Detection of Hyperthermophilic Archaea of the Genus Desulfurococcus by Hybridization with Oligonucleotide Probes

Based on the analysis of nucleotide sequences of 16S rRNA, oligonucleotide probes were designed for the detection and identification of representatives of the genus Desulfurococcus (kingdom Crenarchaeota of the domain Archaea). The detection procedure included obtaining PCR products on DNA isolated from pure cultures, enrichments, or natural samples with a designed Crenarchaeota-specific primer pair: Cren 7F (5"-TTCCGGTTGATCCYGCCGGACC-3") and Cren 518R (5"-GCTGGTWTTACCGCGGCGGCTGA-3"). The PCR products were hybridized with Dig-11-dUTP–labeled oligonucleotide probes targeting the genus Desulfurococcus (Dco 198, 5"-CGTTAACYCCYGCCACACC-3") and its species D. mobilis (Dco_mob 198, 5"-CGTTAACCCCTGCCACACC-3") and D. amylolyticus (Dco_amy 198, 5"-CGTTAACCCCCGCCACACC-3"). With the use of these primers and probes, four new strains isolated from hydrotherms of Kamchatka and Kunashir Island were identified as members of the speciesDesulfurococcus amylolyticus. Desulfurococcus representatives were detected in several natural samples, including a sample taken from a marine hydrotherm at Kunashir Island; this demonstrates that representatives of this genus occur not only in terrestrial but also in marine environments.