Analysis of ultra low genome conservation in Clostridium difficile

Microarray-based comparative genome hybridisations (CGH) and genome sequencing of Clostridium difficile isolates have shown that the genomes of this species are highly variable. To further characterize their genome variation, we employed integration of data from CGH, genome sequencing and putative cellular pathways. Transcontinental strain comparison using CGH data confirmed the emergence of a human-specific hypervirulent cluster. However, there was no correlation between total toxin production and hypervirulent phenotype, indicating the possibility of involvement of additional factors towards hypervirulence. Calculation of C. difficile core and pan genome size using CGH and sequence data estimated that the core genome is composed of 947 to 1,033 genes and a pan genome comprised of 9,640 genes. The reconstruction, annotation and analysis of cellular pathways revealed highly conserved pathways despite large genome variation. However, few pathways such as tetrahydrofolate biosynthesis were found to be variable and could be contributing to adaptation towards virulence such as antibiotic resistance.     

Molecular phylogeny of RPB2 gene reveals multiple origin, geographic differentiation of H genome, and the relationship of the Y genome to other genomes in Elymus species

It has been hypothesized from isozymic and cytological studies of Elymus species that the Old and New World taxa may be of separate origin of the H genome in the StH genome species. To test this hypothesis, and estimate the phylogenetic relationships of polyploid Elymus species within the Triticeae, the second largest subunit of RNA polymerase II (RPB2) sequence of 36 Elymus accessions containing StH or StY genomes was analyzed with those of Pseudoroegneria (St), Hordeum (H), Agropyron (P), Australopyrum (W), Lophopyrum(Ee), Thinopyrum(Eb) and Dasypyrum (V). Our data indicated that the H genome in Elymus species is differentiated in accordance with geographical origin, and that the Eurasian and American StH genome species have independent alloploid origins with different H-genome donors. Phylogenetic analysis of Y genome sequences with other genome donors (St, H, P, W) of Elymus revealed that W and P genomes are sister to Y genome with a 87% bootstrap support, and that StY and StH species group might have acquired their RPB2 St sequences from distinct Pseudoroegneria gene pools. Our data did not support the suggestion that the St and Y genomes have the same origin as put forward in a previous study using ITS data. Our result provides some insight on the origin of Y genome and its relationship to other genomes in Elymus.     

Implication of gene distribution in the bacterial chromosome for the bacterial cell factory

As bacterial genome sequences accumulate, more and more pieces of data suggest that there is a significant correlation between the distribution of genes along the chromosome and the physical architecture of the cell, suggesting that the map of the cell is in the chromosome. Considering sequences and experimental data indicative of cell compartmentalisation, mRNA folding and turnover, as well as known structural features of protein and membrane complexes, we show that preliminary in silico analysis of whole genome sequences strongly substantiates this hypothesis. If there is a correlation between the genome sequence and the cell architecture, it must derive from some selection pressure in the organisms growing in the wild. As a consequence, the underlying constraints should be optimised in genetically modified organisms if one is to expect high product yields. Consequences in terms of gene expression for biotechnology are straightforward: knocking genes out and in genomes should not be randomly performed, but should follow the rules of chromosome organisation.     

Construction of the bacterial genome--the next goal of genetic engineering

The article deals primarily with literature data on the organization and evolution of bacterial genome. An account of the conception of "current" bacterial evolution consisting of periodical inclusions into a bacterial genome of genes having exogenous origin is given. A hypothesis is proposed that rare inversions going through the oriC or terC chromosomal regions could serve as reparative rearrangements providing balanced distribution of the DNA segments of different origin within the circular genome. It is asserted that the progress in the study of the model microorganism Escherichia coli K-12 as well as in genetical methods give rise to a new experimental challenge--the creation of a "rational" bacterial genome on the basis of E. coli cell. It is proposed that a "rational" bacterial genome could be devoid of many genes which provide the evolutionary formed ecological potential of the species and its capacity for rapid adaptation to new environment.     

Episomal amplification or chromosomal integration of the viral genome: alternative pathways in hamster polyomavirus-induced lymphomas.

The state and expression of the hamster polyomavirus genome in a large panel of virus-induced lymphomas have been investigated. The viral genome is present within tumor cells either as abundant nonrandomly deleted extrachromosomal copies or as a single copy integrated into cellular DNA. We show that these two physical states are likely to be functionally equivalent: first, deletion and integration of the viral genome both inactivate the late coding region; second, the amount of viral early RNAs yielded by a single integrated copy appears to be very similar to that associated with several thousands of extrachromosomal copies of the viral genome. These data underline two essential requisites for hamster polyomavirus to become lymphomagenous: suppression of the late coding functions of the viral genome and expression of the viral oncogenes above a threshold level.     

Conditioned genome reconstruction: how to avoid choosing the conditioning genome

Genome phylogenies can be inferred from data on the presence and absence of genes across taxa. Logdet distances may be a good method, because they allow expected genome size to vary across the tree. Recently, Lake and Rivera proposed conditioned genome reconstruction (calculation of logdet distances using only those genes present in a conditioning genome) to deal with unobservable genes that are absent from every taxon of interest. We prove that their method can consistently estimate the topology for almost any choice of conditioning genome. Nevertheless, the choice of conditioning genome is important for small samples. For real bacterial genome data, different choices of conditioning genome can result in strong bootstrap support for different tree topologies. To overcome this problem, we developed supertree methods that combine information from all choices of conditioning genome. One of these methods, based on the BIONJ algorithm, performs well on simulated data and may have applications to other supertree problems. However, an analysis of 40 bacterial genomes using this method supports an incorrect clade of parasites. This is a common feature of model-based gene content methods and is due to parallel gene loss.     

Just click on the mouse!

The mouse genome is being sequenced in an efficient, coordinated manner. The map is complete and an assembly of whole genome shotgun data is available at a click of the mouse. In the final finished sequence phase, it seems a targeted approach would be more useful than random coverage. There are currently rich pickings available for mouse geneticists--that is to say, as long as their particular region of interest has been covered. Hard lessons learnt from the Human Genome Project have been put into practice, resulting in the elucidation of the mouse genome taking place with greater efficiency. The various genome centres have worked with coordination and a common strategy to generate a draft of the mouse genome in under a year, an achievement that would not have been deemed plausible a few years ago.     

The impact of temperature on the inactivation of enteric viruses in food and water: a review

Temperature is considered as the major factor determining virus inactivation in the environment. Food industries, therefore, widely apply temperature as virus inactivating parameter. This review encompasses an overview of viral inactivation and virus genome degradation data from published literature as well as a statistical analysis and the development of empirical formulae to predict virus inactivation. A total of 658 data (time to obtain a first log(10) reduction) were collected from 76 published studies with 563 data on virus infectivity and 95 data on genome degradation. Linear model fitting was applied to analyse the effects of temperature, virus species, detection method (cell culture or molecular methods), matrix (simple or complex) and temperature category (<50 and ≥50°C). As expected, virus inactivation was found to be faster at temperatures ≥50°C than at temperatures <50°C, but there was also a significant temperature-matrix effect. Virus inactivation appeared to occur faster in complex than in simple matrices. In general, bacteriophages PRD1 and PhiX174 appeared to be highly persistent whatever the matrix or the temperature, which makes them useful indicators for virus inactivation studies. The virus genome was shown to be more resistant than infectious virus. Simple empirical formulas were developed that can be used to predict virus inactivation and genome degradation for untested temperatures, time points or even virus strains.     

Drosophila genomes and the development of affordable molecular markers for species genotyping

The recent completion of genome sequencing of 12 species of Drosophila has provided a powerful resource for hypothesis testing, as well as the development of technical tools. Here we take advantage of genome sequence data from two closely related species of Drosophila, Drosophila simulans and Drosophila sechellia, to quickly identify candidate molecular markers for genotyping based on expected insertion or deletion (indel) differences between species. Out of 64 candidate molecular markers selected along the second and third chromosome of Drosophila, 51 molecular markers were validated using PCR and gel electrophoresis. We found that the 20% error rate was due to sequencing errors in the genome data, although we cannot rule out possible indel polymorphisms. The approach has the advantage of being affordable and quick, as it only requires the use of bioinformatics tools for predictions and a PCR and agarose gel based assay for validation. Moreover, the approach could be easily extended to a wide variety of taxa with the only limitation being the availability of complete or partial genome sequence data.     

Weighing the evidence for adaptation at the molecular level

The abundance of genome polymorphism and divergence data has provided unprecedented insight into how mutation, drift and natural selection shape genome evolution. Application of the McDonald-Kreitman (MK) test to such data indicates a pervasive influence of positive selection, particularly in Drosophila species. However, evidence for positive selection in other species ranging from yeast to humans is often weak or absent. Although evidence for positive selection could be obscured in some species, there is also reason to believe that the frequency of adaptive substitutions could be overestimated as a result of epistatic fitness effects or hitchhiking of deleterious mutations. Based on these considerations it is argued that the common assumption of independence among sites must be relaxed before abandoning the neutral theory of molecular evolution.     

Obtaining the sequence of the rice genome and lessons learned along the way

Rice holds the record for the largest number of separate genome projects and for having the genome of two subspecies sequenced. This might be a short-lived record in the genomics era, but it highlights the significance of rice as a food staple and as a model plant for cereal species. Clearly, obtaining the genome sequence four times seems redundant, yet the rationale and motivation for each of these projects is valid; whether it is serving corporate shareholders or the general scientific community. Although the multiple projects resulted in some duplicated efforts, the value of data sharing was obvious and the winner in the end will be the global public.     

Extrapolating ENCODE data to the whole human genome

The ENCODE (ENCyclopedia Of DNA Elements) project was launched three years ago with the purpose of identifying all of the functional elements in the human genome. ENCODE was started with 44 target sequences, which comprise 1% of the human genome. A crucial question about ENCODE is how representative it is of the human genome. Indeed, this is not a negligible problem if one considers that only 1% of the genome was selected for the project, and, more importantly, that the choice of the large DNA segments was based on two major criteria, namely the presence of extensively characterized genes and/or other functional elements, and the availability of a substantial amount of comparative sequence data. We found that the ENCODE data lead to an unbalanced representation of the compositional pattern of the human genome, especially for the GC-poorest and GC-richest regions. This unbalanced representativity of ENCODE can, however, be corrected by multiplying ENCODE data by a G/E factor (the ratio of whole genome data over ENCODE data), so amplifying the potential interest of ENCODE.     

Genomes as geography: using GIS technology to build interactive genome feature maps

Background  

Many commonly used genome browsers display sequence annotations and related attributes as horizontal data tracks that can be toggled on and off according to user preferences. Most genome browsers use only simple keyword searches and limit the display of detailed annotations to one chromosomal region of the genome at a time. We have employed concepts, methodologies, and tools that were developed for the display of geographic data to develop a Genome Spatial Information System (GenoSIS) for displaying genomes spatially, and interacting with genome annotations and related attribute data. In contrast to the paradigm of horizontally stacked data tracks used by most genome browsers, GenoSIS uses the concept of registered spatial layers composed of spatial objects for integrated display of diverse data. In addition to basic keyword searches, GenoSIS supports complex queries, including spatial queries, and dynamically generates genome maps. Our adaptation of the geographic information system (GIS) model in a genome context supports spatial representation of genome features at multiple scales with a versatile and expressive query capability beyond that supported by existing genome browsers.     

The revolution of the biology of the genome

Sequence data of entire eukaryotic genomes and their detailed comparison have provided new evidence on genome evolution. The major mechanisms involved in the increase of genome sizes are polyploidization and gene duplication.Subsequent gene silencing or mutations, preferentially in regulatory sequences of genes, modify the genome and permit the development of genes with new properties. Mechanisms such as lateral gene transfer, exon shuffling or the creation of new genes by transposition contribute to the evolution of a genome, but remain of relatively restricted relevance.Mechanisms to decrease genome sizes and, in particular, to remove specific DNA sequences, such as blocks of satellite DNAs, appear to involve the action of RNA interference (RNAi). RNAi mechanisms have been proven to be involved in chromatin packaging related with gene inactivation as well as in DNA excision during the macronucleus development in ciliates.     

The genome and the nucleus: a marriage made by evolution

Genomes are housed within cell nuclei as individual chromosome territories. Nuclei contain several architectural structures that interact and influence the genome. In this review, we discuss how the genome may be organised within its nuclear environment with the position of chromosomes inside nuclei being either influenced by gene density or by chromosomes size. We compare interphase genome organisation in diverse species and reveal similarities and differences between evolutionary divergent organisms. Genome organisation is also discussed with relevance to regulation of gene expression, development and differentiation and asks whether large movements of whole chromosomes are really observed during differentiation. Literature and data describing alterations to genome organisation in disease are also discussed. Further, the nuclear structures that are involved in genome function are described, with reference to what happens to the genome when these structures contain protein from mutant genes as in the laminopathies. Review related to the 15th International Chromosome Conference (ICC XV), held in September 2004, Brunel University, London, UK     

Phylogenetic reconstruction based on low copy DNA sequence data in an allopolyploid: the B genome of wheat.

Study of bread wheat (Triticum aestivum) may help to resolve several questions related to polyploid evolution. One such question regards the possibility that the component genomes of polyploids may themselves be polyphyletic, resulting from hybridization and introgression among different polyploid species sharing a single genome. We used the B genome of wheat as a model system to test hypotheses that bear on the monophyly or polyphyly of the individual constituent genomes. By using aneuploid wheat stocks, combined with PCR-based cloning strategies, we cloned and sequenced two single-copy-DNA sequences from each of the seven chromosomes of the wheat B genome and the homologous sequences from representatives of the five diploid species in section Sitopsis previously suggested as sister groups to the B genome. Phylogenetic comparisons of sequence data suggested that the B genome of wheat underwent a genetic bottleneck and has diverged from the diploid B genome donor. The extent of genetic diversity among the Sitopsis diploids and the failure of any of the Sitopsis species to group with the wheat B genome indicated that these species have also diverged from the ancestral B genome donor. Our results support monophyly of the wheat B genome.     

Structural organization of the genome of paramyxoviruses

The review summarizes the recent papers on the studies of primary structure of genome of a number of paramyxoviruses from the three genera of a family. The cited data demonstrate that despite the common principles of the genetic material arrangement shared by paramyxoviruses, they are variable in the genome, the primary structure of intragenic region, as well as the strategy of coding for some proteins. The data on the arrangement of the genetic material is discussed as useful as a criterion for classification of single stranded viruses with unsegmented genome.     

基于马尔可夫链的基因芯片特异性探针选择方法

对于基因表达芯片,特异性探针的选择是探针设计的重要环节,由于基因组序列数据量极大,不可能对每个候选探针都在全序列中进行特异性评价并进行取舍。对此问题,提出了一种采用马尔可夫链概率准则的探针特异性选择方法,即把基因组序列看作马尔可夫链,任何探针序列的互补序列作为它的一个子序列,都具有一定的出现概率,概率越小,越可能具有特异性。据此,选择其中概率最小的N个候选探针,能够大大减少进行特异性评价的探针数量,缩短探针设计的计算时间。对实际数据的测试结果表明,该方法选择的探针具有很高的特异性。     

Sampling,sequencing and the SAD

The Species Abundance Distribution (SAD) is a common metric for characterizing macroscopic ecological communities. Recently, this metric has been applied to analysis of microbial communities as well. However, as compared to macroscopic communities, sampling of microscopic communities is different. In particular, most microbial communities are studied using sequencing techniques. These techniques have known biases that result in certain taxa being detected more often than others, even if the taxa are present in the sample at equivalent abundances. There are, for example, amplification biases that result in some sequences being amplified more than others. Likewise, differences in genome size across organisms can result in different numbers of reads from different taxa, again resulting in biased detection. A number of bioinformatics methods have been devised to account for biases in sequencing data, allowing for more accurate estimates of relative taxon abundances. However, because the sampling process itself is affected by biased detection, and because sampling (and under-sampling in particular) can influence the shape of the SAD, it is possible that, even when corrected for through re-scaling, detection biases can affect SAD predictions from sequencing data. To test this hypothesis, we construct a simulation model of the sampling process, focusing on biased detection in shotgun sequencing that arises from genome size differences across microbial taxa. Interestingly, we find that, although genome size itself does not impact SAD predictions, predictions can vary depending on the range of genome sizes that are represented in a community, as well as how genome size is distributed (i.e., whether the majority of species have small versus large genomes). Our results suggest that care should be taken when comparing SADs across environments, particularly when those environments might have taxa with different genome size distributions. Furthermore, our results indicate that relatively deep sequencing might be required to avoid drawing spurious inferences about ecological differences across microbial communities.