Understanding Polycyclic Aromatic Hydrocarbon Degradation by Soil Microbial Communities:
The Concentration and Distribution of Aromatic Dioxygenase Genes
Faculty Chair: Dr. Henry Spratt
The ability to individually identify and enumerate bacterial genes in the environment is an important development in the field of microbial ecology. Researchers are now able to quantify and map populations and communities, the presence of which could only be inferred until recently. Knowledge of this ecology is critical to understanding microbial responses to environmental pollutants, thereby contributing information useful in devising and monitoring bioremediation strategies. The following thesis therefore compares two aromatic dioxygenase genes, nidA and nahAc, which confer the ability for the bacteria carrying the genes to metabolize polycyclic aromatic hydrocarbons (PAHs), persistent pollutant byproducts of coal coking. The deposition of PAHs influences the development of soil microbiological communities, creating selective pressure for the development of microbial communities able to survive. To confirm that the mineralization of PAHs correlates with gene copy number, 14C radiorespireometery was used to detect the mineralization of radiolabled phenanthrene. Due to culture bias, the naphthalene-targeting Pseudomonas proxy gene nahAc was the first discovered and the longest studied. The degradation of larger, more persistent PAHs has been found to rely on the Mycobacterium- specific gene nidA. Soil sampling was conducted at three locations in the Chattanooga region during the summer of 2007. The samples were analyzed using quantitative real-time PCR, and the distribution patterns of nidA and nahAc in copies/gDry/gCarbon were mapped and compared. Differences in gene distribution and concentration were examined between genes as a broad survey, and between sites to examine response to PAH exposure. Prior exposure, indicated by active14C phenanthrene mineralization, was determined by radiorespireometry. The nidA gene was locally predominant over nahAc (p<0.01), and was detected at every sample site, while nahAc was much more variable in concentration, with no detection in 17 of 22 samples. The nidA gene showed an elevated response to prior exposure, occurring at higher levels in the Chattanooga Creek Superfund site than in sites with no history of industrial activity (p<0.05). Additionally, nidA copy number had a weakly positive correlation with the mineralization of 14C phenanthrene (r=0.5594, p<0.1). This establishes nidA as a more reliable indicator than nahAc of the presence of dioxygenase activities under current soil conditions. Mapping the concentration and distribution of a reliable and responsive member of the soil community using GIS technology provides a concise illustration of the current distribution of targeted genes and their response to PAH contamination. Advances in the understanding of microbial ecology underlying contaminated sites may lead to faster, more accurately targeted remediation strategies.