16S rRNA analysis and the phylogenetic position of Wolinella succinogenes

The 16S ribosomal RNA of Wolinella succinogenes ATCC29543 was analyzed by the RNase T1 oligonucleotide cataloguing approach. In contrast to its present classification, W. succinogenes is related neither to members of the genus Bacteroides nor to any other genus of the family Bacteroidaceae. As derived from the similarity coefficients (S_AB values) calculated on the basis of more than 350 eubacterial species, W. succinogenes appears to be a distantly related member of the division of purple photosynthetic bacteria and their relatives; however, S_AB values do not indicate that this species is preferentially related to any representative of the 4 subdivisions.     

A molecular approach to search for diversity among bacteria in the environment

Molecular 16S rDNA-based techniques were applied to a peat sample from northern Germany in order to investigate the bacterial diversity present and compare the clone sequences with those obtained from similar studies on other terrestrial samples. Genomic DNA was extracted from the peat matrix by a direct lysis procedure. 16S rRNA genes were amplified using PCR primers targeting conserved regions of bacterial 16S rDNA. 16S rDNA fragments were blunt end cloned into a plasmid vector and the resulting clone library of 262 sequences was screened by hybridization with different oligonucleotide probes and sequence analysis of randomly selected clones. The 16S rDNA insert of 76 clones was partially sequenced. Clones identified either by hybridization or by sequence analysis fell into three phyla. As judged by hybridization with a specific oligonucleotide probe, 42% of the clones represented members of the alpha subclass of Proteobacteria. Twenty-five of these clones were selected randomly for sequence analysis; none could be assigned to any of the known genera of this subclass. The second largest clone group comprises 15% of the clones and clusters aroundAcidimicrobium ferrooxidans andRubrobacter radiotolerans, both of which are remotely related to members of the order Actinomycetales. The third major clone cluster (10%) was moderately to remotely related to theAcidobacterium capsulatum phylum. Of the additional clones sequenced, a few could be assigned to other subclasses ofProteobacteria, theVerrucomicrobium phylum and the phylum of spirochetes. Comparison of the results presented here with those from other environments reveals a significant number of common clone clusters. As the vast majority of sequences retrieved from any of the marine and terrestrial samples investigated so far by molecular methods indicate the presence of novel bacterial species it can be assumed that a huge, as yet untapped biotechnological potential is present in the environment.     

Cryptoendolithic actinomycetes from antarctic sandstone rock samples: Micromonospora endolithica sp. nov. and two isolates related to Micromonospora coerulea Jensen 1932

Three cryptoendolithic, aerobic actinomycetes (AA-459T, AA-319 and AA-321) from antarctic sandstone were characterised phenotypically and by molecular taxonomic methods. The isolates had single spores on substrate mycelium, meso-diaminopimelic acid (m-DAP) and glycine (cell wall type II), a whole cell sugar pattern D (galactose, xylose, arabinose, glucose or rhamnose) and phospholipids of type PII (diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol). Their predominant fatty acids were iso-16:0 and iso-15:0 or 17:1omega8c, the menaquinone profile was complex with mainly MK10 (H4) and MK10 (H6). A wide variety of sugars and several acids were utilised for growth. The isolates were sensitive to a few antibiotics, but formation and excretion of antibiotics was not observed. Phenotypically, isolates AA-319 and AA-321 were similar. Phylogenetic analysis of 16S rRNA gene sequences revealed close relationship of strains AA-319 and AA-321 with each other (99.5%) and clustering (98.5%) with Micromonospora coerulea DSM 43143T. DNA-DNA hybridisation showed both strains to be genomically highly similar to strain DSM 43143T. Phenotypically they could be viewed as separate taxa, but presently they will be considered as strains of Micromonospora coerulea. Strain AA-459T was phylogenetically close to Micromonospora chersina DSM 44151T (99.1%) and to Micromonospora rosaria DSM 803T, but DNA-DNA similarity with M. chersina DSM 44151T was low with 28.9/33.5 %, indicating the presence of a different and new species. Consequently, isolate AA-459T (DSM 44398T NRRL B-24248T) is described as the type strain of Micromonospora endolithica sp. nov.     

Designation of Streptomycete 16S and 23S rRNA-based target regions for oligonucleotide probes

The 16S and 23S rRNA of various Streptomyces species were partially sequenced and screened for the presence of stretches that could define all members of the genus, groups of species, or individual species. Nucleotide 929 (Streptomyces ambofaciens nomenclature [J.L. Pernodet, M.T. Alegre, F. Boccard, and M. Guerineau, Gene 79:33-46, 1989]) is a nucleotide highly unique to Streptomyces species which, in combination with flanking regions, allowed the designation of a genus-specific probe. Regions 158 through 203 of the 16S rRNA and 1518 through 1645 of the 23S rRNA (helix 54 [Pernodet et al., Gene 79:33-46, 1989]) have a high potential to define species, whereas the degree of variation in regions 982 through 998 and 1102 through 1122 of the 16S rRNA is less pronounced but characteristic for at least certain species. Alone or in combination with each other, these regions may serve as target sites for synthetic oligonucleotide probes and primers to be used in the determination of pure cultures and in the characterization of community structures. The specificity of several probes is demonstrated by dot blot hybridization.     

Anoxygenic phototrophy across the phylogenetic spectrum: current understanding and future perspectives

The phylogenetic heterogeneity of anoxygenic phototrophic bacteria has been revealed by 16S rRNA sequence analysis, the results of which have led to extensive taxonomic rearrangements within previously defined taxa of phototrophs and stimulated interest in this group of organisms. Anoxygenic photosynthetic bacteria can be found within 4 of the 12 phylogenetic lineages, and in some cases are highly related to non-photosynthetic members of these groups. The largest number of phototrophs are found in the class Proteobacteria. Comparative phylogenetic analysis using 23S rDNA sequences generally supports the topology obtained from 16S rDNA sequences. The photosynthetic reaction centers are conserved in all photosynthetic bacteria, and are of two types. One is shared by the Proteobacteria and Chloroflexus aurantiacus and is similar to Photosystem II of cyanobacteria, while heliobacteria and Chlorobium and relatives possess a reaction center similar to the cyanobacterial Photosystem I. These similarities are supported by sequence analysis of core reaction center peptides, but contradict phylogenies reconstructed from rRNA sequence analysis. Genome analysis by means of physical mapping has been performed for only three species of anoxygenic phototrophs. Some conservation of operon structure and gene sequence has been found within the Proteobacteria, but does not extend to other phototrophs. Received: 29 December 1995 / Accepted: 19 July 1996     

Bacterial Diversity in the Haloalkaline Lake Elmenteita, Kenya

Lake Elmenteita is one of the alkaline saline lakes within the Kenyan Rift valley. The lake is situated on the floor of the Kenyan Rift Valley at 1,776?m above sea level and has no direct outlet. The microbial diversity of the lake was investigated using a culture-independent approach. Five different sampling points were selected randomly within the lake. Wet sediments and water samples were collected from each sampling point. In addition, dry mud cake was collected from three points where the lake had dried. DNA was extracted from the samples and the 16S rRNA genes amplified using universal primers for Bacteria. Thirteen clone libraries were constructed using the PCR amplified 16S rRNA genes. A total of 1,663 clones were picked. Representative clones were selected using ARDRA technique for sequencing. 655 partial and non-chimeric clone sequences indicated the presence of 37 orders in the Domain Bacteria. Cyanobacteria were the most abundant clones in terms of numbers whereas members of the phylum Firmicutes group were the second in terms of numbers but the most diverse in terms of genera represented. All clones affiliated to the class Betaproteobacteria originated from DNA obtained from the water samples. Analysis using BLAST showed that 93.1% of the sequenced clones had similarity values below 98% to both cultured and as yet uncultured bacteria, resulting in 596 phylotypes. Therefore, it can be concluded that Lake Elmenteita harbours phylogenetically diverse groups of bacteria involved in complex metabolic interactions within the Lake??s ecosystem.     

Phylogenetic and chemotaxonomic characterization of Acidaminococcus fermentans

The phylogenetic position of Acidaminococcus fermentans was determined by comparative sequence analysis of the 16S rRNA. This Gram-negative bacterium is a member of the Sporomusa cluster that is defined by other Gram-negative bacteria, i.e. Sporomusa, Megasphaera, Selenomonas, Butyrivibrio, Pectinatus, and Zymophilus. The branching point of this group within the radiation of Gram-positive bacteria of the Clostridium/Bacillus subphylum and adjacent to Peptococcus niger could be confirmed. Chemotaxonomic data were provided for a more detailed characterization of A. fermentans.     

Disproving the hypothesis of a common ancestry for the Ochromonas danica chrysoplast and Heliobacterium chlorum

The phylogenetic position of the golden-yellow alga Ochromonas danica chrysoplast was investigated by comparison of the 16S rRNA catalogue and two long 16S rRNA stretches (804 and 454 bases) with catalogues from eubacteria and chloroplasts and with homologoes 16S rRNA regions from Escherichia coli, Bacillus subtilis, Heliobacterium chlorum, Anacystis nidulans and chloroplasts from Zea mays, Nicotiana tabacum, Euglena gracilis and Chlamydomonas reinhardii, respectively. Both approaches indicate a closer relationship of the chrysoplast to chloroplasts and cyanobacteria than to the brownish photoheterotrophic Heliobacterium chlorum for which a common ancestry has recently been hypothesized.     

Towards a phylogeny of the actinomycetes and related organisms

The method of 16S ribosomal RNA oligonucleotide cataloging has been used to establish phylogenetic relationships among a number of genera of actinomycetes and related organisms. The findings serve as a starting point for a restructuring of actinomycete taxonomy. Several genera, such asThermoactinomyces andEubacterium, do not belong with the group, and the species of at least one atypical genus,Micrococcus, must be included therein. The general tendency to classify within the group according to morphological complexity was shown to be phylogenetically invalid.