<Emphasis Type="Italic">In silico</Emphasis> comparison of bacterial strains using mutual information

Fast-sequencing throughput methods have increased the number of completely sequenced bacterial genomes to about 400 by December 2006, with the number increasing rapidly. These include several strains. In silico methods of comparative genomics are of use in categorizing and phylogenetically sorting these bacteria. Various word-based tools have been used for quantifying the similarities and differences between entire genomes. The simple di-nucleotide frequency comparison, codon specificity and k-mer repeat detection are among some of the well-known methods.In this paper, we show that the Mutual Information function, which is a measure of correlations and a concept from Information Theory, is very effective in determining the similarities and differences among genome sequences of various strains of bacteria such as the plant pathogen Xylella fastidiosa, marine Cyanobacteria Prochlorococcus marinus or animal and human pathogens such as species of Ehrlichia and Legionella. The short-range three-base periodicity, small sequence repeats and long-range correlations taken together constitute a genome signature that can be used as a technique for identifying new bacterial strains with the help of strains already catalogued in the database.There have been several applications of using the Mutual Information function as a measure of correlations in genomics but this is the first whole genome analysis done to detect strain similarities and differences.     

Exploration and grading of possible genes from 183 bacterial strains by a common protocol to identification of new genes: Gene Trek in Prokaryote Space (GTPS).

A large number of complete microorganism genomes has been sequenced and submitted to the public database and then incorporated into our complete genome database, Genome Information Broker (GIB, http://gib.genes.nig.ac.jp/). However, when comparative genomics is carried out, researchers must be aware that there are protein-coding genes not confirmed by homology or motif search and that reliable protein-coding genes are missing. Therefore, we developed a protocol (Gene Trek in Prokaryote Space, GTPS) for finding possible protein-coding genes in bacterial genomes. GTPS assigns a degree of reliability to predicted protein-coding genes. We first systematically applied the protocol to the complete genomes of all 123 bacterial species and strains that were publicly available as of July 2003, and then to those of 183 species and strains available as of September 2004. We found a number of incorrect genes and several new ones in the genome data in question. We also found a way to estimate the total number of orthologous genes in the bacterial world.     

The microbial pan-genome

A decade after the beginning of the genomic era, the question of how genomics can describe a bacterial species has not been fully addressed. Experimental data have shown that in some species new genes are discovered even after sequencing the genomes of several strains. Mathematical modeling predicts that new genes will be discovered even after sequencing hundreds of genomes per species. Therefore, a bacterial species can be described by its pan-genome, which is composed of a "core genome" containing genes present in all strains, and a "dispensable genome" containing genes present in two or more strains and genes unique to single strains. Given that the number of unique genes is vast, the pan-genome of a bacterial species might be orders of magnitude larger than any single genome.     

细菌比较基因组学和进化基因组学

通过比较不同细菌基因组间差异性与相似性,进而深入研究其分子机理,最终与其表型特征联系起来,是为比较基因组学;不同细菌经过长期进化,其基因组在结构与功能上存在着明显的分化,并构成表型进化的遗传基础,大量细菌全基因组测序的完成,细菌进化基因组学应运而生;以比较基因组学为研究手段,细菌进化基因组学可从基因组水平深入认识物种分化、生境适应、毒力进化、耐药性产生蔓延等表型进化过程。     

Study of statistical correlations in DNA sequences

Here we present a study of statistical correlations among different positions in DNA sequences and their implications by directly using the autocorrelation function. Such an analysis is possible now because of the availability of large sequences or even complete genomes of many organisms. After describing the way in which the autocorrelation function can be applied to DNA-sequence analysis, we show that long-range correlations, implying scale independence, appear in several bacterial genomes as well as in long human chromosome contigs. The source for such correlations in bacteria, which may extend up to 60 kb in Bacillus subtilis, may be related to massive lateral transfer of compositionally biased genes from other genomes. In the human genome, correlations extend for more than five decades and may be related to the evolution of the ’neogenome’, a modern evolutionary acquisition composed by GC-rich isochores displaying long-range correlations and scale invariance.     

Bacterial genomics

Abstract: During the last decade, great advances have been made in the study of bacterial genomes which is perhaps better described by the term bacterial genomics. The application of powerful techniques, such as pulsed-field gel electrophoresis of macro-restriction fragments of genomic DNA, has freed the characterisation of the chromosomes of many bacteria from the constraints imposed by classical genetic analysis. It is now possible to analyse the genome of virtually every microorganism by direct molecular methods and to construct detailed physical and gene maps. In this review, the various practical approaches are compared and contrasted, and some of the emerging themes of bacterial genomics, such as the size, shape, number and organisation of chromosomes are discussed.     

Analysis of a Leptospira interrogans locus containing DNA replication genes and a new IS,IS1502

Comparative bacterial genomics shows that even different isolates of the same bacterial species can vary significantly in gene content. An effective means to survey differences across whole genomes would be highly advantageous for understanding this variation. Here we show that suppression subtractive hybridization (SSH) provides high, representative coverage of regions that differ between similar genomes. Using Helicobacter pylori strains 26695 and J99 as a model, SSH identified approximately 95% of the unique open reading frames in each strain, showing that the approach is effective. Furthermore, combining data from parallel SSH experiments using different restriction enzymes significantly increased coverage compared to using a single enzyme. These results suggest a powerful approach for assessing genome differences among closely related strains when one member of the group has been completely sequenced.     

Prophage genomics.

The majority of the bacterial genome sequences deposited in the National Center for Biotechnology Information database contain prophage sequences. Analysis of the prophages suggested that after being integrated into bacterial genomes, they undergo a complex decay process consisting of inactivating point mutations, genome rearrangements, modular exchanges, invasion by further mobile DNA elements, and massive DNA deletion. We review the technical difficulties in defining such altered prophage sequences in bacterial genomes and discuss theoretical frameworks for the phage-bacterium interaction at the genomic level. The published genome sequences from three groups of eubacteria (low- and high-G+C gram-positive bacteria and gamma-proteobacteria) were screened for prophage sequences. The prophages from Streptococcus pyogenes served as test case for theoretical predictions of the role of prophages in the evolution of pathogenic bacteria. The genomes from further human, animal, and plant pathogens, as well as commensal and free-living bacteria, were included in the analysis to see whether the same principles of prophage genomics apply for bacteria living in different ecological niches and coming from distinct phylogenetical affinities. The effect of selection pressure on the host bacterium is apparently an important force shaping the prophage genomes in low-G+C gram-positive bacteria and gamma-proteobacteria.     

Mining the genomes of plant pathogenic bacteria: how not to drown in gigabases of sequence

Hundreds of bacterial genomes including the genomes of dozens of plant pathogenic bacteria have been sequenced. These genomes represent an invaluable resource for molecular plant pathologists. In this review, we describe different approaches that can be used for mining bacterial genome sequences and examples of how some of these approaches have been used to analyse plant pathogen genomes so far. We review how genomes can be mined one by one and how comparative genomics of closely related genomes releases the true power of genomics. Databases and tools useful for genome mining that are publicly accessible on the Internet are also described. Finally, the need for new databases and tools to efficiently mine today's plant pathogen genomes and hundreds more in the near future is discussed.     

Comparative Analysis of the Mosaic Genomes of Tailed Archaeal Viruses and Proviruses Suggests Common Themes for Virion Architecture and Assembly with Tailed Viruses of Bacteria

Tailed double-stranded DNA viruses (order Caudovirales) represent the dominant morphotype among viruses infecting bacteria. Analysis and comparison of complete genome sequences of tailed bacterial viruses provided insights into their origin and evolution. Structural and genomic studies have unexpectedly revealed that tailed bacterial viruses are evolutionarily related to eukaryotic herpesviruses. Organisms from the third domain of life, Archaea, are also infected by viruses that, in their overall morphology, resemble tailed viruses of bacteria. However, high-resolution structural information is currently unavailable for any of these viruses, and only a few complete genomes have been sequenced so far. Here we identified nine proviruses that are clearly related to tailed bacterial viruses and integrated into chromosomes of species belonging to four different taxonomic orders of the Archaea. This more than doubled the number of genome sequences available for comparative studies. Our analyses indicate that highly mosaic tailed archaeal virus genomes evolve by homologous and illegitimate recombination with genomes of other viruses, by diversification, and by acquisition of cellular genes. Comparative genomics of these viruses and related proviruses revealed a set of conserved genes encoding putative proteins similar to virion assembly and maturation, as well as genome packaging proteins of tailed bacterial viruses and herpesviruses. Furthermore, fold prediction and structural modeling experiments suggest that the major capsid proteins of tailed archaeal viruses adopt the same topology as the corresponding proteins of tailed bacterial viruses and eukaryotic herpesviruses. Data presented in this study strongly support the hypothesis that tailed viruses infecting archaea share a common ancestry with tailed bacterial viruses and herpesviruses.     

How genomics has affected the concept of microbiology

Genomics influences multiple areas of microbiology, and thus affects key microbiological concepts. Recent reports that describe the large genome and unusual coding capacity of mimivirus, the minimized fungal genomes that contain elements of bacterial metabolism, and the 'signature' eukaryotic proteins in bacteria are introducing grey shades into the black-and-white distinctions between microbial domains. The concept of the 'universal' minimal genome is being challenged, and the ability of minimal genomes to support cellular complexity is under investigation. There have been intriguing insights into microbe-microbe relationships, for example conflict mediation in competing bacteriophages that rapidly evolve survival mechanisms when cooperation is experimentally enforced. Genomics has given birth to metagenomics, but has also stimulated the development of improved cultivation techniques. Lastly, the taxonomic potential of genomics is emerging, as studies of multiple strains allow us to revise and refine the bacterial species concept as well as the idea of a static genome.     

Rickettsial evolution in the light of comparative genomics

Rickettsia are best known as strictly intracellular vector‐borne bacteria that cause mild to severe diseases in humans and other animals. Recent advances in molecular tools and biological experiments have unveiled a wide diversity of Rickettsia spp. that include species with a broad host range and some species that act as endosymbiotic associates. Molecular phylogenies of Rickettsia spp. contain some ambiguities, such as the position of R. canadensis and relationships within the spotted fever group. In the modern era of genomics, with an ever‐increasing number of sequenced genomes, there is enhanced interest in the use of whole‐genome sequences to understand pathogenesis and assess evolutionary relationships among rickettsial species. Rickettsia have small genomes (1.1–1.5 Mb) as a result of reductive evolution. These genomes contain split genes, gene remnants and pseudogenes that, owing to the colinearity of some rickettsial genomes, may represent different steps of the genome degradation process. Genomics reveal extreme genome reduction and massive gene loss in highly vertebrate‐pathogenic Rickettsia compared to less virulent or endosymbiotic species. Information gleaned from rickettsial genomics challenges traditional concepts of pathogenesis that focused primarily on the acquisition of virulence factors. Another intriguing phenomenon about the reduced rickettsial genomes concerns the large fraction of non‐coding DNA and possible functionality of these “non‐coding” sequences, because of the high conservation of these regions. Despite genome streamlining, Rickettsia spp. contain gene families, selfish DNA, repeat palindromic elements and genes encoding eukaryotic‐like motifs. These features participate in sequence and functional diversity and may play a crucial role in adaptation to the host cell and pathogenesis. Genome analyses have identified a large fraction of mobile genetic elements, including plasmids, suggesting the possibility of lateral gene transfer in these intracellular bacteria. Phylogenetic analyses have identified several candidates for horizontal gene acquisition among Rickettsia spp. including tra, pat2, and genes encoding for the type IV secretion system and ATP/ADP translocase that may have been acquired from bacteria living in amoebae. Gene loss, gene duplication, DNA repeats and lateral gene transfer all have shaped rickettsial genome evolution. A comprehensive analysis of the entire genome, including genes and non‐coding DNA, will help to unlock the mysteries of rickettsial evolution and pathogenesis.     

Positive Selection and Horizontal Gene Transfer in the Genome of a Male-Killing Wolbachia

Wolbachia are a genus of widespread bacterial endosymbionts in which some strains can hijack or manipulate arthropod host reproduction. Male killing is one such manipulation in which these maternally transmitted bacteria benefit surviving daughters in part by removing competition with the sons for scarce resources. Despite previous findings of interesting genome features of microbial sex ratio distorters, the population genomics of male-killers remain largely uncharacterized. Here, we uncover several unique features of the genome and population genomics of four Arizonan populations of a male-killing Wolbachia strain, wInn, that infects mushroom-feeding Drosophila innubila. We first compared the wInn genome with other closely related Wolbachia genomes of Drosophila hosts in terms of genome content and confirm that the wInn genome is largely similar in overall gene content to the wMel strain infecting D. melanogaster. However, it also contains many unique genes and repetitive genetic elements that indicate lateral gene transfers between wInn and non-Drosophila eukaryotes. We also find that, in line with literature precedent, genes in the Wolbachia prophage and Octomom regions are under positive selection. Of all the genes under positive selection, many also show evidence of recent horizontal transfer among Wolbachia symbiont genomes. These dynamics of selection and horizontal gene transfer across the genomes of several Wolbachia strains and diverse host species may be important underlying factors in Wolbachia’s success as a male-killer of divergent host species.     

A close relationship between primary nucleotides sequence structure and the composition of functional genes in the genome of prokaryotes

Comparative genomics is an essential tool to unravel how genomes change over evolutionary time and to gain clues on the links between functional genomics and evolution. In prokaryotes, the large, good quality, genome sequences available in public databases and the recently developed large-scale computational methods, offer an unprecedent view on the ecology and evolution of microorganisms through comparative genomics. In this work, we examined the links among genome structure (i.e., the sequential distribution of nucleotides itself by detrended fluctuation analysis, DFA) and genomic diversity (i.e., gene functionality by Clusters of Orthologous Genes, COGs) in 828 full sequenced prokaryotic genomes from 548 different bacteria and archaea species. DFA scaling exponent α indicated persistent long-range correlations (fractality) in each genome analyzed. Higher resolution power was found when considering the sequential succession of purine (AG) vs. pyrimidine (CT) bases than either keto (GT) to amino (AC) forms or strongly (GC) vs. weakly (AT) bonded nucleotides. Interestingly, the phyla Aquificae, Fusobacteria, Dictyoglomi, Nitrospirae, and Thermotogae were closer to archaea than to their bacterial counterparts. A strong significant correlation was found between scaling exponent α and COGs distribution, and we consistently observed that the larger α the more heterogeneous was the gene distribution within each functional category, suggesting a close relationship between primary nucleotides sequence structure and functional genes composition.     

Spectrum of heart diseases in a new cardiac service in Nigeria: An echocardiographic study of 1441 subjects in Abeokuta

Background

The recent determination of the complete nucleotide sequence of several Mycobacterium tuberculosis (MTB) genomes allows the use of comparative genomics as a tool for dissecting the nature and consequence of genetic variability within this species. The multiple alignment of the genomes of clinical strains (CDC1551, F11, Haarlem and C), along with the genomes of laboratory strains (H37Rv and H37Ra), provides new insights on the mechanisms of adaptation of this bacterium to the human host.

Findings

The genetic variation found in six M. tuberculosis strains does not involve significant genomic rearrangements. Most of the variation results from deletion and transposition events preferentially associated with insertion sequences and genes of the PE/PPE family but not with genes implicated in virulence. Using a Perl-based software islandsanalyser, which creates a representation of the genetic variation in the genome, we identified differences in the patterns of distribution and frequency of the polymorphisms across the genome. The identification of genes displaying strain-specific polymorphisms and the extrapolation of the number of strain-specific polymorphisms to an unlimited number of genomes indicates that the different strains contain a limited number of unique polymorphisms.

Conclusion

The comparison of multiple genomes demonstrates that the M. tuberculosis genome is currently undergoing an active process of gene decay, analogous to the adaptation process of obligate bacterial symbionts. This observation opens new perspectives into the evolution and the understanding of the pathogenesis of this bacterium.     

Heterogeneity of genome sizes among natural isolates of Escherichia coli.

Comparisons of the genetic maps of Escherichia coli K-12 and Salmonella typhimurium LT2 suggest that the size and organization of bacterial chromosomes are highly conserved. Employing pulsed-field gel electrophoresis, we have estimated the extent of variation in genome size among 14 natural isolates of E. coli. The BlnI and NotI restriction fragment patterns were highly variable among isolates, and genome sizes ranged from 4,660 to 5,300 kb, which is several hundred kilobases larger than the variation detected between enteric species. Genome size differences increase with the evolutionary genetic distance between lineages of E. coli, and there are differences in genome size among the major subgroups of E. coli. In general, the genomes of natural isolates are larger than those of laboratory strains, largely because of the fact that laboratory strains were derived from the subgroup of E. coli with the smallest genomes.     

Identification and localization of differences between Escherichia coli and Salmonella typhimurium genomes by suppressive subtractive hybridization

The availability of bacterial genome sequences raises an important new problem - how can one move from completely sequenced microorganisms as a reference to the hundreds and thousands of other strains or isolates of the same or related species that will not be sequenced in the near future? An efficient way to approach this task is the comparison of genomes by subtractive hybridization. Recently we developed a sensitive and reproducible subtraction procedure for comparison of bacterial genomes, based on the method of suppression subtractive hybridization (SSH). In this work we demonstrate the applicability of subtractive hybridization to the comparison of the related but markedly divergent bacterial species Escherichia coli and Salmonella typhimurium. Clone libraries representing sequence differences were obtained and, in the case of completely sequenced E. coli genome, the differences were directly placed in the genome map. About 60% of the differential clones identified by SSH were present in one of the genomes under comparison and absent from the other. Additional differences in most cases represent sequences that have diverged considerably in the course of evolution. Such an approach to comparative bacterial genomics can be applied both to studies of interspecies evolution - to elucidate the "strategies" that enable different genomes to fit their ecological niches - and to development of diagnostic probes for the rapid identification of pathogenic bacterial species.     

Microbial ecology in the age of genomics and metagenomics: concepts, tools, and recent advances

Microbial ecology examines the diversity and activity of micro-organisms in Earth's biosphere. In the last 20 years, the application of genomics tools have revolutionized microbial ecological studies and drastically expanded our view on the previously underappreciated microbial world. This review first introduces the basic concepts in microbial ecology and the main genomics methods that have been used to examine natural microbial populations and communities. In the ensuing three specific sections, the applications of the genomics in microbial ecological research are highlighted. The first describes the widespread application of multilocus sequence typing and representational difference analysis in studying genetic variation within microbial species. Such investigations have identified that migration, horizontal gene transfer and recombination are common in natural microbial populations and that microbial strains can be highly variable in genome size and gene content. The second section highlights and summarizes the use of four specific genomics methods (phylogenetic analysis of ribosomal RNA, DNA-DNA re-association kinetics, metagenomics, and micro-arrays) in analysing the diversity and potential activity of microbial populations and communities from a variety of terrestrial and aquatic environments. Such analyses have identified many unexpected phylogenetic lineages in viruses, bacteria, archaea, and microbial eukaryotes. Functional analyses of environmental DNA also revealed highly prevalent, but previously unknown, metabolic processes in natural microbial communities. In the third section, the ecological implications of sequenced microbial genomes are briefly discussed. Comparative analyses of prokaryotic genomic sequences suggest the importance of ecology in determining microbial genome size and gene content. The significant variability in genome size and gene content among strains and species of prokaryotes indicate the highly fluid nature of prokaryotic genomes, a result consistent with those from multilocus sequence typing and representational difference analyses. The integration of various levels of ecological analyses coupled to the application and further development of high throughput technologies are accelerating the pace of discovery in microbial ecology.     

Functional genomics of bacterial pathogens: from post-genomics to therapeutic targets

A wealth of new data have become available to the scientific community as a result of the sequencing of many pathogen genomes. A recent meeting devoted to functional genomics of pathogenic microorganisms confirmed the notion that bacterial genomes are not static, because large blocks of genes can be acquired or deleted. Less complex environments usually result in reduction in genome size, while genome expansion is usually associated with environmental change and complexity. During the meeting, pathogenicity and evolutionary aspects were illustrated for enteric pathogens, as well as the microevolution of the plague bacillus Yersinia pestis. New clues for evolution and pathogenicity were derived from comparative genomics of Listeria species. The genomic organization of Bartonellae, an emerging human pathogen, was also discussed in an evolutionary context. Population and functional genomics of Anthrax-causing bacteria highlighted current scientific interest in this potential biothreat.