Characterization of the Aerobic Hydrocarbon-Oxidizing Enrichments from a High-Temperature Petroleum Reservoir by Comparative Analysis of DNA- and RNA-Derived Clone Libraries

Enrichment cultures of aerobic hydrocarbon-oxidizing bacteria obtained from the injection and production wells of the Dagang oil field (China) were studied by molecular biological and microbiological methods. This work is the first to report simultaneous isolation of DNA and RNA from enrichment cultures of microorganisms from oil strata with further construction of clone libraries of 16S rRNA genes and 16S crDNA (complementary rDNA). Comparative analysis of the DNA- and RNA-derived clone libraries made it possible to determine the total genomic diversity of microorganisms, as well as to reveal metabolically active microorganisms in these cultures. Phylotypes of bacteria of the genus Geobacillus were found to be dominant in the DNA and RNA clone libraries of the enrichment cultures from the production well. Phylotypes of bacteria belonging to Geobacillus, Pseudomonas, Tepidiphilus, and other genera were detected in the DNA and RNA libraries obtained from the culture from the injection well. Phylotypes of bacteria of the genus Geobacillus were predominant in the RNA library and represented the second-largest group (after pseudomonads) in the DNA library. In the RNA libraries of the alkB genes of both enrichments, three homologs close to alkB-geo1, alkB-geo2, and alkB-geo4 of bacteria of the genus Geobacillus were detected. The occurrence pattern of the alkB transcripts, ribosomal RNA, and the 16S rRNA genes of bacteria of the genus Geobacillus indicates the predominance and functional activity of geobacilli in the enrichment cultures of hydrocarbon-oxidizing bacteria from high-temperature petroleum reservoir.