Bioremediation of volatile oil hydrocarbons by epiphytic bacteria associated with American grass (Cynodon sp.) and broad bean (Vicia faba) leaves

Fresh leaves of American grass and broad beans grown in pristine soil were naturally colonized with cultivable volatile oil hydrocarbon-utilizing bacteria, whose numbers increased significantly in plants grown in oily soil. According to their 16S rRNA gene sequences those bacteria were affiliated to various species of the genera Rhodococcus and Pseudomonas. Qualitative growth studies revealed that pure cultures of these phyllospheric bacteria could grow successfully on a solid mineral medium containing individual alkanes with chain lengths of C₉ through C₄₀ and the aromatics phenanthrene, naphthalene, and biphenyl as sole sources of carbon and energy. Quantitative measurements showed that the individual pure bacterial isolates degraded between about 20 and 30% of crude oil, n-hexadecane, or phenanthrene in batch culture after a one-week incubation. These results reflect the high hydrocarbon degradation potential of those bacteria. The isolates were diazotrophic (nitrogen fixers), meaning that they were self-dependent in covering their nitrogen requirements. Incubating fresh leaves in closed microcosms containing volatile oil hydrocarbons resulted in up to more than 80% attenuation of these compounds after two weeks. Experimental evidence was provided that the leaf tissues did not contribute to this attenuation, which was exclusively due to the bacterial activity.