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theses

At deep-sea hydrothermal vents, the flux of energy is mediated by microbial oxidations, through the conversion of the chemical energy stored in reduced compounds (e.g., sulfide, hydrogen, hydrocarbons) into biochemical energy. Natural hydrocarbons are largely formed by the thermal decomposition of organic matter (thermogenesis) or by microbial processes. However, hydrocarbons can also have an abiotic origin and may form, in hydrothermal systems, by water-rock interactions, for example involving Fisher-Tropsch reactions and the serpentinization of ultramafic rocks. Despite the observation that hydrocarbons are enriched in deep-sea hydrothermal vents, our knowledge of the taxonomic and functional diversity of alkane-oxidizing bacteria from these environments remains very limited. In this dissertation, I investigated the diversity of alkane oxidizing bacteria and the genes alkB, CYP153 and almA by performing enrichment cultures for hydrocarbon oxidizing microorganisms from fluid samples and biomass collected from experimental microcolonizers that were deployed on diffuse flow vents on the East Pacific Rise at 9°N and in the Guaymas Basin. These enrichments led to the successful isolations of pure cultures of aerobic, mesophilic organisms capable of using n-alkanes as their carbon sources. Our isolates were, for the most part, Gammaproteobacteria of the genus Acinetobacter and Alcanivorax, but some rare occurring bacteria that were numerically relevant in the environments were also isolated. The PCR amplification of the alkB, CYP153 and almA gene fragments from these isolates, and a phylogenetic analysis of these genes was carried out. The alkB, CYP153 and almA genes encode for the alkane hydroxylase, cytochrome P450 and flavin binding monooxygenase respectively, which are enzymes that catalyze the first reaction in the stepwise oxidation of n-alkanes. Furthermore, transcripts of the alkB gene were detected in two model organisms from the laboratory culture collection, Alcanivorax sp. strain EPR 7 and Acinetobacter sp. strain EPR 111. Reverse Transcription PCR (RT-PCR) experiments showed that alkB transcripts could be detected in the presence of dodecane but not in acetate, confirming that, in these strains, the alkB gene is induced. Finally, a functional gene survey of alkB genes in vent natural microbial populations showed that the majority of the detected sequences derived from Gammaproteobacteria and Alphaproteobacteria.

ETD_4615

Ph.D.

Includes bibliographical references

Includes vita

by Ramaydalis Keddis

http://hdl.rutgers.edu/1782.1/rucore10001600001.ETD.000068893

doi:10.7282/T3VX0F3W

ETD doctoral

The author owns the copyright to this work.