Hydrocarbon contamination emerging from the crude oil industrial-related activities has led to severe environmental issues. Prolonged contamination with the constant infiltration of crude oil into the soil is a severe problem in remediating contaminated soils. Hence, the current study focuses on comparing various bioremediation strategies, thereby isolating native bacteria competent to reduce TPH in both liquid and microcosm environments in an old-aged petroleum hydrocarbon contaminated soil. Assays in the modified 6SW-Vit medium after 7 days of incubation revealed that Bacillus altitudinis strain HRG-1 was highly hydrophobic and had a suitable ability to decrease surface tension (40.98%) and TPH (73.3%). The results of biodegradation in the microcosm proved that among the designated treatments, including bio-stimulated microcosm (SM), bacterialized microcosm (BM), a combined bio-stimulated microcosm and bacterialized microcosm (SB), and natural attenuation (NA), the SB treatment was the most effective in mitigating TPH (38.2%). However, the SM treatment indicated the lowest TPH biodegradation (18%). Pearson correlation coefficient among microcosm biological indicators under investigation revealed that soil basal respiration had the highest correlation with the amount of residual TPH (r = −0.73915, P < 0.0001), followed by the microbial population (r = −0.65218, P < 0.0001), catalase activity (r = 0.48323, P = 0.0028), polyphenol oxidase activity (r = −0.43842, P = 0.0075), and dehydrogenase activity (r = −0.34990, P = 0.0364), respectively. Nevertheless, considering the capability of strain HRG-1 and the higher efficiency of the combined technique, their use is recommended to diminish the concentration of petroleum hydrocarbons in hot and dry contaminated areas.

Graphical abstract

This research aimed to optimize a lipopeptide biosurfactant produced from Bacillus sp. SHA302 due to its high efficiency of heavy metal release in soil. The results demonstrated that the metal release capacity of the lipopeptide biosurfactant alone increased with increasing the biosurfactant concentration. Among treatments with different biosurfactant concentrations plus acid, the highest metal release rates of 53.8% ± 1.4 and 39.3% ± 1.7 for Zn and Pb, respectively, were observed in the critical micelle concentration (CMC) + HCl treatment. The results of a factorial experiment designed for optimizing biosurfactant production showed that among five inexpensive carbon sources and six mineral nitrogen sources, sugar beet molasses (1%) and ammonium chloride (0.1%) were the most efficient sources in lowering the surface tension (ST) of the culture media to 32.2 ± 0.76 mN/m. The second step of the experiment was a Plackett–Burman design with 11 factors and showed that the four factors of pH, ammonium chloride, magnesium sulfate, and molasses significantly affected (P < 0.05) the changes in ST and biosurfactant production. The third step of the experiment was done using the response surface methodology (RSM) with a central composite design. The results showed that a pH of 7.3, 1.5 g/l of ammonium chloride, 0.3 g/l of magnesium sulfate, and 10% of sugar beet molasses yielded values of 29.2 ± 0.71 mN/m and 5.74 ± 0.52 g/l for the two variables of ST and biosurfactant production, respectively, which reached their most optimal levels.

Adsorption of old-aged petroleum hydrocarbons to the soil solid phase, which causes biosurfactant loss of performance, is among the limiting factors for the remediation of the saline-sodic soils contaminated with petroleum. Therefore, to find a functional biosurfactant in oil-contaminated saline-sodic soils, the efficiency of 39 bacteria isolated from petroleum-contaminated soils was evaluated. The strains were cultured in the Bushnell–Haas medium, and the produced biosurfactants and bioemulsifiers in this medium were extracted using chloroform/methanol and ethyl acetate extraction methods, respectively. Their partial purification was performed by column chromatography, and eventually, their performance in releasing TPH from the contaminated soil was evaluated. The soil test results revealed that the highest TPH releases due to the effects of the biosurfactants and bioemulsifier produced from SHA302, SH21, and SH72 isolates were 42.4% ± 0.2, 21.6% ± 0.15 and 24.3% ± 0.91, respectively. Based on the 16S rRNA gene sequence, the SHA302 strain showed 93.98% phylogenetic similarity with Bacillus pumilus strain ATCC 7061. The Fourier transform infrared spectroscopy and thin-layer chromatography results proved that the biosurfactants produced by isolates SHA302, SH21 and SH72 showed lipopeptide, glycolipoprotein and glycoprotein natures, respectively. The performance of the biosurfactant produced by SHA302 isolate indicated that it could be used as a good candidate for releasing TPH from saline-sodic soils with old contamination and facilitating the degradation of hydrocarbons.