Evaluation of Screening Methods for the Isolation of Biosurfactant Producing Marine Bacteria

Hemolytic assay, bacterial adhesion to hydrocarbons (BATH) assay, drop collapse assay, oil spreading assay, emulsification assay and surface tension measurement were compared for their reliability and ease of use to screen biosurfactant producing bacteria. 105 bacterial strains were screened for their biosurfactant production. Results from screening methods revealed that hemolytic and BATH assays are not reliable methods to check biosurfactant production because hemolytic assay relies on lysis of blood cells, which can be caused by compounds produced by microbes other than the biosurfactants. In that case hemolytic assay may include strains that do not produce biosurfactant. BATH assay relies on hydrophobicity or adherence of cells with hydrophobic compounds. Results obtained in BATH assay for three bacterial isolates, Branhamella catarrhalis, Citrobacter intermedius and Klebsiella ozaenae were positive, but when the same isolates screened for drop collapse, oil spreading, and surface tension measurement the results were negative. This indicates that these bacterial cells are either acting as biosurfactant themselves (cell surface) or they don’t have the ability to produce biosurfactants. In conclusion, drop collapse and oil spreading assays are reliable methods to screen large number of samples. However, for confirmation surface tension measurement can be used at the end. Another interesting finding in this study was the use of crude oil as hydrophobic substrate for the isolation of biosurfactant producing bacterial strains; this approach may be useful during the initial isolation of biosurfactant producing bacteria to reduce the number of strains to be screened. Evaluation of Screening Methods for the Isolation of Biosurfactant Producing Marine Bacteria