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A halothermotolerant Gram-positive spore-forming bacterium was isolated from petroleum reservoirs in Iran and identified as Bacillus licheniformis sp. strain ACO1 by phenotypic characterization and 16S rRNA analysis. It showed a high capacity for bioemulsifier production and grew up to 60 degrees C with NaCl at 180 g l(-1). The optimum NaCl concentration, pH and temperature for bioemulsifier production were 4% (w/v), 8.0, and 45 degrees C, respectively. Although ACO1 did not utilize hydrocarbons, it had a high emulsifying activity (E (24) = 65 +/- 5%) on different hydrophobic substrates. Emulsification was optimal while growing on yeast extract as the sole carbon source and NaNO(3) as the nitrogen source. The efficiency of the residual oil recovery increased by 22% after in situ growth of B. licheniformis ACO1 in a sand-pack model saturated with liquid paraffin.

In this study we investigated the phenanthrene degradation by a halophilic consortium obtained from a saline soil sample. This consortium, named Qphe, could efficiently utilize phenanthrene in a wide range of NaCl concentrations, from 1% to 17% (w/v). Since none of the purified isolates could degrade phenanthrene, serial dilutions were performed and resulted in a simple polycyclic aromatic hydrocarbon (PAH)-degrading culture named Qphe-SubIV which was shown to contain one culturable Halomonas strain and one unculturable strain belonging to the genus Marinobacter. Qphe-SubIV was shown to grow on phenanthrene at salinities as high as 15% NaCl (w/v) and similarly to Qphe, at the optimal NaCl concentration of 5% (w/v), could degrade more than 90% of the amended phenanthrene in 6 days. The comparison of the substrate range of the two consortiums showed that the simplified culture had lost the ability to degrade chrysene but still could grow on other polyaromatic substrates utilized by Qphe. Metabolite analysis by HPLC and GC-MS showed that 2-hydroxy 1-naphthoic acid and 2-naphthol were among the major metabolites accumulated in the Qphe-SubIV culture media, indicating that an initial dioxygenation step might proceed at C1 and C2 positions. By investigating the growth ability on various substrates along with the detection of catechol dioxygenase gene, it was postulated that the uncultured Marinobacter strain had the central role in phenanthrene degradation and the Halomonas strain played an auxiliary role in the culture by utilizing phenanthrene metabolites whose accumulation in the media could be toxic.

Biodegradation of petroleum compounds in saline environments seems intricate and needs more attention. In this study, tetracosane was used to enrich alkane-degrading bacteria from oil-contaminated saline soils. Among the isolates, strain Qtet3, with the highest 16s rRNA gene sequence similarity to Alcanivorax dieselolei B-5(T), was able to grow at a wide range of NaCl concentrations and was shown by GC analysis to degrade more than 90% of tetracosane in 10 days. This strain has at least two alkB genes and could grow on crude oil and diesel fuel, and utilize various pure aliphatic hydrocarbon substrates (from C(12) to C(34)). Highly hydrophobic cell surfaces and lack of significant surface tension reduction in the media suggest that the main mechanism of the cells for accessing substrate is to attach directly to hydrocarbon particles. Application of this strain for remediating crude oil-contaminated soils irrigated with defined saline water demonstrated that this halotolerant bacterium could survive and grow in saline soils irrigated with NaCl solutions up to 5% w/v, with the highest hydrocarbon degradation of 26.1% observed at 2.5% NaCl. This strain is promising for future industrial applications especially in bioremediation of saline soils and wastes.

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