 84% of total n-alkanes, indicating that volatilization or microbial degradation is slow in these soils. In the peat horizons the measured organic carbon partition coefficients (K _oc ) for the higher molecular weight PAHs were consistently smaller than those estimated by empirical equations by up to two orders of magnitude. The dissolved organic carbon of these peat soils seems to influence this behavior. At sites of old spills, partition coefficients for the PAHs were larger than at sites of recent spills.     

Enhancing the biodegradation of total petroleum hydrocarbons in oily sludge by a modified bioaugmentation strategy

The effect of successive inoculation with hydrocarbon-degrading bacteria on the dynamics of petroleum hydrocarbons degradation in soil was investigated in this study. Oily sludge was used as a source of mixed hydrocarbons pollutant. Two bacterial consortia composed of alkanes and polycyclic aromatic hydrocarbon degraders were constructed from bacteria isolated from soil and oily sludge. These consortia were applied to incubated microcosms either in one dose at the onset of the incubation or in two doses at the beginning and at day 62 of the incubation period, which lasted for 198 days. During this period, carbon mineralization was evaluated by respirometry while total petroleum hydrocarbons and its fractions were gravimetrically evaluated by extraction from soil and fractionation. Dosing the bacterial consortia resulted in more than 30% increase in the overall removal of total petroleum hydrocarbons from soil. While alkane removal was only slightly improved, aromatic and asphaltic hydrocarbon fraction removal was significantly enhanced by the addition of the second consortium. Polar compounds (resins) were enriched only as a result of aromatics and asphaltene utilization. Nonetheless, their concentration declined back to the original level by the end of the incubation period.     

Stimulation of Diesel Fuel Biodegradation by Indigenous Nitrogen Fixing Bacterial Consortia

Abstract Successful stimulation of N₂ fixation and petroleum hydrocarbon degradation in indigenous microbial consortia may decrease exogenous N requirements and reduce environmental impacts of bioremediation following petroleum pollution. This study explored the biodegradation of petroleum pollution by indigenous N₂ fixing marine microbial consortia. Particulate organic carbon (POC) in the form of ground, sterile corn-slash (post-harvest leaves and stems) was added to diesel fuel amended coastal water samples to stimulate biodegradation of petroleum hydrocarbons by native microorganisms capable of supplying a portion of their own N. It was hypothesized that addition of POC to petroleum amended water samples from N-limited coastal waters would promote the growth of N₂ fixing consortia and enhance biodegradation of petroleum. Manipulative experiments were conducted using samples from coastal waters (marinas and less polluted control site) to determine the effects of POC amendment on biodegradation of petroleum pollution by native microbial consortia. Structure and function of the microbial consortia were determined by measurement of N₂ fixation (acetylene reduction), hydrocarbon biodegradation (¹⁴C hexadecane mineralization), bacterial biomass (AODC), number of hydrocarbon degrading bacteria (MPN), and bacterial productivity (³H-thymidine incorporation). Throughout this study there was a consistent enhancement of petroleum hydrocarbon degradation in response to the addition of POC. Stimulation of diesel fuel biodegradation following the addition of POC was likely attributable to increases in bacterial N₂ fixation, diesel fuel bioavailability, bacterial biomass, and metabolic activity. Toxicity of the bulk phase water did not appear to be a factor affecting biodegradation of diesel fuel following POC addition. These results indicate that the addition of POC to diesel-fuel-polluted systems stimulated indigenous N₂ fixing microbial consortia to degrade petroleum hydrocarbons. Received: 29 December 1998; Accepted: 6 April 1999