Solid medium for the direct isolation of bacterial colonies growing with polycyclic aromatic hydrocarbons or 2, 4, 6-trinitrotoluene (TNT) as the sole carbon source

Isolation of hydrocarbon-degrading bacteria is a key step for the study of microbiological diversity, metabolic pathways and bioremediation, however current methods lack simplicity and versatility. We developed an easy method that allows the screening and isolation of bacterial colonies capable of degrading hydrocarbons, such as diesel or polycyclic aromatic hydrocarbons (PAHs), as well as the pollutant explosive, 2, 4, 6-trinitrotoluene (TNT). The method uses a two-layer solid medium, with a layer of M9 medium, with a second layer containing the carbon source deposited trough the evaporation of ethanol. Using this medium we grew hydrocarbon-degrading strains, using diesel, phenanthrene or anthracene as the sole carbon sources, as well as three TNT-degrading isolates. Using this medium we isolated PAHs-degrading bacterial colonies directly from diesel polluted soils. Analysis revealed that bacteria grown in medium using PAHs as carbon source maintain their morphological characteristics when compared to cells grown on traditional media with glucose.