Novel bacterial groups dominate in a thermophilic methanogenic hexadecane-degrading consortium

A methanogenic hexadecane-degrading consortium designated SK originating from the Shengli oil field was cultured at 55 degrees C. The structure and dynamics of the microbial community during successive transfers were examined using terminal restriction fragment length polymorphism (T-RFLP) fingerprinting and sequencing of 16S rRNA gene fragments. The archaeal community was mainly composed of hydrogenotrophic methanogens affiliated with Methanothermobacter crinale and acetoclastic methanogens related to Methanosaeta thermophila. Over four-fifths of the bacterial clones in the hexadecane-degrading subcultures exhibited < 90% similarity to sequences of known type strains, and clones were mainly grouped into unclassified bacteria (66.3-66.7%), Firmicutes (9.6-10.6%), Thermotogae (7.0-7.7%), and Nitrospira (5.3-5.8%). The dominant operating taxonomic unit (OTU) (41.3-43.0% of all clones) representing terminal restriction fragment (T-RF) 125 bp exhibited only 82.6% sequence similarity to Thermotoga maritime and clustered in a monophyletic, deep-branching lineage (designated Shengli cluster). Two other OTUs (T-RFs 66 and 67 bp) were assigned to uncultured members of the candidate phylum OP8 and Firmicutes, respectively. These novel bacterial assemblages are likely to be involved in the process of hexadecane degradation because of their high abundance in the enrichments. These result substantially expand the knowledge of the extent of bacterial diversity associated with the anaerobic degradation of alkanes under thermophilic conditions.