Complete genome sequence of Staphylothermus marinus Stetter and Fiala 1986 type strain F1

Staphylothermus marinus Fiala and Stetter 1986 belongs to the order Desulfurococcales within the archaeal phylum Crenarchaeota. S. marinus is a hyperthermophilic, sulfur-dependent, anaerobic heterotroph. Strain F1 was isolated from geothermally heated sediments at Vulcano, Italy, but S. marinus has also been isolated from a hydrothermal vent on the East Pacific Rise. We report the complete genome of S. marinus strain F1, the type strain of the species. This is the fifth reported complete genome sequence from the order Desulfurococcales.


Introduction
Strain F1 is the type strain of the species Staphylothermus marinus. It was isolated from geothermally heated sediments at Vulcano, Italy [1], and was the strain sequenced. S. marinus was also isolated from a hydrothermal vent on the East Pacific Rise. There is one other species within the genus, Staphylothermus hellenicus, which was isolated from a hydrothermal vent at Milos, Greece [2]. Four other complete genomes from the order Desulfurococcales have been published, but S. marinus is not closely related to any of these organisms. (Figure 1) We describe here the properties of the complete genome sequence of S. marinus strain F1 (DSM 3639, ATCC 43588).

Classification and features
S. marinus is a nonmotile coccus with a diameter of 0.5-1.0 μm. At low nutrient concentrations it forms clumps of up to 100 cells, while at higher nutrient concentrations single cells or pairs of cells are observed. At high concentrations of yeast extract, giant cells with a diameter of up to 15μm are formed [1]. The optimum and maximum growth temperatures also depend on the nutrient concentration. At low nutrient concentration the optimum growth temperature is 85°C and the maximum is 92°C, while at higher nutrient concentration the optimum growth temperature is 92°C and the maximum is 98°C [1]. The optimum pH for growth is 6.5, but growth is observed within a range of 4.5 to 8.5. S. marinus is a heterotroph, growing on complex media but not on simple carbohydrates or amino acids. Elemental sulfur is required for growth, and it can not be substituted by other sulfur compounds [1]. In the absence of sulfur, cells can survive while producing hydrogen [5]. Metabolic products are CO2, H2S, acetate, and isovalerate, suggesting a metabolism similar to that of Pyrococcus species [1]. Several features suggest that S. marinus is a typical member of the Archaea. Its growth was not inhibited by vancomycin, kanamycin, streptomycin, or chloramphenicol, but it is sensitive to diphtheria toxin [1]. Its cell wall lacks murein, and it contains typical archaeal membrane lipids [1]. Other features of the organism are presented in Table 1.

Genome sequencing and annotation
Genome project history S. marinus was selected for sequencing based upon its phylogenetic position relative to other sequenced archaeal genomes. It is part of a 2006 Joint Genome Institute Community Sequencing Program (CSP) project that included six diverse archaeal genomes. The complete genome sequence was finished in February, 2007. The Gen-Bank accession number for the chromosome is CP000575. The genome project is listed in the Genomes OnLine Database (GOLD) [17] as project Gc00511. Sequencing, finishing and annotation were performed by the DOE Joint Genome Institute (JGI). A summary of the project information is shown in Table 2.  Altitude not applicable Evidence codes -IDA: Inferred from Direct Assay (first time in publication); TAS: Traceable Author Statement (i.e., a direct report exists in the literature); NAS: Non-traceable Author Statement (i.e., not directly observed for the living, isolated sample, but based on a generally accepted property for the species, or anecdotal evidence). These evidence codes are from the Gene Ontology project [16]. If the evidence code is IDA, then the property was observed for a living isolate by one of the authors or an expert mentioned in the acknowledgements.

Growth conditions and DNA isolation
The methods for DNA isolation, genome sequencing and assembly for this genome have previously been published [18].

Genome annotation
Protein-coding genes were identified using a combination of Critica [19] and Glimmer [20] followed by a round of manual curation using the JGI Gene-PRIMP pipeline [21]. The predicted CDSs were translated and used to search the National Center for Biotechnology Information (NCBI) nonredundant database, UniProt, TIGRFam, Pfam, PRIAM, KEGG, COG, and InterPro databases. The tRNAScan-SE tool [22] was used to find tRNA genes. Additional gene prediction analysis and manual functional annotation was performed within the Integrated Microbial Genomes Expert Review (IMG-ER) platform [23].

Genome properties
The genome of S. marinus F1 consists of a single circular chromosome ( Table 3 and Figure 2 About 59% of predicted genes begin with an AUG codon, 33% begin with UUG, and only 8% begin with GUG. There is one copy of each ribosomal RNA. The properties and statistics of the genome are shown in Table 3, and the distribution of proteins in COG categories is shown in Table 4.

Insights from genome sequence
The genome of S. marinus has several novel features compared to other Crenarchaeota. It is the first crenarchaeote found to have a sodium iontranslocating decarboxylase, which is probably involved in energy generation from amino acid degradation [18]. In addition it is the first crenarchaeote found to have proteins related to multisubunit cation/proton antiporters, although the S. marinus proteins probably do not function as antiporters. These antiporter-related proteins belong to larger operons similar to the mbh and mbx operons of Pyrococcus furiosus [24,25], therefore, they may play a role in sulfur reduction or hydrogen production. S. marinus appears to use different proteins for sulfur reduction than the other anaerobic, sulfur-reducing Crenarchaeota. Both Thermofilum pendens and Hyperthermus butylicus appear to have molybdenum-containing sulfur/polysulfide reductases and NADPH:sulfur oxidoreductases, but these are not present in S. marinus [18]