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The Aquificales are widespread in marine and terrestrial hydrothermal environments. Here, we report the complete and draft genome sequences of six new members of the Aquificales: two marine species, Persephonella marina strain EX-H1 and Hydrogenivirga strain 128-5-R1 (from the East Pacific Rise, 9°50.3′N, 104°17.5′W, and the Eastern Lau Spreading Center, 176°11.5′W, 20°45.8′S, respectively), and four terrestrial isolates, Sulfurihydrogenibium azorense strain Az-Fu1, Sulfurihydrogenibium yellowstonense strain SS-5, and Sulfurihydrogenibium strain Y03AOP1 (from Furnas, Azores, Portugal, and Calcite Springs and Obsidian Pool in Yellowstone National Park, United States, respectively), and the only thermoacidophilic isolate, Hydrogenobaculum strain Y04AAS1 (from a stream adjacent to Obsidian Pool). Significant differences among the different species exist that include nitrogen metabolism, hydrogen utilization, chemotaxis, and signal transduction, providing insights into their ecological niche adaptations.
The Aquificales are marine and terrestrial chemolithoautotrophic thermophiles. Like Aquifex aeolicus (2), Hydrogenivirga strain 128-5-R1 and Persephonella marina Ex-H1 (4) are marine isolates, and Sulfurihydrogenibium azorense Az-Fu1 (1), Sulfurihydrogenibium yellowstonense SS-5 (9), and Sulfurihydrogenibium strain Y03AOP1 were isolated from near-neutral-pH terrestrial hot springs whereas Hydrogenobaculum strain Y04AAS1 is a thermoacidophile. The terrestrial Aquificales are found in high abundance in hot spring outflow channels and play an important role in the biogeochemical cycling in these hot springs (10).
The complete (S. azorense, P. marina, Sulfurihydrogenibium strain Y03AOP1, and Hydrogenobaculum strain Y04AAS1) and draft (S. yellowstonense and Hydrogenivirga) genome sequences were generated by whole-genome random shotgun sequencing at The Institute for Genomic Research (S. azorense, P. marina, and S. yellowstonense), The Venter Institute (Hydrogenivirga), and the DOE Joint Genome Institute (Sulfurihydrogenibium strain Y03AOP1 and Hydrogenobaculum strain Y04AAS1). The genomes were sequenced and initially annotated as described in references 6 and 3 and on the DOE Joint Genome Institute Sequencing website (http://www.jgi.doe.gov/sequencing/index.html), respectively. Comparative gene prediction analysis was done using the Integrated Microbial Genomes (IMG; http://img.jgi.doe.gov/) platform (8).
The S. azorense, P. marina, Sulfurihydrogenibium strain Y03AOP1, and Hydrogenobaculum strain Y04AAS1 chromosome sequences contain about 1.64, 1.98, 1.84, and 1.56 Mbp with 1,708, 2,048, 1,774, and 1,640 putative open reading frames, respectively. P. marina has a megaplasmid (53,682 bp, 74 genes), the G+C content of the genomes varies from 31.9% (Y03AOP1) to 43% (Hydrogenivirga), and most of the genomes have two rRNA operons.
Although some members of the Aquificales can grow heterotrophically, all members are able to fix CO2 using the reductive tricarboxylic acid (TCA) cycle. Like all Aquificaceae, Hydrogenivirga and Hydrogenobaculum have the “B-type” reductive TCA cycle (7), whereas all other isolates have the “A-type” reductive TCA cycle. Although hydrogen oxidation (“knallgas” reaction) was thought to be universal in this group, several isolates such as S. yellowstonense and Sulfurihydrogenibium strain Y03AOP1 are unable to oxidize hydrogen, and consequently no hydrogenases are detected in their genomes. What seems more diagnostic of the Aquificales is their ability to oxidize sulfur, and all genomes have a suite of sox genes and a sulfide:quinone oxidoreductase (sqr), which catalyzes the oxidation of sulfide to elemental sulfur. A homolog of SreABC, a sulfur reductase in A. aeolicus (5), was found in Hydrogenivirga.
The marine isolates, P. marina and Hydrogenivirga, are also autotrophic nitrate reducers and have cytoplasmic membrane-bound nitrate reductases (Nar) and nitrite reductases (Nir), nitric oxide reductases (NOR), and nitrous oxide reductases (Nos). However, although Hydrogenobaculum strain Y04AAS1 is unable to reduce nitrate in the laboratory, it contains Nir and Nap genes. Curiously, S. azorense, Sulfurihydrogenibium strain Y03AOP1, and Hydrogenobaculum strain Y04AAS1 all code for the quinone NOR, thought to be exclusively involved in denitrification. Given their redox reactive environment in deep-sea hydrothermal sulfide deposits or along iron-sulfide chemical gradients in terrestrial hot springs, it is no surprise that all the genomes contain a large number of different cytochromes. Additionally, unlike Aquifex aeolicus, whose genome lacks chemotaxis/motility genes (2), all the genomes reported here have a suite of the bacterium-like chemotaxis genes, numerous monocyte chemoattractant proteins, and FliM. Further comparative analysis of these genomes will enable future studies of the physiological ecology and phylogenetic position of the Aquificales.
The sequences are deposited under the project identification numbers as follows (GenBank accession number): Hydrogenivirga strain 128-5-R1, 19313 (NZ_ABHJ00000000); Hydrogenobaculum strain Y04AAS1, 18891 (NC_011126); Sulfurihydrogenibium strain Y03AOP1, 18889 (NC_010730); S. azorense, 12529 (CP001229); S. yellowstonense, 19071 (ABZS00000000); P. marina, 12526 (CP001230 and CP001231).
The funding for these sequencing projects was provided by the NSF-0236856 (to A.-L.R. and J.H.), The Gordon and Betty Moore Foundation, and the Joint Genome Institute as part of the Community Sequencing Program. Sequencing, assembly, and annotation efforts for Hydrogenivirga were supported by the GBMF as part of its Marine Microbial Sequencing Project (www.moore.org/marinemicro). The JCVI software team, under the leadership of Saul A. Kravitz, manages annotation and web delivery of data for the GBMF-funded project (http://research.jcvi.org/moore).
We thank Jennifer Meneghin, Karl Rustenholtz, Isabel Ferreira, and all those who assisted in many different ways to see these genome sequences be made available to the scientific community. Many thanks in particular to Nikos Kyrpides and many others at JGI who were instrumental in facilitating the deposition of the data into IMG and data analysis. Thanks to Loren Hauser at Oak Ridge National Laboratory and the JGI annotation team for their help. We thank Jason Miller and Granger Sutton and their team for the ongoing development and maintenance of the Celera Assembler and related tools. We thank Robert Friedman for his leadership of the GBMF-funded project. We acknowledge the J. Craig Venter Institute (JCVI) Joint Technology Center, under the leadership of Yu-Hui Rogers, for producing the genomic libraries and the sequence data.
Published ahead of print on 9 January 2009.