W. Ford Doolittle

The complete genome of the crenarchaeon Sulfolobus solfataricus P2

The genome of the crenarchaeon Sulfolobus solfataricus P2 contains 2,992,245 bp on a single chromosome and encodes 2,977 proteins and many RNAs. One-third of the encoded proteins have no detectable homologs in other sequenced genomes. Moreover, 40% appear to be archaeal-specific, and only 12% and 2.3% are shared exclusively with bacteria and eukarya, respectively. The genome shows a high level of plasticity with 200 diverse insertion sequence elements, many putative nonautonomous mobile elements, and evidence of integrase-mediated insertion events. There are also long clusters of regularly spaced tandem repeats. Different transfer systems are used for the uptake of inorganic and organic solutes, and a wealth of intracellular and extracellular proteases, sugar, and sulfur metabolizing enzymes are encoded, as well as enzymes of the central metabolic pathways and motility proteins. The major metabolic electron carrier is not NADH as in bacteria and eukarya but probably ferredoxin. The essential components required for DNA replication, DNA repair and recombination, the cell cycle, transcriptional initiation and translation, but not DNA folding, show a strong eukaryal character with many archaeal-specific features. The results illustrate major differences between crenarchaea and euryarchaea, especially for their DNA replication mechanism and cell cycle processes and their translational apparatus.

Use of 16S rRNA and rpoB Genes as Molecular Markers for Microbial Ecology Studies

ABSTRACT Several characteristics of the 16S rRNA gene, such as its essential function, ubiquity, and evolutionary properties, have allowed it to become the most commonly used molecular marker in microbial ecology. However, one fact that has been overlooked is that multiple copies of this gene are often present in a given bacterium. These intragenomic copies can differ in sequence, leading to identification of multiple ribotypes for a single organism. To evaluate the impact of such intragenomic heterogeneity on the performance of the 16S rRNA gene as a molecular marker, we compared its phylogenetic and evolutionary characteristics to those of the single-copy gene rpoB. Full-length gene sequences and gene fragments commonly used for denaturing gradient gel electrophoresis were compared at various taxonomic levels. Heterogeneity found between intragenomic 16S rRNA gene copies was concentrated in specific regions of rRNA secondary structure. Such “heterogeneity hot spots” occurred within all gene fragments commonly used in molecular microbial ecology. This intragenomic heterogeneity influenced 16S rRNA gene tree topology, phylogenetic resolution, and operational taxonomic unit estimates at the species level or below. rpoB provided comparable phylogenetic resolution to that of the 16S rRNA gene at all taxonomic levels, except between closely related organisms (species and subspecies levels), for which it provided better resolution. This is particularly relevant in the context of a growing number of studies focusing on subspecies diversity, in which single-copy protein-encoding genes such as rpoB could complement the information provided by the 16S rRNA gene.

Pattern pluralism and the Tree of Life hypothesis

Darwin claimed that a unique inclusively hierarchical pattern of relationships between all organisms based on their similarities and differences [the Tree of Life (TOL)] was a fact of nature, for which evolution, and in particular a branching process of descent with modification, was the explanation. However, there is no independent evidence that the natural order is an inclusive hierarchy, and incorporation of prokaryotes into the TOL is especially problematic. The only data sets from which we might construct a universal hierarchy including prokaryotes, the sequences of genes, often disagree and can seldom be proven to agree. Hierarchical structure can always be imposed on or extracted from such data sets by algorithms designed to do so, but at its base the universal TOL rests on an unproven assumption about pattern that, given what we know about process, is unlikely to be broadly true. This is not to say that similarities and differences between organisms are not to be accounted for by evolutionary mechanisms, but descent with modification is only one of these mechanisms, and a single tree-like pattern is not the necessary (or expected) result of their collective operation. Pattern pluralism (the recognition that different evolutionary models and representations of relationships will be appropriate, and true, for different taxa or at different scales or for different purposes) is an attractive alternative to the quixotic pursuit of a single true TOL.

Differences between Tissue-Associated Intestinal Microfloras of Patients with Crohn's Disease and Ulcerative Colitis

ABSTRACT A leading hypothesis for the role of bacteria in inflammatory bowel diseases is that an imbalance in normal gut flora is a prerequisite for inflammation. Testing this hypothesis requires comparisons between the microbiota compositions of ulcerative colitis and Crohns disease patients and those of healthy individuals. In this study, we obtained biopsy samples from patients with Crohns disease and ulcerative colitis and from healthy controls. Bacterial DNA was extracted from the tissue samples, amplified using universal bacterial 16S rRNA gene primers, and cloned into a plasmid vector. Insert-containing colonies were picked for high-throughput sequencing, and sequence data were analyzed, yielding species-level phylogenetic data. The clone libraries yielded 3,305 sequenced clones, representing 151 operational taxonomical units. There was no significant difference between floras from inflamed and healthy tissues from within the same individual. Proteobacteria were significantly (P = 0.0007) increased in Crohns disease patients, as were Bacteroidetes (P < 0.0001), while Clostridia were decreased in that group (P < 0.0001) in comparison with the healthy and ulcerative colitis groups, which displayed no significant differences. Thus, the bacterial flora composition of Crohns patients appears to be significantly altered from that of healthy controls, unlike that of ulcerative colitis patients. Imbalance in flora in Crohns disease is probably not sufficient to cause inflammation, since microbiotas from inflamed and noninflamed tissues were of similar compositions within the same individual.

Archaea and the Origin(s) of DNA Replication Proteins

Replication thus joins transcription and translation in compounding the central conundrum of cellular evolution, illustrated in Figure 1Figure 1. The archaeal molecular components responsible for these fundamental cellular processes are more similar to their eukaryotic than their bacterial counterparts. We expect this in a quantitative sense if the structure of the universal tree shown in Figure 1Figure 1 is correct—in fact, such quantitative similarity is the basis of the tree. But there are also qualitative differences between bacteria and archaea–eukaryotes—whole suites of proteins and protein complexes present in the one lineage and absent from the other. Often, the archaeal–eukaryotic system seems more complex (involves more components). And, for much of the latter half of this century we have thought of such complexity as part and parcel of the greater complexity in cellular ultrastructure (in particular, the presence of a cytoskeleton and endomembrane system) that distinguishes eukaryotes from all prokaryotes, archaea included.Perhaps there is no conundrum—perhaps we were just wrong to think that way. But still, the uncoupling of molecular complexity in transcription, translation, and replication from complexity in cell structure is most peculiar. The eukaryotic cytoskeletal system seems to have arisen full blown at the origin of the eukaryotes. Although bacterial and archaeal FtsZ is a likely candidate for the prokaryote homolog of tubulin, amino acid alignments between these proteins are not very convincing. Russell Doolittle (Doolittle 1995xDoolittle, R.F. Philos. Trans. R. Soc. (Lond.) B. 1995; 349: 235–240Crossref | PubMedSee all ReferencesDoolittle 1995) has noted that the evolution of the eukaryotic cytoskeleton would require an abrupt change in the rate of sequence evolution for tubulins and their relatives that is of the order of “10- to 100-fold higher” than the observed rate of sequence evolution of tubulins and FtsZ proteins within the domains Bacteria, Archaea, and Eucarya. This rate of sequence evolution might also be invoked if DNA replication proteins of the three domains are in fact homologs and diverged from a common ancestral set of proteins.

The role of lateral gene transfer in the evolution of isoprenoid biosynthesis pathways

Lateral gene transfer (LGT) is a major force in microbial genome evolution. Here, we present an overview of lateral transfers affecting genes involved in isopentenyl diphosphate (IPP) synthesis. Two alternative metabolic pathways can synthesize this universal precursor of isoprenoids, the 1‐deoxy‐d‐xylulose 5‐phosphate (DOXP) pathway and the mevalonate (MVA) pathway. We have surveyed recent genomic data and the biochemical literature to determine the distribution of the genes composing these pathways within the bacterial domain. The scattered distribution observed is incompatible with a simple scheme of vertical transmission. LGT (among and between bacteria, archaea and eukaryotes) more parsimoniously explains many features of this pattern. This alternative scenario is supported by phylogenetic analyses, which unambiguously confirm several cases of lateral transfer. Available biochemical data allow the formulation of hypotheses about selective pressures favouring transfer. The phylogenetic diversity of the organisms involved and the range of possible causes and effects of these transfer events make the IPP biosynthetic pathways an ideal system for studying the evolutionary role of LGT.

On the chimeric nature, thermophilic origin, and phylogenetic placement of the Thermotogales

Since publication of the first Thermotogales genome, Thermotoga maritima strain MSB8, single- and multi-gene analyses have disagreed on the phylogenetic position of this order of Bacteria. Here we present the genome sequences of 4 additional members of the Thermotogales (Tt. petrophila, Tt. lettingae, Thermosipho melanesiensis, and Fervidobacterium nodosum) and a comprehensive comparative analysis including the original T. maritima genome. While ribosomal protein genes strongly place Thermotogales as a sister group to Aquificales, the majority of genes with sufficient phylogenetic signal show affinities to Archaea and Firmicutes, especially Clostridia. Indeed, on the basis of the majority of genes in their genomes (including genes that are also found in Aquificales), Thermotogales should be considered members of the Firmicutes. This result highlights the conflict between the taxonomic goal of assigning every species to a unique position in an inclusive Linnaean hierarchy and the evolutionary goal of understanding phylogenesis in the presence of pervasive horizontal gene transfer (HGT) within prokaryotes. Amino acid compositions of reconstructed ancestral sequences from 423 gene families suggest an origin of this gene pool even more thermophilic than extant members of this order, followed by adaptation to lower growth temperatures within the Thermotogales.

Lateral gene transfer

Summary The four disparate images shown in Figure 1 have this in common: each represents a radical adaptation that would not have happened had lateral gene transfer (LGT), also known as horizontal gene transfer (HGT), not been the powerful evolutionary force we now know it to be. Those who study the phenomenon are still struggling to quantitatively assess LGT as a process or processes and accommodate its implications for how patterns in nature should be represented — such as the existence of definable species or a meaningful universal Tree of Life. But all agree that the exchange of genetic information across species lines — which is how we will define LGT in this primer — is far more pervasive and more radical in its consequences than we could have guessed just a decade ago. Both prokaryotes (bacteria and archaea) and eukaryotes have experienced LGT, though its potential as a source of novel adaptations and as a challenge to phylogenetics are so far more obvious and better understood for prokaryotes, as are the mechanisms by which it is effected.

Molecular Biological Mechanisms of Speciation

Growing recognition that much of the evolutionary history of eukaryotic genomes reflects the operation of turnover processes involving repetitive DNA sequences has led to the recent formulation of models describing speciation as a consequence of such turnover. These models are of three general kinds: those attributing hybrid infertility to the process of transposition, those attributing hybrid infertility to mispairing between chromosomes of divergent repetitive DNA composition, and those assuming that change in repetitive DNAs can reset coordinated gene regulation. These models are discussed with respect to the kinds of evidence needed for their corroboration and to their significance for questions related to macroevolutionary punctuated equilibria and genetic revolutions.

Reconstructing/Deconstructing the Earliest Eukaryotes: How Comparative Genomics Can Help

We could reconstruct the evolution of eukaryote-specific molecular and cellular machinery if some living eukaryotes retained primitive cellular structures and we knew which eukaryotes these were. Its not clear that either is the case, but the expanding protist genomic database could help us in several ways.