Complete Genome Sequence of Hyperthermophilic Pyrococcus sp. Strain NA2, Isolated from a Deep-Sea Hydrothermal Vent Area

Abstract

Pyrococcus sp. strain NA2, isolated from a deep-sea hydrothermal vent sample, is a novel marine hyperthermophilic archaeon that grows optimally at 93°C. The complete genome sequence of the strain contains all the genes for the tricarboxylic acid cycle except for succinate dehydrogenase/fumarate reductase, but the genome does not encode proteins involved in polysaccharide utilization.

GENOME ANNOUNCEMENT

Pyrococcus sp. strain NA2 was isolated from a sample collected from a hydrothermal vent area in the Papua New Guinea-Australia-Canada-Manus (PACMANUS) field of the western Pacific Ocean at a depth of 1,650 m in 2002. It is noteworthy that this strain had been coisolated with Thermococcus onnurineus NA1, whose complete genome sequence revealed novel CO- and formate-dependent hydrogenogenic metabolisms (1, 5, 7). Pyrococcus sp. NA2 is a sulfur-reducing heterotrophic archaeon belonging to the order Thermococcales (encompassing the genera Thermococcus, Pyrococcus, and Palaeococcus) and is a hyperthermophile growing at temperatures ranging from 70°C to 100°C and optimally at 93°C. To date, the complete genome sequences of three Pyrococcus species, P. horikoshii (4), P. furiosus (9), and P. abyssi (2), have been determined and showed variability of those strains at the level of genomic content and organization (3).

Whole-genome sequencing of Pyrococcus sp. NA2 was performed using GS-FLX Titanium (Roche Diagnostics, Basel, Switzerland) pyrosequencing. The contigs were ordered using Newbler assembler (Roche Diagnostics, Basel, Switzerland), and gap closure was achieved by combinatorial multiplex PCR and sequencing. Protein-coding regions were predicted and annotated using the EMFAS annotation system (Ensoltek, Daejeon, South Korea), and tRNA genes and rRNA genes were predicted using tRNAscan-SE 1.21 and rRNA genes using RNAmmer 1.2 (6, 8).

The Pyrococcus sp. NA2 genome was found to be a single circular 1,861,320-bp chromosome with a G+C content of 42.7%. A total of 1,980 protein-coding sequences, 4 rRNA loci, and 46 tRNA genes were detected. Over 82% of genes were assigned to specific Clusters of Orthologous Groups (COG) Database functional groups, and 45% were assigned an enzyme classification number. Initial analysis of the whole-genome sequence of Pyrococcus sp. NA2 indicated that 80.7%, 80.0%, and 79.7% of the proteins showed similarity (E value lower than 1e−10 with over 80% coverage) to those of P. abyssi, P. horikoshii, and P. furiosus genomes, respectively.

Pathway analysis of the Pyrococcus sp. NA2 genome revealed the presence of all the genes that encode enzymes of the Embden-Meyerhof glycolytic pathway. The strain possesses an incomplete tricarboxylic acid (TCA) cycle in which succinate dehydrogenase/fumarate reductase is missing. Anaerobic operation of an incomplete TCA cycle is proposed to generate extra reducing equivalents. Phosphoenolpyruvate (PEP) carboxylase, PEP carboxykinase, and pyruvate carboxylase carry out anaplerotic reactions that produce oxaloacetate as a TCA cycle intermediate. In comparison with other sequenced Pyrococcus species, the Pyrococcus sp. NA2 genome lacks genes involved in the extracellular degradation of α- or β-glucans, transport of oligosaccharides into the cells through the cytoplasmic membrane, and the subsequent intracellular cleavage of oligosaccharides to monosaccharides, indicating relatively low capacity for polysaccharide degradation. Comparative genomic studies of several Pyrococcus species will provide a foundation for examining the strategies of adaptation to diverse environmental conditions.

Nucleotide sequence accession numbers.

The genome sequence of Pyrococcus sp. NA2 is available in GenBank under accession number CP002670.