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PubMed2004

Comparative genomics identifies genes shared by distantly related insect-transmitted plant pathogenic mollicutes.

Phytoplasmas and spiroplasmas are distantly related insect-transmitted plant pathogens within the class Mollicutes. Genome sequencing projects of phytoplasma strain Aster Yellows-Witches' Broom (AY-WB) and Spiroplasma kunkelii are near completion. Complete genome sequences of seven obligate animal and human pathogenic mollicutes (Mycoplasma and Ureaplasma spp.), and OY phytoplasma have been reported. Putative ORFs predicted from the genome sequences of AY-WB and S. kunkelii were compared to those of the completed genomes. This resulted in identification of at least three ORFs present in AY-WB, OY and S. kunkelii but not in the obligate animal and human pathogenic mollicutes. Moreover, we identified ORFs that seemed more closely related between AY-WB and S. kunkelii than to their mycoplasma counterparts. Phylogenetic analyses using parsimony were employed to study the origin of these genes, resulting in identification of one gene that may have undergone horizontal gene transfer. The possible involvement of these genes in plant pathogenicity is discussed.

Animals↗

PubMed2006

[Analysis of the secreted proteins encoded by genes in genome of filamental fungus (Neurospora crassa)].

The completed genome sequence of filamental fungus Neurospora crassa has completed and 10,082 open reading frames (ORFs) were predicted in whole genome. Among them, 437 proteins were secreted ones based on the combination of prediction algorithms SignalP v3.0 and PSORT, transmembrane domains prediction algorithms TMHMM v2.0 and THUMBUP, potential GPI-anchor sites prediction algorithm big-PI predictor and the subcellular protein location prediction algorithm TargetP v1.01. The minimum ORF length of the encoding sequences for 437 secreted proteins was 252 bp, the maximum ORF length was 6 604 bp and average was 1 433 bp. The length of signal peptide of secreted proteins ranged from 15 to 59 amino acids. Among the 437 secreted proteins, 205 were provided with function description. The function of secretory proteins were involved in kinds of enzyme, cell energy, transmission, cell recovered itself and defense mechanism et al. These results implied that processes catalyzed by these proteins probably occurred outside of cells, at least partially.

Amino Acid Sequence↗

PubMed2006

Identification and mapping of paralogous genes on a known genomic DNA sequence.

The completion of whole genome sequencing projects offers the opportunity to examine the organization of genes and the discovery of evolutionarily related genes in a given species. For the beginners in the field, through a specific example, this chapter provides a step-by-step procedure for identifying paralogous genes, using the genome browser at UCSC (http://genome.ucsc.edu/). The example describes identification and mapping in the human genome, the paralogs of TCF12/HTF4. The example identifies TCF3 and TCF4 as paralogs of the TCF12/HTF4 gene. The example also identifies a related sequence, corresponding to a pseudogene, in one of the introns of the JAK2 gene. The procedure described should be applicable to the discovery and creation of maps of paralogous genes in the genomic DNA sequences that are available at the genome browser at UCSC.

Amino Acid Sequence↗

PubMed2000

Identification of a highly diverged class of S-adenosylmethionine synthetases in the archaea.

S-adenosylmethionine is the primary alkylating agent in all known organisms. ATP:L-methionine S-adenosyltransferase (MAT) catalyzes the only known biosynthetic route to this central metabolite. Although the amino acid sequence of MAT is strongly conserved among bacteria and eukarya, no homologs have been recognized in the completed genome sequences of any archaea. In this study, MAT has been purified to homogeneity from the archaeon Methanococcus jannaschii, and the gene encoding it has been identified by mass spectrometry. The peptide mass map identifies the gene encoding MAT as MJ1208, a hypothetical open reading frame. The gene was cloned in Escherichia coli, and expressed enzyme has been purified and characterized. This protein has only 22 and 23% sequence identity to the E. coli and human enzymes, respectively, whereas those are 59% identical to each other. The few identical residues include the majority of those constituting the polar active site residues. Each complete archaeal genome sequence contains a homolog of this archaeal-type MAT. Surprisingly, three bacterial genomes encode both the archaeal and eukaryal/bacterial types of MAT. This identification of a second major class of MAT emphasizes the long evolutionary history of the archaeal lineage and the structural diversity found even in crucial metabolic enzymes.

Amino Acid Sequence↗

PubMed2002

Sequence diversity of Jeryl Lynn strain of mumps virus: quantitative mutant analysis for vaccine quality control.

The Jeryl Lynn strain of mumps vaccine live (MVL) was developed in 1966 by Merck Co. and has been widely used in the U.S. and other countries since the early 1970s. Partial sequencing has recently shown that the vaccine contains a mixture of two substrains with substantially different nucleotide sequences. We have determined the complete genomic sequences of both substrains and identified 414 nucleotide differences (2.69%), leading to 87 amino acid substitutions (1.67%). We used this information to develop methods for quantification of the substrain components in vaccine samples based on PCR and restriction enzyme cleavage and oligonucleotide microarray hybridization and monitored their dynamics in viral populations propagated in different conditions. Passaging Jeryl Lynn strain in Vero or CEF cell cultures resulted in rapid selection of the major component JL1, while growth in embryonated chicken eggs (ECE) favored accumulation of the minor component JL2. Based on the findings presented here, it is proposed that the substrain composition of Jeryl Lynn vaccine can be monitored as a part of its quality control to ensure consistency of the vaccine.

Amino Acid Sequence↗

PubMed2003

Analysis of conserved non-rRNA genes of Tropheryma whipplei.

The causative agent of Whipple's disease, Tropheryma whipplei, is a slow-growing bacterium that remains poorly-understood. Genetic characterization of this organism has relied heavily upon rRNA sequence analysis. Pending completion of a complete genome sequencing effort, we have characterized several conserved non-rRNA genes from T. whipplei directly from infected tissue using broad-range PCR and a genome-walking strategy. Our goals were to evaluate its phylogenetic relationships, and to find ways to expand the strain typing scheme, based on rDNA sequence comparisons. The genes coding for the ATP synthase beta subunit (atpD), elongation factor Tu (tuf), heat shock protein GroEL (groEL), beta subunit of DNA-dependent RNA polymerase (rpoB), and RNase P RNA (rnpB) were analyzed, as well as the regions upstream and downstream of the rRNA operon. Phylogenetic analyses with all non-rRNA marker molecules consistently placed T. whipplei within the class, Actinobacteria. The arrangement of genes in the atpD and rpoB chromosomal regions was also consistent with other actinomycete genomes. Tandem sequence repeats were found upstream and downstream of the rRNA operon, and downstream of the groEL gene. These chromosomal sites and the 16S-23S rRNA intergenic spacer regions were examined in the specimens of 11 patients, and a unique combination of tandem repeat numbers and spacer polymorphisms was found in each patient. These data provide the basis for a more discriminatory typing method for T. whipplei.

Actinomycetales↗

PubMed1998

DNA sequence sampling of the Streptococcus pneumoniae genome to identify novel targets for antibiotic development.

We initiated a survey of the Streptococcus pneumoniae genome by DNA sequence sampling. More than 9,500 random DNA sequences of approximately 500 bases average length were determined. Partial sequences sufficient to identify approximately 95% of the aminoacyl tRNA synthetase genes and ribosomal protein (rps) genes were found by comparing the database of partial sequences to known sequences from other organisms. Many genes involved in DNA replication, repair, and mutagenesis are present in S. pneumoniae. Genes for the major subunits of RNA polymerase are also present, as are genes for two alternative sigma factors, rpoD and rpoN. Many genes necessary for amino acid or cofactor biosynthesis and aerobic energy metabolism in other bacteria appear to be absent from the S. pneumoniae genome. A number of genes involved in cell wall biosynthesis and septation were identified, including six homologs to different penicillin binding proteins. Interestingly, four genes involved in the addition of D-alanine to lipoteicoic acid in other gram positive bacteria were found, even though the lipoteicoic acid in S. pneumoniae has not been shown to contain D-alanine. The S. pneumoniae genome contains a number of chaperonin genes similar to those found in other bacteria, but apparently does not contain genes involved in the type III secretion commonly observed in gram negative pathogens. The G+C content of S. pneumoniae genomic DNA is approximately 43 mole percent and the size of the genome is approximately 2.0 Mb as determined by pulsed-field gel electrophoresis. Many of the genes identified by sequence sampling have been physically mapped to the 19 different SmaI fragments derived from the S. pneumoniae genome. The database of random genome sequence tags (GSTs) provides the starting material for determining the complete genome sequence, gene disruption analysis, and comparative genomics to identify novel targets for antibiotic development.

Amino Acyl-tRNA Synthetases↗

PubMed2004

Cereulide, the emetic toxin of Bacillus cereus, is putatively a product of nonribosomal peptide synthesis.

AIMS: To determine if cereulide, the emetic toxin produced by Bacillus cereus, is produced by a nonribosomal peptide synthetase (NRPS). METHODS AND RESULTS: NC Y, an emetic strain of Bacillus cereus, was examined for a NRPS gene using PCR with primers recognizing a fragment of a NRPS gene from the cyanobacterium Microcystis. The amplicon was sequenced and compared with other gene sequences using BLAST analysis, which showed that the amplicon from strain NC Y was similar in sequence to peptide synthetase genes in other micro-organisms, including Bacillus subtilis and B. brevis, while no such sequence was found in the complete genome sequence of a nonemetic strain of B. cereus. Specific PCR primers were then designed and used to screen 40 B. cereus isolates previously implicated in outbreaks of foodborne illness. The isolates were also screened for toxin production using the MTT cell cytotoxicity assay. PCR and MTT assay screening of the B. cereus isolates revealed a high correlation between the presence of the NRPS gene and cereulide production. CONCLUSIONS: The results indicate that cereulide is produced by a NRPS complex. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study to provide evidence identifying the mechanism of production of cereulide, the emetic toxin of B. cereus. The PCR primers developed in the study allow determination of the potential for cereulide production among isolates of B. cereus.

Bacillus cereus↗

PubMed2003

Revamp a model-status and prospects of the Dictyostelium genome project.

International efforts are underway that aim at determining the complete genome sequence of the social amoeba Dictyostelium discoideum. As strategy, a whole chromosome shotgun (WCS) approach was chosen and each of the six Dictyostelium chromosomes was assigned to project partners. The project is well advanced, chromosome 2 was recently published, and it is expected that the sequences of chromosomes 1 and 6 and a gene catalogue for the complete genome will be available at the end of this year. The genome sequence, together with powerful molecular genetic tools, will undoubtedly further accelerate Dictyostelium research into a number of fundamental biological processes that are common to a wide range of eukaryotes. Furthermore, it will constitute the basis for genome-wide functional analyses. The integration of results from these studies should ultimately lead to a better understanding of the complex networks that govern cellular behavior and development.

Animals↗

PubMed2005

Comparison of the full-length genome sequence of avian metapneumovirus subtype C with other paramyxoviruses.

We determined the nucleotide (nt) sequence of the small hydrophobic (SH), attachment glycoprotein (G), and RNA polymerase (L) genes, plus the leader and trailer regions of the Colorado strain of Avian metapneumovirus subtype C (aMPV/C) in order to complete the genome sequencing. The complete genome comprised of 13,134 nucleotides, with a 40 nt leader at its 3' end and a 45 nt trailer at its 5' end. The aMPV/C L gene was the largest with 6173 nt and consisting of a single open reading frame encoding a 2005 amino acids (aa) protein. Comparison of the aMPV/C SH, G, and L nt and predicted aa sequences with those of Human metapneumoviruses (hMPV) revealed higher nt and aa sequence identities than the sequence identities between the aMPV subtypes A, B, C, and D, supporting earlier finding that aMPV/C was closer evolutionary to hMPV than the other aMPV subtypes.

Amino Acid Sequence↗

PubMed2001

Comparative genomics and bioenergetics.

Bacterial and archaeal complete genome sequences have been obtained from a wide range of evolutionary lines, which allows some general conclusions about the phylogenetic distribution and evolution of bioenergetic pathways to be drawn. In particular, I searched in the complete genomes for key enzymes involved in aerobic and anaerobic respiratory pathways and in photosynthesis, and mapped them into an rRNA tree of sequenced species. The phylogenetic distribution of these enzymes is very irregular, and clearly shows the diverse strategies of energy conservation used by prokaryotes. In addition, a thorough phylogenetic analysis of other bioenergetic protein families of wide distribution reveals a complex evolutionary history for the respective genes. A parsimonious explanation for these complex phylogenetic patterns and for the irregular distribution of metabolic pathways is that the last common ancestor of Bacteria and Archaea contained several members of every gene family as a consequence of previous gene or genome duplications, while different patterns of gene loss occurred during the evolution of every gene family. This would imply that the last universal ancestor was a bioenergetically sophisticated organism. Finally, important steps that occurred during the evolution of energetic machineries, such as the early evolution of aerobic respiration and the acquisition of eukaryotic mitochondria from a proteobacterium ancestor, are supported by the analysis of the complete genome sequences.

Archaea↗

PubMed2004

VirGen: a comprehensive viral genome resource.

VirGen is a comprehensive viral genome resource that organizes the 'sequence space' of viral genomes in a structured fashion. It has been developed with the objective of serving as an annotated and curated database comprising complete genome sequences of viruses, value-added derived data and data mining tools. The current release (v1.1) contains 559 complete genomes in addition to 287 putative genomes of viruses belonging to eight viral families for which the host range includes animals and plants. Viral genomes in VirGen are annotated using sequence-based Bioinformatics approaches. The genomic data is also curated to identify 'alternate names' of viral proteins, where available. VirGen archives the results of comparisons of genomes, proteomes and individual proteins within and between viral species. It is the first resource to provide phylogenetic trees of viral species computed using whole-genome sequence data. The module of predicted B-cell antigenic determinants in VirGen is an attempt to link the genome to its vaccinome. Comparative genome analysis data facilitate the study of genome organization and evolution of viruses, which would have implications in applied research to identify candidates for the design of vaccines and antiviral drugs. VirGen is a relational database and is available at http://bioinfo. ernet.in/virgen/virgen.html.

Antigens, Viral↗

PubMed2005

Comparative genomics of Dictyostelium discoideum and Entamoeba histolytica.

Amoebozoa represent one of the earliest branches from the last common ancestor of all eukaryotes and contain some of the most dangerous human pathogens. Two amoebozoan genomes -- from the model organism Dictyostelium discoideum and the human pathogen Entamoeba histolytica -- have been published this year. Owing to their high A+T content, both genomes were difficult to sequence. In addition to nine amoebozoan expressed sequence tag projects, efforts are underway for comparative sequencing of four additional Entamoeba species. The completed genome sequences of D. discoideum and E. histolytica revealed unusual telomere structures, a high percentage of repetitive elements and a remarkably high gene content that is close to the one of Drosophila melanogaster. Finally, both organisms are brilliant examples of the influence of the lifestyle of an organism on its genome.

Adaptation, Biological↗

PubMed2002

Complete nucleotide sequence and likely recombinatorial origin of bacteriophage T3.

We report the complete genome sequence (38,208 bp) of bacteriophage T3 and provide a bioinformatic comparative analysis with other completely sequenced members of the T7 group of phages. This comparison suggests that T3 has evolved from a recombinant between a T7-like coliphage and a yersiniophage. To assess this, recombination between T7 and the Yersinia enterocolitica serotype O:3 phage phiYeO3-12 was accomplished in vivo; coliphage progeny from this cross were selected that had many biological properties of T3. This represents the first experimentally observed recombination between lytic phages whose normal hosts are different bacterial genera.

Amino Acid Sequence↗

PubMed1993

Canine distemper terminal and intergenic non-protein coding nucleotide sequences: completion of the entire CDV genome sequence.

Sequences critical for the transcription and replication functions of canine distemper virus (CDV) RNA polymerase were analyzed. The sequence was obtained from polymerase chain reaction (PCR) products using either c-DNA clones from a genomic library as template or in most instances genomic CDV RNA. Clones coding for the precise 3'- and 5'-ends of the CDV genome were sequenced and the results confirmed by additional PCR experiments. The virtual identity of terminal sequences and spacing at the two noncoding ends speak to the importance of these areas in replication and transcription. The sequence for each of the CDV gene boundaries was defined and all were compared to related viruses. This report completes the sequence determination of the CDV genome.

Base Sequence↗

PubMed2006

Psittacid herpesvirus 1 and infectious laryngotracheitis virus: Comparative genome sequence analysis of two avian alphaherpesviruses.

Psittacid herpesvirus 1 (PsHV-1) is the causative agent of Pacheco's disease, an acute, highly contagious, and potentially lethal respiratory herpesvirus infection in psittacine birds, while infectious laryngotracheitis virus (ILTV) is a highly contagious and economically significant avian herpesvirus which is responsible for an acute respiratory disease limited to galliform birds. The complete genome sequence of PsHV-1 has been determined and compared to the ILTV sequence, assembled from published data. The PsHV-1 and ILTV genomes exhibit similar structural characteristics and are 163,025 bp and 148,665 bp in length, respectively. The PsHV-1 genome contains 73 predicted open reading frames (ORFs), while the ILTV genome contains 77 predicted ORFs. Both genomes contain an inversion in the unique long region similar to that observed in pseudorabies virus. PsHV-1 is closely related to ILTV, and it is proposed that it be assigned to the Iltovirus genus. These two avian herpesviruses represent a phylogenetically unique clade of alphaherpesviruses that are distinct from the Marek's disease-like viruses (Mardivirus). The determination of the complete genomic nucleotide sequences of PsHV-1 and ILTV provides a tool for further comparative and functional analysis of this unique class of avian alphaherpesviruses.

Amino Acid Sequence↗

PubMed2002

Complete nucleotide sequence of a Chilean hantavirus.

We have determined the genomic sequence of an Andes virus (ANDV) strain isolated from an infected Oligoryzomys longicaudatus rodent trapped in Chile in 1997. This strain, for which we propose the designation Chile R123, reproduces essential attributes of hantavirus pulmonary syndrome (HPS) when injected intramuscularly into laboratory hamsters (Hooper et al., Virology 289 (2001) 6-14). The L, M, and S segment sequences of Chile R123 are 6562, 3671, and 1871 nt long, respectively, with an overall G+C content of 38.5%. These respective genome segments could encode a 247 kd RNA-dependent RNA polymerase (RdRP), 126 kd glycoprotein precursor (GPC), and 48 kd nucleocapsid (N) protein, in line with other Sigmodontine rodent-associated hantaviruses. Among hantaviruses for which complete genomic sequences are available, Chile R123 is most closely related to Sin Nombre virus (SNV) strain NM R11, with greater than 85% amino acid identity between translated L and S segments and 78% amino acid identity between translated M segments. Because Chile R123 shares essentially 100% amino acid identity in regions of overlap with partially sequenced Argentinian and Chilean ANDV strains, Syrian hamster pathogenicity and the potential for interhuman transmission are features likely common to all ANDV strains.

Animals↗

PubMed2005

Genome sequence of the thermostable Newcastle disease virus (strain I-2) reveals a possible phenotypic locus.

The complete genome sequence of the Australian I-2 heat-tolerant Newcastle disease virus (NDV) vaccine (master seed stocks) was determined and compared to the sequence of the parent virus from which it had been derived after exposure of the parent stock at 56 degrees C for 30 min. Nucleotide changes were observed at a number of positions with synonymous mutations being greater than those observed for non-synonymous mutations. Sequence data for the HN gene of a parental culture of V4 and two heat-tolerant variants of V4 were obtained. These were compared with the data for the I-2 viruses and with published sequences for parental V4 and for a number of ND vaccine strains. Sequence analyses did not reveal the ARG(303) deletion in the HN protein, previously claimed to be responsible for the thermostable phenotype. No consistent changes were detected that would indicate involvement of the HN protein in heat resistance. The majority of alterations were observed in the L protein of the virus and it is proposed that these alterations were responsible for the heat-tolerant phenotype of the I-2 NDV vaccine.

Amino Acid Sequence↗