AEM Accepts, published online ahead of print on 6 April 2007

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Appl. Environ. Microbiol. doi:10.1128/AEM.01372-06
Copyright (c) 2007, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Microbial Diversity in Natural Asphalts of the Rancho La Brea Tar Pits

Jong-Shik Kim and David E. Crowley^*

Dept. Environmental Sciences, University of California, Riverside, CA

^* To whom correspondence should be addressed. Email: Crowley{at}ucr.edu.

Bacteria commonly inhabit subsurface oil reservoirs, but almost nothing is known yet about microorganisms that live in naturally occurring terrestrial oil seeps and natural asphalts that are comprised of highly recalcitrant petroleum hydrocarbons. Here, we report the first survey of microbial diversity in ca 14,000 year old samples of natural asphalts from the Rancho La Brea Tar Pits in Los Angeles, California. Microbiological studies included analyses of 16S rRNA gene sequences and DNA encoding aromatic ring hydroxylating dioxygenases from two tar pits differing in chemical composition. Our results revealed a wide range of phylogenetic groups within the Archaea and Bacteria domains, in which individual taxonomic clusters were comprised of sets of closely related species within novel genera and families. Fluorescent staining of asphalt-soil particles using phylogenetic probes for Archaea, Bacteria, and Pseudomonas showed coexistence of mixed microbial communities at high cell densities. Genes encoding dioxygenases included three novel clusters of enzymes. The discovery of life in the tar pits provides an avenue for further studies on the evolution of enzymes, and catabolic pathways for bacteria that have been exposed to complex hydrocarbons for millennia. These bacteria also should have application for industrial microbiology and bioremediation.

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