Novel aromatic ring-hydroxylating dioxygenase genes detected in coastal marine sediments of patagonia

LOZADA, M.; RIVA MERCADAL, J.P.; DI MARZIO, W.; FERRERO, M.; DIONISI, H.

Buenos Aires, Argentina

Congreso; IV Congreso Argentino de Microbiología General (SAMIGE 2007); 2007

Sociedad Argentina de Microbiología General (SAMIGE)

Polycyclic aromatic hydrocarbons (PAHs) are organic compounds composed of two or more fused aromatic rings, many of them considered toxic, mutagenic and/or carcinogenic. Due to their low water solubility, PAHs tend to adsorb to sediment particles, decreasing their bioavailability and affecting rates of biodegradation by indigenous bacteria. In the Patagonian coast, the diversity of these PAH-degrading bacterial populations, as well as the environmental factors affecting their biodegradation rate, remains largely unknown. The aim of this work was to explore the diversity of PAH-degrading microorganisms in coastal sediments of Patagonia. We used a functional culture-independent approach, and specific catabolic genes as the target. Intertidal sediment samples were collected at different sites along the Patagonian coastline. Gene fragments encoding the catalytic alpha subunit of aromatic ring-hydroxylating dioxygenases (ARHDs), the enzyme responsible for the first step of the aerobic biodegradation of PAHs, were amplified from DNA isolated from intertidal sediment samples, and cloned. In seven catabolic gene libraries, 362 out of 479 analyzed clones were identified as alpha-subunit ARHD gene fragments. These clones were screened by restriction fragment length polymorphism (RFLP) analysis, and randomly chosen for sequencing. Libraries contained one to five ARHD types, the more diverse library corresponding to the sample with a greater number of PAHs. Overall, seven different dioxygenase gene types were detected, only two of them closely related to previously described ARHD genes: archetypical nahAc-like genes (Pseudomonas type) and phnAc-like genes as identified in Alcaligenes faecalis AFK2. The other five gene types formed deeply rooted branches (less than 70% identity at the amino acid level) with previously described ARHD sequences. Importantly, they contain the consensus pattern of bacterial ring-hydroxylating dioxygenase alpha-subunits, suggesting that these gene fragments are novel types of ARHDs. These results suggest that a number of yet unidentified PAH-degrading bacteria may be present in coastal sediments from Patagonia. Further experiments are needed to identify the key players for PAH biodegradation in cold-temperate polluted marine ecosystems. e-mail: hdionisi@cenpat.edu.ar