Characterization and identification of long-chain hydrocarbon-degrading bacterial communities in long-term chronically polluted soil in Ogoniland: an integrated approach using culture-dependent and independent methods.

Escalating oil consumption has resulted in an increase in accidental spills of petroleum hydrocarbons, causing severe environmental degradation, notably in vulnerable regions like the Niger Delta. Complex mixture of these hydrocarbons particularly long-chain alkanes presents unique challenges in restoration of polluted environment due to their chemical properties. This study aimed to investigate the long-chain hydrocarbon-degrading bacterial communities within long-term chronically polluted soil in Ogoniland, by utilizing both traditional cultivation methods and modern culture-independent techniques. Results revealed that surface-polluted soil (SPS) and subsurface soil (SPSS) exhibit significantly higher total organic carbon (TOC) ranging from 5.64 to 5.06% and total petroleum hydrocarbons (TPH) levels ranging from 36,775 ppm to 14,087 ppm, compared to unpolluted soil (UPS) with 1.97% TOC and 479 ppm TPH, respectively. Analysis of carbon chain lengths reveals the prevalence of longer-chain alkanes (C20-28) in the surface soil. Culture-dependent methods, utilizing crude oil enrichment (COE) and paraffin wax enrichment (PWE), yield 47 bacterial isolates subjected to a long-chain alkane degradation assay. Twelve bacterial strains demonstrate significant degradation abilities across all enriched media. Three bacterial members, namely Pseudomonas sp. (almA), Marinomonas sp. (almA), and Alteromonas (ladA), exhibit genes responsible for long-chain alkane degradation, demonstrating efficiency between 50 and 80%. Culture-independent analysis reveals that surface SPS samples exhibit greater species richness and diversity compared to subsurface SPSS samples. Proteobacteria dominates as the phylum in both soil sample types, ranging from 22.23 to 82.61%, with Firmicutes (0.2-2.22%), Actinobacteria (0.4-3.02%), and Acidobacteria (0.1-3.53%) also prevalent. Bacterial profiles at genus level revealed that distinct variations among bacterial populations between SPS and SPSS samples comprising number of hydrocarbon degraders and the functional predictions also highlight the presence of potential catabolic genes (nahAa, adh2, and cpnA) in the polluted soil. However, culture-dependent analysis only captured a few of the dominant members found in culture-independent analysis, implying that more specialized media or environments are needed to isolate more bacterial members. The findings from this study contribute valuable information to ecological and biotechnological aspects, aiding in the development of more effective bioremediation applications for restoring oil-contaminated environments.

Functional Gene Diversity of Selected Indigenous Hydrocarbon-Degrading Bacteria in Aged Crude Oil.

Crude oil pollution has consistently deteriorated all environmental compartments through the cycle of activities of the oil and gas industries. However, there is a growing need to identify microbes with catabolic potentials to degrade these pollutants. This research was conducted to identify bacteria with functional degradative genes. A crude oil-polluted soil sample was obtained from an aged spill site at Imo River, Ebubu, Komkom community, Nigeria. Bacteria isolates were obtained and screened for hydrocarbon degradation potential by turbidometry assay. Plasmid and chromosomal DNA of the potential degraders were further screened for the presence of selected catabolic genes (C230, Alma, Alkb, nahAC, and PAHRHD(GP)) and identified by molecular typing. Sixteen (16) out of the fifty (50) isolates obtained showed biodegradation activity in a liquid broth medium at varying levels. Bacillus cereus showed highest potential for this assay with an optical density of 2.450 @ 600 nm wavelength. Diverse catabolic genes resident in plasmids and chromosomes of the isolates and, in some cases, both plasmid and chromosomes of the same organism were observed. The C230 gene was resident in >50% of the microbial population tested, while other genes occurred in lower proportions with the least observed in nahAC and PAHRHD. These organisms can serve as potential bioremediation agents.

Shift in microbial group during remediation by enhanced natural attenuation (RENA) of a crude oil-impacted soil: a case study of Ikarama Community, Bayelsa, Nigeria.

Acute and chronic pollution of environments with crude oil does not bode well for biota living within the vicinity of polluted environments. This is due to environmental and public health concerns on the negative impact of crude oil pollution on living organisms. Enhancing microbial activities by adding nutrients and other amendments had proved effective in pollutant removal during bioremediation. This study was carried out to determine how microbial group respond during remediation by enhanced natural attenuation (RENA) during a field-scale bioremediation. Crude oil-polluted soil samples were collected (before, during, and after remediation) from a site undergoing remediation by enhanced natural attenuation (RENA) at Ikarama Community, Bayelsa State, Nigeria, and were analyzed for total petroleum hydrocarbon (TPH), polyaromatic hydrocarbon (PAH), and a shift in microbial community. The gas chromatography-flame ionization detector (GC-FID) results showed that the pollutant concentrations (TPH and PAH) reduced by 98 and 85%, respectively, after the remediation. Culturable hydrocarbon utilizing bacteria (CHUB) was highest (8.3 × 104 cfu/g) for sample collected during the remediation studies, whilst sample collected after remediation had low CHUB (6.1 × 104 cfu/g) compared to that collected before remediation (7.7 × 104 cfu/g). Analysis of 16S rRNA of the isolated CHUB showed they belonged to eight bacterial genera namely: Achromobacter, Alcaligenes, Azospirillus, Bacillus, Lysinibacillus, Ochrobactrum, Proteus, and Pusillimonas, with Alcaligenes as the dominant genus. In this study, it was observed that the bacterial community shifted from mixed group (Gram-positive and -negative) before and during the remediation, to only the latter group after the remediation studies. The betaproteobacteria groups were the dominant isolated bacterial phylotype. This study showed that RENA is an effective method of reducing pollutant concentration in crude oil-polluted sites, and could be applied to other polluted sites in the Niger Delta region of Nigeria to mitigate the devastating effects of crude oil pollution.

Bioremediation techniques-classification based on site of application: principles, advantages, limitations and prospects.

Environmental pollution has been on the rise in the past few decades owing to increased human activities on energy reservoirs, unsafe agricultural practices and rapid industrialization. Amongst the pollutants that are of environmental and public health concerns due to their toxicities are: heavy metals, nuclear wastes, pesticides, green house gases, and hydrocarbons. Remediation of polluted sites using microbial process (bioremediation) has proven effective and reliable due to its eco-friendly features. Bioremediation can either be carried out ex situ or in situ, depending on several factors, which include but not limited to cost, site characteristics, type and concentration of pollutants. Generally, ex situ techniques apparently are more expensive compared to in situ techniques as a result of additional cost attributable to excavation. However, cost of on-site installation of equipment, and inability to effectively visualize and control the subsurface of polluted sites are of major concerns when carrying out in situ bioremediation. Therefore, choosing appropriate bioremediation technique, which will effectively reduce pollutant concentrations to an innocuous state, is crucial for a successful bioremediation project. Furthermore, the two major approaches to enhance bioremediation are biostimulation and bioaugmentation provided that environmental factors, which determine the success of bioremediation, are maintained at optimal range. This review provides more insight into the two major bioremediation techniques, their principles, advantages, limitations and prospects.

Bioreactor-based bioremediation of hydrocarbon-polluted Niger Delta marine sediment, Nigeria.

Crude oil-polluted marine sediment from Bonny River loading jetty Port Harcourt, Nigeria was treated in seven 2.5 l stirred-tank bioreactors designated BNPK, BNK5, BPD, BNO(3), BUNa, BAUT, and BUK over a 56-day period. Five bioreactors were biostimulated with either K(2)HPO(4), NH(4)NO(3), (NH(4))(2)SO(4), NPK, urea or poultry droppings while unamended (BUNa) and heat-killed (BAUT) treatments were controls. For each bioreactor, 1 kg (wet weight) sediment amended with 1 l seawater were spiked with 20 ml and 20 mg of crude oil and anthracene which gave a total petroleum hydrocarbons (TPH) range of 106.4-116 ppm on day 0. Polycyclic aromatic hydrocarbons (PAH) in all spiked sediment slurry ranged from 96.6 to 104.4 ppm. TPH in each treatment was ≤14.9 ppm while PAH was ≤6.8 ppm by day 56. Treatment BNO(3) recorded highest heterotrophic bacterial count (9.8 × 10(8) cfu/g) and hydrocarbon utilizers (1.15 × 10(8) cfu/g). By day 56, the percentages of biodegradation of PAHs, as measured with GC-FID were BNK5 (97.93%), BNPK (98.38%), BUK (98.82%), BUNa (98.13%), BAUT (93.08%), BPD (98.92%), and BNO(3) (98.02%). BPD gave the highest degradation rate for PAH. TPH degradation rates were as follows: BNK5 (94.50%), BNPK (94.77%), BUK (94.10%), BUNa (94.77%), BAUT (75.04%), BPD (95.35%), BNO(3) (95.54%). Fifty-six hydrocarbon utilizing bacterial isolates obtained were Micrococcus spp. 5 (9.62%), Staphylococcus spp. 3 (5.78%), Pseudomonas spp. 7 (13.46%), Citrobacter sp. 1 (1.92%), Klebsiella sp. 1 (1.92%), Corynebacterium spp. 5 (9.62%), Bacillus spp. 5 (9.62%), Rhodococcus spp. 7 (13.46%), Alcanivorax spp. 7 (13.46%), Alcaligenes sp. 1 (1.92%), Serratia spp. 2 (3.85%), Arthrobacter spp. 7 (13.46%), Nocardia spp. 2 (3.85%), Flavobacterium sp. 1 (1.92%), Escherichia sp. 1 (1.92%), Acinetobacter sp. 1 (1.92%), Proteus sp. 1 (1.92%) and unidentified bacteria 10 (17%). These results indicate that the marine sediment investigated is amenable to bioreactor-based bioremediation and that abiotic factors also could contribute to hydrocarbon attenuation as recorded in the heat-killed (BAUT) control.

Monitoring of microbial hydrocarbon remediation in the soil.

Bioremediation of hydrocarbon pollutants is advantageous owing to the cost-effectiveness 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. Provided the polluted soil has requisite values for environmental factors that influence microbial activities and there are no inhibitors of microbial metabolism, there is a good chance that there will be a viable and active population of hydrocarbon-utilizing microorganisms in the soil. Microbial methods for monitoring bioremediation of hydrocarbons include chemical, biochemical and microbiological molecular indices that measure rates of microbial activities to show that in the end the target goal of pollutant reduction to a safe and permissible level has been achieved. Enumeration and characterization of hydrocarbon degraders, use of micro titer plate-based most probable number technique, community level physiological profiling, phospholipid fatty acid analysis, 16S rRNA- and other nucleic acid-based molecular fingerprinting techniques, metagenomics, microarray analysis, respirometry and gas chromatography are some of the methods employed in bio-monitoring of hydrocarbon remediation as presented in this review.