PHYLOGENETIC-RELATIONSHIPS OF THIOMICROSPIRA SPECIES AND THEIR IDENTIFICATION IN DEEP-SEA HYDROTHERMAL VENT SAMPLES BY DENATURING GRADIENT GEL-ELECTROPHORESIS OF 16S RDNA FRAGMENTS

Denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S rD NA fragments was used to explore the genetic diversity of hydrothermal vent microbial communities, specifically to determine the importance of sulfur-oxidizing bacteria therein. DGGE analysis of two different h ydrothermal vent samples revealed one PCR band for one sample and thre e PCR bands for the other sample, which probably correspond to the dom inant bacterial populations in these communities. Three of the four 16 S rDNA fragments were sequenced. By comparison with 16S rRNA sequences of the Ribosomal Database Project, two of the DGGE-separated fragment s were assigned to the genus Thiomicrospira. To identify these 'phylot ypes' in more detail, a phylogenetic framework was created by determin ing the nearly complete 16S rRNA gene sequence (approx. 1500 nucleotid es) from three described Thiomicrospira species, viz., Tms, crunogena, Tms. pelophila, Tms. denitrificans, and from a new isolate, Thiomicro spira sp. strain MA2-6. All Thiomicrospira species except Tms. denitri ficans formed a monophyletic group within the gamma subdivision of the Proteobacteria. Tms. denitrificans was assigned as a member of the ep silon subdivision and was distantly affiliated with Thiovulum, another sulfur-oxidizing bacterium. Sequences of two dominant 16S rDNA fragme nts obtained by DGGE analysis fell into the gamma subdivision Thiomicr ospira. The sequence of one fragment was in all comparable positions i dentical to the 16S rRNA sequence of Tms. crunogena. Identifying a dom inant molecular isolate as Tms. crunogena indicates that this species is a dominant community member of hydrothermal vent sites. Another 'ph ylotype' represented a new Thiomicrospira species, phylogenetically in an intermediate position between Tms. crunogena and Tms. pelophila. T he third 'phylotype' was identified as a Desulfovibrio, indicating tha t sulfate-reducing bacteria, as sources of sulfide, may complement sul fur- and sulfide-oxidizing bacteria ecologically in these sulfide-prod ucing hydrothermal vents.