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Published October 2010 | Supplemental Material + Published
Journal Article Open

Identification of Novel Methane-, Ethane-, and Propane-Oxidizing Bacteria at Marine Hydrocarbon Seeps by Stable Isotope Probing

Abstract

Marine hydrocarbon seeps supply oil and gas to microorganisms in sediments and overlying water. We used stable isotope probing (SIP) to identify aerobic bacteria oxidizing gaseous hydrocarbons in surface sediment from the Coal Oil Point seep field located offshore of Santa Barbara, California. After incubating sediment with ^(13)C-labeled methane, ethane, or propane, we confirmed the incorporation of ^(13)C into fatty acids and DNA. Terminal restriction fragment length polymorphism (T-RFLP) analysis and sequencing of the 16S rRNA and particulate methane monooxygenase (pmoA) genes in ^(13)C-DNA revealed groups of microbes not previously thought to contribute to methane, ethane, or propane oxidation. First, ^(13)C methane was primarily assimilated by Gammaproteobacteria species from the family Methylococcaceae, Gammaproteobacteria related to Methylophaga, and Betaproteobacteria from the family Methylophilaceae. Species of the latter two genera have not been previously shown to oxidize methane and may have been cross-feeding on methanol, but species of both genera were heavily labeled after just 3 days. pmoA sequences were affiliated with species of Methylococcaceae, but most were not closely related to cultured methanotrophs. Second, ^(13)C ethane was consumed by members of a novel group of Methylococcaceae. Growth with ethane as the major carbon source has not previously been observed in members of the Methylococcaceae; a highly divergent pmoA-like gene detected in the ^(13)C-labeled DNA may encode an ethane monooxygenase. Third, ^(13)C propane was consumed by members of a group of unclassified Gammaproteobacteria species not previously linked to propane oxidation. This study identifies several bacterial lineages as participants in the oxidation of gaseous hydrocarbons in marine seeps and supports the idea of an alternate function for some pmoA-like genes.

Additional Information

© 2010 American Society for Microbiology. Received 1 February 2010/Accepted 22 July 2010. Published ahead of print on 30 July 2010. We thank Michael Friedrich and Bianca Pommerenke for early assistance with the SIP technique. Heather Coleman, Blair Paul, and Cristoph Pierre collected samples, and Lichun Zhang performed the fatty acid extractions and GC/MS analyses. This work was funded by a National Science Foundation award (OCE-0447395) and a DOE award (DE-NT0005667) to D.L.V. and a National Science Foundation award (EAR-0645502) to A.L.S.

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