Nucleotide sequence of 5S ribosomal RNA of Rhodococcus fascians.
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Biomedical subjects
Publications and source records attributed to C R Woese.
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Through comparative analysis of 16S ribosomal RNA sequences, it can be shown that two seemingly dissimilar types of eubacteria Deinococcus and the ubiquitous hot spring organism Thermus are distantly but specifically related to one another. This confirms an earlier report based upon 16S rRNA oligonucleotide cataloging studies (Hensel et al., 1986). Their two lineages form a distinctive grouping within the eubacteria that deserved the taxonomic status of a phylum. The (partial) sequence of T. aquaticus rRNA appears relatively close to those of other thermophilic eubacteria. e.g. Thermotoga maritima and Thermomicrobium roseum. However, this closeness does not reflect a true evolutionary closeness; rather it is due to a "thermophilic convergence", the result of unusually high G+C composition in the rRNAs of thermophilic bacteria. Unless such compositional biases are taken into account, the branching order and root of phylogenetic trees can be incorrectly inferred.
Comparative analysis of small subunit ribosomal RNA sequences suggests the existence of two new higher order interactions: (i) a double-helical structure involving positions 505-507 and 524-526 (Escherichia coli numbering) and (ii) an interaction between the region of position 130 and the helix located approximately between positions 180 and 195. In the first of these, one of the strands of the helix exists in the bulge loop, and the other strand exists in the terminal loop of a previously recognized compound helix involving positions 500-545. Therefore, the new structure formally represents a pseudoknot. In the second, the insertion/deletion of a nucleotide in the vicinity of position 130 correlates with the length of the helix in the 180-195 region, the latter having a 3-base-pair stalk when the base in question is deleted and a stalk of approximately 10 pairs when it is inserted.
Small subunit rRNA sequences have been determined for representative strains of six species of the family Rickettsiaceae: Rickettsia rickettsii, Rickettsia prowazekii, Rickettsia typhi, Coxiella burnetii, Ehrlichia risticii, and Wolbachia persica. The relationships among these sequences and those of other eubacteria show that all members of the family Rickettsiaceae belong to the so-called purple bacterial phylum. The three representatives of the genus Rickettsia form a tight monophyletic cluster within the alpha subdivision of the purple bacteria. E. risticii also belongs to the alpha subdivision and shows a distant yet specific relationship to the genus Rickettsia. However, the family as a whole is not monophyletic, in that C. burnetii and W. persica are members of the gamma subdivision. The former appears to show a specific, but rather distant, relationship to the genus Legionella.
Critical analysis of the recently proposed alternative to the normal archaebacterial tree, the new eocyte tree, shows that the latter's central topology, in which the eubacteria branch from an entirely different section of the unrooted archaebacterial tree than the eukaryotes, is consistent with an artifact. The effects of the alignment used and the particular composition of the sequence quartets analyzed to infer this tree are discussed in detail.
Sequences for the spacer regions that separate the 16S and 23S ribosomal RNA genes have been determined for four more (strategically placed) archaebacteria. These confirm the general rule that methanogens and extreme halophiles have spacers that contain a single tRNAala gene, while tRNA genes are not found in the spacer region of the true extreme thermophiles. The present study also shows that the spacer regions from the sulfate reducing Archaeglobus and the extreme thermophile Thermococcus (both of which cluster phylogenetically with the methanogens and extreme halophiles) contain each a tRNAala gene. Thus, not only all methanogens and extreme halophiles show this characteristic, but all organisms on the "methanogen branch" of the archaebacterial tree appear to do so. The finding of a tRNA gene in the spacer region of the extreme thermophile Thermococcus celer is the first known phenotypic property that links this organism with its phylogenetic counterparts, the methanogens, rather than with its phenotypic counterparts, the sulfur-dependent extreme thermophiles.
The sequence of the 16S ribosomal RNA gene from the archaebacterium Thermococcus celer shows the organism to be related to the methanogenic archaebacteria rather than to its phenotypic counterparts, the extremely thermophilic archaebacteria. This conclusion turns on the position of the root of the archaebacterial phylogenetic tree, however. The problems encountered in rooting this tree are analyzed in detail. Under conditions that suppress evolutionary noise both the parsimony and evolutionary distance methods yield a root location (using a number of eubacterial or eukaryotic outgroup sequences) that is consistent with that determined by an "internal rooting" method, based upon an (approximate) determination of relative evolutionary rates.
Until recently all archaebacteria isolated conformed to one of three basic phenotypes: they were either methanogens, extreme halophiles, or ('sulphur-dependent') extreme thermophiles. However, a novel phenotype, that fits none of these categories, has recently been described. The organism, strain VC-16 (tentatively called "Archaeoglobus fulgidus") reduces sulphate--the only archaebacterium so far known to do so--and makes very small quantities of methane, although it lacks some of the cofactors normally associated with methanogenesis. These characteristics suggest that strain VC-16 might represent a transition form between an anaerobic thermophilic sulfur-based type of metabolism (which seems to be the ancestral metabolism for archaebacteria and methanogenesis (which somehow then derives from it). We here show that the lineage represented by strain VC-16 arises from the archaebacterial tree precisely where such an interpretation would predict that it would, between the Methanococcus lineage (which is the deepest of the methanogen branchings) and that of Thermococcus (the deepest of all branchings on the methanogen side of the tree).
Thermotoga maritima is one of the more unusual eubacteria: It is highly thermophilic, growing at temperatures higher than any other eubacterium; its cell wall appears to have a unique structure and its lipids a unique composition; and the organism is surrounded by a loose-fitting sheath of unknown function. Its phenotypic uniqueness is matched by its phylogenetic position; Thermotoga maritima represents the deepest known branching in the eubacterial line of descent, as measured by ribosomal RNA sequence comparisons. T. maritima also represents the most slowly evolving of eubacterial lineages. The fact that the two deepest branchings in the eubacterial line of descent (the other, the green non-sulfur bacteria and relatives, i.e. Chloroflexus, Thermomicrobium, etc.) are both basically thermophilic and slowly evolving, strongly suggests that all eubacteria have ultimately arisen from a thermophilic ancestor.
Ribosomal RNA sequence comparisons define a phylogenetic grouping, the green non-sulfur bacteria and relatives (GNS), known to contain the genera Chloroflexus, Herpetosiphon and Thermomicrobium--organisms that have little phenotypic similarity. The unit is phylogenetically deep, but entirely distinct from any other eubacterial division (phylum). It is also relatively ancient--branching from the common eubacterial stem earlier than any other group of eubacteria reported thus far. The group phenotype is predominantly thermophilic, and its thermophilic members, especially Thermomicrobium, are more slowly evolving than Herpetosiphon, a mesophile. The GNS unit appears significantly older than either the green sulfur bacteria or the cyanobacteria--making it likely that organisms such as Chloroflexus, not the cyanobacteria, generated the oldest stromatolites, which formed over three billion years ago.
The phylogenetic incoherency of the genus Bacillus as presently described is demonstrated by analysis of both published and new data from comparative 16S rRNA cataloguing of nine Bacillus species and a number of related non-Bacillus taxa, i.e. Caryophanon latum, Filibacter limicola and Planococcus citreus. While the ellipsoidal-spore-forming bacilli, e.g. B. subtilis and allied species, formed a coherent cluster, the round-spore-forming bacilli showed a higher degree of relationship to the non-spore-forming organisms than these bacilli show among each other. Thus B. sphaericus clustered with C. latum, B. globisporus grouped with F. limicola, B. pasteurii with Sporosarcina ureae, and 'B. aminovorans' with P. citreus, respectively. These organisms formed two related subclusters which, in their phylogenetic depth, are comparable to that of the B. subtilis subline. With the exception of 'B. aminovorans', the 16S rRNA phylogeny was entirely consistent with the distribution of murein types. Even more distantly related to and grouping outside the main Bacillus cluster was B. stearothermophilus, which displayed a moderate relationship to Thermoactinomyces vulgaris. Taxonomic problems arising from the new insights into the intrageneric relationships of Bacillus are discussed.
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The only reliable general method currently available for determining precise higher order structure in the large ribosomal RNAs is comparative sequence analysis. The method is here applied to reveal 'tertiary' structure in the 16S-like rRNAs, i.e. structure more complex than simple double-helical, secondary structure. From a list of computer-generated potential higher order interactions within 16S rRNA one such interaction considered likely was selected for experimental test. The putative interaction involves a Watson-Crick one to one correspondence between positions 570 and 866 in the molecule (E. coli numbering). Using existing oligonucleotide catalog information several organisms were selected whose 16S rRNA sequences might test the proposed co-variation. In all of the (phylogenetically independent) cases selected, full sequence evidence confirms the predicted one to one (Watson-Crick) correspondence. An interaction between positions 570 and 866 is, therefore, considered proven phylogenetically.
Comparisons of complete 16S ribosomal RNA sequences have been used to confirm, refine and extend earlier concepts of archaebacterial phylogeny. The archaebacteria fall naturally into two major branches or divisions, I--the sulfur-dependent thermophilic archaebacteria, and II--the methanogenic archaebacteria and their relatives. Division I comprises a relatively closely related and phenotypically homogeneous collection of thermophilic sulfur-dependent species--encompassing the genera Sulfolobus, Thermoproteus, Pyrodictium and Desulfurococcus. The organisms of Division II, however, form a less compact grouping phylogenetically, and are also more diverse in phenotype. All three of the (major) methanogen groups are found in Division II, as are the extreme halophiles and two types of thermoacidophiles, Thermoplasma acidophilum and Thermococcus celer. This last species branches sufficiently deeply in the Division II line that it might be considered to represent a separate, third Division. However, both the extreme halophiles and Tp. acidophilum branch within the cluster of methanogens. The extreme halophiles are specifically related to the Methanomicrobiales, to the exclusion of both the Methanococcales and the Methanobacteriales. Tp. acidophilum is peripherally related to the halophile-Methanomicrobiales group. By 16S rRNA sequence measure the archaebacteria constitute a phylogenetically coherent grouping (clade), which excludes both the eubacteria and the eukaryotes--a conclusion that is supported by other sequence evidence as well. Alternative proposals for archaebacterial phylogeny, not based upon sequence evidence, are discussed and evaluated. In particular, proposals to rename (reclassify) various subgroups of the archaebacteria as new kingdoms are found wanting, for both their lack of proper experimental support and the taxonomic confusion they introduce.
A method of analyzing 16S rRNA catalog data has been developed in which groupings at various taxonomic levels can be characterized in terms of specific "signature" oligonucleotides. This approach provides an alternative means for evaluating higher order branching possibilities and can be used to assess the phylogenetic position of isolates that are poorly placed by the usual clustering procedures. This signature approach has been applied to forty archaebacterial catalogs and every oligonucleotide with significant signature value has been identified. Sets of specific oligonucleotides were identified for every major group on a dendrogram produced by cluster analysis procedures. Signatures that would establish between group relationships were also sought and found. In the case of the Methanobacteriaceae the clustering methods suggest a specific relationship to the Methanococcaceae. This inclusion is in fact supported by six strong signature oligonucleotides. However there are also significant numbers of signature oligonucleotides supporting a specific relationship of the Methanobacteriaceae to either the Halobacteriaceae or the Methanomicrobiaceae. Thus the placement of the Methanobacteriaceae is less certain than the usual dendrograms imply. The signature approach also was used to assess the phylogenetic position of Thermoplasma acidophilum which is found to be more closely related to the methanogen/halophile Division than to the sulfur dependent Division of the archaebacteria. This does not imply however that Thermoplasma acidophilum is properly regarded as being in the methanogen/halophile Division.
The sequence of the 16S rRNA gene from Chlamydia psittaci was determined. Comparison of this sequence with other 16S rRNA sequences showed the organism to be eubacterial. The organism represents a hitherto unrecognized major eubacterial group. However, this group may be peripherally related to the planctomyces and relatives. Although these two groups seem to have very little in common phenotypically (they have been studied in very different ways), cell walls in both cases contain no peptidoglycan.
On the basis of ribosomal RNA sequence comparisons, the rickettsia Rochalimaea quintana has been found to be a member of subgroup 2 of the alpha subdivision of the so-called purple bacteria, which is one of about ten major eubacterial divisions. Within subgroup alpha-2, R. quintana is specifically related to the agrobacteria and rhizobacteria, organisms that also have close associations with eukaryotic cells. This genealogical grouping of the rickettsiae with certain plant pathogens and intracellular symbionts suggests a possible evolution of the rickettsiae from plant-associated bacteria.