Green sulfur bacteria from hypersaline Chiprana Lake (Monegros, Spain): habitat description and phylogenetic relationship of isolated strains.

The 'Salada de Chiprana' (Chiprana Lake) is a hypersaline (30-73 per thousand), permanent and shallow lake of endorheic origin in a semi-arid region of the Ebro depression (Aragon, Spain). Magnesium sulfate and sodium chloride represent the main salts of this athalassohaline environment. Anoxic conditions occurred periodically in the bottom layers of the lake during the study period. When stratified, high sulfide concentrations (up to 7 mM) were measured in the hypolimnion. Physical and chemical conditions gave rise to the development of very dense green sulfur bacteria blooms (10.7 mg l(-1) of BChl c and 16.7 mg l(-1) of BChl d) at 0.5-1 m from the bottom. Microscopic observations revealed that cells morphologically similar to Chlorobium vibrioforme were dominant in the phototrophic bacterial community, but Prosthecochloris aestuarii was also found sometimes at lower concentrations, as revealed by both microscopic observation and flow cytometric analyses. Deep agar dilution series allowed to obtain several axenic cultures of phototrophic bacteria. They were identified according to their morphology, pigment composition and phylogenetic relationships (16S rDNA sequence analysis). Two of the sequenced strains (CHP3401 and CHP3402) belonged to the green sulfur bacteria and were related to Prosthecochloris aestuarii SK413(T) and Chlorobium vibrioforme DSM260(T), respectively. HPLC analyses of both natural samples and Chlorobium vibrioforme isolates indicated that these strains contained both BChl c and BChl d. Phylogenetic results suggested that Chlorobium vibrioforme strains DSM260(T) and CHP3402, all sequenced strains of Prosthecochloris aestuarii and strain CIB2401 constitute a separate cluster of green sulfur bacteria, all of them isolated from marine to hypersaline habitats.

Phylogeny of the PscB reaction center protein from green sulfur bacteria.

The reaction center (RC) of green sulfur bacteria has iron-sulfur clusters as terminal acceptors and is related to the Type I RC found in Heliobacter sp. and in Photosystem I (PS I) of green plants and cyanobacteria. Degenerate primers were used to retrieve the genes coding for one of the RC proteins, PscB, from 11 strains of green sulfur bacteria. PCR using the same primers gave no product with a second group of strains and the protein from these strains did not crossreact with antibodies raised against purified PscB from the first group, suggesting the presence of a high degree of variability. The sequences shared a high degree of similarity in the region coding for the binding motif for the 4Fe-4S centers. However, the N-terminal portion of the deduced protein sequences was highly variable and contained a highly positively charged, low-complexity region with repeated tetrapeptides with two alanines flanked by proline or lysine. The PscB sequences obtained fell into two major groups, and the results suggested a lack of correlation between the pigmentation of the chlorosome antenna system and the reaction center protein. There is also a lack of correlation between the grouping of the pscB sequences and the phylogeny deduced from 16S rRNA.

Phylogenetic diversity of a bacterial community determined from Siberian tundra soil DNA.

Genomic DNA was isolated from the active layer of tundra soil collected from the Kolyma lowland, Northeast Eurasia, near the Arctic Ocean coast. The SSU (small subunit) rRNA genes were amplified with eubacterial primers from the bulk genomic community DNA and cloned into plasmid vectors. Forty-three SSU rDNA clones were obtained, and all of them had different RFLP patterns. Phylogenetic analysis based on partial sequences (about 300 bp) established with the maximum likelihood method revealed the presence of three major and several minor groups that fell into 11 of the established lines of bacteria, and one sequence that could not be assigned to any of the described groups. Most of the clones belonged to the alpha (20.9%) and delta (25.6%) subdivisions of the Proteobacteria, with lesser proportions in the beta (9.3%) and gamma (4.7%) subdivisions, groups typically isolated from soil by culture methods. Fewer than 12% of the clones belonged to Gram-positive bacteria, and 16% of the clones were related to Fibrobacter. The majority of the clones (70%) had sequences that were 5-15% different from those in the current databases, and 7% of the clones had sequences that differed by more than 20% from those in the database. The results suggest that these tundra-derived clones are very diverse in phylogeny, and that many probably reflect new genera or families. Hence, most of the tundra soil bacterial community has never been isolated and thus the physiology and function of its dominant members appears to be unknown.