Measurement of heat evolution and correlation with oxygen consumption during microbial growth.
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We examined the distributions of short tandemly repeated DNAs (microsatellites) in nine complete microbial genomes (Saccharomyces cerevisiae, Archaeoglobus fulgidus, Escherichia coli, Haemophilus influenzae, Helicobacter pylori, Methanococcus jannaschii, Mycoplasma pneumoniae, M. genitalium, and Synechocystis PCC6803.) These repeats contribute differently to the global features of these genomes, and we explore the evolutionary implications of these differences by empirical examination of length polymorphisms at 20 long triplet-repeats repeats in S. cerevisiae, and by comparison of observed and expected repeat distributions. All of a sample of 20 microsatellites found in S. cerevisiae are highly polymorphic in length, suggesting that mutation pressure overcomes overall selection for small genome size that will tend to shorten or eliminate unnecessary DNA. By comparison, prokaryotes have fewer long repeats than expected, except for a few statistically improbable repeats that appear to function in gene regulation. Finally, we find that in all these genomes there is an excess of repeats shorter than those traditionally considered to be microsatellites. This finding suggests that even in prokaryotes these repeats are being generated by mutational pressures. These results have important potential implications for understanding genome stability and evolution in these microbial species.
Recent geophysical analyses suggest the presence of a late Paleozoic oxygen pulse beginning in the late Devonian and continuing through to the late Carboniferous. During this period, plant terrestrialization and global carbon deposition resulted in a dramatic increase in atmospheric oxygen levels, ultimately yielding concentrations potentially as high as 35% relative to the contemporary value of 21%. Such hyperoxia of the late Paleozoic atmosphere may have physiologically facilitated the initial evolution of insect flight metabolism. Widespread gigantism in late Paleozoic insects and other arthropods is also consistent with enhanced oxygen flux within diffusion-limited tracheal systems. Because total atmospheric pressure increases with increased oxygen partial pressure, concurrently hyperdense conditions would have augmented aerodynamic force production in early forms of flying insects. By the late Permian, evolution of decompositional microbial and fungal communities, together with disequilibrium in rates of carbon deposition, gradually reduced oxygen concentrations to values possibly as low as 15%. The disappearance of giant insects by the end of the Permian is consistent with extinction of these taxa for reasons of asphyxiation on a geological time scale. As with winged insects, the multiple historical origins of vertebrate flight in the late Jurassic and Cretaceous correlate temporally with periods of elevated atmospheric oxygen. Much discussion of flight performance in Archaeopteryx assumes a contemporary atmospheric composition. Elevated oxygen levels in the mid- to late Mesozoic would, however, have facilitated aerodynamic force production and enhanced muscle power output for ancestral birds, as well as for precursors to bats and pterosaurs.
The introduction of methods for automated DNA sequence analysis nearly a decade ago, together with more recent advances in the field of bioinformatics, have revolutionized biology and medicine and have ushered in a new era of genomic science, the study of genes and genomes. These new technologies have had an impact on many areas of research, including the association between genes and disease, in DNA-based diagnostics, and in the sequencing of genomes from human and other model organisms. The demonstration in 1995, that automated DNA sequencing methods could be used to decipher the entire genome sequence of a free-living organism, Haemophilus influenzae, was a milestone in both the genomics and microbial fields [1]. Since the first report of the complete sequence of H. influenzae, these methodologies have been adopted by laboratories around the world. The complete genomic sequence of five eubacterial species [1-5], one archaea [6], and the eukaryote, Saccharomyces cerevisiae [7], have been reported in the last 18 months. At the beginning of 1997 more than a dozen microbial genome projects are at or near completion, with many others in progress. It is likely that in the next few years we will see the complete sequence of perhaps as many as 30-40 microbial genomes. In this article, we will review methods for whole genome sequencing and analysis and examine how this information can be exploited to better understand microbial physiology and evolution.
The human intestinal microbiota is a dynamic ecosystem shaped by extensive horizontal gene transfer, particularly in individuals from industrialized populations. In this review, we discuss recent advances in our understanding of how mobile genetic elements (MGEs) contribute to microbial ecology and evolution in this diverse community, focusing on MGEs carrying fitness-conferring genes. Bacteroidales species can colonize individuals for decades and serve as major hubs for MGE exchange. Most MGEs are highly variable across individuals and geographies. Occasionally, conserved MGEs can spread across geography and lifestyles. Functional characterizations of MGEs reveal their roles in antibiotic resistance, interbacterial antagonism, biofilm formation, immune evasion, and nutrient acquisition, among others. Substantive progress in our understanding of MGEs in the gut microbiome offers promising avenues for therapeutic microbiome interventions. However, major challenges remain in functional prediction, host-MGE linkage, and experimental characterization.
Most mutations are likely to be deleterious, and so the spontaneous mutation rate is generally held at a very low value. Nonetheless, evolutionary theory predicts that high mutation rates can evolve under certain circumstances. Empirical observations have previously been limited to short-term studies of the fates of mutator strains deliberately introduced into laboratory populations of Escherichia coli, and to the effects of intense selective events on mutator frequencies in E. coli. Here we report the rise of spontaneously originated mutators in populations of E. coli undergoing long-term adaptation to a new environment. Our results corroborate computer simulations of mutator evolution in adapting clonal populations, and may help to explain observations that associate high mutation rates with emerging pathogens and with certain cancers.
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The Ribosomal RNA Operon Copy Number Database (rrndb) is an Internet-accessible database containing annotated information on rRNA operon copy number among prokaryotes. Gene redundancy is uncommon in prokaryotic genomes, yet the rRNA genes can vary from one to as many as 15 copies. Despite the widespread use of 16S rRNA gene sequences for identification of prokaryotes, information on the number and sequence of individual rRNA genes in a genome is not readily accessible. In an attempt to understand the evolutionary implications of rRNA operon redundancy, we have created a phylogenetically arranged report on rRNA gene copy number for a diverse collection of prokaryotic microorganisms. Each entry (organism) in the rrndb contains detailed information linked directly to external websites including the Ribosomal Database Project, GenBank, PubMed and several culture collections. Data contained in the rrndb will be valuable to researchers investigating microbial ecology and evolution using 16S rRNA gene sequences. The rrndb web site is directly accessible on the WWW at http://rrndb.cme. msu.edu.
The effect of seeding on the thermophilic composting of sewage sludge was examined by measuring the changes in CO2 evolution rates and microbial numbers. Although the succession of thermophilic bacteria and thermophilic actinomycetes clearly reflected the effect of seeding, no clear difference was observed in the overall rate of composting or quality of the composted product.
A Flavobacterium sp. (strain 50001), capable of degrading 2,4-dichlorophenoxyacetate (2,4-D), 2-methyl-4-chlorophenoxyacetate, and 2-chlorobenzoate and imparting resistance to mercury, harbored a degradative plasmid, pRC10. Cured strains of the Flavobacterium sp. lost the plasmid as well as the ability to degrade these chlorinated compounds. Comparison of this plasmid with the well-characterized 2,4-D-degradative plasmid pJP4 from Alcaligenes eutrophus showed regions of homology between the two plasmids. Restriction fragments of plasmid pRC10 which shared homology with the regions conferring 2,4-D-degradative genes (tfd) of plasmid pJP4 were cloned into a broad-host-range plasmid and studied in Pseudomonas putida. From the results obtained, the cloned DNA fragment expressed the genes for 2,4-D monooxygenase (tfdA) and 2,4-dichlorophenol hydroxylase (tfdB). In spite of the similarity in function, the size (45 kilobases) and restriction pattern of plasmid pRC10 were considerably different from those of pJP4 (80 kilobases). This may be due to the difference in the microbial background during evolution of the two plasmids.
The last decade has seen an explosion of interest in the use of molecular genetic techniques to diagnose infectious diseases. Specific molecular probes have been developed for nearly all of the significant, known microbial pathogens. The evolution of incrementally more powerful technologies, particularly gene amplification, has made it possible to detect pathogens with exquisite sensitivity, as well as specificity, based on their nucleic acids. As increasingly rapid and automated methods have become incorporated into successive generations of probe tests, these diagnostics have gained increasing acceptance for routine clinical use.
Fresh and wilted samples of Lolium italicum A. Br., Dactylis glomerata L., Medicago sativa L., have been ensiled in laboratory microsilos for the study of microflora evolution. The principal microbial groups (coliforms, proteolytic bacteria, clostridia, lactic acid bacteria, yeasts) have been detected and enumerated, in anaerobic atmosphere, after 8, 15, 22, 29, 36, 180 days of ensiling. Although the number of lactic acid bacteria resulted to the high in all the samples studied, they failed to perform a quick pH decrease and silage spoiling bacteria were not inhibited. This suggests the possibility of a cohabitation between useful and noxious bacteria, not affected by the chemical composition of ensiled plants.
In Drosophila, the Toll family of proteins are central to innate defense against microbial pathogens. Conserved throughout evolution, mammalian Toll-like receptors (TLRs) participate in innate immunity. TLRs mediate activation by microbial ligands including lipoproteins, resulting in the activation of IL-12 and nitric oxide synthase. Microbial lipoproteins also induce host cell apoptosis. In this manner, the ability of microbial lipoproteins to activate TLRs can contribute to host defense and immunopathology during infection.
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Microbial carbamoyl phosphate synthetases (CPS) use glutamine as nitrogen donor and are composed of two subunits (or domains), one exhibiting glutaminase activity, the other able to synthesize carbamoyl phosphate (CP) from bicarbonate, ATP, and ammonia. The pseudodimeric organization of this synthetase suggested that it has evolved by duplication of a smaller kinase, possibly a carbamate kinase (CK). In contrast to other prokaryotes the hyperthermophilic archaeon Pyrococcus furiosus was found to synthesize CP by using ammonia and not glutamine. We have purified the cognate enzyme and found it to be a dimer of two identical subunits of Mr 32,000. Its thermostability is considerable, 50% activity being retained after 1 h at 100 degrees C or 3 h at 95 degrees C. The corresponding gene was cloned by PCR and found to present about 50% amino acid identity with known CKs. The stoichiometry of the reaction (two ATP consumed per CP synthesized) and the ability of the enzyme to catalyze at high rate a bicarbonate-dependent ATPase reaction however clearly distinguish P. furiosus CPS from ordinary CKs. Thus the CPS of P. furiosus could represent a primeval step in the evolution of CPS from CK. Our results suggest that the first event in this evolution was the emergence of a primeval synthetase composed of subunits able to synthesize both carboxyphosphate and CP; this step would have preceded the duplication assumed to have generated the two subdomains of modern CPSs. The gene coding for this CK-like CPS was called cpkA.
Clinically and economically, penicillins and cephalosporins are the most important class of the beta-lactam antibiotics. They are produced by a wide variety of microorganisms including numerous species of Streptomyces, some unicellular bacteria and several filamentous fungi. A key step common to their biosynthetic pathways is the conversion of a linear, cysteine-containing tripeptide to a bicyclic beta-lactam antibiotic by isopenicillin N synthase. Recent successes in the cloning and expression of isopenicillin N synthase genes now permit production of a plentiful supply of this enzyme, which may be used for structural and mechanistic studies, or for biotechnological applications in the creation of novel beta-lactam compounds from peptide analogues. New ideas concerning the evolution and prevalence of the penicillin and cephalosporin biosynthetic genes have emerged from studies of isopenicillin N synthase genes.