ABSTRACT:The isolation and characterization of hydrocarbon-utilizing bacteria from petroleum oily sludge collected from crude oil processing plant in Rivers State, Nigeria was carried out. Microbiological analysis of the sludge sample showed that the microbial load consisted average of 2.5 x 10 6 cfu/g total heterotrophic bacterial (THB) count, while the hydrocarbon utilizing bacterial (HUB) count was average of 2.0 x 10 3 cfu/g. Phenotypic characteristics and phylogenetic analysis of the 16S rRNA gene sequence of the isolates revealed that they were related to members of the species Klebsiella pneumonia, Klebsiella oxytoca and Alcaligenes faecalis. The 16S rRNA of the isolates showed percentage similarities to the type strains (99% sequence similarities) of the following; Klebsiella pneumonia strain B21 (gi: 922317936), Klebsella pneumonia strain ICB-C183 (gi: 908478837), Klebsella oxytoca strain BCNAI (gi: 846993354), Klebsella oxytoca strain BC4 (gi: 884060138), Alcaligenes faecalis strain IOU PMR (gi: 686028963) and Alcaligenes faecalis strain AQ-I (gi: 816845513). Of the six hydrocarbon utilizing bacteria identified, 4(67%) were Klebsiella species while 2(33%) were identified as Alcaligenes sp of different strains. The result suggest that these isolated organisms from the petroleum sludge can be explored and used to promote environmentally friendly technology clean-up of petroleum hydrocarbon polluted sites in the Niger Delta region of Nigeria.© JASEM https://dx.doi.org/10.4314/jasem.v21i2.17

The study was aimed at exploring the use of cow dung (CD) and sawdust (SD) as bio stimulants for enhanced ex-situ bioremediation of diesel fuel contaminated soil. Results of the microbial populations in a gram of soil showed that diesel polluted soil amended with CD had the highest total culturable heterotrophic bacterial (TCHB) population range (1.42 x 10 6-1.56 x 10 7 Cfu/g), total fungal (TF) (1.0 x 10 6-1.60x10 7 Cfu/g), hydrocarbon utilizing bacteria (HUB) (4.5 x 10 6-2.05 x 10 7 Cfu/g) and hydrocarbon utilizing fungi (HUF) (3.5 x 10 6-1.3 x 10 7 Cfu/g) respectively during the study. Diesel polluted soil amended with SD had lower TCHB populations (1.32 x 10 6-2.0 x 10 7 Cfu/g), TF (6.0 x 10 6-3.6 x 10 7 Cfu/g), HUB (5.4 x 10 6-1.28 x 10 7 Cfu/g) and HUF (1.2 x 10 6-2.0 x 10 6 Cfu/g) respectively, while the microbial populations in option C (control) were TCHB (1.

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.

Petroleum oil sludge resulting from crude oil storage, illegal crude oil refining and bunkering activities constitutes environmental hazards and pollution in the crude oil producing communities in the Niger Delta region of Nigeria. Biostimulation with N.P.K. fertilizer option C, bioargumentation with indigenous hydrocarbon utilizing bacteria (HUB) option B, combination of biostimulation and bioaugmentation as option A and option D was without any bioremediation treatment were employed in the bioremediation of brackish water artificially polluted with petroleum oil sludge. Brackish water sample was obtained from Elechi Creek, Port Harcourt Rivers State. Petroleum oil sludge sample was obtained from Crude Oil Processing Plant in Obegi community, Rivers State. Bioremediation was monitored for 56 days using the percentage ratio of total petroleum hydrocarbon (TPH) losses for each period to TPH loss at day 0. The result of physicochemical analysis of the petroleum sludge showed that aliphatic hydrocarbon (n-alkanes) ranged from C 13 -C 35, with concentrations ranging from 26.12-7,713.62ppmwith TPH of 89,509.9ppm. The polycyclic aromatic hydrocarbon (PAH) range was 0.03-5.36ppm with total concentration of 24.21ppm. Heavy metal analysis showed; iron (49.42mg/kg), Zinc (3.79mg/kg), Nickel (4.53 mg/kg) and manganese (6.90 mg/kg). The average total heterotrophic bacterial (THB) and (HUB) counts for petroleum sludge were; 2.5 x 10 5 cfu/g and 2.0 x10 5 cfu/g and for the brackish water sample were 1.39 x 106cfu/ml and 1.1 x 10 4 cfu/ml respectively. Statistical analysis (ANOVA) showed that the THB and HUB counts were significantly different at 5 percent levels (P<0.05) in the different treatment options during the bioremediation period. Changes in physico-chemical parameters showed that pH, alkalinity, conductivity, chemical oxygen demand, nitrate and phosphate were significantly different (P<0.05) while there were no significant differences (P>0.05) in the following parameter; salinity biochemical oxygen demand and total hydrocarbon continent.Using least significant difference (LSD), treatment option D and the control option E were different from treatments A, B and C. The phylogenetic analysis identification of the HUB isolates implicated in the degradation process revealed a closely related ness to the following organisms, Lysinibacillus sphaericus, Klebsiella pneumonia and Alcaligenes faecalis of different strains. The bacterial sequences submitted to Genbank were assigned Accession Number KX817218-KXV7225. The percentage losses in TPH from Gas Chromatography (GC) results showed the following; option A (91.8%), option B (92.5%), C (95%) D (57.8%) and option E control (39.5%) respectively. The results suggest that the application of biostimulation with N.P.K fertilizer, bioaugmentation with indigenous HUB or a combination of both will enhance the bioremediation of petroleum sludge polluted brackish water system in the Niger Delta of Nigeria.