Bioremediation Of Crude Oil Polluted Soil Research Articles

Use of Cassava Peel as Biostimulant in Bioremediation of Crude Oil-Polluted Soil

Use of Cassava Peel as Biostimulant in Bioremediation of Crude Oil-Polluted Soil

Remediation of petroleum hydrocarbon contaminated soil using hydrocarbonoclastic rhizobacteria, applied through Azadirachta indica rhizosphere

Crude oil/petroleum hydrocarbons (PHs) are major pollutants worldwide. In the present study, three bacterial isolates -Pseudomonas aeruginosa BB-BE3, P. aeruginosa BBBJ, and Gordonia amicalis BB-DAC were selected for their efficient hydrocarbon degradation and plant growth promotion (PGP) abilities. All three isolates were positive for siderophore production, phosphate solubilization, and IAA production, even in the presence of crude oil. The rhizoremediation ability was validated through pot trials where all three isolates promoted the growth of the Azadirachta indica plant in crude oil-contaminated soils. Treatment with the combination of the plant (A. indica) and bacteria, i.e., Pseudomonas aeruginosa BB-BE3; P. aeruginosa BBBJ; Gordonia amicalis BB-DAC showed 95.71, 93.28, and 89.88% removal of TPHs respectively, while the treatment with the plant (only) resulted in 13.44% removal of TPHs whereas, in the control (Sterile bulk soil + Crude oil), the hydrocarbon removal percentage was only 5.87%. The plant tissues were analyzed for catalase (CAT) and peroxidase (POX) activities, and the plants augmented with bacterial strains had significantly low CAT and POX activities as compared to uninoculated control. Therefore, the results suggest that the A. indica plant, in symbiotic association with these hydrocarbonoclastic rhizobacteria, could be used for bioremediation of crude oil-polluted soil.

Application of nitrogen fixing bacteria and poultry droppings for enhanced bioremediation of crude oil polluted-soil

The exploration, production and refining of crude oil has led to severe environmental degradation in the oil producing communities of the Niger Delta region of Nigeria. Enhanced bioremediation of tropical rainforest soil artificially polluted with crude oil, bioaugmented with nitrogen fixing bacteria (NFB) and biostimulated with poultry droppings was carried out ex situ. Soil sample was collected at 15cm depth from tropical rainforest soil of the University of Port Harcourt, Nigeria. The NFB was isolated from roots of leguminous plant Arachis hypogea, identified as Nitrobacter species. Bioaugmentation by application of NFB served as option A, option B (biostimulation by application of poultry droppings), option C (No amendment) served as the control. Bioremediation was monitored for 28 days for interval of 14 days, and determined using the percentage ratio of total petroleum hydrocarbon (TPH) losses for each period to TPH at initial day (day zero). Results of total culturable heterotrophic bacterial (TCHB) counts showed that highest range in option B (1.9×104- 2.4×109Cfu/g) than in option A (7.8×106 -2.29×107Cfu/g) and C (6.75×106 -2.6×107Cfu/g) respectively. Similarly, hydrocarbon utilizing bacterial (HUB) counts had higher range in option B (1.20×105 - 1.9×107Cfu/g) than in option A (8.30×104 - 2.30×105Cfu/g) and option C control (4.3×104 −1.69×105 Cfu/g) respectively. Changes in physicochemical parameters during the study showed reductions in nitrate, phosphate and TPH in all the options expect pH which showed slight increase in option C (6.20-6.24). Characterization and identification for bacteria revealed the following HUB genera Pseudomonas, Citrobacter, Bacillus, Corynebacterium, Micrococcus, Klebsiella, Staphylocuccus and Nitrobacter). The percentage losses in TPH from gas chromatography (GC) results showed the following; option A (44.24%) option B (61.08%) and option C - control (27.28%) respectively. The results from this study showed that option B, the application of poultry droppings as biostimulant was more efficient than the application of NFB in enhanced bioremediation of crude oil polluted soil, hence the use of poultry droppings which is available as organic waste, eco-friendly and cost-effective is recommended asbiostimulant for enhanced bioremediation in environmental cleanup of crude oil impacted-sites of the Niger Delta region of Nigeria.
 Key Words: Bioremediation, Crude oil polluted-soil, Biostimulation, Bioaugumentation, Poultry droppings, Nitrogen fixing bacteria.

Attenuation of petroleum hydrocarbon fractions using rhizobacterial isolates possessing alkB, C23O, and nahR genes for degradation of n-alkane and aromatics

This work assessed the catabolic versatility of functional genes in hydrocarbon-utilizing bacteria obtained from the rhizosphere of plants harvested in aged polluted soil sites in Ogoni and their attenuation efficacy in a bioremediation study. Rhizosphere soil was enumerated for its hydrocarbon-utilizing bacteria. The bacteria were in-vitro screened and selected through the quantification of their total protein and specific intermediate pathway enzyme (catechol 2,3-dioxygenase) activity in the metabolism of hydrocarbon. Thereafter, agarose gel electrophoresis technique was deployed to profile the genome of the selected strains for catechol 2,3-dioxygenase (C23O), 1,2-alkane monooxygenase (alkB), and naphthalene dioxygenase (nahR). Four rhizobacterial isolates namely Pseudomonas fluorescens (A3), Achromobacter agilis (A4), Bacillus thuringiensis (D2), and Staphylococcus lentus (L1) were selected based on the presence of C23O, alkB, and nahR genes. The gel electrophoresis results showed an approximate molecular weight of 200 bp for alkB, 300 bp for C23O, and 400 bp for nahR. The gas chromatogram for residual total petroleum hydrocarbon (TPH) revealed mineralization of fractions C8–C17, phytane, C18–C30. TPH for in-vitro bioremediation of crude oil-polluted soil was observed to have an optimal reduction/loss of 97% within the 56th day of the investigation. This study has further revealed that the microbiome of plants pre-exposed to crude oil pollution could serve as a reservoir for mining group of bacterial with broad catabolic potentials for eco-recovery and waste treatment purposes.

Percentage Bioremediation Assessment of Spent Mushroom Substrate (SMS) and Mucor racemosus in Hydrocarbon Contaminated Soil

Aim: The aim of the study was to assess Percentage Bioremediation of Spent Mushroom Substrate (SMS) and Mucor racemosus in hydrocarbon contaminated soil
 Place and Duration of Study: A portion of Rivers State University demonstration farmland in Nkpolu-Oroworukwo, Mile 3 Diobu area of Port Harcourt, Rivers State was used for this study. The piece of land is situated at Longitude 4°48’18.50’’N and Latitude 6o58’39.12’’E measuring 5.4864 m x 5.1816 m with a total area of 28.4283 m2. Bioremediation monitoring lasted for 56 days, analysis carried out weekly (per 7 days’ interval).
 Methodology: Five (5) experimental plots employing the Randomized Block Design were used each having dimensions of 100 x 50 x 30 cm (Length x Breadth x Height) = 150,000cm3. Baseline study of the uncontaminated and the deliberately contaminated agricultural soil was investigated for its microbiota and physico-chemical properties. Two of these plots were designated as pristine (Unpolluted soil) (CTRL 1) and crude oil contaminated soil without nutrient organics and bioaugmenting microbes (CTRL 2); these two serve as controls. Each of the experimental plots, except the control (CTRL 1), was contaminated with 2500 cm3 (2122.25 g) of crude oil giving initial Total Petroleum Hydrocarbon (TPH) value of 8729.00 mg/kg. The crude oil polluted soil in Plot 3 was further treated with 750 ml of Mucor racemosus broth (CS+Muc), Plot 4 was treated with 3000 g of Spent Mushroom Substrate (CS+SMS) while plot 5 was treated with the combination of both (CS+Muc+SMS). The plots were left for 7 days to ensure even distribution and soil-oil bonding. Sampling was done at seven-day interval (Day 1, 7, 14, 21, 28, 35, 42, 49, 56). Physicochemical parameters monitored were pH, Temperature, Nitrogen, Phosphorus, Potassium, and Total Petroleum Hydrocarbon (TPH) throughout the experimental period. Microbial parameters monitored were Total Heterotrophic Bacteria (THB), Total Heterotrophic Fungi (THF), Hydrocarbon Utilizing Bacteria (HUB) and Hydrocarbon Utilizing Fungi (HUF). Percentage (%) Bioremediation was estimated from percentage (%) reduction of Total Petroleum Hydrocarbon (TPH) from day 1 to day 56 in relation to control plots. Net % Bioremediation were also assessed to ascertain the actual potential of treatment agents singly or combined.
 Results: Total Heterotrophic Bacteria (THB) (CFU/g) recorded on day 7 and day 56 of the bioremediation were; day 7; CTRL 1 – US (1.07 x109), CTRL- CS (5.4 x108), CS+Muc (3.0 x108), CS+SMS (4.6 x108) and CS+Muc+SMS (5.0 x108). On day 56, data obtained were CTRL 1 –US (9.4 x108), CTRL 2 –CS (7.2 x109), CS+Muc (3.7 x108), CS+SMS (8.1x108) and CS+Muc+SMS (6.8 x108). The increase in number in the treated plots is a depiction of an increase in activity of the organism and the stimulating effect of bio-organics SMS while the untreated plot CTRL 1-US showed decrease in population at day 56. Similar trend showed for Total Heterotrophic Fungi. Generally, it was observed that the highest growth/ count was recorded at the 7th and 8th week (day 42 or day 49), at the 9th week there was an observable decrease; probably due to depletion of nutrients and other factors such as rainfall and seepage. The Net Percentage Hydrocarbon Utilizing Bacteria and Fungi (Net %HUB and Net %HUF) were highest in Crude Oil contaminated plot treated with Spent Mushroom Substrate (SMS) singly; that is (CS+SMS) (11.02% and 12.07%) and lowest in the uncontaminated soil – Control (CTRL 1 –US) (5.41% and 9.26%) respectively. The trend in decreasing order of Net % Hydrocarbon Utilizing Bacteria were as follows: CS+SMS (11.02%) > CS+Muc+SMS (10.14%) > CS+Muc (9.43%) > CTRL 2 –CS (8.1%) > CTRL 1 –US (5.41%) while Net % Hydrocarbon Utilizing Fungi followed similar trend and were: CS+SMS (12.07%) > CS+Muc+SMS (11.76%) = CS+Muc (11.76%) > CTRL 2 –CS (11.05%) > CTRL 1 –US (9.26%). Evaluation of Amount of Crude Oil or Hydrocarbon remediated and Net %Bioremediation revealed Crude Oil contaminated plot augmented with Mucor racemosus broth singly (CS+Muc) as having the highest bioremediation potential while the least is the untreated soil. The trend is as follows: CS+Muc (8599.19 mg/kg; 33.93%) > CS+Muc+SMS (8298.95 mg/kg; 32.74%) > CS+SMS (8197.03 mg/kg; 32.34%) > CTRL 2 –CS (166.54 mg/kg; 0.66%) > CTRL 1 –US (85.48 mg/kg; 0.34%)
 Conclusion: This shows that a single nutrient substrate or augmenting microorganism applied appropriately may have a more positive result, that is; higher bioremediation potential than combined or multiple mixed treatments. It was further observed that microbial counts decreased with time in treatments with augmenting organisms alone but increased considerably in treatments supplement with organics having its peak on the 49th day. It is therefore recommended that bioremediation of crude oil-polluted soil using bio-augmenting microorganism should be applied appropriately noting the volume: area ratio and be supplemented with efficient nutrient organics after every 49-day interval.

Enhanced microbial hydrocarbon biodegradation as stimulated during field-scale landfarming of crude oil-impacted soil

The study investigated microbial population dynamics and petroleum hydrocarbon degradation during 56-day landfarming bioremediation of crude oil-polluted soil at a Community in Niger Delta Nigeria. Landfarming involved the addition of fresh top soil from unpolluted soil to the polluted soil followed by periodic ploughing. Active oleophilic bacteria identified by 16S rRNA sequencing were screened with 2,6-dichlorophenol indophenol (DCPIP) redox indicator for their degradation profile. Physicochemical parameters monitored during landfarming included the extractable Total Petroleum Hydrocarbons (TPH), Polycyclic Aromatic Hydrocarbons (PAHs), pH and heavy metals. TPH was more than 9000 mg/kg which exceeded the Department of Petroleum Resources’ (DPR's) intervention value of 5000 mg/kg. Total viable counts of culturable heterotrophic and hydrocarbon utilizing bacteria in the polluted soil pre-landfarming were between 0.2 and 0.5 × 103 cfu/g. Monitoring of microbial population dynamics on days 0, 9, 18, 36 and 56 revealed an increase in hydrocarbon utilizing bacterial population from 104 up to 107 cfu/g which correlated significantly (R = 0.88; p = 0.05) with TPH reduction to below 700 mg/kg by day 56 when bioremediation experiment ended. Pseudomonas aeruginosa was the most efficient diesel degrader which showed 100% DCPIP decolourization. Acinetobacter, Myroides, Pseudomonas and Bacillus were central in the degradation of the long chain hydrocarbon fraction in the spill site. The fungal classes recovered included Eurotiomycetes, Saccharomycetes and Sordariomycetes. The findings from this study showed that the addition of virgin top soil to a compromised and chemically stressed soil is a green and eco-friendly strategy for enriching microbial degradative activities without addition of chemical fertilizers.

Fungal Diversity and Dynamics During Bioremediation of Crude Oilpolluted Soil

Fungal Diversity and Dynamics During Bioremediation of Crude Oilpolluted Soil

Field Metagenomics of Bacterial Community Involved in Bioremediation of Crude Oil-Polluted Soil

Field Metagenomics of Bacterial Community Involved in Bioremediation of Crude Oil-Polluted Soil

Effect of Woodchips on Bioremediation of Crude Oil-polluted Soil

Effect of Woodchips on Bioremediation of Crude Oil-polluted Soil

Optimization of reed-specific degrading bacteria by response surfaces for remediation of crude oil-polluted soil in Xinjiang, China

This paper discussed the optimization of conditions for remediation of crude oil-polluted soil based on pot experiment by applying reed-specific degrading bacteria, and using response surfaces methodology. We took the initial crude oil concentration, the amount of inoculation, the ratio of nitrogen and phosphorus, and the use of surfactant (Tween-80) as independent variables (factors), and the degrading ratio of crude oil as the dependent variable (response) after a 90-day experiment. The experiment explored the impacts of each independent variable and their interactions on the bioremediation of crude oil-polluted soil using the Box-Behnken design. Working with a simulated forecasting model the study obtained optimization values for the treatment parameters of 200 g/kg of the reed+specific degrading bacteria, a nitrogen to phosphorus ratio of about 6.0, and 0.2% surfactant. Under experimental conditions, for crude oil concentrations of 10, 30 and 50 g/kg, the optimal effects of the treatments achieved 71.87%, 66.61% and 54.52% degradation of the crude oil, respectively. The results can provide a basis for the technical development of plant-microorganism combined bioremediation of crude oil-polluted soil.

Culture-Independent Analysis of Bacterial Community Composition during Bioremediation of Crude Oil-Polluted Soil

Culture-Independent Analysis of Bacterial Community Composition during Bioremediation of Crude Oil-Polluted Soil

Phylogenetic diversity of dominant bacterial communities during bioremediation of crude oil-polluted soil

Bioremediation of hydrocarbon pollutants is advantageous owing to the costeffectiveness of the technology and the ubiquity of hydrocarbon degrading microorganisms in the soil. Soil microbial diversity is affected by hydrocarbon perturbation thus selective enrichment of hydrocarbon utilizers occurs. Hydrocarbons interact with the soil matrix and soil microorganisms determining the fate of the contaminants relative to their chemical nature and microbial degradative capabilities respectively. Bacterial dynamics in crude oil-polluted soil microcosms undergoing bioremediation were investigated over a 42-day period. Four out of the five microcosms containing 4kg of pristine soil each were contaminated with 4% Arabian light crude oil. Three microcosms were amended with either 25g of NPK fertilizer, calcium ammonium nitrate or poultry droppings respectively while the fourth designated oilcontaminated control was unamended. The fifth microcosm had only pristine soil and was set up to ascertain indigenous bacterial community structure pre-contamination. Biostimulated soils were periodically tilled and watered. Hydrocarbon degradation was measured throughout the experimental period by gas chromatography. Gas chromatographic tracing of residual hydrocarbons in biostimulated soils showed marked attenuation of contaminants starting from the second (day 14) till the sixth (day 42) week after contamination whereas no significant reduction in hydrocarbon peaks was seen in the oil contaminated control soil throughout the 6-week experimental period. Molecular fingerprints of bacterial communities involved in aerobic biodegradation of crude oil hydrocarbons in biostimulated soils and controls were generated with DGGE using PCR-amplification of 16S rRNA gene obtained from extracted total soil community DNA. DGGE fingerprints demonstrated that NPK, calcium ammonium nitrate and poultry droppings selected different bacterial populations during the active phase of oil degradation. Cluster analysis of DGGE bands using simple matching group average setting revealed that poultry droppings-amended soils and calcium ammonium nitrateamended soils formed distinct clades meaning that the treatment selected similar bacterial populations for each of the treatments whereas NPK soils showed less association. Excision, reamplification and sequencing of dominant DGGE bands in biostimulated soils revealed the presence of distinct hydrocarbon degraders like Corynebacterium spp., Dietzia spp., low G+C Gram positive bacteria and some uncultured bacterial clones. Phylogenetic analysis of the 16S rRNA gene sequences of these dominant bacterial communities was conducted using the neighbour joining method of PHYLIP. Two distinct clades appeared in the tree clustered members of the Actinobacteria and Firmicutes separately. The overall data suggested that Gram positive bacteria especially members of the Actinobacteria may have a key role in bioremediation of crude oil-polluted soil.

Bioremediation of Crude Oil-Polluted Soil by Sewage Sludge(Symposium 3.5.2 Risk Assessment and Risk Based Remediation,<Special Issue>International Symposium: Soil Degradation Control, Remediation, and Reclamation, Tokyo Metropolitan University Symposium Series No.2, 2010)

Bioremediation of Crude Oil-Polluted Soil by Sewage Sludge(Symposium 3.5.2 Risk Assessment and Risk Based Remediation,<Special Issue>International Symposium: Soil Degradation Control, Remediation, and Reclamation, Tokyo Metropolitan University Symposium Series No.2, 2010)

Bacterial diversity in a tropical crude oil-polluted soil undergoing bioremediation

The bacterial diversity in a tropical soil experimentally polluted with crude oil during a 57 days bioremediation was investigated in five 1 m2 plots using total culturable hydrocarbon utilizing bacteria, heterotrophic bacteria and gas chromatographic analyses. Four out of the five experimental plots received each 4 L of Bonny light crude oil while three treatment plots received 3 kg of NPK, urea fertilizers or poultry droppings with periodic tilling. Two plots, oil-contaminated and pristine served as controls. Bacterial counts increased 200 fold and 2 fold in the NPK treated and poultry-dropping-treated plots respectively, by day 31 post-inoculation. Detectable hydrocarbons in the treatment plots decreased by 84 - 95% and 96 - 99%, 31 and 57 days post-inoculation, respectively, compared with the petroleum contaminated control. Bacterial strains isolated included Rhodococcus sp., Nocardia sp., Arthrobacter sp., Gordonia sp., Mycobacterium sp., Corynebacterium sp., Bacillus sp., Micrococcus sp., Flavobacterium sp., Pseudomonas sp. and Alcaligenes sp. The overall data suggest an important contribution of Actinobacteria during bioremediation of crude oil-polluted soil.

238 POSTER The selective proteasome inhibitor carfilzomib in combination with chemotherapeutic agents improves anti-tumor response in solid tumor xenograft models

The study investigated microbial population dynamics and petroleum hydrocarbon degradation during 56-day landfarming bioremediation of crude oil-polluted soil at a Community in Niger Delta Nigeria. Landfarming involved the addition of fresh top soil from unpolluted soil to the polluted soil followed by periodic ploughing. Active oleophilic bacteria identified by 16S rRNA sequencing were screened with 2,6-dichlorophenol indophenol (DCPIP) redox indicator for their degradation profile. Physicochemical parameters monitored during landfarming included the extractable Total Petroleum Hydrocarbons (TPH), Polycyclic Aromatic Hydrocarbons (PAHs), pH and heavy metals. TPH was more than 9000 mg/kg which exceeded the Department of Petroleum Resources’ (DPR's) intervention value of 5000 mg/kg. Total viable counts of culturable heterotrophic and hydrocarbon utilizing bacteria in the polluted soil pre-landfarming were between 0.2 and 0.5 × 103 cfu/g. Monitoring of microbial population dynamics on days 0, 9, 18, 36 and 56 revealed an increase in hydrocarbon utilizing bacterial population from 104 up to 107 cfu/g which correlated significantly (R = 0.88; p = 0.05) with TPH reduction to below 700 mg/kg by day 56 when bioremediation experiment ended. Pseudomonas aeruginosa was the most efficient diesel degrader which showed 100% DCPIP decolourization. Acinetobacter, Myroides, Pseudomonas and Bacillus were central in the degradation of the long chain hydrocarbon fraction in the spill site. The fungal classes recovered included Eurotiomycetes, Saccharomycetes and Sordariomycetes. The findings from this study showed that the addition of virgin top soil to a compromised and chemically stressed soil is a green and eco-friendly strategy for enriching microbial degradative activities without addition of chemical fertilizers.

Role of diazotrophic bacteria in the bioremediation of crude oil-polluted soil

Nitrogen is one of the most limiting inorganic nutrients in the process of bioremediation of crude oil-polluted environments. Enhanced remediation of crude oil polluted soil was achieved, in situ, by accelerating the biodegradation process through seeding with adapted Azotobacter which not only acted as supplier of fixed nitrogen to the indigenous crude oil-degrading bacteria, but also performed some co-metabolic activities. This work describes the capability of Azotobacter in providing activities that are useful in the bioremediation of crude oil-polluted soil and biological nitrogen fixation when in association with indigenous oil-degrading bacteria. © 1999 Society of Chemical Industry