Bioremediation Of Crude Oil Research Articles

Optimization of Bioremediation of Crude Oil Polluted Soil through Variation in Moisture Content and Proportion of Augmenting Bacterium and Fungus

Aim: The aim of this study was to enhance bioremediation of crude-oil polluted soil through optimization procedures. Study Design: The Box-Benkhen coded values format and experimental design were used in the design of the study. Place and Duration of Study: The study was carried out in the Microbiology laboratory of the Department of Microbiology, Rivers State University, Nigeria, between February 2024 and July 2024. Methodology: Hydrocarbon utilizing bacteria (HUB) & fungi (HUF) were isolated from crude-oil polluted soil, and screened for hydrocarbon degrading ability (HCDA). HUB and HUF with the highest HCDA were identified and used for bioremediation optimization experiment. In the experiment, different combination of moisture content (MC), and proportion of HUB (pHUB) & HUF (pHUF) were investigated for their effect on extent of hydrocarbon reduction (EoHR). EoHR obtained from the different combinations were fitted using a generalized polynomial model so as to obtain a polynomial equation for predicting EoHR. The equation was used in generating prediction profiles from which the combined values of the 3 parameters that will lead to the highest EoHR was determined. The predicted combined values were implemented in a new setup. A control and a setup enhanced with fertilizer were also prepared. The setups were maintained for 3 weeks. On day 0, 14, and 21 samples were collected and analyzed for total hydrocarbon concentration (THC). Results: The results obtained showed that among the coded isolated HUB and HUF, HB5 and HF1 had the highest HCDA; 36.4% and 4.1% respectively, and were identified phenotypically as Klebsiella ornithinolytica and Aspergillus flavus respectively. The results of the optimization experiment and prediction profiles showed that the highest (68.6%) EoHR was achievable at MC = 20%, pHUB = 10%, and pHUF = 1%. The actual EoHR on day 21 in the optimized, enhanced optimized, and control setups were 60.3, 58.9, and 39.9% respectively. Conclusion: Bioremediation optimization studies is advantageous and should be carried out on crude-oil polluted sites before carrying out bioremediation.

Crude oil degrading efficiency of formulated consortium of bacterial strains isolated from petroleum-contaminated sludge.

Crude oil contamination has been widely recognized as a major environmental issue due to its various adverse effects. The use of inhabitant microorganisms (native to the contaminated sites) to detoxify/remove pollutants owing to their diverse metabolic capabilities is an evolving method for the removal/degradation of petroleum industry contaminants. The present study deals with the exploitation of native resident bacteria from crude oil contaminated site (oil exploration field) for bioremediation procedures. Fifteen (out of forty-four) bioremediation-relevant aerobic bacterial strains, belonging to the genera of Bacillus, Stenotrophomonas, Pseudomonas, Paenibacillus, Rhizobium, Burkholderia, and Franconibacter, isolated from crude oil containing sludge, have been selected for the present bioremediation study. Crude oil bioremediation performance of the selected bacterial consortium was assessed using microcosm-based studies. Stimulation of the microbial consortium with nitrogen or phosphorous led to the degradation of 60-70% of total petroleum hydrocarbon (TPH) in 0.25% and 0.5% crude oil experimental sets. CO2 evolution, indicative of crude oil mineralization, was evident with the highest evolution being 28.6mgmL-1. Ecotoxicity of treated crude oil-containing media was assessed using plant seed germination assay, in which most of the 0.25% and 0.5% treated crude oil sets gave positive results thereby suggesting a reduction in crude oil toxicity.

Differential biodegradation of alkanes in crude oil by three oleophilic strains

Ex situ bioremediation of crude oil in bioreactors by oleophilic bacteria permits to regulate temperature, nutrients and oxygen and the results in petroleum removal are much more effective than in situ bioremediation. This article quantify the biodegradation of alkanes which conform crude oil in bioreactors by three oleophilic strains. Samples were collected from the bioreactors and extracted in dichloromethane (10:1 aqueous/organic v/v) for amplification in GC-FID analysis of individual alkanes. 94% of crude oil (initial concentration 8.6 g l−1) is removed in 24 days by Bacillus licheniformis and in 38 days by Pseudomonas putida and Paenibacillus glucanolyticus. Higher biodegradation rates (9.0–26.5 mg l−1d−1) are reached for the lightest fraction of crude oil by Bacillus licheniformis (C10–C16), and much lower for C22–C25 alkanes (4.4–8.9 mg l−1d−1). Pseudomonas putida and Paenibacillus glucanolyticus degrade a wide range of alkanes at 1.6–9.6 mg l−1d−1, with preference over C8–C16 hydrocarbons. Heptane and octane (alkanes cited to be toxic) are difficult for biodegradation in two strains (B. licheniformis and P. putida) and a recalcitrant “window” C18–C21 is observed for the three oleophilic strains in the alkane range studied.

Bioremediation of crude oil polluted surface water using specialised alginate-based nanocomposite beads loaded with hydrocarbon-degrading bacteria and inorganic nutrients

This study explored the elimination of petroleum from river water using matrix-bound composites of iron oxide nanoparticles (IONPs) decorated on biochar with immobilized hydrocarbon-degrading bacteria (HCB) and monoammonium phosphate (MAP) at 10%v/v pollution levels. The beads containing biochar and IONPs (BCNP) showed better pollutant removal than those with biochar alone (BC). The treatment with beads containing biochar, IONPs, HCB and MAP had the highest removal efficiency for TPH (98.5%) and PAH (93.7%) and the lowest biodegradation half-lives. The removal efficiency for TPH and PAH was greater in the treatment microcosms than in the controls with significant differences (p ≤ 0.05) between the two groups. TPH and PAH elimination levels differed significantly (p ≤ 0.05) between systems with the BC bead and those with the BCNP beads. The study revealed a steady increase in the abundance of the total cultivable heterotrophic bacteria and cultivable hydrocarbon utilizing bacteria in the treatment microcosms after an initial decline, with strong negative correlation between bacterial abundance and TPH and PAH concentrations based on r values (0.5 − 1.0) and shared variances obtained. Bacillus, Pseudomonas and Klebsiella pneumoniae dominated as the drivers of the bioremediation process while organic acids and esters (25.0%) and ketones (14.3%) were the main biodegradation metabolite groups.

Using PyCaret to model Chlorella vulgaris's growth response to salinity and oil contamination for crude oil bioremediation

ABSTRACT Crude oil spills significantly impact aquatic ecosystems, necessitating innovative remediation strategies. Microalgae-based bioremediation, particularly with Chlorella vulgaris, offers a promising solution. This study introduces a novel framework that evaluates the combined effects of selected environmental stressors on microalgal adaptability, advancing beyond traditional isolated factor analyses. By integrating a factorial experimental design with a machine learning approach using PyCaret AutoML and SHAP values, we provide a detailed examination of how crude oil concentration, salinity, and exposure duration affect C. vulgaris growth. The Extra Trees Regressor model emerged as highly accurate in predicting biomass concentration, a crucial adaptability indicator, achieving an MAE of 0.0202, RMSE of 0.029, and an R² of 0.8875. SHAP analysis highlighted salinity and crude oil as significant growth influencers, with exposure duration playing a minor role. Notably, C. vulgaris exhibited more sensitivity to salinity than to crude oil, indicating potential high-salinity challenges but also a strong tolerance to oil pollutants. These findings enhance our understanding of microalgal responses in polluted environments and suggest improved bioremediation approaches for saline waters affected by oil spills, leveraging the synergy of environmental factors and machine learning insights.

The use of pigeon droppings in bioremediation of crude oil contaminated open farm

The use of pigeon droppings in bioremediation of crude oil contaminated open farm

The potential of Serratia marcescens in Bioremediation of Crude-oil Polluted Soil

Bioremediation, which involves the use of microorganisms, is an environmentally friendly approach in restoring crude-oil polluted environments. The use of Serratia marcescens (SM) in bioremediation of crude-oil polluted soil was investigated in this study. Soil from a farm was inoculated in nutrient broth for bacterial enrichment. The broth culture was inoculated on nutrient agar and incubated at ambient temperatures (27 – 32 °C). Bacterial isolates with red colonies were selected and identified. Identified SM was used in preparing bacterial-mineral-salts suspension. The bioremediation experiment consisted of two setups: Control (CT) and setup treated with SM (TSM). Both setups contained crude-oil polluted soil. Mineral-salt solution and the bacterial-mineral-salt suspension were added to setup CT and TSM, respectively. Total hydrocarbon concentration (THC), total heterotrophic bacteria (THB), hydrocarbon utilizing bacteria (HUB), total fungi (TF), and hydrocarbon utilizing fungi (HUF) in the setups were determined at various times. About 38.3 % and 46.5 % THC reduction were achieved in setup CT and TSM, respectively. The percentage of THB that are HUB was higher in setup TSM on day 7 (8.8 %) and day 28 (4.4 %) but higher in setup CT on day 14 (1.7 %) and day 21 (6.6 %). The percentage of TF that are HUF was higher in setup TSM on day 14 (98.0 %) and on day 28 (41.6 %) but higher in setup CT on day 21 (85.3 %). It is concluded that SM has a potential for use in bioremediation of crude-oil polluted soil

Microbial remediation of petroleum-contaminated soil focused on the mechanism and microbial response: a review.

The environmental pollution caused by petroleum hydrocarbons has received considerable attention in recent years. Microbial remediation has emerged as the preferred method for the degradation of petroleum hydrocarbons, which is experiencing rapid development driven by advancements in molecular biology. Herein, the capacity of different microorganisms used for crude oil bioremediation was reviewed. Moreover, factors influencing the effectiveness of microbial remediation were discussed. Microbial remediation methods, such as bioaugmentation, biostimulation, and bioventilation, are summarized in this review. Aerobic and anaerobic degradation mechanisms were reviewed to elucidate the metabolic pathways involved. The impacts of petroleum hydrocarbons on microorganisms and the environment were also revealed. A brief overview of synthetic biology and a unique perspective of technique combinations were presented to provide insight into research trends. The challenges and future outlook were also presented to stimulate contemplation of the mechanisms involved and the development of innovative techniques.

Bioremediation of Crude Oil Contaminated Saline Soil Using a Bacterial Consortium and Different Carriers

AbstractBioremediation of crude‐oil‐contaminated soils is critical to supporting soil ecosystem health and functioning. Here, to explore the potential for different remediation strategies to restore soil microbial functioning and enhance remediation, three strains of bacteria, Bacillus toyonensis, Bacillus sp. and Bacillus cereus, were inoculated to saline, crude oil‐contaminated soil collected from Azadegan Oil Field, Iran. Bacteria were added using different carriers, including corn biochar, humic acid, chitosan and kaolin, and free bacteria as a control. Changes in soil metal concentrations and residual crude oil were determined after 60 days of incubation. Soil respiration, microbial biomass, and enzyme (dehydrogenase and catalase) activities were measured. The carriers accompanying a bacterial consortium were more efficient than free bacteria in removing crude oil and heavy metals from soil, and they performed better in removing n‐alkanes with carbon chains of C12–24. The oil removal rate and metal immobilization efficiencies were greatest in soils with chitosan‐bacteria treatment, with removal of crude oil, Al, Pb, Sr, and Cr (93%, 63%, 37%, 54%, and 15%, respectively) greater than for soils with free cells without treatment (61%, 54%, 9%, 50%, and 3%, respectively). Soil electrical conductivity decreased after application of bacterial mixtures with and without carriers. The inoculation of bacteria and application of soil amendments increased soil microbial biomass carbon and the activity of catalase and dehydrogenase. Overall, our results indicated the greatest potential remediation was achieved by applying immobilized microorganisms in chitosan to accelerate the biodegradation of crude oil and the removal of heavy metals.

Exploring Hydrocarbon Degradation Capacity and Mathematical Modeling of Mixotrophic Growth Kinetics in Tetradesmus obliquus and Desmodesmus armatus for Crude Oil Bioremediation

Exploring Hydrocarbon Degradation Capacity and Mathematical Modeling of Mixotrophic Growth Kinetics in Tetradesmus obliquus and Desmodesmus armatus for Crude Oil Bioremediation

Crude oil biodegradation potential of lipase produced by Bacillus subtilis and Pseudomonas aeruginosa isolated from hydrocarbon contaminated soil

Microbial biodegradation of oil pollutants and their derivatives has become the most environmental-friendly method in the developing world. The aim of this study was to evaluate crude oil biodegradation potential of lipase produced by indigenous bacteria from oil contaminated soil. Indigenous bacteria isolates were identified as species of Bacillus subtilis and Pseudomonas aeruginosa, the isolates were able to produce lipase as revealed in their zone of clearance on tween 80 agar plates and the presence of lipase produced by the two bacteria were further confirmed using spectrophotometric analyses. Lipase produced by B. subtilis showed maximal lipase activity at pH 8 and 40°C while the enzyme produced by P. aeruginosa showed maximal lipase activity (U/mL) at pH 8 and 50°C when subjected to various pH and temperature respectively. Lipase produced by B. subtilis recorded 8.11 ± 0.70% of crude oil degradation in mineral salt medium within 28 days, while that of P. aeruginosa recorded 15.6 ± 0.03% of crude oil biodegradation. The GC–MS analysis of the crude oil treatment showed complete mineralization of several compounds, and also showed peak reduction which indicates lipase efficiency in the degradation of hydrocarbons. As revealed by GC–MS analysis, out of the 8 hydrocarbons identified in an undegraded oil, 5 were completely degraded by the enzyme activities while 2 (toluene and methyl, cyclopentane) were identified with hydrocarbons treated with lipase. The enzymes produced by B. subtilis and P. aeruginosa can serve as useful product for bioremediation of crude oil contaminated soil.

Enhancing Bioremediation of Crude Oil Contaminated Soil from Kokori, Delta State, Nigeria Using Cow Dung as Biostimulant

This study was aimed at assessing the bioremediation of crude oil contaminated soil from kokori, Delta state, Nigeria by indigenous bacteria flora using cow dung as biostimulants. The activities of commercial agricultural investors, small-scale farmers, and the indigenous people of the Nigerian Niger Delta, whose livelihood has been heavily dependent on agriculture, have all been severely impeded and even destroyed by the crude oil spill. Therefore, a solution to the issue of oil spills in these places is required. Hydrocarbon contaminated soil was collected from a crude oil polluted sites and a non-contaminated soil was collected from Agricultural cultivable farmland in kokori, Delta state and its physicochemical properties were determined before the bioremediation period. The nutrients composition of cow dung used for the study was determined. After adding different amounts of cow dung to 200g of soil, the mixture was left to incubate at room temperature for six (6) weeks. Throughout the course of the experiment, measurements of the total petroleum hydrocarbon eliminated and the hydrocarbon-utilizing bacteria count (HUB) were made at one-week intervals. Increased in the HUB counts with corresponding decrease in the amount of hydrocarbons were observed. While highest HUB count was observed for the various amendment at week six, 5g/kg cow dung treatments had the highest count (3.35 x103cfu/g) with residual hydrocarbon (519.56mg/kg) while treatments 2.5g/kg cow dung had 2.91x103cfu/g with residual hydrocarbon of 541.06mg/kg. The result further revealed the degree of degradation likewise the bioremediation efficiency with C having the highest degree of degradation of (49.04%) at week six and bioremediation efficiency of (75.26%) at the first week of study. The introduction of cow dung into soil that has been contaminated with hydrocarbons promotes the growth of naturally occurring bacteria that break down hydrocarbons, leading to a greater amount of hydrocarbon degradation.

Bioremediation of Crude Oil Polluted Soil by Cyanobacteria and Bacterial Consortium in Izombe Community of Imo State, Nigeria.

Bioremediation of Crude Oil Polluted Soil by Cyanobacteria and Bacterial Consortium in Izombe Community of Imo State, Nigeria.

Evaluation of Organic Treatments for Enhanced Bioremediation of Crude Oil Impacted Soil in Bayelsa State, Nigeria

The aim of this study was to evaluate organic treatments for enhanced bioremediation of crude oil impacted soil in Bayelsa State, Nigeria. Koroama community in Gbarain clan, Yenagoa local government area, Bayelsa State, Nigeria was randomly selected for the study. The experimental research design was adopted for this study, which was carried out for 28 days. The required soil samples were collected at a depth of 0 – 15cm from a 3 x 3m experimental plot developed in a farmland in Koroama community. Five sampled points, in the form of flat beds A, B, C, D and E respectively were randomly selected. Sampled beds A to D were simulated (contaminated) with 2.25kg of crude oil. The objective is to simulate conditions of a major crude oil spill. The crude oil contaminated soil samples were then allowed to condition for 6 days before treatment with 2kg organic treatments like goat manure (GM), poultry droppings (PD) and the combination of goat manure and poultry droppings (GM & PD). Bed E was unpolluted and untreated (control A) while Bed D was crude oil impacted and untreated soil (control B). The study showed that the impact of crude oil on the soil affects both the physical properties of the soil. The study also showed that the sampled organic treatments were all effective in restoring the physical properties of the crude oil impacted soil. At the end of the study, GM showed the highest TPH degradation rate (62.1%) followed by PD (57.1%) and the least, the combination of GM & PD (52.0%). Hence, the application of GM, PD and the combination of GM & PD are highly recommended for bioremediation of crude of impacted soil with special preference to GM.

Plant biomass extracted eco-friendly natural surfactant enhanced bio-electrokinetic remediation of crude oil contaminated soil

The current work investigates bioremediation (BIO) and electrokinetic (EK) remediation of crude oil hydrocarbons utilizing the biomass-electrokinetic (BIO-EK) approaches. The use of natural surfactants derived from plant biomass may improve remediation capacity by enhancing the solubility of organic pollutants. Sapindus mukorossi, a natural surfactant producer, was extracted from plant biomass in this study. The crude oil biodegradation efficiency was reported to be 98 %. In nature, FTIR confirms that plant biomass is lipopeptide. GCMS revealed that the crude oil (C7 – C23) was efficiently bio-degraded from lower to higher molecular weight. The application of natural surfactants in electokinetic remediation increased the plant biomass degradation of crude oil polluted soil by 98% compared to electrokinetic 55% in 2 days. Natural surfactant improves hydrocarbon solubilization and accelerates hydrocarbon electro migration to the anodic compartment, as confirmed by the presence of greater total organic content than the electrokinetic. This study proves that BIO-EK compared with a natural surfactant derived from plant biomass may be utilized to improve in situ bioremediation of crude oil polluted soils.

Integrated approach for efficient crude oil bioremediation: Bacterial consortium development, mathematical modelling and scalable bioprocess design

Crude oil pollution has a significant impact on the surrounding ecosystem, making its remediation an urgent necessity due to the waste discharge and seepage from petroleum industries. To address this issue, we developed an efficient bacterial consortium from strains isolated from petroleum refinery waste. These strains were identified as genera Pseudomonas, Ochrobactrum, Alcaligenes, and Bacillus, specifically chosen for their ability to biodegrade crude oil. Through the optimization of process parameters, we enhanced the biodegradation efficiency of the bacterial consortium. Furthermore, the scalability of the biodegradation process was established in 6L batch bioreactor, which featured a total TPH removal of 80.10 %. The independently obtained substrate inhibition data and bioreactor batch kinetics data were utilized to develop a batch mathematical-model. The mathematical model accurately simulated batch bioreactor kinetics and was further extrapolated to design a fed-batch nutrient feeding strategy. Implementing this strategy, based on the model, led to a higher TPH removal rate of 86.86 % in a constant feed batch-fed-batch approach, as validated through experimental implementation. Validation of the biodegradation process was confirmed through Gas Chromatography-Mass Spectrometry (GC–MS) analysis of degraded samples, which confirmed the biodegradation of petroleum hydrocarbons. The increased rates of oil biodegradation achieved through our developed consortium and advanced bioprocess offer a promising solution for the effective clean-up of oily pollutants present in petroleum wastewater and oil spillage scenarios.

Use of Wastewater from Legume Cooking in Bioremediation of Crude-Oil Polluted Soil

Wastewaters generated from the cooking of foods contain nutrients. Such wastewaters may have the potential to be used as nutrients in bioremediation of hydrocarbon polluted environments. In this study, use of wastewaters generated from the cooking of two legumes as nutrient source during bioremediation of hydrocarbon polluted soil was investigated. Crudeoil polluted soils were treated with wastewater from cooking of cowpea (setup B), wastewater from cooking of groundnut-pods (setup G), a mixture of the wastewaters (setup B+G), and tap-water (control setup: setup C). Selected microbial group and physicochemical parameters were determined at seven days interval. The results obtained showed that the extent of hydrocarbon degradation in setup B, G, B+G, and C were 45.5, 23.2, 1.3, and 37.9 % respectively. There was decrease in total organic carbon (TOC), phosphorus, and nitrogen in the setups. However, in setup G there was increase in TOC. Setup C and B had the highest decrease in phosphorus, and setup B had the highest decrease in nitrogen. There was decrease and subsequent increase in percent hydrocarbon-utilizing bacteria in all the setups; the subsequent increase was highest in setup B. There was generally a decrease in percent hydrocarbon-utilizing fungi, except in setup C. The hydrocarbon utilizing microorganisms isolated from the treated soils include Bacillus, Micrococcus, Acinetobacter, Staphylococcus, Pseudomonas, Penicillium, Aspergillus, Candida and Fusarium. Their presence indicates that hydrocarbon degradation was due to biodegradation. It is concluded that wastewater from cooking of cowpeas can be applied in bioremediation of crude-oil polluted soil.

Studies on the ecological succession of microorganisms during bioremediation of hyper-toxic crude oil polluted soil using five amendments

Dangerous impact of crude oil in the environment is a common knowledge and its spillage has been controlled using both chemical and biological methods. Bioremediation however, is most advocated because of its environmental friendliness and sustainability. The present study sought to identify and monitor successions, and diversity of microorganisms elicited by five amendments in the bioremediation of hypertoxic crude oil pollution. Five amendment patterns which incorporated 200 g each of saw dust, wood chip, compost, N.P.K. fertilizer and poultry droppings; were used to stimulate biodegradation in 50% crude oil contaminated soil, and their respective microbial counts and microbial successions were monitored over a period of eight months. Statistical analyses were conducted using SPSS version 21 software. Poultry droppings amendment had the highest significant (p<0.05) bacterial and fungal counts. Bacillus subtilis, Stenotrophomas maltophilia and Pseudomonas aeruginosa were the predominant bacteria observed during the microbial succession studies across the amendment microcosms, while Aspergillus flavus, Penicillium citrinnum and Fusarium solani were the predominant fungal isolates. It was observed that the hyper-toxic concentration of the crude oil impacted the diversity and succession of the microbes across all the amendments used when compared to the control sample.

Effect of glucose on biodegradation of hydrocarbons in crude-oil polluted soil undergoing bioremediation

Bioremediation of crude oil polluted soil can be enhanced by addition of easily assimilated carbon sources such as glucose. The aim of this study was to determine the effect of increasing concentration of glucose on biodegradation of hydrocarbons in crude-oil polluted soil undergoing bioremediation. Some hydrocarbon utilizing bacteria and fungi were used as bioremediation agents in the study. Soil artificially polluted with crude-oil was divided into 5 setups: CT (control), S1, S2, S4, and S8. Suspension of the bioremediation agents and nutrients were added to the setups. About 5, 10, 20, and 40 g glucose were added to S1, S2, S4, and S8 respectively: equivalent to 1, 2, 4, and 8 % glucose concentration. Soil moisture in the setups was maintained at 10-20 % during the experimental period. Total hydrocarbon concentration (THC), and population of total heterotrophic bacteria (THB), hydrocarbon utilizing bacteria (HUB), total fungi (TF), & hydrocarbon utilizing fungi (HUF) were determined on day 1 and 21. Results obtained showed that increase in HUB occurred only in setup S1, and setup S1 had the highest extent of hydrocarbon degradation (18.93 %). Setup S8 had the lowest extent of hydrocarbon degradation (0.79 %). It is concluded that glucose concentration of 1 % and below will enhance biodegradation of hydrocarbons in crude-oil polluted soils, whereas high glucose concentration will retard hydrocarbon biodegradation. Also, the study indicates the possibility of fungi not partaking in hydrocarbon degradation when glucose is made available.

Biodegradation of saturate fraction of crude oil and production of signature carboxylic acids

Bacteria degrading large portion of saturated hydrocarbons are important for crude oil bioremediation. This study investigates Novosphingobium sp. S1, Gordonia amicalis S2 and Gordonia terrae S5 capability of degrading wide range of saturated hydrocarbons from Congo Bilondo crude oil and discusses the degradation pathway. A parallel analytical approach combining GC-MS and LC-HRMS enabled characterization of saturated hydrocarbons and comprehensive determination of carboxylic acid metabolites produced during biodegradation, respectively. Results showed that the three strains could efficiently degrade the n-alkanes (C10–C28) as well as methyl-substituted alkanes (C11–C26). The series of mono-, hydroxy- and dicarboxylic acids identified in this study confirmed the active biodegradation of the saturate fraction and suggest their degradation was via the bi-terminal oxidation pathway. This is the first study linking these bacterial species to bi-terminal oxidation of the saturated hydrocarbons. The study highlights the potential application of the bacterial strains in the bioremediation of crude oil contaminated sites. Additionally, while carboxylic acids is indicated as a suitable and valuable metabolic biomarker, its application is considered feasible and cost effective for rapid monitoring and evaluation of hydrocarbon biodegradation.