© 2002 Society for Molecular Biology and Evolution
The Paradox of the "Ancient" Bacterium Which Contains "Modern" Protein-Coding Genes
Department of Ecology and Evolutionary Biology,Department of Veterinary Science and Microbiology, University of Arizona; andDepartment of Biology, West Chester UniversityThe isolation of microorganisms from ancient materials and the verification that they are as old as the materials from which they were isolated continue to be areas of controversy. Almost without exception, bacteria isolated from ancient material have proven to closely resemble modern bacteria at both morphologicaland molecular levels. This fact has historically been used by critics to argue that these isolates are not ancient but are modern contaminants introduced either naturally after formation of the surrounding material (for further details, see Hazenand Roeder 2001
and the reply by Powers, Vreeland, and Rosenzweig 2001) or because of flaws in the methodology of sample isolation (reviewed recently in Vreeland and Rosenzweig 2002 ). Such criticism has been addressed experimentally by the development of highly rigorous protocols for sample selection, surface sterilization, and contamination detection and control procedures. Using the most scrupulous and well-documented sampling procedures and contamination-protection techniques reported to date, Vreeland,Rosenzweig, and Powers (2000) reported the isolation of a sporeformingbacterium, Bacillus strain 2-9-3, from a brine inclusion within a halite crystal recovered from the 250-Myr-old Permian Salado Formation in Carlsbad, NM.As had been noted in earlier studies, a striking observation by Vreeland, Rosenzweig, and Powers (2000)
was that the 16S rDNA of isolate 2-9-3 is 99% identical to that of Salibacillus marismortui, a bacterium isolated from the Dead Sea in 1936 (Arahal et al. 1999 ). In fact, Arahal et al. (1999) identified as S. marismortui three strains with 16S rDNA sequences differing by 0.01%, suggesting that isolate 2-9-3 might also be classified as S. marismortui.Two groups have since used phylogenetic analyses of 16S rDNA sequences to argue that isolate 2-9-3 is unlikely to be 250 Myr old. Graur and Pupko (2001)
used a relative rate test to compare evolutionary rates of 16S rDNA on the branches leading to isolate 2-9-3 and S. marismortui and found no differences between the evolutionary rates. Considering the possibility that S. marismortui may also be ancient (Arahal et al. 1999 ; Vreeland, Rosenzweig, and Powers 2000 ), they also compared the evolutionary rates of isolate 2-9-3, S. marismortui and Virgibacillus proomi, a close relative of S. marismortui, and again found similar rates of evolution (Graur and Pupko 2001 ). More recently, Nickle et al. (2002) also performed relative rate tests using 16S rDNA with the same result; the branch leading to isolate 2-9-3 is not extraordinarily short, as would be expected of an organism that has not been evolving for millions of years.Nickle et al. (2002) used evolutionary rates derived from enteric bacteria to argue that if isolate 2-9-3 has not been evolving for 250 Myr, then S. marismortui must itself have been evolving 5–10 times more slowly than did aphid endosymbionts on which the rate calculations were based. We note that although the evolutionary rates calculated from enterics and endosymbionts are the best estimates we currently possess, it is entirely likely that rates of sporeformer evolution may indeed be slower by several orders of magnitude. Sporeformers have been shown to remain in the metabolically dormant spore state, thus not replicating their DNA, for conservative estimates of anywhere from 102 to 104years between times of growth (Kennedy, Reader, and Swierczynski 1994 ; Nicholson et al. 2000 ).As the analyses discussed above used 16S rDNA genes, the evolution of which may not be representative of the organism as a whole, we wanted to know if the similarities between isolate 2-9-3 and S. marismortui are seen with protein-coding genes as well as with 16S rDNA genes. We therefore analyzed phylogenetic relationships between strain 2-9-3 and S. marismortui, using the spore-forming bacteria as our comparison group. The rationale for this design was that the evolutionary rate among the sporeformers would more closely approximate that of 2-9-3. We used amino acid data from two genes, recA and splB. The recA gene is found throughout all bacteria, and its product is required for homologous recombination and DNA repair. Because of the functional constraints on recA evolution, it can be used to resolve the older evolutionary relationships. The splB gene, on the other hand, has to date only been reported in gram-positive spore-forming bacteria and is important in the repair of spore-specific DNA damage resulting from UV radiation during spore dormancy (Nicholson et al. 2000
). Because splB is only found in gram-positive spore-forming bacteria, it can be assumed to have a more recent origin than recA has and might be useful in resolving closer evolutionary relationships.The results of our analyses are consistent with the phylogenetic relationships shown by Graur and Pupko (2001)
and Nickle et al. (2002) . At the nucleotide level, isolate 2-9-3 and S. marismortui differed by two nucleotides out of the 404 recA nucleotides examined. Both of these substitutions are synonymous, making these two taxa identical at the amino acid level. The phylogenetic reconstruction (Swofford 1998 ) using amino acid sequences of recA (amino acids were used because of site saturation at the nucleotide level across distantly related taxa) places 2-9-3 and S. marismortui in a more recent clade, instead of their occupying a more basal position as one would predict if the clade had not been evolving for 250 Myr (fig. 1 ).+++++
+++++
Vote neste blog para o prêmio TOPBLOG 2010.
