Genomic analysis of the causative agents of coccidiosis in domestic chickens.

Global production of chickens has trebled in the past two decades and they are now the most important source of dietary animal protein worldwide. Chickens are subject to many infectious diseases that reduce their performance and productivity. Coccidiosis, caused by apicomplexan protozoa of the genus Eimeria, is one of the most important poultry diseases. Understanding the biology of Eimeria parasites underpins development of new drugs and vaccines needed to improve global food security. We have produced annotated genome sequences of all seven species of Eimeria that infect domestic chickens, which reveal the full extent of previously described repeat-rich and repeat-poor regions and show that these parasites possess the most repeat-rich proteomes ever described. Furthermore, while no other apicomplexan has been found to possess retrotransposons, Eimeria is home to a family of chromoviruses. Analysis of Eimeria genes involved in basic biology and host-parasite interaction highlights adaptations to a relatively simple developmental life cycle and a complex array of co-expressed surface proteins involved in host cell binding.

Comparative genomic analysis of three Leishmania species that cause diverse human disease.

Leishmania parasites cause a broad spectrum of clinical disease. Here we report the sequencing of the genomes of two species of Leishmania: Leishmania infantum and Leishmania braziliensis. The comparison of these sequences with the published genome of Leishmania major reveals marked conservation of synteny and identifies only approximately 200 genes with a differential distribution between the three species. L. braziliensis, contrary to Leishmania species examined so far, possesses components of a putative RNA-mediated interference pathway, telomere-associated transposable elements and spliced leader-associated SLACS retrotransposons. We show that pseudogene formation and gene loss are the principal forces shaping the different genomes. Genes that are differentially distributed between the species encode proteins implicated in host-pathogen interactions and parasite survival in the macrophage.

The genome of the blood fluke Schistosoma mansoni.

Schistosoma mansoni is responsible for the neglected tropical disease schistosomiasis that affects 210 million people in 76 countries. Here we present analysis of the 363 megabase nuclear genome of the blood fluke. It encodes at least 11,809 genes, with an unusual intron size distribution, and new families of micro-exon genes that undergo frequent alternative splicing. As the first sequenced flatworm, and a representative of the Lophotrochozoa, it offers insights into early events in the evolution of the animals, including the development of a body pattern with bilateral symmetry, and the development of tissues into organs. Our analysis has been informed by the need to find new drug targets. The deficits in lipid metabolism that make schistosomes dependent on the host are revealed, and the identification of membrane receptors, ion channels and more than 300 proteases provide new insights into the biology of the life cycle and new targets. Bioinformatics approaches have identified metabolic chokepoints, and a chemogenomic screen has pinpointed schistosome proteins for which existing drugs may be active. The information generated provides an invaluable resource for the research community to develop much needed new control tools for the treatment and eradication of this important and neglected disease.

Comparative genomics of the fungal pathogens Candida dubliniensis and Candida albicans.

Candida dubliniensis is the closest known relative of Candida albicans, the most pathogenic yeast species in humans. However, despite both species sharing many phenotypic characteristics, including the ability to form true hyphae, C. dubliniensis is a significantly less virulent and less versatile pathogen. Therefore, to identify C. albicans-specific genes that may be responsible for an increased capacity to cause disease, we have sequenced the C. dubliniensis genome and compared it with the known C. albicans genome sequence. Although the two genome sequences are highly similar and synteny is conserved throughout, 168 species-specific genes are identified, including some encoding known hyphal-specific virulence factors, such as the aspartyl proteinases Sap4 and Sap5 and the proposed invasin Als3. Among the 115 pseudogenes confirmed in C. dubliniensis are orthologs of several filamentous growth regulator (FGR) genes that also have suspected roles in pathogenesis. However, the principal differences in genomic repertoire concern expansion of the TLO gene family of putative transcription factors and the IFA family of putative transmembrane proteins in C. albicans, which represent novel candidate virulence-associated factors. The results suggest that the recent evolutionary histories of C. albicans and C. dubliniensis are quite different. While gene families instrumental in pathogenesis have been elaborated in C. albicans, C. dubliniensis has lost genomic capacity and key pathogenic functions. This could explain why C. albicans is a more potent pathogen in humans than C. dubliniensis.

Characterization of a novel wood mouse virus related to murid herpesvirus 4.

Two novel gammaherpesviruses were isolated, one from a field vole (Microtus agrestis) and the other from wood mice (Apodemus sylvaticus). The genome of the latter, designated wood mouse herpesvirus (WMHV), was completely sequenced. WMHV had the same genome structure and predicted gene content as murid herpesvirus 4 (MuHV4; murine gammaherpesvirus 68). Overall nucleotide sequence identity between WMHV and MuHV4 was 85 % and most of the 10 kb region at the left end of the unique region was particularly highly conserved, especially the viral tRNA-like sequences and the coding regions of genes M1 and M4. The partial sequence (71 913 bp) of another gammaherpesvirus, Brest herpesvirus (BRHV), which was isolated ostensibly from a white-toothed shrew (Crocidura russula), was also determined. The BRHV sequence was 99.2 % identical to the corresponding portion of the WMHV genome. Thus, WMHV and BRHV appeared to be strains of a new virus species. Biological characterization of WMHV indicated that it grew with similar kinetics to MuHV4 in cell culture. The pathogenesis of WMHV in wood mice was also extremely similar to that of MuHV4, except for the absence of inducible bronchus-associated lymphoid tissue at day 14 post-infection and a higher load of latently infected cells at 21 days post-infection.

Artemis and ACT: viewing, annotating and comparing sequences stored in a relational database.

MOTIVATION: Artemis and Artemis Comparison Tool (ACT) have become mainstream tools for viewing and annotating sequence data, particularly for microbial genomes. Since its first release, Artemis has been continuously developed and supported with additional functionality for editing and analysing sequences based on feedback from an active user community of laboratory biologists and professional annotators. Nevertheless, its utility has been somewhat restricted by its limitation to reading and writing from flat files. Therefore, a new version of Artemis has been developed, which reads from and writes to a relational database schema, and allows users to annotate more complex, often large and fragmented, genome sequences. RESULTS: Artemis and ACT have now been extended to read and write directly to the Generic Model Organism Database (GMOD, http://www.gmod.org) Chado relational database schema. In addition, a Gene Builder tool has been developed to provide structured forms and tables to edit coordinates of gene models and edit functional annotation, based on standard ontologies, controlled vocabularies and free text. AVAILABILITY: Artemis and ACT are freely available (under a GPL licence) for download (for MacOSX, UNIX and Windows) at the Wellcome Trust Sanger Institute web sites: http://www.sanger.ac.uk/Software/Artemis/ http://www.sanger.ac.uk/Software/ACT/

GeneDB: a resource for prokaryotic and eukaryotic organisms.

GeneDB (http://www.genedb.org/) is a genome database for prokaryotic and eukaryotic organisms. The resource provides a portal through which data generated by the Pathogen Sequencing Unit at the Wellcome Trust Sanger Institute and other collaborating sequencing centres can be made publicly available. It combines data from finished and ongoing genome and expressed sequence tag (EST) projects with curated annotation, that can be searched, sorted and downloaded, using a single web based resource. The current release stores 11 datasets of which six are curated and maintained by biologists, who review and incorporate information from the scientific literature, public databases and the respective research communities.

Integration of tools and resources for display and analysis of genomic data for protozoan parasites.

Centralisation of tools for analysis of genomic data is paramount in ensuring that research is always carried out on the latest currently available data. As such, World Wide Web sites providing a range of online analyses and displays of data can play a crucial role in guaranteeing consistency of in silico work. In this respect, the protozoan parasite research community is served by several resources, either focussing on data and tools for one species or taking a broader view and providing tools for analysis of data from many species, thereby facilitating comparative studies. In this paper, we give a broad overview of the online resources available. We then focus on the GeneDB project, detailing the features and tools currently available through it. Finally, we discuss data curation and its importance in keeping genomic data 'relevant' to the research community.

Sequencing and analysis of chromosome 1 of Eimeria tenella reveals a unique segmental organization.

Eimeria tenella is an intracellular protozoan parasite that infects the intestinal tracts of domestic fowl and causes coccidiosis, a serious and sometimes lethal enteritis. Eimeria falls in the same phylum (Apicomplexa) as several human and animal parasites such as Cryptosporidium, Toxoplasma, and the malaria parasite, Plasmodium. Here we report the sequencing and analysis of the first chromosome of E. tenella, a chromosome believed to carry loci associated with drug resistance and known to differ between virulent and attenuated strains of the parasite. The chromosome--which appears to be representative of the genome--is gene-dense and rich in simple-sequence repeats, many of which appear to give rise to repetitive amino acid tracts in the predicted proteins. Most striking is the segmentation of the chromosome into repeat-rich regions peppered with transposon-like elements and telomere-like repeats, alternating with repeat-free regions. Predicted genes differ in character between the two types of segment, and the repeat-rich regions appear to be associated with strain-to-strain variation.

Common inheritance of chromosome Ia associated with clonal expansion of Toxoplasma gondii.

Toxoplasma gondii is a globally distributed protozoan parasite that can infect virtually all warm-blooded animals and humans. Despite the existence of a sexual phase in the life cycle, T. gondii has an unusual population structure dominated by three clonal lineages that predominate in North America and Europe, (Types I, II, and III). These lineages were founded by common ancestors approximately10,000 yr ago. The recent origin and widespread distribution of the clonal lineages is attributed to the circumvention of the sexual cycle by a new mode of transmission-asexual transmission between intermediate hosts. Asexual transmission appears to be multigenic and although the specific genes mediating this trait are unknown, it is predicted that all members of the clonal lineages should share the same alleles. Genetic mapping studies suggested that chromosome Ia was unusually monomorphic compared with the rest of the genome. To investigate this further, we sequenced chromosome Ia and chromosome Ib in the Type I strain, RH, and the Type II strain, ME49. Comparative genome analyses of the two chromosomal sequences revealed that the same copy of chromosome Ia was inherited in each lineage, whereas chromosome Ib maintained the same high frequency of between-strain polymorphism as the rest of the genome. Sampling of chromosome Ia sequence in seven additional representative strains from the three clonal lineages supports a monomorphic inheritance, which is unique within the genome. Taken together, our observations implicate a specific combination of alleles on chromosome Ia in the recent origin and widespread success of the clonal lineages of T. gondii.

ACT: the Artemis Comparison Tool.

The Artemis Comparison Tool (ACT) allows an interactive visualisation of comparisons between complete genome sequences and associated annotations. The comparison data can be generated with several different programs; BLASTN, TBLASTX or Mummer comparisons between genomic DNA sequences, or orthologue tables generated by reciprocal FASTA comparison between protein sets. It is possible to identify regions of similarity, insertions and rearrangements at any level from the whole genome to base-pair differences. ACT uses Artemis components to display the sequences and so inherits powerful searching and analysis tools. ACT is part of the Artemis distribution and is similarly open source, written in Java and can run on any Java enabled platform, including UNIX, Macintosh and Windows.

A comprehensive survey of the Plasmodium life cycle by genomic, transcriptomic, and proteomic analyses.

Plasmodium berghei and Plasmodium chabaudi are widely used model malaria species. Comparison of their genomes, integrated with proteomic and microarray data, with the genomes of Plasmodium falciparum and Plasmodium yoelii revealed a conserved core of 4500 Plasmodium genes in the central regions of the 14 chromosomes and highlighted genes evolving rapidly because of stage-specific selective pressures. Four strategies for gene expression are apparent during the parasites' life cycle: (i) housekeeping; (ii) host-related; (iii) strategy-specific related to invasion, asexual replication, and sexual development; and (iv) stage-specific. We observed posttranscriptional gene silencing through translational repression of messenger RNA during sexual development, and a 47-base 3' untranslated region motif is implicated in this process.

The genome of the kinetoplastid parasite, Leishmania major.

Leishmania species cause a spectrum of human diseases in tropical and subtropical regions of the world. We have sequenced the 36 chromosomes of the 32.8-megabase haploid genome of Leishmania major (Friedlin strain) and predict 911 RNA genes, 39 pseudogenes, and 8272 protein-coding genes, of which 36% can be ascribed a putative function. These include genes involved in host-pathogen interactions, such as proteolytic enzymes, and extensive machinery for synthesis of complex surface glycoconjugates. The organization of protein-coding genes into long, strand-specific, polycistronic clusters and lack of general transcription factors in the L. major, Trypanosoma brucei, and Trypanosoma cruzi (Tritryp) genomes suggest that the mechanisms regulating RNA polymerase II-directed transcription are distinct from those operating in other eukaryotes, although the trypanosomatids appear capable of chromatin remodeling. Abundant RNA-binding proteins are encoded in the Tritryp genomes, consistent with active posttranscriptional regulation of gene expression.

Genome of the host-cell transforming parasite Theileria annulata compared with T. parva.

Theileria annulata and T. parva are closely related protozoan parasites that cause lymphoproliferative diseases of cattle. We sequenced the genome of T. annulata and compared it with that of T. parva to understand the mechanisms underlying transformation and tropism. Despite high conservation of gene sequences and synteny, the analysis reveals unequally expanded gene families and species-specific genes. We also identify divergent families of putative secreted polypeptides that may reduce immune recognition, candidate regulators of host-cell transformation, and a Theileria-specific protein domain [frequently associated in Theileria (FAINT)] present in a large number of secreted proteins.

The mei3 region of the Schizosaccharomyces pombe genome.

Expression of the mei3 gene is sufficient to induce meiosis in the fission yeast Schizosaccharomyces pombe. The mei3 gene is located 0.64 Mb from the telomere of the left arm of Sz. pombe chromosome II. We have sequenced and analysed 107 kb of DNA from the mei3 genomic region. The sequence includes 14 known genes (bag1-B, csh3, dps1, gpt1, mei3, mfm3, pac1, prp31, rpl38-1, rpn3, rti1, spa1, spm1 and ubc4) and 26 other open reading frames (ORFs) longer than 100 codons: a density of one protein-coding gene per 2.7 kb. Twenty-one of the 40 ORFs (53%) have introns. In addition there is one lone Tf1 transposon long terminal repeat (LTR), tRNA(Trp) and tRNA(Ser) genes and a 5S rRNA gene. 14 of the novel ORFs show sequence similarities which suggest functions of their products, including a coatomer alpha-subunit, a catechol O-methyltransferase, protein kinase, asparagine synthetase, zinc metalloprotease, acetyltransferase, phosphatidylinositol 4-kinase, inositol polyphosphate phosphatase, GTPase-activating protein, permease, pre-mRNA splicing factor, 20S proteasome component and a thioredoxin-like protein. One predicted protein has similarity to the human Cockayne syndrome protein CSA and one with human GTPase XPA binding protein XAB1. Three ORFs are likely to code for proteins because they have sequence similarity with hypothetical proteins, three encode predicted coiled-coil proteins and four are sequence orphans.

Insight into the genome of Aspergillus fumigatus: analysis of a 922 kb region encompassing the nitrate assimilation gene cluster.

Aspergillus fumigatus is the most ubiquitous opportunistic filamentous fungal pathogen of human. As an initial step toward sequencing the entire genome of A. fumigatus, which is estimated to be approximately 30 Mb in size, we have sequenced a 922 kb region, contained within 16 overlapping bacterial artificial chromosome (BAC) clones. Fifty-four percent of the DNA is predicted to be coding with 341 putative protein coding genes. Functional classification of the proteins showed the presence of a higher proportion of enzymes and membrane transporters when compared to those of Saccharomyces cerevisiae. In addition to the nitrate assimilation gene cluster, the quinate utilisation gene cluster is also present on this 922 kb genomic sequence. We observed large scale synteny between A. fumigatus and Aspergillus nidulans by comparing this sequence to the A. nidulans genetic map of linkage group VIII.