Pyrofollis japonicus gen. nov. sp. nov., a novel member of the family Pyrodictiaceae isolated from the Iheya North hydrothermal field.

A novel hyperthermophilic, heterotrophic archaeon, strain YC29T, was isolated from a deep-sea hydrothermal vent in the Mid-Okinawa Trough, Japan. Cells of strain YC29T were non-motile, irregular cocci with diameters of 1.2-3.0 µm. The strain was an obligatory fermentative anaerobe capable of growth on complex proteinaceous substrates. Growth was observed between 85 and 100 °C (optimum 90-95 °C), pH 4.9-6.4 (optimum 5.1), and in the presence of 1.4-4.0% (w/v) NaCl (optimum 3.0%). Inorganic carbon was required as a carbon source. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that the isolate was a member of the family Pyrodictiaceae. The genome size was 2.02 Mbp with a G+C content of 49.4%. The maximum values for average nucleotide identity (ANI), average amino acid identity (AAI), and in silico DNA-DNA hybridization (dDDH) value of strain YC29T with relatives were 67.9% (with Pyrodictium abyssi strain AV2T), 61.1% (with Pyrodictium occultum strain PL-19T), and 33.8% (with Pyrolobus fumarii strain 1AT), respectively. Based on the phylogenetic, genomic, and phenotypic characteristics, we propose that strain YC29T represents a novel genus and species, Pyrofollis japonicus gen. nov., sp. (type strain YC29T = DSM 113394T = JCM 39171T).

Pyrodictium delaneyi sp. nov., a hyperthermophilic autotrophic archaeon that reduces Fe(III) oxide and nitrate.

A hyperthermophilic, autotrophic iron and nitrate reducer, strain Su06T, was isolated from an active deep-sea hydrothermal vent chimney on the Endeavour Segment in the north-eastern Pacific Ocean. It was obligately anaerobic, hydrogenotrophic and reduced Fe(III) oxide to magnetite and NO3- to N2. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the strain was more than 97 % similar to other species of the genera Pyrodictium and Hyperthermus. Therefore, overall genome relatedness index analyses were performed to establish whether strain Su06T represents a novel species. For each analysis, strain Su06T was most similar to Pyrodictium occultum PL-19T. Relative to this strain, the average nucleotide identity score for strain Su06T was 72 %, the genome-to-genome direct comparison score was 13-19 % and the species identification score at the protein level was 89 %. For each analysis, strain Su06T was below the species delineation cutoff. Based on its whole genome sequence and its unique phenotypic characteristics, strain Su06T is suggested to represent a novel species of the genus Pyrodictium, for which the name Pyrodictium delaneyi is proposed. The type strain is Su06T (=DSM 28599T=ATCC BAA-2559T).

Study of the distribution of autotrophic CO2 fixation cycles in Crenarchaeota.

Two new autotrophic carbon fixation cycles have been recently described in Crenarchaeota. The 3-hydroxypropionate/4-hydroxybutyrate cycle using acetyl-coenzyme A (CoA)/propionyl-CoA carboxylase as the carboxylating enzyme has been identified for (micro)aerobic members of the Sulfolobales. The dicarboxylate/4-hydroxybutyrate cycle using oxygen-sensitive pyruvate synthase and phosphoenolpyruvate carboxylase as carboxylating enzymes has been found in members of the anaerobic Desulfurococcales and Thermoproteales. However, Sulfolobales include anaerobic and Desulfurococcales aerobic autotrophic representatives, raising the question of which of the two cycles they use. We studied the mechanisms of autotrophic CO(2) fixation in the strictly anaerobic Stygiolobus azoricus (Sulfolobales) and in the facultatively aerobic Pyrolobus fumarii (Desulfurococcales). The activities of all enzymes of the 3-hydroxypropionate/4-hydroxybutyrate cycle were found in the anaerobic S. azoricus. In contrast, the aerobic or denitrifying P. fumarii possesses all enzyme activities of the dicarboxylate/4-hydroxybutyrate cycle. We conclude that autotrophic Crenarchaeota use one of the two cycles, and that their distribution correlates with the 16S rRNA-based phylogeny of this group, rather than with the aerobic or anaerobic lifestyle.

The complete genome sequence of Staphylothermus marinus reveals differences in sulfur metabolism among heterotrophic Crenarchaeota.

BACKGROUND: Staphylothermus marinus is an anaerobic, sulfur-reducing peptide fermenter of the archaeal phylum Crenarchaeota. It is the third heterotrophic, obligate sulfur reducing crenarchaeote to be sequenced and provides an opportunity for comparative analysis of the three genomes. RESULTS: The 1.57 Mbp genome of the hyperthermophilic crenarchaeote Staphylothermus marinus has been completely sequenced. The main energy generating pathways likely involve 2-oxoacid:ferredoxin oxidoreductases and ADP-forming acetyl-CoA synthases. S. marinus possesses several enzymes not present in other crenarchaeotes including a sodium ion-translocating decarboxylase likely to be involved in amino acid degradation. S. marinus lacks sulfur-reducing enzymes present in the other two sulfur-reducing crenarchaeotes that have been sequenced -- Thermofilum pendens and Hyperthermus butylicus. Instead it has three operons similar to the mbh and mbx operons of Pyrococcus furiosus, which may play a role in sulfur reduction and/or hydrogen production. The two marine organisms, S. marinus and H. butylicus, possess more sodium-dependent transporters than T. pendens and use symporters for potassium uptake while T. pendens uses an ATP-dependent potassium transporter. T. pendens has adapted to a nutrient-rich environment while H. butylicus is adapted to a nutrient-poor environment, and S. marinus lies between these two extremes. CONCLUSION: The three heterotrophic sulfur-reducing crenarchaeotes have adapted to their habitats, terrestrial vs. marine, via their transporter content, and they have also adapted to environments with differing levels of nutrients. Despite the fact that they all use sulfur as an electron acceptor, they are likely to have different pathways for sulfur reduction.

The genome of Hyperthermus butylicus: a sulfur-reducing, peptide fermenting, neutrophilic Crenarchaeote growing up to 108 degrees C.

Hyperthermus butylicus, a hyperthermophilic neutrophile and anaerobe, is a member of the archaeal kingdom Crenarchaeota. Its genome consists of a single circular chromosome of 1,667,163 bp with a 53.7% G+C content. A total of 1672 genes were annotated, of which 1602 are protein-coding, and up to a third are specific to H. butylicus. In contrast to some other crenarchaeal genomes, a high level of GUG and UUG start codons are predicted. Two cdc6 genes are present, but neither could be linked unambiguously to an origin of replication. Many of the predicted metabolic gene products are associated with the fermentation of peptide mixtures including several peptidases with diverse specificities, and there are many encoded transporters. Most of the sulfur-reducing enzymes, hydrogenases and electron-transfer proteins were identified which are associated with energy production by reducing sulfur to H(2)S. Two large clusters of regularly interspaced repeats (CRISPRs) are present, one of which is associated with a crenarchaeal-type cas gene superoperon; none of the spacer sequences yielded good sequence matches with known archaeal chromosomal elements. The genome carries no detectable transposable or integrated elements, no inteins, and introns are exclusive to tRNA genes. This suggests that the genome structure is quite stable, possibly reflecting a constant, and relatively uncompetitive, natural environment.