Isolation and characterization of oligotrophic acido-tolerant methanogenic consortia from a Sphagnum peat bog.

Two dense and highly enriched (up to 10(9) cells ml(-1), <10% of bacterial satellites) acido-tolerant (pH 4.0-6.5) methanogenic consortia, '26' and 'K', were isolated from the peat beneath a Sphagnum-Eriophorum-Carex community in West Siberia. Both consortia produced methane from CO2:H2 on chemically defined, diluted N-free media containing Ti(III)citrate as reducing agent. The phylogenetic analysis of 16S ribosomal DNA revealed three archaeal and nine bacterial sequence types. Consortium '26' contained single archaea Methanobacterium sp., represented by rods of 1.5-10x0.5-1.0 microm. In consortium 'K', there were two archaeal phylotypes, the respective methanogens were further differentiated morphologically with the fluorescence in situ hybridization technique: one less abundant (<2%) population of the long-curved rods (50-100x0.3-0.4 microm) fell into the order of Methanomicrobiales, while the dominant organism ( approximately 98%), represented by straight rods with abrupt rectangular ends (3-9x0.5 microm), was affiliated with earlier uncultured 'Rice cluster I'. The main bacterial satellite used citrate as a single carbon and energy source; it was similar in both consortia, and after isolation in pure culture, it was identified as a new member of the alpha-subclass of Proteobacteria. The other bacterial satellites were distributed among four taxonomic groups: the delta-subclass of Proteobacteria, the Flavobacterium-Bacteroides-Cytophaga line of descent, the Acidobacterium-Fibrobacter line of descent and the Green non-sulfur bacteria line of descent. At least 11 out of 12 components of acido-tolerant consortia are new to science at the species, genus and order levels; their existence until now was evident only from environmental gene retrievals. The Sphagnum wetlands, attracting attention only recently because of their global environmental role, are shown to be an especially valuable source of novel prokaryotic organisms.