Background: Biosurfactants are biocompatible surface-active compounds with environmental and industrial applications. Besides, biosurfactants are not cost-competitive to their chemical counterparts. Cost effective technology such as the use of low-cost substrates is a promising approach to reduce the production cost. Objective: Production of biosurfactants by the novel strain Rhodococcus sp. SP1d was monitored by using different low-cost substrates. Moreover, their possible exploitation in the bioremediation field or as biocontrol agent was evaluated. Methods: 16S rDNA gene analysis was performed. Growth curves using n-hexadecane, diesel, exhausted oil, motor oil and butter was carried out. Biosurfactant production and activity was tested by oil-displacement assay and emulsification activity. The biofilm production was evaluated by MBEC Assay® Kit. Chemical structure of biosurfactants was investigated by Nuclear Magnetic Resonance spectroscopy. Results: Rhodococcus sp. SP1d evidence the ability to grow and biosynthesize surfactants using exhausted vegetable oil, mineral oil, butter, n-hexadecane, and diesel. The maximum production of crude biosurfactants was reached after 168 h of on n-hexadecane and diesel with a final yield of approximately 2.48 and 1.75 g L-1 respectively. SP1d produced biosurfactants on diesel when grown at 10 and 18°C maintaining the activity over a wide range of NaCl concentration, pH, and temperatures. The emulsification activity of the crude biosurfactant at 1000 mg/l was 55% towards xylene and olive oil. Nuclear Magnetic Resonance spectroscopy indicated that the biosurfactant is formed by trehalolipid. Biosurfactants extracted from Sp1d enhanced the biofilm production of fungal-antagonistic P. protegens MP12. Conclusion: The use of low-cost substrates reduces the cost of biosurfactant synthesis, and the environmental pollution due to their inappropriate disposal. The high production at different temperature, stability and emulsification properties using diesel as substrate, make it a sustainable biocompound for bioremediation purpose. Eventually, this biosurfactant improve the adherence to plant surfaces enhancing the antifungal activity of MP12
