Klebseilla Oxytoca; An Efficient Pyrene-Degrading Bacterial Strain Isolated from Petroleum Contaminated Soil

Polycyclic aromatic hydrocarbons (PAHs) are the hazardous xenobiotic agents of oil production. One of the methods to eliminate hazardous compounds is bioremediation, which is the most efficient and cost-effective method to eliminate the harmful byproducts of crude petroleum processing. In this study, five pure bacterial isolates were isolated from petroleum-contaminated soil, four of which showed a robust growth on pyrene as a sole carbon source. Various methods viz mass spectroscopy, biochemical assays, and 16s RNA sequencing employed to identify the isolates ascertained the consistent identification of Klebsiella oxytoca by all three methods. Scanning electron microscopy and Gram staining further demonstrated the characterization of the K. oxytoca. High-performance liquid chromatography of the culture supernatant of K. oxytoca grown in pyrene containing media showed that the cells started utilizing pyrene from the 6th day onwards and by the 14th day of growth 3/4th of the pyrene was completely degraded. Genome search for the genes predicted to be involved in pyrene degradation using Kyoto Encyclopedia of Genes and Genomes (KEGG) confirmed their presence in the genome of K. oxytoca. These results suggest that K. oxtoca would be a suitable candidate for removing soil aromatic hydrocarbons.

Low-cost production and application of lipopeptide for bioremediation and plant growth by Bacillus subtilis SNW3

At present time, every nation is absolutely concern about increasing agricultural production and bioremediation of petroleum-contaminated soil. Hence, with this intention in the current study potent natural surfactants characterized as lipopeptides were evaluated for low-cost production by Bacillus subtilis SNW3, previously isolated from the Fimkessar oil field, Chakwal Pakistan. The significant results were obtained by using substrates in combination (white beans powder (6% w/v) + waste frying oil (1.5% w/v) and (0.1% w/v) urea) with lipopeptides yield of about 1.17 g/L contributing 99% reduction in cost required for medium preparation. To the best of our knowledge, no single report is presently describing lipopeptide production by Bacillus subtilis using white beans powder as a culture medium. Additionally, produced lipopeptides display great physicochemical properties of surface tension reduction value (SFT = 28.8 mN/m), significant oil displacement activity (ODA = 4.9 cm), excessive emulsification ability (E24 = 69.8%), and attains critical micelle concentration (CMC) value at 0.58 mg/mL. Furthermore, biosurfactants produced exhibit excellent stability over an extensive range of pH (1–11), salinity (1–8%), temperature (20–121°C), and even after autoclaving. Subsequently, produced lipopeptides are proved suitable for bioremediation of crude oil (86%) and as potent plant growth-promoting agent that significantly (P < 0.05) increase seed germination and plant growth promotion of chili pepper, lettuce, tomato, and pea maximum at a concentration of (0.7 g/100 mL), showed as a potential agent for agriculture and bioremediation processes by lowering economic and environmental stress. Graphical Abstract

Soil Microcosms for Bioaugmentation With Fungal Isolates to Remediate Petroleum Hydrocarbon-contaminated Soil

The aim of this work was to isolate indigenous PAH degrading-fungi from petroleum contaminated soil and exogenous ligninolytic strains from decaying-wood, with the ability to secrete diverse enzyme activity. A total of ten ligninolytic fungal isolates and two native strains, has been successfully isolated, screened and identified. The phylogenetic analysis revealed that the indigenous fungi (KBR1 and KB8) belong to the genus Aspergillus niger and tubingensis. While the ligninolytic exogenous PAH-degrading strains namely KBR1-1, KB4, KB2 and LB3 were affiliated to different genera like Syncephalastrum sp, Paecilomyces formosus, Fusarium chlamydosporum, and Coniochaeta sp., respectively. Basis on the taxonomic analysis, enzymatic activities and the hydrocarbons removal rates, single fungal culture employing the strain LB3, KB4, KBR1 and the mixed culture (LB3+KB4) were selected to be used in soil microcosms treatments. The Total petroleum hydrocarbons (TPH), fungal growth rates, BOD5/COD ratios and GC-MS analysis, were determined in all soil microcosmos treatments (SMT) and compared with those of the control (SMU). After 60 days of culture incubation, the highest rate of TPH degradation was recorded in SMT[KB4] by approximately 92±2.35% followed by SMT[KBR1] then SMT[LB3+KB4] with 86.66±1.83% and 85.14±2.21%, respectively.

Bioengineered Approach for Ex situ Vermiextraction of Acenaphthylene, Benzo (E)Pyrene and Benzo (Ghi) Perylene Soil Contamination

The ex situ study of vermiextraction of Acenaphthylene (AcPY), Benzo(e)pyrene (BeP) and Benzo(ghi)perylene (BP) form constructed vermiculture containing petroleum contaminated soil (8.00±0.01, 9.80±0.00 and 5.02±0.00 mg/kg respectively) and vermiaccumulation (AcPY, 1.05±0.00, BeP, 2.01±0.00 and BP, 1.73±0.00 mg/kg) by Esenia fetida squirms with mean vermiremoval efficiency of 100% while vermiconversions were AcPY, 86.88, BeP, 79.49 and BP, 65.54%. The identification and quantification of the 3 polycyclic aromatic hydrocarbons (3PAHs) were performed by GS/MD in accordance with analytical procedure of US. EPA 8270; 625. The bioengineered approach by E. fetida squirms in the vermiculture proved effective to detoxify and remove the persistent organic pollutants of the 3PAHs. The ex situ study of vermiextraction of Acenaphthylene (AcPY), Benzo(e)pyrene (BeP) and Benzo(ghi)perylene (BP) form constructed vermiculture containing petroleum contaminated soil (8.00±0.01, 9.80±0.00 and 5.02±0.00 mg/kg respectively) and vermiaccumulation (AcPY, 1.05±0.00, BeP, 2.01±0.00 and BP, 1.73±0.00 mg/kg) by Esenia fetida squirms with mean vermiremoval efficiency of 100% while vermiconversions were AcPY, 86.88, BeP, 79.49 and BP, 65.54%. The identification and quantification of the 3 polycyclic aromatic hydrocarbons (3PAHs) were performed by GS/MD in accordance with analytical procedure of US. EPA 8270; 625. The bioengineered approach by E. fetida squirms in the vermiculture proved effective to detoxify and remove the persistent organic pollutants of the 3PAHs.

Paradesulfitobacterium ferrireducens gen. nov., sp. nov., a Fe(III)-reducing bacterium from petroleum-contaminated soil and reclassification of Desulfitobacterium aromaticivorans as Paradesulfitobacterium aromaticivorans comb. nov.

A strictly anaerobic bacterium, strain PLL0T, was isolated from petroleum-contaminated soil sampled in Gansu Province, PR China. Cells were rods, non-motile and Gram-stain-positive. The strain grew at 25–37 °C (optimum, 30 °C) in the presence of 0–3 % (w/v) NaCl (optimum, 2 %). Strain PLL0T was able to reduce ferrihydrite, Fe(III) citrate and thiosulphate. The 16S rRNA gene analysis revealed that this strain clustered with the genus Desulfitobacterium , and showed highest similarity to Desulfitobacterium aromaticivorans UKTLT (95.4 %) followed by Desulfitobacterium chlororespirans Co23T (93.9 %). However, strains PLL0T and UKTLT showed no more than 94.0 % similarity to other species of the genus Desulfitobacterium , and formed an independent group in the phylogenetic tree. The average nucleotide identity (ANI) and digital DNA–DNA hybridization (dDDH) values between strain PLL0T and Desulfitobacterium species (except for D. aromaticivorans ) were 67.4–68.5 % and 12.6–12.7 %, respectively, which are far below the threshold for delineation of a new species. Based on ANI, dDDH, average amino acid identity, phylogenetic analysis and physiologic differences from the previously described taxa, we suggest that strain PLL0T represents a novel species of a novel genus, for which the name Paradesulfitobacterium ferrireducens gen. nov. sp. nov. is proposed. The type strain is PLL0T (=MCCC 1K05549=KCTC 25248). We also propose the reclassification of D. aromaticivorans as Paradesulfitobacterium aromaticivorans comb. nov.